THYMIC B CELL ACTIVATION IN MYASTHENIA GRAVIS

重症肌无力(MG)是一种以肌肉为特征的自身免疫性疾病 软弱。MG患者由以下原因引起的神经肌肉传递受损 与烟碱型乙酰胆碱受体(NAChR)反应的抗体。 根据临床、病理和免疫学的证据， 长期以来，胸腺一直被认为在发病机制中起关键作用。 这种疾病。重症肌无力患者胸腺是B细胞的异常部位 活动。生发中心(GC)增生见于超过75%的 病人。回收的B细胞具有与前驱一致的特性 在体内激活，并分泌自身抗体抗AChR。Achr 在胸腺中的表达可能在 自身敏化过程或耐受机制的崩溃 正常情况下可防止自身免疫的表达。这个项目的总体目标是 项目是加强我们对免疫学和分子生物学的理解 胸腺B细胞和胸腺nAchRpha的特性。具体目标是： L)测定MG胸腺GC B的功能和分子性质 细胞2)测定胸腺抗AChRpha抗体的VH和VL基因利用率 抗体，以及3)进一步鉴定nAChR阿尔法链(NAChRpha) A)蛋白在正常人和肌无力患者胸腺中的表达 表达，b)定量m RNA c)m RNA的组织定位，d) 细胞因子对信使核糖核酸的调节e)信使核糖核酸的个体发育，L) 胸腺nAChR诱导特异性免疫无反应的能力。 这些目标将部分通过使用胸腺和血细胞来实现。 取自胸腺切除的MG患者和心脏手术的对照组。 研究对象。需要检验的一个假设是，抗AChR是GC的产物 B细胞。我们还将测试胸腺GC B细胞将显示 抗原刺激和V基因偏斜的分子特征 如果GC是由常见的病理事件引起的，则利用。一个 丝状噬菌体系统将用于克隆和测序抗AChRpha 胸腺B细胞使用的VH和VL基因。新生的小鼠将经历 胸腺内注射携带AChR的细胞以确定是否导入 进入胸腺的AChR可以导致对这种自身抗原的无反应 当它随后以免疫原性方式给药时。我们会 并对胸腺AChRpha表达的个体发育进行了分析 确定自身反应性T细胞是否可能在表达之前逃逸 胸腺AChR。胸腺nAChRpha的定量和定性特征 将使用最近开发的定量RT-PCR和在 原位杂交。这些发现具有很大的临床应用潜力。 重要的是因为抗AChR抗体在MG和胸腺切除术中是致病的 通常对这种疾病有益。

Myasthenia gravis, (MG) is an autoimmune disorder characterized by muscle weakness. MG patients have impaired neuromuscular transmission caused by antibodies reactive with the nicotinic acetylcholine receptor (nAChR). Based on clinical, pathologic, and immunologic lines of evidence, the thymus has long been considered to play a pivotal role in the pathogenesis of this disease. The thymus in MG is an anomalous site of B cell activity. Germinal center (GC) hyperplasia is seen in greater than 75% of patients. The recovered B cells have properties consistent with antecedent in vivo activation and they secrete the autoantibody anti-AChR. AChR expressed in the thymus may play an important role in the autosensitization process or in the breakdown of tolerance mechanisms that normally prevent the expression of autoimmunity. The overall goal of this project is to enhance our understanding of the immunologic and molecular properties of thymic B cells and thymic nAchRalpha. Specific aims are to: l) determine the functional and molecular properties of MG thymic GC B cells 2) determine VH and VL gene utilization of thymic anti-AChRalpha antibodies, and 3) further characterize nAChR alpha chain (nAChRalpha) expressed in normal and myasthenic human thymus with regard to a) protein expression, b) quantitative mRNA c) tissue localization of mRNA, d) regulation of mRNA by cytokines e) ontogeny of mRNA expression, and l) the capacity of thymic nAChR to induce specific immunologic unresponsiveness. These aims will be approached, in part, by using thymic and blood cells obtained at thymectomy of MG patients and cardiac surgery of control subjects. One hypothesis to be tested is that anti-AChR is a product of GC B cells. We will also test the hypothesis that thymic GC B cells will show the molecular features of antigen stimulation and show skewed V-gene utilization if GC's are induced by a common pathologic event. A filamentous phage system will be used to clone and sequence anti-AChRalpha VH and VL genes used by thymic B cells. Neonatal mice will undergo intrathymic injection with AChR-bearing cells to determine if introduction of AChR into the thymus can induce unresponsiveness to this autoantigen when it is subsequently administered in an immunogenic manner. We will also carry out an analysis of the ontogeny of thymic AChRalpha expression to determine if autoreactive T cells might escape prior to expression of thymic AChR. Quantitative and qualitative features of thymic nAChRalpha will be determined using a recently developed quantitative RT-PCR and in situ hybridization. These findings are of great potential clinical importance since anti-AChR antibody is pathogenic in MG and thymectomy is often beneficial in this disorder.