Epigallocatechin-3-gallate Protects Neuron Against Rotenone-induced Parkinsonism-like Neurotoxicity Via Activation Of The Nrf2/ARE Signaling Pathway

Objective:Epigallocatechin-3-gallate(EGCG)is the main component of catechol,accounting for about 70% of the total tea polyphenols,possesses a potent antioxidant capacity.Epidemiological investigation shows that green tea is a protective factor of Parkinson's disease(PD).The pathogenesis of PD not yet controversial,studies indicated that oxidative stress in substantia nigra compacta may have contributed to the onset and development of PD.The nuclear factor erythroid-2p45 related factor 2/antioxidant response element signal pathway(Nrf2/ARE pathway)is critical in regulating the redox level in biological mechanism,while the redox sensitive transcription factor Nrf2 is a core regulator against oxidative stress.In this study,we investigated the protective effects of EGCG's on neurological impairments among PD model mice in vitro and in vivo and explored the inherent mechanism of Nrf2/ARE pathway.Method:1.Human neuroblastoma cells(SH-SY5Y)were incubated with rotenone to prepared as the PD cellular model and incubated with various dosage of EGCG(10mmol/L,20 mmol/L,40 mmol/L)for 24 h.The CCK-8 and LDH assays were used to measure cell viability and membrane integrity.The effects of EGCG on cell apoptosis were measured by caspase-3 activity and Bax expression level of cells.Biochemical indexes of oxidative stress such as SOD,MDA,ROS,GPx were assessed with kits.The expression of tyrosine hydroxylase(TH)in rotenone or/and EGCG treated SH-SY5 Y cells was determined using western blot analysis.2.si RNA is used to knock down the Nrf2 expression in cells,and the interfering efficiency was detected by RT-PCR.If the Nrf2 mRNA expression significantly different between the si-control and siNrf2 group,the si Nrf2 group would be treated by rotenone or/and EGCG,followed by measured cell viability and ROS production measurement.3.The PD mouse model was randomly divided into groups: the positive control group,EGCG high(80 mg/kg),middle(60 mg/kg),and low(40 mg/kg)group,drugs were chronically administered to animals at 9:00 AM for 21 days,i.g.,and weighing the body weight every 7 days.Animals were tested the stick climbing and the exercise ability at day 21.Result:1.Compared with the PD cell model group,all EGCG-treated groups cell survivalrate increased(p<0.01),and reduced the cell LDH spillage rate(p<0.01).The group of 20mmol/L EGCG significantly reduced the relative activity of Caspsae 3 and up regulate the Bax expression(p<0.05).Both 10 mmol/L and 20 mmol/L EGCG groups upregulated the TH expression(p<0.05),whilst 40 mmol/L EGCG had no significant effect(p>0.05).2.Compared with the PD cell model group,the GPx activity of all EGCG groups were increased significantly(p<0.01),MDA and ROS decreased significantly(p<0.05),and the SOD units of the 20 mmol/L and 40 mmol/L EGCG groups increased significantly(p<0.05).EGCG treatment could reduce the rotenone-induced neurotoxicity.3.Compared with the control group,EGCG could upregulated Nrf2 protein expression(p<0.05).Compared with the rotenone model group,there was no significant difference on cell viability and ROS in the siNrf2 EGCG treatment groups(p>0.05).4.Compared with the control group,the exercise ability of the model group decreased significantly(p<0.05).Compared with the rotenone model group,the exercise ability of the positive group,the high and middle dose EGCG groups increased significantly(p<0.05),but for low dose group,there was no statistical significance(p>0.05).Conclusion:EGCG protects neuron against rotenone-iduced Parkinsonism-like neurotoxicity in SH-SY5 Y cells and improve the sports performance in mice via activation of Nrf2/ARE signaling pathway.