Estrogen attenuates oxidative stress-induced apoptosis in C6 glial cells: In memory of Dr. Pratima Sur, 1946-2002, who started this work and contributed significantly. We will deeply miss her

We examined the mechanism of 17 beta -estradiol (estrogen)-mediated inhibition of apoptosis in C6 (rat glioma) cells following exposure to hydrogen peroxide (H2O2). Cells were preincubated with 4 mu M estrogen for 2 h and then exposed to 100 mu M H2O2 for 24 h. Exposure to H2O2 caused significant increases in intracellular calcium (Ca super(2+)), as determined by fura-2, which was attenuated by preincubation with estrogen. H2O2 and ionomycin caused cell death in a dose-dependent manner, as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Preincubation with estrogen restored viability in cells exposed to H2O2 but not in cells exposed to ionomycin. Western blot analysis showed an increase in Bax/Bcl-2 ratio, calpain activity, and caspase-3 activity following treatment with H2O2, and estrogen pretreatment decreased levels of all three. Cell morphology, as evaluated by Wright staining, indicated apoptosis in cells treated with H2O2, and pretreatment with estrogen reduced apoptosis. Results from MTT and Wright staining were further supported by the terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP Nick End Labeling (TUNEL) assay. These results indicate a role for estrogen in preventing apoptosis in C6 glial cells exposed to H2O2. Our results suggest that estrogen may have a protective role in minimizing glial cell apoptosis in neurological diseases such as demyelinating disease or central nervous system trauma.