RESUMO
Transforming growth factor-ß1 (TGF-ß1)induced epithelialmesenchymal transition (EMT) is one of the important cellular and molecular mechanisms involved in renal fibrosis. Smad3 and miR-192 (a Smad3-dependent microRNA) are involved in TGF-ß1-mediated EMT. Vascular endothelial growth factor (VEGF) is a renal tubular epithelial survival factor. Therefore, in the present study, we investigated the role of Smad3 and miR192 in the effects of VEGF on TGFß1mediated tubular EMT. A human kidney cortex (HKC) cell line stably overexpressing VEGF (HKC-SOEV) was established. The normal HKC cells and HKCSOEV cells were treated with TGF-ß1 (5 µg/l) or/and LY294002 (20 µmol/l) for 24 and 48 h (LY294002 blocks the effect of VEGF). The protein expression of Smad2, Smad3, Smad4 and phosphorylated Smad3 (pSmad3) were measured by western blot analysis. The expression of Smad3 and miR-192 was determined by realtime PCR. E-cadherin and α-smooth muscle actin (α-SMA) expression was detected by western blot analysis and laser scanning confocal microscopy (LSCM). TGF-ß1 was found to induce the expression of α-SMA in the HKC cells. TGF-ß1 also induced Smad3, miR-192 and p-Smad3 expression, but suppressed Ecadherin expression. However, in the HKC-SOEV cells, the expression levels of α-SMA, Smad3, miR-192 and pSmad3 upon TGF-ß1 stimulation were significantly reduced. In these cells, the suppressive effect of TGF-ß1 on Ecadherin was also reduced. Importantly, treatment with LY294002 significantly diminished the effect of VEGF. VEGF suppressed Smad3 and miR192, and subsequently inhibited EMT induced by TGF-ß1.
