Effect Of Bushen Huoxue Fang On α-syn Aggregation In PD Model Mice Induced By MPTP

Objective 1.To investigate the effect of Bushen Huoxue Fang on inhibiting 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)-induced α-synuclein(α-syn)The mechanism of aggregation.2.Study the potential action targets and related pathways of Bushen Huoxue Decoction in the treatment of PD using network pharmacological methods,and explore the possible mechanism of Bushen Huoxue Decoction on cell autophagy.Methods 1.Thirty-six male C57/BL mice were randomly divided into a blank group,a model group,and a traditional Chinese medicine group,with 12 in each group.A mouse model of subacute PD was prepared by intraperitoneal injection of MPTP.After successful modeling,the traditional Chinese medicine group was given Bushen Huoxue Fang for 14 consecutive days.The blank group and the model group were only given the same amount of normal saline.The immunohistochemical method was used to detect the positive expression of tyrosine hydroxylase(TH)and α-syn in the brain substantia of each group of mice,and the expression of Lamp2a,Hsc70,α-syn in the brain substantia of each group was detected by WB.2.In the TCMSP database,the oral active ingredients of Bushen Huoxue Fang were retrieved with oral availability(OB)≥30%and drug-like properties(DL)≥0.18,and the corresponding targets of each active ingredient were retrieved.Search GeneCards and OMIM databases for disease-related targets in PD.By intersecting the target of Bushen Huoxue Recipe with the PD target,you can obtain the potential active ingredients and action targets of Bushen Huoxue Recipe for the treatment of PD.The"component-target" network diagram was constructed using Cytoscape software,and the results were visualized.The effect target of Bushen Huoxue Decoction for PD was input into STRING data,and the protein interaction network(PPI)of key targets was obtained.The network was analyzed to obtain highly connected targets,which was defined as the key to Bushen Huoxue Decoction Target.The R software was used to analyze GO and KEGG of the target of PD treated by Bushen Huoxue Fang to obtain biological processes and cellular molecular pathways related to the target.Finally,the BATMAN database was used to explore the mechanism of the effect of Bushen Huoxue Fang on cell autophagy.The threshold value of the "drug-target" similarity model was set to Score cutoff≥20 and P≤0.05.The system could obtain its possible effect by scoring each active ingredient The target point is to analyze the physiological processes and diseases that may be affected by Bushen Huoxue Fang through analysis,and retrieve the results related to PD and cell autophagy in the analysis results.Results 1.Compared with the blank group,the mice in the model group showed obvious PD syndrome characteristics,and their behavioral scores were significantly increased(p<0.01).Compared with the model group,the PD symptoms of the mice in the Chinese medicine group were alleviated,and their behavioral scores were significantly reduced.(P<0.01).Immunohistochemistry showed that the positive expressions of TH and α-syn in the model group were significantly lower than those in the blank group(p<0.01);the positive expressions of TH and α-syn in the Chinese medicine group were significantly higher than those in the model group(p<0.01).WBt showed that compared with the blank group,the expression levels of Lamp2A,Hsc70,and α-syn in the model group were significantly increased(p<0.01),and the expression levels of TH in the model group were significantly reduced(p<0.01).Compared with the model group,the expression levels of Lamp2A,Hsc70,and TH in Chinese medicine group were significantly increased(p<0.01),and the expression levels of a-syn were significantly decreased(p<0.01).2.After searching the TCMSP database,a total of 133 oral active ingredients of Bushen Huoxue Fang were obtained,of which 65 active ingredients had potential effects in the treatment of PD.Take the intersection to obtain the 87 active targets of Bushen Huoxue Fang for PD,and enter them into the STRING database to obtain the PPI network.Among them,MAPK8,CCND1,EGFR,CASP3,IL6,MYC,VEGFA,ESR1 and other targets have high connectivity and are PPI The target that plays a central role in the network is the key target for Bushen Huoxue Fang to treat PD.According to GO and KEGG enrichment analysis,it was found that Bushen Huoxue Fang may exert its therapeutic effect on PD through biological processes such as neurotransmitter receptor active nerve and neurotransmitter binding.KEGG enrichment analysis revealed that the PI3K-Akt signaling pathway is a potential pathway for Bushen Huoxue Recipe to treat PD.According to BATMAN data analysis,Bushen Huoxue Fang may play a therapeutic role in PD through 16 targets such as ACHE,ADORA2A,UCHL1,and may affect cell autophagy through 14 targets such as LRRK2,MTOR,and PARK2.Conclusion 1.Bushen Huoxue Fang may inhibit the accumulation of a-syn in the substantia nigra of PD mice by increasing the levels of Lamp2a and Hsc70,and then enhance the molecular chaperone-mediated autophagy,thereby inhibiting the dopaminergic neuron apoptosis.2.TCMSP-based network pharmacological analysis found that Bushen Huoxue Fang may inhibit the apoptosis of DA neurons by affecting the PI3K/Akt pathway,thereby exerting a therapeutic effect on PD.Network pharmacological analysis based on BATMAN-TCM found that Bushen Huoxue Fang may affect CMA by acting on UCHL1,thereby inhibiting the abnormal aggregation of α-syn and protecting DA neurons and inhibiting the development of PD.