电针抑制PI3K/Akt/mTOR通路促进分子伴侣介导的自噬保护帕金森病研究OA

Objective To investigate the regulatory mechanism of electroacupuncture(EA)at"Baihui"(GV20)and"Shenshu"(BL23)on chaperone-mediated autophagy(CMA)in a mouse model of Parkinson's disease(PD).Methods Twenty-seven C57BL/6 mice were randomly assigned to three groups:the control group,the model group,and the EA group(n=9 per group).PD models were established in the model group and the EA group by intraperitoneal injection of MPTP.Mice in the EA group received EA treatment at GV20 and BL23(continuous wave,2 Hz,2 mA,15 min/day)for 14 con-secutive days.Motor performance was evaluated using the rotarod test,suspension test,pole test,and gait analysis.Tyrosine hydroxylase(TH)immunofluorescence and double staining for neuronal nuclei(NeuN)with lysosome-associated membrane protein 2A(LAMP2A)or heat shock cognate protein 70(HSC70)were used to assess dopaminergic neuronal survival and CMA activity in the substantia nigra.The method of western blotting was performed to determine the protein levels of TH,α-synuclein(α-syn),phosphatidylinositol 3-kinases(PI3K),protein kinase B(Akt),p-Akt,mammalian target of rapamycin(mTOR),p-mTOR,LAMP2A,HSC70,glial fibrillary acidic protein(GFAP),and transcription factor EB(TFEB)in the substantia nigra,while RT-qPCR was used to detect mRNA expression of PI3K,Akt,and mTOR.Results Compared with the control group,the model group showed a significantly decreased rotarod latency(P＜0.01),decreased suspension test scores(P＜0.01),prolonged pole-climbing time(P＜0.01),and decreased stride length and increased step frequency(P＜0.01).TH fluorescence intensity in the substantia nigra was markedly reduced(P＜0.01),and the colocalization of NeuN with LAMP2A or HSC70 was decreased(P＜0.05,P＜0.01).The expression of PI3K,p-Akt/Akt,p-mTOR/mTOR,α-syn,and GFAP were increased(P＜0.05,P＜0.01,P＜0.05,P＜0.05,P＜0.01),while TFEB expression were decreased(P＜0.01);mRNA levels of PI3K,Akt,and mTOR were elevated(P＜0.01).Compared with the model group,EA significantly prolonged rotarod latency(P＜0.01),increased suspension test scores(P＜0.05),shortened pole-climbing time(P＜0.01),and improved gait coordination(P＜0.05).EA increased TH-positive neuronal fluorescence intensity(P＜0.01),enhanced NeuN colocalization with LAMP2A and HSC70(P＜0.05,P＜0.01),reduced PI3K,p-Akt/Akt,p-mTOR/mTOR,α-syn and GFAP expression(P＜0.05,P＜0.01,P＜0.01,P＜0.05,P＜0.05)and up-regulated TFEB(P＜0.01).Moreover,EA decreased mRNA levels of PI3K,Akt,and mTOR(P＜0.01,P＜0.01,P＜0.05).Conclusion EA may exert neuroprotective effects against PD by modulating the PI3K/Akt/mTOR signaling pathway to activate neuron-specific CMA,thereby promoting α-syn clearance and alleviating neuroinflammation.