Oxidative stress inhibits growth and induces apoptotic cell death in human U251 glioma cells via the caspase-3-dependent pathway

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 µM H2O2) and CI group (treated with 300 µM H2O2 and 15 µM 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 (O2–). 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 O2– 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 µM 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.

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