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N. Agarwal, I. Charles, D.M. Kumar, R.R. Krishnamoorthy, R.S. Roque; Serum Deprivation Induced Apoptosis Signaling Pathways of RGC–5 Retinal Ganglion Cells. . Invest. Ophthalmol. Vis. Sci. 2004;45(13):2165.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To determine the signaling pathways of apoptosis in retinal ganglion cells deprived of growth factors. Methods: An established line of transformed rat retinal ganglion cells, RGC–5, was subjected to serum deprivation for 3–6 days. Control cells were cultured in growth medium containing 10% fetal calf serum. Cell viability was measured by neutral red assay, apoptotic signaling by immunoblot analyses, electrophoretic mobility shift assay, and biochemical methods. Results: Serum deprivation of RGC–5 cells for three days resulted in 50% cell loss due to apoptosis as established by DNA laddering and propidium iodide staining. Serum deprivation also resulted in increased oxidative stress as revealed by lowering of reduced glutathione (GSH) and increased levels of malonydialdehyde (MDA). Reduced levels of NF–kB binding activity was observed upon serum deprivation of RGC–5 cells. Serum deprivation was also associated with a loss of mitochondrial function as revealed by cytochrome–c release and rhodamine 123 staining. The RGC–5 cell death was further augmented by PI3K inhibitor (LY 294002), MAPK inhibitor (PD 098059), and a Trk– receptor inhibitor (K252a). Serum deprivation also resulted in increased phosphorylation of ERK proteins. Conclusions: These results indicate that serum deprivation of RGC–5 cells result in apoptotic cell death by means of mitochondrial pathways involving oxidative stress, PI3K, MAPK and trk receptors. Supported by American Health Assistance Foundation–National Glaucoma Program (NA).
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