Erythropoietin Protects Adult Retinal Ganglion Cells against NMDA-, Trophic Factor Withdrawal-, and TNF-α-Induced Damage

PURPOSE: This study aimed to evaluate the neuroprotective effect of EPO in the presence of N-methyl-d-aspartate (NMDA)-, trophic factor withdrawal (TFW)-, and tumor necrosis factor-alpha (TNF-α)-induced toxicity on total, small, and large retinal ganglion cells (RGCs). METHODS: Retinal cells from adult rats were cultured in a medium containing brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF), basic fibroblast growth factor (bFGF), and forskolin. Expression of RGC markers and EPOR was examined using immunocytochemistry. RGCs were classified according to their morphological properties. Cytotoxicity was induced by NMDA, TFW, or TNF-α. RGC survival was assessed by counting thy-1 and neurofilament-l double-positive cells. RESULTS: EPO offered dose-dependent (EC(50) = 5.7 ng/mL) protection against NMDA toxicity for small RGCs; protection was not significant for large RGCs. Time-course analysis showed that the presence of EPO either before or after NMDA exposure gave effective protection. For both small and large RGCs undergoing trophic factor withdrawal, EPO at concentrations of 1, 10, or 100 ng/mL improved survival. However, EPO had to be administered soon after the onset of injury to provide effective protection. For TNF-α-induced toxicity, survival of small RGCs was seen only for the highest examined concentration (100 ng/mL) of EPO, whereas large RGCs were protected at concentrations of 1, 10, or 100 ng/mL of EPO. Time-course analysis showed that pretreatment with EPO provided protection only for large RGCs; early post-treatment with EPO protected both small and large RGCs. Inhibitors of signal transduction and activators of transcription such as (STAT)-5, mitogen-activated protein kinases (MAPK)/extracellular-regulated kinase (ERK), and phosphatidyl inositol-3 kinase (PI3K)/Akt impaired the protective effect of EPO on RGCs exposed to different insults. CONCLUSION: EPO provided neuroprotection to cultured adult rat RGCs; however, the degree of protection varied with the type of toxic insult, RGC subtype, and timing of EPO treatment.