The Role And Mechanism Of CD40 Immunoglobulin Chimera Protein In Mice With Viral Myocarditis

ObjectivesThis study was sought to investigate the effect of CD40 immunoglobulin chimera protein (CD40Ig) and its influence on the balance of Thl/Th2 in murine viral myocarditis induced by Coxsackievirus B3 (CVB3).MethodsA total of 106 four to six week old BALB/c male mice were used in the experiments, which were divided into CD40Ig group (n=16), CVB3 group (n=40), IgG group (n=40) and normal control group(n=10) randomly. The mice in CVB3 group, IgG group and CD40Ig group were inoculated intraperitoneally with 0.15ml CVB3 (TCID5010-3/L) and the mice in normal control group with 0.15ml eagle. The mice in IgG group and CD40Ig group were inoculated with IgG(0.1mg/kg) and CD40Ig(0.1mg/kg) on 6 hours and 72 hours post inoculation(p.i.), respectively. The surplus mice in each group were sacrificed on day 7 p.i.. The enzyme linked immunosorbent assay (ELISA) was used to measure the concentration of sera IFN-y, IL-4. The heart were removed aseptically, half of each heart was used for hematoxylin and eosin (HE) staining. The myocardial histopathologic scores were determined by optical microscope. Half of each heart was freezed in -80℃refrigerator. The expression of IFN-y, IL-4 and CVB3 mRNA were detected by Real time polymerase chain reaction(RQ-PCR). The expression of CD40 protein in myocardium was determined by immunohistochemistry. Results1. The incidence of myocarditis were 100%. There were intensive myocyte degeneration, necrosis and inflammatory cells infiltration in the myocardium of mice inoculated by CVB3.2. The mice mortality in CD40Ig group were lower than that in CVB3 group (50% vs.80%; P<0.05) and IgG group (50% vs.77.5%; P<0.05). The histopathologic scores were much lower than that in CVB3 group (0.85±1.01 vs.1.78±1.05; P<0.05). The expression of CVB3 mRNA in CD40Ig group were much lower than CVB3 group (0.91±0.75 vs.3.48±2.89; P<0.05).3. The serum level of IFN-y of mice in CVB3 group were significantly higher than that in normal control group (196.30±16.89 vs.26.20±7.61; P<0.05).The serum level of IL-4 of mice in CVB3 group were much lower than that in normal control group (50.80±6.04 vs.146.50±21.27; P<0.05). The serum level of IFN-γof mice in CD40Ig group were much lower than that in CVB3 group (40.11±12.04 vs. 196.30±16.89; P<0.05).The serum level of IL-4 of mice in CD40Ig group were significantly higher than that in CVB3 group (84.96±23.55 vs.50.80±6.04; P<0.05).4. The expression of IFN-ymRNA in myocardium of mice in CVB3 group were significantly higher than that in normal control group (61.29±68.24 vs.6.61±15.09; P<0.05). The expression of IL-4 mRNA in myocardium of mice in CVB3 group were no significant difference with normal control group (P>0.05). The expression of IL-4 mRNA in myocardium of mice in CD40Ig group were significantly higher than that in CVB3 group (128.58±55.89 vs.1.38±1.69; P<0.05). The expression of IFN-y was no significant difference in these groups(P>0.05)5. CD40 expressions were observed in both myocardium of CVB3 group and CD40Ig group. There was strong expression of CD40 on the membrane of infiltrating cells, and on the membrane and cytoplasm of cardiac myocyte in the inflammatory areas of myocardium.Conclusions1. Treatment with CD40Ig reduced the mice mortality, relieved myocardial inflammation and reduced viral replication. It suggested that CD40Ig have therapeutical role in murine viral myocarditis induced by CVB3.2. The peripheral serum level of IFN-y increased and the serum level of IL-4 decreased significantly. The expression of IFN-y mRNA in myocardium increased. It suggested that the Thl response were predominated during the acute stage of VMC.3. CD40Ig could decrease of IFN-γlevel and IL-4 level in peripheral serum, and up-regulate IL-4 mRNA in myocardium. It suggested that CD40Ig could decrease Thl response, reinforce Th2 response, and switch the balance of Thl/Th2. Therefore it can relive myocardial immune injury.