Exploring The Mechanism Of Ginsenoside Rg1 On Myocardial Fibrosis Based On The Pyroptotic NLRP3/Caspase-1 Signaling Pathway

ObjectiveIn this study,myocardial fibrosis model in mice was constructed by using isoproterenol(ISO).We observed the effect of ginsenoside Rg1 on myocardial fibrosis and cell pyroptosis in myocardial tissue.cardiac fibroblasts were extracted,which aims to explore the effect of ginsenoside Rg1 on pyroptosis and the NLRP3/Caspase-1 pathway of cell pyroptosis in CFs.These can provide a new therapeutic target for ginsenoside Rg1 in the treatment of myocardial fibrosis.Methods1.In vivo experiment:C57BL/6 mice were randomly divided into control group(Control group),model group(ISO group)and ginsenoside Rg1 treatment group(Rg1 group).The myocardial fibrosis model was constructed by ISO,and the Rg1 group was given ginsenoside Rg1 intervention.HE staining,Masson staining and Sirius Red staining were used to observe the pathological changes of myocardial tissue and the degree of myocardial fibrosis;ELISA was used to detect the level of IL-1β in serum;Western blot and qRT-PCR was used to detect the protein and mRNA expression of Col-I,α-SMA,NLRP3,Caspase-1,and GSDMD in myocardial tissue.2.In vitro experiments:cardiac fibroblasts were extracted from C57BL/6 neonatal mice,and the myocardial fibroblasts were identified by immunofluorescence labeling of α-SMA after stimulation with TGF-β1.CCK-8 detected the effect of different concentrations of ginsenoside Rg1 on the survival rate of cardiac fibroblasts to determine the appropriate concentration of ginsenoside Rg1 for subsequent cell experiments.CFs were divided into control group(Control group),model group(Model group),ginsenoside Rg1 intervention group(Rg1 group)and NLRP3 inhibitor group(MCC950 group).Model group used LPS and ATP to induce the pyroptosis of CFs,Rg1 group added ginsenoside Rg1 for intervention before modeling,and MCC950 group added MCC950 before modeling.ELISA was used to detect the level of IL-1β in the supernatant of CFs cells;Western blot was used to detect the protein and mRNA expression of Col-I,α-SMA,NLRP3,Caspase-1,and GSDMD in each group.Results1.In vivo experiment:Compared with the Control group,the deposition of collagen fibers and the activation of myocardial fibroblasts in myocardial tissue of C57BL/6 mice which suffered myocardial fibrosis was significantly induced in the ISO group(P<0.05).The levels of serum IL-1β was obviously decreased(P<0.05).The protein and mRNA expressions of Col-I and α-SMA were significantly increased(P<0.05);meanwhile,the protein and mRNA expressions of pyroptosis-related proteins NLRP3,Caspase-1 and GSDMD were significantly increased(P<0.05).Compared with the ISO group,the collagen deposition and the activation of myocardial fibroblasts in the myocardial tissue of the mice in Rg1 group was significantly reduced,the protein and mRNA expressions of Col-I and α-SMA were significantly reduced(P<0.05),the levels of IL-1β in serum was significantly decreased(P<0.05),and the protein and mRNA expressions of pyroptosis-related proteins NLRP3,Caspase-1,GSDMD were significantly decreased(P<0.05).2.In vitro experiments:Compared with the Control group,the protein expressions of Col-I,α-SMA,NLRP3,Caspase-1 and GSDMD in CFs cells were significantly increased(P<0.05),and the level of IL-1β in the cell supernatant was significantly increased(P<0.05).Compared with the Model group,the expressions of Col-I,α-SMA,NLRP3,Caspase-1 and GSDMD proteins in the cells of the Rg1 group were significantly decreased(P<0.05),and the level of IL1β in the cell supernatant was significantly decreased(P<0.05).Compared with the Model group,the protein expressions of Col-I,α-SMA,NLRP3,Caspase-1 and GSDMD in the CFs cells of the MCC950 group were significantly decreased(P<0.05),and the level of IL-1β in the supernatant of the cells was significantly decreased(P<0.05).Conclusion1.Ginsenoside Rg1 significantly improves ISO-induced myocardial fibrosis and reduces collagen deposition in myocardial tissue;2.Ginsenoside Rg1 attenuates the level of inflammation and the expression of NLRP3/Caspase-1 pathway which related to pyroptosis in mice’s myocardial tissue with ISOinduced myocardial fibrosis.3.Inhibition of NLRP3 inflammasome-mediated pyroptosis of CFs can reduce the activation of CFs and the synthesis of Col-I;4.Ginsenoside Rg1 inhibits CFs pyroptosis,and reduces CFs activation and Col-I synthesis,and its mechanism may be related to the down-regulation of NLRP3/Caspase-1 signaling pathway.