DOCSLIB.ORG

HYOU1 Promotes Cell Growth and Metastasis Via Activating PI3K/AKT Signaling in Epithelial Ovarian Cancer and Predicts Poor Prognosis

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
HYOU1 Promotes Cell Growth and Metastasis Via Activating PI3K/AKT Signaling in Epithelial Ovarian Cancer and Predicts Poor Prognosis European Review for Medical and Pharmacological Sciences 2019; 23: 4126-4135 HYOU1 promotes cell growth and metastasis via activating PI3K/AKT signaling in epithelial ovarian cancer and predicts poor prognosis X. LI1, N.-X. ZHANG2, H.-Y. YE3, P.-P. SONG3, W. CHANG3, L. CHEN1, Z. WANG1, L. ZHANG1, N.-N. WANG4 1Department of Gynecology, 2Department of Hospital Acquired Infection Control, 3Department of Endocrinology, 4Department of Cadre Health Care; Qingdao Hiser Medical Group, Qingdao, Shandong, China. Abstract. – OBJECTIVE: Hypoxia upregulat- Introduction ed 1 (HYOU1) has been reported to be abnor- mally expressed in different malignancies, es- Ovarian cancer is a leading cause of death pecially in breast cancer. However, the role of among gynecological cancers worldwide, and the HYOU1 in epithelial ovarian cancer (EOC) re- mains largely unclear. This study aimed to ex- fatality rate of ovarian cancer is the highest of plore the expression and function of HYOU1 in all types of gynecological tumors, causing about EOC progression. 140,200 deaths in the world annually1,2. The epi- PATIENTS AND METHODS: HYOU1 levels thelial ovarian cancer (EOC) is a common entity in EOC tissues and cell lines were investigat- accounting for 80 to 90% of ovarian cancer cases3. ed by RT-PCR. The clinical and prognostic The current standard of therapy is the debulking significance of HYOU1 in 127 cases of EOC 4 was analyzed using the Chi-square analysis, surgery combined with chemotherapy . In spite Kaplan-Meier analysis, and the Cox propor- of continuous efforts to improve the therapeutic tional hazards regression model. We have al- response, the prognosis of EOC patients remains so performed multiple cells experiments to poor, with a 5-year survival rate of only 30%5,6. In evaluate the effects of HYOU1 on EOC cell addition, although growing researches have been proliferation, apoptosis, migration, and inva- published in recent years and have contributed sion. The protein levels of associated PI3K/ Akt signaling pathway was detected using to the understanding of the basic and clinical Western blot assay. knowledge, the complex molecular mechanism of RESULTS: We found that the expression lev- EOC is still insufficiently understood7. Thus, it is els of HYOU1 were significantly upregulated necessary to further study the molecular mech- in both EOC tissues and cell lines. A higher anisms involved in EOC carcinogenesis and to expression of HYOU1 was associated with ad- identify the prognostic markers for the design of vanced FIGO stage, LN metastasis, and short- er overall survival. In addition, univariate and the individual therapy, and the targeted treatment multivariate analysis identified high HYOU1 ex- of EOC. Hypoxia upregulated 1 (HYOU1), also pression as an unfavorable prognostic factor known as Grp170 and HSP12A, is encoded by for overall survival. Functional assays revealed this HYOU1 gene and belongs to the heat shock that the inhibition of HYOU1 suppressed the protein 70 family8,9. Up to date, the biological tumor proliferation and colony formation, as function of HYOU1 in cell progress remains well as the migratory and invasive capacity. Fi- nally, when HYOU1 was silenced, the results of largely unclear. Previously, evidence has shown Western blot showed that the levels of p-PI3K, that HYOU1 may be involved in the regulation p-Akt, as well as cell cycle and EMT genes, were of apoptosis and its inhibition is correlated with respectively downregulated. accelerated apoptosis10,11. In addition, there are CONCLUSIONS: Our findings highlighted the cellular experiments which indicate that this gene targeting of HYOU1 as a novel therapeutic ap- has an important cytoprotective role in hypox- proach for the treatment of EOC. ia-induced cellular perturbation. Recently, with Key Words: the development of sequencing technology, the HYOU1, Epithelial ovarian cancer, PI3K/AKT signal- dysregulation of HYOU1 was reported in several ing pathway, Prognosis, Tumorigenesis. tumors, which suggested that HYOU1 may play a 4126 Corresponding Author: Na-Na Wang, MD; e-mail: [email protected] HYOU1 expression in epithelial ovarian cancer functional effect in the development and progres- normal tissue samples were immediately frozen sion of tumors12,13. Zhou et al14 firstly reported that using liquid nitrogen and subsequently stored at HYOU1 was highly expressed in nasopharyngeal -80°C for further use. No patients received any carcinoma and associated with poor prognosis chemotherapy, immunotherapy or radiotherapy of the patient with nasopharyngeal carcinoma, before the surgery. The informed consents were suggesting that HYOU1 may serve as a tumor obtained from all patients. promoter in this disease, although in vitro and in vivo experiments were not been performed. How- Cell Lines and Cell Transfection ever, whether HYOU1 was abnormally expressed We obtained IOSE80 (ovarian epithelial cells) in EOC, as well as its potential biological func- and three EOC cells, HEY, SKOV3, OVCAR-3, tion in the progression of EOC, remains largely from Uban Biotechnology Co., Ltd. (Jiading, unclear. In this study, we analyzed microarray Shanghai, China). The cells were maintained in data from GEO datasets, finding that HYOU1 RPMI 1640 medium (BioSUN, Xuhui, Shang- expression was significantly up-regulated in EOC hai, China) supplemented with fetal bovine serum tissues, which was consistent with the expression (FBS; 10% concentration) and penicillin-strepto- trend of HYOU1 in breast cancer9 and naso- mycin antibiotics solution (1%; HY Stem Cells, pharyngeal carcinoma14. Furthermore, RT-PCR Shapingba, Chongqing, China). For cell trans- assays of clinical samples from our hospital also fection, a LipoFiter 3 transfection reagent (Han- confirmed that HYOU1 was overexpressed in Bio, Jiading, Shanghai, China) was employed to EOC tissues. Given the possibility of HYOU1 as transfect small interfering RNAs (siRNAs) into a potential positive regulator in EOC, we further SKOV3 and OVCAR-3 cells. The siRNAs target- analyzed its prognostic value in EOC patients ing HYOU1 (si-HYOU1-1 and si-HYOU1-2) and and performed a lost-function assay to explore negative control siRNAs (si-NC) were purchased its specific biological function in the progression from ZoonBio Co., Ltd. (Nanjing, Jiangsu, China). of EOC. Finally, we also tried to explore the potential mechanism of HYOU1 involved in the RNA Isolation and Quantitative proliferation and metastasis. Real Time-PCR The total RNA from EOC tissues or cells was extracted using TRIzol reagent, and the quantity Patients and Methods of the RNA was determined by NanoDrop ND- 1000 spectrophotometer (Thermo Fisher Scien- Clinical Specimens tific, Waltham, MA, USA). Then, the cDNA was A total of 127 EOC patients (relevant clini- reverse-transcribed using Fermentas RevertAid cal features were summarized in Table I) were first strand cDNA synthesis kit (Think-far, Haid- enrolled in this research, which was approved ian, Beijing, China). Afterward, qRT-PCR as- by the Ethics Committee of Qingdao Hiser Med- says for HYOU1 determination were conducted ical Group from April 2010 to March 2013. using SYBR Premix Ex Taq real time-PCR kit The EOC specimens and adjacent non-cancerous (TaKaRa, Dalian, Liaoning, China). Glyceralde- Table I. Correlation between HYOU1 expression with clinicopathologic features of EOC. Parameters Category No. HYOU1 expression p-value High Low Age (y) ≤50 63 30 33 0.657 >50 64 33 31 FIGO stage I-II 80 34 46 0.037 III-IV 47 29 18 Differentiation 1/2 72 32 40 0.183 3 55 31 24 Tumor size ≤5 cm 89 45 44 0.384 >5 cm 48 28 20 LN Metastasis No 87 37 50 0.019 Yes 40 26 14 4127 X. Li, N.-X. Zhang, H.-Y. Ye, P.-P. Song, W. Chang, L. Chen, Z. Wang, L. Zhang, N.-N. Wang Table II. Microorganism isolated from patients with PJI. EdU Assays Genes Primer sequences (5’-3’) EdU (5-Ethynyl-2’-deoxyuridine; SolarBio, Tongzhou, Beijing, China) assays were applied HYOU1: Forward GAGGAGGCGAGTCTGTTGG to examine the proliferation of SKOV3 and OV- HYOU1: Reverse GCACTCCAGGTTTGACAATGG CAR-3 cells. In short, after transfection with GAPDH: Forward TGGCCTTCCGTGTTCCTAC HYOU1 siRNAs or negative control siRNAs, GAPDH: Reverse GAGTTGCTGTTGAAGTCGCA the SKOV3, and OVCAR-3 cells were harvested and re-plated in 48 well plates. Forty-eight hours later, the EdU reagent (100 μl per well) was added hyde-3-phosphate dehydrogenase (GAPDH) was into the cells, and the plates were continued to be used as the reference gene. Relative HYOU1 ex- cultured for 2 h. Then, the Phosphate-buffered pression was calculated using the 2−DDCt method. saline (PBS) was used to wash the cells, and the The primers for HYOU1 are listed in Table II. DAPI reagent was added into the cells. After rinsing with PBS twice, a NIB100F fluorescence Western Blot Assays microscope (Micro-Science, Jinan, Shandong, HYOU1 siRNAs-transfected SKOV3 and OV- China) was applied to observe the cellular flu- CAR-3 cells were lysed using RIPA lysis buffer orescence. (SeeBio, Pudong, Shanghai, China), and a BCA assay kit (HuaMaiKe, Changping, Beijing, China) Cell Colony Formation Assays was applied to measure the concentration of pro- For colony formation assays, the SKOV3 and teins. Subsequently, 20 μg proteins (per lane) were OVCAR-3 cells were first transfected with HY- separated by 8-10% sodium salt-polyacrylamide OU1 siRNAs or negative control siRNAs. Then, gel electrophoresis (SDS-PAGE) using a Bio-Rad the treated cells were harvested and re-placed into Western blot apparatus and then, transferred to 6-well plates (500 cells per well). The transfections Millipore polyvinylidene difluoride (PVDF) mem- were re-conducted every 3 days. Two weeks later, branes. Next, non-fat milk (5%) was employed to the cell colonies were stained using 0.2% crystal block the membranes, and sequentially the mem- violet solution (in 20% methanol) and subsequent- branes were probed by primary antibodies target- ly photographed by a NIB100F microscope (Mi- ing GAPDH, vimentin, N-cadherin, phosphorylat- cro-Science, Jinan, Shandong, China). ed-PI3K (p-PI3K), phosphorylated-AKT (p-AKT), PI3K, and AKT for 12 h at 4°C.
Load more

Recommended publications
  • Human ORP150 / HYOU1 / HSP12A Protein (His Tag)
    Human ORP150 / HYOU1 / HSP12A Protein (His Tag) Catalog Number: 11342-H08H General Information SDS-PAGE: Gene Name Synonym: GRP-170; Grp170; HSP12A; ORP-150; ORP150 Protein Construction: A DNA sequence encoding the C-terminal segment of human HSP12A (NP_001124463.1) (Met 695-Leu 994) was expressed, fused with a polyhistidine tag at the C-terminus and a signal peptide at the N-terminus. Source: Human Expression Host: HEK293 Cells QC Testing Purity: > 97 % as determined by SDS-PAGE Endotoxin: Protein Description < 1.0 EU per μg of the protein as determined by the LAL method Hypoxia up-regulated protein 1, also known as 150 kDa oxygen-regulated protein, 170 kDa glucose-regulated protein, ORP-150, GRP-170 and Stability: HYOU1, is a member of theheat shock protein 70 family. Seven members from four different heat shock protein (HSP) families were identified Samples are stable for up to twelve months from date of receipt at -70 ℃ including HYOU1 (ORP150), HSPC1 (HSP86), HSPA5 (Bip), HSPD1 (HSP60), and several isoforms of the two testis-specific HSP70 Predicted N terminal: Met 695 chaperones HSPA2 and HSPA1L. HYOU1 is highly expressed in tissues Molecular Mass: that contain well-developed endoplasmic reticulum and synthesize large amounts of secretory proteins. It is highly expressed in liver and pancreas. The recombinant human HSP12A consists of 311 amino acids and HYOU1 is also expressed in macrophages within aortic atherosclerotic predictes a molecular mass of 35.2 kDa. In SDS-PAGE under reducing plaques, and in breast cancers. HYOU1 has a pivotal role in cytoprotective conditions, the apparent molecular mass of rh HSP12A is approximately cellular mechanisms triggered by oxygen deprivation.
    [Show full text]
  • A Master Autoantigen-Ome Links Alternative Splicing, Female Predilection, and COVID-19 to Autoimmune Diseases
    bioRxiv preprint doi: https://doi.org/10.1101/2021.07.30.454526; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. A Master Autoantigen-ome Links Alternative Splicing, Female Predilection, and COVID-19 to Autoimmune Diseases Julia Y. Wang1*, Michael W. Roehrl1, Victor B. Roehrl1, and Michael H. Roehrl2* 1 Curandis, New York, USA 2 Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, USA * Correspondence: [email protected] or [email protected] 1 bioRxiv preprint doi: https://doi.org/10.1101/2021.07.30.454526; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. Abstract Chronic and debilitating autoimmune sequelae pose a grave concern for the post-COVID-19 pandemic era. Based on our discovery that the glycosaminoglycan dermatan sulfate (DS) displays peculiar affinity to apoptotic cells and autoantigens (autoAgs) and that DS-autoAg complexes cooperatively stimulate autoreactive B1 cell responses, we compiled a database of 751 candidate autoAgs from six human cell types. At least 657 of these have been found to be affected by SARS-CoV-2 infection based on currently available multi-omic COVID data, and at least 400 are confirmed targets of autoantibodies in a wide array of autoimmune diseases and cancer.
    [Show full text]
  • Expression of HYOU1 Via Reciprocal Crosstalk Between NSCLC Cells and Huvecs Control Cancer Progression and Chemoresistance in Multicellular Tumor Spheroids
    Expression of HYOU1 via reciprocal crosstalk between NSCLC cells and HUVECs control cancer progression and chemoresistance in multicellular tumor spheroids Minji Lee Institut Pasteur de Coree Yeonhwa Song Institut Pasteur de Coree Inhee Choi Institut Pasteur de Coree Su-Yeon Lee Institut Pasteur de Coree Sanghwa Kim Institut Pasteur de Coree Se-Hyuk Kim Institut Pasteur de Coree Jiho Kim Institut Pasteur de Coree Haengran Seo ( [email protected] ) Institut Pasteur de Coree Research Keywords: Hypoxia up-regulated protein1 (HYOU1), Lung cancer, Multicellular Tumor Spheroids (MCTS), Endothelial cells (ECs), chemoresistance Posted Date: August 20th, 2020 DOI: https://doi.org/10.21203/rs.3.rs-61542/v1 License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License Page 1/26 Abstract Background Among all cancer types, lung cancer ranks highest worldwide in terms of both incidence and mortality. The crosstalk between lung cancer cells and their tumor microenvironment (TME) has begun to emerge as the “Achilles heel” of the disease and thus constitutes an attractive target for anticancer therapy. We previously revealed that crosstalk between lung cancer cells and endothelial cells (ECs) induces chemoresistance in multicellular tumor spheroids (MCTSs). Methods In this study, we idientied that factors secreted in response to crosstalk between ECs and lung cancer cells play pivotal roles in the development of chemoresistance in lung cancer spheroids. Results We determined that the expression of hypoxia up-regulated protein 1 (HYOU1) in lung cancer spheroids was increased by factors secreted in response to crosstalk between ECs and lung cancer cells. Direct interaction between lung cancer cells and ECs also caused an elevation in the expression of HYOU1 in MCTSs.
    [Show full text]
  • Anti-HYOU1 / ORP150 Antibody (ARG42870)
    Product datasheet [email protected] ARG42870 Package: 100 μl anti-HYOU1 / ORP150 antibody Store at: -20°C Summary Product Description Rabbit Polyclonal antibody recognizes HYOU1 / ORP150 Tested Reactivity Hu, Ms, Rat Tested Application FACS, IHC-P, WB Host Rabbit Clonality Polyclonal Isotype IgG Target Name HYOU1 / ORP150 Antigen Species Human Immunogen Synthetic peptide derived from Human HYOU1 / ORP150. Conjugation Un-conjugated Alternate Names GRP-170; Hypoxia up-regulated protein 1; Grp170; ORP150; 150 kDa oxygen-regulated protein; 170 kDa glucose-regulated protein; HSP12A; ORP-150 Application Instructions Application table Application Dilution FACS 1:50 IHC-P 1:50 - 1:200 WB 1:500 - 1:2000 Application Note * The dilutions indicate recommended starting dilutions and the optimal dilutions or concentrations should be determined by the scientist. Positive Control MCF7 Calculated Mw 111 kDa Observed Size ~ 140 kDa Properties Form Liquid Purification Affinity purified. Buffer PBS (pH 7.4), 150 mM NaCl, 0.02% Sodium azide and 50% Glycerol. Preservative 0.02% Sodium azide Stabilizer 50% Glycerol Storage instruction For continuous use, store undiluted antibody at 2-8°C for up to a week. For long-term storage, aliquot and store at -20°C. Storage in frost free freezers is not recommended. Avoid repeated freeze/thaw www.arigobio.com 1/2 cycles. Suggest spin the vial prior to opening. The antibody solution should be gently mixed before use. Note For laboratory research only, not for drug, diagnostic or other use. Bioinformation Gene Symbol HYOU1 Gene Full Name hypoxia up-regulated 1 Background The protein encoded by this gene belongs to the heat shock protein 70 family.
    [Show full text]
  • Sequence and Expression Analysis of HSP70 Family Genes in Artemia
    www.nature.com/scientificreports OPEN Sequence and expression analysis of HSP70 family genes in Artemia franciscana Received: 5 March 2019 Wisarut Junprung1,2, Parisa Norouzitallab1,3, Stephanie De Vos 1, Anchalee Tassanakajon2, Accepted: 24 May 2019 Dung Nguyen Viet1,4, Gilbert Van Stappen1 & Peter Bossier1 Published: xx xx xxxx Thus far, only one gene from the heat shock protein 70 (HSP70) family has been identifed in Artemia franciscana. Here, we used the draft Artemia transcriptome database to search for other genes in the HSP70 family. Four novel HSP70 genes were identifed and designated heat shock cognate 70 (HSC70), heat shock 70 kDa cognate 5 (HSC70-5), Immunoglobulin heavy-chain binding protein (BIP), and hypoxia up-regulated protein 1 (HYOU1). For each of these genes, we obtained nucleotide and deduced amino acid sequences, and reconstructed a phylogenetic tree. Expression analysis revealed that in the juvenile state, the transcription of HSP70 and HSC70 was signifcantly (P < 0.05) higher in a population of A. franciscana selectively bred for increased induced thermotolerance (TF12) relative to a control population (CF12). Following non-lethal heat shock treatment at the nauplius stage, transcription of HSP70, HSC70, and HSC70-5 were signifcantly (P < 0.05) up-regulated in TF12. In contrast, transcription of the other HSP70 family members in A. franciscana (BIP, HYOU1, and HSPA4) showed no signifcant (P > 0.05) induction. Gene expression analysis demonstrated that not all members of the HSP70 family are involved in the response to heat stress and selection and that especially altered expression of HSC70 plays a role in a population selected for increased thermotolerance.
    [Show full text]
  • HYOU1 Purified Maxpab Mouse Polyclonal Antibody (B01P)
    HYOU1 purified MaxPab mouse avoid repeated freezing and thawing. polyclonal antibody (B01P) Entrez GeneID: 10525 Catalog Number: H00010525-B01P Gene Symbol: HYOU1 Regulatory Status: For research use only (RUO) Gene Alias: DKFZp686N08236, FLJ94899, FLJ97572, Grp170, HSP12A, ORP150 Product Description: Mouse polyclonal antibody raised against a full-length human HYOU1 protein. Gene Summary: The protein encoded by this gene belongs to the heat shock protein 70 family. This gene Immunogen: HYOU1 (AAH72436.1, 1 a.a. ~ 678 a.a) uses alternative transcription start sites. A cis-acting full-length human protein. segment found in the 5' UTR is involved in Sequence: stress-dependent induction, resulting in the MADKVRRQRPRRRVCWALVAVLLADLLALSDTLAVMS accumulation of this protein in the endoplasmic reticulum VDLGSESMKVAIVKPGVPMEIVLNKESRRKTPVIVTLKE (ER) under hypoxic conditions. The protein encoded by NERFFGDSAASMAIKNPKATLRYFQHLLGKQADNPHV this gene is thought to play an important role in protein ALYQARFPEHELTFDPQRQTVHFQISSQLQFSPEEVL folding and secretion in the ER. Since suppression of the GMVLNYSRSLAEDFAEQPIKDAVITVPVFFNQAERRAV protein is associated with accelerated apoptosis, it is LQAARMAGLKVLQLINDNTATALSYGVFRRKDINTTAQ also suggested to have an important cytoprotective role NIMFYDMGSGSTVCTIVTYQMVKTKEAGMQPQLQIRG in hypoxia-induced cellular perturbation. This protein has VGFDRTLGGLEMELRLRERLAGLFNEQRKGQRAKDV been shown to be up-regulated in tumors, especially in RENPRAMAKLLREANRLKTVLSANADHMAQIEGLMDD breast tumors, and thus it is associated with tumor
    [Show full text]
  • Admixture Facilitates Genetic Adaptations to High Altitude in Tibet
    ARTICLE Received 21 Oct 2013 | Accepted 17 Jan 2014 | Published 10 Feb 2014 DOI: 10.1038/ncomms4281 Admixture facilitates genetic adaptations to high altitude in Tibet Choongwon Jeong1, Gorka Alkorta-Aranburu1, Buddha Basnyat2, Maniraj Neupane3, David B. Witonsky1, Jonathan K. Pritchard1,4,w, Cynthia M. Beall5 & Anna Di Rienzo1 Admixture is recognized as a widespread feature of human populations, renewing interest in the possibility that genetic exchange can facilitate adaptations to new environments. Studies of Tibetans revealed candidates for high-altitude adaptations in the EGLN1 and EPAS1 genes, associated with lower haemoglobin concentration. However, the history of these variants or that of Tibetans remains poorly understood. Here we analyse genotype data for the Nepalese Sherpa, and find that Tibetans are a mixture of ancestral populations related to the Sherpa and Han Chinese. EGLN1 and EPAS1 genes show a striking enrichment of high-altitude ancestry in the Tibetan genome, indicating that migrants from low altitude acquired adaptive alleles from the highlanders. Accordingly, the Sherpa and Tibetans share adaptive hae- moglobin traits. This admixture-mediated adaptation shares important features with adaptive introgression. Therefore, we identify a novel mechanism, beyond selection on new mutations or on standing variation, through which populations can adapt to local environments. 1 Department of Human Genetics, University of Chicago, Chicago, Illinois 60637, USA. 2 Oxford University Clinical Research Unit, Patan Hospital, Lal Durbar marg, GPO Box 3596, Kathmandu, Nepal. 3 Mountain Medicine Society of Nepal, Maharajgunj, Kathmandu, Nepal. 4 Howard Hughes Medical Institute, Chevy Chase, Maryland 20815, USA. 5 Department of Anthropology, Case Western Reserve University, Cleveland, Ohio 44106-7125, USA.
    [Show full text]
  • Supplemental Information
    Flach et al. Immunity, Volume 33 Supplemental Information Mzb1 Protein Regulates Calcium Homeostasis, Antibody Secretion, and Integrin Activation in Innate-like B Cells Henrik Flach, Marc Rosenbaum, Marlena Duchniewicz, Sola Kim, Shenyuan L. Zhang, Michael D. Cahalan, Gerhard Mittler, and Rudolf Grosschedl Inventory: Supplemental Figures: Figure S1 – related to Figure 1 Figure S2 – related to Figure 2 Figure S3 – related to Figure 3 Figure S4 – related to Figure 4 Figure S5 – related to Figure 5 Figure S6 – related to Figure 6 Figure S7 – related to Figure 7 Table S1 – related to Figure S1 Table S2 – related to Figure 3 Supplemental Figure Legends Legends for Tables S1 and S2 Supplemental Experimental Procedures Supplemental References Flach et al. Flach et al. Flach et al. Flach et al. Flach et al. Flach et al. Flach et al. Flach et al. Flach et al. beads -EBNA -MZB1 Gelband 1 Protein not detected Hyou1 Hyou1 Uniprot Q9JKR6 Q9JKR6 MW (kDa) 111.2 111.2 EmPAI 0.52 9.4 Unique peptides 24 64 Sequence cover. (%) 27 55 Gelband 2 Protein not detected Grp94 Grp94 Uniprot P08113 P08113 MW (kDa) 92.5 92.5 EmPAI 0.07 1.46 Unique peptides 3 25 Sequence cover. (%) 3 34 Protein not detected BiP BiP Uniprot P20029 P20029 MW (kDa) 72.4 72.4 EmPAI 0.27 2.97 Unique peptides 8 31 Sequence cover. (%) 16 49 Protein not detected not detected Calreticulin Uniprot P14211 MW (kDa) 48.0 EmPAI 0.62 Unique peptides 10 Sequence cover. (%) 25 Protein not detected not detected Calnexin Uniprot P35564 MW (kDa) 67.3 EmPAI 0.28 Unique peptides 7 Sequence cover.
    [Show full text]
  • Detection of Serum Mirnas Affecting Liver Apoptosis in a Periodontitis
    in vivo 34: 117-123 (2020) doi:10.21873/invivo.11752 Detection of Serum miRNAs Affecting Liver Apoptosis in a Periodontitis Rat Model 2 1 1 1 1 YOSHIO SUGIURA , TOSHIKI YONEDA , KOHEI FUJIMORI , TAKAYUKI MARUYAMA , HISATAKA MIYAI , TERUMASA KOBAYASHI1, DAISUKE EKUNI1, TAKAAKI TOMOFUJI3 and MANABU MORITA1 1Department of Preventive Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan; 2Center for Innovative Clinical Medicine, Okayama University Hospital, Okayama, Japan; 3Department of Community Oral Health, Asahi University School of Dentistry, Gifu, Japan Abstract. Background/Aim: miRNA molecules have been have suggested that periodontitis affects systemic diseases, attracting attention as genetic modifiers between organs. We such as diabetes mellitus (1), cardiovascular (2), liver (3), and examined the relationship between serum miRNA and kidney disease (4). Other studies have shown that periodontitis targeted liver mRNA profiles in a periodontitis rat model, and may cause liver lesions (5-8), while recently it was also the influence of periodontitis on the liver. Materials and reported to cause slight cell degeneration, inflammatory foci Methods: Male Wistar rats (n=16, 8 weeks old) were (8) and hepatocyte apoptosis in a rat periodontitis model (9). randomly divided into two groups (8 rats each): control and Despite all these reports, it still remains unclear what mediates periodontitis (ligature placement for 4 weeks). Serum miRNA the communication between periodontitis and liver pathology. and liver mRNA profiles were compared. Results: Periodontal MicroRNAs (miRNAs) are endogenous noncoding destruction and hepatocyte apoptosis were induced in the regulatory RNAs comprising 17-25 nucleotides, which have periodontitis group. Microarray analysis indicated that 52 critical functions in post-transcriptional gene regulation (10).
    [Show full text]
  • Evidence for Activation of the Unfolded Protein Response in Collagen IV Nephropathies
    BASIC RESEARCH www.jasn.org Evidence for Activation of the Unfolded Protein Response in Collagen IV Nephropathies Myrtani Pieri,* Charalambos Stefanou,* Apostolos Zaravinos,* Kamil Erguler,* † ‡ ‡ Kostas Stylianou, George Lapathitis, Christos Karaiskos, Isavella Savva,* | Revekka Paraskeva,* Harsh Dweep,§ Carsten Sticht,§ Natassa Anastasiadou, | †† Ioanna Zouvani, Demetris Goumenos,¶ Kyriakos Felekkis,** Moin Saleem, Konstantinos Voskarides,* Norbert Gretz,§ and Constantinos Deltas* *Molecular Medicine Research Center, Department of Biological Sciences, University of Cyprus, Nicosia, Cyprus; †Department of Nephrology, Heraklion University Hospital, Crete, Greece; ‡The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus; §Medical Research Center, University of Heidelberg, Mannheim, Germany; |Department of Histopathology, Nicosia General Hospital, Nicosia, Cyprus; ¶Departments of Internal Medicine-Nephrology, University Hospital of Patras, Patras, Greece; **Department of Life and Health Sciences, University of Nicosia, Nicosia, Cyprus; and ††Children’s and Academic Renal Unit, Southmead Hospital-University of Bristol, Bristol, United Kingdom ABSTRACT Thin-basement-membrane nephropathy (TBMN) and Alport syndrome (AS) are progressive collagen IV nephropathies caused by mutations in COL4A3/A4/A5 genes. These nephropathies invariably present with microscopic hematuria and frequently progress to proteinuria and CKD or ESRD during long-term follow-up. Nonetheless, the exact molecular mechanisms by which these mutations exert their deleterious
    [Show full text]
  • Peripheral Nerve Single-Cell Analysis Identifies Mesenchymal Ligands That Promote Axonal Growth
    Research Article: New Research Development Peripheral Nerve Single-Cell Analysis Identifies Mesenchymal Ligands that Promote Axonal Growth Jeremy S. Toma,1 Konstantina Karamboulas,1,ª Matthew J. Carr,1,2,ª Adelaida Kolaj,1,3 Scott A. Yuzwa,1 Neemat Mahmud,1,3 Mekayla A. Storer,1 David R. Kaplan,1,2,4 and Freda D. Miller1,2,3,4 https://doi.org/10.1523/ENEURO.0066-20.2020 1Program in Neurosciences and Mental Health, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada, 2Institute of Medical Sciences University of Toronto, Toronto, Ontario M5G 1A8, Canada, 3Department of Physiology, University of Toronto, Toronto, Ontario M5G 1A8, Canada, and 4Department of Molecular Genetics, University of Toronto, Toronto, Ontario M5G 1A8, Canada Abstract Peripheral nerves provide a supportive growth environment for developing and regenerating axons and are es- sential for maintenance and repair of many non-neural tissues. This capacity has largely been ascribed to paracrine factors secreted by nerve-resident Schwann cells. Here, we used single-cell transcriptional profiling to identify ligands made by different injured rodent nerve cell types and have combined this with cell-surface mass spectrometry to computationally model potential paracrine interactions with peripheral neurons. These analyses show that peripheral nerves make many ligands predicted to act on peripheral and CNS neurons, in- cluding known and previously uncharacterized ligands. While Schwann cells are an important ligand source within injured nerves, more than half of the predicted ligands are made by nerve-resident mesenchymal cells, including the endoneurial cells most closely associated with peripheral axons. At least three of these mesen- chymal ligands, ANGPT1, CCL11, and VEGFC, promote growth when locally applied on sympathetic axons.
    [Show full text]
  • The Hsp70 Gene Family in Boleophthalmus Pectinirostris: Genome-Wide Identification and Expression Analysis Under High Ammonia Stress
    animals Article The Hsp70 Gene Family in Boleophthalmus pectinirostris: Genome-Wide Identification and Expression Analysis under High Ammonia Stress Zhaochao Deng †, Shanxiao Sun †, Tianxiang Gao and Zhiqiang Han * Fishery College, Zhejiang Ocean University, Zhoushan 316002, Zhejiang, China; [email protected] (Z.D.); [email protected] (S.S.); [email protected] (T.G.) * Correspondence: [email protected]; Tel.: +86-580-2089333 † These authors contributed equally to this work. Received: 12 December 2018; Accepted: 23 January 2019; Published: 26 January 2019 Simple Summary: Heat shock proteins 70 is a family of proteins, which were expressed in response to a wide range of biotic and abiotic stressors. The development of genomic resources and transcriptome sequences makes it practical to conduct a systematic analysis of these genes. In this study, exhaustive searches of all genomic resources for Boleophthalmus pectinirostris Hsp70 genes were performed and their responses to high environmental ammonia stress were investigated. Besides, selection test was implemented on those duplicated genes, and the phylogenetic tree, gene structure, and motif analysis were also constructed to assign names of them. The result showed that there were 20 Hsp70 genes within the genome of Boleophthalmus pectinirostris, and some sites in the duplicated genes may experience positive selection, and most of Hsp70 genes were downregulated after exposure to high concentration ammonia. The present results of this study can be used as a reference for further biological studies on mudskippers. Abstract: Heat shock proteins 70 have triggered a remarkable large body of research in various fishes; however, no genome-wide identification and expression analysis has been performed on the Hsp70 gene family of Boleophthalmus pectinirostris.
    [Show full text]