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Oxidative Stress Induces Synthesis of TGF-beta2 and Increases ECM Gene-expression in Retinal Pigment Epithelial Cells
Alge C., Priglinger S., Neubauer A. S., Kampik A., Welge-Lüßen U.,
Ludwig-Maximilians-Universität München, Klinikum Innenstadt, Augenklinik (München)
Purpose: Age related macular degeneration (AMD) is the leading cause of severe vision loss in the western world. Early AMD is characterized in part by the deposition of extracellular matrix (ECM) material between the retinal pigment epithelium (RPE) and the choroid. Oxidative stress and an increased expression of the fibrogenic cytokine transforming growth factor beta 2 (TGF-beta2) have been discussed to contribute to AMD pathogenesis. The present study was designed to explore the causal relationship between oxidative stress and the expression of TGF-beta2 and ECM proteins in the RPE.
Materials and Methods: Monolayer cultures of RPE cells from eyes of 5 human donors were exposed to oxidative stress (50 - 200 mM for 1 h) or treated with TGF-beta2 (1ng/ml for 2 - 24 h). mRNA-induction of TGF-beta2 and the ECM proteins fibronectin, laminin and osteonectin was investigated by Northern blot analysis. Total TGF-beta2 protein (latent and active) was measured in RPE culture supernatants using a specific sandwich ELISA.
Results: Treatment of cultured RPE cells with oxidative stress led to a substantial increase in TGF-beta2 mRNA and protein (2 - 4 fold). Northern blot analysis revealed significantly increased steady state levels of ECM proteins following oxidative stress and treatment with TGF-beta2, respectively. Pretreatment with Actinomycin D completely abrogated this effect.
Conclusions: Oxidative stress induces synthesis of TGF-beta2 and increases ECM gene expression in cultured RPE cells. These results suggest that oxidative stress and increased expression of TGF-beta2 in macular RPE of AMD patients might be causally linked. This cellular response might contribute to the deposition of ECM material in the aging macula.