Effect of erythropoietin on the survival of retinal neurocytes in culture upon serum withdrawal

OBJECTIVE: To clarify whether erythropoietin (EPO) could substitute for the serum component in cultured retinal neurocytes suffering from serum withdrawal.

METHODS: The study was performed in the Shanghai Institute of Traumatology and Orthopedics, Shanghai, China between April 2008 and March 2009. A total of 160 postnatal 2-3 day-old Sprague-Dawley rats were used for this study. After the retinal neurocytes were cultured for 48 hours, the culture media was replaced with serum- free media, and the cells were exposed to 1 U/ml, 3 U/ml, and 6 U/ml EPO for another 24 or 48 hours, the cell body diameter was then assessed using a computerized image-analysis system, and the survival and apoptosis rates of those cells were estimated by method of transcription and translation assay and flow cytometry. Immunocytochemistry was used to detect EPO and erythropoietin receptor (EPOR) expression.

RESULTS: The retinal neurocytes had obvious EPO/EPOR expression. The early (p=0.002) and total (p=0.049) apoptosis rates of retinal neurocytes cultured with serum withdrawal were significantly higher than that of neurocytes cultured with serum, and the cell viability of neurocytes cultured with serum withdrawal was significantly lower than that of neurocytes cultured with serum (p=0.047). The EPO had no effect on the cell body diameter of cultured retinal neurocytes. The cell viability and the apoptosis rates of retinal neurocytes were not significantly different from that of simple serum-withdrawal culture at any EPO concentration.

CONCLUSION: As the addition of EPO immediately after serum withdrawal had no effect in preventing retinal neurocytes apoptosis induced by serum withdrawal, EPO cannot substitute for the serum component.