Acupuncture DU20 Through GB7 Regulates Neurocyte Autophagy In A Rat Model Of Hemorrhagic Stroke Based On MTOR Signaling Pathway

Object:Explore the effect on acupuncture Baihui and Qubin autophagy and mTOR signaling pathway in hemorrhagic stroke rat.Method:Divided 126 healthy male Sprague-Dawley rats(300±10g)into 7 groups randomly including sham operation group(sham group),hemorrhagic stroke model group(model group),model+acupuncture group(acupuncture group),3-M A group,3-M A+acupuncture group,rapamycin group and rapamycin+acupuncture group.Each group was divided into 3 subgroups according to the time points(1d,3d,7d),each subgroup have 6 rats.According to the Stereotaxic Atlas of rats,injecting 50 ?L autologous blood into rat caudate nucleus,intervention respectively by acupuncture Baihui and Qubin in the affected side of rat,3-MA(ivc),and rapamycin(ivc),score rat in neurological deficit(Complex neurological function score,Forelimb placing test,Suspension test,Balanced walking test)at each time point,at the end of the score take brain tissue testing the protein expression changes on autophagy related protein LC3A/B,p62 and mTOR signaling related protein p-mTOR,p-S6K.Result:1.Acupuncture Baihui,Qubin could alleviate the neurological deficits after hemorrhagic stroke in rats(vs.Model group,p<0.01);3-MA group can aggravate neurological deficits in rats(vs.Model group,p<0.01),3-MA+acupuncture group has no significant improvement of neurological deficits in rats(vs.Model group,p>0.01);rapamycin group and rapamycin+acupuncture group can improve the neurological function score of cerebral hemorrhage rats(vs.Model group,p<0.01).2.Immunohistochemical staining and Western blot analysis of LC3A/B protein in rats brain tissue with intracerebral hemorrhage showed that the expression of LC3A/B protein in rats brain tissue with 1,3,7 days of intracerebral hemorrhage was positive,there was a significant increase in LC3A/B protein expression(vs.Sham group,p<0.05).Compared with model group,acupuncture group can increase the expression of LC3A/B protein in 3,7 days(p<0.05),the difference was statistically significant.The expression of LC3A/B protein was significantly down regulated by autophagy inhibitor 3-MA group,at the same time,acupuncture could increase the protein expression in 3,7 days(p<0.05),the difference was statistically significant.The autophagy agonist rapamycin group can increase the content of LC3A/B protein at different time points,at the same time,the intervention of acupuncture could decrease LC3A/B protein content in 3,7 days(p<0.05),and the difference was statistically significant.3.Western blot analysis of p62 protein in rats brain tissue with intracerebral hemorrhage showed that the expression of p62 protein in model group was significantly increased at 1,3,7 days(vs.sham group,p<0.05).Compared with model group,the expression of p62 protein was decreased in acupuncture group(p<0.05),and the difference was statistically significant.Autophagy inhibitor 3-MA group can significantly improve the expression of p62 protein,while acupuncture intervention can regulate the expression of p62 protein in 1,3 days(p<0.05),the difference was statistically significant.The autophagy agonist rapamycin group at all time points can be regulated p62 protein content down,while the intervention of acupuncture can significantly increase the p62 protein content at 1,3 days(p<0.05),the difference was statistically significant.4.The results of p-mTOR and p-S6K1 protein expression in rats brain tissue in each group showed that,in model group,the expression of p-mTOR and p-S6K1 protein increased significantly in 1,3,7 days(vs.Sham group,p<0.05).Compared with model group,acupuncture group could regulated the expression down of p-mTOR and p-S6K1(p<0.05),the difference was statistically significant.Rapamycin group at all time points can be reduced p-mTOR,p-S6K1 protein content,while the intervention of acupuncture can increase the expression of p-mTOR,p-S6K1 protein,the difference was statistically significant.Conclusion:1.Acupuncture Baihui and Qubin could alleviate neurological deficit with intracerebral hemorrhage rats.2.Acupuncture Baihui and Qubin can promote the expression of autophagy related protein LC3A/B and p62 after hemorrhagic stroke in rats;3.Acupuncture Baihui and Qubin restrain the expression of p-mTOR and p-S6K 1 protein in mTOR signaling pathway;4.Acupuncture Baihui and Qubin may promote levels of autophagy after hemorrhagic stroke,and then improve nerve function impairment after hemorrhagic stroke in rats through regulation of mTOR signaling pathway.