DOCSLIB.ORG

Open Full Page

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Open Full Page Published OnlineFirst December 17, 2015; DOI: 10.1158/0008-5472.CAN-15-0884 Cancer Tumor and Stem Cell Biology Research Eva1 Maintains the Stem-like Character of Glioblastoma-Initiating Cells by Activating the Noncanonical NF-kB Signaling Pathway Naoki Ohtsu1, Yuka Nakatani2, Daisuke Yamashita3, Shiro Ohue3, Takanori Ohnishi3,and Toru Kondo1,2 Abstract Glioblastoma (GBM)–initiating cells (GIC) are a tumorigenic as Eva1 overexpression enhanced these properties. Eva1 deficien- subpopulation that are resistant to radio- and chemotherapies cy was also associated with decreased expression of stemness- and are the source of disease recurrence. Therefore, the identifi- related genes, indicating a requirement for Eva1 in maintaining cation and characterization of GIC-specific factors is critical GIC pluripotency. We further demonstrate that Eva1 induced GIC toward the generation of effective GBM therapeutics. In this study, proliferation through the activation of the RelB-dependent non- we investigated the role of epithelial V-like antigen 1 (Eva1, also canonical NF-kB pathway by recruiting TRAF2 to the cytoplasmic known as myelin protein zero-like 2) in stemness and GBM tail. Taken together, our findings highlight Eva1 as a novel tumorigenesis. Eva1 was prominently expressed in GICs in vitro regulator of GIC function and also provide new mechanistic and in stem cell marker (Sox2, CD15, CD49f)-expressing cells insight into the role of noncanonical NF-kB activation in GIC, derived from human GBM tissues. Eva1 knockdown in GICs thus offering multiple potential therapeutic targets for preclinical reduced their self-renewal and tumor-forming capabilities, where- investigation in GBM. Cancer Res; 76(1); 171–81. Ó2015 AACR. Introduction nance of neural stem cells (NSC; refs. 2–5). NSCs exist only in the subventricular zone and hippocampus (6, 7), both of which Gliomas are brain tumors possessing the characteristics of glial contain a special microenvironment (niche) for the maintenance cells, astrocytes, and oligodendrocytes, and have been classified of NSCs, whereas GBM arises in many areas in the brain. It into four grades (WHO grade I–IV) based on their pathologic currently remains unknown how GICs maintain their stemness features. Glioblastoma (GBM) is the most malignant glioma in the brain; whether GICs generate their preferable niche any- (WHO grade IV), and patients with GBM have a median survival where or use an unknown mechanism for their maintenance in of approximately one year. Despite tremendous efforts to effec- non-NSC niches. tively treat GBM, the overall survival rates of patients with GBM We previously established the mouse GIC lines (mGIC), have remained unchanged over the past few decades. NSCL61 and OPCL61, by overexpressing an oncogenic HRasL61 The discovery of GBM-initiating cells (GIC) has had a signif- in p53-deficient NSCs and oligodendrocyte precursor cells (OPC), icant impact on GBM research (1). GICs have a strong self-renewal respectively (8, 9). These mGICs formed transplantable GBM with capability, express stem cell markers, such as CD133 (also known hypercellularity, pleomorphism, multinuclear giant cells, mitosis, as Prominin1), Sox2, CD15 (also known as Stage-Specific Embry- and necrosis, even when as few as ten cells were injected into the onic Antigen 1 and Lewis X), and CD49f (also known as integrin a brains of nude mice. These findings indicated that they were 6), and are more resistant to radiotherapy and chemotherapy highly enriched in bona fide GICs. Using DNA microarray anal- higher than non-GICs (2–5). GICs have also been shown to ysis, we compared the gene expression profiles of mGICs with exploit the signaling pathways that are involved in the mainte- those of their parental cells and identified genes that increased and decreased in mGICs. By evaluating the candidate genes using 1 human GICs (hGICs) and GBM tissues, we have successfully Division of Stem Cell Biology, Institute for Genetic Medicine, Hokkaido fi – University, Sapporo, Hokkaido, Japan. 2Laboratory for Cell Lineage selected potential GIC-speci c genes (8 11). Modulation, RIKEN Center for Developmental Biology, Kobe, Hyogo, Among the candidate genes evaluated, we focused on epithelial 3 Japan. Department of Neurosurgery, Ehime University Graduate V-like antigen 1 (Eva1, also known as myelin protein zero-like 2). School of Medicine, To-on, Ehime, Japan. Eva1 was originally identified as an immunoglobulin superfamily Note: Supplementary data for this article are available at Cancer Research member expressed on the developing thymus epithelial cell Online (http://cancerres.aacrjournals.org/). membrane and disappeared in the developed one (12–14). Eva1 Current address for Y. Nakatani: Division of Bio-Function Dynamics Imaging, was also shown to be involved in the T-cell development through Center for Life Science Technology, RIKEN, Kobe, Hyogo, Japan. the Eva1–Eva1 homophilic interaction between CD4/CD8 dou- Corresponding Author: Toru Kondo, Division of Stem Cell Biology, Institute for ble-positive cells and thymus epithelial cells in the early embryo Genetic Medicine, Hokkaido University, Kita-15, Nishi-7, Kita-ku, Sapporo 060- (12–14); however, there was no detectable phenotype, including 0815, Japan. Phone: 81-11-706-6082; Fax: 81-11-706-7870; E-mail: the hematopoietic development, in the Eva1 knockout mice [email protected] (12–15; Ohtsu and colleagues, unpublished observation). doi: 10.1158/0008-5472.CAN-15-0884 Notably, it has not been shown that Eva1 was involved in either Ó2015 American Association for Cancer Research. tumorigenesis or stemness. These findings led us to investigate www.aacrjournals.org 171 Downloaded from cancerres.aacrjournals.org on September 27, 2021. © 2016 American Association for Cancer Research. Published OnlineFirst December 17, 2015; DOI: 10.1158/0008-5472.CAN-15-0884 Ohtsu et al. whether Eva1 can be a novel GIC marker and/or a potential polyclonal anti-Eva1 (5 mg/mL), mouse monoclonal anti-CD15 therapeutic target. (5 mg/mL; BD Pharmingen) and biotinylated mouse monoclonal Here, we show the evidence that Eva1 is expressed on GICs and anti-CD49f (Affymetrix; 1:100). Antibodies were detected with its modulation impacts GIC characteristics, including stemness- APC-conjugated goat anti-mouse IgG (Santa Cruz Biotechnology; related gene expression, side population, self-renewal activity and 1:200) and PE-conjugated goat anti-rabbit IgG (Santa Cruz Bio- tumorigenesis, through the noncanonical NF-kB signaling path- technology; 1:200). The cells were analyzed in an Aria II (Becton way, providing a new molecular mechanism that maintains GIC Dickinson) using a dual-wavelength analysis (488 nm solid-state characteristics. laser and 638 nm semiconductor laser). Propidium iodide– positive (i.e., dead) cells were excluded from the analysis. The SP was analyzed as shown previously (11). Reserpine Materials and Methods (10 mmol/L), an inhibitor of some ABC transporters, was used Animals and chemicals to identify SP. Mice were obtained from the Laboratory for Animal Resources and Genetic Engineering at the RIKEN Center for Developmental Human brain tumors Biology (Hyogo, Japan) and from Charles River Japan, Inc. All Human GICs were used according to the research guidelines of mouse experimental protocols were approved by the Animal Care the Ehime University Graduate School of Medical Science and the and Use Committees of RIKEN Center for Developmental Bio- Hokkaido University Institute for Genetic Medicine. Poly(A)þ logy, Ehime University (Ehime, Japan), and Hokkaido University RNA was prepared using a QuickPrep mRNA Purification Kit (GE (Hokkaido, Japan). Chemicals and growth factors were purchased Healthcare). Control human brain total mRNA was purchased from Sigma-Aldrich and Peprotech, respectively, except where from Invitrogen. cDNA was synthesized using a Transcription First otherwise indicated. Strand cDNA Synthesis Kit (Roche). Cell culture Intracranial cell transplants in the NOD/SCID mouse brain and Mouse primary NSCs, NSCL61 cells, human NSCs (hNSCs, brain-tumor histopathology Invitrogen), and GICs (hGICs, E2, E3 and E6) were cultured in NSCL61 cells or hGICs were suspended in 5 mL of culture DMEM/F12 (Gibco, BRL) supplemented with bFGF (10 ng/mL) medium and injected into the brains of 5- to 8-week-old female and EGF (10 ng/mL; NSC medium), as described previously NOD/SCID mice under anesthesia with 10% pentobarbital. The – (8 11, 16). For immunostaining, cells were cultured in chamber stereotactic injection coordinates were 2 mm forward from the fi slides (Nunc) precoated with bronectin and poly-D-lysine, as lambda, 2 mm lateral from the sagittal suture, and 5 mm deep. described previously (16). Mouse brains were dissected, fixed in 4% paraformaldehyde at Immunochemistry 4 C overnight, transferred to 70% ethanol, processed on Tissue- fi Immunostaining of paraffin-embedded human brain-tumor Tek VIP (Sakura Finetek Japan), and embedded in paraf n. sections (6 mm thick) and mouse cells or brain sections was Coronal sections (6-mm thick) from the cerebral cortex were performed as described previously (8). Eva1 was retrieved by prepared on a microtome and stained with hematoxylin and HistoVT One according to the supplier's instructions (Nacalai eosin (H&E). Tesque). The sections were permeabilized with 0.3% TritonX-100 in PBS for penetration, treated with a blocking solution (2% skim RT-PCR RT-PCR was performed as described previously (16), with the milk, 0.3% Triton X-100, and PBS) for 1 hour, and incubated with primary antibodies for 16 hours at 4C. Cells were fixed and cycle parameters of 20 seconds at 94 C, 30 seconds at 57 C, and 60 seconds at 72C for 35 cycles (GICs) or 40 cycles (GBM tissues). immunostained as described previously (16). The following antibodies were used to detect antigens: rabbit polyclonal anti- Cycles for gapdh were 15 seconds at 94 C, 30 seconds at 53 C, and 90 seconds at 72C for 22 cycles. The following oligonucleotide DNA Eva1 (10 mg/mL) produced by immunizing a rabbit with a 0 0 synthetic peptide (CPMSGRFKDRVSWDGNPE) using a standard primers were synthesized: for eva1,asfollows:5 primer, 5 -TTCT- CCAGCTTTGCCCCTGT-30;30 primer, 50-CCGCCCATCGCTTTTTC- method (Supplementary Fig.
Load more

Recommended publications
  • The Intrinsically Disordered Proteins of Myelin in Health and Disease
    cells Review Flexible Players within the Sheaths: The Intrinsically Disordered Proteins of Myelin in Health and Disease Arne Raasakka 1 and Petri Kursula 1,2,* 1 Department of Biomedicine, University of Bergen, Jonas Lies vei 91, NO-5009 Bergen, Norway; [email protected] 2 Faculty of Biochemistry and Molecular Medicine & Biocenter Oulu, University of Oulu, Aapistie 7A, FI-90220 Oulu, Finland * Correspondence: [email protected] Received: 30 January 2020; Accepted: 16 February 2020; Published: 18 February 2020 Abstract: Myelin ensheathes selected axonal segments within the nervous system, resulting primarily in nerve impulse acceleration, as well as mechanical and trophic support for neurons. In the central and peripheral nervous systems, various proteins that contribute to the formation and stability of myelin are present, which also harbor pathophysiological roles in myelin disease. Many myelin proteins have common attributes, including small size, hydrophobic segments, multifunctionality, longevity, and regions of intrinsic disorder. With recent advances in protein biophysical characterization and bioinformatics, it has become evident that intrinsically disordered proteins (IDPs) are abundant in myelin, and their flexible nature enables multifunctionality. Here, we review known myelin IDPs, their conservation, molecular characteristics and functions, and their disease relevance, along with open questions and speculations. We place emphasis on classifying the molecular details of IDPs in myelin, and we correlate these with their various functions, including susceptibility to post-translational modifications, function in protein–protein and protein–membrane interactions, as well as their role as extended entropic chains. We discuss how myelin pathology can relate to IDPs and which molecular factors are potentially involved. Keywords: myelin; intrinsically disordered protein; multiple sclerosis; peripheral neuropathies; myelination; protein folding; protein–membrane interaction; protein–protein interaction 1.
    [Show full text]
  • Ɑ6ß1 Andɑ7ß1 Integrins Are Required in Schwann Cells to Sort
    The Journal of Neuroscience, November 13, 2013 • 33(46):17995–18007 • 17995 Cellular/Molecular ␣6␤1 and ␣7␤1 Integrins Are Required in Schwann Cells to Sort Axons Marta Pellegatta,1,2 Ade`le De Arcangelis,3 Alessandra D’Urso,1 Alessandro Nodari,1 Desire´e Zambroni,1 Monica Ghidinelli,1,2 Vittoria Matafora,1 Courtney Williamson,2 Elisabeth Georges-Labouesse,3† Jordan Kreidberg,4 Ulrike Mayer,5 Karen K. McKee,6 Peter D. Yurchenco,6 Angelo Quattrini,1 Lawrence Wrabetz,1,2 and Maria Laura Feltri1,2 1San Raffaele Scientific Institute, Milano 20132, Italy, 2Hunter James Kelly Research Institute, University at Buffalo, State University of New York, New York 14203, 3Development and Stem Cells Program, Institut de Ge´ne´tique et de Biologie Mole´culaire et Cellulaire, Centre National de la Recherche Scientifique, Unite´ Mixte de Recherche 7104, Institut National de la Sante´ et de la Recherche Me´dicale U964, Universite´ de Strasbourg, Illkirch 67404, France, 4Department of Medicine, Children’s Hospital Boston and Department of Pediatrics, Harvard Medical School, Boston, Massachusetts 02115, 5Biomedical Research Centre, School of Biological Sciences, University of East Anglia, Norwich NR4 7TJ, United Kingdom, and 6Robert Wood Johnson Medical School, Piscataway, New Jersey, New Jersey 08854 During development, Schwann cells extend lamellipodia-like processes to segregate large- and small-caliber axons during the process of radial sorting. Radial sorting is a prerequisite for myelination and is arrested in human neuropathies because of laminin deficiency. Experiments in mice using targeted mutagenesis have confirmed that laminins 211, 411, and receptors containing the ␤1 integrin subunit are required for radial sorting; however, which of the 11 ␣ integrins that can pair with ␤1 forms the functional receptor is unknown.
    [Show full text]
  • How Does Protein Zero Assemble Compact Myelin?
    Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: 13 May 2020 doi:10.20944/preprints202005.0222.v1 Peer-reviewed version available at Cells 2020, 9, 1832; doi:10.3390/cells9081832 Perspective How Does Protein Zero Assemble Compact Myelin? Arne Raasakka 1,* and Petri Kursula 1,2 1 Department of Biomedicine, University of Bergen, Jonas Lies vei 91, NO-5009 Bergen, Norway 2 Faculty of Biochemistry and Molecular Medicine & Biocenter Oulu, University of Oulu, Aapistie 7A, FI-90220 Oulu, Finland; [email protected] * Correspondence: [email protected] Abstract: Myelin protein zero (P0), a type I transmembrane protein, is the most abundant protein in peripheral nervous system (PNS) myelin – the lipid-rich, periodic structure that concentrically encloses long axonal segments. Schwann cells, the myelinating glia of the PNS, express P0 throughout their development until the formation of mature myelin. In the intramyelinic compartment, the immunoglobulin-like domain of P0 bridges apposing membranes together via homophilic adhesion, forming a dense, macroscopic ultrastructure known as the intraperiod line. The C-terminal tail of P0 adheres apposing membranes together in the narrow cytoplasmic compartment of compact myelin, much like myelin basic protein (MBP). In mouse models, the absence of P0, unlike that of MBP or P2, severely disturbs the formation of myelin. Therefore, P0 is the executive molecule of PNS myelin maturation. How and when is P0 trafficked and modified to enable myelin compaction, and how disease mutations that give rise to incurable peripheral neuropathies alter the function of P0, are currently open questions. The potential mechanisms of P0 function in myelination are discussed, providing a foundation for the understanding of mature myelin development and how it derails in peripheral neuropathies.
    [Show full text]
  • Supplementary Table 1: Adhesion Genes Data Set
    Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like,
    [Show full text]
  • Differential Gene Expression in Oligodendrocyte Progenitor Cells, Oligodendrocytes and Type II Astrocytes
    Tohoku J. Exp. Med., 2011,Differential 223, 161-176 Gene Expression in OPCs, Oligodendrocytes and Type II Astrocytes 161 Differential Gene Expression in Oligodendrocyte Progenitor Cells, Oligodendrocytes and Type II Astrocytes Jian-Guo Hu,1,2,* Yan-Xia Wang,3,* Jian-Sheng Zhou,2 Chang-Jie Chen,4 Feng-Chao Wang,1 Xing-Wu Li1 and He-Zuo Lü1,2 1Department of Clinical Laboratory Science, The First Affiliated Hospital of Bengbu Medical College, Bengbu, P.R. China 2Anhui Key Laboratory of Tissue Transplantation, Bengbu Medical College, Bengbu, P.R. China 3Department of Neurobiology, Shanghai Jiaotong University School of Medicine, Shanghai, P.R. China 4Department of Laboratory Medicine, Bengbu Medical College, Bengbu, P.R. China Oligodendrocyte precursor cells (OPCs) are bipotential progenitor cells that can differentiate into myelin-forming oligodendrocytes or functionally undetermined type II astrocytes. Transplantation of OPCs is an attractive therapy for demyelinating diseases. However, due to their bipotential differentiation potential, the majority of OPCs differentiate into astrocytes at transplanted sites. It is therefore important to understand the molecular mechanisms that regulate the transition from OPCs to oligodendrocytes or astrocytes. In this study, we isolated OPCs from the spinal cords of rat embryos (16 days old) and induced them to differentiate into oligodendrocytes or type II astrocytes in the absence or presence of 10% fetal bovine serum, respectively. RNAs were extracted from each cell population and hybridized to GeneChip with 28,700 rat genes. Using the criterion of fold change > 4 in the expression level, we identified 83 genes that were up-regulated and 89 genes that were down-regulated in oligodendrocytes, and 92 genes that were up-regulated and 86 that were down-regulated in type II astrocytes compared with OPCs.
    [Show full text]
  • Antigen Interaction with B Cells in Two Proliferative Disorders
    Linköping University Medical Dissertations No. 1158 Antigen interaction with B cells in two proliferative disorders - CLL and MGUS Eva Hellqvist Akademisk avhandling som för avläggande av medicine doktorsexamen offentligen försvaras i Aulan, Katastrofmedicinskt Centrum, Linköping, fredagen den 15 januari 2010 kl. 9.00 Fakultetsopponent Dr Anne Mette Buhl Department of Hematology, Rigshospitalet Copenhagen, Denmark Division of Cell Biology Department of Clinical and Experimental Medicine Linköping University, SE-581 85 Linköping, Sweden Linköping 2010 All text, tables and illustrations are © Eva Hellqvist, 2010 The original papers included in this thesis have been reprinted with the permission of respective copyright holder: Paper I: © the American Society of Hematology Paper IV: © the Ferrata Storti Foundation ISSN: 0345-0082 ISBN: 978-91-7393-475-6 Printed in Sweden by LiU-Tryck, Linköping 2010 “This world, after all our science and sciences, is still a miracle; wonderful, inscrutable, magical and more, to whosoever will think of it. ” – Thomas Carlyle TO MY GREAT SURPRISE SUPERVISOR Anders Rosén, Professor Division of Cell Biology Department of Clinical and Experimental Medicine Linköping University, Sweden CO-SUPERVISOR Jan Ernerudh, Professor Division of Clinical Immunology Department of Clinical and Experimental Medicine Linköping University, Sweden OPPONENT Anne Mette Buhl, Associate Professor Department of Hematology Rigshospitalet Copenhagen, Denmark COMMITTEE BOARD Hodjattalah Rabbani, Associate Professor Immune and Gene Therapy
    [Show full text]
  • (12) United States Patent (10) Patent No.: US 6,939,952 B2 Zhao (45) Date of Patent: Sep
    USOO69399.52B2 (12) United States Patent (10) Patent No.: US 6,939,952 B2 Zhao (45) Date of Patent: Sep. 6, 2005 (54) PURIFIED AND ISOLATED PROTEIN ZERO Bowie et al. Deciphering the message in protein Sequences: RELATED (PZR) POLYPEPTIDE tolerance to amino acid substitutions. Science (1990) vol. 247, pp. 1306–1310.* (75) Inventor: Zhizhuang Zhao, Franklin, TN (US) GenBank Accession if AAD55347. (73) Assignee: Vanderbilt University, Nashville, TN Burshtyn et al., “Regulation Through Inhibitory Receptors: (US) Lessons from Natural Killer Cells”, Trends in Cell Biology, (*) Notice: Subject to any disclaimer, the term of this vol. 7, p. 473–479, (Dec., 1997). patent is extended or adjusted under 35 Choe, “Packing of Myelin Protein Zero, Neuron, vol. 17, p. U.S.C. 154(b) by 69 days. 363-365, (Sep., 1996). Harding, “From the Syndrome of Charcot, Marie and Tooth (21) Appl. No.: 10/095,131 to Disorders of Peripheral Myelin Proteins,” Brain, vol. 118, (22) Filed: Mar 11, 2002 p. 809–818, (Nov., 1995). Saxton et al., “Abnormal Mesoderm Patterning in Mouse (65) Prior Publication Data Embryos Mutant for the SH2 Tyrosine Phosphatase Shp-2.” US 2003/0171565 A1 Sep. 11, 2003 The EMBO Journal, vol. 16 (No. 9), p. 2352–2364, (Nov., 1997). Related U.S. Application Data Tidow et al., “SH2-Containing Protein Tyrosine Phos (62) Division of application No. 09/430.503, filed on Oct. 29, phatases SHP-1 and SHP-2 are Dramatically Increased at 1999, now Pat. No. 6,355,786. the Protein Level in Neutrophils from Patients with Severe (60) Provisional application No.
    [Show full text]
  • Peripheral Myelin Protein 22 (Pmp22) Is in a Complex with Alpha6 Beta4 Integrin and the Absence of Pmp22 Affects Schwann Cell Adhesion and Migration
    PERIPHERAL MYELIN PROTEIN 22 (PMP22) IS IN A COMPLEX WITH ALPHA6 BETA4 INTEGRIN AND THE ABSENCE OF PMP22 AFFECTS SCHWANN CELL ADHESION AND MIGRATION By STEPHANIE ANN AMICI A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2006 Copyright 2006 by Stephanie Ann Amici I dedicate this work to my family for their constant love and support. ACKNOWLEDGMENTS Most importantly, I would like to thank my advisor, Dr. Lucia Notterpek, for the inspiration and support she has given to me. She has always had my best interests in mind and has been essential in my growth as a scientist and as a person. I also express my gratitude to my committee members, Dr. Brad Fletcher, Dr. Dena Howland, Dr. Harry Nick and Dr. Susan Frost, for their insights and suggestions for my project. Similarly, Dr. William Dunn deserves acknowledgement for his advice and assistance with the electron microscopy studies. Heartfelt thanks also go to the past and present members of the Notterpek lab for their amazing friendships over the years. Finally, my deepest gratitude goes to my family, especially my parents, for their continuous love and encouragement throughout my life. iv TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................................................................................. iv LIST OF FIGURES .......................................................................................................... vii LIST OF ABBREVIATIONS...........................................................................................
    [Show full text]
  • Mutation Update for Myelin Protein Zero-Related Neuropathies and the Increasing Role of Variants Causing a Late-Onset Phenotype
    Journal of Neurology (2019) 266:2629–2645 https://doi.org/10.1007/s00415-019-09453-3 ORIGINAL COMMUNICATION Mutation update for myelin protein zero‑related neuropathies and the increasing role of variants causing a late‑onset phenotype Ilaria Callegari1,2,7 · C. Gemelli2 · A. Geroldi2 · F. Veneri2 · P. Mandich2,3 · M. D’Antonio4 · D. Pareyson5 · M. E. Shy6 · A. Schenone2,3 · V. Prada2 · M. Grandis2,3 Received: 13 March 2019 / Revised: 21 June 2019 / Accepted: 26 June 2019 / Published online: 5 July 2019 © Springer-Verlag GmbH Germany, part of Springer Nature 2019 Abstract Mutations of myelin protein zero gene (MPZ) are found in 5% of Charcot–Marie–Tooth patients. In 2004, Shy et al. identi- fed two main phenotypes associated with them: an early-onset subtype with mainly demyelinating features and a late-onset subgroup with prominent axonal impairment. We evaluated whether novel MPZ mutations described in literature during the last 14 years could still ft with this classifcation. We collected and revised reports of 69 novel MPZ mutations. Almost 90% of them could be alternatively classifed as responsible for: (a) an early-onset phenotype, with frst limitations starting before 3 years (2.5 ± 0.50 years), motor milestones delays, frequently severe course and upper limb MNCVs below 15 m/s; (b) late-onset neuropathy, with mean age of onset of 42.8 ± 1.5 years and mean upper limbs motor nerve conduction velocities (MNCVs) of 47.2 ± 1.4 m/s; (c) a phenotype more similar to typical CMT1A neuropathy, with onset during the 2nd decade, MNCV in the range of 15–30 m/s and slowly progressive course.
    [Show full text]
  • Supplementary Table 1: Differentially Methylated Genes and Functions of the Genes Before/After Treatment with A) Doxorubicin and B) FUMI and in C) Responders Vs
    Supplementary Table 1: Differentially methylated genes and functions of the genes before/after treatment with a) doxorubicin and b) FUMI and in c) responders vs. non- responders for doxorubicin and d) FUMI Differentially methylated genes before/after treatment a. Doxo GENE FUNCTION CCL5, CCL8, CCL15, CCL21, CCR1, CD33, IL5, immunoregulatory and inflammatory processes IL8, IL24, IL26, TNFSF11 CCNA1, CCND2, CDKN2A cell cycle regulators ESR1, FGF2, FGF14, FGF18 growth factors WT1, RASSF5, RASSF6 tumor suppressor b. FUMI GENE FUNCTION CCL7, CCL15, CD28, CD33, CD40, CD69, TNFSF18 immunoregulatory and inflammatory processes CCND2, CDKN2A cell cycle regulators IGF2BP1, IGFBP3 growth factors HOXB4, HOXB6, HOXC8 regulation of cell transcription WT1, RASSF6 tumor suppressor Differentially methylated genes in responders vs. non-responders c. Doxo GENE FUNCTION CBR1, CCL4, CCL8, CCR1, CCR7, CD1A, CD1B, immunoregulatory and inflammatory processes CD1D, CD1E, CD33, CD40, IL5, IL8, IL20, IL22, TLR4 CCNA1, CCND2, CDKN2A cell cycle regulators ESR2, ERBB3, FGF11, FGF12, FGF14, FGF17 growth factors WNT4, WNT16, WNT10A implicated in oncogenesis TNFSF12, TNFSF15 apoptosis FOXL1, FOXL2, FOSL1,HOXA2, HOXA7, HOXA11, HOXA13, HOXB4, HOXB6, HOXB8, HOXB9, HOXC8, regulation of cell transcription HOXD8, HOXD9, HOXD11 GSTP1, MGMT DNA repair APC, WT1 tumor suppressor d. FUMI GENE FUNCTION CCL1, CCL3, CCL5,CCL14, CD1B, CD33, CD40, CD69, immunoregulatory and inflammatory IL20, IL32 processes CCNA1, CCND2, CDKN2A cell cycle regulators IGF2BP1, IGFBP3, IGFBP7, EGFR, ESR2,RARB2
    [Show full text]
  • Urinary Proteomics for the Early Diagnosis of Diabetic Nephropathy in Taiwanese Patients Authors
    Urinary Proteomics for the Early Diagnosis of Diabetic Nephropathy in Taiwanese Patients Authors: Wen-Ling Liao1,2, Chiz-Tzung Chang3,4, Ching-Chu Chen5,6, Wen-Jane Lee7,8, Shih-Yi Lin3,4, Hsin-Yi Liao9, Chia-Ming Wu10, Ya-Wen Chang10, Chao-Jung Chen1,9,+,*, Fuu-Jen Tsai6,10,11,+,* 1 Graduate Institute of Integrated Medicine, China Medical University, Taichung, 404, Taiwan 2 Center for Personalized Medicine, China Medical University Hospital, Taichung, 404, Taiwan 3 Division of Nephrology and Kidney Institute, Department of Internal Medicine, China Medical University Hospital, Taichung, 404, Taiwan 4 Institute of Clinical Medical Science, China Medical University College of Medicine, Taichung, 404, Taiwan 5 Division of Endocrinology and Metabolism, Department of Medicine, China Medical University Hospital, Taichung, 404, Taiwan 6 School of Chinese Medicine, China Medical University, Taichung, 404, Taiwan 7 Department of Medical Research, Taichung Veterans General Hospital, Taichung, 404, Taiwan 8 Department of Social Work, Tunghai University, Taichung, 404, Taiwan 9 Proteomics Core Laboratory, Department of Medical Research, China Medical University Hospital, Taichung, 404, Taiwan 10 Human Genetic Center, Department of Medical Research, China Medical University Hospital, China Medical University, Taichung, 404, Taiwan 11 Department of Health and Nutrition Biotechnology, Asia University, Taichung, 404, Taiwan + Fuu-Jen Tsai and Chao-Jung Chen contributed equally to this work. Correspondence: Fuu-Jen Tsai, MD, PhD and Chao-Jung Chen, PhD FJ Tsai: Genetic Center, China Medical University Hospital, No.2 Yuh-Der Road, 404 Taichung, Taiwan; Telephone: 886-4-22062121 Ext. 2041; Fax: 886-4-22033295; E-mail: [email protected] CJ Chen: Graduate Institute of Integrated Medicine, China Medical University, No.91, Hsueh-Shih Road, 404, Taichung, Taiwan; Telephone: 886-4-22053366 Ext.
    [Show full text]
  • Electrospun Hyaluronan-Gelatin Nanofibrous Matrix for Nerve Tissue Engineering
    Hindawi Publishing Corporation Journal of Nanomaterials Volume 2013, Article ID 613638, 9 pages http://dx.doi.org/10.1155/2013/613638 Research Article Electrospun Hyaluronan-Gelatin Nanofibrous Matrix for Nerve Tissue Engineering Hau-Min Liou,1 Lih-Rou Rau,1 Chun-Chiang Huang,1 Meng-Ru Lu,1 and Fu-Yin Hsu1,2 1 Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 20224, Taiwan 2 Department of Life Sciences, National Taiwan Ocean University, Keelung 20224, Taiwan Correspondence should be addressed to Fu-Yin Hsu; [email protected] Received 23 July 2013; Revised 25 September 2013; Accepted 26 September 2013 Academic Editor: In-Kyu Park Copyright © 2013 Hau-Min Liou et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Schwanncellsplayacriticalroleintherepairoftheperipheralnerve.Thegoalofthisstudywastofabricateelectrospungelatin (Gel) and hyaluronan-gelatin (HA-Gel) composite nanofibers to provide a suitable growth environment for Schwann cells. The fiber diameters of Gel, 0.5 HA-Gel, 1 HA-Gel, and 1.5 HA-Gel were 130 ± 30 nm, 294 ± 87 nm, 362 ± 129 nm, and 224 ± 54 nm, respectively. The biological performance of Gel and HA-Gel was evaluated using an in vitro culture of RT4-D6P2T rat Schwann cells. We found that the cell attachment and proliferation rates were not significantly different on these matrices. However, the Schwann cells displayed better organized F-actin on HA-Gel than on Gel. Moreover, the expression levels of several genes, including Nrg1, GFAP, and P0, were significantly higher on HA-Gel than on Gel.
    [Show full text]