Influence of Ginkgo Biloba extract on beta-secretase in rat hippocampal neuronal cultures following chronic hypoxic and hypoglycemic conditions

BACKGROUND: Preparation of Ginkgo leaf has been widely used to improve cognitive deficits and dementia, in particular in Alzheimer's disease patients. However, the precise mechanism of action of Ginkgo leaf remains unclear. OBJECTIVE: To explore the effect of Ginkgo Biloba extract (Egb761), Ginaton, on beta-secretase expression in rat hippocampal neuronal cultures following chronic hypoxic and hypoglycemic conditions. DESIGN,TIME AND SETTNG: Completely by randomized, grouping study. The experiment was performed at the Laboratory Of molecular Imaging, Southeast University between August 2006 and August 2007. MATERIALS: A total of 128 Wistar rats aged 24 hours were selected, and hippocampal neurons were harvested for primary cultures. METHODS: On clay 7, primary hippocampal neuronal Cultures were treated with Egb761 (0, 25, 50, 100, 150, and 200 mu g/mL) under hypoxic/hypoglycemic or hypoglycemic Culture conditions for 12, 24, and 36 hours, respectively. Hippocampal neurons cultured in primary Culture medium served as control. MAIN OUTCOME MEASURES: Cell viability was assayed using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT); fluorescence detection of beta-secretase activity was performed: Western Blot was used to measure beta-secretase expression. RESULTS: Cell viability under hypoxic/hypoglycemic or hypoglycemic Culture conditions was significantly less than control cells (P < 0.05). Under hypoxic/hypoglycemic or hypoglycemic culture conditions, treatment with 25 mu g/mL Egb761 did not alter cell viability. However, > 25 P g/mL, Egb761 induced greater cell viability (P < 0.05). No differences were observed between hypoxic/hypoglycemic or hypoglycemic cells (P > 0,05). beta-secretase activity was increased after 12 hours in hypoxic/hypoglycemic Culture (P < 0.01). There were no significant differences between the 12-, 24-, or 36-hour Egb761 groups and the hypoxic/hypoglycemic groups (P > 0.05). beta-secretase activity was greater after 12, 24, and 36 hours in hypoxic/hypoglycemic Culture conditions, compared with control conditions (P < 0.05). beta-secretase activity was significantly decreased in neurons treated with Egb761 for 12, 24, or 36 hours, compared with the hypoxic/hypoglycemic group (P < 0.05). beta-secretase protein expression Was Significantly up-regulated in neurons cultured in hypoxic/hypoglycemic conditions for 12, 24, or 36 hours, compared to control cells (P < 0.05), and was decreased compared to neurons treated with Egb761 (P < 0.05). CONCLUSION: beta-secretase expression and activity in rat neonatal hippocampal neurons were influenced by hypoxic and hypoglycemic Culture. Egb761 played a protective role in hippocampal neurons damaged by chronic hypoxic and hypoglycemic culture conditions, possibly through its effect on beta-secretase expression and activity.