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S. He, P. Zhou, N. Chan, J. Xu, S. Ryan, D. Hinton; The Effects of Knock Down of Mecp2 on TGFβ Signaling Involved in RPE Transdifferentiation. Invest. Ophthalmol. Vis. Sci. 2010;51(13):503.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal pigment epithelial (RPE) cell transdifferentiation is a critical step in the wound healing response in proliferative vitreoretinopathy (PVR). Our previous studies have shown that human PVR membranes are rich in methyl CpG binding protein 2 (MeCP2). In current experiments, we investigated the effects of knocking down MeCP2 by siRNA on TGFβ signaling involved in RPE transdifferentiation.
Early passage human fetal RPE cells were used in the study. MeCP2 expression in scratch wounded RPE monolayer was studied by immunocytochemistry. The effect of methylation inhibitor on the expression of proteins MeCP2, TGFβ II receptor, and PPAR-γ was evaluated by western blot after 48 hours 5-aza-2’-deoxycytidine (1, 2 and 6 uM) treatment. For the analysis of the role of MeCP2 in TGFβ mediated fibrosis, RPE were pretreated with siRNA MeCP2 (0.1 or 10nM) and then with or without TGFβ (10ng/ml) for 48 hours ,the response of MeCP2 knockdown on the expression of Smad/2/3, MeCP2, alpha Smooth muscle actin (a-SMA), and fibronectin (FN) induced by TGFβ were determined by western blot.
MeCP2 was prominently expressed at the wounded edge of the scratched RPE monolayer, however MeCP2 expression was low away from wounded area. In the presence of 5-AZA (2-6 uM), the expression of MeCP2 and TGFβ receptor II were significantly inhibited. A strong increase of FN and α-SMA stimulated by TGFβ is seen when RPE were treated with control (scrambled) siRNA, while up regulation of FN ,α-SMA and activation of Smad2/3 by TGFβ was much attenuated by pretreatment with MeCP2 specific siRNA. Inhibition of MeCP2 also up-regulated Smad 7 and PPAR-γ.
These results support the idea that RPE transdifferentiation may be under epigenetic control and that MeCP2 may be a critical factor in the regulation of the PVR pathogenesis.
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