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Alice L. Yu, Rudolf Fuchshofer, Daniel Kook, Anselm Kampik, Hans Bloemendal, Ulrich Welge-Lüssen; Subtoxic Oxidative Stress Induces Senescence in Retinal Pigment Epithelial Cells via TGF-β Release. Invest. Ophthalmol. Vis. Sci. 2009;50(2):926-935. doi: 10.1167/iovs.07-1003.
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purpose. The goal of the present study was to determine whether oxidative stress and transforming growth factor (TGF)-β induce cellular senescence in human retinal pigment epithelial (RPE) cells.
methods. Cultured human RPE cells were exposed to 50 to 150 μM hydrogen peroxide (H2O2) for 1 and 2 hours or treated with 1.0 ng/mL TGF-β1 or -β2 for 12, 24, and 48 hours. Senescence-associated β-galactosidase (SA-β-Gal) activity was detected by histochemical staining. Expression of senescence-associated genes (apolipoprotein J [Apo J], connective tissue growth factor [CTGF], fibronectin, and SM22) was examined by real-time PCR and induction of signal transduction proteins (p21, p16, and pRb) by Western blot analysis. The effects of TGF-β blocking on the oxidative stress-induced expression of senescence-associated biomarkers were investigated by simultaneous incubation with neutralizing antibodies against the TGF-β1, -β2, and -β3 isoforms and the TGF-βII receptor.
results. H2O2 markedly increased the number of SA-β-Gal-positive cells to up to 89% and the expression of Apo J, CTGF, fibronectin, and SM22 by approximately three to fourfold. Treatment with TGF-β1 and -β2 showed similar changes. H2O2and TGF-β1 and -β2 markedly enhanced the expression of p21 but downregulated pRb. In contrast, they had no effect on p16 expression. Simultaneous treatment with neutralizing antibodies against the TGF-β1, -β2, and -β3 isoforms and the TGF-βII receptor prevented the oxidative stress–mediated elevation of senescence-associated biomarkers.
conclusions. Oxidative stress, TGF-β1, and TGF-β2 are capable of inducing cellular senescence in cultured human RPE cells. Therefore, reduction of oxidative stress and minimizing TGF-β may help to prevent senescence-associated changes in the RPE as seen in early age-related macular degeneration.
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