Combined silencing of TGF-β2 and Snail genes inhibit epithelial-mesenchymal transition of retinal pigment epithelial cells under hypoxia

The formation of scar-like fibrous tissue in age-related macular degeneration (AMD) is associated with hypoxia. Under hypoxia, retinal pigment epithelial (RPE) cells can secret more transforming growth factor-beta(2) (TGF-beta(2)), which is determined to induce epithelial-mesenchymal transition (EMT) at certain concentrations. Whether hypoxia can induce EMT by stimulating RPE cell line secrets TGF-beta(2) or not remains unknown. To gain a better understanding of the signaling mechanisms of fibrosis in AMD under hypoxic conditions, we investigated EMT in retinal pigment epithelial (RPE) cells and the effect of TGF-beta(2) and Snail in this process. Human RPE cell line (ARPE-19) was incubated with 5 % O-2 for different periods of time. The expression of N-cadherin, alpha-smooth muscle actin (alpha-SMA), TGF-beta(2) , and Snail were determined by Western blot and real-time PCR. Cell proliferation was assessed by CCK8 kit. RNA interference was used for multi-gene silencing of TGF-beta(2) and Snail genes. N-cadherin was decreased and mesenchymal cell marker alpha-SMA was increased after the ARPE-19 cell line was incubated with 5 % O-2. Meanwhile, the proliferation capability of the cell line was increased. TGF-beta(2) and Snail expression were increased in a time-dependent manner under hypoxia. After multi-silencing TGF-beta(2) and Snail genes, N-cadherin was increased and alpha-SMA was reduced. Meanwhile, the proliferation of the cell line was suppressed. Under hypoxic conditions, RPE cells undergo EMT. Endogenic TGF-beta(2) and Snail are involved in this process. Furthermore, knockdown of both TGF-beta(2) and Snail inhibited EMT to a greater extent than knockdown of either gene individually.