The Role Of MiR-223 In The Pyroptosis Of Myocardium In Mice With Viral Myocarditis Induced By CVB3

IVObjective:1.By establishing the mice model of viral myocarditis induced by CVB3,detecting the molecules associated with pyroptosis in the mice myocardium,and understanding the level of the molecules associated with pyroptosis in the myocardium of mice with viral myocarditis;2.By detecting the level of mi R-223 in the myocardium of mice with viral myocarditis induced by CVB3,the role of mi R-223 in the pyroptosis of myocardial in mice with viral myocarditis induced by CVB3 was explored preliminarily.Methods:1.Establishment of the mice model of viral myocarditis induced by CVB31.1 Thirty BALB/c mice were selected and randomly divided into the control group(Normal group)and the viral myocarditis group(CVB3 group),each with 15 mice.In the CVB3 group,the mice were injected intraperitoneally with CVB3 to establish the mice model of viral myocarditis induced by CVB3.The control group was intraperitoneally injected with the same amount of phosphate buffer saline.The general condition of mice was observed every day,whether there were shrink,activity reduction,clustering,insensitive to stimulation and other symptoms.The changes of body weight and survival status of mice was recorded for7 days.1.2 After 7 days of intraperitoneal injection of CVB3,blood was collected from the intraocular canthus,and the levels of c Tn I in the serum of mice in each group of mice was detected by ELISA.1.3 After 7 days of intraperitoneal injection of CVB3,the mice were sacrificed to extract myocardial tissue samples,which were routinely fixed with 4% paraformaldehyde,dehydrated,and paraffin-embedded,then sectioned,and HE stained to observe the pathological changes of myocardial tissue in each group.The changes of body weight,survival rate,the levels of c Tn I and myocardial histopathology of mice were evaluated to judge whether the mice model of viral myocarditis induced by CVB3 was successfully established.2.Molecular detection of pyroptosis in mice with viral myocarditis induced by CVB3Blood from inner canthus was detected for ELISA,and myocardial tissue preserved with liquid nitrogen was detected for q RT-PCR and Western blot.2.1 After 7 days of intraperitoneal injection of CVB3,the q RT-PCR was used to detect the expression levels of NLRP3 and caspase-1m RNA in the myocardium of the control group and CVB3 group of mice.2.2 After 7 days of intraperitoneal injection of CVB3,the Western blot was used to detect the changes in the ratio of caspase-1/pro-caspase-1 in the myocardium of the control group and the CVB3 group.2.3 After 7 days of intraperitoneal injection of CVB3,the ELISA was used to detect the levels of IL-1β in the serum of mice in the control group and the CVB3 group.3.Detection of mi R-223 in mice with viral myocarditis induced by CVB3After 7 days of intraperitoneal injection of CVB3,the q RT-PCR method was used to detect the expression level of mi R-223 in the myocardium of mice in the control group and the CVB3 group.Results:1.Establishment of the mice model of viral myocarditis induced by CVB31.1 The mice in the CVB3 group began to show symptoms such as shrink,activity reduction,clustering and insensitive to stimulation on the 3rd day;the weight of the mice in the control group gradually increased from the 1st day;the weight of the mice in the CVB3 group decreased from the 3rd day and continued until the 7th day.1.2 Survival of mice within 7 days after infection with CVB3: the survival rate of mice in the control group was 100%,and the mice in the CVB3 group died from the 2nd day,and the survival rate was 47% by the 7th day.1.3 Detection of the levels of c Tn I in the serum of mice in each group by ELISA showed that the levels of c Tn I in the serum of mice in the CVB3 group were significantly higher than those in the control group,the difference was statistically significant(P<0.05).1.4 The HE staining microscope showed that the arrangement of myocardial fibers was disordered,myocardial cells were edema in the CVB3 group,and there was inflammatory cell infiltration in the myocardial interstitium.The cardiomyocytes of the control group were arranged regularly without inflammatory cell infiltration.2.Molecular detection results of pyroptosis in mice with viral myocarditis induced by CVB32.1 The expression levels of NLRP3 and caspase-1 m RNA in the myocardium of mice in each group: the expression levels of NLRP3 and caspase-1 m RNA in the myocardium of mice in the CVB3 group were significantly higher than those in the control group,the difference was statistically significant(P<0.05).2.2 The ratio of caspase-1/pro-caspase-1 in the myocardium of mice in each group: the ratio of caspase-1/pro-caspase-1 in the myocardium of mice in the CVB3 group was significantly higher than that in the control group,the difference was statistically significant(P<0.05).2.3 The levels of IL-1β in the serum of mice in each group: the levels of IL-1β in the serum of mice in the CVB3 group were significantly higher than those in the control group,the difference was statistically significant(P<0.05).3.The results of mi R-223 test results in mice with viral myocarditis induced by CVB3The expression levels of mi R-223 in the myocardium of mice in each group were detected by q RT-PCR,the expression levels of mi R-223 in the myocardium of mice in the CVB3 group were significantly lower than that in the control group,the difference was statistically significant(P<0.05).Conclusion1.Through intraperitoneal injection of CVB3 into BALB/c mice,the mice model of viral myocarditis induced by CVB3 was successfully established;2.The molecules of the associated with pyroptosis increased in the myocardium of mice with viral myocarditis induced by CVB3;3.The levels of mi R-223 decreased in mice with viral myocarditis induced by CVB3,and the mi R-223 may be involved in the regulation of pyroptosis in the myocardium of mice.