Inhibitory Effect Of MicroRNA-200b Down-regulation Of DNMT3A Expression On Myocardial Fibrosis In SD Rats

Objective:Atrial fibrillation(AF)is a relatively common,chronic and persistent type of arrhythmia with high-risk and high-risk characteristics.From the perspective of treatment and prognosis,the current treatment of atrial fibrillation still has some recurrence.The rate of stroke mortality caused by atrial fibrillation is as high as 25%.Studies have shown that the mechanism of occurrence and development of atrial fibrillation mainly includes electrical remodeling and mechanical remodeling.Electrical remodeling generally shows changes in ion channels,while structural remodeling is characterized by myocardial fibrosis.DNA methylation refers to the process by which an organism transfers a methyl group to a specific base under the catalysis of DNA methyltransferase,thereby causing phenomena such as embryo differentiation,gene inactivation,and organ aging in organisms..At present,more and more literature studies have shown that DNMT3 A,which is one of the important members of DNA methyltransferase,has an important influence on the development of cardiac fibroblasts in myocardial fibrosis.Micro RNAs(mi RNAs or mi Rs)are a small class of non-coding RNAs that regulate the expression of target genes at the transcriptional level.Previous studies have shown that mi R-200 b can participate in pathophysiological processes such as cell proliferation,apoptosis,carcinogenesis,and fibrosis by regulating the corresponding target genes.It has been reported in the literature that the expression of DNMT3 A regulated by mi R-200 b affects the development of non-small cell lung cancer.However,whether the effect of DNMT3 A on the development of myocardial fibrosis is also regulated by mi R-200 b is still unclear.Therefore,this study provides a theoretical basis for the exploration of myocardial fibrosis by observing the expression of mi R-200 b and its potential target gene DNMT3 A in myocardial fibrosis in cardiac fibroblasts and its regulatory relationship in myocardial fibrosis.Methods:In vivo experiments: 60 male SD rats of 8 weeks old were randomly divided into model group and control group.Rats in the model group were given subcutaneous injection of the inducer ISO once/d [5 mg/(kg.d)],and the control group was given an equal dose of physiological saline for 7 days.After successful modeling,the animals were sacrificed and the cardiac specimens were taken out.HE staining and Masson staining were used to observe the degree of myocardial fibrosis and collagen volume fraction(CVF)in each group.The myocardial tissue Col1A1 was detected by immunohistochemistry.?-SMA and DNMT3 A expression;q RT-PCR detection of mi R-200 b expression;Western blot analysis of ?-SMA,Col1A1,DNMT3 A expression.In vitro experiment: Cultured SD milk rat cardiac fibroblasts.mi R-200 b promoters(inhibitors)and inhibitors were transfected into CFs,respectively,and negative and blank controls were established.The expression of RNMT3 A,Col1A1 and ?-SMA in CFs transfected with different reagents was detected by q RT-PCR.The proliferation of CFs after dyeing.Results:In vivo experiments: HE staining and Masson staining showed significant collagen fibrosis in the interstitial of SD rats in the experimental group.The myocardial interstitial collagen volume fraction(CVF)in the experimental group increased significantly.The results of immunohistochemistry showed that the protein expression levels of Col1A1,?-SMA and DNMT3 A in the model group were significantly higher than those in the control group.The results of q RT-PCR showed that the expression level of mi RNA-200 b in the total RNA extracted from the model group was significantly lower than that in the normal group.Western blot analysis showed that the levels of ?-SMA,Col1A1 and DNMT3 A in the model group were significantly higher than those in the control group.In vitro experiments: q RT-PCR results showed that the expression of ?-SMA,Col1A1,and DNMT3 A in the CFs of the mi R-200 b promoter group was significantly lower than that of the normal group and the negative control group,while in the CFs of the mi R-200 b inhibitor group.on the contrary.Western blot results showed that the expression of DNMT3 A,Col1A1,and ?-SMA protein was decreased in the promoter group,but increased in the inhibitor group.MTT results indicated that mi R-200 b inhibited the growth of CFs.Conclusion:In summary,mi R-200 b may affect the methylation of CFs by inhibiting DNMT3 A,control the activation of CFs,and reduce the level of myocardial fibrosis.DNMT3 A may be one of the epigenetic therapeutic targets for myocardial fibrosis,and it also provides new ideas for the prevention and study of myocardial fibrosis.