OBJECTIVE: To investigate the possible pathway involved in hydrogen peroxide (H2O2) induced apoptosis in U251 glioma cells. MATERIALS AND METHODS: The cultured U251 glioma cells were used in this study. The cells were divided into three groups: control group (untreated glioma cells), H2O2 group (treated with 100, 300 and 500 mu M H2O2) and CI group (treated with 300 mu M H2O2 and 15 mu M caspase inhibitor, CI). The cellular viability was determined by MTT [3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide] assay. A flow cytometer was used for measuring the cell cycles. The mode of cell death was assessed by Annexin-V/PI-flow cytometry analysis. Fluorescence dihydroethidium (DHE) method was conducted to detect the oxygen species (O-2(-)). Western blot analysis was performed to confirm the pro-caspase-3, caspase-3 and PARP (poly-ADP-ribose polymerase) protein expression. RESULTS: The oxidative stress to U251 glioma cells exhibited in a dose-dependent manner with H2O2 concentrations increasing. The cell viability was considerably decreased and apoptosis occurred in H2O2 treated cells. A G1 cell cycle arrest and O-2(-) level increase were found in H2O2 group. Western blot analysis showed a decrease of pro-caspase-3 protein level and an increase of caspase-3 and PARP level in 300 mu M H2O2 treated cells. The H2O2 induced apoptosis depicted above was significantly restrained by CI. CONCLUSIONS: Oxidative stress inhibits growth and induces apoptotic cell death in human U251 glioma cells via the caspase-3-dependent pathway. Mitochondrial pathway might involve in this signaling conduction. These findings are favorable for understanding the mechanisms of oxidative stress-induced apoptosis in U251 glioma cells.
[1]Jilin Univ, Hosp 1, Dept Neurosurg, Changchun 130023, Peoples R China.
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