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Fang Wang, Min Li, Hui Li, Chen Sun, Hao Wang, Jun Yao, ; MicroRNA-29b Regulates TGF-β1-Mediated Epithelial-Mesenchymal Transition of Retinal Pigment Epithelial Cells by Targeting AKT2. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4994.
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© ARVO (1962-2015); The Authors (2016-present)
The role of microRNA (miRNA) in proliferative vitreoretinopathy (PVR) progression has not been studied extensively, especially in retinal pigment epithelial-mesenchymal transition (EMT) which is the main reason for formation of PVR. The aim of this study was to investigate and validate unidentified miRNAs that regulate EMT and to reveal the function in PVR clinical samples.
MiRNA expression profile in ARPE-19 cells EMT model was performed by microRNA microarray. Immunofluorescence staining, Q-PCR, western blot were conducted to determine the level of three epithelial and mesenchymal markers zona occludin-1 (ZO-1), E-cadherin and α-smooth muscle actin (α-SMA) in ARPE-19 cells and PVR membranes. The migration ability of treated ARPE-19 cells was accessed by Transwell migration assay. The target of miR-29b was predicted by computational approach and identified by Dual luciferase activity assay.
MiRNA microarray detected that miR-3138, miR-22*, and miR-455-3p were upregulated, while miR-550a, miR-135b and miR-29b were significantly downregulated in TGF-β1 mediated EMT of ARPE-19 cells. Among the five changed miRNAs, miR-29b showed the most significant downregulation. Enhanced expression of miR-29b could reverse TGF-β1 induced EMT through targeting Akt2(Figure1 and 2). Akt2 downregulation could inhibit TGF-β1 induced EMT. Furthermore, inhibition of miR-29b in ARPE-19 cells directly triggered EMT process, which characterized by the phenotypic transition and the upregulation of α-SMA and downregulation of E-cadherin and ZO-1 with increased cell migration. Akt2-shRNA also inhibited miR-29 inhibitor induced EMT process. In epiretinal membrane and vitreous of PVR patients, miR-29b expression was downregulated and positively correlated with E-cadherin and ZO-1 mRNA expression, while an inverse correlation was observed between miR-29b and α-SMA or Akt2 mRNA expression.
Our findings suggest that miR-29b plays an important role in TGF-β1-mediated EMT in ARPE-19 cells and PVR pathological process by targeting Akt2.
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