The Role Of IL-11 On Myocardial Fibrosis In Mice With Viral Myocarditis

Objective: During developing of viral myocarditis(VMC)induced by Coxsackie virus B3(CVB3)in mice,the changes of interleukin 11(IL-11)and interleukin 11 receptor(IL-11R)were investigated,and the relationships with myocardial tissue fibrosis were analysed.The aim was to study the effect of IL-11 on myocardial fibrosis in mice with VMC.Methods:(1)Study group: Sixty-four specific pathogen free(SPF)male BALB/c mice were randomly divided into the experimental group(n=32)and control group(n=32).Each group was divided into four subgroups: 1 week(W1),2 weeks(W2),4 weeks(W4),and 6 weeks(W6).There were eight mice in each subgroup.(2)Mice model of VMC was established by intraperitoneal injection of CVB3 venom in the experimental group,and the same amount of phosphate-buffered saline(PBS)was intraperitoneally injected in the control group.The mice were sacrificed at the corresponding time points,blood was collected,plasma was isolated,and heart tissue samples were collected aseptically.(3)The myocardial tissue was paraffin sectioned,and hematoxylin and eosin(H&E)staining was performed to determine the myocardial histopathological changes and to calculate the myocardial pathological scores.Masson’s staining was used to determine the degree of myocardial fibrosis.The relative expression of interleukin-11 receptor α chain(IL-11Rα),collagen type I-A1(COL1-A1),and collagen type Ⅲ-A1(COL3-A1)in myocardial tissue were detected by immunohistochemistry.The real time quantitative polymerase chain reaction(q RT-PCR)was used to determine the relative expression levels of mRNA of IL-11,IL-11Rα,COL1-A1,COL3-A1,MMP9,and TIMP-1.The expressions of IL-11,MMP9,and TIMP-1 proteins in the plasma were determined by an enzyme-linked immunosorbent assay(ELISA).Results:(1)H&E staining of myocardial tissue:Cardiomyocytes in each subgroup of the control group were arranged in a regular and orderly manner,their morphology was normal,and no inflammatory cell infiltration or myocardial cell necrosis was observed in the myocardial interstitium and perivascular areas.In the experimental group,inflammatory cell infiltration and focal necrosis in myocardial tissue were observed in the W1 subgroup,there was extensive inflammatory cell infiltration and myocardial necrosis in W2 subgroup.The extent of myocardial cell necrosis and interstitial inflammatory cell infiltration significantly decreased and was accompanied by obviously myocardial fibrosis in the W4 subgroup.The extent of myocardial fibrosis progressively increased in the W6 subgroup.The scores of myocardial histopathology in each subgroup of the experimental groups were significantly higher than that in the control group(P all<0.05),and the myocardial histopathological scores of the W2 subgroup in the experimental group were the highest.(2)Masson’s staining of myocardial tissue: a small amount of fibrous tissue was evenly distributed in the myocardial interstitium and around the blood vessels in each subgroup of the control group.In the experimental group,fibrous tissue increased in the W1 subgroup.There was a small amount of fibrous tissue in the periphery of the myocardial necrosis in the W2 subgroup.A large amount of fibrous tissue observed in the W4 subgroup;and fibrous tissue further increased in the W6 subgroup.The collagen volume fractions(CVF)of myocardial tissue in each subgroup of the experimental group were significantly higher than that in the control group(P all<0.01).In the experimental group,the CVF increased gradually in the W1,W2,W4,and W6 subgroups(P all<0.01).(3)The expression of COL1-A1,COL3-A1,and IL-11Rα in myocardial tissue of control group was the lowest.In the experimental group,the expression of COL1-A1,COL3-A1,and IL-11Rα in myocardial tissue of mice increased gradually with the evolution of the disease(P all<0.01).The expression of COL1-A1,COL3-A1,and IL-11Rα in myocardial tissue of mice in the W6 subgroup were the highest.(4)The mRNA expression of IL-11 and IL-11 R in myocardial tissue of all subgroup from experimental group were significantly higher than that in the corresponding subgroup from control group(P all<0.05),and gradually increased with the disease evolution(P all<0.05).The mRNA expression of COL1-A1,COL3-A1 and MMP9 in myocardial tissue of W2 subgroup,W4 subgroup and W6 subgroup from experimental group were significantly higher than that in the corresponding subgroup from control group(P all<0.05),and gradually increased with the disease evolution(P all<0.05),while The mRNA expression of TIMP-1 significantly decreased(P all<0.05).(5)Compared with the corresponding subgroup in control group,the levels of plasma IL-11 and MMP9 significantly increased in all subgroup from experimental group(P all<0.01),and increased gradually with the course of disease(all P <0.01),while TIMP-1 level significantly decreased(P all<0.05).Conclusion: Pathogenesis process of VMC induced by CVB3 in mice was accompanied by progressively myocardial fibrosis.The expressions of mRNA and protein of IL-11,IL-11 R,COL1-A1,COL3-A1,and MMP9 were gradually upregulated in myocardial tissue,while TIMP-1 was downregulated at the mRNA and protein levels.The plasma levels of IL-11 and MMP9 gradually increased,while TIMP-1 gradually decreased.These results suggested that IL-11 could promote myocardial fibrosis in VMC mice.