CHARACTERIZATION OF AUTOANTIBODIES IN IMMUNE CYTOPENIAS

慢性自身免疫性细胞减少症样特发性的病因 血小板减少性紫癜(AITP)和温反应性溶血性贫血(AIHA) 都是未知的。这项提议将检验自身抗体 在这些疾病中产生的每一种疾病都源于有限的 免疫球蛋白可变区生殖系基因，以及这种偏好 基因的利用将通过频繁共享的存在来反映 中国人血小板或红细胞反应性自身抗体的独特型 AITP或AIHA患者的人群。建议的实验方法 是产生多种抗独特型试剂，使用AITP作为免疫原 和由两种不同技术产生的AIHA自身抗体。第一 是通过一种新的方法培育分泌人源自身抗体的单抗细胞系 系统：SCID(严重联合免疫缺陷)小鼠 突变将稳定地植入AITP或AIHA患者的淋巴细胞和 用人的血小板或红细胞免疫，这可能会激活植入的 人类细胞，而不是遗传反应迟钝的SCID小鼠淋巴细胞； 植入并免疫的SCID小鼠将用于产生人的单克隆 产生自身抗体的杂交瘤细胞系。第二种方法是 AITP和AIHA患者自身抗体的直接纯化 抗原亲和技术可获得高度浓缩的多克隆 制备血小板或红细胞反应性自身抗体。 将使用单克隆和多克隆自身抗体来产生 BALB/c小鼠单抗独特型特异性抗体。这些AITP和AIHA 反独特型将与现有的反独特型一起使用，以 其他人类自身抗体(RF和抗DNA)建立的模式 多发性AITP或AIHA患者及其家系的独特型表达 成员，以及随机的无关的未受影响的对象。此外，抗体 从不同捐赠者制备的独特型阳性蛋白将被分析 对于免疫球蛋白同种异型和与相关自身抗原的体外结合， 提供有关独特型表达与弗兰克的关系的数据 自身免疫性疾病。最后，鼠抗独特型试剂和人类 由提议的活动产生的自体反应性B细胞系将 用于表征编码特定免疫球蛋白基因的工具 AITP和AIHA中的自身抗体。这些研究将提供重要的新的 关于人类自身免疫性疾病的遗传参数的信息。

The etiologies of chronic autoimmune cytopenias like idiopathic thrombocytopenic purpura (AITP) and warm-reactive hemolytic anemia (AIHA) are unknown. This proposal will test the hypothesis that autoantibodies produced in each of these diseases originate from limited repertoires of immunoglobulin variable region germline genes, and that such preferential genetic utilization will be reflected by the presence of frequently shared idiotypes among platelet or red cell reactive autoantibodies from populations of AITP or AIHA patients. The proposed experimental approach is to generate multiple anti-idiotypic reagents, using as immunogens AITP and AIHA autoantibodies generated by two different techniques. The first is to develop human monoclonal autoantibody-secreting cell lines by a novel system: Mice homoxygous for the scid (severe combined immunodefiency) mutation will be stably engrafted with AITP or AIHA patient lymphocytes and immunized with human platelets or red cells, which may activate engrafted human cells but not the genetically unresponsive scid mouse lymphocytes; engrafted and immunized scid mice will be used to generate human monoclonal autoantibody producing hybridoma cells lines. The second approach is direct purification of autoantibodies from AITP and AIHA patients, using antigen-affinity techniques to yield highly enriched polyclonal preparations of platelet or red cell reactive autoantibodies. Both monoclonal and polyclonal autoantibodies will be used to produce BALB/c mouse monoclonal idiotype-specific antibodies. These AITP and AIHA anti-idiotypes will be used in conjunction with extant anti-idiotypes to other human autoantibodies (RF and anti-DNA) to establish the pattern of idiotype expression among multiple AITP or AIHA patients, their family members, and random unrelated unaffected subjects. Additionally, antibody idiotype-positive proteins prepared from various donors will be analyzed for immunoglobulin isotype and binding to relevant autoantigens in vitro, providing data regarding the association of idiotype expression with frank autoimmune disease. Finally, both murine anti-idiotypic reagents and human autoreactive B cell lines generated by the proposed activities will be tools for characterizing specific immunoglobulin genes that encode autoantibodies in AITP and AIHA. These studies will provide important new information regarding genetic parameters in human autoimmune diseases.