Fig. 3. Hypoxia induces HIF-1α and HIF-2α expression in 661W photoreceptor cells.

(A) HIF-1α and HIF-2α protein immunoreactivity on western blots of homogenates from 661W cells incubated in normoxic conditions or hypoxic (1% O₂, 5% CO₂, 94% N₂) conditions over times indicated. GAPDH served as a loading control. The ratios of HIF-2α and GAPDH levels based on densitometry of respective bands from western blots are shown. Control sample, C, were from cells grown in normoxic conditions. Images are representative of 9 independent experiments with cells between passages 4 and 6. The overall effect of hypoxia on HIF-2α protein levels was assessed by ANOVA followed by Bonferroni’s test p<0.05. (B) Immunohistochemical analysis of cultured 661W photoreceptor cells under hypoxic conditions for 24 hours and normoxic controls. Cells were fixed and stained with antibodies recognizing HIF-1α or HIF-2α. DAPI-stained nuclei are shown in blue. Control images of cells from which primary antibodies are omitted are shown.