Protective effects of ginkgo biloba leaves extract on peroxide-induced oxidative stress damage in PC12 cells

Abstract

Background Extracts of ginkgo biloba leaves (EGB) and its metabolites have been reported to enhance brain function and nerve behavior. It has also been hypothesized that they can protect neurons from oxidative stress. Objective To investigate protective effects of EGB on peroxide (H 2O 2)-induced oxidative stress damage in PC12 cells. Design Observational contrast study. Setting Department of Pathophysiology, Guangdong Pharmacological College. Materials EGB was provided by Xi'an Fujie Biotechnological Development Company; 1640 culture medium, methylthiazolyl tetrazolium (MTT), trypsin and dimathyl sulfoxide (DMSO) by Sigma Company; PC12 cell strain by Cell Center of Medical College of Zhongshan University; calf serum by Hangzhou Sijiqing Bioengineering Company; lactate dehydrogenase (LDH) kit by Nanjing Jiancheng Bioengineering Research Institute. Methods The experiment was carried out in Department of Cell Biology of Guangdong Pharmacological College from June to December 2005. ▪ Cell culture PC12 cells were cultured in 1640 medium containing 200 g/L fetal calf serum. The cells were diluted to 1×10 7 L −1 and washed every two days. Those cells were used to experiment until they grew in logarithm on solid wall. ▪ Grouping and intervention: PC12 cells (1×10 8 L −1) were plated in 96-well plates with the density of 200 μL/hole and divided into three groups: normal control group (routinely adding media), H 2O 2 group (treating with media and H 2O 2 for 20 hours) and EGB group (adding media, 100 μmol/L EGB and 100 μmol/L H 2O 2). ▪ MTT assay: PC12 cells (1× 10 8 L −1) were plated in 96-well plates and divided into three groups with 8 holes for each group. Under sterile condition, cells were added with 5 g/L MTT (100 μL) and cultured for 4 hours. And then, 200 μL DMSO fluid was added and shaken for 30 minutes until blue crystal products formed were dissolved soundly. ▪ Experimental evaluation: Absorbance ( A) at 630 nm was measured and LDH activity was measured at the same time. Main Outcome Measures Results of MTT assay and LDH activity. Results ▪Results of MTT assay A value was lower in the H 2O 2 group than that in the normal control group ( P < 0.01), while A value was higher in the EGB group than that in the H 2O 2 group ( P < 0.01). ▪ LDH activity: LDH activity was higher in the H 2O 2 group than that in the normal control group ( P < 0.01), while LDH activity was lower in the EGB group than that in the H 2O 2 group ( P < 0.01). Conclusion EGB can inhibit H 2O 2-induced oxidative stress damage in PC12 cells possibly by preventing damage to the cell membrane.