title: Tumor suppressor function of ezrin-radixin-moesin-binding phosphoprotein-50 through β-catenin/E-cadherin pathway in human hepatocellular cancer.
abstract: OBJECTIVE
  To determine the effect and molecular mechanism of ezrin-radixin-moesin-binding phosphoprotein-50 (EBP50) in hepatocellular carcinoma (HCC).
  
  
  METHODS
  Three human HCC cell lines, i.e., SM-MC7721, HepG2 and Hep3B, were used. We transfected the Pbk-CMV-HA-EBP50 plasmid into SMMC7721 cells with Lipofectamine 2000 to overexpress EBP50. Western blotting were performed to determine the effects of the plasmid on EBP50 expression and to detect the expression of β-catenin and E-cadherin before and after the transfection of the plasmid into SMMC7721 cells. In vitro cell proliferation was assessed with a Cell Counting Kit-8 (CCK-8) assay. Cell cycle distribution was assessed with flow cytometry. Invasion and migration ability of before and after the transfection were determined with a transwell assay. Cell apoptosis was demonstrated with Annexin V-FITC. The effect of EBP50 overexpressing on tumor growth in vivo was performed with a xenograft tumor model in nude mice.
  
  
  RESULTS
  The transfection efficiency was confirmed with Western blotting (1.36 ± 0.07 vs 0.81 ± 0.09, P < 0.01). The CCK8 assay demonstrated that the growth of cells overexpressing EBP50 was significantly lower than control cells (P < 0.01). Cell cycle distribution showed there was a G0/G1 cell cycle arrest in cells overexpressing EBP50 (61.3% ± 3.1% vs 54.0% ± 2.4%, P < 0.05). The transwell assay showed that cell invasion and migration were significantly inhibited in cells overexpressing EBP50 compared with control cells (5.8 ± 0.8 vs 21.6 ± 1.3, P < 0.01). Annexin V-FITC revealed that apoptosis was significantly increased in cells overexpressing EBP50 compared with control cells (14.8% ± 2.7% vs 3.4% ± 1.3%, P < 0.05). The expression of β-catenin was downregulated and E-cadherin was upregulated in cells overexpressing EBP50 compared with control cells (0.28 ± 0.07 vs 0.56 ± 0.12, P < 0.05; 0.55 ± 0.08 vs 0.39 ± 0.07, P < 0.05). In vivo tumor growth assay conﬁrmed that up-regulation of EBP50 could obviously slow the growth of HCC derived from SMMC7721 cells (28.9 ± 7.2 vs 70.1 ± 7.2, P < 0.01).
  
  
  CONCLUSIONS
  The overexpression of EBP50 could inhibit the growth of SMMC7721 cells and promote apoptosis by modulating β-catenin, E-cadherin. EBP50 may serve as a potential therapeutic target in HCC.
authors.affiliation: Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: Xiu-Lan
authors.initials: XL
authors.lastname: Peng
authors.forename: Meng-Yao
authors.initials: MY
authors.lastname: Ji
authors.forename: Zi-Rong
authors.initials: ZR
authors.lastname: Yang
authors.forename: Jia
authors.initials: J
authors.lastname: Song
authors.forename: Wei-Guo
authors.initials: WG
authors.lastname: Dong
mesh_terms: D000818:Animals; D015703:Antigens, CD; D017209:Apoptosis; D015820:Cadherins / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D009361:Neoplasm Invasiveness; D009368:Neoplasm Transplantation; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D047368:Tumor Burden; D051176:beta Catenin / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; C513384:CDH1 protein, human; C495270:CTNNB1 protein, human; D015820:Cadherins; D010750:Phosphoproteins; D017923:Sodium-Hydrogen Exchangers; D051176:beta Catenin; C093376:sodium-hydrogen exchanger regulatory factor
keywords: Ezrin-radixin-moesin-binding phosphoprotein-50; Growth; Hepatocellular carcinoma; Invasion; Migration