A study on autoimmune mechanism in the pathogenesis of myocarditis of cardiomyopathy analysis by anti-heart monoclonal antibody

为了阐明抗心脏抗体在心肌炎和心肌病发病机制中的作用，我们通过研制单克隆抗体对抗心脏抗体进行分析。脑心肌炎（EMC）病毒感染的Balb/c心肌炎小鼠血清中检测到抗心脏自身抗体。未感染小鼠的心脏冷冻切片与小鼠血清孵育，用抗小鼠lgG和lgM染色。第4天首次出现阳性颗粒染色，并持续到第90天；第21天滴度最高。将感染后第21天抗心脏抗体阳性的小鼠脾细胞与浆细胞瘤细胞系融合。采用免疫荧光法对小鼠心脏切片进行杂交瘤上清的筛选。获得了2个产生lgm的杂交瘤克隆；一个克隆IB5染色肌细胞呈条纹状，另一个克隆IB5染色内皮细胞。western免疫印迹分析结果表明，IA5与心肌肌球蛋白反应。这些研究提示肌球蛋白是EMC病毒性心肌炎的主要自身抗原之一。

In order to clarify the role of antiheart antibody in the pathogenesis of myocarditis and cardiomyopathy, we analyzed anti-heart antibody by developing monocional antibodies. Anti-heart autoantibodies were found in the sera from Balb/c mice with myocarditis infected with encephalomyocarditis (EMC) virus. Frozen sections of the hearts of uninfected mice were incubated with mouse sera and stained with anti-mouse lgG and lgM. Positive granular staining was first seen on day 4 and persisted to day 90 ; the titer was highest on day 21. Spleen cells from mice with positive anti-heart antibody on day 21 at post-infection were fused with a cell line of plasmacytoma. Supernatants of hybridoma were screened by immunoflurerescence against murine heart sections. Two clones of lgM-producing hybridoma were developed ; one clone IA5, stained myocytes with striated pattern and the other clone IB5, stained endothelial cells. Results from western immunoblotting analyses demonstrated that IA5 reacted with cardiac myosin. These studies suggest that myosin is one of the major autoantigens in EMC virus myocarditis.