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M. Yamasaki, T. Inaba, K. Murata, Y. Tsumamoto, K. Okada, K. Kashiwagi, H. Yamashita, H.K. Mishima; Erythropoietin Prevents Glutamate–induced Retinal Ganglion Cell Death . Invest. Ophthalmol. Vis. Sci. 2004;45(13):855.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Erythropoietin (Epo) has been reported to protect not only erythroid progenitors but also neurons from apoptosis. Because retinal ganglion cell (RGC) death is one of the common causes of the major ocular diseases, the potential of Epo to prevent RGCs from neurotoxicity was explored using the primary cultured RGCs isolated from neonatal rats by a two–step panning method. Methods: Expression of Epo receptor (EpoR) was examined by RT–PCR and by double staining method using Thy–1.1 and EpoR antibodies in normal and ischemic conditions. Isolated RGCs were pretreated with Epo (0.5, 1.5U/ml) for 12 hours in a serum–free medium and then were cultured in the medium with glutamate (50mM∼10mM). The survival of RGCs was examined using XTT assay. The expression of Bcl–2, Bcl–xL and Bim, members of the Bcl–2 superfamily, in cultured RGCs was evaluated by real time quantitive PCR. BDNF was used as a positive control to evaluate the potential of Epo in neuroprotection. Results: More than 90% cells obtained by this procedure were proved to be RGCs by DiI, a retrograde fluorescent tracer, injected at superior colliculi. mRNA of EpoR was proved to be expressed in isolated RGCs by RT–PCR. EpoR protein was expressed in the RGCs in the normal retina, and the expression level was enhanced by ischemia. Glutamate decreased the survival rate of RGCs in a dose–dependent manner. Epo significantly reduced neurotoxicity induced by glutamate. Simultaneous upregulation of Bcl–2 and downregulation of Bim was observed in RGCs cultured without cytokines. Epo, like BDNF, reversed this expression change of these Bcl–2 superfamily members. Conclusions: Epo protected primary cultured RGCs from glutamate cytotoxicity. Epo regulated the expression of Bcl–2 and Bim in RGCs. These results suggest that Epo acts directly on RGCs to promote their survival.
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