Richter W, Endl J, Eiermann T H, Brandt M, Kientsch-Engel R, Thivolet C, Jungfer H, Scherbaum W A
Department of Internal Medicine 1, Univerity of Ulm, Federal Republic of Germany.
Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8467-71. doi: 10.1073/pnas.89.18.8467.
The autoimmune phenomena associated with destruction of the beta cell in pancreatic islets and development of type 1 (insulin-dependent) diabetes mellitus (IDDM) include circulating islet cell antibodies. We have immortalized peripheral blood lymphocytes from prediabetic individuals and patients with newly diagnosed IDDM by Epstein-Barr virus transformation. IgG-positive cells were selected by anti-human IgG-coupled magnetic beads and expanded in cell culture. Supernatants were screened for cytoplasmic islet cell antibodies using the conventional indirect immunofluorescence test on cryostat sections of human pancreas. Six islet cell-specific B-cell lines, originating from a patient with newly diagnosed IDDM, could be stabilized on a monoclonal level. All six monoclonal islet cell antibodies (MICA 1-6) were of the IgG class. None of the MICA reacted with human thyroid, adrenal gland, anterior pituitary, liver, lung, stomach, and intestine tissues but all six reacted with pancreatic islets of different mammalian species and, in addition, with neurons of rat cerebellar cortex. MICA 1-6 were shown to recognize four distinct antigenic epitopes in islets. Islet cell antibody-positive diabetic sera but not normal human sera blocked the binding of the monoclonal antibodies to their target epitopes. Immunoprecipitation of 35S-labeled human islet cell extracts revealed that a protein of identical size to the enzyme glutamate decarboxylase (EC 4.1.1.15) was a target of all MICA. Furthermore, antigen immunotrapped by the MICA from brain homogenates showed glutamate decarboxylase enzyme activity. MICA 1-6 therefore reveal glutamate decarboxylase as the predominant target antigen of cytoplasmic islet cell autoantibodies in a patient with newly diagnosed IDDM.
与胰岛β细胞破坏及1型（胰岛素依赖型）糖尿病（IDDM）发生相关的自身免疫现象包括循环胰岛细胞抗体。我们通过爱泼斯坦-巴尔病毒转化使糖尿病前期个体和新诊断的IDDM患者的外周血淋巴细胞永生化。通过抗人IgG偶联磁珠选择IgG阳性细胞，并在细胞培养中进行扩增。使用人胰腺冰冻切片上的传统间接免疫荧光试验筛选上清液中的细胞质胰岛细胞抗体。来自一名新诊断的IDDM患者的6个胰岛细胞特异性B细胞系可在单克隆水平上稳定下来。所有6种单克隆胰岛细胞抗体（MICA 1-6）均为IgG类。MICA均不与人甲状腺、肾上腺、垂体前叶、肝脏、肺、胃和肠道组织反应，但所有6种均与不同哺乳动物物种的胰岛反应，此外，还与大鼠小脑皮质的神经元反应。MICA 1-6被证明可识别胰岛中的4个不同抗原表位。胰岛细胞抗体阳性的糖尿病血清而非正常人血清可阻断单克隆抗体与其靶表位的结合。对35S标记的人胰岛细胞提取物进行免疫沉淀显示，一种大小与谷氨酸脱羧酶（EC 4.1.1.15）相同的蛋白质是所有MICA的靶标。此外，MICA从脑匀浆中免疫捕获的抗原显示出谷氨酸脱羧酶活性。因此，MICA 1-6揭示谷氨酸脱羧酶是一名新诊断的IDDM患者中细胞质胰岛细胞自身抗体的主要靶抗原。
