Regulatory Effect And Mechanism Research Of Warming Yang-Benefiting Heart Therapy On Mirna Expression Of Atherosclerosis

Objective: 1.To screen differential expression of atherosclerosis(AS)mi RNA and confirm the regulatory effect of Warming Yang-Benefiting Heart Therapy on mi RNA expression of atherosclerosis;2.To explore the pathway of Warming Yang-Benefiting Heart Therapy and verify the therapeutic mechanism,and provide experimental support for the further study.Methods: 1.AS Model: This experiment used C57BL/6J male mice as the experimental model by vitamin D3 abdominal injection,lard and high fat feeding methods.Model evaluation was conducted at week 9th.Half mice before the module at week 10 th were randomly divided into 5 groups(model group,low-dose decoction group,medium-dose decoction group,high-dose decoction group and western medicine group),with pure water,Warming-Heart Decoction(low,medium and high dose),western medicine(aspirin and lipitor)respectively.Six weeks after administration,all mice were killed and the blood taken from the eyeball;2.mi RNA gene sequencing and Expression Verification: Different genes were screened through high-throughput sequencing of gene chips,using QT-PCR technology to verify the expression of key genes.Enter mi RNA into Targetscan and Miranda gene databases for target gene and pathway prediction;3.Network Pharmacology Analysis:Retrieving the TCMSP database retrieval compounds(OB ? 30%,DL ? 0.18),The target protein information is imported into the Uniprot site for genoname conversion.Import integrated data into Cytoscape 3.5.1 software to build Drug-Compounds-Target visual network diagram.Log in the Dis Ge NET,Genecard,Drug Bank,OMIM,GAD,TTD database to retrieve the arteriosclerosis target gene.Input the intersection target of disease and drug into the String website and Cytoscape3.5.1 to build the PPI network,import the target gene into the Metascape website for GO and KEGG enrichment analysis,and predict the pathway;Proteins(IGF1R,ERK2,PIK3 CA,PRKCA,PRKCA,VEGFA)on the key pathway(PI3K-AKT)were tested by the Western Blot experiment.Results: Blank group mice are generally in good condition,with smooth blood vessels without lesion formation.The model group showed obvious arteriosclerosis and scattered plaque formation.HE staining showed significant AS lesions,no improvement in low-dose Warming-Heart Decoction group,but obvious in other groups.The difference in blood lipids level was not significant(P > 0.05).In this study,30 "lesion" mi RNA genes were selected through high-throughput sequencing,and Warming Yang-Benefiting Heart Therapy effectively regulated 18 of these genes.Three genes(mi R-6984-5p,mi R-497-5p,mi R-187-3p)were confirmed by QT-PCR.Network pharmacology study selected 3,127 disease target genes and 196 cardiac targets.?-glusterol,legosterol,berberine,and quercetin are the highest rated warm-square median compounds.It may be regulated by PI3K-Akt,NF-B,HIF-1.Western Blot experiment showed that the Warming Yang-Benefiting Heart Therapy regulated the expression of IGF1 R,ERK2,PIK3 CA and PRKCA protein on PI3K-Akt pathway,which was statissignificant for regulation of IGF1 R and PRKCA protein(P<0.05).Conclusions: 1.Warming Yang-Benefiting Heart Therapy can effectively improve the condition of arterial lesions in AS mice;2.The method can improve the expression of some AS "lesions" mi RNA;3,?-glusterol,legosterol and berberine may play a key role,while PI3K-AKT may be a key pathway;4.Warming Yang-Benefiting Heart Therapy can regulate the expression of IGF1 R and PRKCA on PI3K-AKT pathway.