Study On Protective Effects And Mechanisms Of Ginsenoside Re On β-Amyloid Induced Neuronal Injury

Objective:To investigate the effect of ginsenoside Re on the damage of SH-SY5Y cells induced by Aβ_25-35,and to explore the protective effect of ginsenoside Re on neurons and its possible mechanism.Methods:1.SH-SY5Y cells were treated with Aβ_25-355-35 and ginsenoside Re.Cell viability was detected by CCK-8 method,and the concentration,duration of action and concentration of ginsenoside Re in the subsequent experiment were determined.2.Each group of SH-SY5Y cells were treated with Aβ_25-355-35 and ginsenoside Re,flow cytometry was used to detect the apoptosis rate;Detection of Caspase activity by Caspase activity assay kit.3.Flow cytometry was used to determine intracellular ROS content and mitochondrial membrane potential;UV spectrophotometry was used to detect ATP content.The cytoplasm and mitochondria of SH-SY5Y cells were isolated,and the expression of cytochrome c,Bcl-2 and Bax protein were detected by Western blotting.4.Each group of SH-SY5Y cells were treated with Aβ_25-355-35 and ginsenoside Re,the nucleus and cytoplasm were separated,the expression level of target protein and nuclear transfer of Nrf2 cells were detected by Western blotting;total RNA was extracted by TRIzol method;Detection of changes in the expression level of the target gene at the transcription level.Results:1.Ginsenoside Re has a significant protective effect on Aβ_25-355-35 induced cytotoxicity.The most effective concentration is 25μM（P<0.0001）.At the same time,flow cytometry results suggest that it can significantly inhibit Aβ_25-355-35 induced apoptosis（P<0.0001）.2.Ginsenoside Re significantly inhibited the increase of Caspase-3 and Caspase-9activities induced by Aβ_25-35（P<0.01,P<0.001）,but had no significant effect on Caspase-8and Caspase-12 activities.3.Ginsenoside Re exerts cytoprotection by inhibiting mitochondrial apoptosis pathway:25μM ginsenoside Re significantly inhibits MMP reduction in SH-SY5Y cells induced by Aβ_25-35（P<0.0001）,Aβ_25-355-35 induced decrease in ATP production in SH-SY5Y cells was significantly attenuated（P<0.0001）;Ginsenoside Re attenuated Aβ_25-355-35 induced Bax protein expression compared to Aβ_25-355-35 induced SH-SY5Y cells up,significantly increased the proportion of Bcl-2/Bax protein expression（P<0.01）,effectively preventing the release of cytochrome c from mitochondria into cytoplasm（P<0.05）.4.Ginsenoside Re inhibits ASK1/JNK signaling pathway in a ROS-dependent manner:25μM ginsenoside Re can significantly attenuate the phosphorylation of ASK1 and JNK induced by Aβ_25-35（P<0.05）;Ginsenoside Re intervention it can significantly inhibit the increase of ROS level induced by Aβ_25-355-35 induction,and it shows a significant concentration-dependent;After DPI intervention,the effect of ginsenoside Re on ASK1/JNK signaling pathway is invalid.5.Inhibition of ROS-dependent ASK1/JNK signaling pathway attenuated the neuroprotective effect of ginsenoside Re:After DPI or SP600125 intervention,the neuroprotective effect of ginsenoside Re was significantly eliminated（P<0.01）.Meanwhile,the inhibitory effect of ginsenoside Re on the expression of Bax protein induced by Aβ_25-35disappeared（P<0.05）.6.Ginsenoside Re slows down the oxidative stress induced by Aβ_25-355-35 by activating Nrf2 antioxidant pathway:25μM ginsenoside Re treatment significantly increased the level of Nrf2 protein in the nucleus,accompanied by a decrease in cytoplasm（P<0.01,P<0.05）,thereby increasing the expression of the antioxidant enzyme gene（P<0.05）.Conclusion:1.Ginsenoside Re has protective effect on Aβ_25-355-35 induced SH-SY5Y cytotoxicity.2.Ginsenoside Re can significantly alleviate mitochondrial dysfunction induced by Aβ_25-355-35 in SH-SY5Y cells,and play its role by inhibiting mitochondria-mediated apoptosis pathway.3.Ginsenoside Re inhibits Aβ_25-355-35 induced ASK1/JNK signaling pathway through ROS-dependent manner and exerts neuroprotective effects.4.Ginsenoside Re reduces the oxidative stress induced by Aβ_25-355-35 by activating the Nrf2 antioxidant signaling pathway.