Effects of Hypoxic Preconditioning and Vascular Endothelial Growth Factor on the Survival of Isolated Primary Retinal Ganglion Cells

Abstract

This study aimed to investigate the effect of hypoxic preconditioning (HPC) on primary retinal ganglion cell (RGC) survival and the associated mechanism, including the role of vascular endothelial growth factor (VEGF). Retinas were separated from the enucleated eyeballs of Sprague-Dawley rats on postnatal days 1-4. RGCs were harvested using an immunopanning-magnetic separation system and maintained for 24 h in a defined medium. Hypoxic damage (0.3% O2) was inflicted on the cells using a CO₂ chamber. Anti-VEGF antibody (bevacizumab) was administered to RGCs exposed to hypoxic conditions, and RGC survival rate was compared to that of non-anti-VEGF antibody-treated RGCs. HPC lasting 4 h significantly increased RGC survival rate. In the RGCs exposed to hypoxic conditions for 4 h, VEGF mRNA and protein levels were significantly increased. Treatment with high dose bevacizumab (>1 mg/mL) countered HPC-mediated RGC survival. Protein kinase B and focal adhesion kinase levels were significantly increased in 4-h hypoxia-treated RGCs. HPC showed beneficial effects on primary RGC survival. However, only specifically controlled exposure to hypoxic conditions rendered neuroprotective effects. Strong inhibition of VEGF inhibited HPC-mediated RGC survival. These results indicate that VEGF may play an essential role in promoting cell survival under hypoxic conditions.