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Y. Wang, G. Dvoriantchikova, D. Ivanov, V. Shestopalov, N. Agarwal, E. Fini, M. L. Bajenaru; Activation of β1-Integrin Signaling Pathway Increases Retinal Ganglion Cells Resistance to Glutamate Induced Oxidative Stress. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5204.
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Extracellular matrix (ECM) integrity in the CNS is essential for neuronal homeostasis and ECM-survival signals are transmitted to neurons through the integrin survival pathway. The purpose of this study is to explore the hypothesis that laminin-integrin survival signaling pathway can modify RGC vulnerability in glaucoma.
As an experimental paradigm of RGC apoptosis in glaucoma we exposed the RGC-5 cell line in culture to glutamate induced oxidative stress. RGC-5 cells were plated without (plastic), or on laminin substrate and treated with high concentrations (5, 10mM) of glutamate, characteristic to glutamate oxidative stress for 24 hours. Cell survival was examined by the MTT assay. Expression levels of components of the integrin survival pathway were examined by western blotting and immunohistochemistry. Pharmacological inhibitors: genistein and PP2 for FAK, and LY294002 for PI3K/Akt were used to demonstrate that the laminin protective effect is specifically mediated by the integrin survival pathway.
We have previously shown that 10 mM glutamate treatment of RGC-5 cells cultured on plastic results in 75% cell death. Laminin confers 25% resistance to RGC-5 cells against glutamate induced oxidative stress, and the protective effect is specifically mediated by β1 integrin receptors. Western blotting analysis showed a significant decrease in the expression level of several focal adhesions (FA) components: β1 integrin, talin, paxillin, and dephosphorylation of FAK in RGC-5 cells as a result of oxidative stress. In addition, oxidative stress results in Akt dephosphorylation, and reduced expression of the anti-apoptotic protein bcl-xL. Moreover, the protective effect of laminin was reversed when RGC-5 cells were treated with different pharmacological inhibitors of FAK, and Akt.
These results suggest that laminin conferred significant RGC-5 protection against glutamate oxidative stress and this protection is mediated specifically via β1 integrin receptors and is attributed to increased integrin survival signaling.
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