Effects And Mechanisms Of Ginkgolide A Against Alzheimer’s Disease

Alzheimer’s disease (AD) is a neurodegenerative disease of the central nervous system in old age or early old, with the main feature including cognitive impairment of the memory and personality changes. Its typical pathological changes include beta-amyloid protein (Aβ) deposition, tau protein hyperphosphorylation and extensive neurons loss in the hippocampus. With the global acceleration of aging, the incidence of AD becomes increasingly pronounced, which takes heavy burden to family and society. AD has become the concerned research topics of neuroscience and geriatrics, that is imminent to develop AD therapy and explore the molecular mechanism.Objective:(1) To detect the protection of ginkgolide A on N2a cells apoptosis induced by okadaic acid.(2) To investigate the effects of ginkgolide A on tau hyperphosphorylation induced by okadaic acid and PI3K-Akt signaling pathway in N2a cells.(3) To study the impact of ginkgolide A on the damage of brain neurons and tau hyperphosphorylation induced by (3-amyloid protein (Aβ) in the proposed AD mice.Methods:(1) The toxic of ginkgolide A on N2a cells and the protection of ginkgolide A on okadaic acid-induced cell apoptosis were detected by MTT assay; The protection of ginkgolide A on N2a cells cultured with okadaic acid was observed by fluorescence microscopy with acridine orange/ethidium bromide (AO/EB) staining method; The effects of ginkgolide A on okadaic acid-induced cell apoptosis were determined by flow cytometry with Annexin V-FITC/PI staining method.(2) Tau hyperphosphorylation in N2a cell was induced using okadaic acid before ginkgolide A was given. The content of p-Tau in N2a cell lysates were detected using enzyme-linked immunosorbent assay (ELISA), and the expression of p-Tau, p-GSK3p, p-PI3K and p-Akt were determined using western blot assay.(3) Establish an animal model of Alzheimer’s disease (AD) by unilateral injection of Aβ1-40into hippocampus of mice. The phosphorylation levels of tau protein in the hippocampus of mice were detected by ELISA and western blot, and the morphological changes of mouse brain cell pathology were observed using HE staining method.Results:(1) Okadaic acid-induced N2a cell viability decrease was prevented after treatment using different concentrations of GA (10-4-10-7M). Compared with normal controls, apoptosis and necrosis of OA model group were significantly increased, however, apoptosis and necrosis situation has improved after treatment using different concentrations of GA, cells of GA high-dose group (10-4M) almost restored to normal state, indicating that GA could protect and improve the OA-induced apoptosis.(2) GA low-dose (10-7M) had little effect, but tau protein phosphorylation was significantly improved with the increase of GA concentration. The phosphorylation of GSK-3β at Ser9site was inhibited, meanwhile PI3K and Akt were phosphorylated at Tyr508and Ser473respectively.(3) Compared with model group, the phosphorylation levels of tau protein in proposed AD mice declined after treatment with GA. The number of neurons significantly increased and the cell morphology was relatively complete, indicating that GA could against the toxic induced by A(3.Conclusion:Neuronal apoptosis is an important mechanism in the process of AD, it was found that ginkgolide A could protect N2a cells against OA-induced apoptosis with a certain degree; NFTs is a typical pathological feature in the brain of AD patients, and abnormal phosphorylated tau protein is the main components of NFTs, the results of this study showed that ginkgolide A improved tau hyperphosphorylation with dose-dependent, and the mechanism may be the activation of PT3K-Akt signaling pathway; In addition, the results of the proposed AD mice experiments showed that ginkgolide A significantly inhibited Aβ-induced neurotoxicity.