Research On The Mechanism Of Immune In The Experimental Autoimmune Myocarditis

Objective:To study the immuno-costimulatory molecules CTLA-4, CD80, CD86, and anti-cardiac myosin antibodies in the development process of EAM, and explore the pathogenesis of EAM by EAM rat model.Methods:Male Lewis rats were randomly divided into two groups: (1) experimental group consisted of 40 rats. Porcine cardiac myosin and FA mixed emulsion 0.1ml were subcutaneously injected into both of hinds foot pad of rats in 1d and 8d, total pig cardiac myosin 2mg per rat every time, and then the 40 rats divided into different subgroups in accordance with 14d, 28d, 42d, 56d after the first immunization; (2) normol control group consisted 20 rats, which were injected with saline emulsion in the same way. Transthoracic echocardiography were performed in two groups in 28, 56d after the first immunization. HE staining was used to observe the change of the histopathological characteristics. Myocardial tissue pathological changes of Immunohistochemical staining and real-time fluorescence quantitative PCR detection were used to positioned and qualitative analysis the expression of CTLA-4, CD80, CD86. ELISA was used to evaluate the anti-cardiac myosin antibodies in blood plasma in 14, 28, 42, 56d after the first immunization.Results:1,Model preparation: All rats were modeled successfully.2,Echocardiography: Compared with normal group, LVEDD, LVESD increased significantly, LVEF decreased significantly (P<0.05), Pericardial effusion was observed in 28d after the first immunization; LVEDD, LVESD, IVST, LVPWT increased significantly, FS, LVEF decreased significantly (P<0.05), pericardial effusion disappeared, wall motion reduced and chambers of the heart enlarged. 3,Pathology testing: HE staining of myocardial cells in normal control group arranged in uniform. In experimental group, myocarditis began to performance as inflammatory cell infiltration, myocardial hypertrophy associated with increases of the adjacent cell gap, disorganized, and large nuclear stained deeply in 14d after the first immunization, and then inflammatory cells decreased in 42d, fibroblast proliferated significantly in 56d.4,Immunohistochemistry: CTLA-4, CD80, CD86 mainly expressed in cardiac cells or cardiac interstitial connective tissue. Compared with normal group, the expression of those proteins were higher at different times (P<0.05). After the first immunization, expression of proteins increased gradually from 14d to 28d, but decreased from 42d to 56d.5,RT-PCR: Compared with normal group, the expression of CTLA-4,CD80,CD86 mRNA were higher in experimental group (P<0.05). The expression level in 14, 28 d after the first immunization was higher than 42, 56 d (P<0.05). And the increased expression of CD80, CD86, CTLA-4 mRNA was associated with the decreased levels of cardiac function.6,Anti-cardiac myosin antibodies in serum: antibody levels in the experimental group was significantly higher than which in normal control group (P<0.05), and increased gradually with progression, began to decrease in 42d after the first immunization.Conclusion:1,EAM rats model can be made by injecting with porcine cardiac myosin.2,The expression of CD80, CD86, CTLA-4 and anti-myosin antibody promotes heart tissue inflammation, and closely related to the occurrence and development of EAM.