Study On The Mechanism Of Prevention And Treatment Of Hypertensive Myocardial Fibrosis By Tianma Gouteng Decoction Based On MiR-150-3p Regulation Of Wnt/β-catenin Pathway

Objective: To study the mechanism of miR-150-3p regulating Wnt/β-catenin pathway and the mechanism of Tianma Gouteng Decoction in hypertensive myocardial fibrosis.To find new targets for prevention and treatment of traditional Chinese medicine in hypertensive myocardial fibrosis.Methods Part Ⅰ: We used Bioinformatics methods to search for miRNA datasets related to hypertensive myocardial fibrosis in the GEO database to screen differentially expressed miRNAs in hypertensive myocardial fibrosis.The miR-150-3p target genes were predicted,and their signal pathway analysis was performed.Part Ⅱ: Clinically verified the expression of serum miR-150-3p in patients with hypertensive myocardial fibrosis.We collected serum from hypertensive patients and healthy volunteers who met the inclusion criteria for miR-150-3p expression detection by RT-PCR,and then compared the expression of miR-150-3p in hypertensive myocardial fibrosis patients and healthy volunteers.Part III: Research on the production of hypertensive myocardial fibrosis machine.To elucidate the mechanism by which miR-150-3p regulates Wnt/β-catenin signaling pathway in hypertensive myocardial fibrosis,we transfected miR-150-3p mimics and inhibitors into cardiac fibroblasts by transient transfection.Used RT-PCR assay for miR-150-3p expression,CCK8 assay for cell proliferation,ELISA assay to detect the expression of type I and III collagen and α-SMA protein in cell culture supernatant,and the expression of β-catenin was detected by western-blot method.Part Ⅳ: The mechanism of Tianma Gouteng Decoction in preventing and treating hypertensive myocardial fibrosis.Myocardial fibrosis model was induced by TGF-β1 stimulation of cardiac fibroblasts,and SD rats were gavage to prepare drug-containing serum,and the myocardial fibrosis model was intervened in groups.To investigate the effects of serum containing Tianma Gouteng Decoction on hypertensive myocardial fibrosis and wnt/β-catenin pathway.We detected the expression of miR-150-3p by RT-PCR;the cell proliferation was detected by CCK8 method,the expression of type I and III collagen and α-SMA in cell culture supernatant were detected by ELISA,and β-catenin expression was detected by western blot.Results 1.Bioinformatics analysis: miR-150-3p is differentially expressed in serum of patients with hypertension or chronic heart failure combined with myocardial fibrosis.The KEGG pathway enrichment analysis of the target genes showed the closely relationship between those and the Wnt/β-catenin pathway.2.Compared with healthy controls,the expression of serum collagen I,III and α-SMA was significantly higher in patients with hypertensive left ventricular hypertrophy(P<0.05).While the expression of serum miR-150-3p in patients with hypertensive myocardial fibrosis was down- regulated(P<0.05).3.After miR-150-3p mimics and inhibitors transfection,the inhibitory group showed higher levels of cell proliferation and increased expression of I,III collagen and α-SMA protein than the control group and the mock group.At the same time,the expression of β-catenin was increased,and the results were statistically significant(P<0.05).4.Compared with the control group,the expression of miR-150-3p in the model group was decreased,the proliferation of cardiac fibroblasts,and the secretion of I,III collagen α-SMA,β-catenin increased.The results were statistically significant(P< 0.05).Compared with the model+SD group,the expression of miR-150-3p was increased in the model+Chinese medicine group,the secretion of I,III collagen and α-SMA protein was decreased,while the expression of β-catenin was down-regulated.The results were statistically significant(P< 0.05).Conclusion: 1.miR-150-3p was down-regulated in patients with hypertensive myocardial fibrosis 2.The inhibition of miR-150-3p could up-regulate β-catenin expression leading to activation of cardiac fibroblasts;3.The drug-containing serum of Tianma Gouteng Decoction might inhibit hypertensive myocardial fibrosis by up-regulating miR-150-3p to inhibit Wnt/β-catenin pathway.