Alpha-lipoic acid protects against hypoxia/reoxygenation induced injury in rat C6 glioma cells

Abstract

Objective To study the protective effects ofαt-lipoic acid (LA) on rat C6 glioma cells (C6 cells) exposed to hypoxia/reoxygenation and its mechanism.Methods Rat glioma cells C6 were conventionally in vitro cultured and cells at logarithmic growth phase were used in the experiment.Normal control group,oxygen-glucose deprivation/reoxygenation (OGD/ROG) model group,and OGD/ROG plus 10,25 and 50 μmol/L LA treatment groups were chosen.C6 cells were exposed to OGD/ROG to simulate ischemia/reperfusion in vitro.Different concentrations of LA (10,25 and 50μmol/L) were added to the culture medium in the later three groups 12 h before hypoxia.Twelve h after C6 cells reperfusion,cell viability was analyzed by MTT assay; flow cytometry was used to evaluate the level of cellular reactive oxygen species (ROS) and Annexin V/PI staining to examine cell apoptosis rate;the expression of cleaved Caspase-3 protein was determined by Western blotting.Results As compared with normal control group,OGD/ROG model group,and OGD/ROG plus 10,25 and 50 μmol/L LA treatment groups had significantly decreased cell viability and increased cell apoptosis rate (P＜0.05); as compared with that in the normal control group,ROS level in the model group was statistically increased,while that in the 50 μmol/L LA treatment group was significantly decreased (P＜0.05); as compared with that in the normal control group,Caspase-3 expression in the model group and 10 μmol/LA treatment group was significantly increased (P＜0.05).As compared with the model roup,the 25 and 50 μmol/L LA treatment groups had significantly higher cell viability,the 10,25 and 50 μmol/L LA treatment groups had significantly lower ROS level,cell apoptosis rate and Caspase-3 expression (P＜0.05).Conclusion LA has a protective effect on injury in C6 cells induced by hypoxia/reoxygenation,whose mechanism may be related to LA eliminating excess cellular ROS,preventing occurrence of oxidative damage and thereby inhibiting Caspase-3 pathway-dependent apoptosis in C6 cells. Key words: Alpha-lipoic acid; Glioma cell; Hypoxia/reoxygenation; Cell apoptosis