被动免疫抗LRP4抗体及抗agrin抗体致重症肌无力的研究

Myasthenia gravis (MG) is an autoimmune disease with severe impairments in the neuromuscular junction (NMJ). Agrin-LRP4-MuSK signaling pathway plays critical roles in promoting AChR clustering, NMJ formation and function. Even though anti-AChR antibodies is the main cause of MG, there is still a part of MG patients showing AChR antibodies negative. We have detected the anti-LRP4 and anti-agrin antibodies in the serum of MG patients with AChR antibodies negative. Furthermore, immunization of LRP4 and agrin to mice induced MG-related deficits. However, whether the anti-LRP4 and anti-agrin antibodies can induce MG directly is unclear. In this project, we will purify the anti-LRP4 and anti-agrin antibodies from MG patients with AChR antibodies negative, and inject the purified antibodies to the mice. We will test if the injected mice exhibit MG-related deficits, like: decreased muscle strength, fragmented NMJs morphology, compromised neuromuscular transmission and repetitive nerve stimulation. Moreover, to clarify underlying molecular mechanisms, we will map the epitopes in Lrp4 or agrin binding with the anti-LRP4 and anti-agrin antibodies, and analyze if blocking the epitopes by antibodies breakdown agrin-LRP4-MuSK signaling. These results will be helpful for illuminating the effects of anti-LRP4 and anti-agrin antibodies on the pathogenesis of MG, and could possibly lead to strategy for effective diagnose and treatment of MG.

重症肌无力(MG)是一种神经肌肉接头(NMJ)功能严重受损的自身免疫性疾病。经典信号通路agrin-LRP4-MuSK介导AChR聚集对NMJ的形成和功能具有关键作用。虽然AChR抗体是导致MG的主要病因，但仍有部分患者为AChR抗体阴性。我们在这些阴性患者血清中检测到LRP4及agrin抗体，并且在小鼠中免疫LRP4及agrin蛋白成功建立了MG小鼠模型，但LRP4及agrin抗体是否直接导致MG尚不清楚。为此，本项目拟从AChR抗体阴性患者血清中纯化LRP4及agrin抗体，注射至免疫缺陷小鼠，通过检测肌力、NMJ形态和电生理，结合重复性电刺激实验，鉴定被注射小鼠是否表现MG样症状。进一步鉴定LRP4及agrin抗体与各自抗原结合的表位，分析是否阻碍agrin-LRP4-MuSK关键通路，从而在机制上阐明LRP4及agrin抗体导致MG发病的分子机制，为临床诊断及治疗提供了新的思路。

重症肌无力（Myasthenia gravis,MG）是一种自身免疫性疾病，可造成神经肌肉接头（NMJ）突触传递功能障碍。MG的发病与抗体的产生密切相关，80-85%MG患者血清中可以检测到抗AChR抗体，但仍有约20%的患者血清并不能检测到抗AChR抗体，提示这些患者可能是由于产生了其它对NMJ形成和维持关键蛋白的抗体而致病。在NMJ处AChR聚集受损将扰乱神经肌肉之间的信息传递，导致MG和先天性肌无力综合症（congenital myasthenic syndromes）等病症的发生。本项目筛选了抗agrin抗体阳性的MG患者，纯化该种血清并注射入小鼠构建MG模型。取得以下结果：1）注射抗agrin抗体的小鼠体重下降，翻转悬网时间减少，肌肉收缩力减弱；2）神经肌肉接头AChR聚集功能受损。AChR形态呈现碎片化改变，AChR面积减少，AChR聚集碎片化数量增多；3）抗agrin抗体影响了agrin-LRP4-MuSK信号通路的传导。本项目结果丰富了抗agrin抗体致MG的分子机制研究，也为临床上治疗MG提供了科学依据。

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