Study On Neuroprotective Effect Of 6-gingerol And Its Mechanism To Alzheimer’s Disease

Backgroud and Objective:Alzheimer’s disease (AD) is the most common human age-related progressive neurodegenerative disorder characterized by disordered cognitive function, progressive memory impairment and altered behavior, and has been put into the list of the five primary diseases of 21st century by WHO. Currently, no effective anti-AD drugs are available to either stop or reverse the progression of AD, although the development of anti-AD drugs has been slightly successful in aspects of symptomatic improvement. Our previous study suggests that ginger root extract can reverse behavioral dysfunction and prevent Alzheimer’s disease (AD)-like symptoms induced by amyloid β-protein (Aβ) in a rat model.6-gingerol, a phenolic compound, is the major ingredient of ginger with diverse pharmacological activities, but its effect on the treatment of AD remains unclear. In this study, we aimed to investigate the effect of 6-gingerol on Aβ-42 aggregation and to determine if 6-gingerol had a protective effect on Aβ1-42-induced damage in PC 12 cells. Based on these findings, we further investigated the underlying mechanisms by which 6-gingerol may exert its neuroprotective effects.Methods:(1) Aβ1-42 was incubated with or without 6-gingerol at different concentrations for 5 d, and thioflavine-T (ThT) fluorometric assay was performed to determine the effect of 6-gingerol on Aβ1-42 aggregation in vitro; (2) After 5 d of induced by nerve growth factor (NGF), the differentiated PC 12 cells were treated with different concentrations of Ap 1-42 for 24,48 and 72 h, and cell viability was detected by MTT assay; MTT assay was also performed to determine the protective effect of 6-gingerol (40,80,120,200, and 300 μM) on PC 12 cells apoptosis induced by Aβ1-42; (3) To detect the effect of 6-gingerol on Aβ1-42-induced cell apoptosis, the chromosomal condensation and morphological changes of the cell nucleus were assessed by staining with Hoechst 33258; (4) The anti-apoptotic effects of 6-gingerol in PC 12 cells were further studied by flow cytometry; (5) DCFH-DA method was performed To evaluate the effect of 6-gingerol on Aβ1-42-induced ROS; (6) Effect of 6-gingerol on Aβ1-42-induced lactate dehydrogenase (LDH) and superoxide dismutase (SOD) activity in the medium, malondialdehyde (MDA) and nitric oxide (NO) generation were detected with their assay kit on the basis of the manufacturer’s instructions respectively; (7) Western blot was performed to evaluate the expressions of glycogen synthase kinase-3β (GSK-3β), phosphorylated GSK-3β (Ser9), Akt and p-Akt(Ser473).Results:(1) ThT fluorometric assay showed that the 6-gingerol treatment group significantly reduced the fluorescence intensity compared with the AP1-42 treatment group, indicating that 6-gingerol could attenuate Aβ1-42 aggregation; (2) The MTT assay revealed that the PC 12 cells viability were decreased with the increasing concentrations of Aβ-42 and the duration of interventions, and the cell model of AD was ideally established after stimulation for 48 h by 10 μM Aβ1-42. (3) The MTT assay showed that pretreatment with 6-gingerol (80,120 and 200 μM) variably reduced cell apoptosis compared with the Aβ-42 group, of which 120μM 6-gingerol treatment showed the best effect; (4) Hoechst 33258 and Annexin V-FITC/PI staining also revealed that pretreatment with 6-gingerol (80, 120 and 200 μM) variably reduced cell apoptosis compared with the Aβ1-42 treatment group; (5) Treatment with Aβ1-42 markedly increased the level of intracellular ROS and MDA, the leakage of LDH and the production of NO and decreased the SOD activity compared with the Aβ1-42 treatment group, whereas 6-gingerol could reverse these changes; (6) Treatment with Aβ1-42 significantly suppressed the expression of p-GSK-3(3 and p-Akt, whereas pretreatment with 6-gingerol (80 and 120 μM) significantly inhibited the decreased expression of p-GSK-3β and p-Akt caused by Aβ1-42.Conclusions:(1) 6-gingerol may exert neuroprotective effects by inhibiting Aβ1-42 aggregation; (2) 6-jingerol exhibited protective effects on apoptosis induced by Aβ-42 in cultured PC 12 cells by reducing oxidative stress and inflammatory responses, suppressing the activation of GSK-3β and enhancing the activation of Akt, thereby exerting neuroprotective effects. Therefore, 6-gingerol may be useful in the prevention and/or treatment of AD. In addition, further studies are needed to investigate the underlying molecular mechanism.