Article ID Journal Published Year Pages File Type
4012899 Experimental Eye Research 2007 11 Pages PDF
Abstract

VEGF secretion by the human retinal pigment epithelium (hRPE) plays an important role in retinal and choroidal neovascularization. In this study, transforming growth factor-β2 (TGF-β2)-induced vascular endothelial growth factor (VEGF) gene expression was investigated in hRPE cells. Treatment of hRPE cells with TGF-β2 for 24 and 48 h as compared to 8 h resulted in markedly increased VEGF secretion by fivefold and nine-fold, respectively. Induced VEGF mRNA peaked within 3 h of stimulation and remained above the basal at 36 h. Stimulation of VEGF expression by TGF-β2 was blocked by cycloheximide, suggesting that de novo protein synthesis is required. Induced VEGF production was strongly inhibited by anti-inflammatory agents, dexamethasone and cyclosporin A. Despite of the weak stimulation of VEGF expression by TNF-α or bFGF alone, co-administration of either of these two cytokines synergized the effect of TGF-β2 on VEGF mRNA expression and protein production. Quantitative RT-PCR revealed that the synergy was predominantly at the level of VEGF transcription. Moreover, TGF-β2-induced RPE VEGF secretion was significantly reduced by inhibitors of mitogen-activated protein (MAP) kinase (MEK) (U0126), p38 (SB202190), c-Jun NH2-terminal kinase (JNK), Sp600125, protein tyrosine kinase (PTK) (Genistein), and phosphatidylinositol 3-kinase (PI3K) (Ly294002). Induced VEGF expression was completely abrogated by inhibitors of protein kinase C (PKC) (Ro318220), nuclear factor-κB (NF-κB) [caffeic acid phenethyl ester (CAPE)], and reactive oxygen species (ROS) [N-acetyl-cysteine (Nac) and diphenyleneiodonium (DPI)]. These results suggest that MEK, p38, JNK, PI3K, and NF-κB as well as multiple essential signaling intermediates, including PKC, PTK and ROS, are involved in hRPE VEGF up regulation by TGF-β2.

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