title: MicroRNA-26a inhibits angiogenesis by down-regulating VEGFA through the PIK3C2α/Akt/HIF-1α pathway in hepatocellular carcinoma.
abstract: OBJECTIVE
  microRNAs (miRNAs) have been reported to regulate angiogenesis by down-regulating the expression of pro-angiogenic or anti-angiogenic factors. The aims of this study were to investigate whether miR-26a inhibited angiogenesis by down-regulating vascular endothelial growth factor A (VEGFA) and its clinical relevance in hepatocellular carcinoma (HCC).
  
  
  METHODS
  The expression of miR-26a was modified in HepG2 and HCCLM3 cell lines respectively, and a panel of angiogenic factors was measured by real-time PCR in the cells. A luciferase reporter assay was used to validate the target gene of miR-26a. Specific inhibitors of signal transduction pathway and siRNA approaches were used to explore the regulatory mechanism of miR-26a. Migration and tube forming assays were conducted to show the changes of angiogenesis induced by miR-26a and its target genes. Finally animal studies were used to further validate those findings.
  
  
  RESULTS
  Ectopic expression of miR-26a exhibited decreased levels of VEGFA in HepG2 cells. Migration and tube forming of human umbilical vein endothelial cells (HUVECs) were decreased in the conditioned medium from ectopic expression of miR-26a in HepG2 cells compared to control HepG2 cells. The pro-angiogenic effects of the conditioned medium of HepG2 cells on HUVECs were specifically decreased by LY294002, YC-1, and bevacizumab. Integrated analysis disclosed PIK3C2α as a downstream target gene of miR-26a. Ectopic expression of miR-26a suppressed ectopic and orthotopic tumor growth and vascularity in nude mice. The results in HCCLM3 were consistent with those in HepG2. miR-26a expression was inversely correlated with VEGFA expression in HCC patients.
  
  
  CONCLUSIONS
  miR-26a modulated angiogenesis of HCC through the PIK3C2α/Akt/HIF-1α/VEGFA pathway. The expression of VEGFA was inversely correlated with miR-26a expression in HCC tumors.
authors.affiliation: Liver Cancer Institute, Zhongshan Hospital, Fudan University; Key Laboratory of Carcinogenesis and Cancer Invasion (Fudan University), Ministry of Education, Shanghai, P.R. China.
authors.forename: Zong-Tao
authors.initials: ZT
authors.lastname: Chai
authors.forename: Jian
authors.initials: J
authors.lastname: Kong
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Zhu
authors.forename: Yuan-Yuan
authors.initials: YY
authors.lastname: Zhang
authors.forename: Lu
authors.initials: L
authors.lastname: Lu
authors.forename: Jia-Min
authors.initials: JM
authors.lastname: Zhou
authors.forename: Long-Rong
authors.initials: LR
authors.lastname: Wang
authors.forename: Ke-Zhi
authors.initials: KZ
authors.lastname: Zhang
authors.forename: Qiang-Bo
authors.initials: QB
authors.lastname: Zhang
authors.forename: Jian-Yang
authors.initials: JY
authors.lastname: Ao
authors.forename: Miao
authors.initials: M
authors.lastname: Wang
authors.forename: Wei-Zhong
authors.initials: WZ
authors.lastname: Wu
authors.forename: Lu
authors.initials: L
authors.lastname: Wang
authors.forename: Zhao-You
authors.initials: ZY
authors.lastname: Tang
authors.forename: Hui-Chuan
authors.initials: HC
authors.lastname: Sun
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D056945:Hep G2 Cells; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism*; D008156:Luciferases; D051379:Mice; D008819:Mice, Nude; D035683:MicroRNAs / Q000494:pharmacology* / Q000627:therapeutic use; D009389:Neovascularization, Pathologic / Q000188:drug therapy*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
chemical_list: D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008156:Luciferases; D035683:MicroRNAs; D019869:Phosphatidylinositol 3-Kinases; D034741:RNA, Small Interfering; D042461:Vascular Endothelial Growth Factor A; C497442:HIF1A protein, human; C528163:MIRN26 microRNA, human; C467484:VEGFA protein, human; C546621:PIK3C2A protein, human