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D. Hunt, P. Kothary, M. Del Monte; PD 98059 Inhibits Insulin Stimulated Synthesis of Connective Tissue Growth Factor in Human Retinal Pigment Epithelial Cells . Invest. Ophthalmol. Vis. Sci. 2006;47(13):3824.
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Insulin is known to stimulate hRPE cell proliferation. Connective Tissue Growth Factor (CTGF) is associated with fibrosis and has recently been found in high levels in the vitreous of diabetic retinopathy (DR) patients, suggesting that CTGF may contribute to the formation of proliferative membranes. Because the mechanism of action of CTGF has been shown to involve the MAP kinase pathway in cardiac fibroblasts, we investigated the effect of the MAP kinase inhibitor PD 98059 (PD) on insulin stimulated CTGF synthesis and cell proliferation in human retinal pigment epithelial cells (hRPE).
Primary hRPE cell cultures were established from human eyes, and then treated with insulin ± PD. Cell proliferation was assayed using 3H–thymidine incorporation and by direct cell counting using the trypan blue exclusion method. 14C–methionine labeled CTGF synthesis was assessed using CTGF–specific immunoprecipitation. 14C–met and 3H–thymidine assays were expressed as cpm per well and cell viability results as cells per 1.0 µl. Data were analyzed using the Student ‘t’ test.
Insulin increased hRPE cell proliferation and CTGF synthesis in a dose dependant manner. PD inhibited insulin stimulated cell proliferation as measured by 3H–thymidine incorporation (1545 ± 83 vs. 5529 ± 352, p<0.05, mean ± SEM, n=6) and by the trypan blue exclusion method (9.27 ± 0.59 vs. 16.5 ± 0.70, p<0.05, mean ± SEM, n=6). PD also inhibited insulin stimulated 14C–met CTGF synthesis (319.97 ± 55.79 vs. 707.14 ± 119.66, p<0.05, mean ± SEM, n=6).
PD inhibits insulin stimulated hRPE cell proliferation and CTGF synthesis. These findings suggest that if CTGF is involved in proliferative eye disease and traction membrane formation then PD may have therapeutic value in the treatment of this disease.
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