Proceedings of the International Dental Materials Congress 2016

I D MC2016 http://idmc2016.com/http://idmc2016.com/

Dental Materials and Technology to the Next Generation

In conjunction with 68th Meeting of 2016 Annual Meeting of the Japanese Society the Indonesian Society for for Dental Materials Dental Materials Science and Devices and Technology

Chairperson November 4-6, 2016 Professor Anita Yuliati Legian, Bali, Indonesia Airlangga University TheTh StStones HHotel t l

Table of Contents

Welcome Messages ……………………………..……………………………………………… 1

Organizing Committee ………………………………………….….………………………….. 3

Contributors ……………………………………….…………………………………………… 5

Map of Conference Site ………………………………………………………………...……… 6

Meeting Schedule …………………….………………………………………….…….…….… 7

General Session Program Oral Presentation ……………………………………….…………………………….. 10 Poster Presentation …………………………………………………………………… 15

Invited Lecture 1 “Surface treatment and modification of metals to add biofunctions” Takao Hanawa (Tokyo Medical and Dental University, Japan) ……………………… 26

Invited Lecture 2 “Surface modifications for enhanced integration of titanium implant with soft and hard tissues and prevention of peri-implantitis” Kazuhiko Endo (Health Sciences University of Hokkaido, Japan) …..……………. 27

Invited Lecture 3 “Carbonate apatite bone replacement” Kunio Ishikawa (Kyusyu University, Japan) ……………………………….………….. 28

Invited Lecture 4 “Carbonate apatite ceramics for drug delivery and synthetic scaffold in tissue engineering: a review of the development and application in dentistry” Ika Dewi Ana (Gadjah Mada University, Indonesia) …………………………….…… 29

Invited Lecture 5 “Prosthetic rehabilitations using pure titanium to avoid metal combinations” Chikahiro Ohkubo (Tsurumi University, Japan) ……………….……………………… 30

Invited Lecture 6 “Non-biodegradable polymer particles for drug delivery: A new technology for ‘bio-active’ restorative materials” Satoshi Imazato (Osaka University, Japan) ……………..…….……………………… 31 Invited Lecture 7 “Reaction supporting dental tissue on endosseous dental implants using titanium materials, based on immunoexpression of laminin” Nina Djustiana (Padjajaran University, Indonesia) ………………….……………….. 32

Invited Lecture 8 “Design and fabrication of orthodontic bracket using metal injection molding (MIM) process” Bambang Irawan (University of Indonesia, Indonesia) ………..…………………….. 33

Invited Lecture 9 “Properties of super-elastic Ni-Ti alloy for dental applications” Takayuki Yoneyama (Nihon University, Japan) …………………………………….….. 34

Luncheon Seminar ………………………………………………………………………….…. 35

Abstracts Oral Presentation …………………………………………………….……………….. 36 Poster Presentation …………………………………………………………………… 72

Welcome to IDMC2016

Dear Colleagues,

It is my great honor to welcome you to the beautiful island of Bali, Indonesia and to International Dental Materials Congress 2016 (IDMC). IDMC 2016 is the first international congress of dental materials organized under collaboration between the Japanese Society for Dental Materials and Devices and the Indonesian Society for Dental Materials Sciences and Technology. The part of our obligation as country members and as an executive committee of IDMC 2016 is to foster advancement and improvement services/education of dental materials in Asia and particularly in Indonesia.

The theme of this IDMC 2016 is “Dental Materials and Technology to the Next Generation”. This addresses to meet our aims and objective, which are to provide our next generation of professional and skillful researchers with a sound and continuously updated knowledge.

We sincerely hope that the benefits of this meeting would cover the aspect of knowledge sharing, networking, friendship, which eventually would strengthen our close ties and presence. To our sponsors and trade exhibitors, we give our token of appreciation for your support to make this conference a success.

We hope that you would also have some time to enjoy the beautiful sceneries of Bali with its worldwide known beaches, lakes and cultural heritage.

I hope this event will bring you faithful and enjoyable memories.

Enjoy your stay in Bali as one of the best Indonesian islands of culture.

Sincerely,

Prof. Dr. Anita Yuliati, DDS, MSc. Chairperson, IDMC 2016 Airlangga University

Welcome to IDMC2016

It is a great honor and an enormous pleasure to hold the International Dental Materials Congress 2016 (IDMC) here in Bali, Indonesia.

First, on behalf of the Japanese Society for Dental Materials and Devices, I would like to express special thanks to the members of the Indonesian Society for Dental Materials Science and Technology, especially to Prof. Anita Yuliati, Chairperson of Airlangga University. This meeting is a joint meeting of the Indonesian Society for Dental Materials Science and Technology and the Japanese Society for Dental Materials and Devices. The preparatory committees of both countries collaborated to organize the IDMC.

The Japanese Society for Dental Materials and Devices has been holding international meetings since 1989. The initial meeting was in Hawaii, and subsequent meetings have been held roughly every 4 years in several countries. This is our first International Congress in Indonesia. We will have more than 150 presentations and an estimated attendance of more than 250, due to the great efforts of the preparatory committees.

The main theme of this meeting is “Dental Materials and Technology to the Next Generation”, as Professor Anita Yuliati stated. From this perspective, there are many novel studies appearing as abstracts, especially those by remarkable young researchers who will be the main thrust for new, evolving scientific studies in the next generation.

The serendipitous exchange of scientific information among researchers here in the colorful, bright atmosphere of Bali should lead to new investigations.

I hope this meeting will be fruitful and contribute not only to collaboration between the Indonesian and Japanese Societies but also to worldwide collaboration.

Thank you.

Tatsushi Kawai, DDS, Ph.D. President, The Japanese Society for Dental Materials and Devices Aichi Gakuin University

Organizing Committee

Organizers The Japanese Society for Dental Materials and Devices The Indonesian Society for Dental Materials Science and Technology Universitas Airlangga The Indonesian Organizing Comitte for IDMC 2016

Chairperson Prof. Dr. Anita Yuliati, DDS., MSc. (Airlangga University, Indonesia)

Vice Chairman Prof. Seno Pradopo, DDS., Ph.D. (Airlangga University, Indonesia) Udijanto Tedjosasongko, DDS., Ph.D.

Secretary Mega Moeharyono P, DDS., Ph.D. Irma Josephina, DDS., Ph.D.

Treasurer Asti Meizarini, drg., M.S Tania Saskianti, drg., Sp.KGA., Ph.D.

Scientific Commitee Dr. Intan Nirwana, drg., M.Kes Ketut Suardita, drg., Ph.D., Sp.KG Priyawan Rachmadi, drg., Ph.D. Andra Rizqiawan, drg., Ph.D., Sp.BM

Event / program Endanus Harijanto, drg., M.Kes Sri Yogyarti, drg., M.S

Fund & Exhibition Dr. Elly Munadziroh, drg., M.S Harry Laksono, drg., Sp.Pros., M.Kes

Registration Devi Rianti, drg., M.Kes Titien H.Agustantina, drg., M.Kes

Banquet Nurina F Ayuningtyas, drg., M.Kes., Ph.D.

Accommodations & Materials R.M.Yogiartono, drg., M.Kes Soebagio, drg., M.Kes

Publication & Documentation R.Helal Soekartono, drg., M.Kes

Indonesia Liaison Officer for Japan Maretaningtyas Dwi Ariani, DDS., Ph.D.

The Japanese Society for Dental Materials and Devices

Co-chair, Organizing Committee Thoru Hayakawa, Ph.D. (Tsurumi University) Koichi Kato, Ph.D. (Hiroshima University) Osamu Suzuki, Ph.D. (Tohoku University)

Organizing Committee Tatsushi Kawai, DDS, Ph.D. (Aichi Gakuin University) Satoshi Imazato, DDS, Ph.D. (Osaka University) Motohiro Uo, Ph.D. (Tokyo Medical and Dental University) Masayuki Hattori, DDS, Ph.D. (Iwate Medical University) Shozo Tsuruta, DDS, Ph.D. (Aichi Gakuin University) Yoshiya Hashimoto, DDS, Ph.D. (Osaka Dental University) Isao Hirata, Ph.D. (Hiroshima University) Kanji Tsuru, Ph.D. (Kyushu University) Yusuke Tsutsumi, Ph.D. (Tokyo Medical and Dental University) Takashi Saito, DDS, Ph.D. (Health Sciences University of Hokkaido) Chikahiro Ohkubo, DMD, Ph.D. (Tsurumi University) Guang Hong, MD, DDS, Ph.D. (Tohoku University)

Contributors

Map of Conference Site

The Stones Hotel – Legian Bali Jl. Raya Pantai Kuta, Banjar Legian Kelod, Legian, Bali 80361, Indonesia

Meeting Schedule

Friday, November 4

12:00 - 20:00 Registration 13:00 - Opening Remarks (Ballroom 2) 13:20 - 13:50 Invited Lecture 1 (Ballroom 2) “Surface treatment and modification of metals to add biofunctions” Coordinator: Takayuki Yoneyama (Nihon University, Japan) Speaker: Takao Hanawa (Tokyo Medical and Dental University, Japan) 13:50 - 14:20 Invited Lecture 2 (Ballroom 2) “Surface modifications for enhanced integration of titanium implant with soft and hard tissues and prevention of peri-implantitis” Coordinator: Takayuki Yoneyama (Nihon University, Japan) Speaker: Kazuhiko Endo (Health Sciences University of Hokkaido, Japan) 14:20 - 14:50 Coffee Break 14:50 - 16:00 Oral Presentation (O-01– O-07) (Ballroom2) 16:10 - 16:40 Invited Lecture 3 (Ballroom 2) “Carbonate apatite bone replacement” Coordinator: Osamu Suzuki (Tohoku University, Japan) Speaker: Kunio Ishikawa (Kyusyu University, Japan) 16:40 - 17:10 Invited Lecture 4 (Ballroom 2) “Carbonate apatite ceramics for drug delivery and synthetic scaffold in tissue engineering: a review of the development and application in dentistry” Coordinator: Osamu Suzuki (Tohoku University, Japan) Speaker: Ika Dewi Ana (Gadjah Mada University, Indonesia) 17:10 - 17:40 Invited Lecture 5 (Ballroom 2) “Prosthetic rehabilitations using pure titanium to avoid metal combinations” Coordinator: Thoru Hayakawa (Tsurumi University, Japan) Speaker: Chikahiro Ohkubo (Tsurumi University, Japan) 18:00 - 20:00 Welcome Reception (Ballroom 3)

Saturday, November 5

8:30 - 17:00 Registration 9:30 - 10:00 Invited Lecture 6 (Ballroom 2) “Non-biodegradable polymer particles for drug delivery: A new technology for ‘bio-active’ restorative materials” Coordinator: Takuya Matsumoto (Okayama University, Japan) Speaker: Satoshi Imazato (Osaka University, Japan) 7

10:00 - 10:50 Oral Presentation (O-08–O-12) (Ballroom 2) 10:50 - 11:10 Coffee Break 10:00 - 11:30 Poster Presentation (P-01–P-60) (Ballroom 3) 11:10 - 12:10 Oral Presentation (O-13–O-18) (Ballroom 2) 12:20 - 13:20 Lunchon seminar (Ballroom 2) Sponsor: SHOFU INC. “Possibility of bioactive materials containing Surface Pre-Reacted Glass-ionomer Filler” Coordinator: Thoru Hayakawa (Tsurumi University, Japan) Speaker: Shuichi Ito (Health Sciences University of Hokkaido, Japan) 13:30 - 14:00 Invited Lecture 7 (Ballroom 2) “Reaction supporting dental tissue on endosseous dentalimplants using titanium materials, based on immunoexpression of laminin” Coordinator: Koichi Kato (Hiroshima University, Japan) Speaker: Nina Djustiana (Padjajaran University, Indonesia) 14:00 - 14:30 Invited Lecture 8 (Ballroom 2) “Design and fabrication of orthodontic bracket using Metal Injection Molding (MIM) process” Coordinator: Koichi Kato (Hiroshima University, Japan) Speaker: Bambang Irawan (University of Indonesia, Indonesia) 14:30 - 15:00 Invited Lecture 9 (Ballroom 2) “Properties of super-elastic Ni-Ti alloy for dental applications” Coordinator: Takao Hanawa (Tokyo Medical and Dental University, Japan) Speaker: Takayuki Yoneyama (Nihon University, Japan) 15:00 - 15:30 Coffee Break 15:30 - 17:00 Poster Presentation (P-61–P-116) (Ballroom 3) 15:30 - 16:10 Oral Presentation (O-19–O-22) (Ballroom 2) 16:20 - 17:00 Oral Presentation (O-23–O-26) (Ballroom 2)

Sunday, November 6

9:00 - 11:30 Registration 9:30 - 10:10 Oral Presentation (O-27–O-30) (Ballroom 2) 10:10 - 10:30 Coffee Break 10:30 - 11:30 Oral Presentation (O-31–O-36) (Ballroom 2) 11:40 - Closing Remarks (Ballroom 2)

General Session Program

Oral Presentation Ⅰ(O-01 - O-07) (Ballroom2): Friday, November 4, 14:50 - 16:00 (Biological Reaction) Chair: Tsukasa Akasaka (Hokkaido University), Isao Hirata (Hiroshima University)

O-01 Possibility of bone regeneration of bone-like tissue induced by recombinant human BMPs in vitro *Hayashi T, Kawase M, Hiroyasu K, Tomino M and Kawai T (Aichi Gakuin Univ., Nagoya, Japan)

O-02 Cell Proliferation Ability of Mouse Fibroblast-Like Cells and Osteoblast-Like Cells on Pure Titanium Film Manufactured by Electron Beam Melting *Kawase M, Hayashi T, Asakura M, Mieki A, Tomino M and Kawai T (Aichi Gakuin Univ., Nagoya, Japan)

O-03 Stem cell behavior on micro porous titanium implant Bei Chang1, Tianxiao Han1, Fuping Li2, Hongchao Kou2, *Yumei Zhang1 (1The Fourth Military Medical University, Xi’an, People’s Republic of China, 2Northwestern Polytechnical University, Xi'an, People’s Republic of China)

O-04 Chondrocyte differentiation ability of dedifferentiated fat cells compared with adipose-derived stem cells derived from the human buccal fat pad *Kubo H, Nishio A, Miya Y, Hashimoto Y, Kishimoto N ( Osaka Dental University, Osaka, Japan)

O-05 In vivo evaluation of H2O2 hydrothermal surface modification with FGF-2 of mini-implants to enhance new bone formation *Matsuno T1, Yoneyama2 Y, Miki T1, Asano K1 and Mataga I1 (1 The Nippon Dental Univ. School of Life Dentistry, Tokyo, Japan, 2 Tokyo Medical Univ. Tokyo, Japan)

O-06 withdraw

O-07 withdraw

Oral Presentation Ⅱ(O-08 - O-12) (Ballroom2): Saturday, November 5, 10:00 - 10:50 (Calcium Phophate) Chair: Kanji Tsuru (Kyushu University), Yoshiya Hashimoto (Osaka Dental University)

O-08 Analysis of dissolution behaviour of octacalcium phosphate and β-tricalcium phosphate bone substitute materials *Suzuki O1, Sakai S1, Anada T1, Tsuchiya K1, Yamazaki H2, Margolis HC2 (1Tohoku Univ., Sendai, Japan, 2 The Forsyth Institute., Cambridge, MA, USA)

O-09 Fabrication of carbonate apatite foam from the set calcium sulfate hemihydrate foam via solution mediated phase conversion *Sugiura Y, Tsuru K, and Ishikawa K (Kyushu Univ., Fukuoka, Japan)

O-10 Effect of porous alpha-tricalcium phosphate with immobilized basic fibroblast growth factor on bone regeneration in a canine mandibular bone defect model *Kobayashi N1, Hashimoto Y1, Otaka A2, Yamaoka T2 and Morita S1 (1Osaka Dent Univ., Osaka, Japan, 2NCVC, Osaka, Japan)

O-11 Preparation of calcium hydrogen phosphate loaded with protamine and its anti-bacterial property *Abe K1, Honda M1, Iohara K2, Tajima Y2, Kamata A2, Kokaji S2, Yamamoto T3, Hayakawa T3 and Aizawa M1 (1Meiji Univ., Kanagawa, Japan, 2Maruha Nichiro Corporation, Tokyo, Japan, 3Tsurumi Univ., Kanagawa, Japan)

O-12 Characteristics of experimental calcium silicate as a pulp capping material *Hayashi Y1, Kawaki H2, Hori M1, Hasagawa T1, Tanaka M1, Kawano S1, Yoshida T1, and Tamaki Y3 (1~3Asahi Univ., Mizuho, Japan)

Oral Presentation Ⅲ(O-13 - O-18) (Ballroom2): Saturday, November 5, 11:10 - 12:10 (Ceramics) Chair: Motohiro Uo (Tokyo Medical and Dental University), Masahiro Okada (Okayama University)

O-13 withdraw

O-14 Three-dimensional characterization and distribution of fabrication pore defects in bilayered lithium disilicate glass-ceramic molar crowns Zi-Hua He 1, Li Dao 1, Yu-Tao Jian 1, Michael V. Swain2 and *Ke Zhao 1 (1Sun Yat-sen University, Guangzhou, China, 2the University of Sydney, NSW 2006, Australia)

O-15 Evaluation of Newly Developed Investment Material for Press Ceramics *Mori D, Yoshinaga M, Fujimoto T, Mashio G, Yokohara H, Hoshino T, Miyake T, Sato T, Kumagai T (GC corporation, Tokyo, Japan)

O-16 Effects of rapid heating and shape of semi-sintered zirconia crown on marginal fit accuracy *Ohkuma K1, Kameda T1, Ebihara Y2, Mizuguchi Y2and Miyagawa Y1 (1Nippon Dental Univ., Niigata, Japan, 2GC Corp., Tokyo, Japan)

O-17 Influence of different finishing procedures on the wear behavior of zirconia ceramics *Hata U1, Doi Y1, Nakatsuka T2, Mori D1, Yamamura O1 and Fujiwara S1 (1Asahi Univ., Gifu, Japan, 2Shofu Co. Inc., Kyoto, Japan)

O-18 Change in surface of yttria-stabilized zirconia in water and Hanks’ solution characterized using XPS *Oishi M, Tsutsumi Y, Chen P, Doi H and Hanawa T (Tokyo Medical and Dental Univ., Tokyo, Japan)

Oral Presentation Ⅳ(O-19 - O-22) (Ballroom2): Saturday, November 5, 15:30 - 16:10 (Composites & Dental Adhesives 1) Chair: Shinji Takemoto (Tokyo Dental College), Masanori Hashimoto (Osaka University)

O-19 Aspects of Silicon Release from Resin Composites and Glass Ionomer Cement Yon MJY, *Tsoi JKH, Matinlinna JP (The University of Hong Kong, Hong Kong SAR, China)

O-20 Relationship between mechanical properties, interfacial characteristics and bond fatigue strength of short fiber-reinforced resin composite with universal adhesive *Tsujimoto A1,2, Barkmeier WW2, Takamizawa T1, Latta MA2 and Miyazaki M1 (1Nihon Univ., Tokyo, Japan, 2Creighton Univ., Omaha, Nebraska, USA)

O-21 Evaluation of GIC-Surface Treatment on Bond Strength to Resin Composite *Sakamoto M, Arita A, Fukushima S, Kumagai T (Research & Development, GC Corporation, Tokyo, Japan)

O-22 Bond strength of experimental epoxy resin-based sealer cement with submicron-size zirconium oxide filler particles *Pane E1,2, Messer HH1, Palamara J1 (1Melbourne Dental School, University of Melbourne, Australia, 2Faculty of Dentistry, Universitas Sumatera Utara, Indonesia)

Oral Presentation Ⅳ(O-23 - O-26) (Ballroom2): Saturday, November 5, 16:20 - 17:00 pm (Composites & Dental Adhesives 2) Chair: Katsushi Okuyama (Asahi University), Takashi Nezu (Health Sciences University of Hokkaido)

O-23 Bond strength of self-adhesive flowable resin composite to artificial tooth erosion by scratch test and tensile bond strength test *Murase Y1, Kotake H1, Kusakabe S1, Okuyama K2, Tamaki Y2 and Hotta M１ (1,2 Asahi Univ., Mizuho, Japan)

O-24 Dynamic mechanical analysis of dentin-bonding interface prepared by a self-etching primer *Zhou J1,2, Tanaka R1, Shibata Y1, Gao P2, Miyazaki T1 (1Showa Univ., Tokyo, Japan, 2Tianjin Medical Univ., Tianjin, China)

O-25 withdraw

O-26 Regulation of degradation rate and pattern of dental collagen membrane in vivo Jian-min Han1, *Hong Lin11, Xianchang Sun2, Jiahuan Dong2, Yang Wu1, Youdong Zhang1, Meng Yang1 (1Peking University School and Hospital of Stomatology, Beijing, China, 2Yantai Zhenghai Bio-tech Co., Ltd, Shandong, China)

Oral Presentation Ⅴ（O-27 - O-30) (Ballroom2): Sunday, November 6, 9:30 - 10:10 (Clinical Application) Chair: Tomotaro Nihei (Kanagawa Dental College), Masayuki Hattori (Iwate Medical University)

O-27 withdraw

O-28 Effect of Ca(OH)2 treated root canal dentin on retention of posts *Someya T, Harada R, Kinoshita H, Takemoto S, Kawada E (Tokyo Dental College, Tokyo, Japan)

O-29 Fatigue resistance of CAD/CAM crowns restored on premolars *Tsoi JKH1, Homaei E1,2, and Matinlinna JP1 (1The University of Hong Kong, Hong Kong SAR, P.R. China, 2 Ferdowsi University of Mashhad, Mashhad, Iran)

O-30 Development of a cross platform program to analyze cephalometric radiographs by Python *Kawai T, Ohno Y, Egashira M, Iwata J, Miyazawa K, and Goto S (Aichi-Gakuin Univ., Nagoya, Japan)

Oral Presentation Ⅵ（O-31 - O-36) (Ballroom2): Sunday, November 6, 10:30 - 11:30 (Metals) Chair: Toshikazu Akahori (Meijo University), Yusuke Tsutsumi (Tokyo Medical and Dental University)

O-31 withdraw

O-32 Discoloration of titanium in response to sulfide released by P. gingivalis *Harada R, Kinoshita H, Takemoto S, Kawada E (Tokyo Dental College, Tokyo, Japan)

O-33 Application of the solution plasma surface modification technology to the formation of thin hydroxyapatite film on titanium implants *Akashlynn Badruddoza Dithi, Takashi Nezu, Futami Nagano-Takebe, Kazuhiko Endo (Health Sciences University of Hokkaido, Hokkaido, Japan)

O-34 Chitosan/siCkip-1 biofunctionalized titanium implant for improved osseointegration in the osteoporotic condition Li Zhang, Wen Song, *Yumei Zhang (The Fourth Military Medical University, Xi’an, China)

O-35 Design and fabrication orthodontic bracket using Metal Injection Molding (MIM) process *Supriadi S, Irawan B, and Suharno B, Prasetyadi T, Gratis A, Zulkifly F F (Universitas Indonesia, Jakarta, Indonesia)

O-36 Effects of heat treatment on the microstructure and mechanical property of selective laser melted Co-Cr-Mo alloys *Kajima Y1, Takaichi A1, Kittikundecha N1, Nakamoto T2, Doi H1, Takahashi H1, Nomura N3，Hanawa T1 and Wakabayashi N1 (1Tokyo Medical and Dental Univ., Tokyo, Japan, 2Technology Research Institute of Osaka Prefecture, Osaka, Japan, 3Tohoku Univ., Miyagi, Japan) 13

Poster Presentation Ⅰ（P-01 - P-60) (Ballroom3): Saturday, November 5, 10:00 - 11:30

P-01 Fatigue Resistance of GUMMETAL Clasps *Shimamoto K, Tamaki Y, Wakamatsu N, Iwahori M, Nigauri A, Miyao M, Kajimoto T (Asahi Univ. ,Gifu,Japan)

P-02 Correlation between magnetic susceptibility and phase constitution of Au–Nb alloys for MRI artifact-free biomedical applications. *Inui S, Uyama E, Honda E, and Hamada K (Institute of Biomedical Sciences, Tokushima Univ., Tokushima, Japan)

P-03 The effect of ice-quenching on change in hardness during porcelain firing simulation in a Pd-Au-Zn-In-Sn alloy Shin HJ, Kwon YH, Seol HJ, *Kim HI (Pusan National University, Yangsan, Korea)

P-04 Effects of support structure on the fatigue strength of selective laser melted Co-Cr-Mo clasps *Takaichi A1, Kajima Y1, Kittikundecha N1, Nakamoto T2, Nomura N3, Takahashi H1, Hanawa T1, and Wakabayashi N1 (1Tokyo Medical and Dental Univ., Tokyo, Japan, 2Technology Research Institute of Osaka Prefecture., Osaka, Japan, 3Tohoku Univ., Miyagi, Japan)

P-05 Effect of low concentration of sulfide on corrosion of Ti-Cr alloy *Takemoto S, Harada R, Kinoshita H, and Kawada E (Tokyo Dental College, Tokyo, Japan)

P-06 Relationship between Microstructure and Mechanical Strength of Newly Developed Ag-Pd-Cu-Au System Alloy Subjected to Solution Treatment *Akahori T1, Fukui H2, and Niinomi M1 (1Meijo Univ., Nagoya, Japan, 2Aichi-gakuin Univ., Nagoya, Japan)

P-07 In vitro evaluation of H2O2 hydrothermal treatment of aged titanium surface to enhance biofunctional activity *Yoneyama Y1, Matsuno T2, Hashimoto Y3, and Chikazu D1 (1Tokyo Medical Univ., Tokyo, Japan, 2Nippon Dental Univ., Tokyo, Japan, 3Osaka Dental Univ., Osaka, Japan.

P-08 NaOCl-treatment of titanium implant enhances cytocompatibility *Ichioka Y, Kado T, Kanada K, Endo K, and Furuichi Y (Health Sciences University of Hokkaido., Hokkaido, Japan)

P-09 Monitoring of protein adsorption and apatite precipitation on titanium and zirconia by QCM method *Yoshida E and Hayakawa T (Tsurumi Univ., Kanagawa, Japan)

P-10 Application of porous titanium in tailored post-core systems using a moldless process: evaluation of the physical and mechanical properties with different particle sizes *Prananingrum W1, 3, Naito Y2, Sekine K1, Hamada K1, Ichikawa T1 (1,2Tokushima University, Tokushima, Japan, 3Hang Tuah University, Surabaya, Indonesia)

P-11 Molecular orientation and surface properties of gold-deposited titanium following alkanethiol immersion treatment *Saitoh S, Sasaki K, Taira M, and Hattori M (Iwate Medical Univ., Iwate, Japan)

P-12 Construction and biological evaluation of multi-factor-functionalized coating on titanium implant Luo QJ1, Wang Y2, Zha GY2, Chen YD2, *Li XD2 (1The First Affiliated Hospital, Zhejiang Univ., Hangzhou, China, 2The Affiliated Stomatology Hospital, Zhejiang Univ., Hangzhou, China)

P-13 Interactions between the nano-micro-hierarchical titanium and platelet Chen YD1, Luo QJ2, Huang Y3, *Li XD1 (1The Affiliated Stomatology Hospital, Zhejiang Univ., Hangzhou, China, 2 The First Affiliated Hospital, Zhejiang Univ., Hangzhou, China, 3Peking University School and Hospital of Stomatology, Beijing, China)

P-14 Cortical Bone Response of MDF Titanium Implant *Suzuki H1, Hirota M1, Ohkubo C1, Hoshi N2, Kimoto K2, Miuar H3 and Hayakawa T1 (1Tsurumi Univ., Kanagawa, Japan, 2Kanagawa Dental Univ., Kanagawa, Japan, 3Toyohashi University of Technology, Aichi, Japan)

P-15 Nanosecond-Pulsed Laser-Treated Titanium Implants Enhanced Bone and Gingival Tissue attachment * 1 1 2 3 4 2 2 Fukayo Y , Horie A , Amemiya T , Komotori J , Mizutani M , Hamada Y , and Hayakawa T (1Kanto Rosai Hospital, Kanagawa, Japan, 2Tsurumi Univ., Kanagawa, Japan, 3Keio Univ., Kanagawa, Japan, 4Tohoku Univ., Sendai, Japan)

P-16 withdraw

P-17 A Study on Wear Resistance of Flowable Composite Resin Material *Teramae M, Takahashi S, Uchida J, Fujimura H, and Nakatsuka T (Research and Development Dept., Shofu Inc., Kyoto, Japan)

P-18 Discoloration and microbial adhesion to composite resin artificial denture teeth containing fluoropolymers and nano silica filler *Oike K, Fujii K, Kusakabe S, Kotake H, Tamaki Y, Hotta M (Asahi Univ., Mizuho, Japan)

P-19 Fluidity quantification of flowable resin composites through measurement of yield stress using viscometer *Asakura M, Hayashi T, Mieki A, Aimu K and Kawai T (Aichi Gakuin Univ., Nagoya, Japan)

P-20 Wear Resistant of Composites Containing Filler Treated with a Novel Hydrophobic Silane *Nihei T1, Kunzelmann KH2, Ohashi K1, Miyake K1，Yoshino N3 (1Kanagawa Dental Univ., Kanagawa, Japan, 2Dental School of LMU, Munich, Germany, 3Tokyo Univ. of Science, Tokyo, Japan)

P-21 Degradation behaviour of elements from resin composite blocks for CAD/CAM system *Iwata J, Asakura M, Hamajima S, Hasegawa A, Kawai T, Ban S (Aichi Gakuin Univ., Nagoya, Japan)

P-22 Polymerization shrinkage of various core build-up resin composites *Maseki T, Maeno M, Ogawa S and Nara Y (The Nippon Dental University, Tokyo, Japan)

P-23 Effect on the flexural test methods of dental hard resins after accelerated deterioration *Miura D, Miyasaka T, Aoki H, Aoyagi Y, Ishida Y (The Nippon Dental University, Tokyo, Japan)

P-24 Influence of ethanol solution on fatigue strength of resin composite with hybrid cluster filler *Sakaguchi T1, Nishikawa I1, Kato T2, Anraku T2 , Takahashi H3 (1Osaka Institute of Technology Univ.,Osaka ,Japan, 2Yamamoto Precious Metal Co., Ltd.,Osaka ,Japan, 3Tokyo Medical and Dental Univ.,Tokyo ,Japan)

P-25 Study on Surface Properties of Composite Resin and Discoloration *Ishikawa A (The Nippon Dental University, Tokyo, Japan)

P-26 The influence of water aging on surface hardness of low shrinkage light-cured composite resins *Zhu Song (School and Hospital of Stomatology, Jilin University, Changchun , China)

P-27 Influence of the hypochlorous-acid water on properties of light-cured glazing agent membranes *Aoki H, Miyasaka T, Aoyagi Y, Ishida Y, Miura D (The Nippon Dental University, Tokyo, Japan)

P-28 Development of a quantitative method of residual substances in the autopolymerizing acrylic resin *Hongo T, Wada T, and Uo M (Tokyo Medical and Dental Univ., Tokyo, Japan)

P-29 Hydrophilicity of MMA-HEMA resin investigated by fluorescence spectroscopy *Nezu T, Nagano-Takebe F, and Endo K (Health Sciences Univ. Hokkaido, Hokkaido, Japan)

P-30 Effect of bleaching after the discoloration immersion test on color difference and mechanical properties of CAD/CAM resin block *Aoyagi Y, Miyasaka T, Aoki H, Ishida Y, Miura D (The Nippon Dental University, Tokyo, Japan)

P-31 Flexural properties of denture base resin for 3D printing *Park DR, Bae JM (College of Dentistry, Wonkwang Univ., Iksan, Korea)

P-32 Influence of a plasma treatment using a different gas on the bond strength between composite resin and fiber post *Ito Y, Okawa T, Yamamura T, Fukumoto T, Tanaka M (Osaka Dental Univ., Osaka, Japan)

P-33 Influence of plasma treatment to the three-point bending strength of the fiber post *Yamamura T, Okawa T, Ito Y,Yosie S and Tanaka M (Osaka Dental Univ., Osaka, Japan)

P-34 withdraw

P-35 withdraw

P-36 Manufacture of the metal dies for the experiment in the adhesion examination *Igarashi K, Moriue T, Ohki T, Saitou R, Ishida Y, Okada H, and Terada Y (Ohu Univ., Koriyama, Japan)

P-37 Shear bond strength of “PANAVIA V5” between dentin and resin blocks for CAD/CAM Kashiki N and *Okada K (Kuraray Noritake Dental Inc., Tokyo, Japan)

P-38 The effect of application conditions of core resin on the push-out bond strength to root canal dentin *Niitsuma A1, Shinya A1, 2, Shimizu S1, Kuroda S1 and Gomi H1 (1The Nippon Dental Univ., Tokyo, Japan, 2Turku Univ. , Turku, Finland)

P-39 Dentin bond strengths of a new one-bottle universal adhesive containing an amide monomer irradiated by QTH and LED *Hanabusa M, Yamamoto T, Kimura S and Momoi Y (Tsurumi Univ., Yokohama, Japan)

P-40 Bond strengths of a one-bottle universal adhesive containing a new amide monomer to various dental restorative materials *Kimura S, Ito M, Hanabusa M, Yamamoto T and Momoi Y (Tsurumi Univ., Yokohama, Japan)

P-41 Effect of surface treatment after the flowable composite base on the bond strength of the restorative ceramic block *Haruyama A, Kameyama A (Tokyo Dental College, Tokyo, Japan)

P-42 Influence of solvent evaporation on ultimate tensile strength of cured dental adhesives *Kameyama A, and Haruyama A (Tokyo Dental College, Tokyo, Japan)

P-43 Effect of different dental bonding agent application method to bonding thickness analyze with periapical radiography *Rosalina Tjandrawinata, Lyvia Juliana (Trisakti University, Jakarta, Indonesia)

P-44 Comparison of microtensile strength and failure mode under saliva contamination in class V restorations *Shimazu K1, 2, Karibe H1, Yamada H1, 3, Ogata K1, 3 (1Nippon Dental University School of Life Dentistry at Tokyo, Japan, 2Momose Dental Clinic, Tokyo, Japan, 3Tokyo Metropolitan Children’s Medical Center, Japan)

P-45 Evaluation method of the adaptability of machined dental restorations *Nagase Y1, Asakura M1, Ohkuma K2 and Kawai T1 (1Aichi Gakuin Univ., Nagoya, Japan, 2Nippon Dental Univ., Niigata, Japan)

P-46 Educational evaluation of a drilling simulator for dental implant surgery *Kinoshita H, Takemoto S, and Kawada E (Tokyo Dental College, Tokyo, Japan)

P-47 Dimensional accuracy of mandibular models created by personal 3D printers and CT data *Ishida Y1, 2, Miyasaka T2, Aoki H2, Aoyagi Y2, Miura D2, Kawai T3 and Asaumi R3 (1, 2, 3 Nippon Dent. Univ., Tokyo, Japan)

P-48 The accuracy of intraoral and extraoral scanning *Shimizu S1, Shinya A1, 2, Niitsuma A1, Kuroda S1, Gomi H1 (1Nippon Dental Univ, Tokyo, Japan, 2Truk Univ., Truk, Finland)

P-49 Evaluation of adhesion of bonding agents for CAD/CAM processing *Niizuma Y, Kobayashi M, Miyazaki T, and Manabe A (Showa Univ., Tokyo, Japan)

P-50 Fracture strengths of composite resin crowns made with dental CAD/CAM system *Okada R, Ando A, Asakura M, Ozawa S, Takebe J, Kawai T, Ban S (Aichi Gakuin Univ., Nagoya, Japan)

P-51 Strain Analysis of Mandibular Implant Overdentures in Different Dislodging Position with Different Implant Number *Kaidiliya Y, Kanazawa M, Takeshita S, Oda K, Tanoue M, Miyayasu A, ThuyVL, Thu KM, Asami M, Shimizu H, Shimada R, Minakuchi S (Tokyo Medical and Dental University, Tokyo, Japan)

P-52 Clasp fabrication using one-process molding by repeated laser sintering and high-speed milling *Nakata T, Kurihara D, Ohkubo C (Tsurumi University, Yokohama, Japan)

P-53 Supramolecular Sulfonated Polyrotaxanes complexed with Bone Morphogenetic Protein-2 for Bone Tissue Engineering *Terauchi M, Tamura A, and Yui N (Tokyo Medical and Dental Univ., Tokyo, Japan)

P-54 withdraw

P-55 Release of FGF-2 from 4-META/MMA-based adhesive resins containing non-biodegradable FGF-2-loaded-polymer particles *Tsuboi R, Kitagawa H, Takeda K, Sasaki JI, Takeshige F, Imazato S (Osaka Univ., Osaka, Japan)

P-56 Comparison of antibacterial activities of Xanthorrhizol and Bakuchiol with and without fluoride varnish *Son JL1, Kim AJ2, Oh SH1, Bae JM1 (1College of Dentistry, Wonkwang Univ., Iksan, Korea, 2College of Sorabol, Gyengju, Korea)

P-57 Effect of Mesenchymal Stem Cells on Mixed Self Assembled Monolayer *Hirata I1, Veronica SR1, Tania S1,2, Yamauchi Y1, Kanawa M1, Kato Y1,3, Kato K1 (1Hiroshima Univ., Hiroshima, Japan, 2Airlangga Univ., Surabaya, Indonesia, 3TWO CELLS Co.,Ltd., Hiroshima, Japan)

P-58 Evaluation of degradability of EGCG-conjugated gelatin sponge *Honda Y1, Tanaka T2, Hashimoto Y1, Imai K1, and Arita K1 (1Osaka Dental Univ., Osaka, Japan, 2Kyoto Institute of Technology., Kyoto, Japan)

P-59 Redox-Responsive Supramolecular Polymeric Prodrug for Cancer Chemotherapy Wang Y1, Sun R2, Luo QJ3, *Li XD1 (1-3 Zhejiang Univ., Hangzhou, China)

P-60 The expression of NF-kB on rat tooth mechanically exposured after administration of pomegranate extract *Intan Nirwana, Titien Hary Agustantina and Helal Soekartono (Universitas Airlangga, Surabaya, Indonesia)

Poster Presentation Ⅱ(P-61 - P-116) (Ballroom3): Saturday, November 5, 15:30 - 17:00

P-61 Reasons for failures of posterior restorations *Kubo S, Sumi T, and Hayashi Y (Nagasaki Univ., Nagasaki, Japan)

P-62 Improvement in oral environment of the denture wearer by neutral electrolyzed water *Shimizu H1, Nagamatsu Y1, Nagamatsu H1, Murakami S2, Nakamura K1, Ikeda H1, Komagata Y1, Masumi S1, Kozono Y1 (1Kyushu Dental Univ., Kitakyushu, Japan, 2Kyushu Univ. of Nursing and Social Welfare, Tamana, Japan)

P-63 Bone response to multilayered DNA/protamine coated titanium implants *Sakurai T1, Yoshinari M,2 Toyama T,3 Hayakawa, T1, and Ohkubo C1 (1Tsurumi Univ., Kanagawa, Japan, 2Tokyo Dental College, Tokyo, Japan, 3Nihon University, Tokyo, Japan)

P-64 Bond strength of restoration handling instruments with adhesive and restoration materials *Ishida Y1, Igarashi K2, Terada Y2, Okada H1 (1,2Ohu Univ., Fukushima, Japan)

P-65 Wear Properties of CAD/CAM Hybrid Resin Block *Kai T, Teramae M, and Nakatsuka T (Research and Development Dept., Shofu Inc., Kyoto, Japan)

P-66 Highly-viscous glass-ionomer cement for filling: Interfacial Gap-formation in Class I restoration and mechanical properties. *Irie M, Matsumoto T, Taketa H, Torii Y, Maruo Y, Nishigawa G, Minagi S, Nagaoka N, Yoshihara K (Okayama Univ., Okayama, Japan)

P-67 Influence of constant strain on elastic modulus of thermoplastic orthodontic materials *Inoue S, Yamaguchi S, Uyama H, Yamashiro T, Imazato S (Osaka Univ., Suita, Japan)

P-68 Wear of dental restorative materials under low occlusal force *Kakuta K, and Miyagawa Y (Nippon Dental Univ., Niigata, Japan)

P-69 Influence of Shades of Resin Cement on Shades of Laminate Veneer Restorations Evaluated Using Shade Models *Hara D, Kitada N, Nagafuji A, Shinno K, and Nakatsuka T (Research and Development Dept., Shofu Inc., Kyoto, Japan)

P-70 Effect of the use of commercial cleaning liquid in thermoforming process of mouthguard materials *Tanabe G1, Churei H1, Wada T2, Uo M2, Takahashi H3, and Ueno T1 (1-3Tokyo Medical and Dental Univ., Tokyo, Japan)

P-71 Stress-strain behavior of bleached enamel stored in a high ionic strength solution *Tanaka R, Zhou J, Shibata Y, and Miyazaki T (Showa Univ., Tokyo Japan)

P-72 Evaluation of the acid resistance of root dentin when applying fluoride containing materials incorporating Ca using in-air micro- PIXE/PIGE. *Yagi K1, Yamamoto H1, Uemura R1, Okuyama K 2, Matsuda Y 3, Suzuki K4, and Hayashi M1 (1Osaka Univ., Osaka, Japan, 2Asahi Univ., Gifu, Japan, 3Health Sciences Univ. of Hokkaido, Hokkaido, Japan, 4The Wakasa Wan Energy Research Center, Japan)

P-73 Influences of various dentin desensitizers on the effect of tooth whitening *Yoshikawa K , Yasuo K, Furusawa K, Iwasa K , Finger WJ, and Yamamoto K (Osaka Dental University, Osaka, Japan)

P-74 The Effects of Food and Drink on the Color Stability of Various Thermoplastic Resins *Kuroda S1, Yokoyama D1, Shinya A1,2, Gomi H1 (1The Nippon Dental Univ., Tokyo, Japan, 2Truk Univ., Truk, Finland)

P-75 Experimental immediate evaluation system for endodontic lesion using light-induced red fluorescence *Takino H, Isatsu K, and Hasegawa T (Showa Univ., Tokyo, Japan)

P-76 withdraw

P-77 withdraw

P-78 Fabrication of a faceguard by incorporating a glass-fiber reinforced thermoplastic and shock escape space and this shock-absorption ability *Wada T, Churei H, Fukasawa S, Shirako T, Hongo T, Ueno T, and Uo M (Tokyo Medical and Dental Univ., Tokyo, Japan)

P-79 withdraw

P-80 The influence of naval nano calcium and nano chitosan contents toothpaste on enamel surface microhardness *Octarina, Rahma Meilita, Komariah (Trisakti University, Jakarta, Indonesia)

P-81 Bond Characterization of Various Framework Materials and Ceramic *Kawata K, Teramae M, and Nakatsuka T (Research and Development Dept., Shofu Inc., Kyoto, Japan)

P-82 Development of thin apatite coated zirconia implant *Hirota M1, Ohkubo C1, Sato M2, Toyama T3, Tanaka Y4, and Hayakawa T1 (1Tsurumi Univ., Kanagawa, Japan, 2Kogakuin Univ., Tokyo, Japan, 3Nihon Univ., Tokyo, Japan, 4Kagawa Univ., Kagawa, Japan)

P-83 Influence of bilayer ceria-stabilized zirconia/alumina nanocomposite-based crowns with supporting structures on fracture properties *Sawada T, Spintzyk S, Schille C, Schweizer E, and Geis-Gerstorfer J (University Hospital Tübingen, Tübingen, Germany) P-84 The effect of surface modification on the bioactivity of zirconia materials *Hong G1, Liu J1,2, Wu YH1, Endo K1, Han JM3, Wada T1, Kato H1, and Sasaki K1 (1Tohoku Univ., Sendai, Japan, 2Tianjin Medical Univ., Tianjin, China, 3Peking Univ., Beijing, China)

P-85 Three-dimensional finite element analysis of the effect of load patterns on stress distribution in a zirconia all-ceramic crown *Miura S, Kasahara S, Yamauchi S, Egusa H (Tohoku Univ., Miyagi, Japan)

P-86 The effect of the cone angle and space between inner and outer crowns of a cone crown telescope with nano zirconia on retentive force and the fit of the copings *Nakagawa S, Torii K, Yoshikawa Y, Okawa T, Ito Y, Fukumoto T, Tanaka J and Tanaka M (Osaka Dental Univ., Osaka, Japan)

P-87 Charpy impact test using single-edge notched zirconia specimens *Tsuruta S, Mizuno M, Uematsu Y, Kawai T, Kurita S and Yamamoto I (Aichi Gakuin Univ., Nagoya, Japan)

P-88 Development of tetragonal zirconia nanocrystals with high aspect ratio for composite resin fillers *Okada M, Taketa T, and Matsumoto T (Okayama Univ., Okayama, Japan)

P-89 Effects of zirconia surface synthesized polycrystalline particulates on the bonding strength of porcelain *Zhang ZT, Tian YM, Zhuge RS, Zhang LL, Ding N (School of Stomatology, Capital Medical Univ., Beijing, China)

P-90 Effects of zirconia particle addition on mechanical properties of calcium phosphate cement *Bae JY, Ida Y, Sekine K, Kawano F, and Hamada K (Tokushima University, Tokushima, Japan)

P-91 Change in the retentive force of Akers clasp for zirconia crown *Tanaka A, Miyake N, Hotta H, Takemoto S, Yoshinari M, Yamashita S (Tokyo Dental Collage, Tokyo, Japan)

P-92 Effect of surface treatment on the bond strength between zirconia ceramic and core resin *Uno M1, Kawaki H2, Nonogaki R1, Sawada T1, Nishikawa M3 , Kurachi M4, Ishigami H1, and Doi Y5 (1-5Asahi University, Gifu, Japan)

P-93 Zirconia surface coating with silk fibroin electrogels *Qu YY1, 2, Hong G1, Liu L2, and Sasaki K1 (1Tohoku Univ., Sendai, Japan, 2Dalian Stomatological Hosp., Dalian, China)

P-94 XRD and FT-IR analysis on fluoride-treated hydroxyapatite *Okuyama K1, Komada Y1, Abe M1, Matsuda Y2, Yamamoto H3 and Tamaki Y1 (1Asahi Univ., Gifu, Japan, 2Health Science Univ. of Hokkaido, Hokkaido, Japan, 3Osaka Univ., Osaka, Japan)

P-95 Hydroxyapatite nanoparticle-assembled powder as a pulp-capping agent in rats Imura K1, *Hashimoto Y1, Okada M2, Yoshikawa K1, Imai K1, Matsumoto T2 and Yamamoto K1 (1Osaka Dental Univ., Osaka, Japan, 2Okayama Univ., Okayama, Japan)

P-96 Bone response to apatite paste derived from Ca-amino acid complex *Waki T1, Mochizuki C2, Sato M2, Hayakawa T1, and Ohkubo C1 (1Tsurumi Univ., Kanagawa, Japan, 2 Kogakuin Univ., Tokyo, Japan)

P-97 Bone regeneration with a collagen model polypeptide/porous alpha-tricalcium phosphate sponge in a canine mandibular bone defect model Matsuse K, *Hashimoto Y, Imai K, Baba S and Morita S (Osaka Dent Univ, Osaka, Japan)

P-98 Effect of water structure on bone mineral formation I:Kosmotropes *Kobayashi D, Kai S,Okada M, and Matsumoto T (Okayama Univ., Okayama, Japan)

P-99 Effect of water structure on bone mineral formation II:Chaotropes *Kai S, Kobayashi D, Okada M, and Matsumoto T (Okayama Univ., Okayama, Japan)

P-100 Fabrication of biphasic bone substitute consisting of calcite and carbonate apatite *Tsuru K, Munar M, and Ishikawa K (Kyushu Univ., Fukuoka, Japan)

P-101 Clinical application of octacalcium phosphate collagen composite in sinus floor elevation *Tadashi Kawai1, Keiko Matsui1, Fumihiko Kajii1,2, Yoshinaka Shimizu1, Hiroyuki Kumamoto1, Osamu Suzuki1, Shinji Kamakura1, Tetsu Takahashi1 (1Tohoku Univ., Sendai, Japan, 2TOYOBO CO., LTD, Otsu, Japan)

P-102 The application of oxygen-permeable spheroid culture chip (Oxy chip) in bone tissue engineering *Anada T, Kamoya T, Sato T, Shiwaku Y, Takano-Yamamoto T, Sasaki K, and Suzuki O (Tohoku Univ., Sendai, Japan)

P-103 Influence of surface physicochemical modification on wettability of zirconia (TZP) and behaviour of oral keratinocytes *Yoshinari M, Takemoto S (Tokyo Dental College, Tokyo, Japan)

P-104 withdraw

P-105 Induced pluripotent stem cells: A potential cell source for regenerative dentistry *Abdullah AN1, 2, Tanimoto K2, and Kato K1 (1,2Hiroshima University, Hiroshima, Japan)

P-106 Designed SDF-1 chimeric protein for bone tissue regeneration *Date T, Hirata I, Hattori S, Matsuda C, Nakano A, Yamakado N, Tanimoto K, and Kato K (Hiroshima University, Hiroshima, Japan)

P-107 The surface treatment to realize both antibacterial property and bioactivity *Tsutsumi Y, Shimabukuro M, Ashida M, Chen P, Doi H, and Hanawa T (Tokyo Medical and Dental Univ., Tokyo, Japan) P-108 Cytotoxicity of methacrylate-based resin monomers as evaluate by an anti-oxidant responsive element-luciferase reporter assay *Egashira M, Suzuki T, Fujimoto K, Kanamori T, and Kawai T (Aichi-Gakuin Univ., Nagoya, Japan)

P-109 Preparation of antibacterial high-strength dental stone using with titanium dioxide nanopowder as photocatalyst *Muraoka K, Yoshida K, Miyahara H, Ikeda H, Nagamatsu Y, Awano S, Shimizu H (Kyushu Dental Univ., Kitakyushu, Japan)

P-110 Optimization of culture conditions for the efficient differentiation of iPS cells into dental epithelial cells *Miyauchi S1, 2, Abdullah AN1, Nikawa H2, and Kato K1 (1,2Hiroshima University, Hiroshima, Japan)

P-111 Influence of Organic Materials on HAp Crystal Formation in Calcium Phosphate-based Cements *Tai Y, Sekimizu T, Asada Y, Chiba T, Momoi Y, Hosoya N, Shimoda S (Tsurumi Univ., Kanagawa, Japan)

P-112 withdraw

P-113 Cell aggregation of periodontal ligament fibroblasts controlled by micro-patterns *Akasaka T, Kaga N, Yokoyama A, Abe S, Yoshida Y (Hokkaido Univ., Sapporo, Japan)

P-114 Assessment of cytocompatibility of nano-sized ceramics particles on several cells Seitoku E, *Abe S. Era Y, Kiyama N, Tsuchiya S, Nakanishi K, Nakagawa Y, Akasaka T, Iida J, Yoshida Y, Sano H (Hokkaido University, Sapporo, Japan)

P-115 Proliferation of mesenchymal stem cells seeded on biodegradable hydroxyapatite–chitosan porous scaffolds *Maretaningtias Dwi Ariani1, Sherman Salim1, Isao Hirata2, Koichi Kato2 (1Airlangga University, Surabaya, Indonesia, 2Hiroshima University, Hiroshima, Japan)

P-116 Mechanical properties of coating layer modification chitosan-TiAlCr on calcium phosphate cement with sol-gel method *Atia Nurul Sidiqa1, Bambang Sunendar2, Nina Djustiana 2, Renny Febrida2 (1Jenderal Achmad Yani University, Indonesia, 2Advanced Material Department,Institute Technology of Bandung, Indonesia, 3Dental Materials Department, Dentistry Faculty, Padjadjaran University, Indonesia)

Invited Lecture 1

Surface treatment and modification of metals to add biofunctions

Hanawa T* Tokyo Medical and Dental Univ., Tokyo, Japan * [email protected]

Introduction: A disadvantage of using metals as biomaterials is that they are typically artificial materials and have no biofunction. To add biofunction to metals, surface modification is necessary. In dentistry, dental implants require hard tissue compatibility for osseointegration and bone formation, soft tissue compatibility for adhesion of gingival epithelium, and antibacterial property for the inhibition of biofilm formation. These biofunctional properties consist of conflict two properties: the inhibition and enhancement of protein adsorption or cell adhesion. In this paper, especially electrochemical techniques to add biocompatibility and biofunction to metals are reviewed.

Surface treatment and modification techniques: A tremendous number of surface treatment and modification techniques to improve the hard tissue compatibility of titanium have been developed. Among these techniques, I summarized techniques investigated in our laboratory in Fig. 1. Recently, we are concentrating the development electrochemical techniques such as cathodic polarization, micro arc oxidation (MAO), and electrodeposition of biofunctional molecules. In particular, both bone formation and antibacterial properties are acquired simultaneously by a modified MAO technique.

MAO Zr cathodic polarization Zr white oxide layer J. Y. Ha et al: Surf Coat Technol 205 (2011) 4948 Y. Tsutsumi et al: Act Biomater 6 (2010) M. Yu et al: Dent Mater J 33 (2014) 490 C. Ma et al: Act Biomater 8 (2012) 860 4161 Y. Tsutsumi et al: Appl Surf Sci 262 (2012) 34

Hydrothermal treatment in Ca-containg solution Zr-coated Ti K. Hamada et al.: Biomaterials 23 (2002) 2265 E. Kobayashi et al.: Mater Trans 48 (2007) 301

Immersion in Ca-containing solution Ca-P formation on Ti T. Hanawa et al.: J Biomed Mater Res 34 (1997) 273 T. Hanawa: J Jpn Dent Mater Dev 8 (1989) 832 T. Hnanawa et al.: Biomaterials 12 (1991) 767

Repassivation in Hanks’ solution T. Hanawa al.: Mater Trans 43 (2002) 3000 Ca-ion-implanted Ti Hanawa et al.: J Jpn Soc Biomater 12 (1994) 209 Amorphous CaTiO3 layer Hanawa et al.: J Biomed Mater Res 36 (1997) 131 Low-voltage anodic polarization Crystalline CaTiO3 layer in Hanks’ solution Y. Tanaka et al.: J Collagen coating＋He-ion N. Ohtsu et al: Mater Trans 45 (2004) 1778 Mater Sci Mater Med 18 (2007) 797 N. Ohtsu et al: J Biomed Mater Res 82A (2007) 304 implantationS. Nakajima et al.: N-ion-implante Co-Cr alloy Trans Mater Res Soc Jpn 28 (2003) 499 T. Hanawa et al.: Mater Trans 46 (2005) 1593 PEG electrodeposition S. Hiromoto et al: Mater Trans 46 (2005) 1627 Y. Tanaka et al.: Mater Sci Eng C27 (2007) 206 SPU- Y. Tanaka et al.: Mater Trans 48 (2007) 287 electrodeposition Y. Tanaka et al: Mater Trans 49 (2008) 805 SPU-Ti composite on Ti Ｘ Y. Tanaka et al: J Biomed Mater Res 92A (2010) 350 H. Sakamoto et al: J Biomed Mater Res 82A (2007) 52 Y. Tanaka et al: J Biomed Mater Res 95A (2010) 1105 H. Sakamoto et al: Dent Mater J 27 (2008) 81 A. Kawabe et al: Dent Mater J 33 (2014) 638 H. Sakamoto et al: Dent Mater J 27 (2008) 124 Collagen electrodeposition J. H. Seo et al: Soft Mater 11 (2015) 936 H. Kamata et al: Mater Trans 52 (2011) 81 SPU-Ti alloy composite RGD/PEG/Ti J. Hieda et al: Surf Coat Technol 206 MPC electrodeposition Y. Tanaka et al: J Colloid Interface Sci 330 (2009) 138 (2012) 3137 Y. Fukuhara et al: J Biomed Mater Res Appl Biomater K. Oya et al: Biomaterials 30 (2009) 1238 Y. Fukuhara et a: Appl Surf Sci J. W. Park et al: Act Biomater 7 (2011) 3222 Fig. 1 Surface treatment and modification techniques investigated in our laboratory.

Acknowledgements: I would like to thank Dr. Tsutsumi to perform the researches introduced in this paper.

Invited Lecture 2

Surface modifications for enhanced integration of titanium implant with soft and hard tissues and prevention of peri-implantitis Endo, K* Health Sciences University of Hokkaido (Ishikari-tobetsu, Japan) e-mail ([email protected]) Introduction: The use of dental implants has been recognized as an effective treatment of partially and fully edentulous patients due to their high level of predictability and a wide variety of treatment options. While in many cases, dental implants have been reported to achieve long-term success, one of the most frequent complications in dental implants is peri-implantitis. In order to enhance integrations of the titanium implant with the surrounding soft and hard tissues, and also to prevent peri-implantitis, we investigated the surface modifications of titanium using physical coating technologies and biochemical techniques.

Materials and Methods: The surface structures of titanium after surface modifications were characterized by XRD, XPS, FT-IR, SEM/EDX and SPM. Physical, chemical, and biological properties of the surface were studied by nano-indentation tests, abrasion tests, electrochemical corrosion tests, various cell culture experiments, bacterial adhesion tests, and implant tests using animals.

Results and Discussion: We found that the chemical modification of the titanium surface with collagen significantly enhanced initial attachment, spreading, and differentiation of HPDLCs and HBMMSCs, suggesting the achievement of rapid soft and hard tissues-titanium integration [1]. For zirconia abutment, the insulin-like growth factor 1(IGF-1) immobilized on the yttria-stabilized tetragonal zirconia polycrystal surfaces enhanced the extension and adhesion of human gingival epithelial cells without enhancing S. gordornii adhesion on the surfaces [2]. We also demonstrated that the adsorption of human lactoferrin, an antibacterial protein in saliva, on the titanium surface inhibited bacterial adhesion and exhibited bactericidal effects [3]. The initial bacterial attachment and subsequent plaque formation can also be lowered by the hard and chemically stable TiN coating since smooth surfaces were maintained even in the presence of abrasion and chemical attack. The combination of the TiN film coating and lactoferrin application to titanium surface could be effective in inhibiting biofilm formation and preventing peri-implantitis around the titanium abutment.

Conclusions: These findings suggested that surface modifications we employed can be effective in enhancing integrations of the titanium and zirconia implants with the surrounding soft and hard tissues, and also to prevent peri-implantitis.

References: [1] T. Kado, T. Hidaka, H. Aita, K. Endo, Y. Furuichi: Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells. Applied Surface Science, 262: 240-247, 2012. [2] D. Ito, T. Kado, F. Takebe, T. Hidaka, K. Endo, Y. Furuichi: Biological activation of zirconia surface by chemical modification method with IGF-1. J Biomed Mater Res, Part A 103, 3659-3665, 2015. [3] F. Nagano-Takebe, H. Miyakawa, F. Nakazawa, K. Endo: Inhibition of initial bacterial adhesion on titanium surfaces by lactoferrin coating. Biointerphases 9, 1-7, 2014.

Invited Lecture 3

Carbonate apatite bone replacement

Ishikawa K* Kyushu Univ., Fukuoka, Japan [email protected]

Bone apatite is not hydroxyapatite (HAp: Ca10(PO4)6(OH)2) but carbonate apatite (CO3Ap: Ca10-a(PO4)6-b(CO3)c(OH)2-d) that contains 6-9% of carbonate. However, CO3Ap is decomposed thermally at high temperature since carbonate is liberated at high temperature. Thus, sintered HAp has been used as an artificial bone substitute. We have proposed a fabrication method of CO3Ap block by compositional transformation reaction based on dissolution-precipitation method using a precursor block such as calcium carbonate and tricalcium phosphate. For example, CaCO3 block is made as a precursor by exposing Ca(OH)2 compact to carbon dioxide. Then, CaCO3 block is immersed in sodium phosphate solution. Sodium phosphate solution is undersaturated with respect to CaCO3 block, and thus 2+ CaCO3 is dissolved and supplied Ca and 2- CO3 to the sodium phosphate solution. Then, the solution became supersaturated 2+ 2- with respect to CO3Ap, thus, Ca and CO3 are precipitated as CO3Ap with the phosphate ions in the sodium phosphate solution. The CO3Ap thus fabricated contains 4- 12wt% carbonate in its apatitic structure and the content of the carbonate can be regulated by the fabrication condition such as solution Fig. 1 SEM images of osteoclasts when temperature and its pH. incubated on the surface of a) HAp and b) Figure 1 shows the typical SEM pictures CO3Ap. when osteoclasts were incubated on the surface of a) HAp and b) CO3Ap. No resorption pits were formed in the case of HAp disk. In contrast, resorption pits were formed in the case of CO3Ap. When proliferation of the osteoblastic cells were evaluated, all makers of differentiation such as type I collagen, ALP, osteopontin, osteocalcin were up-regulated Fig. 2 The CT images of rabbit femur bone on the surface of CO Ap when compared to 3 defect 18 months after reconstructed with HAp, indicating possible higher HAp and CO3Ap granules. osteoconductivity of CO3Ap at cell level. Histological results were consistent to the cell evaluation. As a result of osteoclastic resorption of CO3Ap, CO3Ap is replaced to bone whereas HAp remained at the bone defect as shown in Fig. 2. As a result of up-regulated proliferation of osteoblastic cells, CO3Ap showed higher osteoconductivity than HAp as shown in Fig. Fig. 3 Histological picture of beagle 3 where beagle mandibular bone defect was mandibular bone defect 3 months after reconstructed with HAp and CO3Ap granules. reconstructed with HAp and CO3Ap granules.

28 Invited Lecture 4

Carbonate apatite ceramics for drug delivery and synthetic scaffold in tissue engineering: a review of the development and application in dentistry

ID Ana* Department of Dental Biomedical Sciences, Faculty of Dentistry, Universitas Gadjah Mada, Yogyakarta 55282, Indonesia *[email protected]

Introduction: When there is a defect in the body, both cells and ECM are lost. Tissue regeneration can be achieved by single or combined use of two or three elements in an appropriate way. However, tissue regeneration cannot happen only by simple combination of the three elements. To achieve tissue regeneration successfully, proper biomedical mechanism is needed [1]. The basic technology is by preparing scaffold to act as an artificial ECM (extracellular matrix). So far, calcium phosphate ceramics are very popular materials for diverse clinical application in dentistry. The version of calcium phosphate called calcium hydroxyapatite constitutes the inorganic and mineral phases of normal calcified tissues, but are also found in some pathologic calcification such as dental calculus, heart calcification, urinary stones, soft tissues calcification, etc. [2]. Previous researches introduced ceramics incorporation to improve mechanical properties of the scaffold [3,4] even calcium from ceramics was also noted to influence axonal growth [5]. This article aims to review the development of carbonate apatite ceramics for drug delivery vehicle and scaffold in tissue engineering, from research to its clinical application, especially the current status in Indonesia.

Materials and Methods: The review is based on the published articles and on-going research works done in our laboratory.

Results and Discussion: Carbonate apatite based bone substitute has been developed in our laboratory and it is commercially available for the clinical used in dentistry [5]. Another work have been being done to investigate peripheral nerve scaffold designed by the incorporation of carbonated apatite (CHA) on gelatin hydrogel system. Glucose permeability, hydrophilicity, degradation and weight loss rates as well as calcium released from the scaffolds increased by the increment of CHA concentration. It is also known that CHA incorporated into the system influence PC12 growth. In another study to investigate metronidazole loading capacity of periodontal membrane, it was found that the incorporation of CHA into the system in a proper concentration did improve loading capacity of metronidazole.

Conclusions: It was concluded from several studies done in our laboratory that CHA is potential candidate to modulate better properties of drug delivery vehicle and scaffold to be used in dentistry.

References: [1] Langer R, Vacanti JP. 1993. Science 260: 920; [2] Tabata Y, Nagano A, Ikada Y. 1999. Tissue Eng 5(2): 127; [3] Le Geros. 1991. Calcium Phosphate in Oral Biology and Medicine. Monograph in Oral Sciences Volume 15. Myers H (Ed.). Karger: Basel; [4] Le Geros RZ. 2002. Clin Orthop Rel Res 395: 81; [5] Zheng JQ. 1999. Nature 403: 89

Invited Lecture 5

Prosthetic rehabilitations using pure titanium to avoid metal combinations

Ohkubo C* Department of Removable Prosthodontics Tsurumi University School of Dental Medicine, Yokohama, Japan *[email protected]

Introduction: Although all restorations ideally should be made without metal, frameworks for removable partial dentures (RPDs) and some implant superstructures will be made, reluctantly, with metal even in the future. To avoid the presence of various ―metal combinations‖ in the patients’ oral cavities, the use of a single metal for all restorations would be necessary because it protects against metal corrosion caused by the contact of different metals. For this ―one-metal rehabilitation‖ concept, nonalloyed commercially pure (CP) titanium should be used for all restorations on the assumption that implant treatment will be carried out if a tooth is lost. This presentation will describe prosthetic rehabilitations using only CP titanium based on the ―one-metal rehabilitation‖ concept.

Materials and Methods: The reactivity of titanium causes the development of a brittle reaction layer with increasing hardness toward the surface during the casting process. Thus, titanium implant abutments, crowns, and bridge prostheses have been recently fabricated using computer-aided design/computer-aided manufacturing (CAD/CAM). However, the frameworks of removable partial dentures (RPDs) are very complicated and consist of many component parts; the milling process may not be capable of machining such complex structures. Currently, we have been challenging the fabrication of RPD frameworks using one-process molding by repeated laser sintering and high-speed milling. Additionally, laser welding has been used typically for titanium framework repair and reform. Even broken titanium clasps can be rejoined using laser welding, and the retentive force does not decrease after clasp repair.

Results and Discussion: Even now, careful selection of processing methods and laboratory skill are necessary to ensure casting success. In a clinical setting, CP titanium crown copings and frameworks for removable dentures could be cast with acceptable accuracy. However, discoloration and severe wear of titanium occlusal surfaces were frequently observed in vivo. Plaque tended to adhere to titanium frameworks more easily as compared to those made of conventional alloys. CAD/CAM fabrication of titanium RPD frameworks is unlikely to be realized soon, whereas most of the components of implant superstructures and fixed prosthetic appliances have been used with great success. Repeated laser sintering and high-speed milling can be expected for the fabrication of titanium RPD frameworks.

Conclusions: Although laboratory and clinical problems still remain, the one-metal rehabilitation concept, using CP titanium as a bioinert metal, would be recommended for all restorations.

References: Ohkubo C, Hanatani S, Hosoi T. Present status of titanium removable dentures—a review of the literature. J Oral Rehabil. 2008;35(9):706–714.

Invited Lecture 6

Non-biodegradable polymer particles for drug delivery: A new technology for “bio-active” restorative materials

Imazato S* Osaka University Graduate School of Dentistry, Suita, Japan [email protected]

Dental restorative materials on the market nowadays show excellent clinical performance with improved mechanical, physical, aesthetic, and bonding properties. Hence, a target of their advancement is being shifted to exhibition of “bio-active” functions to prevent primary/secondary diseases or promote healing of surrounding tissue1. One of the effective approaches to achieve “bio-active” restorative materials is addition of the capability to release agents. However, for reconstructive materials, such function needs to be attained without compromising their basic properties mentioned above. We developed novel non-biodegradable small particles for drug delivery which are applied to polymer-based materials. The particles are made of a hydrophilic monomer 2-hydroxyethyl methacrylate (HEMA) and a cross-linking monomer trimethylolpropane trimethacrylate (TMPT), and can be a reservoir of active agents to be released in a wet environment. We found that the polymer particles consisting of 50% HEMA/50% TMPT were useful for loading and release of an antimicrobial, cetylpyridinium chloride (CPC)2. Especially, loading of CPC by the pre-mixing method enabled long-term release of CPC from the polymer particles. Therefore, the experimental resin-based root canal sealer containing CPC- loaded poly-HEMA/TMPT particles demonstrated antibacterial effects even after being aged. In addition, recharge of CPC to the poly-HEMA/TMPT particles is possible by exposure to CPC solution, and sustained release of CPC can be obtained. Thus, incorporation of CPC- loaded-particles is useful to provide various resin-based restorative materials with long-lasting antibacterial effects. Another design for usage of poly-HEMA/TMPT particles is a carrier of growth factors3. The 90% HEMA/10% TMPT particles loaded with fibroblast growth factor-2 (FGF-2) promoted proliferation of osteoblast-like cells in vitro. Trials to combine FGF-2-loaded- particles with 4-META/MMA-based resins are in progress to develop novel adhesives with the ability to enhance regeneration of periodontal tissue. In this presentation, the new technology to achieve “bio-active” restorative materials with therapeutic effects will be addressed.

Acknowledgements: This study was supported by a Grant-in-aid for Scientific Research (26293409, 26893140, and 15K20402) from the Japan Society for the Promotion of Science.

References: 1. Imazato S, Ma S, Chen J-H, Xu HH. Therapeutic polymers for dental adhesives: loading resins with bio-active components. Dent Mater 30: 97-104, 2014. 2. Kitagawa H, Takeda K, Kitagawa R, Izutani N, Miki S, Hirose N, Hayashi M, Imazato S. Development of sustained antimicrobial-release systems using poly(2-hydroxyethyl methacrylate)/trimethylolpropane trimethacrylate hydrogels. Acta Biomater 10: 4285-4295, 2014. 3. Takeda K, Kitagawa H, Tsuboi R, Kiba W, Sasaki JI, Hayashi M, Imazato S. Effectiveness of non-biodegradable poly(2-hydroxyethyl methacrylate)-based hydrogel particles as a fibroblast growth factor-2 releasing carrier. Dent Mater 31: 1406-1414, 2015.

Invited Lecture 7

REACTION SUPPORTING DENTAL TISSUE ON ENDOSSEOUS DENTAL IMPLANTS USING TITANIUM MATERIALS, BASED ON IMMUNOEXPRESSION OF LAMININ * N. Djustiana, ** S. Widyaputra

*Lecturer and Researcher in Biomaterial at Department of Dental Material Faculty of Dentistry UniversitasPadjadjaran, Bandung West Java Indonesia. **Lecturer and Researcher in Dental Pathologyat Department of Oral Biology Faculty of Dentistry UniversitasPadjadjaran, Bandung West Java Indonesia.

Introduction :The surface reaction of dental implant is a complex biological phenomenon, which includes development of transmucosal seal as physiological barrier to prevent access of toxic materials into the implant supporting area. Therefore, an early detection is essentially required to avoid possible failures that cause time and cost escalation. Objective :To indicated the existence of the protein laminin that could predict the development of transmucosal seal. Material and Methode :A laboratory experiment research was conducted on New Zealend White Rabbits that were screened throught appropriate research-requirement criteria set by Biofarma- Bandung-West Java-Indonesia. The implant material selected for the research were titanium which than applied on 32 dental implants ( included 16 for negative control). Two implant were respectively inserted into upper and lower jaws of eight male rabbitsthrought appropriate surgery. A control group was collected from peri implant tissues of eight clinically normal rabbits. Immunoexpression of related protein such as laminin as an indicator in this research, based on clinical confirmation. The research method utililizedexperimental and descriptive laboratory mechanism using ABC ( avidin-biotin-complex ) technique for examining protein immunoexpression.

Results :Regression analysis revealed that related protein expression correlated significantly with successrate of a dental implant treatment (p = 0.022). Expression of related protein between dental implant and normal rabbit tooth elicited incignificant different ( P laminin = 0,395 ). Statistical analysis also disclosed that success rate of using titanium implant material was significantly higher (p = 0.01) than using Stainless Steel as negative control.

Conclusions :The research concludes that the immunoexpression examination can provide early detection on success or failure degrees of dental implant treatment. The clinical examination with mobility test can also be trusted to early predict the successfullness of dental implant procedure.

Keywords : Dental Implant, laminin, immunohistochemical, immunoexpression

Invited Lecture 8

Design and fabrication of orthodontic bracket using Metal Injection Molding (MIM) process

Supriadi S1*, Irawan B2, and Suharno B3, Prasetyadi T2, Gratis A1, Zulkifly F F1 1Dept. of Mechanical Eng.,Universitas Indonesia, Jakarta,Indonesia 2Dept. of Dental Material,Universitas Indonesia, Jakarta,Indonesia 3Dept. of Metalurgy and Material Eng.,Universitas Indonesia, Jakarta,Indonesia *[email protected]

Introduction: Indonesian relatively has small jaws which is have tendency of improper teeth alignment called as malocclusion. Malocclusion prevalence in Indonesia reached 80% of the population.Recent years, as the increasing of Indonesian economy level and easier access to dental health services, more people looking for dental treatment especially in order to improve their aesthetic. One of the treatment to correct the malocclusion by using orthodontic appliance which mainly consist of orthodontic bracket and archwires. Until now, orthodontic appliances are still mainly imported from other countries, therefore the price becomes relatively higher and also the diseign is not specified for Indonesian.Meanwhile, the abillity of Indonesianmanufacturer of these orthodontic devices is still inadequate. In order to increase affordability and specific design for Indonesian, the aim of this study is to design orthodontic bracket for Indonesian and fabrication of orthodontic bracket using Metal Injection Molding (MIM) process.

Materials and Methods: In this work, stainless steel 17-4 PH is used which is already mixed with organic binder material to produce feedstock. Targeted bracket design is developed in or previous work. The small injection machine is designed and contructed The feedstock is injected in to a mold cavity toproduce a green part. This part still weak because the strength of the part is strength of polymer binder strength. The next process is debinding. In this process, binder material is removed using thermal by placing the green part in to a furnace to produce a brown part. The brownpart is sintered up to 1100 oC to get dens part. Geometrical analysis is conducted to osberved optimum processing conditions.

Results and Discussion: Small injection molding machine was contructed for small part metal injection molding process. The machine has capability to extrude and to inject . The extruder is useful for feedstcok making and injection mode is useful for injection material into a cavity. The injection temperature is set between melting temperatur and degradation temperature of the feedstock. Sintering. Sintering process lead shringkage that have to be compansated when design the mold. Therefore the mold should be bigger than the target design. Metal injection molding perform better surface finish than orthodontic bracket that produced using invesment casting.

Conclusions: Design and fabrication of orthodontic bracket using metal injection molding process has been presented. Mold design should considering the shrinkage during process. Therefore the mold should be bigger than the design. Even the surface roughness better than orthodontic bracket that produced by investment casting however further polishing process still necesarry.

Acknowledgements : Author would like to thanks to Ministri of Research and Higher Education for the reserach grant on PUPT grant 2016 to conduct this research

Invited Lecture 9

Properties of super-elastic Ni-Ti alloy for dental applications

Yoneyama T* Nihon Univ., Tokyo, Japan *[email protected]

Introduction: Ni-Ti alloy is a functional metallic material because of its special mechanical properties, such as shape memory effect and super-elasticity. Super-elasticity is a phenomenon at a temperature above the reverse transformation temperature range, associated with stress- induced martensitic transformation. The alloy exhibits high flexibility owing to its wide recoverable strain range, in which the stress is kept low. Since the alloy also has high corrosion resistance and good biocompatibility, many applications have been tried in the medical and dental fields. In this paper, the functional properties of Ni-Ti alloy orthodontic wires and endodontic files were evaluated in relation to the phase transformation of the alloy for their clinical performance.

Materials and Methods: Twenty Ni-Ti orthodontic round archwires were used in a bending test. To investigate the bending property of the wires, a three-point bending method was carried out. Four kinds of Ni-Ti alloy orthodontic rectangular archwires were used in a torsional test. The gauge length, torsional speed, and maximum angle of the test were set at 5.0 mm, 0.75 degree/s, 45 degrees, respectively. Five types of rotary Ni-Ti files were used in a cantilever bending test. The maximum deflection was set at 4.0 mm, and the initial loading point was 3.0 mm distant from the file tip. These tests were performed at 310 K. Phase transformation behavior of the Ni-Ti specimens were examined by differential scanning calorimetry (DSC). The scanning range was between 173 and 373 K.

Results and Discussion: In the load-deflection diagrams of the Ni-Ti orthodontic wires, there were conspicuous flat areas for the super-elastic types, where the load changed slightly during the loading and unloading processes. On the other hand, the load for the non super-elastic types was nearly proportional to the increasing and decreasing deflection. The super-elastic type wires exhibited high endothermic energy on the DSC curves, while the non super-elastic types showed very low energy. In the torsional moment-angle diagrams of the Ni-Ti rectangular wires, three kinds of the wires showed hysteresis curves, characteristic of super-elasticity. The reverse transformation finishing temperatures of these wires were below 310 K, while that of the other wire was above 310 K. Since super-elasticity is derived from the stress-induced martensitic transformation, these transformation temperatures are one of the most influential factors for the super-elasticity of the wires. The load-deflection curves of the Ni-Ti rotary files showed super-elastic hysteresis curves without permanent deformation. Although each file had different cross-sectional configuration, the files with low transformation temperatures showed higher bending load than those with high transformation temperatures. In conclusion, the functional properties of Ni-Ti alloy dental appliances were much influenced by the phase transformation behavior of the alloy, which should be carefully controlled for effective and safe application in dental treatment.

Luncheon Seminar

Possibility of bioactive materials containing Surface Pre-Reacted Glass-ionomer filler

Shuichi ITO* Health Sciences University of Hokkaido, Japan. [email protected]

In recent years, the conception that the tooth lifetime is extended by prevention and management of early caries, but not early intervention has established with the spreading of minimal intervention (MI) in restorative dentistry. This means the breakaway from the concept that caries inevitably receives cutting treatment. However, we must still rely to restoration techniques and materials. Hinoura defined a sequence of process, in which the restoration treatment participates, from occurrence of caries through loss of tooth by extraction, as “repeated restoration cycle”. He emphasizes the importance of delaying progress of the process for the extension of tooth lifetime. For that purpose, improvement of restorative materials and advance of restorative technologies are necessary. Recently, new type of adhesion system has been developed using S-PRG technology. In the technology, a stable glass-ionomer phase on the surface of fillers is formed by pre-reaction of reactive glass containing various ions with polyacrylic acid in the presence of water. S-PRG filler has anti-cariogenic activity come from its fluoride-releasing and recharging properties. S-PRG filler releases aluminium, boron, sodium, silicate, and strontium ions as well as fluoride ion. Silicate and fluoride ions have been known as strong inducers of mineralization and remineralization in dentin. Strontium and fluoride ions have also been known to improve acid resistance of hydroxyapatite in enamel and dentin by conversion of hydroxyapatite into strontium-apatite and fluoro-apatite, respectively. Moreover, it has been known that strontium ion inhibits bone resorption and promotes bone formation. Therefore, the clinical use of strontium-containing α-TCP cement is also anticipated in dentistry. By means of addition of the activity for remineralization of dentin collagen in nano-sized space to adhesive restoration materials, they can be maintained and function in the oral cavity for a long time. Consequently, dentin remineralization therapy that we propose would contribute to the remarkable advance of clinical dentistry. Our strategy would enable people to masticate and prevent them from the aspiration, leading to keep their health for a long time.

O-01

Possibility of bone regeneration of bone-like tissue induced by recombinant human BMPs in vitro

Hayashi T*, Kawase M, Hiroyasu K, Tomino M and Kawai T Aichi Gakuin Univ., Nagoya, Japan *[email protected]

Introduction: Recent experiments have attempted to induce bone from immature muscular tissue (IMT) in vitro using crude BMP and e-PTFE as a scaffold. This process induces bone-like tissue; however, this induced bone-like tissue has ossification potential within 2 weeks after implantation. Meanwhile, recombinant human BMP (rhBMP)-2, -4 and -7 has osteogenic potential both in vitro and in vivo. The aim of this study was to induce bone-like tissue from IMT in vitro by rhBMP-2, rhBMP-4 and rhBMP-7 first, and then implant this tissue into a calvarial defect in rats and finally assess the healing.

Materials and Methods: Commercially available rhBMP-2, rhBMP-4 and rhBMP-7 were used in this experiment. IMTs were obtained from forelimbs of the 20-day Sprague-Dawley embryonic rats and placed into a homogenizer with 10ng/μl each of rhBMP-2, rhBMP-4 and rhBMP-7 and then homogenized. The homogenized IMT was placed on ePTFE and cultured for 2 weeks. The analyses of histological observation, electron probe micro analyzer (EPMA), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR) were carried out following culture, respectively. RT-PCR was performed to examine the gene expression of Runx2, collagen type I, osteopontin and osteocalcin on days 7 and 14. Furthermore, specimens were implanted into Sprague-Dawley rats’ calvarial defect up to 3 weeks, and then evaluated by micro CT and histological observation.

Results and Discussion: The bone-like tissue, which was made up of osteoblast-like cells and osteoids, was partially observed by H-E staining. Moreover, strong mineral deposition was observed in the extracellular matrix by von Kossa staining. Relatively strong radiopacity was observed by micro CT at 2 weeks after culture. The expression of all osteoblastic marker genes was confirmed on both days 7 and 14. Ca, P and O were detected in the extracellular matrix by EPMA and were confirmed to be at almost the same position based on the findings of synchronized images. XRD patterns and FT-IR spectra of specimen were found to have typical apatite crystal peaks and spectra, respectively. Furthemore, the strong radiopacity and partial ossification was confirmed at 1 week after implantation, and dominant new bone was observed after 2 weeks in the defect area. These results suggest that rhBMP-2, rhBMP-4 and rhBMP-7 induced IMT differentiation into bone-like tissue in vitro, and this induced bone-like tissue has an ossification potential after implantation and promotes healing of calvarial defect.

Conclusion: Thus, immature muscular tissue has a possibility to be an effective tissue source for bone tissue engineering.

O-02

Cell Proliferation Ability of Mouse Fibroblast-Like Cells and Osteoblast-Like Cells on Pure Titanium Film Manufactured by Electron Beam Melting

Kawase M*, Hayashi T, Asakura M, Mieki A, Tomino M and Kawai T

Aichi Gakuin Univ., Nagoya, Japan

*[email protected] Introduction Electron Beam Melting (EBM) has been used for fabrications of specific net-shaped parts for use within both aerospace and medical implant industries. It has been reported that the characteristics of manufacture by EBM are different from usual casting. However, the physical and biological characteristics are not well known, and still now, there are not many reports about the biological characteristics. In this study, pure Ti was applied as a biomedical material because it has a good biocompatibility, which pure Ti film was manufactured using EBM. Moreover, we tested the cell proliferation and calcification ability of pure Ti film.

Materials and Methods The EBM apparatus (Arcam Q10, Arcam AB, Sweden) was used to manufacture the pure Ti film with a diameter of 10mm. Then, the Wettability of pure Ti surface was assessed by measuring the contact angle of 50 µL of distilled water placed on the pure Ti film. Fibloblast-like cells (L929) and osteoblast-like cells (MC3T3-E1) were suspended at 1×105 cells/mL in each medium and seeded at 400/µL. The cells were seeded directly on a 48-well micro-plate as a negative control. The proliferation of L929 cells was measured each day for 5 days, and measurements of proliferation for MC3T3-E1 cells were carried out on days 2, 4, 6, 8 and 10. Moreover, the morphology of L929 cells and MC3T3-E1 cells was observed using Scanning Electron Microscopy (SEM). Furthermore, Alizarin Red S Staining was performed on days 14, 21 and 28 to detect mineral deposition caused by MC3T3-E1 cells.

Results and Discussion The contact angle on the pure Ti film was 103.0±3.5° and showed slight hydrophobicity. It is reported that biomaterial surfaces with moderate hydrophilicity display improved cell growth and biocompatibility. However the cell number of both L929 and MC3T3-E1 favorably increased on pure Ti films, and especially MC3T3-E1 on pure Ti film increased more than that on control. In addition, the morphology of L929 and MC3T3-E1 cells were polygonal and spindle-shaped and the cytoskeleton was well developed in the pure Ti surface groups. As for Alizarin Red S Staining, a slight calcium deposition was observed from day 14 and this level became gradually remarkable. Thus, pure Ti film manufactured by EBM would be designed to function as true tissue substitutes that satisfy the patient-specific biological, mechanical, and geometrical requirements, and then be widely applied as a biomedical material in the near future.

O-03

Stem cell behavior on micro porous titanium implant

Bei Chang1, Tianxiao Han1, Fuping Li2, Hongchao Kou2, Yumei Zhang1* 1Department of Prosthetic Dentistry, School of Stomatology, The Fourth Military Medical University, Xi’an, People’s Republic of China 2State Key Laboratory of Solidification Processing, Northwestern Polytechnical University, Xi'an 710072, People’s Republic of China *e-mail ([email protected])

Introduction: Titanium are commonly used as artificial bone substitutes.1 However, the Young’s moduli of titanium could not match the trabecular bone commendably, which might lead to surrounding bone resorption.2 To minimize the negative effects of the incompatibility, porous titanium are extensively studied.2,3 The cell ingrowth of porous implant is elegantly dependent on the pore size, which has not been well elucidated yet. In this study, porous titanium with porosity of 70% and average pore size of 188–390 μm were fabricated and the stem cell behavior on the micro porous titanium implant were observed.

Materials and Methods: Porous titanium were fabricated by vacuum diffusion bonding of titanium meshes.4 The structure was characterized by FE-SEM and Micro-CT. The rat mesenchymal stem cells were obtained5 and seeded on the samples mounted in 24 well plates at a density of 5×104 cells/well. The cells proliferation within the samples was evaluated by DNA quantification at day 1, 4, 7, 14 and 21. The cells differentiation was evaluated by alkaline phosphatase activity and bone-related gene (ALP, COL-1, BMP-2, OPN, RUNX2) expressions at day 7 and 14. Mesenchymal stem cells expressing green fluorescent proteinwere used to observe the cell morphology during the culturing period. Photos were taken by fluorescence microscope.

Results and Discussion: The SEM images and Micro-CT scanning images showed the mesh wires are well aligned and the pore structure were homogenous. The average pore sizes of the three groups were 188 μm, 313 μm and 390 μm, which are in the range of appropriate pore size for cell penetration.4 The DNA contents of Ti 313 and Ti 390 were significantly higher than Ti 188 since day 4 while the fluorescence images showed that less cells were observed on the top of Ti 313 and Ti 390. The phenomenon could be attributed to higher permeability of relative larger pores, which results in less resistance for cell suspension when permeating the samples and lower cells seeding efficiency on the superficial layer. There was no significant difference of cells differentiation among the porous titanium at day 7 which was supported by Real Time RT- PCR and ALP activity. However, Ti 313 and Ti 390 exhibited lower ALP activity and bone- related gene expression levels than Ti 188 at day 14. These may stem from different mechano-biological stimuli within the porous structure6 and the reciprocal relationship between cell differentiation and proliferation.7 Our study demonstrates that small pores (Ti 188) is more inclined to promote cells differentiation at the beginning while cell proliferation within the porous titanium are biased to the relative large pores (Ti 313 and Ti 390). The hybrid designed porous titanium which combine the advantages of smaller pores and larger pores may be meaningful and promising for large bone defect restoration.

Acknowledgements: This work was financially supported by Natural Science Foundation of China 81470785&81530051

O-04

Chondrocyte differentiation ability of dedifferentiated fat cells compared with adipose-derived stem cells derived from the human buccal fat pad. Kubo H1, Nishio A1, Miya Y1, Hashimoto Y2, Kishimoto N3 12nd Department of Oral and Maxillofacial Surgery, Osaka Dental Univ., Osaka, Japan 2Department of Biomaterials, Osaka Dental Univ., Osaka, Japan 3Department of Oral and Maxillofacial Surgery, Osaka Dental Univ., Osaka, Japan * [email protected]

Introduction: Dedifferentiated fat cells (DFAT cells) and Adipose-derived stem cells (ASCs) reported to show the same high proliferation ability and multilineage potential as bone marrow mesenchymal stem cells. On the other hand, there are few reports that used human buccal fat pad (BFP), and the report is not seen that compared differentiation ability to cartilage by the three-dimensional culture. The purpose of this study was to evaluate and compare the chondrocyte differentiation ability of DFAT cells and ASCs from the BFP in the 3D construct.

Materials and Methods: We isolated human DFAT cells and ASCs from the BFP of a patient who underwent our hospital. (The ethics committee of Osaka Dental University: Approval No.110714). BFP was digested collagenase solution and centrifugation. The floating top layer containing mature adipocytes and precipitated stromal-vascular fraction (SVF) was collected. We isolated DFAT cells from mature adipocytes by ceiling culture and ASCs from SVF by conventional culture. Furthermore, DFAT cells and ASCs were seeded collagen sponge and cultured three dimensions in chondrogenic induction medium for 7, 14 and 21 days. We performed hematoxylin and eosin (HE) staining, Alcian blue staining and real-time PCR.

Results and Discussion: The cells attached to the upper surface of the flasks by ceiling culture. The cells were exhibited the characteristic views of cytoplasmic extension, reduction and lost the lipid droplets and fibroblast-like morphology. The cells in the 3D construct were evenly seeded in the collagen sponge and the round chondrocyte-like cells were exhibited by HE staining for 7, 14 and 21 days. The cells were also positive for alcian blue staining. Real time-PCR showed increased expression of aggrican, collagen type2, SOX9 for 7, 14 and 21 days. In addition, DFAT cells were significantly higher than ASC with gene expression of aggrican, collagen type2, sox9 for 14 and 21 days in real time-PCR. In this study, we were able to isolated DFAT cells and ASC from human buccal fat pad and exhibited chondrocyte differentiation in the 3D construct. In addition, DFAT cells were higher in the chondrocyte differentiation ability than ASCs.

Conclusions: We were able to isolate DFAT cells and ASCs from the human buccal fat pad and produced chondrocyte differentiation in a 3D construct. In addition, DFAT cells showed higher chondrocyte differentiation ability than ASCs.

Conflict of interest statement: The authors declare that they have no conflict of interest. This work was supported by JSPS KAKENHI Grant No.26463033.

O-05

In vivo evaluation of H2O2 hydrothermal surface modification with FGF-2 of mini-implants to enhance new bone formation

Matsuno T1*, Yoneyama2 Y, Miki T1, Asano K1 and Mataga I1 1 The Nippon Dental Univ. School of Life Dentistry, Tokyo, Japan 2 Tokyo Medical Univ. Tokyo, Japan * [email protected]

Introduction: Surface modification applied on Titanium (Ti) through various methods for creating unique microstructures improves biological responses and surface energy. Generally, the main purpose of physically and chemically modifying surfaces is to promote implant osseointegration. In our previous study, we demonstrated the efficacy of bone bioactivity 1) by H2O2 hydrothermal treatment on Ti disk surface in vitro . Fibroblast growth factor-2 (FGF-2) affects the BMP signaling pathway during bone morphogenetic protein-2 (BMP-2)-induced ectopic bone formation. Furthermore, FGF-2 has already been approved as a clinical pharmaceutical in Japan and is commercially available. Then we designed bio-active Ti surface modification using H2O2 hydrothermal treatment with FGF-2 for reducing healing time and achieving early osseointegration. In this study, we investigated that H2O2 hydrothermal treatment with FGF-2 promotes the bone formation around the mini-implant surface in vivo.

Materials and Methods: Ti-6Al-4V mini-implants with diameters of 1.4 mm and length of 6.0 mm (FCU1406, Re-Joine. Co., Ltd, Korea) were used in this study. All the mini-implants were ultrasonically cleaned in acetone, 100% ethanol, and distilled water for 15 min each. After cleaning, mini-implants were divided into four groups according to the method of surface modification applied as follows: only cleaning (control), control implants dipped into 0.1 μg/mL rhFGF-2 solution (FIBRAST® Spray 250; Kaken Pharmaceutical Co. Ltd, Tokyo, Japan) at room temperature for 24h (FGF-2), control implants hydrothermally treated with 3% H2O2 solution and autoclaved at 121°C and 0.2 MPa for 20 min (TiO2), TiO2 implants treated with FGF-2 (TiO2/FGF-2). These mini-implants were inserted into the respective femur of 15-week old Wister rats by self-drilling. After 4 weeks, femurs with mini-implant were excised, and then histological and histomorphometric analyses were performed.

Results and Discussion: Newly formed bone was observed around all implants. However, bone histomorphometry indicated that the percentage of bone–implant contact for FGF-2, TiO2, and TiO2/FGF-2 implants was consistently greater than that for control implants. On the other hand, bone volume associated with the TiO2/FGF-2 implants was consistently greater than for control, FGF-2, and TiO2 implants. In this study demonstrated that surface modification on the mini-implant using H2O2 hydrothermal treatment with FGF-2 enhanced the newly bone formation and promoted the osseointegration in vivo.

References : 1) Yoneyama Y, Matsuno T, Hashimoto Y, Satoh T. In vitro evaluation of H2O2 hydrothermal treatment of aged titanium surface to enhance biofunctional activity. Dent Mater J, 32, 115-121, 2013.

O-08

Analysis of dissolution behaviour of octacalcium phosphate and β- tricalcium phosphate bone substitute materials

Suzuki O1*, Sakai S1, Anada T1, Tsuchiya K1, Yamazaki H2, Margolis HC2 1Tohoku Univ., Sendai, Japan, 2 The Forsyth Institute., Cambridge, MA, USA *e-mail: [email protected]

Introduction: Hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP) have been used clinically as promising bone substitute materials in various bone defects [1]. Sintered stoichiometric HA is classified as a non-resorbable material, while β-TCP is recognized as a resorbable material [1]. We have reported that octacalcium phosphate (OCP) displays osteoconductive and bioresorbable properties, while OCP is progressively converted to an apatite phase when implanted in bone defects [2]. However, differences in the dissolution behavior between OCP and β-TCP under physiological conditions have not been clarified, which may be associated with the distinct appearance of the osteoconductivity of these materials. The aim of the present study was to compare the dissolution behavior of OCP and β-TCP under physiological conditions in vitro and in vivo.

Materials and Methods: OCP was prepared in a wet synthesis condition according to a method previously reported [3]. β-TCP used was commercially available as a sintered porous material (Osferion, Olympus Terumo Biomaterials Corp., Tokyo, Japan). The filter chamber made of PTFE and Millipore membrane, having an inner diameter of 8mm and 1 mm thickness, was filled with granule forms of OCP or β-TCP and then implanted into Wistar rat abdominal subcutaneous tissue for 8 weeks, to assess material dissolution. All procedures were approved by the Animal Research Committee of the institution. In vitro studies were also conducted by immersing OCP and β-TCP particles in simulated body fluid (SBF) at 37oC for up to 15 days, to similarly assess the dissolution behavior of these materials. X-ray diffraction (XRD), FTIR and chemical analyses were carried out. Degrees of saturation (DS) of the SBF solutions were determined.

Results and Discussion: X-ray peaks characteristic of the OCP structure changed to that of low crystalline HA, suggesting that OCP tends to dissolve or convert to HA after the implantation in the subcutaneous tissue. On the contrary, the structure of β-TCP implanted remained unchanged. The in vitro studies indicated a similar tendency with that seen from the in vivo analysis regarding observed structural changes. However, the SBF solutions in which OCP and β-TCP were immersed remained in a slightly supersaturated state with respect to OCP and β-TCP. The results indicate that OCP and β-TCP do not simply dissolve under physiological conditions [4] and suggest that the biodegradable properties reported in vivo [5] could mainly be caused by cellular resorption processes.

Acknowledgements: This study was supported by Grants-in Aid (23106010, 23390450 and 25670829) from the Ministry of Education, Science, Sports, and Culture of Japan.

References: [1] Chow LC. Dent Mater J, 28 (2009) 1-10, [2] Suzuki O et al., Biomaterials, 27 (2006) 2671-2681, [3] Suzuki O et al., Tohoku J Exp Med, 164 (1991) 37-50, [4] Sakai S et al. Dent Mater J, 35 (2016), [5] Miyatake N et al., Biomaterials, 30 (2009) 1005-1014.

O-09

Fabrication of carbonate apatite foam from the set calcium sulfate hemihydrate foam via solution mediated phase conversion

Sugiura Y1*, Tsuru K1, and Ishikawa K1 1Kyushu Univ., Fukuoka, Japan

* [email protected]

Introduction: Carbonate apatite (CO3Ap) scaffold has been attracted attention because of its high osteoconductivity, which can replace bone through a bone-remodeling process. The interconnected porous structure called as foam enables cells and tissues to intrude into the CO3Ap scaffold. The CO3Ap is fabricated from precursor materials, for example, calcium carbonate, calcium sulfate, and soluble calcium phosphate, through solution-mediated or hydrothermal phase conversion. The main challenge in fabrication of CO3Ap scaffold from precursor is how to promote the phase conversion reaction completely. Sometimes, small amount of precursor remains as residual materials at the core part of CO3Ap compact, due to the fact that the phase conversion rate depends on the difference in solubilities, on the other words, stabilities, of precursor and CO3Ap. Calcium sulfate hemihydrate (CSH) is known as a dental plaster, and its solubility is one of the highest of CO3Ap precursor materials. However, CSH when comes in contact with H2O quickly converted into calcium sulfate dihydrate (CSD); phosphate ion acts as an inhibitor of CSH–CSD conversion. The sodium dihydrogen phosphate (NaDP) can easily be obtained and used as granule particles. It is believed that NaDP granules can act as a phosphate source, a spacer, and a binder in CSH setting through salt precipitation process. In this study, we first fabricated CSH-NaDP porous compact, and then the CO3Ap porous compact was fabricated from CSH-NaDP porous compact through carbonate-phosphate solution immersion.

Materials and Methods: The CSH powder and sieved NaDP granules (0.6–1.0 mm) were mixed with water (L/P ratio~0.15) into separated-type DTS mold. The interior of CSH-NaDP porous compact was measured by microCT and XRD. The fabricated CSH-NaDP porous compacts were immersed into 1 M Na2HPO4–0.5 M NaHCO3 at 80°C for 24 h. The treated CSH-NaDP foams were analyzed by XRD, FT-IR, CHN, and DTS.

Results and Discussion: The microCT analysis showed that the porous structure has been formed inside the CSH- NaDP compact. The NaDP cast was partially connected and formed incomplete interconnected porous structure. The XRD measurement confirms that the setting reaction of the CSH-NaDP porous compact consisted of CSH and NaDP phases without CSD. The NaHCO3-treated CSH-NaDP porous compact exhibits that the CO3Ap maintains whole structure along with CSH-NaDP porous structure. The CO3Ap porous compact was directly fabricated from CSH-NaDP compact. During the setting reaction of the CSH-NaDP compact, CSH was preserved due to the phosphate- binding effect from NaDP granules.

O-10

Effect of porous alpha-tricalcium phosphate with immobilized basic fibroblast growth factor on bone regeneration in a canine mandibular bone defect model

Kobayashi N1*, Hashimoto Y1, Otaka A2, Yamaoka T2 and Morita S1 1Osaka Dent Univ., Osaka, Japan 2NCVC, Osaka, Japan * [email protected]

Introduction: Among the various calcium phosphate materials available, previous studies have highlighted the potential of alpha-tricalcium phosphate (-TCP) particles as bone rebuilding materials because they gradually biodegrade as bone regenerates around them. Basic fibroblast growth factor (bFGF) enhances the proliferation of undifferentiated mesenchymal cells, resulting in the promotion of bone formation. In this study, we evaluated the effect of porous -TCP with immobilized bFGF on bone regeneration in a canine mandibular bone defect model.

Materials and Methods: Porous -TCP particles were prepared by pulverization of an -TCP block with 80% of the pores showing a continuous porous structure. -TCP particles were placed in a peptide solution prepared in distilled water and incubated for 24 h at 50ºC. After rinsing, the powder was dried overnight. Subsequently, the particles were immersed in heparin solution for 8 h at room temperature and then dried overnight in vacuum. Immobilization of b-FGF to the heparin modified α-TCP particles was performed by immersion in 1 mg/mL bFGF (Fiblast Spray) in distilled water for 24 h at 4°C. The surface elements of porous -TCP in each reaction process were analyzed by scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS) analysis. Identical bone defects in the canine mandible (4 mm diameter, 6 mm depth) were made using a bone defects twist drill and were divided into two groups. One group of defects was filled with porous α-TCP bound to bFGF via heparin (bFGF group). The other group of defects was filled with untreated porous α-TCP. Micro- computed tomography and histological evaluation were performed at 2, 4, and 8 weeks after implantation.

Results and Discussion: SEM image shows that the porous -TCP particles had a continuous pore structure, with a pore diameter of approximately 5-10 m. The N 1s peak derived from b-FGF was found on porous α-TCP bound to bFGF through heparin in XPS analysis. The volume density and bone mineral contents of new bone were higher in the bFGF group than in the control group at 4 and 8 weeks. Histological evaluation at 4 weeks after implantation revealed that the porous - TCP had degraded and that newly formed bone was present on the surface of the -TCP particles in both the groups. At 8 weeks, continuous cortical bone formation with a Haversian structure covered the top of the bone defects in the bFGF group. This study demonstrates that porous -TCP with immobilized bFGF is sufficiently adaptable for treatment of bone defects.

O-11

Preparation of calcium hydrogen phosphate loaded with protamine and its anti-bacterial property

Abe K1, Honda M1, Iohara K2, Tajima Y2, Kamata A2, Kokaji S2, Yamamoto T3, Hayakawa T3 and Aizawa M1* 1Meiji Univ., Kanagawa, Japan 2Maruha Nichiro Corporation, Tokyo, Japan 3Tsurumi Univ., Kanagawa, Japan *[email protected]

Introduction: Calcium phosphates are one of major components of bone and teeth, and clinically used as artificial bones. However, bacteria could adhere to these biomaterials and cause in-hospital infection. Thus anti-bacterial processing for these biomaterials has been required. On the other hand, protamine is a series of arginine-rich protein, which can be obtained from fish and has anti-bacterial properties [1]. In this study, we prepared the calcium hydrogen phosphate loaded with protamine and examined its anti-bacterial property.

Materials and Methods: Calcium hydrogen phosphate powders were ground using a planetary ball mill for 3 h. The slurry after ball-milling was freeze-dried to prepare ground powders. Ground powders were immersed into protamine solution for 48 h; and then powders were freeze-dried. Adsorbed amounts of protamine to powders were measured by high-performance liquid chromatography (HPLC). Antibacterial activity of resulting powders was examined using Escherichia coli (E. coli) by scanning electron microscopy (SEM).

Results and Discussion: The adsorption amounts of protamine to the calcium hydrogen phosphate powders before and after ball-milling were at 1.52 and 3.63 mg∙g-1, respectively. Specific surface area of the (a)(a) (b) calcium hydrogen phosphate powder Fig.1 SEM images of E.coli cultured on the calcium hydrogen increased from 3.21 to 6.78 m2∙g-1 after phosphate compacts with(a) and without(b) protamine ball-milling; however, the zeta potential (scale bar : 1 m) of the ball-milled powder was almost the same as that of the powder without ball-milling. Thus, increase of adsorption amounts may be caused by increment of specific surface area. Anti-bacterial tests showed that the number of bacteria decreased on the protamine-loaded calcium hydrogen phosphate compacts. Figure 1 shows the morphologies of E. coli cultured for 24 h on the compact with and without protamine. The morphology of E. coli cultured on the compact with protamine was elongated.

Conclusions: Calcium hydrogen phosphate loaded with protamine showed the anti-bacterial property to E.coli. Protamine may be effective for the development of anti-microbial biomaterials.

References: [1] Johansen C, Gill T, Gram L, J. Appl. Bacteriol., 78, 297-303 (1995).

O-12

Characteristics of experimental calcium silicate as a pulp capping material

1 2 1 1 1 1 1 Hayashi Y , Kawaki H , Hori M , Hasagawa T , Tanaka M , Kawano S , Yoshida T , and Tamaki Y3 1~3Asahi Univ., Mizuho, Japan * [email protected] Introduction: Mineral Trioxide Aggregate (MTA) has been popular in endodontic field; however, the manipulation in clinical use is not advantageous due to its long setting time. Moreover, high cost is not negligible problems as compared with other products. MTA consists of several ingredients such as SiO2, CaO, CaSiO3, Al2O3, CaSO4, and Bi2O3. Particularly, some kinds of calcium silicates (CaSiO3, Ca2SiO4, and Ca3SiO5) have been considered to be useful as a pulp capping material because of their excellent biological abilities. The aim of this research was to syntheses artificially calcium silicates and evaluate their characteristics for a pulp capping material. Materials and Methods:

Referring to chemical equation between CaCO3 and SiO2, both of reagents were individually measured and the mixture was heated up to 1300ºC in an electric furnace. After sintered and powdered, it was analyzed by X-ray diffraction apparatus (Ultima IV, Rigaku) with a range of 10-60 degree. To improve clinically operability, high-purity plaster (CaSO4 ‧ 1/2H2O, Shimomura Gypsum Corp.) was blended to syntheses. Three kinds of samples were prepared with different rates. The addition of plaster to the synthetic substance was designed each of 25, 33, 40wt% (Code: A-C, respectively). After 24 hrs from preparing specimens (diameter: 6mm, height: 12mm), compressive examination was performed by universal testing machine (Autograph, Shimadzu Co.Ltd.). Setting time of each specimen was also measured in accordance with a JIS (T 6603-1994). Then, water/powder ratio was set at 0.4-0.5 from preliminary test. Commercial pulp capping product (Code:D, Dycal, Dentsply Co. Ltd.) was also examined and compared as a control. Furthermore, an influence of high-purity plaster mentioned above was also investigated (Code:P). In addition, we evaluated the cellular responses to our calcium silicates using human dental pulp stem cells (hDPSCs) in culture. Results and Discussion:

XRD results suggested that Ca2SiO4 was artificially formed. When mixing it with deionized water, Ca(OH)2 was newly generated, but its slurry was incompletely hardened. Therefore, the addition of plaster to Ca2SiO4 was investigated. As results, added groups (A-C) showed larger compressive strength (8.7 – 12.2 MPa) than that of D, and had acceptable setting time within 24 - 27 minutes. Particularly, hardening behavior was remarkably improved and setting time was obviously shortened comparing with that of MTA reported by previous article1). In addition, synthesized Ca2SiO4 materials showed little cytotoxicity and did not inhibit cell proliferation of hDPSCs. In summary, newly synthesized Ca2SiO4 materials was considered to be hopeful as the pulp capping material. References: 1) Shahi S et al. The Effect of Different Mixing Methods on Working Time, Setting Time, Dimensional Changes and Film Thickness of Mineral Trioxide Aggregate and Calcium- Enriched Mixture. Iran Endod J. 2015. 10:248-251.

O-14

Three-dimensional characterization and distribution of fabrication pore defects in bilayered lithium disilicate glass-ceramic molar crowns

Zi-Hua He 1, Li Dao 1, Yu-Tao Jian 1, Michael V. Swain2 and Ke Zhao 1* 1Department of Prosthodontics, Guanghua School of Stomatology, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Stomatology, Guangzhou, China 2Faculty of Dentistry, the University of Sydney, NSW 2006, Australia *e-mail: [email protected]

Introduction: Bilayered dental ceramic have been widely used in prosthetic dentistry. However, fracture and chipping have been reported. Fractographic investigations shown the pores existed in the ceramic largely compromise the mechanical behavior. Clarifying the characteristics and distribution of pores should provide better insight to understand the relationship between pores and crown failure. So far, researches were generally based upon the 2D images analysis, which may not adequately describe pores’ tortuous 3D morphology, which could induce false predictions when considering the influence of the pores. In the present study, bilayered lithium disilicate glass-ceramic (LDG) molar crowns were selected for investigating the characteristic form and distribution of pore defects by means of high resolution micro-CT. The 3D reconstruction and statistical analysis were performed to characterize the pore defects.

Materials and Methods: Ten standardized molar crowns (IPS e.max Press; Ivoclar Vivadent) were fabricated by heat- pressing on a core and subsequent manual veneering. All crowns were scanned by micro-CT and 3D reconstructed for pores analysis using VGStudio Max 2.2. Volume, position and sphericity of each defect was measured in every crown. Each crown was divided into four regions – central fossa (CF), occlusal fossa (OF), cusp (C) and axial wall (AW). Porosity size and number density of each region were calculated. Statistical analyses were performed using Welch two sample t- test, Friedman one-way rank sum test and Nemenyi post-hoc test. The defect volume distribution type was determined based on Akaike information criterion.

Results and Discussion: The 3D spatial morphologies of crown and inner pore defects can be visible after reconstruction as figure below. Pores were detected both in veneer and core in the same manner. The total number of pores was 119,771. The distribution of defect size was ranging in volume from 3,241.00 µm3 to 8.54 × 108 µm3 with the equivalent diameter was between 10 µm to 600 µm. Core ceramic contained fewer, smaller and more regular defects than veneer ceramic because of different fabrication process. Pores in veneer were more likely to occur in the CF region than that of region AW (P < 0.001) and C (P = 0.009), while no difference was found between region CF and OF Large defects were more likely to occur in the core-veneer interface of occlusal fossa. The predicted strengths on the basis of a simple fracture mechanics approach (Eq. 1) are comparable with previously reported values. Small defects (volume ≤ 95% ) size distribution was obeyed a logarithmic normal distribution as Eq. 2 with μ and σ were estimated as 9.15 and 0.95.

2 K1c 1  (ln x  )    (1) f (x)  exp  2  (2) 1.12 c x 2  2 

O-15

Evaluation of Newly Developed Investment Material for Press Ceramics.

Mori D*, Yoshinaga M, Fujimoto T, Mashio G, Yokohara H, Hoshino T, Miyake T, Sato T, Kumagai T GC corporation, Tokyo, Japan * [email protected] Introduction: Newly developed Lithium silicate ceramic system, initial LiSi press (GC Corp, LP) and LiSi press vest (GC Corp, LPV) was introduced into the Japanese market in March 2016. One of the features of this system is the easy removal of the reaction layer due to the product containing Boron Nitride (BN). The purpose of this study was to evaluate the removing property of the reaction layer which was generated during the pressing procedure. Materials and Methods: LPV consists of powder, Liquid and SR liquid. The powder contains BN and SR liquid which contains a high concentration of BN for the easy removal of the reaction layer. The test material which did NOT contain BN was prepared to compare the influence on the reaction layer. Table-1 Combination of Investment materials Powder Liquid SR Liquid Ceramic Combination1 Test powder LPV Liquid Not used LiSi press MT A2 (Without BN) (LP) Combination 2 LPV Powder LPV Liquid Not used LP Combination 3 LPV Powder LPV Liquid SR Liquid is used LP Two Acryl disks (φ 13mm, thickness 2mm) were attached to a Ring base system LiSi press (GC Corp) with sprue. The SR liquid was sprayed on to the acryl disks only for combination3. The disks were invested following the combination in table-1. Invested disks were put into the pre-heated furnace at 850 deg C for a time period of 60 minutes. The LP was inserted into the fired investment and heat-pressed by a Panamat press (GC Corp) according to the schedule below. Table-2 Press schedule Start temperature Heating rate Final temperature Hold time Press Duration 700 deg C 60 deg C 903 deg C 25 minutes 5minutes One heat-pressed ceramic disk was divested carefully to remove the excess investment material except for an area that was in contact with the surface of the disk. The disk was defined as a ‘before sandblasting sample’. The other disk was divested by a sandblaster (Basic quarto, Renfelt) with glass beads at 0.4Mpa pressure to remove all the investment material and was defined as an ‘after sandblasted sample’. Both disks were embedded in an epoxy resin and the cross-sections were observed by a SEM (MINISCOPE TM3000, HITACHI) to make comparisons before and after. Results and Discussion: Combination 1: Approximately 50micrometers thickness of reaction layer between the ceramic layer and the investment layer was observed before and after. Combination 2: Approximately 10 to 30 micrometers of orbicular holes were observed in the reaction layer before sandblasting sample. Most of reaction layer was removed after the sample was sandblasted. A slight reaction layer was still observed. Combination 3: Approximately 50micrometers of orbicular holes were observed along the ceramic layer before sandblasting sample. No reaction layer was observed after the sandblasted the sample. According to the results, it is expected that BN which is contained in the LPV Powder and in the SR Liquid generated the orbicular holes between the ceramic layer and the investment layer and it enabled the reaction layer to easily be broken down.

O-16

Effects of rapid heating and shape of semi-sintered zirconia crown on marginal fit accuracy

Ohkuma K1*, Kameda T1, Ebihara Y2, Mizuguchi Y2and Miyagawa Y1 1Nippon Dental Univ., Niigata, Japan 2GC Corp., Tokyo, Japan *e-mail ([email protected])

Introduction: A normal heating time for zirconia ceramics is very long (6.5 hours). To make a CAD/CAM crown quickly, it is important to know the effect of rapid heating on dimensional accuracy. Furthermore, the effect of unbalanced buccal and lingual thicknesses of zirconia crown has not been clarified yet. The purpose of this study was to evaluate the effects of rapid heating and shape of semi-sintered zirconia crown on marginal fit accuracy.

Materials and Methods: The material used was semi-sintered blank (Aadva Zirconia Disk ST, GC). Three types of crown shape were tested. The buccal thickness of the crown was the same for all types, however three different lingual thicknesses were prepared as shown in

Fig.1. The three-dimensional data were input into the machining device (the cement layer was 0 mm) and ten crowns were milled for each shape using a CAD/CAM system (GM- 1000, GC). The full-sintered crowns were fabricated in two kinds of heating schedule: a) Normal heating (to 1000℃ for 2 hours and to 1450 ℃ for 4.5 hours) and b) Rapid heating (to 1450 ℃ for 1 hour). The Fig.1 Dimensions of three semi-sintered crowns marginal fit accuracy was measured as a discrepancy between the crown margin and the metal die baseline using a digital measuring microscope system.

Results and Discussion: As shown in Fig.2, the discrepancy at normal heating was significantly larger than that at rapid heating for each shape. Significant differences in the discrepancy after rapid heating were observed between the lingual thicknesses of 0.5mm and 1.5mm (p<0.01), and between the lingual thicknesses of 1.0mm and 1.5mm (p<0.05). Fig.2 Effects of heating speed and shape of semi-sintered crown

O-17

Influence of different finishing procedures on the wear behavior of zirconia ceramics

Hata U1*, Doi Y1, Nakatsuka T2, Mori D1, Yamamura O1 and Fujiwara S1 1Asahi Univ., Gifu, Japan 2Shofu Co. Inc., Kyoto, Japan *e-mail [email protected]

Introduction: All-ceramic restorations are widely used in esthetic dentistry. There has been a significant growth in interest for zirconia ceramics in recent years, with a major impact in the field of prosthodontics. Due to its mechanical properties, biocompatibility, and optical properties, zirconia has been championed as a metal free alternative. The purpose of this study was to evaluate the influence of different finishing procedures on the wear behavior of anatomic contour zirconia ceramics against enamel-mimicking specimens.

Materials and Methods: Instead of natural enamel, disk specimens whose hardness number was essentially the same as enamel were made by sintering carbonate apatites (C/P=1) at 800℃ for 2 hours. The surfaces of zirconia specimens (diameter 3mm, thickness 6.00mm) , which were used as the antagonists of enamel-mimicking were fabricated from Kyocera Y-TZP zirconia. The study included 4 groups : polished with the abrasive paper of 4000-gride silicon carbide paper (Struers)(①；ZP), polished with the abrasive paper and grinded with Vitrified dia (Shofu)(②;ZPG), polished with the abrasive paper, grinded with vitrified dia and polished with Zircoshine (coarse, medium, fine )(Shofu)(③;ZPGP) and polished with abrasive paper, grinded with vitrified dia , polished with Zircoshine (coarse, medium, fine) and polished with Zircon-Brite (DVA)(④;ZPGPP). Two-body collision and rotation wear testing was performed according to the Alabama wear testing procedure (vertical force: 75N, 10,000 cycles, rotation: 30°, water temperature:37℃)． The amount of wear was evaluated with different methods for zirconia and enamel- mimicking specimens. For zirconia specimens, the wear was estimated from the decrease in the thickness, while for enamel mimicking specimens, it was estimated with the most surface depth (Rz [μm]).

Results and Discussion: The amount of wear for enamel mimicking specimens (surface depth: Rz) was greatest with ZPG (125.7μm) , followed by ZPGP (13.8μm), ZPGPP(8.7μm) and ZP(1.8μm) . No significant differences were observed in the amount of wear for zirconia specimens under the four conditions.

Conclusions Within the limitations of this study, mirror polished zirconia is wear-friendly to the opposing tooth.

O-18

Change in surface of yttria-stabilized zirconia in water and Hanks’ solution characterized using XPS

Oishi M1*, Tsutsumi Y1, Chen P1, Doi H1 and Hanawa T1

1Tokyo Medical and Dental Univ., Tokyo, Japan *e-mail [email protected]

Introduction: Yttria-stabilized zirconia (YSZ) has been widely used in dentistry. Although YSZ is known as chemically stable, YSZ has been sometimes reported to compatible to tissues in dental implants. To clarify the mechanism of tissue adhesion to YSZ, we investigated the reaction of YSZ surface with water molecule and quasi-biological environment. In addition, titanium was also investigated in the same condition as a well-known reference material.

Materials and Methods: Three kinds of YSZ plates (10mm x 10mm x 0.5mm) containing 13mol% yttria with crystalline planes of (100), (110) and (111) were purchased from Dalian Keri Optoelectronic Technology Co. Ltd. These crystalline planes were confirmed by X-ray diffraction (XRD). Their roughness is less than 0.5nm. The specimens were cleaned by ultra-sonication in acetone and ethanol twice for 5 min, respectively. These specimens are named as (i) “Polished”. The “Polished” specimens were immersed into (ii) ultra-pure water for 1d and 60 d named as “Water” or (iii) Hanks’ solution without glucose for 7 d named as “Hanks”. Then, the specimens were rinsed in ultra-pure water and dried with N2 gas flow. The specimens were characterized using X-ray photoelectron spectroscopy (XPS) through X-ray photoelectron spectrometer (JPS-9010MC, JEOL). CP-Ti disks with 8-mm diameter were evaluated with the same as the above YSZ specimens.

Results and Discussion: 3- XRD patterns showed crystalline planes of exact PO4 (100), (110) and (111). Zr, Y, and O as well as C as YSZ(111) contaminant carbon were detected from “Polished”, 3- 2- PO “Water” and “Hanks”. Zr and Y existed as oxide HPO 4 2- - 4 YSZ(110) states. O was originated from O , OH , and H2O. 3- Relative concentrations of Y in (100) and (111) were PO4 smaller than the nominal composition, while that in YSZ(100) (110) was the same as it. The “Water” with (110) immersed for 60 d contained a large amount of hydroxyl groups. On the other hand, P was detected Binding Energy, Eb / eV from “Hanks”, especially in (110) specimen. P Figure 1 XPS spectra of P 2p electrons existed as phosphate, as shown in Figure 1. from YSZ(100), (110), and (111) According to the above results, surface reaction immersed in Hanks’ solution for 7 d. rates are different among crystalline planes. Real YSZ used in dentistry contains all planes randomly. Therefore, resultant reaction occurs according to the mixture of all plane reactions. The concentration of hydroxyl groups on YSZ increase in water and phosphate ions chemically adsorbed during immersion in Hanks’ solution. These phenomena may accelerate tissue adhesion to YSZ in vivo.

O-19

Aspects of Silicon Release from Resin Composites and Glass Ionomer Cement

Yon MJY, Tsoi JKH*, Matinlinna JP The University of Hong Kong, Hong Kong SAR, China *e-mail: [email protected]

Introduction: Resin composites and glass ionomer cements (GIC) contain silicon, which is a key component of the filler content. Both dental restorative materials are subject to degradation by water in the humid oral environment and their mechanical properties may be adversely affected. In this study, the release profiles of silicon in water from the two types of dental restorative materials were investigated.

Materials and Methods: A total of 60 specimens from resin composite (Filtek Supreme XT Restorative) of 5 different shades (A1B, A2B, A3B, A3.5B & A4B) and a glass ionomer cement (Ketac Molar, 3M ESPE) were prepared, immersed in deionised water and stored in closed containers at 37°C / 99%RH. At 5 different time intervals, a portion of liquid was quenched. The elemental concentrations of silicon were analysed using ion coupled plasma- optical emission spectrometry (ICP-OES, SPECTRO ARCOS, Germany). Cumulative release profiles of silicon were fitted by the second-order least-square regression method.

Results and Discussion: Silicon was found to be released from all dental restorative materials in the study, such that the cumulative release profiles were highly follow second- order kinetics (r2~0.97 to 1.00). For resin composites, silicon release was found to stabilise after some time when immersed in deionised water regardless of shade, whereas that of the GIC was found to accelerate (Fig. 1). The release of silicon in both resin composites and GIC may be detrimental to their mechanical properties after their placement in the oral cavity. In particular, the continuous increase in leaching of silicon from GIC might shorten the service time of this restorative material.

10 Filtek-A1B Filtek-A2B Filtek-A3B 8 Filtek-A3.5B Filtek-A4B GIC-Ketac regression line 6

[Si] / ppm / [Si] 4

0 Fig. 1 Cumulative release of Silicon from Dental 0 200 400 600 800 1000 Restorative Materials stored in deionised water time / hrs Conclusion: Silicon is leachable from dental restorative materials following certain pattern.

Acknowledgements: This work was partially funded by the Undergraduate Research Programme, Faculty of Dentistry, The University of Hong Kong. We have no significant financial interest/arrangement or affiliation with an organisation / institution whose products or services are being discussed in this research.

O-20

Relationship between mechanical properties, interfacial characteristics and bond fatigue strength of short fiber-reinforced resin composite with universal adhesive

Tsujimoto A1,2*, Barkmeier WW2, Takamizawa T1, Latta MA2 and Miyazaki M1 1Nihon Univ., Tokyo, Japan 2Creighton Univ., Omaha, Nebraska, USA *e-mail: [email protected]

Introduction: The purpose of this study was to investigate the relationship between mechanical properties, interfacial characteristics and bond fatigue strength with a universal adhesive used with short fiber-reinforced resin composite (SFRC) compared to other composite restoratives.

Materials and Methods: The resin composites used were a SFRC, micro-hybrid resin composites (MHRCs) and nano-hybrid resin composites (NHRCs) and three universal adhesives used were. The fracture toughness and flexural properties of resin composites, the interfacial characteristics of resin composites and universal adhesives, and shear bond strength (SBS) and shear fatigue strength (SFS) of universal adhesive bonds with resin composite using both total-etch and self-etch modes were determined.

Results and Discussion: SFRC showed a significantly higher fracture toughness value than MHRCs and NHRCs (p<0.05).1 The flexural properties of SFRC were comparable with NHRCs, but they were significantly lower than those of those of MHRCs (p<0.05).1 The interfacial characteristics of SFRC were similar to MHRCs, NHRCs and cured universal adhesives unlike uncured universal adhesives.2 Regardless of etching mode, the SBS values of universal adhesives with SFRC did not show any significant differences from those with MHRCs and NHRCs. The SFS values of universal adhesives with SFRC and MHRCs were significantly higher than those of NHRCs (p<0.05).3 The results of this study suggest that the similarity of the interfacial characteristics of SFRCs, other composite restoratives and cured universal adhesive influence SBS of universal adhesive, and the mechanical properties of SFRC improve their SFS with the use of universal adhesive. The enhanced mechanical properties and bond fatigue strength with universal adhesive of SFRC might improve the clinical longevity of resin composite restorations.

References: 1. Tsujimoto A, Barkmeier WW, Takamizawa T, Latta MA, Miyazaki M. Mechanical properties, volumetric shrinkage and depth of cure of short fiber-reinforced resin composite. Dent Mater J 2016, in press. 2. Tsujimoto A, Barkmeier WW, Takamizawa T, Latta MA, Miyazaki M. Bonding performance and interfacial characteristics of short fiber-reinforced resin composite in comparison with other composite restoratives. Eur J Oral Sci 2016, in press. 3. Tsujimoto A, Barkmeier WW, Takamizawa T, Watanabe H, Johnson WW, Latta MA, Miyazaki M. Relationship between mechanical properties and bond fatigue strength with universal adhesive of short fiber-reinforced resin composite. Eur J Oral Sci 2016, in press.

O-21

Evaluation of GIC-Surface Treatment on Bond Strength to Resin Composite

Sakamoto M, Arita A, Fukushima S, Kumagai T Research & Development, GC Corporation, Tokyo, Japan *e-mail [email protected]

Introduction: Glass ionomer (GI) or resin-modified glass ionomer (RMGI) is frequently used as the base/liner material under resin composites. This “sandwich technique” is suitable to reduce sensitivity and marginal leakage. The purpose of this study was to evaluate the effect of surface treatment on bond strength of GI and RMGI to resin composite.

Materials and Methods: Fuji II LC (F2, RMGI, GC) and Fuji IX GP EXTRA (F9, GI, GC) were filled in mold (4 mm diameter and 6 mm depth hole in center of cylindrical acrylic block) at 37oC / 80% R.H. The surface of F2 was light-cured for 20 sec immediately after filling. F9 was stored in 37oC / 80% R.H. for 2 min 30 sec after starting mix. Afterward, the surface of specimens either ground with #600-grid silicon carbide paper (#600) or unground (UG). The surfaces were treated as follows; 1) G-Premio BOND (GPB, GC), 2) no bond treatment (NB). Then flowable composite (G-aenial Universal Flo, GC) was placed on their surface using plastic button mold (2.38 mm hole diameter using Ultradent jig). The specimens (5 specimens per group) were immersed in distilled water at 37oC for 24 hrs or placed in thermal cycling (10000 cycle, 5-55oC). Shear bond strengths were measured at 1 mm / min crosshead speed (EZ-S 500N, Shimadzu). Statistical analysis were performed using one-way ANOVA followed by Tukey’s Test between 4 group (F9-24h, F9-TC, F2-24h and F2-TC), same letters in each groups indicate no statistically significant difference (Tukey’s test, p < 0.05).

Results and Discussion: Both bond strengths of F9 with G-Premio BOND treatment were higher than with no bond treatment. The #600-grid with no bond treatment of F2 was lower than unground surface. The untreated surface (unground, no bond treatment) of F2 is tightly bonded with resin composite. It is considerable that oxygen inhibition layer forms covalent bond resin composite chemically.

Conclusions: GI might need bonding treatment to obtain their bonding strength. RMGI did not require bonding treatment if air inhibition layer exists on the surface (unground surface).

O-22

Bond strength of experimental epoxy resin-based sealer cement with submicron- size zirconium oxide filler particles

Pane E1,2*, Messer HH1, Palamara J1

1Melbourne Dental School, University of Melbourne, Australia 2Faculty of Dentistry, Universitas Sumatera Utara, Indonesia

* [email protected]

Introduction: The aim of this study was to evaluate the influence of reduced-size filler particles incorporated into an epoxy resin-based sealer cement on tensile, shear and push-out bond strengths to root dentine and its penetration into dentinal tubules.

Materials and Methods: A specially formulated sealer cement containing zirconia filler particles averaging 0.6-0.8 µm in size was prepared, with working properties similar to those of a commercially available epoxy resin-based sealer with filler particles averaging 1.5-8 µm. Thin and thick layers (0.1 or 1.0 mm) of both sealers were bonded to root dentine, using 10 samples for each material and test. To test tensile and shear bond strengths to root dentine, an alignment device was prepared for accurate positioning of 2 mm diameter root dentine cylinders with concentrically positioned 2 mm stainless steel rods fixed in a predefined gap. Push-out strength was measured using 1 mm thick root slices following root filling with either material. Strengths were measured after 7 days using a universal testing machine. Then bond strengths of the two materials were compared. Representative surfaces were examined visually and using SEM and EDS analysis. Data were analysed using ANOVA with the minimum level of significance set at p< 0.05.

Results and Discussion: Thick layers of both sealers produced higher bond strengths compared to thin layer except in tensile bond strength of conventional sealer, with significant differences only in the shear strength of the experimental sealer (P<0.05). Compared to the conventional epoxy resin-based sealer, the reduced-filled experimental sealer was not significantly different, with respect to bond strength, except for the tensile and shear strengths of thick layers (p<0.05). Reduced-size filler particles penetrated the dentinal tubules more deeply than the larger conventional filler particles.

Conclusion: Reduced-size filled epoxy resin-based sealer cement behaved similarly to the conventional epoxy resin-based sealer. The use of reduced-size fillers did not enhance the bond strength to root dentine.

Acknowledgement: The experimental material used in this study was provided by Dentsply, Germany.

O-23

Bond strength of self-adhesive flowable resin composite to artificial tooth erosion by scratch test and tensile bond strength test Murase Y1*, Kotake H1, Kusakabe S1, Okuyama K2, Tamaki Y2 and Hotta M１ Department of Operative Dentistry, Division of Oral Functional Science and Rehabilitation, Asahi University, School of Dentistry, Gifu, Japan 1 Department of Dental Material Science, Asahi University, School of Dentistry, Gifu, Japan 2 *e-mail: [email protected] Introduction: Tooth wear is significant dental disease for us same as caries and periodontal disease. Needs of treatment for tooth wear are increased, however the information about it is not enough to our understanding. The purpose of this study was to evaluate the bond strength between artificial acid erosive tooth surface (one kind of tooth wear) and self-adhesive flowable resin composites as a tooth-coating material.

Materials and Methods: Human extracted teeth were used in this study. Flat colonal enamel or dentin surface was exposed by #600 SiC paper, and then samples were kept into artificial saliva (AS) (pH:7.0) for 2hrs. For artificial acid erosion group (group E), tooth samples were immersed into acidic carbonated beverages (Coca-Cola, Coca-Cola Japan, Japan) at 4ºC for 5 min with shaking. After immersion, samples were rinsed with water at room temperature for 5 min with shaking, and then stored into AS at 37ºC for 6.5hrs. This cycle was alternated three times. For remineralization group (group R), tooth samples were stored into AS for same periods as group E. Ten samples on each surface, group and material were subjected in this study. Two flowable resin composites (zero-step resin composite; fusio Liquid Dentin, Pentron Clinical: fusio, and one-step resin composite; LLB-CR6, Tokuyama dental, Japan: LLB) were used. Each material was applied on enamel or dentin surface following manufacture’s instructions with acrylic plate and pressed with 200g for 30 sec, then light cured for 20 sec. Bond strength of each specimen were measured by scratch test (sample scan speed: 5.245mm/min, load:0.03-15.0N) and tensile bond strength test (cross-head speed: 0.5mm/min). The data were statistically analyzed by one-way ANOVA and Fisher’s PLSD test (p<0.05).

Results and Discussion: On scratch test, for fusio on enamel surface, group R shows higher value than group E. For fusio on dentin surface and LLB, group R shows higher value than group E. There is no significant difference between group R and E. On tensile strength test, for LLB, group R shows higher value than group E on enamel and dentin surface. For fusio, there is no significant difference between group R and E. While, the values are different according to the testing method, the bond strength on erosive surface exhibit similar value compared with remineralized surface. These results of this study suggest that both fusio Liquid Dentin and LLB-CR6 might be useful for coating erosive enamel or dentin surface for the clinical situation.

O-24 Dynamic mechanical analysis of dentin-bonding interface prepared by a self-etching primer

Zhou J1,2*, Tanaka R1, Shibata Y1, Gao P2, Miyazaki T1

1Showa Univ., Tokyo, Japan 2School and Hospital of Stomatology, Tianjin Medical Univ., Tianjin, China

* [email protected] Introduction The longevity of resin-based materials on teeth is entirely associated with the mechanical integrities of a dentin-bonding hybrid interface. The contemporary filling materials have been applied in line with a self-etching primer containing an acid-functionalized monomer such as 10-methacryloxydecyl dihydrogen phosphate (MDP). This study evaluates the nanostructure-function relationship of dentin-bonding interfaces, with and without MDP pre-application by means of Raman microspectroscopy and nanoindentation tests.

Materials and Methods The dentin cavities were treated with or without MDP monomer (SE BOND, CLEARFIL). The bonding agent (SE BOND, CLEARFIL) was subsequently placed in the cavities and was photo-irradiated by a light curing unit (DP-075, Morita). A flowable composite resin (Filteck Supreme Ultra, Flowable Restorative) was filled into the cavities and then photo-irradiated. The degree of conversion ratio and dynamic mechanical properties at the dentin-bonding interfaces on the cross-sectioned samples were assessed by Raman microspectroscopy and nanoindentation, respectively[1]. Five regions at each interface were tested. Data were analyzed by ANOVA followed by a post hoc Tukey test. A p-value less than 0.05 was considered significant.

Results and Discussion Raman spectra revealed that the degree of conversion ratios at the interface regions between control and MDP samples were constant (P >0.05). The average tan δ of the interfaces reduced as a function of frequencies (P <0.05). The MDP treated sample exhibited lower tan δ values than those of control sample (P <0.05). The average storage moduli of MDP sample were higher than those of control sample (P <0.05). Such enhanced stiffness against dynamic strain might well be an important determinant for mechanical integrity of dentin-bonding hybrid layer as it is likely to bare variety of stresses and consequent imposed strains. Although the degree of conversion ratio is a widely accepted concept in assessing polymerization quality of dental composite, solely the function of resin matrix molecular conformation does not always relate the in situ mechanical integrity at the dentin-bonding hybrid interface. Therefore, nanomecahnical testing technique such as nanoindetation is a useful tool for the evaluation of dentin-material interfaces.

References [1] Sato M. et al. Nanoindentation tests to assess polymerization of resin-based luting cement. Dental materials 30:1021-8, 2014.

O-26

Regulation of degradation rate and pattern of dental collagen membrane in vivo

Jian-min HAN1, Hong LIN1＊，Xianchang SUN2, Jiahuan DONG2, Yang Wu1, Youdong 1 1 ZHANG , Meng YANG 1Peking University School and Hospital of Stomatology, Beijing, China 2Yantai Zhenghai Bio-tech Co., Ltd, Shandong, China [email protected] Purpose: The purpose of this study is to regulate the degradation rate and pattern of bovine dermis collagen membrane by adding nature tea-polyphenol in the rat.

Methods: The hair of bovine leather was mechanically removed by hand. The collagen membrane was decellularized by NaOH and TrionX-100, Group A was add tea polyphenol, and group B without tea polyphenol treatment as control group. The surface morphology and inner structure of both collagen membranes were determined by scanning electron morphology (SEM) and frozen section separately. The circular membrane samples with diameter 10mm were placed in the subcutaneous of Wister rat, Rats were sacrificed after 2w, 4w, 12w, 20w, 24w and 36w, and histology was performed. The membrane was stained with HE. The area of collagen and the thickness of the residual membranes were determined by image analysis.

Results: For the degradation rate of both types collagen membrane, the thickness of the residual membrane at various times was show in the following picture. At weeks 2, the degradation of both types collagen membrane can be observed, and the thickness was gradually reduce over the time. However, the control membrane was quickly degraded between 2w and 4w, while the tested group with tea-polyphenol mainly degradation occurred between 4w and 12w. For the degradation pattern, the control group was collapse type degradation, with the surface and inner degradation simultaneous, while the degradation of tested group started from surface, with the inner structure remaining relatively complete.

Conclusion: Tea-polyphenol is a good selection for protecting the collagen membrane structure complete, and regulating the degradation rate and pattern of dental collagen membrane.

O-28

Effect of Ca(OH)2 treated root canal dentin on retention of posts

Someya T*, Harada R, Kinoshita H, Takemoto S, Kawada E Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: Restorations of endodontically treated teeth often require post and core for improved retention of prosthodontic device. The treatment of root canal with irrigators and chemical reagents have shown to reduce the survival rate of the prosthesis (1). Our previous research showed that calcium hydroxide (Ca(OH)2) treatment on root canal dentin adversely affects the bonding strength of core materials depending on the type of resin cements. However, the effect of Ca(OH)2 treatment on the retention of prosthodontic devices has yet to be clarified. The aim of this study was to investigate the effect of Ca(OH)2 treatment on bovine root canal dentin on the retention of prosthodontic devices. Fiber reinforced composite resin post (FRC post) was luted to Ca(OH)2 treated bovine root canal using various adhesive resin cements and post retention was evaluated using the pull-out test.

Materials and Methods: Bovine roots were irrigated with sodium hypochlorite and EDTA, and filled with gutta percha according to endodontical treatment procedure. After the treatment, post space 4 mm in depth were drilled using a 3.0 mm drill and filled with Ca(OH)2 reagent. The specimens were placed in 95 % humidity for 1 week at 37ºC. After 1 week the Ca(OH)2-treated root canals were ultrasonically washed to remove the reagent. FRC post was inserted and the remaining space was filled with resin composite (DC core, Kuraray Noritake Dental). The posts were light cured and luted with Panavia F2.0 (PAF: Kuraray Noritake Dental), RelyX Ultimate Adhesive Resin (ULR: 3M ESPE), RelyX Unicem2 self-adhesive Resin (UNA: 3M ESPE), Clearfil SA luting(CSA: Kuraray Noritake Dental), and G-CEM(GCM: GC). After 1 week of immersion in water at 37ºC, the posts were pulled out from the root canal using a universal testing machine. The post retention force was indicated as the maximum load. After the pull-out test, post specimens were observed under a stereomicroscope. Statistical analysis of post retention force with and without Ca(OH)2 treatment in each adhesive resin cement were performed using the Student’s t test.

Results Fig.1 shows the retention force of post with luting cements. The retention force of PAF and UNA were approximately 200N; that of ULR was 300N; 2 2 that of CSA, and GCM were approximately 150N, respectively. No significant differences between with and without Ca(OH)2 treatment group for all cements were found. Microscope observation revealed that fractures were caused mostly by a cohesive failure in the resin cements. Ca(OH)2 treatment had no effects on retention force of posts. Fig. 1 Retention force

References (1) Nomoto, S., et al., Clin Oral Invest, 2006. 10(4): p. 325-330.

O-29

Fatigue resistance of CAD/CAM crowns restored on premolars

Tsoi JKH1*, Homaei E1,2, and Matinlinna JP1 1The University of Hong Kong, Hong Kong SAR, P.R. China 2 Ferdowsi University of Mashhad, Mashhad, Iran *[email protected]

Introduction: CAD/CAM ceramics become more popular in nowadays dentistry due to the satisfactory aesthetic outcome and good clinical performance. To assess the longevity of the dental crown, one could use fatigue test under various loads in order to simulate various mastication conditions under a controlled manner, and thus logically deduce the clinical outcome. The aim of this study was to investigate and compare the fatigue strength of maxillary premolars restored with two modern CAD/CAM ceramics, lithium disilicate e.max® CAD (LD) and polymer-infiltrated ceramic Enamic® (PIC).

Materials and Methods: Twenty extracted maxillary premolars were selected and prepared for full-coverage restoration (IRB number: UW 15-578). They were randomly divided into two groups, i.e. 10 teeth for each group. Cerec system was used to mill LD and PIC ceramic blocks with a standardized model for the crown restorations. Accordingly, surfaces of ceramic crowns and prepared teeth were etched with HF and phosphoric acid, respectively. The crowns were then restored on the teeth by resin cement. These restored teeth were embedded in acrylic resin and loaded with a 5 mm stainless steel ball-shaped indenter to simulate cyclic chewing (9 Hz) (Instron E3000, USA). The fatigue loading was started with 500 N and increased each 100,000 cycles with a stepwise of 100 N until occurrence of fracture or 1400 N, i.e. 1,000,000 cycles. Kaplan-Meier survival analysis was applied on fatigue loading of each material group and level of significance was set at 0.05.

Results and Discussion: The average fatigue load of LD crowns was 1400 N (904,728 cycles) with just 30% failure rate, whilst 870 N (378,167 cycles) was observed for PIC samples with no survivability. The log-rank test showed that the influence of ceramic crown on fatigue resistance is significant. Most of the failed samples of each group experienced fracture above the CEJ.

Conclusions: The fatigue resistance of LD crowns was significantly higher than PIC restorations. Most of the fractured samples are supposed to be re-restorable.

O-30

Development of a cross platform program to analyze cephalometric radiographs by Python

Kawai T*, Ohno Y, Egashira M, Iwata J, Miyazawa K, and Goto S Aichi-Gakuin Univ., Nagoya, Japan * [email protected]

Introduction: First objective of this experiment is to make a cephalometric analysis program which works on cross platforms, and has flexibility to be changed or to be modified easily by user. Second objective is to code functions to display a result of cephalometric analysis on a polygon diagram immediately when user moves the cephalometric landmarks on a patient X- ray image, and to code functions enables try and error simulation on a cephalometric radiographs. For that purpose, we selected the Python for the software programming language. Materials and Methods: Python was released at around 1990 and progressed rapidly and maturated to script artificial intelligence nowadays. This flexible programming language provides a powerful capability to work on Linux, Windows and Macintosh, therefore, called cross platform programming language. Development environment was as follows. Mac Book Air: 1.7 GHz Intel Core i7 / 8 GB 1600 MHz DDR3 OS X: Yosemite 10.10.5 Python 2.7.10: programming language Text edit (Apple computer) terminal window system(Apple computer) Python launcher Tkinter module was imported to create graphical interface and to produce an event driven source code. Standard patient data were archived in a data module and referred from main program as super class. Main program included 21 methods (functions), and five classes were defined. Results and Discussion: Size of original source code was 541steps. This program works not only on Macintosh but also on Windows and Raspberry Pie (Linux). When an operator moves a triangle point object to each cephalometric landmark, the result of plane angles were calculated immediately and displayed the result on polygon diagram spontaneously. When all cephalometric landmarks were determined, dentist can start simulation to predict better result by means to move patient point objects to clinically good points and can check the result immediately on the polygon diagram. TKinter is a powerful module included in the standard modules of python 2.7.XX, which offers dynamic and flexible event driven program. Python PIL or Open CV are also effective modules for graphical inter face, and especially, Open CV has a potential capability to support automatic recognition for important cephalometric landmarks on X-ray image. However, these module needs pre-install in each computer before a program was launched, and sometimes, it is not easy to include these modules for end users. On the contrary, TKinter is a standard module bundled in the package of Python 2.7.XX and works on almost all platforms without any additional install. Many kinds of AI (artificial intelligence) has been written by Python and getting more easy to use as an open source program, and in near future, this experimental program will be able to mount an AI for automatic diagnostic tool for cephalometric analysis.

O-32

Discoloration of titanium in response to sulfide released by P. gingivalis Harada R1, Kinoshita H1, Takemoto S1, Kawada E1 1Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: A recent literature had reported that discoloration was observed on failed removed dental implants indicating the possibility of titanium corrosion occurring in the oral environment1. Porphyromonas gingivalis (P. gingivalis) is a common culprit of chronic periodontitis that produce large amounts of volatile sulfur compounds (VSCs) as a result of their metabolism. We had recently reported that Na2S alkaline solution causes titanium corrosion indicated through discoloration1. However the effect of sulfides produced by oral bacteria on the corrosion of titanium has yet to be studied. The purpose of this study was to clarify whether sulfides, H2S and mercaptans, produced by P. gingivalis causes titanium discoloration.

Materials and Methods: P. gingivalis strain ATCC 33277 was anaerobically cultured in brain heart infusion broth until stationary phase. Test mediums included the sterile culture medium with no sulfides (BHI), culture medium with P. gingivalis (~1×108 microorganisms/mL) and (BHI-P), and the supernatant layer of the spent culture medium separated from the bacteria by centrifugation (BHI-S). Titanium disks 1.3 mm thick and 13.0 mm in diameter were prepared from wrought titanium rods (Grade 2, Tokyo Titanium) and mirror polished. These were immersed in 3L of each medium in well plates and placed in sealable bags which were closed inside the anaerobic chamber and incubated at 37°C for up to 14 days. The medium was renewed for BHI-P and BHI-S every 3-7 days or 7 days respectively to ensure adequate levels of VSCs. Hydrogen sulfide (H2S) and mercaptan levels were measured using the Gastec Gas Sampling Pump (Zefon International, Inc., Florida, USA) and detector tubes. Titanium corrosion was evaluated through changes in color (ΔE*ab) (MCR-A, Luck Office) and glossiness (Gs(20°)) (GM-26D, Murakami Color Research Laboratory).

Results and Discussion: Over a span of 14 days, H2S and mercaptan levels ranged from 20-70 ppm and 10-50 ppm for BHI-P and 1.5-2.0 ppm and 0.5-5.0 ppm for BHI-S respectively. Neither sulfide products were detected from BHI specimens. The maximum amount of VSC’s detected in this study is 10 to 100 times the amount found in the oral cavity of periodontal patients however, these values seem appropriate since VSCs on implant surface with concentrated biofilms would have high localized levels of VSCs. No significant differences in ΔE*ab and changes in Gs(20°) were observed for any of the specimens placed in the test mediums and values did not change with longer immersion periods. Our past study indicated that titanium discoloration occurs in sulfide alkaline solutions using Na2S, however not in sulfide neutral solutions when the pH was adjusted to 7.5. This is consistent with our current study where discoloration did not occur in a neutral environment. Results indicated that sulfides produced by P. gingivalis and their sulfur metabolic products do not cause titanium discoloration.

References 1) Rodrigues, D. et al. Mater 2013 (6): 5258-5274. 2) Harada, R et al. Mater Sci Eng C 2016(62): 268-273.

O-33

Application of the solution plasma surface modification technology to the formation of thin hydroxyapatite film on titanium implants AkashlynnBadruddozaDithi, Takashi Nezu, Futami Nagano-Takebe, Kazuhiko Endo* Health Sciences University of Hokkaido, Hokkaido, Japan *[email protected]

Introduction: Hydroxyapatite (HA) coatings on titanium implants facilitate rapid bone formation due to their excellent osteoconductive property. The present study was aimed to achieve a thin and uniform HA film coating on titanium implants by a solution plasma surface modification technique.

Materials & Methods: Commercially pure titanium disks (JIS type 2, 14 mm in diameter, 3 mm in thickness) were used. HA film was formed on the titanium disks using three step procedures. First step was to polish the disks with silicon carbide abrasive papers (#240-600, polished-Ti). Second step was to form sodium titanate hydrogel nanofibres by soaking the polished titanium disks in 5 M NaOH solution at 60°C for 24 h (alkaline-treated-Ti). Third step was to apply plasma to the disk surface in mineralizing solution at pH 7.4. A pulse plasma generator (MPP-NV04, Pekuris, Japan.) was used and the titanium disk was placed in 5 mm distance from the electrodes within a glass vial. Plasma was generated at a 5 V, 3 µm pulse width for 30 min. The solution was stirred using a magnetic stirrer (Ion Stir 7d, Central Kagaku Co. Ltd., Japan) at a rotation speed of 240 rpm (sp-Ti). Morphologic and crystallographic features of deposited films were examined by Scanning Electron Microscope (SSX-550, Shimadzu Corporation, Japan) and X-ray diffractometer (JDX-3500, JEOL Ltd., Japan). To evaluate the wettability of the titanium disks before and after surface treatment, contact angle of 20 µl of water droplet on titanium surfaces was measured by contact angle analyzer (Phoenix Alpha, SEO, Republic of Korea) at 0h, 1day, 3days, 7days and 14days after each surface treatment.

Results & Discussion: During solution plasma treatment, a lot of fine crystals were precipitated both in the solution around the two electrodes and on the titanium disk specimens. SEM observation showed that the film composed of fine deposited crystals was formed on both polished and on alkaline treated titanium disk by solution plasma treatment for 30 min in a meta stable calcium phosphate solution. The alkaline treatment contributed to form more uniform film on the titanium disk. XRD demonstrated that crystals precipitated in the solution and on the titanium disk were HA. These results suggested that a thin HA film composed of fine crystals with diameter of approximately 1-5µm were successfully formed on the titanium disk by solution plasma surface modification techniques. The heat generated by plasma increased the temperature of the solution around titanium disk which probably enhanced the HA precipitation. The contact angle of polished-Ti increased with time (θ=37°→51°) indicating that hydrophillic surface became hydrophobic with time. On the contrary, the alkaline- treated-Ti and the sp-Ti remained hydrophilic (θ < 5°) up to 7 days aging. This super hydrophilic characteristic of HA-coated titanium is probably one of the reasons for better biocompatibility than that of titanium.

Conclusion: The present study demonstrated that it is possible to acquire a homogenous precipitation of HA over the entire surface of titanium disk by implementation of solution plasma treatment in mineralizing solution.

O-34

Chitosan/siCkip-1 biofunctionalized titanium implant for improved

osseointegration in the osteoporotic condition

Li Zhang1, Wen Song1, Yumei Zhang1* 1Department of Prosthetic Dentistry, School of Stomatology, The Fourth Military Medical University, Xi’an, China. *e-mail: [email protected]

Introduction: Biofunctionalization with siRNA targeting the key negative modulators of bone turnover involved in the molecular mechanism of osteoporosis, such as casein kinase-2 interacting protein-1 (Ckip-1), may lead to enhanced Ti osseointegration in the osteoporotic condition. In this study, even siRNA loading was accomplished by the thermal alkali (TA) treatment to make the Ti ultrahydrophilic and negatively charged to facilitate the physical adsorption of the positively charged CS/(chitosan)siR complex, designatedas TA-CS/siR. The intracellular uptake of the CS/siR complex and the gene knockdown efficiency were assessed with bone marrow mesenchymal stem cells (MSCs) as well as the green fluorescent protein (GFP) expressing H1299 cells. In vitro osteogenic activity of TA-CS/siCkip-1 targeting Ckip-1 was assessed with MSCs. In vivo osseointegration of TA-CS/siCkip-1 was assessed in the osteoporotic rat model. TA-CS/siR showed excellent siRNA delivery efficiency and gene silencing effect. TA-CS/siCkip-1 significantly improved the in vitro osteogenic differentiation of MSCs, and led to dramatically enhanced in vivo osseointegration in the osteoporostic rat model, showing promising clinical potential for the osteoporotic condition application. TA-CS/siR may constitute a general approach for developing the advanced Ti implants targeting specific molecular mechanism. Materials and Methods: TA-CS/siCkip-1 samples were fabricated via electrostatic interaction. The surface morphology was observed and the surface parameters were assessed. MSCs were used to evaluate the in vitro siRNA delivery efficiency, gene knockdown efficiency and osteogenic differentiation. Osteoporotic rat model, developed by bilateral ovariectomy, was used to test the in vivo osseoingration in osteoporotic condition. Results and Discussion: The novel siRNA biofunctionalized Ti implant TA-CS/siR with an even siRNA loading was successfully developed by the TA treatment to make the Ti ultrahydrophilic and negatively charged to facilitate the physical adsorption of the positively charged CS/siR complex. TA-CS/siR showed excellent siRNA delivery efficiency and gene silencing effect. TA-CS/siCkip-1 significantly improved the in vitro and in vivo osseointegration. TA-CS/siCkip-1 shows promising clinical potential for enhanced implant performance in the osteoporotic bone condition. Furthermore, TA-CS/siR shall provide a general approach for developing advanced Ti implants.

O-35

Design and fabrication orthodontic bracket using Metal Injection Molding (MIM) process

Supriadi S*, Irawan B, and Suharno B, Prasetyadi T, Gratis A, Zulkifly F F Universitas Indonesia, Jakarta, Indonesia *[email protected]

Introduction: Indonesian people has small jaws which is have tendency of impoper teeth arrangement called as maloculation. As increasing GDP of Indonesian people, they start to pay attention on the estetic. One of the technique to repair positioning of the teeth is using bracket orthodontic and shape memory wire. However abillity of Indonesia manufacturer of these orthodontic tool still loe. Bracket ortodontis stil depend on import from other contry. Therefore affordability of this treatment is still low. In order to increase affordability and specific design for Indonesian people in this work describribed or work to design bracket orthodontic for indonesia people and fabrication of bracket orthodontic using Metal Injection Molding (MIM) process

Materials and Methods: In this work, stainless steel 17-4 PH is used which is already mixed with organic binder material to produce feedstock. Targeted bracket design is developed in or previous work. The small injection machine is designed and contructed The feedstock is injected in to a mold cavity to produce a green part. This part still weak because the strength of the part is strength of polymer binder strength. The next process is debinding. In this process, binder material is removed using thermal by placing the green part in to a furnace to produce a brown part. The brown part is sintered up to 1100 oC to get dens part. Geometrical analysis is conducted to osberved optimum processing conditions.

Results and Discussion: Small injection molding machine was contructed for small part metal injection molding process. The machine has capability to extrude and to inject . The extruder is useful for feedstcok making and injection mode is useful for injection material into a cavity. The injection temperature is set between melting temperatur and degradation temperature of the feedstock. Sintering. Sintering process lead shringkage that have to be compansated when design the mold. Therefore the mold should be bigger than the target design. Metal injection molding perform better surface finish than bracket orthodontic that produced using invesment casting.

Conclusions : Design and fabrication of bracket orthodontic using metal injection molding process has been presented. Mold design should considering part shringkage during process. Therefore the mold design shoul be bigger than design. Even the surface roughness better than bracket orthodontic that produced by investment casting however further polishing process still necesarry.

Acknowledgements: Author would like to thanks to Ministri of Research and Higher Education for the reserach grant on PUPT grant 2016 to conduct this research

O-36

Effects of heat treatment on the microstructure and mechanical property of selective laser melted Co-Cr-Mo alloys

Kajima Y1*, Takaichi A1, Kittikundecha N1, Nakamoto T2, Doi H1, Takahashi H1, Nomura N3，Hanawa T1 and Wakabayashi N1 1Tokyo Medical and Dental Univ., Tokyo, Japan 2Technology Research Institute of Osaka Prefecture, Osaka, Japan 3Tohoku Univ., Miyagi, Japan *[email protected]

Introduction: Recently, selective laser melting (SLM), an additive manufacturing process, has been developed for producing components that are difficult to process through conventional casting or subtractive methods. In the SLM process, high temperature gradients lead to high residual stress and rapid cooling rates result in directional grain growth. Consequently, the SLM components show specific oriented microstructures and the anisotropy of mechanical properties. Therefore, post-heat treatment processing is usually necessary for the SLM components to accelerate relief of internal residual stresses and make them homogeneous structures. However the information concerning the microstructural evolution of Co-Cr-Mo (CCM) alloy prepared by SLM during heat treatment is scarce. Thus, the aim of this study was to investigate the effect of post-heat treatment on the microstructure and mechanical properties of CCM alloy prepared by SLM.

Materials and Methods: The CCM powders (MP1, EOS, Krailling, Germany) were used in this study. Cylindrical specimens with a diameter of 6mm were fabricated, with the longitudinal axes rotated by 45° and 90° around the building directions, by an SLM machine equipped with a fiber laser (EOSINT M280, EOS, Krailling, Germany). The SLM machine was operated using the manufacturer’s recommended parameters for MP1 under a nitrogen atmosphere. The specimens were cut and polished into small columnar with1.0mm height. The samples were heat treated at 750 °C, 900 °C and 1050 °C for 2 hours followed by the furnace cooling. The microstructure was investigated by confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Phase identification was performed using an X-ray diffractometer (XRD) with Cu K under 40 kV and 40 mA. The Vickers hardness tests were performed with a load of 4.9N for 15 s.

Results and Discussion: Several molten pool boundaries (MPBs) and numerous fine cellular dendrites were observed in as-SLM alloys. When the alloys were heat treated, the MPBs were disappeared and fine grain boundaries were observed with many precipitates along them. In this heat treatment condition, phase transformation from γ to ε phase occurred only on the surface of the specimen. Although the anisotropy of the hardness decreased after heat treatments, it should be noted that the effect of heat treatment Fig. 1 SEM image of SLM on the hardness was changed according to its temperature, fabricated Co-Cr-Mo alloy 750 °C and 900 °C increased and 1050 °C decreased the heat treated at 1050 °C for hardness, which can be ascribed to the formation of 2 h. precipitates and coarser cellular dendrites respectively.

P-01

Fatigue Resistance of GUMMETAL Clasps Shimamoto K*，Tamaki Y，Wakamatsu N，Iwahori M，Nigauri A，Miyao M，Kajimoto T Asahi Univ. ,Gifu,Japan e-mail;[email protected]

Introduction Materials with a low Young's modulus and a high strength have been developed recently in broad areas including artificial bone and dental materials. A metallic material developed therein is GUMMETAL. GUMMETAL is a β-type titanium alloy with a body-centered cubic structure, and is a metallic material that shows elastic deformability an order of magnitude greater than typical metallic materials. We report the fatigue resistance of a prototype GUMMETAL clasp arm assessed in a constant displacement fatigue test to study the application of GUMMETAL to clasps, supporting devices for partial dentures. Materials and methods Samples were clasps prepared by casting GUMMMETAL in the form and size shown in Figure 1. The number of samples was seven. In the constant displacement fatigue test, samples were secured to face the inner surface of GUMMETAL clasps perpendicularly to the loading rod using a fatigue testing device (clasp fatigue testing device type A, Ito Engineering). The length from the clasp arm base to the clasp tip was 12 mm, and a constant displacement (0.5 mm) corresponding to the amount of undercut was applied 2-mm below the clasp tip (Figure 2). Repeat constant displacements were loaded at 110 cycle/min up to 20000 times corresponding to the use over about five years with an assumed daily number of denture placement/removal of 5 times. The test was halted after every 500 repeat loads to inspect clasp samples with an optical microscope (VHX-200, Keyence) to see if any cracks occurred during the fatigue test.

Figure 1 Size of sample for constant displacement fatigue test Figure 2 Constant displacement fatigue test

Results and Discussion In repeat loads up to 14500 times, no crack or fracture was observed for all samples. However, cracks or fractures were observed for two of seven samples after 15000 loads. In fractured samples, cracks were observed in the clasp arm base after 15500 loads, and then fractures occurred after 16500 loads. In cracked samples, cracks were found in the clasp arm base after 15000 loads, but the cracks did not appear to be extended by 0.5-mm displacement loads up to 20000 times. In this experiment, we prepared a prototype clasp arm with GUMMETAL and assessed its fatigue resistance in the constant displacement fatigue test. When constant displacements corresponding to the 0.5-mm undercut were repeatedly loaded 2-mm below the clasp tip 20000 times corresponding to the about 5-year use with an assumption of denture placement/removal of 5 times a day, cracks or fractures were found in two of seven samples, and these were observed along the air bubble part of the sample surface. Improvements may be possible by processing the GUMMMETAL clasp surface into smooth one in the future. Moreover, even with the cross-sectional shape and thickness of the clasps used in this experiment, the use for about 3 years appeared to be quite possible, and the clasps were suggested to withstand the use for a longer term. Reference 1) Fumiko Takita, et. al., Fatigue resistance of hybrid resin composite/glass fiber reinforced resin composite layered clasps. Ann Jpn Prosthodont Soc, 2009;1:46-54

P-02

Correlation between magnetic susceptibility and phase constitution of Au–Nb alloys for MRI artifact-free biomedical applications.

Inui S*, Uyama E, Honda E, and Hamada K Institute of Biomedical Sciences, Tokushima Univ., Japan *[email protected]

Introduction: Metal devices in human body cause metal artifact in MRI. To reduce the artifact, we have developed Au alloys indicating similar volume magnetic susceptibility (χv) to human tissue, approximately -9 × 10-6 (-9 ppm). A promising candidate is Au–Nb alloys: χv of Au–Nb solid solution with Nb content in mass (CNb) ≤ 12% increases linearly with increasing CNb (3.7 × 1) CNb – 34.0 ppm) and χv value of Au–6Nb alloy is close to -9 ppm . In contrast, χv of Au– 15Nb alloys indicates reduction from Au–12Nb alloy. The phase constitution of Au–15Nb 1) alloy is binary phase of Au–Nb solid solution and Au2Nb (χv = –23.4 ppm) . These facts suggest that χv of Au–Nb alloy is controllable through phase constitution design. In this study, correlation between χv and phase constitution in Au–Nb alloy was investigated quantitatively.

Materials and Methods: χv values of Au–12Nb alloy and Au–15Nb alloy after homogenization at 1273 K for 20 h were 7.8 ppm and 0.9 ppm, respectively1). After aging at 1073 K for 100h, χv values of the alloys reduced and saturated to be –16.6 ppm and –20.4 ppm, respectively. XRD analysis showed phase constitution of both alloy after the aging was binary phase of Au–Nb solid solution and Au2Nb. The saturation of χv values suggested that the deposition of Au2Nb was saturated, and CNb of the solid solution phase (a%) was solid solubility limit at 1073 K. The deposition reaction is; Au–pNb → (1-Mf) × Au–aNb + Mf × Au–19.1Nb (1) where Mf is mass fraction of Au2Nb and 19.1 is CNb of Au2Nb. The material balance is then; p = (1-Mf) × a + Mf × 19.1 (2) Here we assume that χv value of alloy is the a (%) Mf mass average of χv value of constituent phases. Au–12Nb 6.2 0.45 The χv value Au–pNb alloy after aging at 1073 K for 100h is; Au–15Nb 5.6 0.70 (1-Mf) × (3.7 × a – 34.0) + Mf × (–23.4) (3) Table.1 a and Mf of Au–Nb alloys Results and Discussion: From eq. (2) and (3), and the experimental data, a and Mf of Au–12Nb alloy and Au–15Nb alloy were summarized in Table 1, indicating a value of both alloy approximately corresponded. This result confirms the hypothesis that χv value of Au–Nb alloy is the mass average of χv value of constituent phases, and suggests that it can be designed through controlling mass fraction of Au2Nb using heat treatment.

Acknowledgements (Optional): This work was financially supported by JSPS KAKENHI (21500427 and 24500528), Japan Science and Technology Agency, Tanaka Kikinzoku Research Fund and Terumo Life Science Foundation.

References : 1. Inui S., Uyama E. Hamada K., J. J. Dent. Mater. 34:97, 2015.

P-03

The effect of ice-quenching on change in hardness during porcelain firing simulation in a Pd-Au-Zn-In-Sn alloy

Shin HJ, Kwon YH, Seol HJ, Kim HI* BK21 PLUS Project, School of Dentistry, Pusan National University, Yangsan, Korea *[email protected]

Introduction: Degassing treatment in alloys for bonding porcelain is usually conducted at about 1,000 C, where the alloys are possibly softened. Thus, by ice-quenching after degassing, the alloys may remain soft. Thus, if processing for adjustment is performed for the ice-quenched metal substructure after degassing, it will improve the efficiency of the adjustment. Furthermore, the softened alloy by ice-quenching after degassing was expected to be effectively hardened by proper cooling during the rest firing process for bonding porcelain. To examine the effect of ice-quenching on change in hardness during porcelain firing, the change in hardness and microstructure, phase transformation and elemental distribution was examined for a Pd-Au- Zn-In-Sn alloy for bonding porcelain.

Materials and Methods: The specimen used in this study was a Pd-Au-Zn-In-Sn alloy for bonding porcelain (Surefire 10, Aurium research, USA). The specimens were cast using the lost wax casting technique in a standard broken arm centrifugal casting machine (Centrifugal casting machine, Osung, Korea). The as-cast specimens were degassing treated in a dental porcelain furnace (Multimat 2 touch, Dentsply, Germany). Then, various cooling rates were tried after degassing to find out the most effective cooling rate for alloy hardening. Then, for the specimen which was ice-quenched after degassing or cooled at the most effective cooling rate for hardening, the rest porcelain firing process were simulated. At this time, cooling rate during the rest firing process was fixed to the most effective cooling rate for hardening. The changes in hardness during the porcelain firing simulation was examined using a Vickers microhardness tester (MVK-H1, Akashi Co., Japan) with 300 gf load and dwell time of 10 s. The microstructural changes during the porcelain firing simulation were examined by FE- SEM (JSM-6700F, Jeol, Japan). XRD (XPERT-PRO, Philips, Netherlands) profile was recorded at 30 kV and 40 mA using Ni-filtered Cu Kα radiation as the incident beam. The ice- quenched specimen after degassing was porcelain firing simulated. Then, it was polished metallographically and etched in an aqueous solution to examine the elemental distribution using energy dispersive X-ray spectrometer (INCA x-sight, Oxford Instruments Ltd., UK).

Results and Discussion: By ice-quenching after degassing, the specimens were softened due to homogenization without an additional softening heat treatment. The lowered hardness by ice-quenching after degassing was greatly recovered from the first stage of the rest porcelain firing process by cooling rate control. The increase in hardness during cooling after porcelain firing process was resulted from the precipitation of the Pd- and Zn-rich β1 phase of f.c.t structure, which formed severe lattice strains in the interphase boundary between the precipitates of the f.c.t structure and surrounding matrix of the f.c.c structure.

Acknowledgements: This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education, Science and Technology (grant number: NRF-2014R1A1A2055018).

P-04

Effects of support structure on the fatigue strength of selective laser melted Co-Cr-Mo clasps

Takaichi A1*, Kajima Y1, Kittikundecha N1, Nakamoto T2, Nomura N3, Takahashi H1, Hanawa T1, and Wakabayashi N1 1 Tokyo Medical and Dental Univ., Tokyo, Japan 2 Technology Research Institute of Osaka Prefecture., Osaka, Japan 3Tohoku Univ., Miyagi, Japan *[email protected]

Introduction: Selective laser melting (SLM) technology creates 3-dimensional metal parts using laser power, by melting metal powders layer by layer according to the CAD data. This can create complex geometries with undercuts, therefore SLM process could potentially be applied in the preparation of removable partial denture (RPD) alloy frameworks. However, it has been reported that due to the high temperature gradients that occur during the SLM process, something resulting in residual stress that have a significant influence on the fatigue strength. It is known that the support structure is useful to conduct excess heat away from the part and relieve residual stresses trapped inside the part. Therefore, the support structure can be used to improve the fatigue strength. The aim of this study is to investigate the effects of support structure on the fatigue strength of clasps prepared by SLM.

Materials and Methods: Clasp specimens were built using an SLM machine (EOSINT M280, EOS, Krailling, Germany) using commercially available CCM alloy powders (MP1, EOS, Krailling, Germany). To investigate the effects of support structure on the fatigue strength, 2 kinds of specimens (with or without support structure under clasp arm) were prepared (n= 6 for each group). The specimens were fixed to the fatigue-testing machine with screws, and were subjected to a sinusoidal cyclic 0.5 mm deflection generated by the radial direction force at the tip of each clasp arm at 5 Hz. The load/deflection curve was monitored, and the test was terminated when the maximum force was reduced to less than 15% of the initial load. Following the fatigue tests, the fracture surfaces were observed by scanning electron microscopy (SEM). By the use of the preprocessor of the finite element method (FEM) computer software (ANSYS 11.0 FEM; ANSYS Inc, Canonsburg), a 3-dimensional finite element model for each fatigue test clasp was created and thermo-mechanical field analysis was analyzed using ANSYS programme to predict the temperature distribution.

Results and Discussion: SEM images of the fracture surface showed the crack was started near the inner surface of the clasp arms. Specimens with support structure exhibited significantly longer fatigue life than the specimens without it. From the result of FEM analysis, support structure have functions as dissipating heat away from newly melted surface, which can be contributed to decrease the internal residual stress and improve the fatigue strength of SLM CCM clasp. Fig. Changes of permanent deformation for representative specimens from each

group.

P-05

Effect of low concentration of sulfide on corrosion of Ti-Cr alloy

Takemoto S1*, Harada R1 Kinoshita H1, and Kawada E1 1Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: Our previous study demonstrated that the discoloration of commercially pure titanium (TI) in sulfide-containing alkaline solution was affected by sulfide concentration and immersion period1). Yoshida et al. has also showed that 4 types of titanium alloys (titanium, Ti-6Al-4V, Ti-6Al-7Nb, Ni-Ti) undergo discoloration when immersed in sulfide solutions for 3 days2). Ti-20Cr (TCR) has shown to have superior corrosion resistance towards fluoride and peroxide compared to TI, Ti-6Al-4V, and Ti-6Al-7Nb, however corrosion resistance in sulfide has not yet been examined. The objective of this study was to investigate the corrosion behavior of TCR in a low concentration sulfide containing alkaline solution. Materials and Methods: TCR specimens, 13 mm in diameter and 1.1 mm in thickness, were fabricated via casting technique using argon arc melting and casting machines. As a reference, TI (grade 2, Tokyo titanium) specimens were prepared from wrought titanium rods in the same dimensions. All specimens were mirror-polished according to metallographic procedures. Sulfide-containing alkaline solutions were prepared by dissolving 3.12 g of Na2S·9H2O (Wako) in 1 L of ultrapure water (MilliQ®) (MQS-L). TCR and TI specimens were immersed in 20 mL MQS-L and stored at 37ºC for 1, 3, and 7 days. Corrosion was evaluated through changes in color (∆E*ab), measured with a colorimeter (MCR-A, Luck Office), and glossiness, Gs(20º) measured with a glossiness meter (GM-26D, Murakami color research). Results and Discussion: Fig. 1 shows the ∆E*ab for TCR and TI immersed in MQS- L. ∆E*ab values after 7 days of-immersion for TCR and TI were 0.7±0.2 and 10.1±0.6, respectively, and the value of TI increased with increasing immersion periods. ∆E*ab values for TI and TCR immersed in SAL were 0.8 and 0.3, respectively. TCR demonstrated less discoloration compared to TI in sulfide-containing alkaline solution. Gs(20º) of TCR before immersion was similar to that of TI. Fig. 1 Color changes of TCR and TI After 7-days of immersion in MQS-L, Gs(20º) of TI decreased immersed in sulfide-containing by 20% compared to before immersion, while the value for solutions. TCR was similar to that before immersion. Our previous study suggested that TI was oxidized in MQS-L, shown through an increase in thickness of the surface oxide layer1), which was also indicated to be the cause of discoloration. TCR has shown to have superior resistance to fluoride which was dependent on the ratio of chromium to titanium oxides in the passive films4). The similar rationale in relation to the compositional differences of the passive films formed on TCR and TI may be used to explain the differences in corrosion resistance and smaller discoloration of TCR. References: [1] Harada R, et al. Mater Sci Eng C 2016; 62: 268-273., [2] Yoshida T, et al. J Jpn Soc Dent Products 2009; 23: 10-19., [3] Noguchi T, et al. Dent Mater J 2008; 27: 117-123., [4] Takemoto et. al., Dent Mater 2009; 25: 467-472.

P-06

Relationship between Microstructure and Mechanical Strength of Newly Developed Ag-Pd-Cu-Au System Alloy Subjected to Solution Treatment

Akahori T1*, Fukui H2, and Niinomi M1 1Meijo Univ., Nagoya, Japan 2Aichi-gakuin Univ., Nagoya, Japan *[email protected]

Introduction: Silver alloy is one of semi-precious alloys for dental prosthesis, which has been applied remarkably in Japanese dental field. Ag-20Pd-17.7Cu-12Au alloy (G12) with Cu/Ag ratio of 0.367, which was newly developed for commercial dental silver alloy, shows an unique hardening mechanism after a simple solution treatment (ST) at relatively high temperature as well as traditionally applied Ag-20Pd-14.5Cu-12Au alloy (S12) with Cu/Ag ratio of 0.284. However, the relationship between ST at various temperatures and the mechanical strength of G12 has not been investigated fully. Therefore, the relationship between microstructure and mechanical strength of G12 after ST at various temperatures was investigated systematically in this study.

Materials and Methods: Materials used in this study were hot-rolled plates of Ag-20Pd-17.7Cu-12Au alloy (G12) and Ag-20Pd-14.5Cu-12Au alloy (S12). All samples were conducted with the solution treatment (ST) at 1023~1173 K for 3.6 ks in an Ar gas atmosphere followed by water quenching (WQ). Hereafter, the samples of G12 and S12 subjected to ST were designated by using temperature. (e.g. G12/1123 K). In addition, some samples subjected to aging treatment (AT) at 673 K for 1.2 ks in a vacuum followed by air cooling after ST (G12/AT673 K). Back-scattering electron (BSE) images from a scanning electron microscope (SEM) and elemental mapping by a wavelength dispersive X-rays spectrometer (WDS) were used for the observation of microstructures and the identification of phase constitutions. The Vickers hardness (HV) and tensile tests were performed to evaluate the mechanical strength. The corrosion potential was measured in a 3 mass% NaCl solution to evaluate the corrosion resistance.

Results and Discussion: G12/1173 K was composed of three kinds of phases with Ag-rich α2 phase, Cu-rich α1 phase and β phase of Pd-Cu intermetallic compound. Although nominal melting point of G12 was around 1233 K, the high Cu concentration area like α1 phase was partially dissolved and then the α1 and β phases re-precipitated during cooling. On the other hand, the microstructure of the G12/AT673 K was composed of two kinds of phases with α2 and β phases, which is a typical microstructure of AT after ST. The corrosion potential of G12 changed drastically with the temperature of ST. The potential may have been increased by the decrease in the volume fraction of the α1 phase with relatively high Cu content. The Vickers hardness of G12/1173 K and G12/AT673 K were almost the same value each other. It is considered that the Vickers hardness increased remarkably by the precipitation of meta-stable β’ phases of the coherency in α2 phases. The Vickers hardness of G12 subjected to ST at relatively high temperature of 1173 K was identical to that of the same alloy subjected to typical age-hardening.

P-07

In vitro evaluation of H2O2 hydrothermal treatment of aged titanium surface to enhance biofunctional activity

Yoneyama Y1*, Matsuno T2, Hashimoto Y3 and Chikazu D1

1 Dept. of Oral and Maxillofacial Surgery, Tokyo Medical Univ., Tokyo, Japan. 2 Dept. of Oral and Maxillofacial Surgery, Nippon Dental Univ., Tokyo, Japan. 3 Dept. of Biomaterials ,Osaka Dental Univ., Osaka, Japan. *e-mail ([email protected])

Introduction: Surface modification of titanium has been extensively investigated in implant science and technology in an effort to improve its osteoconductivity. The rate of protein adsorption on titanium surfaces is known to vary depending on the chemistry, structure, morphology, and titanium-specific biological aging of the surface. In this study, we have developed a simple surface modification of titanium alloy that improves its biofunctional activity. The surface of a Ti-6Al-4V disk was modified by applying 3% H2O2 hydrothermal treatment using an autoclave. A nanostructured porous network TiO2 was observed on the treated surface. Treated surfaces exhibited higher hydrophilicity, protein adsorption, and cell proliferation than untreated surfaces. 3% H2O2 hydrothermal treatment is thought to provide biofunctional activity for aged titanium surface.

Materials and Methods: Ti-6Al-4V disks with diameters of 15 mm and thicknesses of 1 mm were used in this study. After ultrasonic cleaning, all disks were stored at room temperature in the dark place for 1 month. One month after, the half disks were randomly selected as the control group. The remaining disks were hydrothermally treated with 3% H2O2 solution and autoclaved at 121°C and 0.2 MPa for 20 min. The treated disks were used as the test group. Surfaces were analyzed by scanning electron microscopy （SEM）, scanning probe microscopy（SPM）, X-ray photoelectron spectroscopy（XPS）and contact angle measurements. Bioactivities were analyzed by protein adsorption, MC3T3-E1 cell proliferation on Ti-6Al-4V disks and cell adhesion was observed with a fluorescent immunostaining.

Results and Discussion: The surface of the control disk exhibited a smooth line pattern, whereas the test disk exhibited a relatively irregular surface with a porous network structure. The surface roughnesses (Ra) of the control and test groups were measured by SPM to be 14.132 and 22.315 nm, respectively. Surface composition was coated with a TiO2 film having a thickness of 90nm. Further, the titanium surface by the process significantly increases the hydrophobicity, even decrease over time in the biological activity by aging is suppressed. Protein adsorption and cell proliferation compared to the pre-treatment was significantly increased. Moreover, this treatment reduces the degradation of the hydrophilicity and protein adsorption of titanium surfaces due to aging after surface treatment. Therefore, the 3% H2O2 hydrothermal treatment is thought to provide biofunctional activity to aged titanium surfaces.

P-08

NaOCl-treatment of titanium implant enhances cytocompatibility

Ichioka Y, Kado T, Kanada K, Endo K and Furuichi Y

Health Sciences University of Hokkaido., Hokkaido, Japan *e-mail [email protected]

Introduction: It is well known that the topological and physicochemical properties of Ti implant surfaces affect the osteoconductive capability of implant materials, on the other hands, osteocompatibility of Ti implant degrades over time after surface processing. The aim of this study was to examine whether the chemical conditioning of titanium with NaOCl solution would effectively decompose surface contaminants and enhance cytocompatibility.

Materials and Methods: Commercially pure titanium disk (Ti) was used as a substrate. After polishing, the specimens were cleaned ultrasonically in distilled water for 5 min (As-polished Ti) and then stored under a dark ambient condition for 14 days (2-week-old Ti). The surface contaminants of the aged Ti disks were decomposed by chemical treatment in 5% NaOCl solution for 24 h (NaOCl-treated Ti). The disk immersed in distilled water for 24 h was employed as a control (H2O-treated Ti). The surface characterizations were examined by X-ray photoelectron spectroscopy (XPS). The wetability of the Ti surfaces was examined by measuring the contact angle of 4 μl water droplet using a contact angle analyzer. Initial cell attachment of human bone marrow mesenchymal stem cell on each Ti was evaluated by hemacytometry and scanning electron microscopy (SEM). Confocal laser scanning microscopy was used to examine cell morphology and cytoskeletal arrangement. The area and feret’s diameter of the cells were quantified using an image analyzer (Image J). To confirm adsorption of collagen I, samples were incubated with collagen I-FITC conjugate for 1 h. After washes, samples were observed by confocal laser scanning microscopy. The amounts of adsorbed collagen I was evaluated by measuring the fluorescence intensity of 5 randomly chosen images from three specimens of each groups using Image J.

Results and Discussion: The change of surface characterization of 2-week-old Ti from fresh surface, the level of surface hydrocarbons, especially carboxyl group, and nitrogen were increased. After immersion in NaOCl solution, these contaminants were removed. Furthermore, the amount of hydroxyl group was increased. The surface of NaOCl-treated Ti and H2O-treated Ti substantially was converted to the superhydrophilic surface (< 5˚). The number of attached cells to the NaOCl-treated Ti and H2O-treated Ti was about 2 times as large as that attached to the 2-week-old Ti. These results were consistent with observations under SEM in terms of cell density on the titanium surfaces examined. Cytomorphometric evaluations for the area and Feret’s diameter of the cells showed greater values of these parameters on the NaOCl-treated Ti surface than on the as- polished surfaces. The amounts of adsorbed collagen I on the NaOCl-treated Ti was significantly higher than that on the others.

Conclusions: Bio-functionalization of the Ti surface can be achieved by treatment with 5% NaOCl.

P-09

Monitoring of protein adsorption and apatite precipitation on titanium and zirconia by QCM method

Yoshida E* and Hayakawa T Tsurumi Univ., Kanagawa, Japan *e-mail: [email protected]

Introduction: In recent years, metal-free dental implants using zirconia has been attached. Regarding the osseointegration of zirconia implant compared to titanium implant is still controversy. In the present study, protein adsorption and apatite precipitation in simulated body fluid (SBF) were monitored by using 27M-Hz quartz crystal microbalance (QCM) method as a basic research for biocompatibility of titanium and zirconia. QCM method can detect the mass change with nano-level by measuring the frequency shift. In the present study we aimed to analyze the adsorption behavior of fibronectin (FN) and albumin (BSA) to titanium (Ti) or zirconia (ZrO2) sensor and the precipitation behavior of apatite on titanium or zirconia sensor in the SBF.

Materials and Methods: QCM apparatus: High sensitivity QCM device with the frequency of 27MHz (AFFINIX QNµ, ULVAC, Japan) was used. Ti and ZrO2 sensor were prepared by the sputter coating on an Au electrode. A frequency decrease of 1 Hz corresponds to 0.61 ± 0.1 ng cm-2 adsorption on the sensor in a 27 MHz QCM system. The amount of protein adsorption or apatite precipitation was calculated by Sauerbrey’s equation. Protein adsorption experiment: The 0.5mg/ml concentration of FN/PBS and BSA/PBS solution was prepared. First, 500 µL of PBS (pH=7.4) was injected into the QCM cell equipped with sensor. After stabilization of the cell, a 50 µl of FN/PBS or BSA/PBS solution was injected to the PBS solution in the cell. Frequency shift was measured for 30 minutes Apatite precipitation experiment: The Hank’s balanced solution (pH=7.4) was used as SBF. 500 µL of Hank’s balanced solution was injected into the QCM cell and the frequency shift was measured for 24 hours.

Results and Discussion: Protein adsorption experiment: Adsorbed Table 1 Protein adsorption experiment amounts of FN and BSA to each sensor were Ti ZrO2 show in table 1. Adsorbed amounts of FN FN (ng/cm2) 3.35 (0.42) 2.11 (0.09) and BSA on Ti was significantly higher than BSA (ng/cm2) 1.72 (0.29) 0.99 (0.08) those on ZrO2 (p<0.05). Apatite precipitation experiment: Apatite precipitated amount and initiation time of apatite precipitation were listed in Table 2. Precipitated amounts of apatite were not significantly deference between Ti and ZrO2 (p>0.05). However, initiation time of apatite precipitation on Ti was significantly Table 2 Apatite precipitation experiment shorter than that on Ti ZrO2 ZrO2 p<0.05). 2 In conclusion, Precipitated amount (µg/mm ) 0.30 (0.42) 0.33(0.02) the results of present Initiation time of apatite recipitation (min) 52.0 (31.7) 134.3 (5.9) study revealed that Ti and ZrO2 showed different behavior for protein adsorption and apatite precipitation in SBF.

P-10

Application of porous titanium in tailored post-core systems using a moldless process: evaluation of the physical and mechanical properties with different particle sizes

Prananingrum W1, 3*, Naito Y2, Sekine K1, Hamada K1, Ichikawa T1 1Tokushima University Graduate School, Tokushima, Japan 2Tokushima University Hospital, Tokushima, Japan 3Hang Tuah University, Surabaya, Indonesia *e-mail : [email protected]

Introduction: Cast titanium post-core systems have been successfully utilized because of their superior mechanical properties. However, their higher elastic modulus can result in stress concentration in the surrounding dentin, resulting in root fracture. One of the methods to overcome this issue is introducing porosity to the material. Our research group developed a production method of porous titanium from titanium particles using moldless process on behalf of the biomaterial application. This study aimed to reveal correlation between titanium particles size and physical and mechanical properties of porous titanium, and to control the properties through controlling titanium particle size and its distribution.

Materials and Methods: Commercially pure Ti powders with different particle sizes, group 1: < 45 m (Nilaco Co., Tokyo, Japan) and group 2: <150 m (Osaka Titanium Technologies Co., Tokyo, Japan) were used. A 90:10 wt% mixture of Ti powder and wax binder was prepared manually to make specimens at 70C. After debindering at 380C, the specimen was sintered in Ar at 1100C without mold for 1h. Particle size distribution, linear shrinkage ratio, porosity, bending strength, and elastic modulus were evaluated. Specimens after sintering were observed by SEM.

Results and Discussion: Group 1 presented a narrow particle size distribution with the median particle size 8.14 μm. In contrast, a broad particle size distribution presented by Group 2 with the median particle size 63.8 μm. The elastic modulus (85 GPa to 18 GPa) and bending strength (428 MPa to 106 MPa) decreased with increasing particle size; the linear shrinkage ratio (14% to 2%) also decreased, whereas the porosity (17% to 38%) increased. The lower shrinkage of Group 2 is appropriate for clinical use. Moreover, Group 2 showed significantly lower elastic modulus, similar to the dentin and presented significantly lower bending strength, close to coronal dentin. The higher bending strength of Group 1 is close to the root dentin, however, the elastic modulus is disappointingly higher than that of dentin. SEM images indicated the necking among titanium powder for each particle size.

Conclusions: The physical and mechanical properties of porous titanium using a moldless process were dependent on the particle size of titanium powders. Titanium powders smaller than 150 m diameter offer major advantages for the fabrication of tailored post-cored system. A modless process can be applied to fabricate the tailored post-core systems with low elastic modulus and low shrinkage by optimizing material design.

P-11

Molecular orientation and surface properties of gold-deposited titanium following alkanethiol immersion treatment

Saitoh S, Sasaki K, Taira M, and Hattori M Iwate Medical Univ., Iwate, Japan * [email protected]

Introduction: Methods utilizing the generation of organic molecular membranes have been applied to improve material surface characteristics in a wide range of fields. Previously, we reported that gold vapor deposition on a Ti surface effectively forms an alkanethiol layer having protein- binding functionality. In this study, we examined the relationship of the coating state of vapor-deposited Au with alkanethiol absorption, looking at changes in physical surface properties and electrochemical properties.

Materials and Methods: Sheets of Grade 2 commercially pure Ti were used as the substrate in the experiments. Sheets were subjected to ultrasonic cleaning in acetone, 30% nitric acid, and then distilled water for 10 min each. After drying, sheets were subjected to gold vapor deposition in an Ar gas atmosphere by means of a magnetron sputtering device for 0, 5, 10, 20, 50, 100, or 200 s. Following gold deposition, Ti sheets were immersed in an ethanol solution of an alkanethiol (12-mercaptododecanoic acid: HSCH2(CH2)9CH2COOH) for 2 h. Specimens were then cleaned in ethanol, distilled water, and finally dried before use in the experiments. Static contact angle was measured to evaluate the surface characteristics of the substrate after gold deposition and after alkanethiol treatment. In addition, cyclic voltammetry (CV) measurements were performed to evaluate the electrochemical properties of gold-deposited Ti following alkanethiol treatment.

Results and Discussion: The contact angle of gold-deposited substrates was found to hardly change when Au coverage was low, but to decrease with higher coverage, suggesting that increased Au content contributes to improved wettability. The elemental concentration of S in alkanethiol-treated substrates, on the other hand, increased with increasing Au coverage, suggesting binding of alkanethiol molecules. Elemental S concentration converged at >80% Au coverage. As an explanation of this finding, we suppose that coverage initially proceeded on individual islands, and increased deposition time allowed the extent of deposition to expand and the deposition layer to grow thicker. Contact angle gradually decreased with increasing Au coverage; wettability rose exponentially once coverage exceeded 80%. In the cyclic voltammograms obtained from CV measurements, a peak was detected in the vicinity of –1.0 V when voltage was swept in the cathode direction. With longer deposition times of 50, 100, and 200 s (Au coverages of 86%, 96%, and 98%, respectively), peak area increased and exhibited a sharper shape. This peak derives from the reductive desorption of thiol molecules that had been adsorbed to the surface by alkanethiol treatment. The increase in peak area thus demonstrates high amounts of adsorbed molecules. At deposition times of 20 s (55% Au coverage) and lower, on the other hand, peaks were indistinct and broadly shaped. These results suggest that while alkanethiol molecules bind loosely to substrates having low Au coverage, they bind more tightly with increasing Au coverage, and that their outward- facing, terminal COOH group causes them to exhibit hydrophilicity.

P-12

Construction and biological evaluation of multi-factor-functionalized coating on titanium implant

Luo QJ1, Wang Y2, Zha GY2, Chen YD2, Li XD2* 1The First Affiliated Hospital, Zhejiang Univ., Hangzhou, China 2The Affiliated Stomatology Hospital, Zhejiang Univ., Hangzhou, China *e-mail [email protected]

Introduction: Osseointegration was defined histologically as the direct bone-implant contact without an intervening fibrous connective tissue interface and clinically as the rigid fixation of an implant in bone allowing the functional loading of dental implants. Biochemical surface modification was referred to the modification of an implant surface with signal generating molecules, and regarded as a promising strategy to make implant surfaces with more osteogenic potential. As the advancement in the field of molecular mechanism of bone healing, arginine-glycine-aspartic acid (RGD), basic fibroblast growth factor (bFGF), and bone morphogenetic proteins (BMP) were confirmed to be important signal molecules for bone regeneration. Previous studies have reported that modifying titanium surface with one or two of these bioactive molecules could obtain positive effects on osseointegration [1, 2]. In the present study, we functionalized implant surfaces with all these three biomolecules to find out if it can achieve a synergic effect. Materials and Methods: Commercially pure Titanium (Ti) were polished and treated with a mixture of sulfuric acid and hydrogen peroxide for further modification. Four groups were prepared: Col/HA- RGD group, group modified with collagen/ HA-RGD polyelectrolyte multilayer coating (PEM); Col-BMP2/ HA-RGD group; Col-bFGF/ HA-RGD group and Col-BMP2-bFGF/ HA- RGD group. MC3T3-E1 cells were used to evaluate the osteogenic properties of these PEM- coating modified Ti in terms of cell proliferation, ALP activity and OC production. Results and Discussion: DNA quantification demonstrated that after cultured for three days, the Col-BMP2-bFGF/ HA-RGD group had a promoted cell proliferation when compared with the other three groups (p<0.05). Cell differentiation was also significantly improved on the Col-BMP2-bFGF/ HA- RGD group, characterized by the highest ALP activity and OC level (Fig. 1, p<0.05). These results suggested that, the combination of three bioactive agents in the way presented in the study could achieve a synergetic effect on Ti osteogenesis.

Figure 1. Cell proliferation (A) and differentiation characterization (B,C). References: [1] Huang Y, Luo QJ, Zha GY, et al. Biomater. Sci., 2014, 2, 980 [2] Luo QJ, Huang Y, Deng XL, et al. Nanomedicine , 2013, 8(5), 739 Acknowledgement: This study was financially supported by the National Natural Science Foundation of China (51173163) and the Natural Science Foundation of Zhejiang Province (LZ16H140001).

P-13

Interactions between the nano-micro-hierarchical titanium and platelet

Chen YD1, Luo QJ2, Huang Y3, Li XD1* 1 The Affiliated Stomatology Hospital, Zhejiang Univ., Hangzhou, China 2 The First Affiliated Hospital, Zhejiang Univ., Hangzhou, China 3Peking University School and Hospital of Stomatology, Beijing, China *e-mail [email protected]

Introduction: The majority of studies in the field of titanium implant modification were focused on the response of osteoblastic cell lineage. However, the process of osseointegration in vivo is not only related to implant-osteoblast interactions. Osseointegration, similarly to bone healing, was achieved by a temporal sequence, including haemostasis, inflammatory, proliferative and remodelling. Each plays a unique role and has a significant impact on the subsequent phase as well as the final osseointegration. Before the attachment of osteoblastic cells, other types of cells adhere to the surface of implant and response to the surface properties. The platelets, which contained within blood and play pivotal role in the processes of haemostasis and blood coagulation, are among these other cells. Activated platelets release kinds of cytokines and influence at least two phases of osseointegraton. In the present study, we focused on the effect of the nano-micro-hierarchical surface topography on the activation of platelets. Materials and Methods: Commercially pury titanium (Ti) discs (30 mm in diameter, 1 mm in thickness) were used and divided into two groups according to the preparation. The Smooth Ti group: Ti discs were sequentially polished with SiC paper from 320 # to final 2400 #. The nano-micro-hierarchical Ti group: Ti discs were polished, sandblasted, acid-etched with the mixture of H2SO4 and HCl, and bathed with a mixture H2SO4 and H2O2. Platelet rich plasma, prepared from sodium citrate anticoagulant human blood, was added to each sample and cultured for 30 min at 37°C. Platelet attachment and β-thromboglobulin were detected. Results and Discussion: CCK-8 assay showed that, there were more platelets adhered on the Nano-micro-hierarchical Ti group. Similarly, more β-TG was detected in the Nano-micro-hierarchical Ti group. These results indicated that the nano-micro-hierarchical surface topography could act as a stimulator of platelet activation, and this property might contribute to its osseointegration potential.

Figure 1. Activity of platelets on different surface topographies. Acknowledgement: This study was financially supported by the National Natural Science Foundation of China (51173163) and the Natural Science Foundation of Zhejiang Province (LZ16H140001).

P-14

Cortical Bone Response of MDF Titanium Implant

Suzuki H1*, Hirota M1, Ohkubo C1, Hoshi N2, Kimoto K2, Miuar H3 and Hayakawa T1 1Tsurumi Univ., Kanagawa, Japan 2Kanagawa Dental Univ., Kanagawa, Japan 3Toyohashi University of Technology, Aichi, Japan *[email protected]

Introduction: Titanium and titanium alloy have been widely used for dental implants due to their excellent biocompatibility. It is well known that titanium alloys have better mechanical properties than to pure titanium. Some tried to improve the mechanical properties of pure titanium. Multi-directional forging (MDF) method is one of the useful techniques for improving the mechanical properties of metals and alloys. It is reported that MDF treatment produced significant increase of pure titanium1). In the present study, we aimed to evaluate the cortical bone response of MDF titanium implant by the implantation experiments into the rabbit tibiae.

Materials and Methods: Multi-directionally forged titanium (MDF-Ti, Kawamoto Heavy Industries Co., Ltd) cylinder implants and commercial pure titanium (Ti, Furuuchi Chemical Corp.,) cylinder implants with 3.5 mm in diameter and 7.0 mm in height were used. Half of Ti and MDF-Ti were sandblasted. As an animal, Japanese white rabbit was used (Approval Number. 26A002). The implants were implanted into bone defects of rabbit tibiae. Fluorochrome labeling was performed at 11 weeks after the implantation by calcein injection. At 12 weeks postsurgery, the implants were excised and non-decalcified thin sections were prepared after formalin fixation, dehydration with graded series of ethanol and embedding with methyl methacrylate. After staining with toluidine blue, bone response was histologically and histomorphometrically evaluated.

Results and Discussion: SEM observation showed that sandblasting to MDF-Ti and Ti did not cause any distinct differences. During animal experiments, rabbit remained in good health. No significant difference was observed in length of fluorescence labeling by calcein between Ti-Sandblast and MDF-Ti- Sandblast implants (p>0.05). After 12 weeks of implantation, new bone formation was observed around four different implants. There were no significant differences in bone-to- implant contact ratio (BIC) and bone mass (BM) between Ti-As and MDF-As (p>0.05). BICs between Ti-Sandblast and MDF-Ti-Sandblast were not also significantly different (p>0.05). BICs were not significantly increased by sandblasting (p>0.05), but BMs were increased by sandblasting (p<0.05). In conclusion, it revealed that MDF-Ti showed a cortical bone response same as conventional commercial pure titanium. MDF-Ti will be a candidate for dental implant material instead of titanium alloys.

Acknowledgements (Optional): We are grateful to Kawamoto Heavy Industries Co., Ltd. for supplying MDF titanium.

References 1) Hoshi et al., J J Dent Mater 2013; 32:403.

P-15

Nanosecond-Pulsed Laser-Treated Titanium Implants Enhanced Bone and Gingival Tissue attachment

1* 1 2 3 4 2 Fukayo Y , Horie A , Amemiya T , Komotori J , Mizutani M , Hamada Y , and Hayakawa T2 1Kanto Rosai Hospital, Kanagawa, Japan, 2Tsurumi Univ., Kanagawa, Japan 3Keio Univ., Kanagawa, Japan, 4Tohoku Univ., Sendai, Japan *e-mail:[email protected]

Introduction: Nowadays attachment of gingival connective tissue to titanium implants is insufficient, resulting in soft-tissue inflammations. Hence, implant surface modification should be needed for attaining a tight bonding to connective tissue as well as to bone. We focused on a laser processing procedure with defocus technique for controlling surface topography and chemistry. The aim of this study was to evaluate bone and gingival connective tissue responses towards nanosecond-pulsed laser-treated titanium implants.

Materials and Methods: A Nd:YVO4 nanosecond-pulse laser with a defocus technique was used to modify the surfaces of cylindrical titanium implants. Two types of cylindrical implants were employed. One had a 3.5 mm diameter and 7.0 mm length (3.5 Ti) to assess rabbit bone responses, and the other a 1.0 mm diameter and 4.5 mm length (1.0 Ti) to assess rat gingival connective tissue responses. Laser-treated titanium implants, a 3.5 Laser-Ti and 1.0 Laser-Ti, were obtained. Collagen immobilized 1.0Laser-Ti (1.0 Coll/Laser-Ti) implants were obtained by a tresyl chloride-activated method. The surface morphologies were observed by SEM. The surface roughness and surface elemental analysis were performed. In animal experiment, 3.5 implants were implanted into the bone defects of rabbits to evaluate bone responses and 1.0 implants were implanted into the extracted sockets of rat maxilla to evaluate gingival connective tissue responses. After implantation periods, the specimens were excised and nondecalcified thin sections were prepared to evaluate histological responses. The percentage of BIC and bone mass (BM) around the implant were determined using an image analysis system.

Results and Discussion: Laser-Ti surfaces had micro-scale roughened oxide layers and parallel arranged grooves. Sa and Sdr values of the Laser-Ti were significantly higher than those of Ti implants (p<0.05). In the XPS analysis, the atomic oxygen content of the Laser-Ti implant was higher than that of the Ti implant. After 12 weeks of implantation in the rabbit experiments, bone-to-implant contact for the Laser-Ti implants was significantly higher than for the Ti in both tibia and femoral condyle (p<0.05). Improved attachment of gingival connective tissue to the implant surface was observed for Laser-Ti and Coll/Laser-Ti in the rat maxilla. Polarized light microscopy showed perpendicular rod-like attachments of gingival collagen fibers on the Laser-Ti and Coll/Laser-Ti implant surfaces. Ti implants had no discernible attachments with gingival connective tissue along the implant surface. A roughened titanium surface with parallel grooves and micro-roughened asperities produced by nanosecond-pulsed laser treatment induced better bone responses and enhanced attachment of gingival connective tissue.

P-17

A Study on Wear Resistance of Flowable Composite Resin Material

Teramae M*, Takahashi S, Uchida J, Fujimura H, and Nakatsuka T Research and Development Dept., Shofu Inc., Kyoto, Japan *[email protected]

Introduction: Indirect resin composite materials clinically used in Japan are mainly regular, non-flow paste products, whereas in Europe flowable paste products are predominantly used. Due to its lower filler content, flowable paste is generally considered to have weaker physical properties than regular non-flow paste. Wear properties of either type of paste, however, have not yet been fully elucidated. In this study, we performed reciprocating wear testing on these two types of composite resin materials and compared their wear resistance.

Materials and Methods: Indirect resin composite materials used in this study were two regular paste products (CE: CERAMAGE, Shofu; ES: Estenia, Kuraray Noritake) and two flowable paste products (CU: CERAMAGE UP, Shofu; SI: Sinfony, 3M ESPE), a total of four products. Each composite resin material was filled in a mold (15×20×2.5mm) and light- (or heat-)cured according to the manufacturers’ instructions. The surface of the cured materials was polished with waterproof abrasive paper (#600, #1200), and buff-finished to obtain wear test specimens. The test specimens were subjected to reciprocating wear testing (testing machine: TriboGear, type: 14FW) at a load of 1000g with 5mm-distance of linear reciprocation for 1000 times using SUJ2 steel antagonist (diameter: 5mm). After testing, the maximum wear depth (µm) on the surface of the specimens and wear volume (µm3) were measured using a contact surface roughness tester (SURFCOM1500, ACCRETECH) and a laser scanning microscope (LEXT OLSD4100, OLYMPUS). Data obtained were statistically analyzed by Tukey’s test.

Results and Discussion: 1. CU exhibited the lowest maximum wear depth (CU

P-18

Discoloration and microbial adhesion to composite resin artificial denture teeth containing fluoropolymers and nano silica filler Oike K, Fujii K, Kusakabe S, Kotake H, Tamaki Y, Hotta M* 1Asahi Univ., Mizuho, Japan *e-mail([email protected])

Introduction: Composite denture teeth are frequently used for removable protheses, however, it has been observed that composite denture teeth suffer from unexpected discoloration. The objective of this study was to determine the discoloration, microbial adherence, surface characteristics, mechanical properties of a new composite denture teeth contaiinig fluoropolymers and nano silica filler.

Materials and Methods: We experimentally produced 2 types of composite resin artificial teeth for dentures. In one type, aliphatic monomers containing fluorine were used as the matrix resin of composite resin, and micro-fillers were added as inorganic fillers. In the other type, aliphatic and aromatic monomers containing fluorine were used as the matrix resin, and nano-fillers were added as inorganic fillers. A acrylic resin was also examined as control. Thereafter, before and after alkaline degradation of the 2 types of artificial tooth materials was performed to evaluate the three-point bending strength, Vickers hardness, surface roughness(Ra), specular gloss, water contact angle, SEM observation, coloring(0.04wt% basic fuchsin), and microbial adhesion(Streptococus oralis and Candida albicans). All data were statistically analyzed with ANOVA and Scheffe’s test(α=0.05).

Results and Discussion As a result of alkaline degradation, although the mechanical strength of the 2 types of experimentally produced composite resin artificial teeth decreased, the strength was more than 80% of that before alkaline degradation, and the hydrophobicity increased. Furthermore, the surface roughness (Ra) of the composite resin artificial tooth material containing fluorine aliphatic monomers as the matrix resin and inorganic micro-fillers of 1-15 μm increased to more than 0.3 μm, and coloring and adhesion of S. oralis increased because gaps and dislodgement of fillers from the matrix occurred due to alkaline degradation, whereas Ra of the nano-composite resin artificial tooth material containing fluorine aliphatic and aromatic monomers as the matrix resin and ultra-fine fillers of nano-size (40 nm) was lower than 0.2 μm even if the fillers dislodged due to alkaline degradation, and coloring and adhesion of S. oralis were suppressed. Therefore, it is suggested that the experimentally produced composite resin artificial tooth material containing fluorine aliphatic and aromatic monomers and nano-fillers shows excellent long-term resistance and anti-contamination properties, and may suppress the adhesion of denture plaque on the surface of artificial denture teeth.

P-19

Fluidity quantification of flowable resin composites through measurement of yield stress using viscometer

Asakura M*, Hayashi T, Mieki A, Aimu K and Kawai T Aichi Gakuin Univ., Aichi, Japan *[email protected]

Introduction: Selection of flowable resin composites with appropriate flow properties for specific clinical cases is a crucial part of successful dental restorations. For placement of these materials, flow properties are particularly important. Flowable resin composites have found a wide range of clinical applications and consequently have a wide variety of flow properties. However, because there is no standard test method for evaluating the flow properties of these materials, manufacturers have been unable to conduct absolute assessments of their products, and instead make relative comparisons of their own products. The purpose of this study is to quantify their fluidity, the yield stress of several resin composites prior to setting were measured using a simple cone-and-plate rotational viscometer.

Materials and Methods: Five commercially available composites (AliteFlo LV: Bisco, Flow-it ALC: Jeneric/Pentron, Venus flow: Heraeus-Kulzer, Tetric N-flow: Ivoclar-Vivadent, Revolution Formula2: Kerr) were studied. A cone-and-plate rotational viscometer was used to determine all yield stresses. Composites were pre-sheared under a shear rate of 40 s-1 to simulate extrusion of composites from a syringe (the typical tool for application in a clinical setting). The flow curve was then obtained by measuring shear stresses under varying shear rates at a fixed temperature of 25°C. Yield stress values were obtained by plotting the shear stresses for a range of shear rates, fitting the Casson fluid model to the flow curve data, and extrapolating through the stress axis. Apparent viscosity at the lower shear rate (0.2 s-1) was also calculated from the flow curve, as a check on how well yield stress captured fluidity.

Results and Discussion: The flow curves of the composites were fit well by the Casson model. The yield stresses obtained ranged from 5.4 Pa for Revolution Formula2 to 43.1 Pa for AliteFlo LV, and were found to capture differences in the fluidity of flowable resin composites that were not captured by apparent viscosity measurement at the lower shear rate. This proposed method of calculating yield stress was able to quantify the fluidity of flowable resin composites and was more suitable than apparent viscosity measurement.

Conclusions: The yield stress values relate directly to fluidity, and could serve as a simple and precise indicator for selecting flowable resin composites in various clinical applications.

P-20

Wear Resistant of Composites Containing Filler Treated with a Novel Hydrophobic Silane.

Nihei T1*, Kunzelmann KH2, Ohashi K1, Miyake K1，Yoshino N3 1Kanagawa Dental Univ., Kanagawa, Japan 2Dental School of LMU, Munich, Germany 3 Tokyo Univ. of Science, Tokyo, Japan

*[email protected]

Introduction: The mechanical properties of cured composites crucially depend upon the condition of the interface between surfaces of the inorganic filler particles and the polymerized organic resin in which the filler particles are dispersed. This present study has been planed to investigate the water resistance of the experimental composites containing the filler modified with a novel hydrophobic silane coupling agent by wear test. Materials and Methods: The novel silane coupling agent having hydrophobic phenyl group 3-(3-methoxy- 4-methacryloyloxyphenyl)propyltrimethoxysilane (p-MBS) was synthesized. 3-methacryloyloxypropyltrimethoxysilane (3-MPS) was used as a control. The experimental light-cure hybrid composites containing 85 wt% of filler modified with those silanes were formulated. The samples (10×7×3 mm) of the prepared composites were polymerized by irradiation for 120 seconds. After polymerization, the specimens were attached sample wheel, and then evaluated at three-body-wear test (ACTA wear machine 3, Willytec GmbH, Germany). Ten specimens was tested for each storage condition. Obtained data were statistically analyzed with one-way ANOVA. Post-Hoc Tukey multiple comparison tests were used to identify statistically homogenous subsets (α=0.05). Results and Discussion: The experimental composites containing filler modified with 3-MPS composite showed significant increase of wear loss values after water immersion. On the other hand, the wear loss values of p-MBS composite was significantly low compared with that of 3-MPS composite after water immersion for 180 days, and showed no significant change after water immersion. It was suggested that the resin composite containing filler modified with the novel hydrophobic silane has high water resistance, because of the silanized layers treated with this silane had a good affinity with base resin and formed a high hydrophobic layer.

P-21

Degradation behaviour of elements from resin composite blocks for CAD/CAM system

Iwata J*, Asakura M, Hamajima S, Hasegawa A, Kawai T, Ban S Aichi Gakuin Univ., Nagoya, Japan * [email protected]

Introduction: Recently, CAD/CAM resin composite blocks including various kind of ceramic fillers have been frequently used. However, there are a few reports about degradation behavior of these blocks. Our previous study demonstrated that the ceramic fillers disappeared after the degradation test. However, it is uncertain whether this defect was caused by dissolution or debonding of the fillers. In the present study, the dissolution test was carried out using inductively coupled plasma atomic emission spectrometer (ICP-AES) to clarify the phenomenon.

Materials and Methods: Four commercially available composite blocks (Katana Avencia block, Kuraray; Shofu block HC, Shofu; KZR-CAD HR2, Yamamoto Precious Metal; and Cerasmart, GC) were used in this study. The blocks were sliced to 2 mm thick disc (10 × 12 mm), using a low- speed diamond saw and polished with a sequence of abrasives to 0.3 µm alumina powder. The discs were immersed in 40 ml of ultra pure water at 37˚C, 45˚C, and 60˚C for 30 days. After the immersion, the dissolution amounts of each element in the immersion solution were measured with ICP-AES (Optima 7300 DV PerkinElmer). Surface of the immersed specimens were characterized by scanning electron microscopy (SEM).

Results and Discussion: Silicon (Si), Barium (Ba), Aluminium (Al) and Boron (B) were detected in the solution after the immersion of Cerasmart, and the released amounts of each element increased with increasing of the maintained temperature (Fig. 1). As for the other resin blocks, Si from Katana Avencia block, Si and sodium (Na) from Shofu block HC, and Si, Ba, Al, strontium (Sr) and Na from KZR-CAD HR2 were detected. The leached amounts varied among resin blocks. SEM observations demonstrated that ceramic filler particles in KZR-CAD HR2 and Fig. 1 Amount of elements released from Cerasmart partially disappeared. These Cerasmart at 37, 45 and 60˚C for results suggest that the ceramic fillers of 30days. KZR-CAD HR2 and Cerasmart easily dissolute into the water.

P-22

Polymerization shrinkage of various core build-up resin composites

Maseki T*, Maeno M, Ogawa S and Nara Y Department of Adhesive Dentistry, School of Life Dentistry at Tokyo, The Nippon Dental University, Tokyo, Japan * [email protected]

Introduction: Core build-up resin composites have been used for reinforce of endodontically treated teeth with glass fiber post materials in recent clinical situation. The purpose of this study was to examine the polymerization shrinkage of dual-cure and light-cure core build-up resin composites by different curing methods.

Materials and Methods: Three dual-cure type core build-up resin composites; Clearfil DC Core Plus (DP, Kuraray Noritake Dental), UniFil Core EM (UC, GC), Build-it FR (BI, Pentron) and two light-cure type core build-up resin composites; Clearfil Photo Core (PC, Kuraray Noritake Dental) and MI Core LC (ML, GC) were used. Each core material was filled into a translucent acrylic tube mold (φ4 * 8mm) and then cured by two different light irradiation methods; HP: irradiated with high-power mode (light intensity; 1,600 mw/cm2 * 5 sec) or SS: soft-start mode (light intensity; 1,000 mw/cm2 * 2 sec + 1,600 mw/cm2 * 3 sec) using a LED light source (G-Light Prima 2, GC). Linear polymerization shrinkage was measured using a diode laser displacement sensor (HL-C105B-BK, SUNX) during 0-180 sec (n=5). The data were analyzed using two-way ANOVA and Tukey’s q-test.

Results and Discussion: The mean values of linear polymerization shrinkage (S.D.) in % with HP / SS were DP: 2.37(0.37) / 2.12(0.36), UC: 2.82(0.40) / 2.37(0.26), BI: 3.81(0.40) / 3.63(0.40), PC: 1.04(0.19) / 0.70(0.20) and ML: 1.23(0.28) / 1.17(0.12). Both differences in core resin composite and curing method influenced shrinkage rate significantly (p<0.01, 0.05) and there was no interaction effect between two factors. There was no significant difference in polymerization shrinkage between DP and UC, and also PC and ML. It had a tendency that the linear polymerization shrinkage of light-cure type core build-up resin composite was lower than dual-cure type core build-up resin composite. Meanwhile, the soft-start light irradiation was effective for reduction of polymerization shrinkage. It suggested that light intensity of initial polymerization stage is important factor of polymerization shrinkage at core build-up resin composite.

Conclusions (Optional): The polymerization shrinkage occurred by the high-power mode LED irradiation was significantly greater than that of soft-start mode LED irradiation. The effect of the curing methods with regard to polymerization shrinkage was influenced with the various core build-up resin composites in this study.

Acknowledgements (Optional): This work was supported by JSPS KAKENHI Grant Numbers: 15K11134.

P-23

Effect on the flexural test methods of dental hard resins after accelerated deterioration

Miura D, Miyasaka T, Aoki H, Aoyagi Y, Ishida Y Nippon Dental Univ., Tokyo, Japan * [email protected]

Introduction: Three points bending test is adopted in the ISO standard for dental hard resin. However, it is well- known that the measured bending strength varies significantly due to the defects at the sample preparation. On the other hand, biaxial bending tests were known to be less affected by such defects. In our previous study, it was found that a biaxial bending test was applicable to the bending strength of dental CAD/CAM resin blocks. The purpose of this study was to examine the relationships among three point, four point and biaxial bending strength of dental hard resins. Furthermore, the effects of accelerated deterioration on dental hard resin were investigated.

Materials and Methods: Five kinds of dental hard resins were examined (Table1). Sample preparations and mechanical strength tests were carried out in accordance with ISO 6872-2008. All specimens of each hard resin were prepared according to manufacturer’s instructions. The bar shaped specimens for three points and four points bending tests were prepared using a Teflon mold of 2.0 x 2.0 x 25 mm. The hard resin paste was filled into the mold and polymerized by visual light irradiation for 30 minutes. After photo- polymerization , ES was heated for 15 minutes at 105 degrees Celsius with an electrical oven. All specimens were sequentially polished with #600 and #1000 water-proof abrasive paper. Specimens were divided into two groups, and subjected to the accelerated deterioration test under the standard autoclave sterilization conditions, i.e., 121°C in the water vapor of 0.2 MPa for 24 or 72 hours (n=5). Three points bending tests with the loading rod of 3mm in diameter, the support rollers of 3mm in diameter and supporting span of 20mm, was conducted using a universal testing machine at a crosshead speed of 1.0mm/min. Four points of bending tests were carried out at the similar test condition of the three points bending except that the inner span of the loading rods was 10 mm. Biaxial flexural strengths were measured by the piston-on-three-ball method according to the standard of dental ceramics ( ISO 6872 ). The specimens were placed in a circular center and supported by three symmetrically spaced balls of 1.5 mm in diameter. The bending load was applied by the flat end loading rod of 1.4 mm in diameter. Three flexural strengths for each specimens were statistically analyzed by Two-way analysis of variance, then Tukey’s multiple comparison tests were performed, and Weibull analysis was also performed. Table 1 Dental hard resins used in this study Results and Discussion: Code Product name Shade Maker Lot No Significant differences were observed in CM CERAMAGE A3B Shofu 101576 the main factors, i.e., the materials (factor SH SOLIDEX HARDURA A3B Shofu 121519 A), the accelerated deterioration (factor B), GD GC GRADIA DA3 GC 1512171 and the test method (factor C) (p<0.01). DN GC Dia-Na DA3 GC 1601191 Significant differences were also observed ES ESTENIA ®C&B DA3 Noritake 6F0017 in the interactions between the materials and the test method( A x C ) (p<0.01), the accelerated deterioration and the test method ( B x C ) ( p<0.05 ）. Flexural strength became smaller along with testing time. The accelerated deterioration was found to cause a reduction in flexural strength and affect the result of bending test methods.

P-24

Influence of ethanol solution on fatigue strength of resin composite with hybrid cluster filler Sakaguchi T1, Nishikawa I1, Kato T2, Anraku T2 , Takahashi H3 1Osaka Institute of Technology Univ.,Osaka ,Japan 2Yamamoto Precious Metal Co., Ltd.,Osaka ,Japan 3Tokyo Medical and Dental Univ.,Tokyo ,Japan e-mail:[email protected]

Introduction: Improvement of fatigue strength and wearing property of dental resin composite is an important issue in application of teeth restorative material. An emphasis was put on fatigue property of resin composite under ethanol environment in this study. Special cluster filler was newly produced by sintering micro particles of silica. Fatigue strength and wear property of dental composites with various kinds of cluster fillers, wear investigates.

Materials and Methods: Monomer used in this study was composed of UDMA , TEGDMA and camphorquinone (CQ). 2μm, 3.5μm, 5μm, 10μm in diameter of cluster fillers were prepared for reinforced materials. Table1 shows filler compositions of resin composites used in this study. Specimen made of composites with various compositions of cluster fillers were used for fatigue tests. Test specimens had a cross section of 2mm × 2mm, and a longitudinal length of specimen was 20mm. A span of specimen for applying a cyclic stress was 20mm, and a stress wave was selected as sinusoidal. Stress ratio of fatigue test was setting 0.1.

Results: Fig.1 shows the relationship between applied stress range and number of stress cycles to failure obtained from three point bending fatigue tests in air. A plot with an arrow symbol in Fig.1 indicates a run-out data, which means a sample is not broken until 106 in number of stress cycles. The fatigue strength of a sample made of hybrid composite with two type of cluster filler sizes, was higher than that of composite with a single size of cluster. Dual cluster structure of composite was found to be effective to raise the fatigue strength of composite at air environment. Furthermore, fatigue strength of hybrid composite with small size of fillers was higher than that of composite with larger size of fillers. Next, fatigue strength of composite under ethanol environment was investigated. There was no environmental effect on the degradation of fatigue strength of composite with cluster fillers. It was found that the hybrid composite composed of small cluster fillers had a good fatigue property even under ethanol environment.

P-25

Study on Surface Properties of Composite Resin and Discoloration Ishikawa A The Nippon Dental Univ., Tokyo, Japan e-mail ([email protected],jp )

Introduction: To meet patients increasing aesthetic needs, the use of composite resin treatment is increasing annually. Surface properties of composite resin is that factors affecting the prognosis of the restoration. This study compared the levels of beverage-associated discoloration of composite resins, focusing on their types and surface roughness.

Materials and Methods: ESTELITE Σ QUICK (EST; Tokuyama Dental Corp.) , GC MI GRACEFIL (MIG; GC Corp.) and GC MI FIL( MIF ;GC Corp.) were used as 3different types of composite resin. Their upper and lower surfaces were light-irradiated for 40 seconds each, 80 seconds in total, using the light irradiation device G-Light Prima (GC Corp.) to create 3 sample columns with a diameter of 1 cm and height of 2 mm. Subsequently, their upper and lower surfaces were polished roughly and roughly to finely, respectively, using the following waterproof polishing papers: upper: #600; and lower: #600, #800, and #1,000, in this order, for 10 seconds each. After polishing, 3 samples were soaked in green tea (IEMON TOKUTYA; Suntory Holdings Ltd.) at 37 degrees Celsius for 4 weeks. To examine color differences, their colors on L*, a*, and b* were measured before and after treatment using a colorimeter (Shade Eye NCC; SHOFU Inc.), and 2-way ANOVA was performed.

Results and Discussion: In color differences of composite resin, MIG showed the highest value at 9.49, followed by MIF: 9.06, EST: 4.27. On comparison of the color difference E*ab, significant differences were observed between the combination of two samples. : MIG, EST and MIG, MIF. But there were no significant difference in the value between MIG and MIF. The color difference also markedly varied, depending on the polishing level: rough: 9.14 and fine: 6.03. The level of discoloration time-dependently increased when soaking composite resins in green tea.

Conclusion: Significant differences in the level of beverage-associated discoloration were observed between the different types of composite resin, as it was higher in MIG and MIF than in EST. Furthermore, the discoloration level was lower when polishing them finely compared to roughly.

P-26

The influence of water aging on surface hardness of low shrinkage

light-cured composite resins Zhu Song Department of Prosthetic Dentistry, School and Hospital of Stomatology, Jilin University, Changchun 130021, China e-mail:[email protected] Introduction: Light-cured composite resins have been widely used since they were introduced into dentistry, but there are still many deficiencies. They have a degree of hydrolysis, incomplete monomer polymerization and polymerization shrinkage when they are used in clinic. Reduced the polymerization shrinkage rate is the most significant advantage of low shrinkage composite resins. This experiment investigate the changes of low shrinkage composite resins’ performance during long-term artificial aging process. Materials and Methods: Six kinds of light-cured composite resins (two kinds of universal composite resins: Filtek™ Z250 and Solitaire2; four kinds of low shrinkage composite resins: Clearfil Majesty Posterior, Kalore GC, Filtek™ LS and Admira) were made into circular samples (6mm in diameter and 2mm in thickness) with two energy levels (12J/cm2 and 16J/cm2) and three kinds of light intensity (500mW/cm2, 700mW/cm2 and 900mW/cm2). The samples were immersed in distilled water for 1 year. Vickers hardness of the composites’ top and bottom surface were measured at the fixed time. Results and Discussion: The Vickers hardness of Clearfil Majesty Posterior, Filtek™ Z250 and Solitaire2 continued to decline during the process of 1 year aging (P＜0.05). The values of Kalore GC, Admira and Filtek™ LS have been increasing early and decreasing at last, the final hardness values of Kalore GC and Admira were still higher than their initial hardness of 24h (P＜0.05). The final hardness value of Filtek™ LS was similar to its initial hardness of 24h (P＞0.05). Conclusions: The hardness values of six kinds of commercial composite resins in this experiment presented falling and stable trend after long-term water aging process. Among them, the kinds of prepolymerized, organically modified ceramics and silorane-based composites in low shrinkage composite resins performed better aging resistance. The hardness of two universal composite resins were decreasing during water aging process.

P-27

Influence of the hypochlorous-acid water on properties of

light-cured glazing agent membranes

Aoki H1*, Miyasaka T1, Aoyagi Y1, Ishida Y1, 2, Miura D1 Nippon Dental Univ. at Tokyo, Japan * [email protected] Introduction: Light-cured glazing agent is mainly used for dental resin prosthesis in order to improve surface roughness and wear-resistance. We reported the surface roughness, contact angle of water, gloss, color difference, wear resistance and shear bond strength of light-cured glazing agent applied on the heat-cured denture base acrylic resin1). On the other hand, hypochlorous-acid water shows bactericidal property and disinfection effect. However, it is known that this water cause demineralization of tooth enamel surface and corrosion of dental alloys2, 3). The purpose of this study was to clarify the effect of hypochlorous acid water on light-cured glazing agent membranes.

Materials and Methods: Plate samples were made by curing the heat-cured denture base acrylic resin (ACRON, GC) in the usual manner. The plates were divided into samples of 20 × 10 × 1.5 mm, and then the surface was polished by the waterproof abrasive papers of #400. Five light-cured glazing agents, i.e. RESIN GLAZE Liquid (SHOFU), NANOCOAT Lab (GC), SURFACECOAT (KURARAY), Palaseal (HERAEUS KULZER) and BISCOVER LV (BISCO) were used. Four hypochlorous acid water, i.e. strongly acidic-, mildly acidic-, slightly acidic-, neutral-water, and tap water were adopted as immersion fluids. Coated with each light-cured glazing agent, the samples were light cured for 5 minutes by light irradiating apparatus. The specimens were hung by nylon strings into sample tube holding the immersion fluid of 50 ml for 0, 1, 2, 3, 4 weeks. After immersion for each period, surface gloss, color difference and surface roughness of the specimen were measured. As for statistical treatment, three-way ANOVAs (A: Light-cured glazing agent, B: Immersion fluids, C: Immersion period) and Tukey’s multiple tests were performed.

Results and Discussion: The surface gloss decreased when immersed in strongly acidic- and mildly acidic-water. The values increased as the immersion period became longer. The color differences before and after the immersion in acidic-water tended to be smaller than those immersed into tap water. The surface roughness of specimens immersed in acidic-water also tended to be smaller than those immersed into tap water. It became clear that the hypochlorous acid water had influence on light-cured glazing agent membranes.

References: 1. Aoki H, Miyasaka T, Aoyagi Y, Ishida Y, Ando N: Effect on the application of light-cured glazing agents for denture base acrylic resin, JJ Dent Mater, 31(5): 456, 2012. 2. Aoki H, Miyasaka T, Ando N: Influence of acidic and slightly acidic electrolyzed water on human teeth, JJ Dent Mater, 30(5): 350, 2011. 3. Aoki H, Yoshida K, Okamura H, Nakayama M, Miyasaka T: Influence of acidic and slightly acidic electrolyzed water on dental alloys, JJ Dent Mater, 29(5): 416, 2010.

P-28

Development of a quantitative method of residual substances in the autopolymerizing acrylic resin

Hongo T*, Wada T, and Uo M

Tokyo Medical and Dental Univ., Tokyo, Japan * [email protected]

Introduction: N,N-Dimethyl-p-toluidine (DMPT) is widely used as an accelerator in the autopolymerizing acrylic resin in dentistry and also as the intermediate in dye and pesticide synthesis, indicating that DMPT is a high-production volume chemical with the potential for human exposure. However, there are very few reports about the amounts of residual DMPT in cured resins. The purpose of the present study was to develop the simultaneous analysis of residual substances, especially aromatic amines, remaining in the cured resin using high performance liquid chromatography (HPLC).

Materials and Methods: Cured acrylic resins of Unifast TRAD® (GC), which were made from the paste using Teflon mold (5 mm in diameter and 2 mm in length), were prepared according to the manufacturer’s instructions. These cured resins were stored in dark place at room temperature over time periods. Residual substances in cured resins were extracted with the modification of the method of ISO 20795-1:2013 using tetrahydrofuran containing 20 μM hydroquinone (THF-HQ) as an immersion solvent. The HPLC system was an Agilent 1100 with diode array detection (Agilent Technologies). A reversed phased C18, Capcell Pak C18 MGII (250 mm x 2 mm i.d.; Shiseido) with a guard cartridge, was used. The mobile phase was acetonitrile (AcCN) and 95% distilled water-5% methanol. Gradients were set as follows. Time from 0 to 9 min: 35% AcCN; time from 9 to 40 min: linear gradient of AcCN from 35% to 95%; time from 40 to 45 min: 95% AcCN; time from 45 to 50 min: linear gradient of AcCN from 95% to 35%; time from 50 to 60 min: 35% AcCN. Injection volume was 20 μl, and the UV wavelength used for detection was 217 nm, 238 nm and 254 nm. Peaks for eluted substances were identified primarily by retention time with respect to the calibration standards, and diode array spectral match and peak purity were used to confirm identity.

Results and Discussion: We identified methyl methacrylate (MMA), benzoyl peroxide (BPO), DMPT, N-methyl-p- toluidine (NMPT) and 4-aminotoluene (4AT) from the cured resin. NMPT and 4AT may be degradation products of DMPT. DMPT, NMPT, 4AT, BPO and MMA was all stable over 48 h at 24 ˚C in THF-HQ used as an immersion solvent, indicating that these substances did not decompose over time. In 47.5% AcCN-47.5% H2O-5% THF used as a diluting solvent for the HPLC measurement, aromatic amines were resistant to decomposition beyond 40 h at 6 ˚C. Substances in cured resins were able to identify as 4AT, MMA, NMPT, DMPT, and BPO by comparing the retention time as well as the wavelength analysis of standard substances with those of the residues. However, six other peaks could not be identified. Residual amounts of MMA, BPO and 4AT decreased with time (Days 1, 7 and 60) in cured resins, whereas residual amounts of DMPT did not result in significant change with time. The simultaneous analysis of residual substances, especially aromatic amines, from cured acrylic resins using HPLC was developed. This HPLC method for simultaneously quantifying DMPT, NMPT, 4AT, MMA and BPO was simple, easy-to-use and reliable.

P-29

Hydrophilicity of MMA-HEMA resin investigated by fluorescence spectroscopy

Nezu T*, Nagano-Takebe F, and Endo K Health Sciences Univ. Hokkaido, Ishikari-Tobetsu, Japan *[email protected]

Introduction: Hydrophilicity/hydrophobicity of resin materials is related to their properties as wettability, degradation by water, staining by oleaginous food component, and so on. It becomes important in molecularly homogeneous addition of some functionalizing hydrophilic agent into resin part. Water sorption and contact angle measurement have been common techniques, but their information is mostly limited the material surface. Mechanical strength test is an indirect post hoc method. This study aimed to physico-chemically evaluate the hydrophilicity of acrylic resin by fluorescence spectroscopy using hydrophobic fluorescent probes.

Materials and Methods: As the resin monomers, methyl methacrylate (MMA) and 2-hydroxyethyl methacrylate (HEMA) of reagent grade were used without further purification. MMA and HEMA were mixed at various ratios with the HEMA volume fraction of 0, 25, 50, 75, and 100%. Two typical hydrophobic fluorescent probes, pyrene (py) and N-phenyl-1-naphthylamine (npn) were chosen. Stock solutions of these dyes (1 mM in ethanol) were diluted 100-fold by each MMA-HEMA mixture in a quartz cuvette, which was set in a spectrofluorometer (FP-6300, Jasco, Tokyo, Japan). Excited at 342 nm for py or 340 nm for npn, fluorescence emission spectra were measured (350~650 nm) at 25.0°C.

Results and Discussion: Fluorescence spectrum of py is characterized by a monomer band and an excimer (aggregate) band, and only the former was observed in the current nonaqueous systems. There are four subbands in the py monomer band, and the ratio of the first band (374 nm) to the third band (385 nm) intensities, I1/I3, is a common “hydrophilicity” index (Fig. 1). In this experiment, I1/I3 linearly increased with the HEMA fraction (Fig. 2). On the other hand, npn gives a single band, in which the intensity (quantum yield) becomes larger and the peak shifts to a shorter wavelength in more hydrophobic environment (Fig. 3). Actually, λmax increased almost linearly with the HEMA fraction (Fig. 2). A highly linear correlation between the λmax and the known dielectric constant (εr) values for some liquids enabled to deduce unknown εr values from the observed λmax (data not shown). Furthermore, we previously showed that the water sorption of the MMA-HEMA resin increased with HEMA content (Fig. 2), and both the I1/I3 of py and the λmax of npn showed a good correlation with the water sorption. Thus it was concluded that the hydrophilicity of the resin materials can be numerically evaluated on the physico-chemical basis, using the fluorescent hydrophobic parameters in combination with the tabulated εr values.

max / nm (△) I1/I3 (○)  = 342 nm  = 340 nm

ex 420 5 ex )

● hydrophobic environment ( 4 2 HN monomer 1.3 I1 * I3 3 I e 410 excimer hydrophilic environment 2 1.2

Im 1

Fluorescenceintensity Fluorescenceintensity

400 0 Water/ sorption mg/cm 400 500 600 0 25 50 75 100 400 500 600 em / nm Vol% of HEMA in MMA-HEMA mixture em / nm Fig. 1 Typical fluorescence emission Fig. 2 Specific parameters in MMA- Fig. 3 Typical fluorescence emission spectrum of pyrene (py) in water. HEMA mixture at various HEMA spectrum of N-phenyl-1-naphthylamine Subscripts “m” and “e” denote content (npn). Peak position and intensity vary monomer and excimer, respectively. depending on the hydrophobicity.

100

P-30

Effect of bleaching after the discoloration immersion test on color difference and mechanical properties of CAD/CAM resin block.

Aoyagi Y, Miyasaka T, Aoki H, Ishida Y, Miura D Nippon Dent. Univ., Tokyo, Japan *[email protected]

Introduction: The CAD/CAM composite resin block (CR block) has been in widespread clinical use in recent years, but because of the resin matrix, there is a risk of discoloration. In the previous study, change over time in color and mechanical properties of CR blocks after immersion test were investigated. There has been few studies on the color change and mechanical properties after breaching of CR blocks, which were experienced the discoloration by immersion tests. Therefore, the purpose of this study is to accurate the color change and mechanical properties of CR blocks for CAD/CAM, which are immersed in red wine, then experienced dental beaching. Materials and Methods: Three kinds of dental CR block for CAD/CAM, i.e., HC block (SHOFU), Enamick (VITA) and GRADIA block (GC), were examined. . These prism shape blocks were lathe processing into cylindrical form with diameters of 12 mm, then sliced into 1.5mm thickness by slicing machine (Isomet 1000 Precision Saw, BUEHLER). The disk- shaped CR block sample was subsequently polished with #400 to #1000 water-proof abrasive paper to a thickness of 1.2 mm and a diameter of 12 mm, then buff finished using the alumina powder of 1 ㎛ in diameter until the mirror-like surface was obtained. The red wine (Casa pino, Japan) was used as the immersing discoloration fluid, and tap water was used as a control immersing fluid. The immersing fluid of 50 ml was poured into a screw test tube, and the specimen was completely immersed into the fluid, then stored at 37±1 degrees Celsius for more than four weeks. The specimen experienced discoloration were taken up from the immersion fluid, then bleached by the dental bleach agent (Hi Light, SHOFU) every week. The bleaching procedure was followed the maker instructions. The breaching agent was applied to the specimen, and standstill for two minutes, then visual light was irradiated for 3 minutes, subsequently washed with water. The Vickers hardness of specimen was measured by a Vickers hardness test machine ( AVK, Akashi ). The color of specimens before and after immersion test and after breaching, were measured by a color colorimeter (CR- 200, MINOLTA), then the color changes in NBS unit were calculated. The biaxial bending tests (piston-on-three ball test) before and after bleaching were performed by a universal testing machine ( AGS-X, Shimadzu ) at a crosshead speed of 1.0 mm/min. The data were analyzed by a three-way analysis of variance, then Tukey’s multiple comparison tests were performed. Results and Discussion: Vickers hardness tended to decrease after bleaching. Vickers hardness of Enamic was the highest ( > 150 ), and that of HC showed the lowest value ( <80 ). All CR brocks tends to whiten by bleaching and nearly return to an original color before immersion to the wine. The flexural strength tended to decrease according to the immersion time and became weaker to immerse in the wine. It was found that the immersion into the wine and the bleach process weaken the mechanical strength of CAD/CAM brock and breaching decrease the color change.

101

P-31

Flexural properties of denture base resin for 3D printing Park DR*, Bae JM College of Dentistry, Wonkwang Univ., Iksan, Korea *e-mail ([email protected]) Introduction: The increasing use of 3D printer in dentistry makes it possible to produce prosthesis with higher accuracy easily. Nowadays 3D printing can be applied to fabricate resin coping of PFM, resin crown and bridge, resin framework of removable partial denture to be cast into metal, temporary crown, surgical guide and models for orthodontic and prosthodontic treatment. The aim of this study was to compare the flexural properties of commercial denture base resin, commercial 3D printing resins, and experimental resins for light-curing and 3D printing.

Materials and Methods: Self-curing pour-type denture base resin (PRESS LT, Retec, Germany) was used as a control. Two kinds of commercial 3D printing resins, White prototyping resin (PR57-W, Autodesk, USA) and Clear prototyping resin (PR48, Autodesk) were used. Experimental resins were fabricated using bis-GMA and di(trimethylolpropane) tetraacrylate (DiTMPTA) for light-curing and 3D printing, respectively. DLP type 3D printer (EMBER, Autodesk) was used for 3D printing. The specimen size was 64×10×3.3 mm according to ISO 20795-1. Self-curing and light-curing specimens were made using the mold with the same size made of polyvinyl siloxane. Light-curing was done for 20 sec twice on both upper and lower surfaces with LED light-curing unit (Dr’s light Clever, Good Doctors Co., Korea). The sample size of each group was 9. The flexural strength and flexural modulus were measured by a universal testing machine (Z020, Zwick, Germany) at a crosshead speed of 5mm/min. The results were analyzed with one-way ANOVA, and the Tukey HSD test (α=0.05) with IBM SPSS program (version 22).

Results and Discussion: Flexural strength Flexural modulus Type Fabrication method Name (MPa) (GPa) Self-curing PRESS LT 99.36±9.92e 2.51±0.18d a a Commercial 3D printing White 41.79±5.23 0.65±0.12 3D printing Clear 46.83±8.02ab 0.84±0.16a Light-curing Bis-GMA-based 88.21±6.47d 1.68±0.17c Light-curing DiTMPTA-based 56.31±9.03bc 1.7±0.23c Experimental 3D printing Bis-GMA-based 53.89±6.17b 0.87±0.09a 3D printing DiTMPTA-based 64.47±3.75c 1.18±0.06b

The flexural strengths and moduli of commercial denture base resin (PRESS LT) were significantly higher than other groups and those of commercial 3D printing groups (White and Clear) were significantly lower than other groups (P<0.05). In experimental groups, light-curing groups showed significantly higher flexural modulus than 3D printing groups (P<0.05). In 3D printing groups, DiTMPTA-based experimental group showed significantly higher flexural strength and modulus than other groups (P<0.05).

Conclusions: Commercial denture base resin showed the highest flexural properties than 3D printing groups. Because DiTMPTA-based 3D printing group showed the highest flexural properties among 3D printing groups, it has possibility to be developed as a 3D printed denture base resin.

Acknowledgements (Optional): This work was supported by the Technological Innovation R&D Program (S2311063) funded by the Small and Medium Business Administration (SMBA, Korea).

102

P-32

Influence of a plasma treatment using a different gas on the bond strength between composite resin and fiber post

Ito Y*, Okawa T, Yamamura T, Fukumoto T, Tanaka M Osaka Dental Univ., Osaka, Japan *e-mail ([email protected])

Introduction: We have previously reported on the efficiency of plasma treatment with helium gas as a surface treatment method of fiber posts. However, it was not elucidated how difference in active gas type influences the efficiency. Therefore, in this study we performed the plasma treatment using various gas types, and examined the impact on the bonding strength between the composite resin and fiber post.

Materials and Methods: Fiber post (GC) was used as the fiber post, Unifilcore EM (GC) as the abutment construction resin, G-CEM LinkAce (GD) as the adhesive resin cement, and ceramic primer II (GC) as the saline processing material. The active gas types were helium (He) and argon (Ar). Two types of the surface treatment method of fiber post were used: plasma treatment only (Ps), and a saline processing after plasma treatment (Ps-Pm). After the surface treatment, a frame fabricated using a silicon mold was filled with abutment construction resin or adhesive resin cement, the fiber post was inserted into the center of the frame before photo polymerization was carried out. After leaving at room temperature for 24 hours, the frame/post was sliced to 3mm, which were then used as test samples. An extrusion test was employed as the adhesion test. Statistical analysis was carried out using two way factorial analysis of variance with factors of the gas types and surface treatment method. The significance level was set to 1%.

Results and Discussion: The results of variance analysis showed that statistically significant difference was observed for the abutment construction resin depending on the gas type and surface treatment, but no interaction was observed. In the case of the resin cement, statistically significant difference was only found for the surface treatment method. As compared to the He treatment, the Ar treatment showed higher values for Ps-Pm than for Ps. In the Ps-Pm group, destruction of the cement was observed in all samples. We speculate that since the bonding strength between the post and cement exceeded the strength of the cement, there was no significant difference between the gas species.

Conclusions: The test results of the influence of plasma treatment revealed that higher bond strength can be achieved by the argon plasma treatment followed by the saline processing.

103

P-33

Influence of plasma treatment to the three-point bending strength of the fiber post

Yamamura T*, Okawa T, Ito Y,Yosie S and Tanaka M Osaka Dental Univ., Osaka, Japan *e-mail ([email protected])

Introduction: Since the flexural modulus of fiber posts is close to that of dentin, stress concentration at the post tip portion is unlikely to occur as compared to a metal post. Fiber posts can be applied for patients with metal allergy or prevent discoloration of the gingiva, as well as for aesthetic prosthetic restoration. In recent years, the plasma treatment was reported to enable activation of surface energy and removal of contaminants present in fine gaps without risk of deformation due to physical force and heat. Therefore, the plasma treatment is deemed very effective as a surface treatment method in dental clinical practice dealing with restoration devices made of brittle materials and having fine irregularities. In this study, we examined the effectiveness of the plasma treatment as a surface treatment of fiber posts.

Materials and Methods: In this study, as a fiber post, a fiber post (GC, φ1.4 mm) was used. Surface treatments for the fiber post were categorized into three groups: (1) non-treatment (Cont group), (2) sandblasting (Sb group), and (3) plasma treatment (Ps group) (n = 7). Ps group was irradiated with plasma in remote mode from a distance of 10 mm for 30 seconds. For the Sb group, alumina sandblasting with alumina particles of 50-70 μm in size was performed for 5 seconds at 0.3 Mpa from a distance of 30 mm. A three-point bending test was carried out with the Universal tester ALTOGRAPHE AGSJ- 5kN (Shimadzu) at a crosshead speed of 1mm/min with a span of 10 mm between the support points. The maximum load obtained was considered as the maximum bending strength. SPSS version19 (IBM) was used for statistical analysis and one-way analysis of variance with the surface treatment method as factor was performed. If statistically significant difference was observed, multiple comparisons were carried out using Bonferroni’s correction method. The significance level was set to 1%. Further, each sample surface was observed under a stereomicroscope, SZX-12 (Olympus).

Results and Discussion: Variance analysis results showed statistically significant difference in the surface treatment method. Multiple comparisons confirmed significantly higher maximum bending strength in the Cont and Ps groups than in the Sb group. No statistically significant difference between the Cont and Ps groups was observed. Sample observation with a stereomicroscope after the surface treatment revealed remarkable destruction on the fiber post surface in the Sb group. Fiber posts are manufactured by embedding resin into fibers tightly bundled in the longitudinal direction. In the Sb group, decrease in bending strength is attributable to damage to the resin and fiber caused during the sandblasting treatment. Thus, the plasma treatment to the fiber post was found to have no adverse impact on the flexural strength of the fiber post.

104

P-36

Manufacture of the metal dies for the experiment in the adhesion examination

Igarashi K*, Moriue T, Ohki T, Saitou R, Ishida Y, Okada H, and Terada Y Ohu Univ., Koriyama, Japan

* [email protected]

Introduction: We generally use the abutment tooth metal die in measuring the bond strength of dental cements (temporary luting materials, luting materials). However, most dies are for the single crown, and some of them are unsuitable for real clinical practice. This times, we report the status setting of the abutment teeth metal dies of the bridge type to use in the adhesion examination to in new temporary luting material which developed in our school.

Materials and Methods: The means of the anatomical width diameters of human mandibular teeth were investigated. And the appropriate tapers of bridge abutment teeth were investigated. Stainless steel blocks were cut with CAD/CAM, processed and completed. In addition, the metal dies for the temporary superstructures.

Results and Discussion: The abutment teeth dies became the reproducible high anatomically. However, removing is difficult after polymerization, and the metal dies for the temporary superstructures need further reformability.

106

P-37

Shear bond strength of “PANAVIA V5” between dentin and resin blocks for CAD/CAM

Kashiki N and Okada K* Kuraray Noritake Dental Inc., Tokyo, Japan * [email protected]

Introduction: Kuraray Noritake Dental Inc. has developed a new adhesive resin cement system “PANAVIA V5”. It consisted of a dual-curing composite resin cement --“PANAVIA V5 Paste”, a one-bottle MDP-based self-etching tooth primer --“PANAVIA V5 Tooth Primer”, & a one-bottle prosthetics primer --“CLEARFIL Ceramic Primer Plus” which bonds to all silica-based ceramics (including lithium disilicate), zirconia, composites & metals. The purpose of this study was to evaluate the shear bond strength (SBS) between bovine dentin and the resin blocks for CAD/CAM cemented with “PANAVIA V5”.

Materials and Methods: Rod-shaped resin block specimens (diameter 5mm) were prepared from the resin blocks commercially available, Katana Avencia Block (KAB; Kuraray Noritake Dental), Lava Ultimate (LU), Shofu Block HC (SHOFU; SHC) and Cerasmart (GC; CS). The surface were polished with 1000-grit SiC paper, and then sandblasted with 50μm alumina. The bovine tooth was embedded in acrylic resin, grinded for the exposure of the dentin surface, polished with 1000-grit SiC paper. SBS to bovine dentin in self-curing mode was measured after storage in 37˚C water for 24 hours and after subsequent thremocycling (4˚C-60˚C, 1min. each, 3,000cycles: TC3000). The bonding procedure was performed according to the instruction for use. For comparison, the bond strength of the previous resin cement “PANAVIA F2.0” was evaluated with KAB.

Results and Discussion: The results were summarized in Table 1.

Table 1 The share bond strength to dentine and the resin blocks Resin cement PANAVIA F2.0 PANAVIA V5 Resin block KAB LU SHC CS 37 ˚C 24 hours 7.7 (1.5) 15.2 (3.6) 15.2 (3.2) 11.8 (3.8) 14.3 (4.7) TC3000 7.6 (4.3) 14.7 (2.4) 14.2 (3.4) 11.5 (6.0) 14.8 (2.0)

The new resin cement system “PANAVIA V5” showed the excellent bond strengths after 24 hours and TC3000 for all resin blocks for CAD/CAM. This study indicates that “PANAVIA V5” would exhibit reliable clinical performance for resin block restoration.

107

P-38

The effect of application conditions of core resin on the push-out bond strength to root canal dentin

Niitsuma A1*, Shinya A1, 2, Shimizu S1, Kuroda S1 and Gomi H1 1The Nippon Dental Univ., Tokyo, Japan 2Turku Univ. , Turku, Finland * [email protected]

Introduction: The core resin replaces carious, fractured, or missing coronal structure and helps to retain for final restoration. The advantages of core resins are adhesive bonding to tooth structure and many posts, ease of manipulation, and translucent formulations. Core resin materials are typically two-paste, dual-cured composites, but light-cured materials are also available. In areas that are not easily penetrable by light, dual-cured resin composites are frequently used as core resin materials for direct restorations. Although, the dual-cured core resin was increased the contraction stress than when used to light-cured core resin. The purpose of this study was to evaluate the regional bond strength of a composite resin core material to root canal dentin using eight pattern application condition including resin cure type and fill method and light irradiation distance.

Materials and Methods: Eighty single -rooted bovine-teeth were used in this study. Crowns were removed under water using a low-speed diamond saw to obtain a root height of 15 mm. After preparing the root canals, the specimens were randomly divided into eight groups. The surface of the root canal dentin was treated with the bonding systems according to the manufacturer’s instructions and composite was applied into the root canal. Their root canal dentin surfaces treated with one of the following polymerization procedure (Dual Cure: DC, Light Cure: LC) / filling method (Batch filling: I, Incremental filling: II) / light irradiation distance (0mm: 0, 5mm: 5). All specimens were then stored in water for 24 hours at 37℃. Each root was embedded in auto-polymerizing acrylic resin by using a plastic mold (12 mm in diameter and 25mm in height). Each specimen was then sectioned horizontally for obtaining 1 ± 0.1 mm thick dentin slices using a diamond blade under water cooling, thus obtaining five slices of each root, for a total of 400 slices. Each slice was submitted to push-out bond strength testing a universal testing machine at a 1 mm/min crosshead speed until failure occurred. Data were compared using two-way ANOVA and Tukey test. The two independent factors were application condition and measurement position. The fractured dentin surfaces were examined after push-out bond testing under a scanning electron microscope (S-4000, Hitachi; Japan)

Results and Discussion: The results of push-out bond strength of the each condition are indicative of DCI5 produced the highest bond strength (13.7 ± 4.3 MPa). In contrast, the lowest bond strength (10.3 ± 3.7 MPa) was produced with LCI0. Two-way ANOVA revealed that push-out bond strength was influenced by both application condition (p<0.05) and measurement position (p<0.05). An interaction was also found between these two factors (α=0.05).

Conclusions: The present study has shown that DCI5 was led to the highest mean bond strength values. Incremental filling is necessary for increasing bond strength for LC.

108

P-39

Dentin bond strengths of a new one-bottle universal adhesive containing an amide monomer irradiated by QTH and LED

Hanabusa M, Yamamoto T, Kimura S and Momoi Y Tsurumi Univ., Yokohama, Japan [email protected]

Introduction: One-bottle universal adhesives have been recently developed to simplify the bonding procedures for various dental materials. Kuraray Noritake Dental developed an experimental one-bottle universal adhesive (SKB-100) containing a new hydrophilic amide monomer, which needs a shortened application time. The purpose of this study was to evaluate the influence of light source for polymerization on micro-tensile bond strength of SKB-100 to dentin. The hypothesis tested was that the bond strengths with different light sources are similar.

Materials and Methods: Flat Dentinal surfaces were made in non-carious extracted human molars using a diamond saw and 600-grit SiC paper. The surfaces were treated with one of four self-etch adhesives (see Table, Kuraray Noritake Dental, Tokyo, Japan). The adhesives were applied to the surfaces according to the manufacturer’s instructions and irradiated by a quartz-tungsten- halogen (QTH) light-curing unit (Optilux 501, Kerr, CA, USA) or a blue light-emitting diode (LED) light-curing unit (Pencure 2000, Morita, Tokyo, Japan). Total radiant exposures for the adhesives were more than 7.6 J/cm2. A resin composite (CLEARFIL AP-X, Kuraray Noritake Dental) was incrementally built on the adhesive and light-cured. Following 24 h storage in distilled water at 37ºC, the bonded specimens were sectioned and subjected to micro-tensile bond strength tests (bonded area: 1.0 mm x 1.0 mm, crosshead speed: 1.0 mm/min). Data were statistically analyzed by a priori comparisons (Tukey-Kramer, α=0.05).

Results and Discussion: Table Micro-tensile bond strength of each adhesive to dentin (MPa, n = 45) QTH LED SKB-100 44.5 (11.0)ab 48.6 (11.4)bc CLEARFIL S3 BOND PLUS 40.8 (11.9)a 42.9 (8.9)ab CLEARFIL SE BOND 2 54.3 (11.1)cd 57.3 (13.4)d CLEARFIL SE BOND 52.2 (9.7)cd 53.1 (10.2)cd Values having the same superscript are not significantly different (p > 0.05).

All of the adhesives demonstrated no significant differences between the bond strengths with QTH and LED. The hypothesis was accepted. Comparing the one-bottle adhesives (SKB-100 and CLEARFIL S3 BOND PLUS) and the two-step adhesives (CLEARFIL SE BOND 2 and CLEARFIL SE BOND), the latter exhibited significantly higher bond strengths with QTH. The bond strength of SKB-100 was similar to the existing one-bottle adhesive CLEARFIL S3 BOND PLUS, and was improved to the level of CLEARFIL SE BOND when irradiated by LED. Under limited condition of this study, we can conclude SKB-100 with LED is comparable with the two-step self-etch adhesives in the bond strength.

109

P-40

Bond strengths of a one-bottle universal adhesive containing a new amide monomer to various dental restorative materials

Kimura S, Ito M, Hanabusa M, Yamamoto T and Momoi Y Tsurumi Univ., Yokohama, Japan [email protected]

Introduction: Demand of universal adhesive systems is spreading because of their simplicity in usage. An experimental one-bottle universal adhesive (SKB-100, Kuraray Noritake Dental, Tokyo Japan) was developed, which contains a new hydrophilic amide monomer and requires shorter application time. The objective of the present study was to evaluate shear bond strengths of SKB-100 to various restorative materials in comparison with those of an existing universal adhesive. The hypothesis tested was that the bond strengths of the adhesives are similar.

Materials and Methods: Plates of five restorative materials (see Table, two resin composites, one feldspar ceramic and two zirconia) were embedded in acrylic resin. Their surfaces were ground with 180-grit SiC paper after 37ºC water immersion for 1 week. The specimens were divided into two groups for each material. Each group received adhesive treatments with SKB-100 or Scotchbond Universal Adhesive (3M ESPE, St. Paul, USA). Total radiant exposures for the adhesives were 7.6 J/cm2. A resin composite (CLEARFIL AP-X, Kuraray Noritake Dental) was placed on the adhesive-treated surfaces using a mold (ø2.38 mm x 2 mm, Ultradent, South Jordan, USA) and irradiated for 40 s at 600 mW/cm2. The specimens were stored in 37ºC water for 24 h, and then subjected to shear load (CHS = 1.0 mm/min) to measure the bond strength. Data were statistically analyzed by Two-way ANOVA and Tukey’s multiple comparisons (α=0.05).

Results and Discussion: Table Shear bond strengths to restorative materials (MPa, n = 5) Restorative Materials SKB-100 Scotchbond Uni. CLEARFIL AP-X (Kuraray Noritake) 23.4 (2.6)ac 29.9 (5.1)cd KATANA AVENCIA (Kuraray Noritake) 22.5 (5.5)ac 24.0 (3.2)abc VITABLOCS Mark II (Vita) 15.5 (2.3)a 17.6 (5.8)ab KATANA ZIRCONIA HT (Kuraray Noritake) 24.8 (4.1)bc 17.3 (2.4)ab KATANA ZIRCONIA UTML (Kuraray Noritake) 20.3 (4.6)ab 15.5 (3.9)a adhesive: p = 0.706; restorative material: p < 0.001; adhesive x restorative material: p = 0.0028. Values having the same alphabet are not statistically different by the post-hoc test (p > 0.05).

ANOVA revealed that the adhesives had no significant influence on the bond strengths, namely the bond strengths of SKB-100 and Scotchbond Universal Adhesive were similar. The hypothesis was accepted. SKB-100 and Scotchbond Universal Adhesive exhibited the highest bond strengths to KATANA ZIRCONIA HT and CLEARFIL AP-X, respectively. We may conclude that the bond strengths of SKB-100 are comparable with those of Scotchbond Universal Adhesive.

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Effect of surface treatment after the flowable composite base on the bond strength of the restorative ceramic block

Haruyama A*, Kameyama A Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: Resin coating technique has recently been applied to the cavity surface of indirect posterior restoration, not only for protection of exposed dentin surface from tooth sensitivity but also enhance the bond strength between dentin and luting cement. On the other hand, undercuts have often been filled with flowable resin composite in order to reduce the volume of restorative material or for creating adequate geometry for inlay/onlay cavity preparation. However, the ideal pretreatment has not yet been clear for prepared cavity surface filled with the base material. Therefore, the purpose of this study was to evaluate the different pretreatments on microtensile bond strength (µTBS) between flowable composite base surface and restorative ceramic block.

Materials and Methods: Flowable composite blocks (Clearfil Majesty ES flow low-A3, Kuraray Noritake Dental) were prepared. After the flat surface was prepared using #600 SiC paper, four different treatments were preformed; Group I: no treatment (control) Group II: silane treatment with the mixture of Porcelain Bond Activator and Clearfil Mega Bond Primer (Kuraray Noritake Dental) Group III: resin coating with Clearfil Mega Bond + Clearfil Majesty ES flow High (Kuraray Noritake Dental, shade A3) Group IV: resin coating prior to silane treatment For Groups II and IV, silane treatment was performed after the removal of the hydraulic temporary sealing material (Caviton EX, GC). Ceramic blocks (IPS Empress CAD, HT/A2, Ivoclar Vivadent) were ground with #600 SiC paper to obtain the flat surface. After the flat ceramic surface was treated with phosphoric acid (K-etchant) and Clearfil Ceramic Primer, the ceramic block was bonded to the flowable composite block using Panavia V5 and light-cured. After 1 week of the water storage at 37ºC , the bonded specimens were sectioned perpendicular to the bonded surface with a cross sectional area of approximately 1 mm2. The stick-shaped bonded specimens were stressed at a crosshead speed of 1 mm/min until failure. The data were analyzed statistically with 1-way ANOVA and Student-Newman-Keuls test (p=0.05).

Results and Discussion: µTBSs of Group II (20.2 ± 4.0 MPa) and Group III (21.3 ± 7.2 MPa) were significantly higher than that of Group I (17.6 ± 5.9 MPa) (p<0.05). Group IV showed the highest µTBS value (24.9 ± 3.8 MPa) and was found the significant difference to both Groups II and III (p<0.05). These results suggested that either immediate resin coating and silane treatment just before bonding procedure to the composite base material affects to improve the bond strength. In the clinical situation, resin coating should be performed entirely both exposed dentin surface and composite base surface immediately after the cavity preparation. It is also recommended to treatment the surface of base material with silane coupling agent just before the bonding procedure.

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Influence of solvent evaporation on ultimate tensile strength of cured dental adhesives

Kameyama A1*, and Haruyama A1 1Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: Durable bonding to dental tissues is one of the ultimate goal of the longevity for the direct composite restoration. In order to achieve this goal, adhesive resin monomer should be penetrated securely into the demineralized dental tissue, and should also be polymerized sufficiently. Generally, self-etching primer contains hydrophilic monomers dissolved in solvents, such as water, ethanol, and acetone. Hydrophilic monomers promote wettability and infiltration of adhesive resin mainly composed of hydrophobic resin monomers. More simplified all-in-one adhesives are dissolved the hydrophobic resin monomers in solvents. Ideally, the water and solvents should be completely eliminated from the adhesive before polymerization, because the residual solvent may inhibit polymerization. Insufficient polymerization might be caused of the degradation of adhesive interface. The composition of commercially-available adhesives is slightly differed in each product. Therefore, the purpose of this study was to evaluate the influence of solvent evaporation on the ultimate tensile sterength (UTS) of commercial adhesives.

Materials and Methods: Two 2-step self-etch adhesives (Clearfil Megabond FA, Kuraray Noritake Dental; OptiBond XTR, Kerr) and two all-in-one adhesives (OptiBond All-In-One, Kerr; G-Premio Bond, GC) were used in this study. Four bottles were used in each adhesive, and two of them in each adhesive were opened the cap, and stored at 37ºC dry oven with silica gel under the shielding from light for 2 weeks, in order to evaporate the contained solvent aggressively (Evaporated). Remained two bottles were stored at the room temperature (Original). A 1 mm- thick glass slide was fixed under the hour-glass shaped metallic matrix mold with the narrowest potion of 0.9-mm and 0.9-mm thickness. The mold was then filled with the adhesive resin and pressed with a second glass slide with a 1-mm thickness, prior to being light-cured for 60 s through the top glass slide using LED light-curing unit (Demi Plus, Kerr). Cured specimen was then carefully removed from the mold, and measured the weight individually. After the immersion in 37ºC water for 1 day or 1 week under the shielding from light, each specimen was weighed again, and measured the cross-sectional area at the narrowest portion of the specimens individually. Thereafter, the UTS was measured at a cross-head speed of 1 mm/min (n=10). The data were analyzed using ANOVA and Tukey’s HSD test at a 5% level.

Results and Discussion: For Original, two all-in-one adhesives could not be hardened even if the sufficient light was exposed. On the other hand, Evaporated OptiBond All-In-One obtained UTS values of 35.6 ± 15.2 MPa (1h) and 28.2 ± 6.2 MPa (1 week), respectively. UTS values of Clearfil Megabond FA was rather higher in Original than Evaporated (p>0.05). No significant difference was found between Original and Evaporated in OptiBond XTR (p>0.05). Both OptiBond All-In-One and Clearfil Megabond FA increased the weight after the 1- week immersion in water. On the other hand, the weight change of G-Premio Bond and OptiBond XTR were slight. The results found that the residual solvent reduces the mechanical strength of adhesive itself. It was also suggested that the hydrophilicity of adhesive resin might also affect the mechanical strength.

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EFECT OF DIFFERENT DENTAL BONDING AGENT APPLICATION METHOD TO BONDING THICKNESS ANALYZE WITH PERIAPICAL RADIOGRAPHY

Rosalina Tjandrawinata, Lyvia Juliana Faculty of Dentistry, Trisakti University Jakarta, Indonesia

Background: In dental clinic, if there is complain about tooth condition, dentist usually use radiograph as diagnostic equipment. Since bonding material is radiolucent, while composite material, dentin and enamel are radiopaque, sometimes, radiograph interpretation of bonding area between composite filling and tooth structure often misunderstood as secondary caries, thereby dentist break down the restoration.

Objective: The objective of this research is to examine the different effects of dental bonding agent application method taken with periapical radiography, thus creating a good thickness and not being mistaken as secondary caries.

Method: Twenty four proximal boxes were made in premolar teeth. Samples were divided into four groups with different treatments; they were bonding that applied maximally without given any air flow, bonding that applied maximally with given air flow, bonding that applied minimally without given any air flow, and bonding that applied minimally with given air flow. Each sample were digitally exposed and the results were then analyzed with SOPRO IMAGING 1.71 system. Bonding thickness was examined at three diagnostic points; #1 at horizontal part of proximal box, #2 at the corner of proximal box and #3 at vertical part of proximal box.

Results: There were statistically significant differences between bonding thickness due to amount of bonding, air flow and points of measurement (ANOVA, p<0,05). Widest radiolucency of bonding was examined at the corner of proximal boxes.

Conclusion: The minimally applied amount of bonding blown with air flow is the best method of bonding application, which will not be misinterpreted as secondary caries

Keywords: Composite filling, bonding thickness, secondary caries, periapical radiograph

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Comparison of microtensile strength and failure mode under saliva contamination in class V restorations Kisaki Shimazu1, 2*, Hiroyuki Karibe1, Hiroyuki Yamada1, 3 and Kiyokazu Ogata1, 3 1Department of Pediatric Dentistry, Nippon Dental University School of Life Dentistry at Tokyo, Japan 2Momose Dental Clinic, Tokyo, Japan 3Division of Dentistry, Tokyo Metropolitan Children’s Medical Center, Japan *e-mail: [email protected]

Introduction: Contamination of the restorative treatment area by saliva may have an adverse effect on positive restoration. The aim of this study was to evaluate the effects of artificial saliva contamination on cervical restorations (class V) with 3 restorative materials, namely, a glass ionomer cement (GIC), a resin-modified GIC (RMGIC), and a composite resin (CR), for which 2 different etching adhesive systems were used.

Materials and methods: Various surface conditions were created on bovine teeth using artificial saliva. The test specimens were divided into the following groups: Group I (control), Group II (mild saliva contamination), and Group III (severe saliva contamination). The microtensile strength of the bond between the materials and dentin/enamel on the cervical area were determined for each condition. In addition, the failure mode between dentin/enamel and the 3 restorative materials was categorized by scanning electron microscopy.

Results and discussion: The dentin bond strength for CR was significantly lower in Group III than in Group I (analysis of variance, p<0.05). However, there were no significant differences between the 3 groups with regard to the bond strength of GIC and RMGIC with both enamel and dentin. Three types of failure were observed: mixed failure, interfacial failure, and cohesive failure. Most of the post-test specimens showed the cohesive failure mode in GIC and RMGIC (46-83%), and the mixed failure mode in CR (67-100%). For CR with dentin, percentages of interfacial failure mode increased with the level of artificial saliva contamination. These results suggest that GIC and RMGIC are suitable for preventing secondary caries after class V restorative treatments when contamination by saliva cannot be avoided.

Conclusions: Artificial saliva contamination does not affect the microtensile bond strength or failure mode of GIC and RMGIC when used for class V restorations.

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Evaluation method of the adaptability of machined dental restorations

Nagase Y1*, Asakura M1, Ohkuma K2 and Kawai T1 1Aichi Gakuin Univ., Nagoya, Japan 2Nippon Dental Univ., Niigata, Japan * [email protected]

Introduction: CAD/CAM systems is frequently used to fabricate dental restorations such as inlays, crowns and bridges. It is important to evaluate not only the final accuracy of restoration but also the processing accuracies, because CAD/CAM system consists of several processes which include acquisition, measuring, filtering, designing, manufacturing. Y-TZP (Yttria- tetragonal partially stabilized zirconia) is used as CAD/CAM blanks, and the machined restorations shrink about 20-30% by the subsequent sintering process. The purpose of this study is to evaluate the adaptability between the metal die used to prepare the STL data which is used as CAM input data and a dental CAD/CAM restoration made of Y-TZP (Yttria-tetragonal partially stabilized zirconia) with or without sintering process.

Materials and Methods: Ceramill Motion 2 Type 179270 (Amann Girrbach AG) was used as the CAM machine, and Ceramill ZI 71L (Amann Girrbach AG) was used as the Y-TZP blank. STL data for a crown and a 4-unit bridge were prepared using the metal die for crown and bridge Table 1 Test specimens restoration with no cement layer thickness. Type of Sintering Thickness of CAM data sets were created using the STL restorations process cement layer data with and without the shrinkage factor CU without for sintering. The sintering process was crown CS with performed using Ceramill Therm (Amann 0 μm BU without Girrbach AG) with the program 3 specified 4-unit bridge for Ceramill ZI. Each 5 test specimens BS with were fabricated for each case (table 1). The machined test specimens (CU and BU) and the sintered test specimens (CS and BS) were placed on the metal die with 1 N and 50 N for 30 s respectively. The discrepancies at the margin of test specimen were measured using the measuring microscope STM6 (Olympus Corp.). The measuring points were four points of the margin for CU and CS, or three points of each crown margin of BU and BS. The dimensions of the restoration and of the impression made of the vinyl silicon material Exafine (GC Corp.) were measured using the profile projector PJ-2500 (Mitutoyo Corp.).

Results and Discussion: The positive discrepancy (the gap between the margin of test specimen and the base plane of metal die) was greater in order of CS, CU, BU and BS. The discrepancies at the crown part and the another crown part were not different remarkably in both BU and BS. The lateral discrepancies of CS and CU were -1.3% and -1.1% respectively; i.e. the internal diameter at marginal plane (bottom plane) were smaller than that of the metal die. The same phenomena were observed in the dimension measurements. The metal die method is available to evaluate the adaptability of CAD/CAM restorations. The adaptability of restoration would be affected by the restoration type as well as by the shrinkage due to the sintering process.

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Educational evaluation of a drilling simulator for dental implant surgery

Kinoshita H*, Takemoto S1 and Kawada E Tokyo Dental College, Tokyo, Japan *[email protected]

Introduction: A recent survey showed that almost 30 percent of accidents during implant surgery occurred while drilling the trabecular bone region involving the mandibular canal. The fundamental reason is due to the clinicians’ lack of anatomical knowledge and surgical experience. In order to serve this problem from an educational standpoint, we proposed and developed a new simulator system for dental students1). The aim of this study was to evaluate a dental implant surgery simulator that allowed students to experience the dynamic sensations experienced during bone drilling in the posterior mandibular bone.

Materials and Methods: A dental implant surgery simulator was developed in order to teach students the correlation between the sensation felt while drilling and the micro-architecture of trabecular bone in various cases1). The simulator system enabled students to experience the reaction force while drilling various regions of the mandible. We gathered a group of 5th year clinical students attending Tokyo Dental College to experience drilling with the simulator. After repetitively experiencing 2 separate cases, a third case was introduced and differences in force sensations were represented on a graph. A questionnaire was also performed to evaluate the effectiveness of the implant simulator.

Results and Discussion: The simulator was rated highly overall. Based on the students’ experience with repetitive drilling, we were able to quantitatively assess and determine the differences in relative forces of various cases. Mean values of responses on the 5-point scale ranged from 4 to 5 (Figure 1), indicating that this simulator has the potential to benefit students and repeated practice can enhance the educational process. From our questionnaire results, we concluded that the simulator provided students with relevant knowledge and experience on the various sensations felt while drilling the posterior mandibular bone.

Figure 1 Students’ responses on questionnaire about the simulator (N=24)

References 1) Kinoshita H, et al. Journal of Dental Education 2016 Jan;80(1):83-90.

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Dimensional accuracy of mandibular models created by personal 3D printers and CT data

Ishida Y1, 2*, Miyasaka T2, Aoki H2, Aoyagi Y2, Miura D2, Kawai T3 and Asaumi R3 1, 2, 3 Nippon Dent. Univ., Tokyo, Japan *[email protected]

Introduction: Recently 3D printing technology has become commonly used and the performance is getting to extremely improved. Along with the reduction of the price, personal 3D printers using thermo-fusion method or stereo-lithograph method became readily available. The official performance of recent personal devices was approaching the performance of high-end industrial devices. Thus, if it could be possible to create dental models with CT data by personal devices, it would be helpful in the surgical planning or the explaination to patients. In this study, the dimensional accuracy of mandibular models created by personal 3D printers and CT data was investigated. The possibility of personal devices to dental applications was examined by comparison with high-performance industrial devices.

Materials and Methods: The dried human mandibular bone (hereafter abbreviated as mandibular) was used for the CT imaging. At that moment, six makers made of radiopaque-resin (Scanning resin, Yamahachi Dental, Aichi, Japan) were attached as the measure points. The makers were attached on the height of contour and in front of the both mental foramen on labial and lingual side. CT imaging of the mandibular with six makers was performed by dental CT (AZ3000CT, Asahi Roentgen, Kyoto Japan, hereafter abbreviated as CBCT) and medical CT (Aquilion, Toshiba Medical Systems, Tochigi, Japan, hereafter abbreviated as MSCT). CT data was processed by the software (Mimics, Materialise, Leuven, Belgium), then STL file was made from CT data. The mandibular models were created by the personal 3D printer, i.e., a thermo- fusion device (MAESTRO, ALT design, Taipei, Taiwan) and a stereo-lithograph device (Nobel 1.0, XYZ printing, New Taipei, Taiwan). These models were scanned by a 3D scanner for the dental technician (ceramill map 400, Koblach, Australia) and a 3D scanner for the oral cavity (Lava C.O.S., 3M, Saint Paul, MN, USA). The software (GOM Inspect V8, GOM, Braunschweig, Germany) was used to measure the distances between makers on mandibular and models from the STL data obtained from the scanner. The difference in distance and change rate between mandibular and models was calculated. In our previous study, models were created by two high performance industrial devices (Objet Eden260V, Stratasys, Minneapolis, MN, USA and Projet 3500HD, 3D Systems, Rock Hill, SC, USA) and investigated similarly. The models made by personal devices were compared to those made by the high performance industrial devices. Three different mandibulars were used to perform the CT imaging and to create the models. For the statistical treatment of the results, a four- way ANOVA were performed. Tukey’s multiple comparisons tests were performed when a significant was observed for main factors or their interaction in the four-way ANOVA.

Results and Discussion: Change rates of the imaging obtained by CBCT were smaller than those obtained by MSCT. It might be caused by the size of voxel. Change rates of models created by two personal 3D printers were on the almost same level with those created by high performance industrial devices. From these result, the personal 3D printers seemed to have enough performance to create the mandibular model.

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The accuracy of intraoral and extraoral scanning Shimizu S1*, Shinya A1, 2, Niitsuma A1, Kuroda S1, Gomi H1 1Nippon Dental Univ, Tokyo, Japan 2Truk Univ., Truk, Finland *[email protected]

Introduction： In recent years, digital scanning with intraoral and extraoral scanner have been significant development in the field of dentistry. The quality of intraoral scanning directly influences the fitting of prosthesis. A good internal and marginal fit seems to be one of the most important clinical factors for the long-term success of a prosthesis. The aim of this study was to evaluate the accuracy of intraoral and extraoral scanning.

Materials and Methods： A master model measured using a 3-dimensional coordinate measuring instrument was used as the reference data. For the control group, the master model was scanned 10 times with an extraoral scanner (D810, 3Shape [D]). The experimental groups comprised the same master model scanned 10 times by using two intraoral scanners (Cerec AC Omnicam, Sirona Dental Systems [AC]; TRIOS, 3Shape [TR]). The data were exporting stereolithography (STL) files. The STL files were analyzed using a 3-dimensional analysis software (GOM-Inspect, GOM). The accuracy consisted of its trueness (the extent to which the test scan data differ from the reference scan data) and precision (the similarity of the data from multiple scans). The STL files were measured to determine the trueness and precision. The measurement points were the margin, axis, and height. The 2.0, 3.0, 4.0, 5.0 mm far from the occlusal were measurement points at the axial wall. The results were analyzed using one-way ANOVA (p < .05).

Results： The axial measurement points obtained smaller than the reference data in all groups. At the margin, AC were smaller than the reference data, while those TR and D were larger. At the height, AC and D were smaller than the reference data, while those TR were larger. The precision was similar value in each group.

Conclusion： Intraoral and extraoral scanning showed similar value in this study. At the axial, all groups showed smaller value than the reference data. At the height, except for TR were smaller than the reference data.

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Evaluation of adhesion of bonding agents for CAD/CAM processing

Niizuma Y1*, Kobayashi M1, Miyazaki T1, and Manabe A1 1Showa Univ., Tokyo, Japan *[email protected]

Introduction: Our research recently reported that sandblasting and silanization significantly increased shear bond strength values for bonding between CAD/CAM blocks and resin composite. The aim of this research was to evaluate the shear bond strengths of different CAD/CAM blocks with six universal type self-etching adhesive agents.

Materials and Methods:

Table 1 CAD/CAM Blocks Code Material Manufacturer Filler wt% Matrix resin LU LavaUltimate 3M ESPE Silica nano filler , Zirconia nano filler 80 BisGMA, UDMA EN VITA ENAMIC VITA Ceramics network(Al) 86 UDMA

Table 2 Surface Treatments Code Material Manufacturer CB Clearfil®photo bond Kurary Noritake Dental SB Scotchbond™Universal Adhesive 3M ESPE GB G-Premio BOND + G-Premio BOND DCA + G-Multi PRIMER GC EB ESTELINK BOND + Universal Primer Tokuyama Dental AB ALL UNIVERSAL BOND BISCO PO Prelude One Danville

The materials used in this research are listed in Table1 and 2. The surface of block sandblasted with alumina-oxide particles of which mean particle size was 110μm using airborne-particle abration for 5 s .Following adhesive application, resin composite (Cleafil Majesty LV) was incrementally built up on the surfaces. Shear bond strength test was done using a universal testing machine (Instron) , after bonding specimens were stored in deionized water at 37℃ for a day(n=5).Shear bond strength values were analyzed with Turkey’s multiple comparison test(p<0.05).

Results and Discussion: EN group, the shear bond strength showed the high value in EB, CB, AB, SB, AB, PO, GB order. There were significantly different between EB and GB. LU group, the shear bond strength showed the high value in AB, CB, SB, EB, GB, PO order. There were significantly different between AB and PO. The effective universal type self-etching adhesive agents was found to differ depending on the composition.

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Fracture strengths of composite resin crowns made with dental CAD/CAM system

Okada R*, Ando A, Asakura M, Ozawa S, Takebe J, Kawai T, Ban S Aichi Gakuin Univ., Aichi, Japan * [email protected] Introduction: CAD/CAM technology has been widely applied to dental clinics. Various materials are employed to the restoratives made with CAD/CAM systems. Especially, CAD/CAM composite resins were rapidly expanded in Japanese Dental Treatment. Because this material was employed as the approved one for health insurance in Japanese Government. However, there are a few reports about the strength in a clinical form. The aim of this study was to investigate the relation between the fracture strength and biaxial flexural strength of CAD/CAM composite resin crowns of the maxillary first molar form before and after the thermal aging test. Materials and Methods: Four kinds of CAD/CAM composite resins (Block HC, KZR-CAD HR, KZR-CAD HR2, Avencia Block) and a lithium disilicate glass ceramic (IPS e.max CAD) were used in this study. The abutment tooth model (SUS304) of the maxillary right first molar form was scanned and created the STL date. Each blocks were milled using each CAD/CAM system. Various crowns were immersed in physiological saline solution for 28 days in thermostatic bath at 90°C. The crowns were cemented to the abutment tooth model using an adhesive resin cement (Estecem, Tokuyama Dental). Fracture strengths were measured by the loading of a steel ball with a diameter of 5.0mm via a transparent polyethylene sheet having a thickness of 0.04mm at the central fissure. It was performed with a universal testing machine (4481, INSTRON). The maximum forces to produce the fracture were recorded. Biaxial flexural strengths were measured according to ISO 6872: 2008 ‘Dentistry–Ceramic materials.’ Fractured surfaces of the specimens were observed using a scanning electron microscope (JXA-8530FA, JEOL). Results and Discussion: Fracture strengths of CAD/CAM composite crowns varied between 2490 and 3600 N after thermal aging test, and slightly decreased with the thermal aging test. On the other hand, biaxial flexural strengths significant decreased with the thermal aging test. It seems that the influence of the thermal aging test on the strength depends on the thickness of specimens.

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Strain Analysis of Mandibular Implant Overdentures in Different Dislodging Position with Different Implant Number

*Kaidiliya Y, Kanazawa M, Takeshita S, Oda K, Tanoue M, Miyayasu A, ThuyVL, Thu KM, Asami M, Shimizu H, Shimada R, Minakuchi S. Gerodontology and Oral Rehabilitation, Tokyo Medical and Dental University (TMDU).Tokyo. Japan ＊E-mail：[email protected]

Introduction: In the case of mandibular implant overdenture (IOD), the number of implants to support the prosthesis is the most important question to be answered. Strain around implant abutments have negative effect on implant overdenture. The aim of this study was to investigate the influence of implant number on strain exerted on implants retaining a mandibular overdenture under dislodging movement.

Material and Methods: An edentulous mandibular test model with artificial mucosa and an experimental overdenture were fabricated. Locator attachment systems with pink nylon inserts were chosen for this study. Five implant replicas were placed in the anterior midline, bilateral lateral incisor regions and bilateral canine regions of the mandibular model. Five metal caps were incorporated to the intaglio surface of the experimental overdenture. Three implant positions were prepared: 1-IOD (midline), 2-IOD (lateral incisor regions) and 3-IOD (the midline and bilateral canine regions). Three chains connected to the denture and 3 kinds of dislodging tests (Anterior dislodgement, Posterior dislodgement and Anterior-Posterior dislodgement) were applied. The strain was recorded using strain gauges attached to the Locator abutment. Six measurements were performed at each dislodging condition with 3 kinds of implant positions (N=6). Resultant strains were converted to the lateral forces using strain-force relations obtained from the calibration test. Lateral forces were compared between the mid-anterior implant of 1-IOD, the left lateral implant of 2-IOD, the mid-anterior implant of 3-IOD and the left canine implant of 3-IOD. Statistical comparison was made using a one-way ANOVA and Tukey’s test (α<.05).

Results and Discussion: On the each of dislodging condition, 3-IOD showed significantly larger lateral force to the both abutments than 1-IOD and 2-IOD.Statistically significant difference was found between all implant positions.

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Clasp fabrication using one-process molding by repeated laser sintering and high-speed milling

Nakata T*, Kurihara D*, Ohkubo C* Department of Removable Prosthodontics Tsurumi University School of Dental Medicine, Yokohama, Japan *[email protected]

Introduction: Regarding the use of CAD/CAM to fabricate a prosthetic framework, the milling process would not match the fabrication of a removable partial denture (RPD) framework with a clasp because it has a long, thin shape and undercut. The surface of a framework manufactured by laser sintering is considerably rougher than one manufactured by milling. To resolve these problems, a single machine platform that integrates repeated laser sintering and high-speed milling for one-process molding has been developed. The purpose of this study was to evaluate the Akers clasp assembly prepared with Co-Cr alloy particles by one-process laser sintering and milling as compared to the conventional casting method.

Materials and Methods: After the 18-8 stainless steel die was digitally scanned using a dental laboratory scanner, an Akers clasp was designed using the CAD system (DWOS Partial Frameworks, Dental Wings) and fabricated using a process molding machine with repeated laser sintering and high-speed milling (LUMEX Advance-25, Matsuura) using 50-m Co-Cr powder (Matsuura Cobalt Chromium, Sandvik) (CAM clasp). As controls, cast Akers clasps of the same sizes were also prepared using conventional casting methods with a Co-Cr alloy and CP titanium. After a nondestructive inspection by taking radiographs, the surface roughness and the gap distance between the clasps and the cask-forming die were measured using the silicone film method. The initial retentive force and changes in retention up to 10,000 cycles were also measured. The data were analyzed by two-way ANOVA and Tukey’s multiple comparison test (α=0.05).

Results and Discussion: CAM clasps exhibited smoother surfaces (0.6 m) than those of cast Co-Cr and CP Ti clasps (p<0.05). The gap distances of the CAM clasps were significantly greater than those of the cast clasps (p<0.05). All clasps indicated similar initial retentive forces (cast Co-Cr: 12.9 ±3.5 N; cast CP Ti: 10.9±3.4 N; and CAM clasp: 12.3±2.6 N) without significant differences (p>0.05). The retentive forces of the cast Co-Cr and CP Ti clasps indicated a remarkable decrease from the initial retentive forces to 2,000 insertion/removal cycles. In contrast, the CAM clasps demonstrated a constant or slight decrease from 1,000 up to 10,000 cycles. These results suggest that the CAM clasp made by repeated laser sintering and high- speed milling can be used effectively as an RPD component.

References: Shimpo H. Effect of arm design and chemical polishing on retentive force of cast titanium alloy clasps. J Prosthodont, 2008;17(4):300-7.

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Supramolecular Sulfonated Polyrotaxanes complexed with Bone Morphogenetic Protein-2 for Bone Tissue Engineering

Terauchi M1, Tamura A1, and Yui N1*

1Tokyo Medical and Dental Univ., Tokyo, Japan *[email protected]

Introduction: Although bone morphogenetic protein-2 (BMP-2) has received considerable attention because of its strong osteoinductivity, the clinical application of BMP-2 is limited due to its degradation and deactivation under physiological conditions. Negatively charged sulfated polymers such as heparin are known to form polyelectrolyte complexes with positively charged BMP-2 to prevent deactivation and to enhance the osteoinduction capability of BMP-2. Herein, we report the development of a new modality of sulfonated polymers based on supramolecular polyrotaxanes (PRX) composed of α-cyclodextrins threaded onto a linear polymer. The sulfonated PRXs (SPE-PRXs) formed nanoparticle-like polyelectrolyte complexes upon mixing with BMP-2, and are expected to exhibit excellent osteoinductivity in both in vitro and in vivo studies.

Materials and Methods: The bioactivity of SPE-PRX/BMP-2 complexes was examined by alkaline phosphatase (ALP) production, the gene expression of RUNX2, osterix and osteocalcin, and mineralization in osteoprogenitor cells (MC3T3-E1 cells) in vitro. For in vivo experiments, the full-thickness calvarial defect with a diameter of 3.5 mm was made on the lateral parietal bone in mice by a biopsy punch and a collagen sponge soaked with the SPE-PRX/BMP-2 was placed in each defect. Bone healing in each mouse was evaluated by scanning the computed X-ray microtomography (μ-CT) at 1, 2, 3, and 4 weeks after the operation. The percentage of the bone healing area for each defect was calculated on a scatter plot for each sample.

Results and Discussion: In the in vitro experiments, the SPE-PRX/BMP-2 complexes showed significantly higher in the ALP production, the osteoinductive gene expression and the deposition of mineralized matrix in MC3T3-E1 cells than the heparin/BMP-2 complexes. In the in vivo experiments, the SPE-PRXs/BMP-2 was found to show higher scores than heparin/BMP-2 and BMP-2 at 4w after the operation.

Conclusions: It is suggested that the SPE-PRXs are promising candidates for enhancing osteoinductive ability of BMP-2 without toxicity and anticoagulant activity and could contribute to clinical bone regeneration. Further studies are now in progress with SPE-PRX hydrogels for implantation.

References: Terauchi M, Ikeda G, Nishida K, Tamura A, Yamaguchi S, Harada K, Yui N. Macromol. Biosci. 2015, 15, 953-964.

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Release of FGF-2 from 4-META/MMA-based adhesive resins containing non-biodegradable FGF-2-loaded-polymer particles

Tsuboi R1*, Kitagawa H1, Takeda K1, Sasaki JI1, Takeshige F1, Imazato S1 1Osaka Univ., Osaka, Japan *e-mail ([email protected])

Introduction: Aiming at achievement of bio-active dental resins which promote healing of the surrounding tissues, we have developed poly-2-hydroxyethyl methacrylate (HEMA)/trimethylolpropane trimethacrylate (TMPT) particles and reported that they were successfully used as the non-biodegradable carrier for fibroblast growth factor-2 (FGF-2). To obtain optimum polymer particles to provide dental resins with the ability to release FGF-2, here we fabricated novel non-biodegradable particles consisting of polyethyleneglycol monomethylether monomethacrylate (PEGMA) and TMPT. In this study, FGF-2 release from PEGMA/TMPT- and HEMA/TMPT-polymer particles loaded with FGF-2 was compared, and the release profiles of FGF-2 from the cured 4-META/MMA-based adhesive resins incorporating each polymer particle were determined.

Materials and Methods: The mixture of PEGMA/TMPT or HEMA/TMPT at the ratio of 90/10 by weight was cured and pulverized into the powders with the average size of 500 μm. To determine the water absorbability, the polymer particles were immersed in distilled water for 4 days, and the water absorption ratio and water contents were calculated by measuring the weight. FGF-2 was loaded to each polymer particle by immersion in 500 µg/mL FGF-2 solution (Fiblast, Kaken) for 24 hours. The particles were then freeze-dried and the concentrations of FGF-2 released into distilled water were measured using a micro BCA protein assay. The PEGMA/TMPT- or HEMA/TMPT-polymer particles loaded with FGF-2 were added to a commercial 4-META/MMA-based adhesive resin (Super-Bond C&B, Sun Medical). The cured resin disc (5 mm diameter and 0.5 mm thickness) was immersed into distilled water, and the release of FGF-2 was determined until 14 days.

Results and Discussion: The PEGMA/TMPT-polymer particles demonstrated greater water absorption ratio and water content than the HEMA/TMPT particles. Due to such difference in hydrophilic nature, the total amount of FGF-2 released from the FGF-2-loaded-PEGMA/TMPT particles were significantly greater (Student’s t-test, p < 0.05) than those from the FGF-2-loaded-HEMA/TMPT particles. While both resins containing FGF-2-loaded particles made of PEGMA/TMPT or HEMA/TMPT exhibited continuous release of FGF-2 up to 14 days, the concentrations of FGF-2 released were greater for the PEGMA/TMPT particles.

Conclusion: It is possible to achieve a sustained release of FGF-2 from the 4-META/MMA-based adhesive resins by incorporating FGF-2-loaded polymer particles. Incorporation of the FGF-2-loaded-PEGMA/TMPT-polymer particles was effective to obtain greater release of FGF-2 than the HEMA/TMPT particles.

This study was supported by a Grant-in-Aid for Scientific Research 26293409 from the JSPS.

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Comparison of antibacterial activities of Xanthorrhizol and Bakuchiol with and without fluoride varnish Son JL1*, Kim AJ2, Oh SH1, Bae JM1 1 College of Dentistry, Wonkwang Univ., Iksan, Korea 2 College of Sorabol, Gyengju, Korea *e-mail ([email protected]) Introduction: Fluoride varnish is known to be the most effective method to prevent dental caries. However, antibacterial activity of fluoride varnish is not well known. Incorporation of natural antibacterial agent into fluoride varnish would increase antibacterial effect against microorganism related to dental caries. Xanthorrizol has been reported to show excellent antibacterial effect against Streptococcus mutans. Bakuchiol was also published to have bactericidal effects on oral microorganism. The purpose of the present study was to investigate the antibacterial effect of Xanthorrihizol and Bakuchiol incorporated into fluoride varnish on S. mutans.

Materials and Methods: Experimental fluoride varnish was fabricated with rosin base, ethanol and 5 wt% NaF. Antibacterial effect against S. mutans was evaluated using agar diffusion test, minimum inhibitory concentraion(MIC) and minimal bacterial concentration(MBC). Antibacterial activities of Xanthorrhizol and Bakuchiol with and without experimental fluoride varnish were evaluated. The diameter of paper disc used in agar diffusion test was 6 mm. Using 1.1 × 1010 CFU/ml of S. mutans, MIC and MBC test were conducted.

Results and Discussion: Agar diffusion test MIC MBC (mM) (mM) 1 mM 10 mM Experimental fluoride varnish (EFV) 7.16 mm Xanthorrhizol 6 mm 7.75 mm 0.125 0.25 Xanthorrhizol+ EFV 9.14mm 10.62 mm 0.0062 0.125 Bakuchiol 8.9 mm 14.5 mm 0.015 0.031 0.031 0.062 Bakuchiol + EFV 8.53 mm 10.10 mm

In agar diffusion test, the antibacterial effect of Xanthorrihizol was significantly increased when incorporated with fluoride varnish compared (P<0.05) with Xanthorrhizol itself and EFV(P<0.05) . MIC and MBC of Xanthorrihizol decreased with fluoride varnish. However, the antibacterial effect of Bakuchiol was not significantly different (P> 0.05 ) or rather decreased when incorporated with fluoride varnish. MIC and MBC of Bakuchiol increased with fluoride varnish.

Conclusions: Xanthorrihizol incorporated in fluoride varnish show significantly higher antibacterial effects when incorporated into fluoride varnish than itself through agar diffusion test, MIC and MBC. Therefore, Xanthorrihizol is considered to be used with fluoride varnish effectively as a natural antibacterial agent to prevent dental caries.

Acknowledgements (Optional): This research was supported by Basic Science Research Program through the National Research Foundation of Korea(NRF) funded by the Ministry of Science, ICT and future Planning (2015R1A2A2A01007393).

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Effect of Mesenchymal Stem Cells on Mixed Self Assembled Monolayer

Hirata I1*, Veronica SR1, Tania S1,2, Yamauchi Y1, Kanawa M1, Kato Y1,3, Kato K1 1Hiroshima Univ., Hiroshima, Japan 2Airlangga Univ., Surabaya, Indonesia 3TWO CELLS Co.,Ltd., Hiroshima, Japan *e-mail : [email protected]

Introduction: Mesenchymal stem cells (MSCs) has the ability to differentiate into a variety of cells such as bone, cartilage, and fat. There are many studies aimed at a variety of tissue regeneration. Serum free media (STK series; TWO CELLS, Hiroshima University-launched venture company) adopt to primary and passage culture of MSCs and they have been used not only at research but also clinical field recently. A common cell culture plate (TCP), however, are used in MSCs culture and an optimization of the culture substrate has not been performed. Self-assembled monolayers (SAMs) made of alkanethiols with a long methylene chain give well-defined model surfaces for studies on interfacial phenomena and intermolecular interactions. In this study, we report the optimization of the surface components for MSCs culture and the differentiation in gene expression and metabolism behavior of MSCs culture on between TCP and SAMs.

Materials and Methods: Gold substrates were cleaned by immersion in piranha solution and washed with ultra pure water. Then, 1 mmol/L of 11-amino-1-undecanethiol (NH2), 11-mercapto-1-undecanol (OH), 11-mercapto-1-undecanoic acid (COOH), and 1-dodecane-thiol (CH3) solutions were prepared in ethanol. These alkanethiol solutions were mixed at various ratios. The gold plates were immersed in these solutions for 24 hours, and the mixed SAMs were formed on the surface of the gold substrates. MSCs were cultured on the mixed SAMs in a medium containing serum and in a serum-free medium (STK1 and 2) and evaluated between surface composition and cell proliferation using a cell counting reagent WST-8. The differentiation in gene expression and metabolism behavior of MSCs culture on between TCP and SAMs were evaluated by DNA micro allay (Agilent SurePrint G3 Human Gene Expression 8x60K v2 Microarray) and total-metabolome analysis (C-Scope; Human Metabolome Technologies).

Results and Discussion: Various kinds of proteins are contained in serum but their concentrations are not completely defined, whereas STKs are well-defined solution. On mixed SAMs, the patterns of cell proliferation were quite different between the two cases, and cell proliferation was more noticable in STK2 than serum contained medium. From the results of gene expression and metabolism analysis, a mitochondrial activity of MSCs on the SAM was lower than on TCP and a glycolytic activity on the SAMs was higher. Many studies reported the activation of glycolysis and the suppression of mitochondrial activity for stemness maintaining of stem cells. Therefore, the SAM was expected to be excellent in the maintenance of MSCs stemness. MSCs culture system using serum-free medium is not only improvement of MSCs proliferation but also maintain a stemness using the SAM compared with TCP. We expect this well-defined culture system is useful for research and clinical application of MSCs.

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Evaluation of degradability of EGCG-conjugated gelatin sponge

Honda Y1*, Tanaka T2, Hashimoto Y1, Imai K1, and Arita K1 1Osaka Dental Univ., Osaka, Japan 2Kyoto Institute of Technology., Kyoto, Japan *[email protected]

Introduction: Epigallocatechin-3-gallate (EGCG), the main component of green tea catechins, is a promising natural polyphenol for application in promoting health care [1]. The molecule is extensively investigated for the treatment of various diseases like cancer [2] and oral diseases [3], because of its low cost and pharmacological effects. Nevertheless, few studies have been performed on the application of EGCG in bone regeneration therapy. Recently, we have fabricated EGCG-conjugated gelatin (EGCG-Gel) sponge, applicable to bone regeneration therapy [4]. The fabricated complex containing the appropriate amount of EGCG could effectively facilitate the bone regeneration even in vivo and can promote the osteoblastogenesis in mouse mesenchymal stem cell line in vitro [4]. In the present study, we have evaluated the degradability of the EGCG-Gel sponge synthesized under multiple conditions to estimate their robustness.

Materials and Methods: EGCG-Gels were synthesized using aqueous chemical synthesis methods with 4-(4,6- dimethoxy-1,3,5-triazin-2-yl)-4-methyl-morpholinium chloride (DMT-MM) and N- methylmorpholine (NMM). Contents of EGCG, DMT-MM, and NMM were altered to prepare the different EGCG-Gels. After the syntheses, the products were subjected to dialysis in water and lyophilized to prepare the spongy morphology. A scanning electron microscope (SEM) was used to observe the porous structure and the surface of the complexes. Fourier- transform infrared (FTIR) spectroscopy was used to identify the existence of EGCG in the complexes. To verify the robustness of the product in water, EGCG-Gels were immersed in distilled water under stirring up to day 28.

Results and Discussion: During synthesis, adding excess amounts of EGCG, DMT-MM, and NMM completely hindered preparation of the sponge structure, producing powder morphology. SEM observation of the EGCG-Gel having spongy structure revealed macropores in the complex. Characteristic spectral features of EGCG could be detected in the FTIR spectra of EGCG-Gel, suggesting that the EGCG was successfully retained in the product even after the dialysis treatment. Degradability of the EGCG-Gel sponges was controlled by the amount of EGCG used during synthesis. These results suggest that the appropriate amount of EGCG in the EGCG-Gel enhanced the robustness of the product, possibly due to the crosslink between the Gels and EGCG.

References: [1] Mereles D et al., Int J Mol Sci, 12, 5592 (2011) [2] Park G et al., J Invest Dermatol., 128, 2429 (2008) [3] Venkateswara B et al. J Indian Soc. Peridontol., 15, 18 (2011) [4] Honda Y et al., Int J Mol Sci, 16, 14143 (2015)

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ｙRedox-Responsive Supramolecular Polymeric Prodrug for Cancer Chemotherapy

Wang Y1, Sun R2, Luo QJ3, Li XD1* 1 The Affiliated Stomatology Hospital, Zhejiang Univ., Hangzhou, China 2 Department of Polymer Science and Engineering, Zhejiang Univ., Hangzhou, China 3 The First Affiliated Hospital, Zhejiang Univ., Hangzhou, China * email: [email protected]

Introduction: Oral squamous cell carcinoma is one of the most common cancers with high rate of mortality and morbidity. Although chemotherapy plays a vital role in its clinical therapy, it still faces some challenges, such as poor tumor selectivity and severe side effect. To date, with the deepening of nanotechnology, self-assembled nano-prodrugs have emerged as novel stimuli responsive biomaterials for drug delivery due to their improved pharmacokinetics via the enhanced permeability and retention effects. Herein, based on biocompatible cyclodextrin terminated poly(ethylene glycol), we constructed redox-responsive core crosslinked supramolecular prodrug nanoparticles by host-guest interaction and in situ crosslinking. Materials and Methods: Camptothecin, one of the most potent chemotherapeutic agents, was used as a model anticancer drug to conjugate with adamantane through a reduction-cleavable disulfide linker, giving the redox-responsive core crosslinked supramolecular prodrug nanoparticles via the host-guest interaction between cyclodextrin and adamantane. The structure, size, morphology and stability of these prodrug nanoparticles were evaluated, and the drug release profiles at different GSH concentrations were determined. The cellular uptake, intracellular drug release and anticancer effect in vitro were also confirmed. Results and Discussion: These nano-prodrug particles had stable nano-scaled spherical structure with narrow distribution. In vitro release study illustrated a minimized release of camptothecin at physiological environment, while a rapid release under reductive environment. Confocal laser scanning microscopy, flow cytometry analysis and CCK-8 assay demonstrated that these nano-prodrug nanoparticles could rapidly enter cancer cells and release drug in response to intracellular reductive environment, showed strong anticancer effect, which may expected to be promising candidates as nano-drug carriers for cancer chemotherapy.

Figure 1. Size, morphology (A) and in vitro release profiles (B) of nano-prodrug particles. Acknowledgements: This study was financially supported by the National Natural Science Foundation of China (51173163), the Natural Science Foundation of Zhejiang Province (LZ16H140001) and China Postdoctoral Science Foundation (2015M580517).

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The expression of NF-kB on rat tooth mechanically exposured after administration of pomegranate extract

Intan Nirwana, Titien Hary Agustantina and Helal Soekartono Dental Materials Department, Faculty of Dental Medicine, Universitas Airlangga Surabaya Indonesia Correspondence : Intan Nirwana, c/o: Dental Materials Departmentnt, Faculty of Dental Medicine, Universitas Airlangga, Jl. Mayjend. Prof. Dr. Moestopo 47 Surabaya 60132, Indonesia. E-mail: [email protected]

Background: The use of calcium hydroxide as a direct pulp capping material still showed inflammation, so the alternative materials are required, one of them is pomegranate extract that has an anti-inflammatory activity. It has the main content of ellagic acid. Nuclear factor kappa beta is a transcription factor that regulates many genes such as IL-1, IL-2, IL-2 receptor, IL-6, IL-8, intercellular adhesion molecule-1, TNF and interferon-γ. Mechanically exposured tooth will show inflammation and enhance NF-kB expression. Pomegranate extract is expected to decrease NF-kB expression. Activity of pomegranate extract was observed by measuring the expression of NF-kB.

Objective: This study was to analyze the activity of pomegranate extract on rat tooth mechanically exposured on the expression of NF-kB.

Methods: Eighteen male rats (Rattus norvegicus), aged 2.5 months and has a weight of about 190-230 grams were divided into 3 groups. The first group (K) is a rat tooth mechanically exposured group which given with gel carboxy methyl cellulose (CMC) 3%, the second group was given with calcium hydroxide, the third group was given with pomegranate extract. The expression of NF-kB was examined using immunohistochemical techniques after one day of treatment. Data were analyzed using ANOVA and HSD.

Result : This study showed a significant difference between the control and the treatment group (p < 0,05).

Conclusion: The administration of Pomegranate extract on the rat tooth mechanically exposured decreased NF-kB expression.

Kata kunci: pomegranate extract, mechanically exposured tooth, NF-kB expression

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Reasons for failures of posterior restorations

Kubo S*, Sumi T, and Hayashi Y Nagasaki Univ., Nagasaki, Japan *e-mail: [email protected]

Introduction: In order to extend the longevity of dental restorations, an investigation of the reasons for failures may be important since it can lead to improvement of restorative materials and techniques. However, with respect to failure modes of posterior restorations, only a few studies have been available. The purpose of this cohort study was to examine the reasons for failures of posterior resin composites (CR) and cast metal restorations (CM). The median ages of failed restorations by failure modes were also estimated.

Materials and Methods: All patients involved in this study were under charge of a principal investigator (SK). Most (84%) of them have been regularly checked up twice or three times a year for more than 10 years. Reasons for failure of posterior restorations (crowns were not included), which were retreated between 1995 and 2015, were retrospectively investigated using the patient records from July to December in 2015. Survival times of failed restorations were estimated by the Kaplan Meier method using retreated restorations placed in Nagasaki University Hospital. The Cox proportional hazards model was used to study the dependence of survival times on potential explanatory variables and controlled confounding factors.

Results and Discussion: Data of 349 failed restorations [125 for resin composites (CR) and 224 for cast metal inlays/onlays (CM)] were obtained and used for the statistical analysis. Two hundred and fifty-eight retreated restorations were placed in 83 patients by 25 dentists in the university hospital and 91 restorations in 28 patients had been placed by unknown dentists in other dental offices. The Fisher’s exact test revealed significant differences in percentage distribution of reasons for retreatment between CR and CM. For CR, caries, loss of restorations and fracture of restorative materials occurred with almost the same frequency. For CM, the loss was the most failure mode though it was often accompanied by caries and/or marginal fracture. CR was more sensitive to variables compared to CM, and the number of restored surface, retreatment risk, tooth type and age at placement had a significant influence on its failure modes. Among them, tooth type had the greatest influence. Caries was a main reason for premolars, whereas fracture and loss were two to three times as many as caries for molars. This is probably due to the easiness of caries detection and masticatory forces. On the other hand, only the number of restored surfaces had a significant influence on the failure modes of CM, showing the more surfaces restored, the more tooth fractures occurred. There was no significant difference in the age of failed restorations between CR (4.0 y) and CM (4.9 y). However, the Cox model indicated that CM tended to show a significant longer survival time of failed restorations as reported by our prospective study on the longevity of posterior restorations. In addition, the Cox model revealed that retreatment risk, reasons for retreatment and Class (number of restored surfaces) had a significant influence of survival time of failed restorations.

Acknowledgements: This study was supported in part by JSPS KAKENHI Grant Number 26462888

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Improvement in oral environment of the denture wearer by neutral electrolyzed water

Shimizu H1, Nagamatsu Y1*, Nagamatsu H1, Murakami S2, Nakamura K1, Ikeda H1, Komagata Y1, Masumi S1, Kozono Y1 1Kyushu Dental Univ., Kitakyushu, Japan 2Kyushu Univ. of Nursing and Social Welfare, Tamana, Japan *[email protected]

Introduction: Several types of electrolyzed water have been widely used in clinical dentistry because of their superior bactericidal activities. We reported that they showed high bactericidal efficacies for disinfection of impression, denture and dental instruments. It was also reported that neutral type is the most appropriate for use in dental practice from the point of no or little metallic corrosion and no decalcification of enamel unlike acid type. In this study, usefulness of neutral electrolyzed water for the improvement in oral environment of healthy or care- required denture wearers aged 50s-90s was examined in terms of disinfection effect on denture.

Materials and Methods: Neutral electrolyzed water (NW, pH:7.0±0.05, residual chlorine 38±0.4ppm) was automatically prepared using tap water containing 5% sodium chloride by an electrolysis apparatus (APaqua21, Asahipretec., Kobe, Japan). Eight dentures in total out of 5 or 6 subjects per group were examined. Group A: Healthy denture wearers taking oral care by own usual method (age: 53-76). Group B: Healthy denture wearers visiting the university hospital (age: 69-84) Group C: Denture wearers requiring long-term care (age: 84-96) Numbers of surviving bacteria on the surface of subject's denture before and after the ultrasonic cleaning in NW were examined according to the method of Taniguchi et al.1). This clinical experiment was conducted by the approval of the Research Ethical Review Committee of Kyushu Dental University． 6 A 5 Results and Discussion: 4 lower In A and B, less than 3-min treatment showed high 3 upper bactericidal efficacy for all denture used by healthy subjects. 2 On the other hand, 3-min treatment was sufficient for only 3 1 0 dentures out of 8, the other 3 dentures needed 4- or 6-min A1 A2 A3 A4 A5 A6 treatment in C. In the remaining 2 dentures, which were 6 5 B observed denture plaque, about 80% of surviving bacteria 4 were removed by 6-min treatment; however, they needed 3 further treatment to get sufficient efficacy. These findings 2 suggest that it is important to set an appropriate treatment 1 0 time in each denture according to the degree of B1 B2 B3 B4 B5 6 independence in denture cleaning. C >6 >6 5 Treatment time (min) 4 Acknowledgements: This study was supported in part by a 3 research grant from Kyushu Dental Association. 2 1 0 References: 1) Taniguchi M. et al.: Sterilization of Denture C1 C2 C3 C4 C5 with Electrolyzed Neutral Water. J.Kyushu.Dent. 62; 29-38, Fig.1 Treatment time removing 2008. more than 95% of surviving bacteria attached to denture

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Bone response to multilayered DNA/protamine coated titanium implants

Sakurai T1*, Yoshinari M,2 Toyama T,3 Hayakawa, T1, and Ohkubo C1 1Tsurumi Univ., Kanagawa, Japan 2 Tokyo Dental College, Tokyo, Japan 3 Nihon University, Tokyo, Japan *e-mail [email protected]

Introduction: DNA is a unique biomolecule for novel biomaterials, irrespective of its genetic information. A DNA/protamine (D/P) complex has been proposed as a candidate bone substitute material1). A DNA coating on dental titanium (Ti) implants is attracting attention due to its osteogenic properties. The aim of the present study was to evaluate the in vitro and in vivo bioactivities of a multilayered D/P coated Ti implant by simulated body fluid (SBF) immersion experiments and implantation experiments into extracted sockets of rat molars.

Materials and Methods: Two types of DNA, 300-base pair (bp) and 7000-bp fragments, were used. Protamine was initially immobilized onto Ti implants using a tresyl chloride-activated method2) and DNA and protamine were then alternatively deposited after the immobilization of protamine by a layer-by-layer technique. A multilayered D/P coating was confirmed by X-ray photoelectron spectroscopy (XPS) and Fourier-transform infrared (FT-IR) spectroscopy measurements. As in vitro study, multilayered D/P coated Ti implant was immersed into SBF and the precipitation behavior on multilayer D/P coating was observed. As in vivo study, multilayered D/P coated Ti implant was implanted into the extracted sockets of rat molars. Nondecalcified thin sections were prepared and were stained with methylene blue and basic fuchsin. Bone response after 3 and 9 weeks implantation was histologically and histomorphometrically evaluated by.

Results and Discussion: XPS and FT-IR measurement identified the presence of D/P coating on titanium implants. The deposition of apatite progressed more on the surfaces of multilayered D/P-coated Ti implants than on those of non-treated Ti implants in SBF immersion experiments. Animal implantation experiments showed that multilayered D/P-coated Ti implants provided a significantly higher bone-to-implant (BIC) contact ratio 3 weeks after implantation. No significant difference was observed in the BIC ratio 9 weeks after implantation. The results of the present study demonstrated that a multilayered D/P coating promoted new bone formation at the early stages of the bone healing process.

Acknowledgements: The authors are grateful to Maruha-Nichiro Holdings, Ltd. for supplying salmon testes DNA and protamine.

References : 1) Mori N, Ohno J, Sakagami R, Hayakawa T, Fukushima T. Cell viabilities and biodegradation rates of DNA/protamine complexes with two different molecular weights of DNA. J Biomed Mater Res B Appl Biomater 2013;101:743–751 2) Hayakawa T. Biochemical surface modifications to titanium implants using the tresyl chlorideactivated method. Dent Mater J. 2015;34:725-39

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Bond strength of restoration handling instruments with adhesive and restoration materials Ishida Y1, Igarashi K2, Terada Y2, Okada H1 1Ohu Univ., Fukushima, Japan 2Ohu Univ., Fukushima, Japan [email protected]

Introduction:

Optra Stick (Ivoclar vivadent), a placement instrument that features a flexible adhesive tip, is an instrument with a sticky gel at the tip for handling restorations, This instrument is expected to reduce the risk of accidental swallowing by fall of restorations in the oral cavity and avoid pollution caused by dropping them outside the mouth.

In this study, we examined the tensile bonding strength between the Optra Stick and restoration materials, and the impact of the surface roughness of the materials on the tensile bonding strength.

Materials and Methods:

Testing samples consisted of Optra Stick (Ivoclar vivadent) and plate specimens to be adhered which were processed from ceramic block IPS Empress CAD (ivoclar vivadent), composite resin for crown and bridge PROSSIMO Enamel E3 (GC) or glass. For the measurement of the tensile bonding strength, a universal testing machine (1310DW, Aiko Engineering) was used. An Optra Stick and a plate piece to be adhered were fixed to the top and bottom of the universal testing machine, respectively. After contacting with the adhesive gel to the plate piece with a load of 20gf, tensile bonding strength test was performed at a cross head speed 5.0 mm/min until the adhesive gel and the plate piece separated. Loading was repeated 10 times for one piece of OptraStick. The test was carried out five times for each testing sample. The surface roughness Ra of each plate specimen was measured by 5 times.

Results and Discussion: It was observed that the tensile bonding strength was 79 gf for the Ag-Pd-Cu-Au alloy, 89 gf for the ceramic block, 101 gf for the composite resin, and 80 gf for the glass plate. These values were about 4 to 5 times the applied load. Upon repeating the load, the tensile bonding strength gradually decreased. After the 10th loading, the bonding strength decreased to equal to, or less than, half of that of the first loading. The largest fall in tensile bonding strength occurred between the first and second loadings for all the materials, resulting in significant difference in tensile bonding strength (p < 0.01). The composite resin exhibited slightly greater tensile bonding strength than the other materials, but there was no significant difference between the materials (p <0.01). The surface roughness (Ra) was 406 μm for the ceramic block, 75 μm for the hard resin, 33 μm for the glass plate, and 91 μm for the gold-silver-palladium alloy, demonstrating a significant difference between the ceramic and the other samples ( p <0.01). The results of the adhesion test revealed that all of the materials showed several times higher holding strength than the force applied at the bonding. Further, it also demonstrated that despite significant differences in surface roughness, the differences gave no large impact on the tensile bonding strength.

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Wear Properties of CAD/CAM Hybrid Resin Block

Kai T*, Teramae M, and Nakatsuka T Research and Development Dept., Shofu Inc., Kyoto, Japan *[email protected]

Introduction: With the growing use of dental CAD/CAM systems, clinical applications of hybrid resin blocks (HRBs) have been increasing. The manufacturing methods of HRBs can be roughly classified into two types: the “filler-dispersed” type where the filler is densely filled in the monomer and polymerized; and the “monomer-injection” type where the monomer is impregnated into glass particles and polymerized. In this study we performed antagonistic enamel wear testing (Alabama method) to evaluate wear resistance of HRBs manufactured by the above-mentioned two different methods.

Materials and Methods: We used three “filler-dispersed” type HRBs (HC: SHOFU BLOCK HC, Shofu; CE: CERASMART, GC; and LU: LAVA ULTIMATE, 3M ESPE) and two “monomer-injection” type HRBs (AV: KATANA AVENCIA Block, Kuraray Notirake Dental; and VE: VITA ENAMIC, VITA). Each HRB was milled into a stylus (diameter: 30mm, length: 4.6mm) using SHOFU S-WAVE CAD/CAM System, and the surface of the stylus was buff-polished. The styluses were subjected to wear testing (Alabama method) under specific conditions (immersed in water at 37°C; vertical load: 75N; horizontal rotation: 30°; performed 10,000 times) using bovine enamel pieces (polished with 600-grit) as antagonist teeth. Wear amounts of antagonist teeth (i.e. maximum wear depth of enamel in µm) and self-wear amounts of the HRBs (i.e. decrease in stylus length in µm) were measured (n=6). Data obtained were statistically analyzed by Tukey’s test.

Results and Discussion: The self-wear amounts of the HRBs were in the following ascending order: HC

)

μm ( c

b a,b a,b

wear amount of HRB of amount wear

- Self

Fig.1 Self-wear amounts of HRBs * The bars marked with different letters are significantly different (p<0.05).

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Highly-viscous glass-ionomer cement for filling: Interfacial Gap-formation in Class I restoration and mechanical properties.

Irie M1*, Matsumoto T1, Taketa H1, Torii Y1, Maruo Y1, Nishigawa G1, Minagi S1, Nagaoka N1, Yoshihara K1 1Okayama Univ., Okayama, Japan *e-mail ([email protected])

Introduction: One of the major concerns with highly-viscous glass ionomer cements (HV-GICs) is their ability to achieve effective initial interfacial gap-formation in restorative cavities 1). This in vitro study examined the initial stage (after one-day storage) of interfacial gap-formation in Class I restoration together with determination of associated mechanical properties (compressive strength and flexural strength).

Materials and Methods: Cavity preparation was made in occlusal surface of premolar teeth. Six HV-GICs were studied (Ketac Universal Aplicap, Ketac Molar Aplicap: 3M Oral Care, Fuji IX GP, Fuji IX EXTRA, EQUIA Forte: GC, Zirconomer improved: Shofu, and two conventional glass- ionomer cements (C-GICs, Ketac Silver Aplicap: 3M Oral Care, Fuji II: GC, as controls), with specimen sub-groups (n = 10 / group) for each property measured. After one-day storage and polishing, the restored teeth were sectioned in a mesio-distal direction through the center of the model Class I restorations. The presence or absence of interfacial-gaps was measured at x 1000 magnification at 14 points (each 0.5-mm apart) along the cavity restoration interface; (n=10; total points measured per group =140) 1). Compressive and flexural strengths were measured (n = 10 / group), as described above.

Results and Discussion: For HV-GICs (except Zirconomer improved) and C-GICs, significant differences (p<0.05) in gap-incidence were observed. In the former case, 4-14 gaps were found. In the latter case, 21-24 gaps were observed. The compressive and flexural strengths of HV-GICs significantly increased compared to C-GICs. After one-day storage, a HV-GIC performed significantly better than its corresponding a C- GIC. Increasing the powder-liquid ratio is the primary reason for improving these results. This improvement is achieved by a reduction in the size of the glass particle. A number of variations led to a HV-GIC with improved sealing and mechanical properties.

Conclusions: For HV-GICs, investigation interfacial gap-formation had values compared to C-GICs. And, it is thought that a HV-GIC is the useful and significant restorative material for some pediatric or geriatric patients.

Acknowledgements: This work was supported by JSPS KAKENHI Grant Number 22592183, 26462950.

References: 1) Irie M et al., Class I gap-formation in highly-viscous glass- ionomer restorations: delayed vs. immediate polishing. Oper Dent 33: 196-202, 2008.

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Influence of constant strain on elastic modulus of thermoplastic orthodontic materials

Inoue S1, Yamaguchi S1, Uyama H1, Yamashiro T1, Imazato S1 1Osaka Univ., Suita, Japan *e-mail ([email protected])

Introduction: Thermoplastic materials for transparent aligners have been widely used for orthodontic treatment with their esthetic advantage. However, because of reduction in the orthodontic forces, commercially available aligners are recommended to be replaced every 2 weeks [1]. It is possible that a constant strain in the aligners during teeth movement causes reduction in elastic moduli related to the orthodontic forces. In this study, we investigated an influence of the constant strain on elastic moduli of the commercial thermoplastic materials.

Materials and Methods: Three commercial thermoplastic materials: Essix type A: EA (Dentsply), Duran: DU (Scheu Dental), and Erkodur: ER (Erkodent Erich Kopp) were used. Thirty dumbbell-shaped specimens of each material were fabricated according to the ISO 527-2 type 5B. Fifteen specimens were immersed in distilled water at 37 °C with no strain for 24 hours. For another fifteen specimens, the load was applied to give 1% strain by using an original loading device and stored in distilled water at 37 °C for 2 weeks. Then, tensile tests were performed with a compact universal testing machine (EZ-SX, Shimazu), and elastic moduli were calculated. The results were statistically analyzed by Mann-Whitey U test between the two immersion conditions.

Results and Discussion: Elastic moduli after 24-hour immersion were 705.754±13.947, 684.179±22.495 and 726.265±31.130 MPa for EA, DU, and ER, respectively. By 2-week immersion with strain, elastic moduli were significantly decreased (p < 0.05) for all materials (598.73±74.79 MPa for EA, 641.44±34.492 MPa for DU, and 619.465±75.809 MPa for ER). These results suggest that the orthodontic forces possibly decrease after 2 weeks in the oral cavity, which is caused by the changes of the relaxation modulus of thermoplastic materials during stress relaxation behavior, and the water absorption may accelerate these phenomena.

Conclusions: Three proprietary thermoplastic materials demonstrated decrease of the elastic moduli by the stress relaxation behavior. Development of novel thermoplastic materials with greater resistance against the stress relaxation behavior could be useful to achieve orthodontic aligners which induce optimal and sustained orthodontic forces.

Acknowledgements: This research was supported by a Grant-in-Aid for Scientific Research (No. 15K11195) from the Japan Society for the Promotion of Science (JSPS).

References: 1. Kwon JS et al., Am J Orthod Dentofacial Orthop 2008; 133: 228-34.

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Wear of dental restorative materials under low occlusal force

Kakuta K* and Miyagawa Y Nippon Dental Univ., Niigata, Japan *[email protected]

Introduction: Authors investigated occlusal wear of dental restorative materials which faced a dental porcelain antagonist. In previous investigation, 20, 40 and 60 N were chosen for testing occlusal forces in the occlusal wear test. However, actual occlusal forces varied with occlusal conditions. Especially in a usual mastication, it can be expected that optimum occlusal force to grind foods was not so high. It is necessary to investigate occlusal wear of dental restorative materials under low occlusal force. On the other hand, it is difficult to evaluate occlusal wear under low occlusal force because wear trace on a wear specimen becomes small. The aim of this study was to evaluate wear of dental restorative materials which faced the dental porcelain antagonist under low occlusal force.

Materials and Methods: Five dental restorative materials, namely, two resin blocks for CAD/CAM (Cerasmart, GC, Japan, and Shofu Resin Block HC, Shofu, Japan), a resin-based restorative material for crown & bridge (Estenia C&B, Kuraray, Japan), a dental porcelain (Vintage ZR, Shofu, Japan) and a type 3 gold alloy (Casting Gold, GC, Japan), were used for wear specimens. The dental porcelain was used for an antagonist too. Wear test was performed using a surface property tester (Type 32, Heidon, Japan) in a both-way wear test mode that a spherical tip of the dental porcelain antagonist slid 2 mm on a flat surface of the wear specimen in a reciprocating motion. A thousand reciprocating cycles was performed under 5 N occlusal force. After wear test, a surface profile of wear track on the specimen was measured using a three-dimensional measuring microscope. Wear volume per 1 mm length at center of wear track was computed from the surface profile data. Average wear depth and average wear width obtained at three points of the wear track were measured. In addition, frictional resistance of final reciprocating cycle and an area of wear of the dental porcelain antagonist tip were measured for evaluation of wear process. Measurements were repeated three times and the data were analyzed using one-way ANOVA and Tukey’s multiple comparisons.

Results and Discussion: Results of wear volume were different among the dental restorative materials statistically. The resin-based restorative material for crown & bridge showed the largest wear volume (P<0.01). Wear volume of the dental porcelain was lower than that of the resin-based restorative material (p<0.01). The smallest wear volume were shown in the type 3 gold alloy and the two resin blocks for CAD/CAM (p<0.05). No significant difference was shown among the type 3 gold alloy and the two resin blocks for CAD/CAM (p>0.05). Results of depth and width of wear track--, frictional resistance and wear area of the porcelain antagonist tip were also similar to the results of wear volume. In spite of similar structure of the resin-based restorative material and the resin blocks for CAD/CAM, wear behavior of these materials were extremely different. Differences of wear behavior among dental restorative materials could be detected under comparatively low occlusal force.

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Influence of Shades of Resin Cement on Shades of Laminate Veneer Restorations Evaluated Using Shade Models

Hara D*, Kitada N, Nagafuji A, Shinno K, and Nakatsuka T Research and Development Dept., Shofu Inc., Kyoto, Japan *[email protected]

Introduction: Laminate veneer (LV) restoration has been widely performed because it can improve the esthetics of natural teeth more effectively in a shorter time period, compared to whitening treatment. However, a shade of a tooth restored with LV is more susceptible to the influence of shades of an abutment tooth and cement. With that in mind, we have developed a novel resin cement system for LV restoration, which enables a clinician to freely control the Value(Lightness) of LV restorations, as well as to control the influence of abutment tooth colors. The purpose of this study was to evaluate basic properties of novel resin cement, and also to investigate the influence of the shades of the novel resin cement paste on the Value of the final restorations.

Materials and Methods: We used the novel resin cement system (SI-R21302) for LV restoration, comprising light- cured resin cement paste (RC paste) of three shades that differ in the levels of value (low, medium, high). Basic properties (bond strength and flexural strength to enamel and dentin, and color stability) of SI-R21302 were evaluated and compared to those of the following commercial resin cement systems: RelyX Veneer (RXV, 3M ESPE) and VarioLink Veneer (VLV, Ivoclar Vivadent), both of which have the same indication for use as SI-R21302. For shade evaluation models of LV restorations, abutment teeth (Ab) with shades equivalent to A1, A2 and A4, and LV (LV, shade: incisal, thickness:0.4mm) were used; RC paste was applied on LV, placed on Ab of each shade, light-cured under the conditions recommended by the manufacturer. The shades of the prepared evaluation models were then measured using a contactless colorimeter (DCCM-1, Shofu; 55mm light path length, halogen light source, 45- degree incident angle) to investigate differences in value of LV restorations caused by RC paste of the respective shades with the three levels of value against each background shade of Ab. The data obtained were statistically analyzed using Tukey’s test.

Results and Discussion: There were no statistically significant differences in the basic properties among SI-R21302, RXV and VLV (p<0.05). The value of the shade evaluation models of LV restorations increased as the value of RC paste increased, regardless of the shades of the abutment teeth; there was, however, a tendency that the degree of change in value of the shade models was smaller with the use of the A1-equivalent shade for the abutment tooth, whereas a larger degree of change in value was observed when the darker, A4-equivalent shade was used for the abutment tooth. The results of this study revealed that the novel resin cement system (SI-R21302) adequately controlled the value of LV restorations by selecting the most optimal value level (low, medium, or high) of the resin cement paste, while its thin cement layer was not influenced by the shades of the abutment teeth. This suggested that the novel resin cement system can facilitate easier control of the shades of LV restorations, and thus can more readily provide a favorable shade match to the adjacent teeth.

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Effect of the use of commercial cleaning liquid in thermoforming process of mouthguard materials Tanabe G1*, Churei H1, Wada T2, Uo M2, Takahashi H3, and Ueno T1 Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental Univ., Tokyo, Japan 1 Department of Sports Medicine/Dentistry, 2 Department of Advanced Biomaterials, 3Department of Oral Biomaterials Development Engineering * [email protected]

Introduction: Recently mouthguard (MG) is made by thermoplastic sheets with thermoforming machine. MG sheets can be used either single-layered or double-layered or sometimes even multilayered. When laminated MG is fabricated, it is important to adhere MG sheets to one another. The existence of any foreign particles such as dust, fingerprint and skin oil on and/or between surfaces of MG sheets makes the bonding strength (BS) weak and there may be risk of delamination of the sheets in a long time use. Recently a degreasing agent or cleaning liquid to laminate strongly MG sheets is available from manufactures, but their effectiveness was not tested objectively. The purpose of this study was to evaluate the effect of the commercial MG cleaning liquids by de-lamination test.

Materials and Methods: Ethylene vinyl acetate copolymer based MG sheets (3 mm) – MOUTHGUARD (Mo) (YAMAHACHI DENTAL MFG., CO., Japan) and Erkoflex® (Er) (ERKODENT, Germany) were used. MG cleaning liquids - Mouthguard Fitter (Mf) (YAMAHACHI DENTAL MFG., CO.,) and ENTFITTER (ENT), (ERKODENT) were used. Each MG sheet was intentionally imprinted with finger print oil and three conditions were tested: 1. no treatment (control) 2. treated with Mf and 3. treated with ENT. Subsequently, 6 types of sheets were made, namely 1. Mo-control, 2. Mo-Mf, 3. Mo-ENT, 4. Er-control, 5. Er-ENT and 6. Er-Mf and each sheet were observed under microscope (Keyence VHX-1000 JAPAN). Each MG test sheet was laminated with same manufacturing company’s MG sheet using vacuum forming machine (Keystone Industries, NJ, USA). After 24 hours, the laminated sheets were cut into a dumbbell shaped cutter according JIS K6251:2004 so that the adhesive area was at the center of the isthmus, and sectioned at the center of the isthmus as the final specimen for delamination having an adhesive area of 4x7.5 mm (n=6). Then, the delamination test was carried out using a universal test machine(Type 1123, Instron, USA) at a crosshead speed of 50 mm/min for determination the BS The statistical analysis was done with two-way ANOVA and Tukey’s HSD test (p<0.05).

Results and Discussion: BS values were Mo-control = 2.90 ±0.14 N/mm (mean±S.D); Mo-Mf = 6.80 ± 0.93 N/mm; Mo-ENT = 3.73 ± 0.27 N/mm; Er-control = 8.44 ± 0.30 N/mm; Er-ENT = 8.46 ± 0.15 N/mm and Er-Mf = 6.60 ± 0.68 N/mm. The significant difference in BS was Mo-control≒Mo-ENT ＜ Er-Mf ≒ Mo-Mf ＜ Er-control ≒ Er-ENT respectively. When Mf was used, BS of Mo increased, whereas BS of Er decreased in a significant manner (p<0.05). On the other hand, there was no significant difference when ENT was used. Accordingly, commercial MG cleaning liquids were not equally effective to MG sheets. The compatibility between cleaning liquid and sheet material should be carefully considered.

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Stress-strain behavior of bleached enamel stored in a high ionic strength solution Tanaka R*., Zhou J., Shibata Y. and Miyazaki T. 1Showa Univ., Tokyo Japan *[email protected] Introduction: Tooth bleaching is a popular esthetic treatment that enables to improve whiteness of discolored tooth via the least destructive procedure. Such bleached enamel surface attracts metal ions so that substitution of hydroxyl ion (X ion) within hydroxyapatite (HAP) lattice may occur. Assuming that the intrinsic mechanical property of tooth enamel is partially associated with HAP crystal structures, nanomechanical property evaluation on X ion substituted enamel must be considered. At nanoscale observation, enamel rods comprise HAP crystals tightly packed together with protein glues. A profound understanding of enamel local structural changes necessitates enamel mechanical characterization at smallest level.

Materials and Methods: Human third molar teeth were extracted for orthodontic indications, under a protocol approved by the ethics committee of the School of Dentistry, Showa University (Ref.: 2014- 031). The cuspal region of the tooth was cut and polished horizontally. As a consequence, keyhole like structures corresponding to microscale enamel rods exposed perpendicularly to the surface. 1 Kg of surface pre-reacted glass ionomer fillers were added into 1 L of pure- distilled water and mixed. A high ionic strength supernatant was obtained by a high-speed 3+ 3- centrifuge machine. The final products has an ion ingredient of Al 20.8 ppm, BO3 1683.8 ppm, Na+ 554.2ppm, Sr2+ 149.6ppm, and F- 137.0 ppm [1]. The polished surface of tooth specimen was exposed to 20 μL of a 35 % hydrogen peroxide (pH 3.4) bleaching agent (Hi- Lite, Shofu, Kyoto, Japan) and photo-irradiated [2]. Specimen was submerged into 100 mL of the above ionic solution and stored for 24 hours. Nanoindentation experiments were performed within the enamel rods using a diamond Berkovich tip and a spherical tip R = 0.5 μm using a quantitative nanomechanical test instrument. At least five nanoindentation regions were assessed and results were merged. The data were analyzed by ANOVA with Tukey post- hoc tests. P-values less than 0.05 were considered significant.

Results and Discussion: Limit of elastic range observed in control enamel was 5 GPa. It was higher than typically observed elastic limit in micron scale enamel characterization measuring multiple enamel “rods”. The limit of elastic range observed in bleaching enamel was observed to be lower than that of control. After storage in the solution, the elastic limit was superior to the point of control enamel. An enhanced tan δ values after storage implied higher mineral density of enamel after storage compared to the control sample. While sole function of bleaching diminishes enamel mechanical properties, subsequent exposure to a high ionic strength solution likely enhances enamel yield strength.

References: [1] Ito S, Iijima M, Hashimoto M, Tsukamoto N, Mizoguchi I, Saito T. Effects of surface pre- reacted glass-ionomer fillers on mineral induction by phosphoprotein. Journal of Dentistry. 2011;39:72-9. [2] Ogura K, Tanaka R, Shibata Y, Miyazaki T, Hisamitsu H. In vitro demineralization of tooth enamel subjected to two whitening regimens. Journal of the American Dental Association. 2013;144:799-807.

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Evaluation of the acid resistance of root dentin when applying fluoride containing materials incorporating Ca using in-air micro- PIXE/PIGE.

Yagi K1*, Yamamoto H1, Uemura R1, Okuyama K 2, Matsuda Y 3, Suzuki K4, and Hayashi M1 1Dept of Restorative Dentistry and Endodontology, Osaka Univ. 2 Dept of Dental Materials Sciences, Asahi Univ. 3 Div. of Clinical Cariology and Endodontology, Health Sciences Univ. of Hokkaido 4The Wakasa Wan Energy Research Center *[email protected]

Introduction: Recently, we established a measuring method of concentrations of calcium (Ca) and fluoride (F) before and after demineralization in dentin using in-air micro-PIXE/PIGE (proton induced X-ray/gamma-ray emission) measurement system. The aim of this study was to evaluate the acid resistance of human root dentin when applying fluoride containing materials (FCMs) incorporating Ca using in-air micro-PIXE/PIGE.

Materials and Methods: A total of 6 human third molars free from caries were used. Three root dentin blocks with a thickness of 200 µm were obtained from each tooth. Exposed dentin surfaces were covered with FCMs (Fuji VII (G-F7, GC) and F7Ca (G-F7Ca; Sr in Fuji VII was replaced with Ca)); while root dentin without FCMs was served as a control. The specimens were kept in saline at 37 °C for 1 month. After removing the FCMs, the specimens were kept in 10 ml of demineralizing solution (pH5.0, 37 °C) for 3 days. Before and after the demineralization, Ca and F concentrations were measured using the (1) PIGE/PIXE system at WERC (Wakasa, Japan) in accordance with the previous report . The results obtained from same specimens before and after the demineralization were compared on the basis of a reference point provided by a cupper foil (Kruskal-Wallis and Scheffe’s tests, α=0.05).

Results and Discussion: FCM groups showed significantly higher F uptake (median (interquartile range); G-F7=319(148-525) ppmF･µm, G-F7Ca=694 (360-906) ppmF･µm) than the control group (1.0 (1.0-1.0)ppmF･µm). G-F7Ca showed lower Ca loss (52 (26-129) wet%･µm) than the control (161 (119-340)wet% ･ µm). The F uptake before and after the demineralization showed significant negative correlations with the Ca loss (p<0.01). G-F7Ca showed significantly higher frequency of keeping outer surfaces compared with outer two groups. FCM incorporating Ca was found to be effective in improving the acid resistance of root dentin. This study was supported by Grants-in-Aid for Scientific Research (15K15700, 26462879) from the JSPS.

References: (1) Yasuda et al. NIM B 2011, 269(20), 2180-2183.

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Influences of various dentin desensitizers on the effect of tooth whitening Yoshikawa K1 , Yasuo K1, Furusawa K1, Iwasa K1 , Finger WJ2 and Yamamoto K1 1 Department of Operative Dentistry Osaka Dental University, Osaka, Japan *[email protected]

Introduction: Office bleaching is advantageous in obtaining a tooth-whitening effect within a short time compared to home bleaching and being applied under the control of a dentist, but agents used for office bleaching contain hydrogen peroxide at a high concentration, for which there are hypersensitivity frequently develops. In this study, we investigated the influence of the concomitant use of dentin desensitizers with tooth whitening on the tooth-whitening effect.

Materials and Methods: The test teeth were immersed in a tea-infused solution as a stainer for 7 days to prepare discolored samples. The color of the labial central region was measured (pretreatment).Tooth whitening was performed using TiON in Office, and the color was measured (first bleaching). In the control group, teeth were stored in saline thereafter and subjected to bleaching followed by color measurement every 7 days (2nd-4th whitening), 4 times in total. The tooth was then immersed in the stainer again, and the color was measured every 7 days (1st-4th relapse). As intra-treatment application groups, teeth were subjected to bleaching 4 times and MS-Coat F (MS), Nanoseal (NS), Teethmate Desensitizer (TD), and Teethmate AP Paste (AP) applied immediately after each bleaching. As post-treatment application groups, teeth were subjected to bleaching 4 times. After the 4th bleaching, dentin desensitizers were applied after the measurement after the 1st-3rd relapse every 7 days. The tooth color was measured before treatment, after the 1st-4th bleaching, and after the 1st-4th relapse. The color differences, ∆E, between the pretreatment value and value after the 4th bleaching and between the value after the 4th bleaching and 4th relapse, were determined. The measurement results were analyzed using one-way layout analysis of variance and Tukey’s test (n=5) (P<0.05).

Results: ∆E after the 4th bleaching compared to the pretreatment value was 32.9±4.5 in the control group and 44.9±6.2,7.7±10.9,40.9±9.6, and 34.6±2.5 in the intra-treatment application groups with MS, NS, TD and AP, respectively, showing no significant difference due to the concomitant use of the dentin desensitizers compared to the control. ∆E after the 4th relapse compared to the value after the 4th bleaching was 42.0±7.8 in the control group and 59.6±9.1, 34.4±2.4, 31.9±10.9, and 27.4±8.1 in the post-treatment application groups, respectively, showing a significant difference due to the concomitant use of MS applied after whitening treatment compared to the control.

Conclusion: The influences of various dentin desensitizers applied during and after whitening treatment on the tooth-whitening effect were investigated. 1. When the dentin desensitizers were applied during whitening treatment, none of them influenced the tooth-whitening effect. 2. When the dentin desensitizers were applied after whitening treatment, NS, TD and AP Paste did not influence the tooth-whitening effect. It was suggested that NS, TD and AP acting on hydroxyapatite and forming a thin capsule, can be concomitantly applied during and after whitening treatment.

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The Effects of Food and Drink on the Color Stability of Various Thermoplastic Resins Kuroda S1*, Yokoyama D1, Shinya A1,2, Gomi H1 1The Nippon Dental Univ., Tokyo, Japan 2Truk Univ., Truk, Finland *[email protected]

Introduction: Thermoplastic resins used as materials in non-metal clasp dentures are clinically adopted for their esthetic, biocompatibility and material biological property benefits. These resins are therefore regarded as materials in new prosthetic treatments. Clarifying how much of an esthetic impact food and drink have on prosthetic appliances fabricated with thermoplastic resins could enable esthetic predictions and the selection of materials tailored to the preferences of individual patients. In the present study, we therefore examined the color stability of thermoplastic resins. Materials and Methods: The thermoplastic resins used in the preparation of test specimens are presented in Table 1. Each thermoplastic resin underwent injection molding, recontouring and final polishing according to the manufacturer’s instructions to fabricate test specimens measuring 2.0 mm × 5.0 mm × 10 mm. The immersion fluids used in the immersion tests are presented in Table 2. Five specimens of each of the resins (total 240 specimens) were immersed in the six different immersion fluids. The immersion tests lasted from 1 to 5 weeks and the surface gloss and color difference were measured every week. Measurements were also taken following re- polishing of each specimen after 5 weeks of immersion. Results and Discussion: Surface gloss tended to decrease in value in all specimens from weeks 1 to 5. Color difference tended to increase slightly from weeks 1 to 5. Although re-polishing was promising in improving surface gloss, it only improved color difference slightly. Conclusions: Surface gloss was found to decrease and color difference was found to increase in thermoplastic resins as a result of exposure to various foods and drinks. These findings suggested the need for sufficient consideration to the properties and maintenance of thermoplastic resins when such resins are adopted for long-term use in non-metal clasp dentures. Table. 1 Thermoplastic Resin Materials Used in the Study Material Code Manufacturer Non-proprietary name EstheShot ESB Bright i-CAST, Kyoto, Japan Polyester EstheShot EST Reigning N REI Toushinyoukou, Niigata, Japan Polycarbonate UNIGUM UNM WELDENZ, Aichi, Japan Polypropylene Acron MC ACR GC, Tokyo, Japan Acrylic Lucitone FRS LCT DENTSPLY SANKIN, Tokyo, Japan Valplast VAL UNIVAL, Kanagawa, Japan Polyamide TCS TCS TCS, California, USA

Table. 2 Solutions Used for the Immersion Tests Immersion Fluid Proprietary name Manufacturer Tea Java Tea Otsuka Pharmaceutical Co., Ltd Coffee Nescafe Instant Coffee Nestle Japan Ltd Soda Coca-Cola Coca-Cola (Japan) Company Ltd. Lemon Pokka Lemon 100 Pokka Sapporo Food & Beverage Ltd. Mouth Wash Listerine Johnson & Johnson K.K. Curry Powder S&B Curry Powder S&B Foods Inc.

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Experimental immediate evaluation system for endodontic lesion

using light-induced red fluorescence

Takino H*, Isatsu K, and Hasegawa T Showa Univ., Tokyo, Japan *[email protected]

Introduction: In endodontic treatment for apical periodontitis, a bacterial infection significantly affects a prognosis and the healing process of apical periodontitis. As a key of success of endodontic treatment, any micro-flora, infected organization and inflammatory exudate should not be remained in the root canal before the root canal filling. For accurate diagnosis of optimum period of a root canal filling, the root canal sterility should be confirmed. Bacterial culture method has been widely used for the confirmation of root canal sterility. Since the bacterial culture method needs 24-48 hours as a culture period to detect bacteria flora, the rapid bacterial recognition system has been required clinically. Authors have already reported that a blue light of wavelength at 405nm cause a light-induced red fluorescence of a caries infected dentin, a plaque, a calculus and colonies of some periodontal pathogenic bacteria. In this study, the accuracy of experimental immediate evaluation system for endodontic lesion using light-induced red fluorescent was determined, and the spectral characteristics of the red fluorescent was examined by the high sensitivity multi-channel photo detector.

Materials and Methods: The experimental immediate evaluation system was assembled from a marketed digital camera, a laser diode light source of maximum output 5mW and an experimental fluorescence imaging box. All the subjects were the ones who visited the Comprehensive Dentistry of Showa University Dental Hospital (Tokyo Japan), required an endodontic treatment without regard of this study and accepted the volunteers of this study. All endodontic treatments were operated under rubber dam isolation. After removing a temporary sealing, drug which had been applied in root canal was removed ultrasonic scaler carefully, and rinsed with EDTA solution. 2 sterile paper points were inserted in the root canal for at least 30 seconds, to absorb periapical exudates and microorganisms from root canal as much as possible. One of paper points was subjected an anaerobic culture inspection, and another one was evaluated by the experimental system before the high sensitivity multi-channel photo detector.

Results and Discussion: 62 root canals in 188 root canals evaluated showed a positive contaminate reaction in anaerobic culture inspection, and paper points of these 62 root canals showed distinctive spectral characteristics which had twin peaks (620, 680 nm). 41 root canals in the 62 root canals were recognized red fluorescent by experimental system. In this study, a positive correlation between infection evaluation by the high sensitivity multi-channel photo detector and anaerobic culture inspection was recognized. In order to establish an accurate diagnostic tools, more effort should be required.

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Fabrication of a faceguard by incorporating a glass-fiber reinforced thermoplastic and shock escape space and this shock-absorption ability

Wada T*, Churei H, Fukasawa S, Shirako T, Hongo T, Ueno T, and Uo M Tokyo Medical and Dental Univ., Tokyo, Japan *e-mail : [email protected]

Introduction: Face guards (FG) are protection device that assists the safe and rapid return of athletes after facial traumatic injuries and orbital fracture. Current FGs have enough shock-absorption ability; however, heavy and thick FGs reduce the field of vision and comfortable wearing feeling. Then, performace of the athlete would be decreased by wearing such FGs. Therefore, thin and light-weight FGs are requested. In this study, thin and light-weight FG was fabricated with using the glass-fiber reinforced plastic (FRP) and incorporating the shock-escape space, and their shock-absorption abilities were estimated.

Materials and Methods: Figure 1 shows a cross sectional view of FG samples. Commercial thermoplastic resins for medical splint were used (Aquaplast, Sakai Medical Co., 3.2 mmt (AP32, Fig.1 (a)) or 1.6 mmt (AP16)). FRP was prepared from AP16 and the glass fiber cloth. Shock escape space (30 mmΦ) was incorporated in the AP16 (APS) and covered by AP16 (AP-APS) and FRP (FRP-APS, Fig.1 (b)). The specimens were covered with cushioning material (Neoprene, Sakai Medical Co.) on both sides. Shock absorption tests were carried out using an impact testing machine (modified IM-201, Tester Sangyo Co.). The impact load and pressure distribution were measured by the load cell sensors and the pressure measurement film. Fig. 1. Cross sectional view of FGs.

Results and Discussion: Figure 2 shows the impact load and maximum pressure. FRP-APS showed the lowest impact load, which was 75% that of the conventional FG (AP32). The pressure distribution of FRP-APS was too low to detect by the pressure measurement film (measurement range > 0.5 MPa). The weight except cushions was decreased to 60% compared to that of conventional FG. Therefore, the experimental FG with FRP and space could be decrease the total thickness and weight with keeping the shock-absorption ability to protect the injured athletes enough Fig. 2. The results of shock absorption safely. tests.

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THE INFLUENCE OF NAVAL NANOCALCIUM AND NANOCHITOSAN CONTENTS TOOTHPASTE ON ENAMEL SURFACE MICROHARDNESS Octarina*, Rahma Meilita**, Komariah*** * Departement of Dental Material, Faculty of Dentistry, Trisakti University, Jakarta 11440, Indonesia ** Undergraduated Program, Faculty of Dentistry Trisakti University, Jakarta 11440, Indonesia *** Departement of Oral Biology, Faculty of Dentistry, Trisakti University, Jakarta 11440, Indonesia Correspondence e-mail to : [email protected]

Introduction : Calcium and chitin derived from Scylla serrata and Xylotrupes gideon exoskeleton can be modified into nanocalcium and nanochitosan. These nanocalcium and nanochitosan has been known being able to remineralize the demineralized tooth. Aim : The study aims to evaluate the using of naval toothpaste containing nanocalcium and nanochitosan as basic ingridients on the enamel surface micohardness. Methode: A total of 10 labial incicivus enamel surface used as sampels in this study. Samples divided into 2 treatment groups (naval toothpaste group and commercial toothpaste group). Each group was brushed repeatedly for 42, 84 and 126 minutes. The enamel surface microhardness was tested before and after treatment by microhardness tester (Buchler). Result : The average of enamel surface microhardness before and after brushed with naval tooth paste for 42, 84 and 126 minutes were (397,80 ± 18,30) VHN, (388,80 ± 19,82)VHN, (401,40 ± 13,26) VHN, (405,80 ± 27,70), while those with comercial toothpaste were (372,40 ± 30,87) VHN, (391,20 ± 27,63) VHN, (416,60 ± 50,44) VHN, (430,20 ± 36,29) VHN respectively. Stastistical analysis with General Linear Model post hoc Bonferoni showed no significant differences (p=0,533 > 0,05). Conclusion : The using naval toothpaste contain nanocalcium and nanochitosan could increase the enamel surface microhardness. Key word : Naval toothpaste contain nanocalcium and nanochitosan, enamel surface microhardness

149

P-81

Bond Characterization of Various Framework Materials and Ceramic

Kawata K*, Teramae M, and Nakatsuka T Research and Development Dept., Shofu Inc., Kyoto, Japan *[email protected]

Introduction: There are various materials, such as metal, zirconia and lithium disilicate (LDS)-based glass ceramics, currently used for frameworks in porcelain restorations for crowns and bridges. Although ISO standards specify requirements for metal-ceramic bond strength in porcelain- fused-to-metal restorations, there are currently no defined standards for zirconia- or LDS- ceramic bond characterization. The purpose of this study was to evaluate bond strength between various framework materials and the ceramic in accordance with the same standard (ISO 9693-1:2012 Dentistry -- Compatibility testing -- Part 1: Metal-ceramic systems), and clarify differences in failure mode of the respective materials.

Materials and Methods: The framework materials used in this study were 3 types, i.e., metal [Me]: COBALTAN MB (Shofu, Co-Cr alloy); zirconia [Zr]: Shofu Disk ZR-SS (Shofu, TZP zirconia); and LDS: VINTAGE LD PRESS (Shofu). The dental porcelain materials used were: VINTAGE MP (Shofu) for Me; VINTAGE ZR (Shofu) for Zr; and VINTAGE LD (Shofu) for LDS. Sample pieces (25.0×3.0×0.5mm) were prepared from the respective framework materials, and their surfaces were alumina-blasted at 0.2MPa (Bl 0.2MPa) or 0.4MPa (Bl 0.4MPa), or mirror-polished (PO). Each corresponding porcelain material was built up on the center of the blasted/polished surface of each piece and fired (8.0×3.0×1.1mm) to obtain test specimens. The testing for debonding/crack-initiation strength of the specimens was carried out using a testing machine (AUTOGRAPH AG-5000B, Shimadzu) with a 20mm-distance between supports at a crosshead speed of 1.0mm/min. The test results were statistically analyzed by two-way ANOVA.

Results and Discussion: Table 1. Debonding/crack-initiation strength (MPa) Table 1 shows the Surface treatment PO Bl 0.2MPa Bl 0.4MPa debonding/crack-initiation strength Material used of the test specimens prepared with for framework the ceramic and the respective Metal (Me) 34.5(2.7) 32.4(3.2) 34.9(2.6) framework materials subjected to Zirconia (Zr) 28.2(1.6) 27.2(2.0) 28.4(0.4) the different surface treatment Lithium disilicate (LDS) 49.8(7.4) 47.2(4.6) 53.1(5.0) methods. * Standard deviation (SD) in brackets Differences in debonding/crack-initiation strength were observed among the framework materials used as in the following order: Zr0.01) although there were differences in surface roughness. The test results suggested that debonding/crack-initiation strength of the zirconia framework and the ceramic can be adequately evaluated by the method specified in the ISO standard for metal-ceramic systems. However, an alternative test method will need to be established for evaluating debonding/crack-initiation strength in LDS-ceramic systems since the LDS framework tended to fracture during the testing in this study.

150

P-82

Development of thin apatite coated zirconia implant

Hirota M1*, Ohkubo C1, Sato M2, Toyama T3, Tanaka Y4 and Hayakawa T1 1Tsurumi Univ., Kanagawa, Japan 2Kogakuin Univ., Tokyo, Japan 3Nihon Univ., Tokyo, Japan 4Kagawa Univ., Kagawa, Japan *e-mail [email protected]

Introduction: In recent years, yttria-stabilized tetragonal zirconia polycrystal (Y-TZP), has became an attractive new material for dental implants. Y-TZP provides high mechanical strength, fracture toughness, aesthetic performance and biocompatibility. In order to establish a metal- free implant system, the aim of the present study was to evaluate the bone responses of thin carbonate-containing hydroxyapatite (CA) film coated Y-TZP (CA-Y-TZP) using a molecular precursor method.

Materials and Methods: The CA film was deposited on the surface of Y-TZP (TZ-3YB-E, Tosoh, Japan) using a molecular precursor solution, which was a mixture of a calcium-EDTA complex and phosphate compounds1). CA thin film was deposited by heat treatment at 600°C for 2 h after molecular precursor solution application. The deposited CA film was observed and characterized by SEM, AFM, XRD, FT-IR and EDX. A scratch test was performed to measure the bond strength to Y-TZP. Biological evaluations of CA-Y-TZP were performed by immersion experiments in simulated body fluid (SBF) and animal implantation experiments in the bone defects of rabbits. The precipitation behavior on the surface of of Y-TZP and CA-Y-TZP after immersion in Hanks’ balanced salt solution was observed using SEM. As animal experiments, implantation into the tibia and femoral condyle of rabbits were performed (certificate No. 24A059). After 12 weeks of implantation, implants and surrounding bone were excised. Non- decalcified sections were prepared and the bone response towards the implants and the percentage of bone-to-implant contact (BIC) ratio and bone mass (BM) were evaluated.

Results and Discussion: The coated film was identified carbonate-containing hydroxyapatite by XRD, FT-IR and EDX. SEM observation with FIB technique revealed that the thickness of the uniform CA film was less than 1.0 μm. Scratch test and AFM analysis suggested that CA film on Y-TZP was strongly adhered. In the SBF immersion experiment, more apatite deposition progressed on CA-Y-TZP at the early stage of immersion than on Y-TZP. Animal experiments revealed that bone formation on CA-Y-TZP was similar with than on Y-TZP. Histomorphometrical evaluations showed a significantly higher BIC and BM on CA-Y-TZP into the femoral trabecular bone. Therefore, CA-Y-TZP appears to be applicable as a metal-free implant.

References: 1) Takahashi K et al. Molecular precursor method for thin calcium phosphate coating on titanium. Thin solid films 2005; 484: 1-9.

Acknowledgments: This work was supported by Grants-in-Aid for Scientific Research (B) (15K20492) from the Japan Society for the Promotion of Science.

151

P-83

Influence of bilayer ceria-stabilized zirconia/alumina nanocomposite-based crowns with supporting structures on fracture properties

Sawada T*, Spintzyk S, Schille C, Schweizer E, and Geis-Gerstorfer J Section Medical Materials Science & Technology, University Hospital Tübingen, Tübingen, Germany *E-mail; [email protected]

Introduction: In bilayer all-ceramic dental systems, chipping of the veneering porcelain is the most common technical complication in clinical practice. Zirconia framework modification is one approach to overcome this problem. The aim of this study was to evaluate the fracture load and failure mode of all-ceramic crowns with different ceria-stabilized tetragonal zirconia/alumina nanocomposite (Ce-TZP/A) framework designs.

Materials and Methods: Four different frameworks (1, anatomical shape (AS); 2, with an additional buccal (BS) or 3, lingual supporting structure (LS); and 4, with both buccal and lingual supporting structures (BLS]) were fabricated by using Ce-TZP/A blanks (C-Pro Nano-Zirconia, Panasonic Healthcare, Japan) with a CAD/CAM system. All frameworks were veneered with porcelain (VITA VM9, VITA Zahnfabrik, Germany) by using the conventional layering technique to fabricate all-ceramic crowns, followed by cementation to tooth analogs with self-adhesive resin cement (RelyX Unicem 2, 3M, Germany). The fracture load of each crown either without or with pre-loading (1.2 million cycles, 49 N) was measured by using a universal testing machine. Failure mode was classified into partial (cracking or chipping of porcelain veneer) or complete (fracture of Ce-TZP/A framework or tooth analog) fracture. Data were analyzed by using two-way analysis of variance followed by Tukey’s test and by Fisher’s exact test, respectively (α = 0.05).

Results and Discussion: Additional buccal and/or lingual supporting structures did not influence the fracture load and failure mode of Ce-TZP/A crowns without pre-loading. The most common failure modes were partial fractures. Pre-loading tended to increase mean fracture load in the LS, BS and BLS groups, while AS was decreased, but the differences were not statistically significant. On the other hand, pre- loading promoted fracture incidence and severity of failure mode from partial to complete fracture. In the AS group, prefailures were observed during pre-loading, and complete fractures were significantly increased after pre-loading. Only for the BLS group, the failure mode remained unchanged, showing only partial fracture even after pre-loading.

Conclusions: Bilayer Ce-TZP/A-based all-ceramic crowns manufactured with a framework consisting of anatomical shape with additional buccal and lingual supporting structures did not influence fracture load and failure mode, irrespective of pre-loading. This framework design has the potential to reduce chipping of the veneering porcelain.

Acknowledgments: The authors thank Dekema Dental-Keramiköfen and Panasonic Healthcare.

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P-84

The effect of surface modification on the bioactivity of zirconia materials

Hong G1*, Liu J1,2, Wu YH1, Endo K1, Han JM3, Wada T1, Kato H1, and Sasaki K1 1Tohoku Univ., Sendai, Japan 2Tianjin Medical Univ., Tianjin, China 3Peking Univ., Beijing, China *e-mail: [email protected]

Introduction: Presently, zirconia is one of the most promising implant materials due to its favorable physical, mechanical, biological and chemical properties.1 The aim of this study was to investigate and compare the initial bioactivity and surface characteristics of ceria-stabilized zirconia/alumina nanocomposite (NANOZR) with those of yttria-stabilized zirconia (3Y-TZP) and pure titanium (CpTi) following the use of three surface modification methods; mirror polishing, sandblasting/acid-etching (SB-E) and electrochemical deoxidation (ECD).

Materials and Methods: NANOZR (Panasonic Health Care Co, Japan), 3Y-TZP (Panasonic Health Care Co, Japan), and CpTi (Nippon Steel Co, Japan) discs (15 mm in diameter and 1.5 mm thick) were used in this study. The surfaces of all specimens were mirror polished using aluminum oxide waterproof abrasive paper (200#, 400#, 600#) prior to surface modification. Then, the specimens were treated for surface modification by ECD and SB-E. The mirror polished specimens prior to surface modifications were used as controls. Physical properties including surface morphology, chemical composition, X-ray diffraction, surface wettability, surface roughness, and hardness were measured. Osteoblast-like MC3T3- E1 cells were used to examine cell morphology and attachment to the surfaces of the materials.

Results and Discussion: ECD treated NANOZR showed a well-arranged, self-organized microporous surface structure with significantly low contact angles when compared with the other specimens (P < 0.05). NANOZR-E also demonstrated a slight decrease in monoclinic phase content (- 4.4 wt %). The morphology and attachment of MC3T3-E1 cells on NANOZR-E were similar to those on mirror polished and SBE-treated CpTi surfaces. Higher cell affinity was observed on NANOZR-E when compared with ECD treated 3Y-TZP.

Conclusions: Within the limitations of this study, our results suggest that the ECD technique is effective for the formation of a microporous surface on NANOZR, and may be useful in improving its biological activity during the initial stages.

Acknowledgements: This research was supported by Translational Research Network Program Seeds A (No. A- 72) for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan. We would like to thank the Panasonic Healthcare Company; and Dr. Yoshinaka SHIMIZU of Tohoku University for their support of this study.

References: 1. Chevalier J. What future for zirconia as a biomaterial? Biomaterials. 2006;27:535-543.

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P-85

Three-dimensional finite element analysis of the effect of load patterns on stress distribution in a zirconia all-ceramic crown Miura S1*, Kasahara S1, Yamauchi S1, Egusa H1 1Tohoku Univ., Miyagi, Japan *e-mail [email protected]

Introduction: The fracture of veneering ceramics is problematic in clinical zirconia all-ceramic restorations 1. One of the important clinical factors in the strength of a zirconia all-ceramic crown is the occlusal parameters. The purpose of this study was to evaluate the effect of different load patterns on the stress distribution in a zirconia all-ceramic crown using a three- dimensional finite element analysis.

Materials and Methods: A three-dimensional model of a zirconia all-ceramic premolar crown (yttria-stabilized zirconia framework with veneering ceramic) was used in the analysis. A cervical margin form of the crown abutment was prepared with a curvature radius R=1.0 mm (rounded shoulder margins). Dentin was used as a model of the abutment material. A force of 1 N was loaded in three different manners (Case 1: point load at the occlusal surface center; Case 2: surface load at the occlusal surface center; Case 3: surface load perpendicular to the angle of the abutment tooth). Stress analysis was performed using a linear static analysis (NX Nastran structural analysis software: MSC software, USA) to calculate the stress distribution that was generated in the crown and abutment.

Results and Discussion: In the Case 1 and 2, the maximum principal stress values of the crowns were equivalent (0.047 MPa). The maximum principal stress value in the Case 3 was higher (0.111 MPa) than that in the Case 1 and 2. In the Case 1 and 2, stress was concentrated on the load area of the crown. The stress distribution of the abutment tooth in the Case 2 decreased compared to the Case 1. In the Case 3, stress distribution was observed particularly in a crown margin area of the load side. In addition, a tendency of leaning of the whole abutment tooth was observed. These results suggest that a load pattern of the Case 2 would provide less burdening on a zirconia all-ceramic crown and the abutment tooth. Therefore, such a load may decrease the risk of fracture in zirconia all-ceramic crowns and protect the abutment tooth.

Acknowledgements: This study was supported by a Grant-in Aid for scientific Research (C: 26462908) from the Japan Society for the Promotion of Science and Tohoku Development Memorial Foundation.

Reference: 1. Takeichi T, Miura S, Kasahara S, Egusa H, Hara M, Sato T, Yoshinari M, Odatsu T, Watanabe I, Sawase T. Update zirconia restorations. J Prosthodont Res 2015; 59; 81-83.

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P-86

The effect of the cone angle and space between inner and outer crowns of a cone crown telescope with nano zirconia on retentive force and the fit of the copings

Nakagawa S*, Torii K, Yoshikawa Y, Okawa T, Ito Y, Fukumoto T, Tanaka J and Tanaka M Osaka Dental Univ., Osaka, Japan *e-mail ([email protected])

Introduction: Nano zirconia has recently been of interest not only for its biological affinity but also the expectedly stable technical accuracy by using CAD/CAM. In this study we conducted a basic research on the effect of the cone angle and space between inner and outer crowns on retentive force and the fit of the copings, with the aim of clinical application of cone crown telescopes with nano zirconia.

Materials and Methods: The shapes of the inner crown at tooth cervix included an ellipse with major radius 8 mm and minor radius 6 mm, a flat top shape with top radius 6.5 mm, and the cone angles 2, 4, and 6 °. One inner crown was made using a dental CAD/CAM system (D700-3SP，3Shape) for each condition, and we polished the products and used them as inner crown samples. We also scanned the inner crowns to make the outer crowns. The conditions for the outer crowns were such that the space between the inner and outer crowns was 0 or 10 μm. Five samples were made for each condition. For the retentive force measure, we first reposed the outer crown on the inner crown and applied a load using a tensile testing machine (EZTest, Shimazu Corp.). We then pulled the outer crown in the perpendicular direction with crosshead speed 40 mm/min and took the maximum value as the retentive force. To measure the fit of the copings, the outer crown first were placed on the inner crown and applied a load. We then used a universal tool maicroscope (TOPCON TUN-200, TOPCON) to measure the fit of the copings for the four borders and took the average value as the representative value. The load used was 50 and 100 N. As a statistical analysis, we conducted a two-way ANOVA analysis (α=0.01) with the cone angle and space as the factors. When the cone angle was significant, we used the Bonferroni method for multiple comparisons.

Results and Discussion: As a result of a two-way ANOVA analysis, for both loads, the cone angle was significant for retentive force, and the cone angle and space were significant for the fit of the copings (p＜ 0.01). For retentive force, the cone angle showed significant differences at all the levels, indicating that retentive force increases with decreasing cone angle. It is clinically established that 5 - 10 N is optimal, and the retentive forces with the cone angle 4° for loads of 50 N and 100 N were 7.9±0.28 N and 15.2±0.95 N, respectively. We did not show retentive force for the cone angle 6°. For the fit of the copings, the cone angle 2° was significantly different from 4° and 6°, and the fit of the copings tended to decrease with increasing cone angle and space. The fits of the copings, with cone angle 4° and space 10 μm, for the loads of 50 and 100 N were 174±51μm and 158±53μm, respectively. The above findings showed the possibility of adjusting the retentive force and the fit of the copings of a cone crown telescope by setting the cone angle and space.

155

P-87

Charpy impact test using single-edge notched zirconia specimens

Tsuruta S*, Mizuno M, Uematsu Y, Kawai T, Kurita S and Yamamoto I

Aichi Gakuin Univ., Nagoya, Japan *[email protected]

Introduction: Zirconia has recently been used for the dental restorations - usually crowns, fixed dentures and implants. A few reports have shown fractures of these zirconia restorations. These studies have demonstrated that the fractures were caused by either thin restorations, thick cements, or a discrepancies of the occlusal relation. It is well known that ceramics are prone to break in accidental impact, such as when they are dropped on the floor. Although zirconia shows high toughness in the static load test, the effect of instantaneous force is not clear. In this study, the CAD/CAM zirconia was evaluated with a Charpy impact test.

Materials and Methods: Two commercial zirconia disks, a CoCr disk (Ceramill Zi, Ceramill Zolid and Ceramill Sintron, Amann Girrbach AG, Austria) and a PMMA disk (Asahi PMMA Disk Temp, Asahi Roentgen Ind. Co., Ltd., Japan) were machined using a CAD/CAM system (Ceramill system, Amann Girrbach AG, Austria). The codes were ZI, ZO, SI and PM respectively. ZI, ZO and SI were sintered according to the manufacturer’s direction. After sintering, SI and PM were polished as usual. However, ZI and ZO were used as is (i.e. unpolished). The surface roughness (Ra) of ZI, ZO, SI and PM were lower than 3 µm, 3 µm, 0.2 µm and 1 µm respectively. Finally, five 3 mm x 4 mm x 40 mm beams were prepared. A straight notch, which was 0.8 - 1.2 mm depth in 4 mm side, was made in the center of the specimen using a low speed diamond saw with a 0.2 mm blade thickness. A Charpy impact testing machine with 1.96 J capacity was used. The specimen placed on the 30 mm-distance support was struck with a hammer behind the notch. The absorbance energy for fracture (E) was calculated from the following formula: E = M((cosb-cosa)-(cosa'-cosa)(a+b)/(a+a')), (M = moment of hammer, a = angle of fall, a' = angle of rise by blank run, b = angle of rise). The impact strength (c) was calculated from the following formula: c = E/2B(W-a), (B = width of specimen, W = thickness of specimen, a = depth of notch).

Results and Discussion: The results of the impact strength (kJ/m2) and mean (SD) are mentioned below. ZI = 1.52 (0.06), ZO = 1.34 (0.15), SI = 29.2 (1.5), PM = 0.504 (0.156) The specimen size and the formula were defined by the standard of the ceramic society of Japan (JCRS 202-1994). The relatively small dimension of the specimen was advantageous due to the size limitation of the CAD/CAM disks and moreover the deformation during the sintering became smaller because of the lower weights. The impact strength varied by size of specimen, shape of the notch, surface roughness and other variables. Therefore the strengths of this study cannot be compared to the results of the other studies indiscriminately. However, the strength of the zirconia specimen was measured at one-twentieth of that of the CoCr alloy. This weakness suggests that even sudden biting may induce fracture of oral zirconia restorations.

156

P-88

Development of tetragonal zirconia nanocrystals with high aspect ratio for composite resin fillers

Okada M1*, Taketa T1, and Matsumoto T1 1Okayama Univ., Okayama, Japan * [email protected]

Introduction: The mechanical properties of dental composite resins are significantly influenced by the nature of fillers (such as morphologies, chemical compositions and crystal phases) as well as their concentration and distribution in resin matrices. Tetragonal zirconia stabilized with dopants (such as Y3+) has exceptional mechanical properties due to crystal phase transformation (i.e., tetragonal to monoclinic transformation) upon loading. In order to incorporate the mechanical properties of tetragonal zirconia into the composite resins, the high-aspect-ratio morphology should be preferable because of its high capacity for bending stress and large interfacial area between a filler and resin matrix. Herein, we describe the development of tetragonal zirconia nanocrystals with high aspect ratio by a novel post-doping method.

Materials and Methods: First, monoclinic zirconia nanocrystals with high aspect ratio were prepared by a hydrothermal crystal growth method [1]. In order to obtain tetragonal zirconia, the monoclinic zirconia nanocrystals were calcined at 600°C for 1 hr after mixing with Y(NO3)3. The morphology was observed by scanning electron microscopy (SEM) using a JSM-6701F microscope (JEOL Ltd., Tokyo, Japan) operated at 5 kV. The number-averaged particle size (N = 50) was determined from SEM photographs with image analysis software (Image J; National Institutes of Health, Bethesda, MD, USA). The crystal phase identification was conducted by X-ray diffraction (XRD) measurements (RINT2500HF; Rigaku Corp., Tokyo, Japan) equipped with a Cu-Kα radiation source.

Results and Discussion: The monoclinic zirconia nanocrystals prepared by the hydrothermal crystal growth method showed high aspect ratio (227±162 nm in long axis; 16±6 nm in short axis; aspect ratio = 14). After calcination at 600°C without additives, the crystal phase of zirconia was not changed. On the other hands, tetragonal zirconia could be obtained after calcination at 600°C with 3+ Y(NO3)3, which indicates that Y ions were doped in zirconia after the calcination. The morphology of the tetragonal zirconia was not significantly changed (232±97 nm in long axis; 17±4 nm in short axis; aspect ratio = 14) as compared with that before calcination. Although the hydrothermal crystal method is one of the feasible methods to prepare nanocrystals with high aspect ratio, only monoclinic (non-doped) zirconia has been prepared because the presence of dopants (Y3+) leads to the formation of spherical nanocrystals. In this study, tetragonal zirconia with high aspect ratio could be successfully prepared by a novel post-doping method at 600°C after mixing with Y(NO3)3.

Acknowledgements This work was supported by JSPS KAKENHI Grant Number 15K13303.

References: 1) Murase Y, Kato E. J Am Ceram Soc 2001;84:2705–2706.

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P-89

Effects of zirconia surface synthesized polycrystalline particulates on the bonding strength of porcelain

Zhang ZT*, Tian YM, Zhuge RS, Zhang LL, Ding N

School of Stomatology, Capital Medical Univ., Beijing, China

* [email protected]

Introduction: Zirconia ceramics have been widely used in dental fixed restoration. This study aimed to improve the shear bond strength of porcelain through in-situ synthesized polycrystalline particulates on zirconia surface. Materials and Methods: Yttrium-stabilized tetragonal zirconia (Y-TZP) discs (16 mm in diameter, 1.5 mm in thickness) were cut from prefabricated zirconia blocks and polished with water-proof silicone- carbide paper until 1200#. Based on the immersion time of zirconia disc in HF solution, zirconia samples were divided into three groups: C, without immersion treatment; I1 and I2, immersed in the HF solution for 30s and 50s, respectively. Then, samples were put into CaCl2 solution (0.1 mol/L) for 90 s, dipped in NaOH solution from 20℃ to 80℃ in a water bath， and kept at 80 ℃ for 2 h. After final sintering(1450℃), surface appearance and chemical components were characterized with scanning electron microscopy (SEM) and energy dispersive spectrometry (EDS), respectively. Cylinder shaped porcelain veneer (zirconia porcelain) to the zirconia was build-up in the center of each specimen. Each specimen was embedded in an acrylic mold using methyl methacrylate resin and subjected to bonding tests. The shear bond strengths were performed using a universal testing machine at a crosshead speed of 1 mm/min. The results of bond strength were statistically analyzed with one-way ANOVA (α= 0.05). Results and Discussion: SEM images showed that regular polycrystalline particulates were formed on the zirconia matrix surface. Polycrystalline particulate distribution density became more connected closely with the increasing of immersion time in HF; EDS analysis showed the polycrystalline particulates consisted of Zr and O. Shear bond strength values of the three groups were

19.01+2.83 MPa, 22.01+2.28 MPa and 26.26+3.12 MPa, respectively, and the value of I2 and were statistically different from that of C and I1 (P<0.05). Due to HF could reacted with presintered zirconia ceramic, after a series of chemical reactions, ZrO2 polycrystalline particulates were synthesized on zirconium ceramic surface, and the polycrystalline particulates could effectively improve the bonding strength of porcelain. This study was supported by the Beijing Natural Science Foundation (Grant No. 7152065) and the National Natural Science Foundation of China (Grant No.81371175).

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P-90

Effects of zirconia particle addition on mechanical properties of calcium phosphate cement

Bae JY*, Ida Y, Sekine K, Kawano F, and Hamada K Tokushima University, Tokushima, Japan *[email protected]

Introduction: Calcium phosphate cements (CPCs) are important materials in several biomedical applications due to their excellent biocompatibility and osteoconductivity. However, their poor mechanical properties limit their applications. Zirconia is a promising additive to improve strength of CPCs without degrading biocompatibility. In this study, two types of ceramic composite of zirconia and CPC were fabricated with various powder-liquid mixing ratio, and their strength were evaluated.

Materials and Methods: β-tricalcium phosphate (control TCP; cβ-TCP) powder was mechano-chemically modified through ball-milling process. One additive was zirconia powder contaminated into β-TCP powder during ball-milling process. The zirconia powder was originated from milling-jar and milling-balls. The mixed powder of β-TCP and zirconia was rinsed in hydrochloric acid and only zirconia powder was filtrated. The extracted powder was then washed in distilled water and dried (evapo-ZR). The other additive was zirconia powder extracted from commercial water-base nano-zirconia dispersion with particle size of approximately 100 nm and mass- fraction of 10%. The dispersion was centrifuged to concentrate zirconia powder, and the dense dispersion was sucked up and dried to remove water (nano-ZR). Zirconia powder was mixed into cβ-TCP powder with mixing ratio from 1% to 6% in mass. The mixed powder was ball-milled for 24 hours. The mixed powder after ball-milling (β- TCP-evapo-ZR and β-TCP-nano-ZR) was mixed with 2.5 mass% CaCl2 solutions, and then 2.5 mass% NaH2PO4 solutions at a powder/liquid ratio of 4:1:1. The mixed paste was stored for setting in an incubator for 1 week, and compressive strength (CS) and diametral tensile strength (DTS) of set specimen was evaluated. Fracture surface of specimen after DTS test were observed by scanning electron microscopy (SEM).

Results and Discussion: The highest CS of β-TCP-evapo-ZR and β-TCP-nano-ZR was 34.67 MPa at a mixing ratio of 2% and 38.21 MPa at a mixing ratio of 1%, respectively. The highest DTS of β-TCP- evapo-ZR and β-TCP-nano-ZR was 7.84 MPa at a mixing ratio of 4% and 7.54 MPa at a mixing ratio of 3%. Both CS and DTS of both composite increased with increasing zirconia content from 1%. However they indicated peaks and changed to decrease with increasing content. These CS and DTS values were significantly higher than those of c-β-TCP.

Conclusions: Zirconia particle addition successfully improved strength of c-β-TCP. The type of zirconia did not affect CS and DTS of CPC significantly in this study.

Acknowledgements: This work was partially supported by the JSPS KAKENHI grant numbers 16K20504 and 16K11626.

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P-91

Change in the retentive force of Akers clasp for zirconia crown

Tanaka A1*, Miyake N1, Hotta H1, Takemoto S1, Yoshinari M1, Yamashita S1

1Tokyo Dental Collage, Tokyo, Japan

*e-mail: [email protected]

Introduction: Recently, high strength ceramic materials, especially, zirconia crown have been used for various prosthetic treatment and a clinical situations are supposed using zirconia crown as an abutment tooth for removable partial denture. However, little study has been reported about the suitability of clasp for zirconia crown. The purpose of this study was to clarify the availability of zirconia crown for the abutment tooth of removable partial denture, by investigating the change in retentive force of clasp fitted on zirconia crown.

Materials and Methods: Porcelain glazed zirconia crowns were designed for mandibular second premolar using zirconia blank and fabricated by a CAD/CAM system (S-WAVE, Shofu, Co., Ltd., Kyoto, Japan), and Akers clasps adapted these crowns were fabricated using 12% gold-silver- palladium alloy. Retentive force of Akers clasps were measured using a universal testing machine (EZ-S, Shimadzu, Tokyo, Japan). Using an insertion/removal testing apparatus(JM100-T, Japan Mecc, Co., Ltd., Tokyo, Japan) designed at the Tsurumi University School of Dental Medicine, the change in retentive force was measured after repeating the tensile and compressive motions (crosshead speed: 950 mm/min) up to 10,000 cycles at 37 oC in distilled water.

Results and Discussion: The initial retentive force was more than 9N. This force didn't change until 3,000 cycles of repeating motions, but the retentive force tended to decrease gradually over 5,000 cycles. Within the limitations of this study, these results suggest the availability of zirconia crown for the abutment tooth of removable partial denture.

Acknowledgements: We greatly appreciate the members of Tsurumi University School of Dental Medicine Department of Removable prosthodontics for assistance and cooperation throughout the study. We also extend our sincere gratitude to Shofu Co., Ltd., for using special facilities.

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Effect of surface treatment on the bond strength between zirconia ceramic and core resin

Uno M1*, Kawaki H2, Nonogaki R1, Sawada T1, Nishikawa M3 , Kurachi M4, Ishigami H1, and Doi Y5 1Department of Prosthodontics, Division of Oral Functional Science and Rehabilitation, Asahi University School of Dentistry, Gifu, Japan 2Department of Oral Biochemistry, Division of Oral Structure, Function and Development, Asahi University School of Dentistry, Gifu, Japan 3Department of Dental Materials, Division of Oral Functional Science and Rehabilitation, Asahi University School of Dentistry, Gifu, Japan 4Dental Education Promotion Center, Asahi University School of Dentistry, Gifu, Japan 5Asahi University School of Dentistry, Gifu, Japan *e-mail [email protected]

Introduction: Zirconia has been used in various dental applications due to its esthetic and biocompatible properties, especially, framework and core restorations. Although zirconia frameworks have proper mechanical property and unique chemical stability, the successful zirconia frameworks depend on the strong bond between the zirconia ceramics and resin substructures. In this study, we evaluated the effects of three different primer systems on the shear bond strength between zirconia-based ceramics and core resin.

Material and Methods: Cylinder shaped zirconia blocks (diameter: 10 mm, length: 30mm) were prepared. (Toso, Japan)

These zirconia specimens were airborne-particle-abraded with 70 μm Al203 particles under 0.3 MPa bar pressure in circular movements at a distance of 10 mm perpendicular to the surface for 20 s. All of zirconia specimens were then treated with AZ primer and divided into 4 groups. Cylinder shaped composite resin specimens were also prepared (3 mm diameter and 3 mm length). We tested 4 groups with following conditions, no primer treatment group as Group 1, AZ primer to zirconia and silane coupling porcelain primer to resin, as Group 2, AZ primer to zirconia and 2-HEMA and 4-AET based AB Primer to resin, as Group 3, and AZ primer to zirconia and both of silane coupling porcelain primer and AB Primer to resin, as Group 4. The resin specimens were bonded to the zirconia specimens with resin cement, fixed to universal testing machine, and bond strength was evaluated. Multiple comparisons were performed using a one-way analysis of variance and the Bonferroni method.

Results and Discussion: The highest shear bond strength, approximately 34 MPa was obtained in Group 3 and 4. Our study showed that the 2-HEMA and 4-AET based AB primer system was promising to effectively bond zirconia-based ceramics and core resin.

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Zirconia surface coating with silk fibroin electrogels Qu YY1, 2*, Hong G1, Liu L2, and Sasaki K1 1Tohoku Univ., Sendai, Japan 2Dalian Stomatological Hosp., Dalian, China *e-mail ([email protected]) Introduction: Excellent biocompatibility, ivory color, and favorable mechanical properties make zirconia a candidate for dental implant material. To achieve better initial stability after implantation, functionalization by the surface modification is one of the key solutions. Silk fibroin, which good performance in controlled drug delivery has been identified by lots of in vivo experiments, is a promising biomaterial for tissue engineering. This research aimed to explore the possibility of functionalization of zirconia surface with silk fibroin by its electrogel coating.

Materials and Methods: Silk fibroin solution was produced from silkworm cocoons. Disks 15mm in diameter and 2mm in thickness of 3 mol% yttria-stabilized zirconia (3Y-TZP) and ceria-stabilized zirconia/alumina nanocomposite (NANOZR) were used as substrates. The zirconia disks were polished, then the surface was sandblasted and etched using 47% HF. For electrogelation, electrodes were immersed in silk fibroin solution at the concentration of 30mg/ml and direct current (20V) was applied to the anode (platinum) and cathode (titanium). Two hours later, electrogels formed at the bottom of the electrophoresis chamber were taken and put into the plate with the zirconia disks. After thoroughly dried, the samples were coated with silk fibroin. The surface wettability of the coated specimens was evaluated by contact angle meter (PCA-1). And the bond strength between silk fibroin coating and zirconia surface was measured by a testing machine (Instron 5565). In addition, the surface morphology was evaluated using SEM (JSM-6390LA). All data were analyzed independently using one-way analysis of variance (ANOVA).

Results and Discussion: Silk fibroin electrogels formed stable coating on the surface of zirconia disks. The density of silk fibroin electrogels increased with the time of electrophoretic deposition, however, after 2 hours of deposition, electrogels were easily changed into clumps and therefore incapable of forming uniform coating on zirconia surface. This could be attributed to the irreversible change into β-sheet structure by the shear raised up while sucking and ejecting. The contact angle of surface coated by silk fibroin was significantly lower than un-coated surface. The tensile bond strength between silk fibroin and NANOZR samples was higher than 3Y-TZP samples. This could be attributed to different surface characters of the two kinds of materials and suggested NANOZR might be more suitable for silk fibroin electrogel coating.

Conclusions: Silk fibroin electrogels were able to form stable coating on the surface of zirconia materials, which may provide possibilities of controlled delivery of bioactive molecules for zirconia materials.

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P-94

XRD and FT-IR analysis on fluoride-treated hydroxyapatite

Okuyama K1*, Komada Y1, Abe M1, Matsuda Y2, Yamamoto H3 and Tamaki Y1 1Asahi Univ., Gifu, Japan 2Health Science Univ. of Hokkaido, Hokkaido, Japan 3Osaka Univ., Osaka, Japan *e-mail: [email protected]

Introduction: It is well-known that fluoride is demonstrated as a significant benefit for preventing caries. Incorporating fluoride in tooth mineral is enhanced the dissoluble acid resistance of tooth structures. This acid resistance ability was performed when hydroxyapatite (HAP) was converted to fluorapatite (FAP) by fluoride application. However, the differences of structures between fluoride-treated HAP (f-HAP) and HAP or FAP were not clear. The purpose of this study was to evaluate HAP, which was treated with various concentrations of fluoride used by X-ray diffraction (XRD) and Fourier Transform Infrared Spectroscopy (FT-IR) analysis.

Materials and Methods: 400 mg HAP powder (Wako chemical, Japan) was soaked into 15 ml remineralizing solution (pH:7.0, including CaCl2, KH2PO4, KCl) adjusted various concentrations of fluoride (10, 100, 1000 ppmF; NaF) for 24 hours. After 24 hours, samples were washed with water and filtered, and then dried at 100ºC for 2 hours. Dried samples were subjected to XRD (UltimaIV, Rigaku, Japan) or FT-IR (FTIR-8400S, Shimadzu, Japan) analysis. FAP (BIO-RAD, USA) or HAP was analyzed as a positive or negative control, respectively.

Results and Discussion: For XRD analysis, both 211(around 31.8º) and 300 (around 32.8º) peaks of FAP shifted to higher angle compared with HAP. These peaks of f-HAP shifted to higher angle slightly. The peak position of 10 ppmF treatment samples was almost same as HAP. For FT-IR analysis, the OH absorption peak (at 3568 cm-1) of FAP was weaker than that of HAP. On f-HAP with 100 or 1000 ppmF treatment, the peak of the same position was weaker than that of HAP, however, larger than that of FAP. These results demonstrate that f-HAP and FAP has less OH group than HAP, because OH- ion was replaced to F- ion on these samples. f-HAP shows smaller shift to high angle on XRD and smaller absorption on FT-IR than FAP. On f-HAP samples, the small amounts of fluoride might be contributed to convert to FAP, therefore, parts of HAP became FAP.

This research was supported by JSPS KAKENHI Grant Number 15K11101

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P-95

Hydroxyapatite nanoparticle-assembled powder as a pulp-capping agent in rats

Imura K1, Hashimoto Y1*, Okada M2, Yoshikawa K1, Imai K1, Matsumoto T2 and Yamamoto K1 1Osaka Dental Univ., Osaka, Japan 2Okayama Univ., Okayama, Japan *E-mail ([email protected])

Introduction: Direct pulp capping is frequently used to preserve the vitality when dental pulp is accidentally exposed during cavity preparation. Recently, nanosized hydroxyapatite (HAp) structures have attracted attention because of their improved bioactivity compared to conventional HAp. In this study, HAp nanoparticle-assembled powder was applied to exposed dental pulp of rat molars, and the tissue reaction was then examined histologically to assess its biocompatibility and ability to promote hard tissue formation.

Materials and Methods: HAp nanoparticles were prepared using a wet precipitation method. Briefly, an aqueous solution of (NH4)2HPO4 was added to an aqueous solution of Ca(NO3)2·H2O, whose initial pH was 10. HAp nanoparticle-assembled blocks were then prepared by drying the aqueous dispersion of HAp nanoparticles (ca. 3 wt%) on a polytetrafluoroethylene (PTFE) dish at 60°C for 18 h. Approximately 50-μm-sized HAp nanoparticle-assembled powders were obtained by grinding HAp nanoparticle-assembled blocks. The surfaces of the sample were observed using scanning electron microscopy (SEM; S- 4800; Hitachi High Technologies Corp., Tokyo, Japan) at 5 kV. Fourier-transform infrared (FT-IR) spectra were obtained using a Spectrum One (Perkin-Elmer Inc., Waltham, MA) device equipped with a diffuse reflectance unit at a resolution of 4 cm–1 with 16 scans. The crystal phase was identified by X-ray diffraction (XRD) measurements (XRD-6100; Shimadzu Corp., Kyoto, Japan). X-ray photoelectron spectroscopy (XPS) measurements were performed to determine the surface Ca/P atomic ratios with a PHI X-tool (Ulvac-Phi, Inc., Kanagawa, Japan) equipped with an Al-K radiation source (15 kV; 25 W). The pulps of maxillary first molars of male Wistar rats were exposed and then capped directly by using either HAp nanoparticle-assembled powders or calcium hydroxide slurries (control). Histologic examinations were performed at 1 and 2 weeks after the surgical procedure.

Results and Discussion: FT-IR and XRD measurements showed that the nanoparticle-assembled powders prepared by the above-described procedure consisted of low-crystallized HAp, and no other calcium phosphate phase was observed. The Ca/P atomic ratio of the powder measured by XPS was 1.45, thereby suggesting that the surface of the powder was calcium-deficient. SEM revealed that the HAp nanoparticles had a spherical morphology with a diameter of approximately 30 nm, and nanosized pores (ca. 10 nm) were also observed between the nanoparticles. Histologic examinations showed that there were few inflammatory cells in the pulp near the exposed sites in both groups at 1 week. At 2 weeks, reparative dentin covered with a layer of odontoblast-like cells was observed in the HAp nanoparticle-assembled powder group. HAp nanoparticle-assembled powder promoted healing of the dental pulp.

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P-96

Bone response to apatite paste derived from Ca-amino acid complex

Waki T1*, Mochizuki C2, Sato M2, Hayakawa T1, and Ohkubo C1 1Tsurumi Univ., Kanagawa, Japan 2 Kogakuin Univ., Tokyo, Japan *[email protected]

Introduction: Hydroxyapatite is well-known bone substitute material. Various methods for preparing apatite have been reported. Previously, we prepared apatite from the calcium (Ca) complexes of ethylenediamine tetraacetic acid (EDTA) and evaluated their bone response1). Further more, the preparation of apatite from Ca-Aspartic acid (Asp) which constituted non-collagen proteins was reported2,3). It was found that apatite from Ca-Asp complex could be formed in a paste after mixing with water. In the present study, we aimed to investigate the influence of the difference of amino acid ligand for calcium on bone response. Apatite from Ca-Asp and Ca-Glutamic acid (Glu) was prepared and evaluated the bone response towards each apatite paste after implantation into a subperiosteal pocket of rat calvaria. As a control, apatite form Ca-EDTA was evaluated.

Materials and Methods: First aqueous mixture of Asp or Glu and Ca(OH)2 was heated and phosphoric acid was added after cooling. Then, L-Alginine was added to reaction mixture. Precipitated apatite powder was obtained by filtration and characterized by x-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR) measurement. Nine-week-old male Sprague-Dawley rats were used (approval number: A27A051). Apatite powder from Ca-Asp, Ca-Glu and Ca-EDAT was mixed with double distilled water to form in a paste. Apatite paste was placed into a polytetrafluoroethylene (PTFE) tube, which was implanted directly into a subperiosteal pocket of rat calvaria. At 4 or 8 weeks after implantation, each specimen was excised, and new bone formation was observed by micro- Computed Tomography (CT, inspeXio SMX-225 Shimadzu).

Results and Discussion: Each apatite powder was characterized by XRD and FT-IR measurements. Both measurements confirmed apatite structure. In micro-CT observation, newly formed bone-like structure was identified. At 4 weeks after the implantation, new bone-like structure was slightly observed and no distinct difference was recognized among three kinds of apatite paste. At 8 weeks, more progress of new bone-like structure was observed. New bone-like structure formed from the calvaria surface, not from the periosteum for all cases. More amount of new bone-like structure was identified for apatite from Ca-Glu. Thus, it could be revealed that the difference of amino acid as a ligand of Ca influence the bone response. Apatite paste has an advantage for the application of any shaped of defects.

References 1) Hayakawa et al., J Maer Sci Mater Med. 21; 251-258: 2010. 2) Mochizuki et al., Special Issue for 21st International SPAC symposium in Tokyo 2014. 3) Hayakawa et al., J Oral Tissue Engin 12; 115-120: 2015.

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P-97

Bone regeneration with a collagen model polypeptide/porous alpha- tricalcium phosphate sponge in a canine mandibular bone defect model

Matsuse K, Hashimoto Y*, Imai K, Baba S and Morita S Osaka Dent Univ, Osaka, Japan * yoshiya＠cc.osaka-dent.ac.jp

Introduction: Mandibular defects originate from various pathologic processes such as congenital malformations, trauma, oral cancer, and infections. Canines have proven to be suitable to study bone regeneration of mandibular defects because they enable the study of large deformations/defects In this study, we used a dehydrothermal cross-linking method to construct collagen model polypeptides comprising a poly(PHG) (a proline-hydroxyproline- glycine sequence) and a porous alpha-tricalcium phosphate (-TCP) sponge as a scaffold for bone regeneration. The effects of the poly(PHG)/-TCP scaffold on bone regeneration were evaluated on surgically formed canine mandibular bone defects.

Materials and Methods: The porous particles were prepared by pulverizing an -TCP block with 80% of its pores having continuous pore structures. These porous -TCP particles were mixed with a poly(PHG) solution; the resultant sponge was then cross-linked and characterized by X-ray diffraction (XRD), Fourier-transform infrared spectroscopy(FT-IR), and scanning electron microscopy (SEM). A twist drill was used to generate identical bone defects (4 mm in diameter and 6 mm in depth) in canine mandibles that were then divided into three groups. Canine mandibles from the poly(PHG)/-TCP group, the poly(PHG) group, and the no implant group were implanted with the poly(PHG)/-TCP sponge, the poly(PHG) sponge, and nothing, respectively. Mandibular bone defects were analyzed using micro-computed tomography and histological evaluation at 2, 4, and 8 weeks after implantation.

Results and Discussion: XRD, FTIR, and SEM revealed that the poly(PHG)/-TCP sponge composites comprised crystallized -TCP particles and a 3D porous structure. The bone volumetric densities (VDs) were determined for each group at three time points. At 2 and 4 weeks post- implantation, the VDs for the poly(PHG)/-TCP group were higher than those for the poly(PHG) and no implant groups (p < 0.05). However, no significant difference in VD was observed among the three groups at 8 weeks post-implantation (p > 0.05). Histological evaluation at 4 weeks after implantation revealed that the poly(PHG) had degraded and a newly formed bone was present on the surface of the -TCP particles. At 8 weeks post- implantation, continuous cortical bone formation with a Haversian structure was observed covering the top of the bone defects in both groups with implants. This study demonstrated that the composite created using porous -TCP particles and poly(PHG) is sufficiently adaptable to treat bone defects.

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P-98

Effect of water structure on bone mineral formation I: Kosmotropes

Kobayashi D1*, Kai S1,Okada M1, and Matsumoto T1 1Okayama Univ., Okayama, Japan *e-mail: [email protected]

Introduction: Bone mineralization involves complex and spatiotemporal dynamic physico-chemical reactions. Despite large number of reports for bone mineralization, there still remain some unclear aspects. For instance, although the bone mineral (apatite) precipitates in the presence of water molecules, there is a lack in the state of water molecules surrounding bone minerals. A water molecule has a strong electrical polarity; i.e., a water molecule has a partial negative charge (–) near the oxygen atom due the unshared pairs of electrons, and partial positive charges (+) near the hydrogen atoms. This electrical polarity causes the water molecules to interact with each other and with ionic solutes (electrolytes) in water media through hydrogen bonding. Ionic solutes in water media are classified into kosmotropes (order-making agents) and chaotropes (disorder-making agents) [1]. Kosmotropes (such as Na+, Mg2+, Sr2+ and Zn2+) contribute to stabilize the hydrogen bonding network of water molecules. In this study, we evaluated the apatite formation in vitro in the presence of kosmotropes, in order to investigate the effect of the water structure (i.e., hydrogen bonding network) surrounding bone minerals.

Materials and Methods: 2+ 3– Aqueous solutions of Ca and PO4 were, respectively, prepared and mixed in an aqueous solution in the presence of kosmotropes (Na+, Mg2+, Sr2+ or Zn2+). The resulting precipitation was then centrifugally washed with pure water, and then redispersed in water. The morphology of the precipitation was observed by scanning electron microscopy (SEM) using a JSM-6701F microscope (JEOL Ltd., Tokyo, Japan) operated at 5 kV. The number-averaged particle size was determined from SEM photographs with image analysis software (Image J; National Institutes of Health, Bethesda, MD, USA). The crystal phase identification was conducted by X-ray diffraction (XRD) measurements (RINT2500HF; Rigaku Corp., Tokyo, Japan) equipped with a Cu-Kα radiation source.

Results and Discussion: In the case of the presence of a small amount of kosmotropes, the particle size and the crystallinity of the precipitation tended to decrease as compared with those of the precipitation obtained without kosmotropes. Kosmotropes exhibit stronger interactions with water molecules than water with itself. Therefore, it seems that the kosmotropes prevent the initial formation of particle nuclei and the subsequent particle growth due to smaller rate of dehydration on the calcium phosphate surfaces. The detailed data and discussion on the basis of the structure of water molecules in the presence of different kinds and concentrations of kosmotropes will be shown in this presentation.

References: [1] Collins KD. Biophys J 1997;72:65–76.

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P-99

Effect of water structure on bone mineral formation II: Chaotropes

Kai S1*, Kobayashi D1, Okada M1, and Matsumoto T1 1Okayama Univ., Okayama, Japan *e-mail: [email protected]

Introduction: Ions in aqueous media are surrounded by water molecules, and the interaction between ions and water molecules depends on the size and charge of the ion. The findings in thermodynamic, spectroscopic and computer-simulation studies indicate that ions can be classified into kosmotropes (structure makers) and chaotropes (structure breakers). Chaotropes (such as K+, Cl– and urea) disrupt the hydrogen bonding network of water molecules [1]. Although it is well known that chaotropes also disrupt the structure of macromolecules such as proteins and nucleic acids by weakening the hydrophobic interaction, there is less knowledge about the effects of chaotropes on mineral (apatite) precipitation and growth in bone tissues. In this study, we evaluated the apatite formation in vitro in the presence of chaotropes, in order to investigate the effect of the water structure (i.e., hydrogen bonding network) surrounding bone minerals.

Materials and Methods: 2+ 3– Aqueous solutions of Ca and PO4 were, respectively, prepared and mixed in an aqueous solution of chaotropes (K+, Cl– or urea) in the presence or absence of water-soluble macromolecules. The resulting product was then centrifugally washed with pure water until the pH of the solution became neutral, and then redispersed in water. The morphology of the product was observed by scanning electron microscopy (SEM) using a JSM-6701F microscope (JEOL Ltd., Tokyo, Japan) operated at 5 kV. The number-averaged particle size was determined from SEM photographs with image analysis software (Image J; National Institutes of Health, Bethesda, MD, USA). The crystal phase identification was conducted by X-ray diffraction (XRD) measurements (RINT2500HF; Rigaku Corp., Tokyo, Japan) equipped with a Cu-Kα radiation source.

Results and Discussion: In the case of the presence of a small amount of chaotropes, hydroxyapatite crystals were precipitated and the particle size of the crystals tended to increase as compared with that obtained in the absence of chaotropes. Chaotropes disrupt the hydrogen-bonding network due to their weak interaction with water. Therefore, it seems that the chaotropes promote the initial formation of crystal nuclei and the subsequent crystal growth due to the disrupted dehydration layers on the crystal surfaces. The detailed data and discussion on the basis of the structure of water molecules in the presence of different kinds and concentrations of chaotropes will be shown in this presentation.

References: [1] Collins KD. Biophys J 1997;72:65–76.

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P-100

Fabrication of biphasic bone substitute consisting of calcite and carbonate apatite

Tsuru K1*, Munar M1, and Ishikawa K1 1Kyushu Univ., Fukuoka, Japan, *e-mail: ([email protected])

Introduction: Carbonate apatite (CO3Ap) granules prepared based on dissolution-precipitation reaction using calcite as precursors were found to be replaced to bone1. However, the replacement of to bone needs longer time. One of methods to fasten the bone replacement may be to separate the role of surface and core materials of the bone substitute. In other words, surface material is expected to show good osteoconductivity and should dissolve at weak acidic condition formed by the osteoblasts but should not dissolve at physiological condition. In contrast, core material should show higher dissolution than surface material. In the present study, CO3Ap coated calcite was fabricated and its behavior at the bone defect was evaluated using rabbits.

Materials and Methods: Calcite block was prepared by exposing Ca(OH)2 compacted uniaxially at 5MPa to CO2 at room temperature for 7days. Calcite granules (600-1000m) were immersed in 1 mol/L Na2HPO4 solution for prescribed period. Composition was evaluated by means of X-ray diffraction. SEM and EDX were used to perform morphological and elemental analyses. Cylindrical bone defects (φ5.0 mm×8.0 mm) formed at rabbit femur was reconstructed with 1) CaCO3, 2) 10wt% CO3Ap coated CaCO3, 3) 30wt% CO3Ap coated CaCO3, 4) CO3Ap. Also sham operation was made for comparison. Rabbits were sacrificed 4 weeks after surgery and analyzed by means of micro computed tomography (microCT; Latheta LCT-200, ALOKA). Histological observation (Villanueva-Goldner staining, Toluidine blue staining) was used to quantify new bone volume at the defect site.

Results and Discussion: Calcite transforms to CO3Ap and the rate of transformation was faster at higher o temperature. 4 and 8 hours at 40 C were chosen to fabricate 10 and 30wt% CO3Ap coated calcite. EDX analysis confirmed that surface and core were CO3Ap and calcite, respectively. New bone formation was observed on the surface of all granular including calcite. However, the amount of newly formed bone was limited in the case of calcite. Interestingly, approximately twice amount of new bone were observed in the case of 10wt% CO3Ap coated calcite and 30wt% CO3Ap coated calcite when compared to CO3Ap. Although the mechanism of the larger amount of bone observed for the CO3Ap coated calcite has not been clarified in the present study, Ca2+ supplied by the dissolution of calcite may elicit the activation of osteoblastic cells. CO3Ap coated calcite seems to have good potential to be ideal bone substitutes since it has higher osteoconductivity and shows quicker replacement to bone when compared to CO3Ap.

Acknowledgements: This study was supported in part by Grant for Strategic Promotion of Innovative Research and Development, Japan Science and Technology Agency, Japan and Grant-in-aid for Scientific Research from the Ministry of Education, Science, Sports and Culture, Japan.

Reference: 1. Ishikawa K. et al., Materials 3:76-83, 2009

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P-101

Clinical application of octacalcium phosphate collagen composite in sinus floor elevation

Tadashi Kawai1*, Keiko Matsui1, Fumihiko Kajii1,2, Yoshinaka Shimizu1, Hiroyuki Kumamoto1, Osamu Suzuki1, Shinji Kamakura1, Tetsu Takahashi1

1Tohoku Univ., Sendai, Japan 2TOYOBO CO., LTD, Otsu, Japan.

*e-mail : [email protected]

Introduction: Synthetic octacalcium phosphate (OCP) has become recognized as a highly osteoconductive bone substitute material based on in vitro and vivo studies 1, 2. It has been shown that OCP combined with collagen (OCP/Collagen) facilitates bone regeneration in comparison with OCP itself 3. The osteoconductivity of OCP/Collagen has been demonstrated also in critical-sized bone defect of dog 4. After that, we examined the effect of OCP/Collagen on bone regeneration in human small bone defect and confirmed the healing at the defect 5. In this study, we performed sinus floor elevation for bone augmentation by OCP/Collagen as a preparation for dental implant treatment in clinical patient.

Materials and Methods: OCP was prepared according to a method of synthesis by mixing calcium and phosphate solution 1. Particle size of OCP was 300 – 500 m in diameter. OCP/Collagen was prepared from pepsin-digested atelocollagen isolated from the porcine dermis and OCP. In this study, we performed sinus elevation in a 41-years old man at left sinus under local anesthesia. OCP/Collagen (approximately 1.8 cc) was used in this surgery. We observed the healing at operative region and confirmed having infection or allergy, or not and radiographic examination or CT scan was performed at 3 or 6 months after operation. Furthermore, we performed bone biopsy at the same time of placement of dental implant, and we examined the newly formed bone histologically.

Results and Discussion: OCP or OCP/Collagen does not have radiopacity under normal X-ray condition. However, radiopacity of OCP/Collagen implanted in sinus increased with time. CT scan showed radiopacity at 3 months after implantation. At 6 months, the radiopacity of bone defect was almost same as surrounding bone. CT value of OCP/Collagen in sinus increased significantly at 3 or 6 months and CT value was almost same as surrounding bone of that in cancellous bone. Histological analysis showed the normal bone tissue including a little remaining OCP. These results showed that OCP/Collagen could convert to bone tissue in clinical use of sinus floor elevation. It was suggested that OCP/Collagen could be a suitable material for bone augmentation.

References : 1. Suzuki O. et al., Tohoku J Exp Med 164: 37-50, 1991. 2. Suzuki O. et al., Biomaterials 27: 2671-2681, 2006. 3. Kamakura S. et al., J Biomed Mater Res B 79: 210-217, 2006. 4. Kawai T. et al., Clin Implant Dent Relat Res 13: 112-123, 2011. 5. Kawai T. et al., Tissue Eng Part A 20: 1336-1341, 2014.

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P-102

The application of oxygen-permeable spheroid culture chip (Oxy chip) in bone tissue engineering

Anada T1*, Kamoya T1, Sato T1, Shiwaku Y1, Takano-Yamamoto T1, Sasaki K1, and Suzuki O1 1Tohoku Univ., Miyagi, Japan *e-mail ([email protected])

Introduction: Three dimensional (3-D) cell cultures have been considered to be a better in vitro model of tissues than monolayer cultures. In order to make large 3-D cellular constructs, spherical cell aggregates (spheroids) have attracted much attention as cellular building blocks. One of the most serious problems of spheroids is oxygen depletion in the inside of cell aggregates. The increase in size of aggregates causes a hypoxia and subsequent necrosis in the core of large aggregates. Therefore, to overcome this problem, we have developed an oxygen-permeable spheroid culture chip (Oxy chip). The Oxy chip enables direct oxygen supply to the cells through the polydimethylsiloxane (PDMS) substrate. We previously reported that the Oxy chip can maintain metabolic functions of hepatoma cells. In addition, the chip can dramatically prevent hypoxia and cell death in the center of spheroids. However, the effects of oxygenation on the osteoblastic differentiation of 3-D cultured mesenchymal stem cells (MSCs) have not been clarified. On the other hand, we previously reported that octacalcium phosphate (OCP), which is one of the synthetic calcium phosphates (CaPs), is effective to the osteoblastic differentiation of stromal cells [1]. The aim of the present study was to examine the effects of oxygen supply by the Oxy chip on the differentiation of 3-D cultured MSCs. Furthermore, we showed the technique for constructing a hybrid spheroid consisting of MSCs and synthetic CaPs including OCP using the Oxy chip.

Materials and Methods: We prepared a spheroid culture chip using a rapid and simple method as previously reported [2]. Mouse bone marrow-derived MSC (D1) cells were cultured in osteogenic differentiation medium. Proliferation of cells was measured by a DNA quantification kit. Osteoblastic differentiation of D1 cells was analyzed by alkaline phosphatase (ALP) activity, real-time PCR, and DNA microarray. Moreover, we compared the formation of aggregates and osteoblastic differentiation of D1 cells with or without three types of CaP materials; hydroxyapatite (HA), β-tricalcium phosphate (β-TCP), and octacalcium phosphate (OCP).

Results and Discussion: Our data demonstrated that the Oxy chip promoted the differentiation of D1 cells into osteoblasts more quickly and efficiently than the conventional non-oxygen permeable chip and monolayer culture. DNA array analysis showed the results that the Oxy chip facilitated osteoblastic differentiation rather than chondrogenic differentiation [3]. Furthermore, the Oxy chip was able to generate CaP/cell hybrid spheroids rapidly and easily. OCP showed the greatest ability to increase the ALP activity of 3-D cultured D1 cells [4]. Together, the Oxy chip and CaP/cell hybrid spheroids generated by the Oxy chip could be useful for bone regenerative therapy.

References: [1] T Anada et al., Tisuue Eng PartA, 14 (2008) 965-978, [2] T Anada et al., Biomaterials, 33 (2012) 8430-8441, [3] T Kamoya et al., Sens Actuators B, (2016) in press. [4] T Anada et al., Regenerative Therapy, (2016) in press.

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P-103

Influence of surface physicochemical modification on wettability of zirconia (TZP) and behaviour of oral keratinocytes

Yoshinari M1*, Takemoto S1 1Tokyo Dental College, Tokyo, Japan *e-mail: [email protected]

Introduction: Surface modification technologies are available for tetragonal zirconia polycrystal (TZP) to enhance its bioactivity. The surface wettability of an implant material is one of the important factors in the process of osseointegration and making a soft tissue barrier, possibly regulating protein adsorption and subsequent cell behavior. The aim of the present study was to clarify the effect of physicochemical modification of TZP on wettability and behavior of oral keratinocytes to determine the potential of such treatment in application to dental implants.

Materials and Methods: Surface physicochemical modifications were carried out with atmospheric-pressure plasma (Plasma) and ultraviolet light (UV) on yttria-stabilized TZP (TZ-3YB-E, Tosoh, Japan). The specimens without physicochemical modifications were used as a control. The surface wettability of the samples was characterized by contact angle measurement with double distilled water using a contact angle meter (Phoenix α, Meiwa-forces, Japan). The obtained specimens were also subjected to storage under various conditions to evaluate their potential to maintain superhydrophilicity. An X-ray photoelectron spectroscopy (XPS) analysis (AXIS- ULTRA, Kratos-Shimadzu, Kyoto, Japan) was performed to determine the composition of the outermost surface and chemical shift, using an X-ray source of Al Kα (monochrometer), 15 kV and 10 mA to determine the intensity of Zr3d, C1s and O1s. The binding energy of each spectrum was calibrated with a C1s of 285.0 eV. Cell culture experiments were performed using human oral keratinocytes (HGEPs, CELLnTEC, Bern, Switzerland). The viability of the attached cells, mRNA expression of laminin γ2 and integrin β4, distribution of laminin γ2 and integrin β4, cell area, and cell morphology were assessed.

Results and Discussion: Surface wettability was greatly increased by surface modification with both Plasma and UV treatments, resulting in superhydrophilicity. Furthermore, superhydrophilicity was maintained, even after immersion in an aqueous solution. XPS analyses revealed that a remarkable decrease in carbon content and the introduction of hydroxyl groups were responsible for the observed superhydrophilicity. The results of cell culture experiments showed that no differences in the viability of attached cells were recognized among the conditions. However, expression of laminin γ2 and integrin β4 as well as cell morphology was promoted only in Plasma specimens even though superhydrophilicity was obtained in both Plasma and UV specimens compared with the untreated control specimen. The photocatalytic activity was believed to be closely involved in the above-mentioned differences. The results of this study suggest that TZP surface treated with plasma promotes the initial attachment capability of human oral keratinocytes with enhancing the extracellular matrix such as lamininγ2, leading to an important consideration in the clinical application of this technology.

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Induced pluripotent stem cells: A potential cell source for regenerative dentistry Abdullah AN1, 2*, Tanimoto K2, and Kato K1 1 Department of Biomaterials, Institute of Biomedical & Health Sciences, Hiroshima University, Japan 2 Department of Orthodontics, Institute of Biomedical & Health Sciences, Hiroshima University, Japan * [email protected]

Introduction: Cell-based therapy is considered as one of the most convenient approach in regenerative dentistry, especially for tooth regeneration. Several research groups have successfully regenerated tooth-like structure by mimicking the reciprocal interactions between mesenchyme and epithelial cells. However, clinical applications of such technologies are limited primarily due to the lack of appropriate sources of dental epithelial cells. The discovery of induced pluripotent stem cells (iPSCs) in 2006 has opened-up an attractive alternative source of dental epithelial cells due to their unique characteristics and less ethical issues. In this study, we examined the differentiation potential of iPSCs into dental epithelial cells using neurotrophin 4 (NT-4). Materials and Methods: The differentiation of iPSCs into dental epithelial was initiated through the formation of embryoid bodies (EBs) using the hanging drop technique with/without the presence of NT-4 for 6 days, and further expanded under serum or serum- free medium for additional 9 days. The morphological changes were observed. mRNA expression of stem cell marker (Oct4), ectoderm marker (K18), epithelial progenitor markers (p63 and CK14) and ameloblast specific markers (AMG, AMBN, and DSPP) were analyzed to assess the differentiation of iPSCs into dental epithelial cells. Immunostaining and western blot for the expression of epithelial progenitor markers (CK14 and p63) and ameloblast marker (AMG) were also carried out. Results: Our findings revealed that the addition of NT-4 significantly triggered up-regulation of p63 and CK14, suggesting that NT-4 provides an inductive microenvironment for the differentiation of iPSCs into dental epithelial cells. Expansion under serum-free culture condition promotes the formation of differentiated cells with cobblestone-like morphology and significantly down-regulated the expression of Oct4 and K18. Formation of iPS-derived dental epithelial-like cells was further confirmed by the high expressions of AMBN, AMG, and DSPP. Conclusion: Based on the findings obtained in this study, we conclude that dental epithelial- like cells can be derived from iPSCs through the stimulation with NT-4 together with serum- free culture condition. Further study in this field will give a significant contribution to the regenerative dental technologies and their clinical applications.

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Designed SDF-1 chimeric protein for bone tissue regeneration

Date T, Hirata I, Hattori S, Matsuda C, Nakano A, Yamakado N, Tanimoto K, and Kato K* Department of Biomaterials, Hiroshima University, Hiroshima, Japan * [email protected]

Introduction: For regeneration of bone tissues in cleft palate, much attention has recently been paid to tissue engineering using biodegradable scaffolds. I the present study, we focused on the cell migration promoting effect of stromal cell-derived factor 1 (SDF-1). It is hypothesized that endogenous bone-forming cells are recruited into SDF-1 immobilized scaffolds, and the rate of bone regeneration is accelerated in the scaffolds. In this study, we designed and synthesized a SDF-1 chimeric protein with a scaffold- binding peptide (SDF1-KL5) and evaluated it for the promotion of bone regeneration in a biodegradable scaffold loaded with the SDF-1 chimeric protein.

Materials and Methods: SDF1-KL5 was synthesized using a bacterial expression system. Molecular size and secondary structure of SDF1-KL5 were analyzed by SDS-PAGE and circular dichroism (CD) spectroscopy. Cell migration was assessed by the Boyden chamber method with a medium containing SDF1-KL5. The expression of SDF-1 receptor (CXCR4) on osteoblast-like cells (MG63) and mesenchymal stem cell-like cells (MSC; UE6E7T-3) was analyzed by RT-PCR. The affinity between SDF1-KL5 and poly(L-lactide-co-ε-caprolactone) (PLACL) was analyzed by surface plasmon resonance (SPR) method. PLACL sponges were prepared by salt-leach technique and were immersed in SDF1-KL5 solution to attach this chimeric protein on the inner surfaces. The amount of immobilized SDF1-KL5 was measured by BCA method. The SDF1-KL5 immobilized scaffolds (SDF1-KL5-Scaffold group) were implanted into bone defects with 5 mm in diameter created in the rat cranial bone. For comparison, scaffolds without SDF1-KL5 (Scaffold group) were implanted into the defect. A sham operation was also done without scaffolds (control group). The volumes of bone defects were measured by micro-CT 0 to 28 weeks post-operation.

Results and Discussion: The molecular weight and the secondary structure of synthesized SDF1-KL5 were coherent to their prediction. SDF1-KL5 enhanced the migration of osteoblasts and MSCs. These results showed that SDF1-KL5 was appropriately synthesized in terms of thes structure and function. The dissociation constant between SDF1-KL5 and PLACL was found to be 9.8 x 10-8 M by SPR and the amount of SDF1-KL5 immobilized on PLACL scaffolds was 1.6 ng/mm2. These results indicated that SDF1-KL5 was strongly bound on the scaffold surface. Although the bone defect volumes increased with growth of rat, the defect volume in SDF1- KL5 Scaffold group was significantly smaller than in Scaffold and control groups. This suggested that the bone regeneration on the scaffold was accelerated by incorporating SDF1- KL5 into the scaffolds.

Conclusion: SDF1-KL5 accelerated bone regeneration in the PLACL scaffolds probably through the enhanced recruitment of endogenous bone-forming cells into the scaffolds.

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The surface treatment to realize both antibacterial property and bioactivity

Tsutsumi Y1*, Shimabukuro M1, Ashida M1, Chen P1, Doi H1, and Hanawa T1 1Tokyo Medical and Dental Univ., Tokyo, Japan *[email protected]

Introduction: Titanium (Ti) is widely used in both orthopaedic and dental fields due to their good mechanical properties, high corrosion resistance, and biocompatibility. Micro-arc oxidation (MAO) is a useful surface treatment based on electrochemical reactions under high voltage in a specific electrolyte. MAO treatment can easily alter the surface properties of metals and is also effective to improve the bioactivity of Ti. Our previous study revealed that certain amounts of extra elements presented in the electrolyte are incorporated in the porous oxide layer during the MAO process. Therefore, in this study, MAO treatment was performed on Ti in silver (Ag)-containing solution. The incorporation of Ag in the porous oxide layer can lead the slight release of Ag ions and the improvement of antibacterial property after implantation in a living body. The surface layer of MAO-treated Ti was characterized by surface analyses. The behavior of the Ag-ion release from the oxide layer was also evaluated by immersing in a simulated body fluid. ICP-AES was performed to determine the concentration of Ag ions in the fluid.

Materials and Methods: A commercially pure Ti rod was sliced to make disk-shaped specimens with 8 mm in diameter and 1.5 mm in thickness. The surfaces of the disks were grinded with up to #800 grid SiC abrasive papers followed by ultrasonic cleaning in acetone and ethanol. The specimen was fixed in a polytetrafluoroethylene holder that allows exposing to an electrolyte only the specimen surface (7.0 mm in diameter). A Type-304 stainless steel plate was used as a counter electrode. The electrolyte for MAO treatment was 0.1-mol L-1 calcium glycerophosphate, 0.15-mol L-1 magnesium acetate, and 0-10 mmol L-1 silver nitrate. After pouring the electrolyte into the electrochemical cell, positive voltage was applied for 8-10 min. The surface morphologies and compositions of the specimens were analyzed using a scanning electron microscope with an energy-dispersive X-ray spectroscope (SEM/EDS). X-ray diffraction (XRD) was also performed with a diffractometer to identify the crystal structure of the oxide layer. 0.9mass% NaCl solution was used as a simulated body fluid to evaluate the Ag ion release from the MAO-treated specimen. The solution was replaced every 7 d to simulate the refreshment of the body fluid inside a living body. ICP-AES was used to measure the concentration of the Ag ions in the solution.

Results and Discussion: The porous oxide layer was formed on the MAO-treated Ti. The morphology of the layer was not changed by the presence of Ag ions in the electrolyte. The EDS results showed that Ag was incorporated in the oxide layer. From the dissolution test, we confirmed that the Ag ion release was kept for at least 4 months. From the result of the antibacterial evaluation test, bacteria (Escherichia coli) could proliferate on both untreated and MAO-treated Ti without Ag. On the other hand, number of the viable bacteria apparently decreased when it was incubated on the MAO-treated Ti with Ag. In addition, initial attachment, proliferation, and calcification of MC3T3-E1 cells were not affected by the presence of Ag when MAO was performed with the electrolyte containing below 0.001 mM of AgNO3. Thus, we confirmed that antibacterial property can be achieved without losing hard-tissue compatibility of Ti by MAO treatment.

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Cytotoxicity of methacrylate-based resin monomers as evaluate by an anti- oxidant responsive element-luciferase reporter assay

Egashira M*, Suzuki T, Fujimoto K, Kanamori T, and Kawai T Aichi-Gakuin Univ., Nagoya, Japan * [email protected]

Introduction: Monomers which contain a methacryloyloxy group are routinely used in dental practice, and methacrylate derivatives with several functional groups have been currently developed for various applications. We previously showed that an anti-oxidant responsible element (ARE)- firefly luciferase reporter system for evaluating cytotoxicity of methyl methacrylate (MMA) and containing a hydroxyl group; hydroxyethyl methacrylate (HEMA). The results suggested that introduction of a hydroxyl group in a methacrylate-based resin monomer increases its intracellular electrophilic reactivity and cytotoxicity. This purpose of this study is to observe the cytotoxicity relationship of (meth)acrylate derives carrying an amid group or another methacrylate group, using the improved ARE-luciferase reporter assay system. Materials and Methods: MMA, HEMA, N-(2-hydroxyethyl) acrylamide (HEAA) and dimethacrylate hydrophilic monomer; triethylene glycol dimethacrylate (TEGDMA) were used. HepG2-AD13 cells, subclonal HepG2 cells stably transfected with an ARE-destabilized luciferase reporter vector (pGL4.24-2E-Neo), were previously established [1]. The HepG2-AD13 cells were seeded in 24- well culture plates (Falcon, Corning, NY, USA) at a density of 6.0 × 105 cells/mL (3.0 × 105 cells/well), and then cells were incubated for 24 hours at 37 °C under 5% CO2. The culture medium was then replaced with 500 µL of the fresh medium with or without various concentrations of resin monomers, which were dissolved directly in D-MEM supplemented with 10% FBS (each solution was used immediately after preparation). Cells were then incubated at 37 °C for 6 hours under 5% CO2. Thereafter, cells in each well were lysed with 100 µL of Passive Lysis Buffer (Promega Corporation, Madison, WI, USA) (the plates were stirred for 15 minutes with a rotary shaker). The lysates were assayed for firefly luciferase activity by using the Single-Luciferase Reporter Assay System (Promega Corporation). Luminescence was measured in relative light units using a luminometer (AB-2200 Ver. 2.61D, ATTO Corporation, Tokyo, Japan) with an integration time of 10 seconds. The ARE activity based on the luminescence intensity is expressed as fold activation to the control value obtained without additives. Results and Discussion: Dose-response curves for ARE activation by HEAA and TEGDMA were almost same shape as HEMA, like a bell. The curve of TEGDMA reached the peak top at lower concentration approximately 2 mM than that of HEMA (between 5 and 10 mM). In contrast, the curve of HEAA reached the peal top at higher concentration approximately 20 mM. This study suggested that the monomers containing amid group are lower cytotoxic monomers than hydroxyl group. References: 1) A. Orimoto, et al. : Effect of 2-hydroxyethyl methacrylate on antioxidant responsive element-mediated transcription: a possible indication of its cytotoxicity, PLoS One. 8 (3) (2013) e58907.

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Preparation of antibacterial high-strength dental stone using with titanium dioxide nanopowder as photocatalyst

Muraoka K, Yoshida K, Miyahara H, Ikeda H, Nagamatsu Y*, Awano S, Shimizu H Kyushu Dental Univ., Kitakyushu, Japan *[email protected]

Introduction: Gypsum model contaminated with surviving bacteria attached impression after insufficient disinfection treatment can cause the infection through the dental technician’s work. However, immersion treatment of model in disinfectant solution might change its surface properties and dimension. We reported that gypsum model made from high-strength dental stone added titanium dioxide nanoparticles at the rate of 7wt% showed antibacterial effect by 16-h UV irradiation. In this study, bactericidal activities were examined in the cases of shorter irradiation and lower rate of contain to high-strength dental stone for prevent influences of mechanical and surface properties of model.

Materials and Methods: Gypsum specimens (20×20mm2, 5mm) were prepared by using with high-strength dental stone (NEW DIAROCK, Ryokadental, Yokkaichi, Japan) added TiO2 P90 (φ:14nm, NIPPON AEROSIL, Tokyo, Japan) at the rates of 0, 1, 2, 3 and 7wt%. Bacteria suspension was prepared to be 1.0×106/mL in Brain Heart Infusion (DIFCO, Detroit, MI, USA) with Staphylococcus aureus NBRC12732. Surface of the gypsum model containing 7wt% TiO2 dropped with 0.15 mL of bacteria suspension was UV-irradiated with Blacklight (FPL27BLB, SANKYODENKI, Tokyo, Japan) for 2, 8 and 16 h from 20 cm distance or not irradiated. After irradiation, surviving bacteria in specimen were extracted to PBS solution by 3-min ultrasonic cleaning and this extract solution or its dilution was added to the agar culturing medium (Netrient Agar, Nissui Pharmaceurical, Tokyo, Japan). Total number of the surviving bacteria was calculated from the number of the colonies in the medium after incubation at 37℃ for 48 h. In specimens containing 1, 2, 3 and 7wt% TiO2, the numbers of surviving bacteria after 8-h UV irradiation were also examined in the same way.

106 Results and Discussion: The numbers of surviving bacteria in the specimens containing 7wt% TiO2 after 104 irradiation are shown in Fig. 1. 8- and 16-h UV irradiations showed high bactericidal activity. No 102 significant difference was observed between these 100 two conditions (p>0.05). Fig. 2 shows the numbers bacteria of Number 0 2 8 16 Irradiation time (h) of surviving bacteria after 8-h irradiation in the case Fig. 1 Number of surviving bacteria in gypsum of change in the contain rate of TiO . In the 2 model (TiO2 :7wt%) after UV-irradiation specimens containing 3 and 7wt% TiO , the number 2 106 of surviving bacteria decreased to less than 102 by irradiation and no significant difference was observed 104 between these contain rates (p>0.05). These contain rates were effective to impart antibacterial properties 102 to gypsum models. These results indicated that high- 100 strength dental stone has the antibacterial effect by bacteria of Number 1 2 3 7 TiO2 (wt%) containing 3wt% TiO2 and 8 h-UV irradiation. Fig. 2 Number of surviving bacteria in gypsum model after UV-irradiation for 8 h

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Optimization of culture conditions for the efficient differentiation of iPS cells into dental epithelial cells

, Miyauchi S1, 2*, Abdullah AN1, Nikawa H2, and Kato K1 1 Department of Biomaterials, Hiroshima University, Japan 2 Department of Oral Biology & Engineering, Hiroshima University, Japan * [email protected]

Introduction: Tooth loss can be considered as one of the major challenges in the dentistry field. Previous studies have revealed that tooth regeneration can be achieved by mimicking the interaction between epithelial and mesenchyme cells. Our current research discovered that the addition of neurotrophin-4 (NT-4) throughout the formation of embryoid bodies (EBs) stimulated differentiation of induced pluripotent stem cells (iPSCs) into dental epithelial-like cells. Concurrent to the finding, the present study was carried out with the aim to identify ideal initial cell numbers and extracellular matrix for efficient differentiation of iPSCs into dental epithelial cells.

Materials and Methods: The formation of EBs from mouse iPSCs was performed with the presence of NT-4 using variable initial cell numbers per drop: 500, 1000, 1500, and 2000 cells per 20 μl of culture media. After 6 days, the EBs were collected and cultured onto three different types of culture dishes i.e. gelatin-coated, laminin-coated, and non-coated culture dishes for 9 days under serum-free culture condition. The distance of migrating cells from the EB was measured and the morphological changes were observed for each case. The differentiation rate of these cells was examined by the expression of CD49f and E-cadherin by flow cytometry.

Results: The results revealed that the number of initial cells showed correlation with the distance of migrating cells from the EB and the expression of CD49f and E-cadherin for all cases except for the case with 2000 cells which displayed inversed relationship. Both cells cultured on gelatin-coated and laminin-coated dishes showed formation of cobblestone-like cells, while cell on the non-coated dish showed formation of fibroblast-like cells.

Conclusion: The results of this study suggest that the altered amounts of initial cell numbers and the types of extracellular matrix coated can affect the differentiation efficiency of iPSCs into dental epithelial cells.

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Influence of Organic Materials on HAp Crystal Formation in Calcium Phosphate-based Cements

Y.Tai*, T.Sekimizu, Y.Asada, T. Chiba, Y.Momoi, N.Hosoya and S.Shimoda Tsurumi Univ., Kanagawa, Japan *e-mail : [email protected]

Introduction: Recently Calcium Phosphate-based Cement (CPC) is used as one of the filling bone prosthetic material corresponding to the shape of a fractured bone part. Advantages of CPC are, 1) doesn’t contain any non-biocompatible material, 2) contains low molecule calcium phosphate materials which react with H2O then decompose, and 3) precipitates biochemically in hydroxyapatite (HAp). However, unlike in the case of bone and dentin, conventional CPC does not contain organic materials. With the goal of development of CPC containing biocompatible organic materials at the beginning we observed HAp crystal formation time- dependently in CPC which collagen fiber or collagen molecule was added to. Materials and Methods: Teeth Mate ®desensitizer (TMD) manufactured by Kuraray Noritake Co Ltd., was used. TMD contains tetra-calcium phosphate and calcium hydrogen phosphate anhydrous as main calcium phosphate chemicals. Specimens were prepared under following 3 conditions. TMD: Powder and liquid of TMD were mixed according to the manufacturer’s instruction. TMD/CF: Collagen fiber (CF) was added to TMD. TMD/CM: Collagen molecule (CM) was added to TMD. All specimens were stored in distilled water and then observed with a transmission electron microscope at 1-week, 2-week and 4-week. Results and Discussion: At 1-week HAp crystals were observed for all specimens where shape of crystals were clearly different among TMD, TMD/CF and TMD/CM. For TMD at 1-week various shape of crystals were observed and at 4-week typical flattened hexagonal crystals of HAp were observed. For TMD/CF long-needle crystals were uniquely seen at 1-week until 4-week. For TMD/CM delay of HAp formation was noted compared with TMD and TMD/CF. It was considered that CF promoted uniform precipitation of HAp crystals where the CF might have to function in regulating the size and shape of the HAp crystals. Conclusions: Organic Materials influenced HAp crystal formation in calcium phosphate-based cements. Acknowledgements: This research was supported by the Reorganization Support Foundation for Private Universities, MEXT (Ministry of Education, Culture, Sports, Science and Technology-Japan). References: Chiba et al., Applicability of Calcium Phosphate Based Paste as a Tooth Care Material. Jpn J Conserv Dent. 58(3):200-211, 2015. Chiba et al., Remineralization Effects of Calcium Phosphate Based Paste for Tooth Enamel. Jpn J Conserv Dent. 59(1):58-64, 2016

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Cell aggregation of periodontal ligament fibroblasts controlled by micro-patterns

Akasaka T1*, Kaga N2, Yokoyama A2, Abe S1, Yoshida Y1 1Hokkaido Univ., Sapporo, Japan *[email protected]

Introduction: Many micro/nano-structures such as bone and teeth in the body play an important role. Bone architecture is proposed that organization of collagen and hydroxyapatite. Nano hydroxyapatites are oriented to organic collagen fibers, and are hierarchically organized with collagen bundles. Dental enamel is highly organized array of apatite nanocrystals. Further, it is possible to see other many examples of micro/nano-structures in the body. We investigated preparation of micro-pattered polystyrene substrates using laser plotting and nano-imprint methods. And then the behaviors of cell aggregation and movement of the cells on micro-pattern were estimated using human periodontal ligament fibroblasts (HPDLF).

Materials and Methods: The resist (OFPR-5000) was coated on a cover glass and baked at 140°C for 3 min. The patterns were plotted using laser direct lithography system (DDB-201-200) and developed. The micro-patterns of the resist were transferred to light-curing resins by light-curing nano- imprinting, and subsequently to polystyrene substrates by thermal nano-imprinting. Cell movement and aggregation were estimated using HPDLF. The cells were seeded on the patterns, and cultured in DMEM with 10% FBS at 37°C. The behavior of the cells was analyzed by optical microscopy, scanning electron microscopy, and fluorescence microscopy.

Results and Discussion: The resulting patterns have groove patterns of concentric circles with 2 mm maximum diameter, 12.5 to 50 µm pitch, 1.1 to 6.1 µm width, and 0.2 to 0.8 µm depth depending on conditions by laser plotting (Fig. 1). As a result, cell aggregation was observed at the center of concentric circles controlled by micro-patterns (Fig. 2). The degree of cell aggregation was influenced by pitch of concentric circular grooves. Efficient cell aggregation was observed at the narrowest pitch of 12.5 µm and wider width of 6 µm in this study. The mechanism of aggregation is currently under consideration. These data supported that the micro-patterns can control the aggregation and movement of HPDLF cells.

Acknowledgements: This work was partly funded by JSPS KAKENHI Grant Number (No. 25463047).

Fig. 1 Laser microscope image (a) and SEM image Fig. 2 Optical microscope image of Giemsa- (b) of concentric circular micro-patterned stained HPDLF cells after 7d culturing. (a) 50 µm polystyrene. and (b) 12.5 µm groove pitch of concentric circles with 2 mm maximum diameter.

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Assessment of cytocompatibility of nano-sized ceramics particles on several cells E. Seitoku, S. Abe*, Y. Era, N. Kiyama, S. Tsuchiya, K. Nakanishi, Y. Nakagawa, T. Akasaka, J. Iida, Y.Yoshida, and H. Sano Graduate School of Dental medicine, Hokkaido University, Sapporo, Japan *e-mail: [email protected]

Introduction: Nano-sized materials have received much attention for their several applications and their biocompatibility because of recent development of nanotechnology. For their application on biological fields, it is very important to evaluate the biodistribution of administered nano- materials. In our previous study, we succeeded in visualizing the biodistribution of administered nano-materials and quantitative observation [1]. The administered nano-particles were observed in the liver, lung and spleen of mice. In addition, some researchers were interested a “nano-sizing” effect on their biocompatibility. For example, when particles reach nano-level, some of them exhibit toxicity in vitro even if they are considered biocompatible at the macro level. Thus, we focused on cytocompatibility of nano-sized ceramics particles. To assess their cytocompatibility, these particles were exposed to human liver cells and osteoblastic cells. Materials and Methods: For evaluation of cytocompatibili1ty of nano-sized ceramics particles, several types of cells exposed to the nano-particles. As nano-sized ceramics particles, TiO2, Fe2O3, ITO (Indium-Tin oxide), CuO, ZnO, In2O3, Y2O3:Eu (Eu-doped Y2O3 nanoparticles [2]) etc. were investigated. These particles were exposed to cells. The concentrations were varied from 0.01 to 30 ppm. The morphology of cultured cells was observed using SEM and time-lapse observation [2]. The proliferation and cell viability was determined using Cell CellTiter-Glo assay kit (Promega, Madison, WI, USA). Results and Discussion: Even the exposure concentration increased (up to 30 ppm), these cells maintained their cell viability more than 80 %. Also, under long-term exposure, they showed high cell viability [2]. Only CuO particles caused the viability decrease. When the concentration of CuO increased, the viability markedly decreased. In addition, the cytotoxicity showed dependence of the particle size. When the particle size changed from micro-level to nano, they expressed the inflammation. Based on the time-lapse observation of cell behavior, it suggested that the inflammation caused when cells were in contact to CuO particles. In same concentration, nanoparticles existed widely on culture dish surface compared with micro-sized ones. It meant that the cell-particles contact probability increased. Therefore, the inflammation under CuO exposure indicated a “nano-sizing effect”. In this study, several nano-sized ceramics particles indicated good biocompatibility without CuO. Although cytotoxicity of CuO increased depending on the size decrease, nano-sizing effect cannot be a dominant factor on the nanotoxicity. References (Optional): [1] F. Watari et al J. Royal Society Interface, 6, S371 (2009). [2] N. Kiyama, S. Abe, S. Yamagata, Y. Yoshida, J. Iida, Nanoscience and Nanotechnology Letters, (in press).

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Proliferation of mesenchymal stem cells seeded on biodegradable hydroxyapatite–chitosan porous scaffolds

Maretaningtias Dwi Ariani1, Sherman Salim1, Isao Hirata2, Koichi Kato2

1 Department of Prosthodontics, Faculty of Dentistry, Airlangga University, Surabaya, Indonesia 2 Department of Biomaterials, Institute of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan

Introduction Needs for bone repair have increased along with an increase in life expectancy. Currently, special attention is paid to polymer scaffolds that have a three-dimensional structure as a bone graft material in tissue engineering approaches. Chitosan [poly(1,4-D- glucosamine)], which is the product of deacetylation of chitin has become a topic of interests in the development of bone graft materials. Chitosan is biodegradable and non-toxic. To improve cell adhesion and bone formation ability of chitosan, we made here an attempt to combine hydroxyapatite (HA) with chitosan. Generally HA dissolves under acidic conditions and grows in supersaturated environments. Due to such these properties, HA has been clinically used as bone substitutes.

Objective The objective of this study is to develop scaffolds made of HA and chitosan (HA- ChSs) and evaluate cell proliferation in HA-ChSs scaffolds using mesenchymal stem cells (MSCs).

Methods Porous HA-ChSs matrix was made through lyophilization. A chitosan gel was prepared by dissolving 200 mg of chitosan powder (98.7% degree of deacetylation, YSK,

Japan) into 5 mL CH3COOH. After neutralization, 25, 50, 100, 125 and 150 mg of HA powders were added to the chitosan gel and then agitated for 15 min. After centrifugation at 10,000 g for 10 min, excess water was removed and then the HA-ChSs gel was packed into a disc-shaped mold (diameter: 5 mm, height: 2 mm). Then frozen at ‒80 °C for 2 hr and freeze- dried at ‒54 °C for 24 hr. The dried HA-ChSs was irradiated with ultraviolet light for 2 hr. The porous structure of HA-ChSs was observed by scanning electron microscopy (SEM). Proliferation of MSCs in HA-ChSs scaffolds was evaluated at 1, 3, 5 and 7 days after cell seeding.

Results and Discussion Three-dimensional porous structures of HA-ChSs with HA powders attachment were clearly observed by SEM. We further observed that proliferation of MSCs was significantly enhanced by incorporating HA into chitosan. Based on these results, we expect that HA-ChSs may be candidates for bone graft materials in bone tissue engineering.

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MECHANICAL PROPERTIES OF COATING LAYER MODIFICATION CHITOSAN-TiAlCr ON CALCIUM PHOSPHATE CEMENT WITH SOL-GEL METHOD

Atia Nurul Sidiqa1, Bambang Sunendar2, Nina Djustiana 2, Renny Febrida2 1Dental Materials Department, School of Dentistry, Jenderal Achmad Yani University 2Advanced Material Department,Institute Technology of Bandung 3Dental Materials Department, Dentistry Faculty, Padjadjaran University

Introduction: Hydroxyapatite is increasingly used for medical aplication because of its biocompatibility. A very similar components with human bone helps HA form an intimate contact and improve a better adhesion to the bone. However, poor mechanical properties of HA have a limited aplication. The common application are used HA as a coating layer on titanium implant or develop it to produce a better properties. The combination of Ti alloy with HA. Therefore, the development is done by modifying the surface of a multilayer coating of Ti-Al-Cr and hydroxyapatite in calcium phosphate cement that produces strength similar to dental implants.

Objectives: to measure the elastic modulus of synthesis calcium phosphate cement products modified by Ti-Al- Cr and hydroxyapatite.

Methods : Specimens were divided into three groups, CPC without coating treatment (Gr I); CPC with coating layer by layer (CPC / CH / Ti / CH / Al / CH / Cr / HA) (Gr II); multilayer coating of Ti-Al-Cr (CPC / Ti-Al-Cr / HA) (Gr III). Elastic modulus evaluated using UTM.

Results: multilayer coating of Ti-Al-Cr and hydroxyapatite group showed the highest ME value (206.394±1.582MPa) with a significant difference compared with calcium phosphate cement treated layer by layer coatings (67.609±3.131Mpa) and a control group of pure calcium phosphate cement (38.28±6.891MPa).

Conclusion: ME as one of the main mechanical properties for dental implants can be improved by coating multilayer modification Ti-Al-Cr on CPC with sol-gel method.

Key words: implant, calcium phosphate cement, coating, Ti-Al-Cr, hydroxyapatite, elastic modulus

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Disinfection Capacity of Mouthwashes using as Admix Solution of Alginate Impression

Sumadhi Sastrodihardjo, Kholidina Imanda Harahap Departemen Ilmu Material dan Teknologi Kedokteran Gigi Fakultas Kedokteran Gigi Universitas Sumatera Utara Jl. Alumni No.2 Kampus USU Medan E-mail : [email protected]

Abstract

Background: Immersion and spraying with disinfectant solution are the effective disinfection methods for alginate impression. However these methods causes dimensional changes on impression. Therefore mouthwash, which also has disinfectant function on microorganism and can be used as mixing solution for alginate impression material that can be learned in this study .

Purpose: The aim of the study is to analyze anti bacterial capacity of mouthwashes that used as admix solution for alginate impression.

Methods: Samples was made from alginate impression material in tablet form with 15mm of diameter and 1mm of thickness. Total samples are 35 which are divided into 7 test groups, admix with aquadest as control, admix with clorhexidine 0.1%, admix with povidon iodine, admix with non-alcohol mouthwash, immerse in clorhexidine 0.1% , immerse in povidon iodine, immerse in non-alcohol mouthwash. Sample was put on incubated Stafilococcus aureus in petri dish and kept in incubator for 24 hours at 37°C. Inhibition zone was measured by using digital caliper. Statistic analyzis was performed by using ANOVA one way and non-pair t-test.

Results: The admix with clorhexidine 0.1% show the inhibition zone by 8.09 mm, non-alcohol mouthwash show 2.91mm and admix with povidon iodine by 0.52 mm. There are significance differences between test group with p value = 0.00. By using immersion methods they show 7.63mm inhibition zone for clorhexidine 0.1%, 1.51mm for povidon iodine and 0.91mm for non-alcohol mouthwashes. There is insignificant difference between admix and immerse with chlorhexidine 0.1% with p= 0.38 (p>0,05) but significance on povidon iodine and non-alcohol mouthwashes.

Conclusion: The chlorhexidine mouthwash 0,1% has the equal anti bacterial capacity when used for admix solution nor immerse the alginate impression.

Keywords: alginate impression, mouthwash, disinfection.

186

P-118

Oshphronemus Gouramy scales is a candidate for periodontal tissue engineering

Chiquita Prahasanti1, Agung Krismariono1, Denny Tri Wulandari2 1Periodontics Department, Faculty of Dental Medicine, Airlangga University Surabaya – Indonesia 2Periodontist, Majene District, West Sulawesi, Indonesia

Abstract Background: Conventional periodontal treatment were all not sufficient to resolve the problem of periodontal bone defect. Recently, tissue engineering is believed can facilitate periodontal regeneration. Treatment of periodontal bone defect using bone graft has been mostly done. Bone graft proven to produce regeneration of bone to achieve the bone volume. Oshphronemus Gouramy scales have potential capacity as bone graft because it’s scales are suspected containing much collagen.

Purpose: This study investigated the collagen content of Oshphronemus Gouramy scales.

Methods: 100gr Oshphronemus Gouramy scales were immersed in acetic acid 6% for 7x24 hours. pH scales that have been expand were neutralized by running the water until collagen fibers appearance. Collagen fibers cloth were formed completely in pH 7. Then, collagen fibers cloth were filtered by filter paper and dried by freeze-dry method. After all procedure was done this material were analized by FTIR (Fourier Transform Infrared Spectroscopy) to characterization and analysis the functional group of it. Control group using collagen which commonly used.

Results: The FTIR result analysis of Oshphronemus Gouramy scales showed that FTIR spectrum pattern was virtually identical between the experiment group and control group. The identical pattern of two groups was assumed that Oshphronemus Gouramy scales and collagen which commonly used have similar collagen content.

Conclusion: Oshphronemus Gouramy scales is a candidate for periodontal tissue engineering because it’s scales containing much collagen.

Key words: Oshphronemus Gouramy scales, collagen, periodontal tissue engineering

187

P-119

The Differences of Lipopolysaccharide And The Whole Bacteria Porphyromonas Gingivalis Characteristics In Rat Immune Response

N. Sindy Cornelia, Nuraini Prawati, Pradopo Seno, Endaryanto Anang, J.M.Harjanto Departement of Paediatric Dentistry Faculty of Dentistry, Airlangga University. Surabaya – Indonesia

Abstract Introduction : Lipopolysaccaride(LPS) is a common material used to represent the existence of bacteria, as Porphyromonas gingivalis (Pg) LPS. In pediatric dentistry, gingivitis is a common disease, where one of the pathogens causing the problem is Porphyromonas gingivalis (Pg). The same research on Pg LPS, as Pg endotoxin, has a controversial results. For that reason, this study eager to know the characteristics immune responses of Pg LPS and Pg whole bacteria.

Objective : To explain the characteristics immune responses of Pg LPS and Pg whole bacteria.

Material and Methods: Six Wistar rats were divided in two groups. The first group were injected with whole bactery Pg 9x1011 CFU and the second group injected with Pg LPS 3g/ml, both intrasulcularly. After eleven days, the innate immune responses were examined the level of TLR-2 and TLR-4, while adaptive immune responses were examined the level of IgE and IgG4 by ELISA (Enzyme Linked Immunosorbent Assay).

Result: After eleven days, for innate immune response, in LPS Pg injected group, TLR-2 was decrease 2,29 + 1,26 ng/ml and TLR-4 was increase, while the opposite occurs in whole bactery Pg group, TLR2 was increase 0,42 + 0,22 ng/ml and TLR-4 was decrease 1,42 + 0,92 ng/ml. The result for adaptive immune response, in LPS Pg injected group, IgE was extremely increase 95,31 + 22,3 ng/ml and IgG was increase 53,6 + 10,8 while the opposite occurs in whole bactery Pg group, IgE was slightly increase 1,03 + 4,57 ng/ml and IgG was decrease 0,77 + 4,87 ng/ml.

Conclusion: The LPS and Pg LPS have different cahracteristic, specialy in this study was showed in the both immune response, innate and adaptive have a contradiction result.

Keywords: Lipopolysaccharide, Porphyromonas gingivalis, Immune response.

188

P-120

APPLICATION NANOPOWDER STICHOPUS HERMANII TO PREVENT RELAPS ORTHODONTIC BY PERIODONTAL LIGAMENT REMODELLING

Noengki Prameswari*, Puguh BP**, Sudibyo*** Orthodontics Laboratory*, Dental Material Laboratory**, Public Health Laboratory***, Dentistry Faculty Hang Tuah University

Abstract : Background:. Relapse tendency occurs 33–90 per cent after at least 10 years post-treatment Relapse Orthodontic is the return, following correction, any change from the final tooth position at the end of treatment relapse, could be a return to the original teeth position, caused by periodontal,occlusal,soft tissue factor and growth. Nanopowder Stichopus hermanii contain various active ingredient such as collagen, flavonoid, chondroitin sulphate, cell growth factor, EPA DHA, that might have role to periodontal ligament remodelling in reducing relapse orthodontic. Objectives: The aim of this study is to investigate application of Nanopowder Stichopus hermanii to prevent relaps orthodontic by periodontal ligament remodelling through MMP-8, Integrin α2β1, and Collagen type 1 parameter. Methods: The experiment was held by Post Test Only Group design. Twenty four male Cavia Cobaya were divided into three groups. K(-) group as negative control group (without treatment), K(+) group as positive control group which were applied with relaps orthodontic forces, and the other groups P, were applied with relaps orthodontic forces and nanopowder Stichopus hermanii 3 %. After treatment the cavia cobaya were sacrificed. MMP-8, collagen type 1 and Integrin α2β1 expression is examined with immunohistochemistry Results: Application nanopowder Stichopus hermanii can reduce relaps until 30%. This study showed MMP-8 expression means and SD in K(-), K(+), P are 4±0,76 ; 2,63±1,4; 1,88±0,8 ; Collagen type 1 expresssion are 16,7±2,54 ; 8±2,07; 26,25±2,38 and Integrin α2β1 expression are 9.9±1,91 ; 5,25±3,24; 16,38±4,96. There was significantly differences in group P compare to K(-) and K(+). Conclusion : Application of Nanopowder Stichopus hermanii 3% have role in periodontal ligament remodelling through decreasing MMP-8, increasing Integrin α2β1 and Collagen type 1 parameter to prevent relaps orthodontic until 30%

Keywords: Nanopowder Stichopus hermanii, MMP-8, Integrin α2β1, collagen type 1, relaps orthodontic.

Correspondence : Noengki Prameswari. Orthodontic Laboratory. Arif Rahman Hakim 150 Surabaya. Email : [email protected]

189

P-121

Application of recombinant secretory leucocyte protease inhibitor amnion membrane on incision of Rattus norvegicus to the number of fibroblasts

Elly Munadziroh1, Cornelia Johana Corputty 2 dan Intan Nirwana3 Staf Pengajar Departemen Material Kedokteran Gigi 1,3 Mahasiswa Program Sarjana Kedokteran Gigi1 Fakultas Kedokteran Gigi Universitas Airlangga Surabaya - Indonesia

Background: Secretory leukocyte protease inhibitor (SLPI) is a protein found in various places the human body including in amnion membrane. Secretory leukocyte protease inhibitor function are to inhibit the protease, control leukocytes activity, regulate TGF-β, anti inflammatory, anti bacterial, anti retroveral (human immunodeficiency virus). The cases of incision on the oral muccosal tissue often found in Dentistry. SLPI can be used as natural biomaterial in accelerating the process of wound healing, which is characterized by an increase in the number of fibroblasts cells through the inhibiton of elastase production.

Purpose: To know the application of recombinant SLPI amnion membrane fluid on incision wounds of Rattus norvegicus in increasing the number of fibroblasts on day 4th.

Methods: Incisions was carried out on the 24 rats (Rattus norvegicus) males, divided to 4 groups, control group (not given anything), P1 (SLPI 0.03 cc), P2 (SLPI 0.045), and P3 group (SLPI 0.06 cc). The 24 rats (Rattus norvegicus) will be executed on day 4th. The samples were tested by histopathological (HPA) test to observe the number of fibroblast cells using microscope with 400x magnification.

Results: The difference of fibroblasts cells number will be evaluated statistically using Kruskal- Wallis test, statistic analistic demonstrated that there are significant differences between the control and treatment groups (p < 0.05, p=0.000). Mann-Whitney statistical test demonstrated that the difference of control group and all treatment group have significant differences (p < 0.05, p = 0.04).

Conclusion: The application of 0,06 cc recombinant SLPI amnion membrane on incision wound Rattus norvegicus showed the highest number of fibroblasts cells on day 4th.

Key words: Recombinant secretory leucocyte protease inhibitor, amnion membrane, wound healing, fibroblasts

190

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130529_advertising_soaric_Ad_En_297x210_sw.indd 1 2013/05/29 13:33:42