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R. J. van Geest, I. Klaassen, I. M. C. Vogels, C. J. F. van Noorden, R. O. Schlingemann; Differential TGF-Beta Signaling Pathways in Retinal Endothelial Cells and Pericytes. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4979.
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We hypothesize that TGF-beta is involved in vascular pro-fibrotic changes in the pathogenesis of diabetic retinopathy. Our aim is to investigate TGF-beta signaling in retinal vascular cells.
Bovine retinal endothelial cells (BRECs) and pericytes (BRPCs) were stimulated with different concentrations of TGF-beta with or without a specific TGF-beta type I receptor-inhibitor, SD-208.To determine TGF-beta-signaling activity, western blotting was used to detect phosphorylated Smad2 protein at different time points. Qualitative RT-PCR was performed to compare gene expression levels of TGF-beta receptors and to determine the effect of TGF-beta on pro-fibrotic gene expression (PAI-1, fibronectin and CTGF) in endothelial cells and pericytes.
TGF-beta receptor-II and ALK5 were equally expressed in both cell types, whereas Endoglin and ALK1 were preferentially expressed in BRECs. In BRECs, TGF-beta dose-dependently induced phospho-Smad2 protein, which was efficiently blocked by SD-208. In contrast, in pericytes, TGF-beta induced phospho-Smad2 protein at low concentration which could be blocked by SD-208. TGF-beta caused upregulation of downstream pro-fibrotic genes PAI-1 and fibronectin in both cell types, which was prevented by SD-208. However, CTGF mRNA expression was only induced by TGF-beta and inhibited by SD-208 in BRPCs.
TGF-beta induces Smad2-phosphorylation in bovine retinal vascular cells through the TGF-beta type I receptor, especially in pericytes, which leads to increased expression of pro-fibrotic genes. This suggests that the pro-fibrotic gene expression observed in DR may be caused by TGF-beta, which may serve as an intervention target.
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