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

为了阐明抗心抗体在心肌炎和心肌病发病机制中的作用，我们通过研制单抗来分析抗心抗体。在感染了脑心肌炎(EMC)病毒的心肌炎Balb/c小鼠血清中发现了抗心脏自身抗体。将未感染小鼠的心脏冰冻切片与小鼠血清孵育，用抗小鼠免疫球蛋白和免疫球蛋白染色。阳性颗粒染色始于第4天，一直持续到第90天，滴度在第21天最高。将感染后第21天抗心脏抗体阳性小鼠的脾细胞与浆细胞瘤细胞系进行融合。用免疫荧光法对杂交瘤细胞上清液进行免疫荧光筛选。建立了两个产生免疫球蛋白的杂交瘤细胞克隆：一个克隆IA5，染有条纹的肌细胞；另一个克隆IB5，染内皮细胞。免疫印迹分析结果表明IA5能与心肌肌球蛋白反应。这些研究表明，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.