Detection of autoantibody against extracellular epitopes of N-methyl-D-aspartate receptor by cell-based assay
Detection of autoantibody against extracellular epitopes of N-methyl-D-aspartate receptor by cell-based assay
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DOI：
10.1016/j.neures.2011.07.1834
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发表时间：
2011-11-01
影响因子：
2.9
通讯作者：
Mori, Hisashi
中科院分区：
文献类型：
--
作者：
Takano, Shiho;Takahashi, Yukitoshi;Mori, Hisashi
提出了抗N-甲基-D-天冬氨酸受体（NMDAR）脑炎的概念，这是一种由抗NMDAR自身抗体介导的严重、潜在致死性、治疗反应性疾病。由于副肿瘤性抗NMDAR脑炎在肿瘤切除和免疫治疗后具有更好的预后，因此需要用于检测针对NMDAR的细胞外表位的功能性自身抗体的快速定量系统。为了检测识别NMDAR的细胞外表位的自身抗体，我们稳定表达了在没有任何拮抗剂的情况下降低异源细胞表面上的Ca 2+渗透性的突变NMDAR。使用表达突变NMDAR亚基的细胞通过免疫细胞化学和使用稳定表达突变NMDAR的活细胞的细胞上蛋白质分析来分析来自患者的血清和CSF样品。此外，我们能够表达突变型GluR zeta 1（NR 1. GluN 1）亚基，并获得了识别GluR zeta 1的细胞外表位的自身抗体的存在以及患者血清和CSF中自身抗体诱导内化的直接证据。在37 ℃下，细胞上蛋白质印迹分析的特异性得到提高。结合我们的on-cell Western分析，详细分析NMDAR的胞外表位，和NMDAR的内化分析，这种快速定量检测将对抗NMDAR脑炎的诊断，临床治疗的评价和随访有价值。(C)2011 Elsevier爱尔兰有限公司和日本神经科学学会。All rights reserved.
The concept of anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis, a severe, potentially lethal, treatment-responsive disorder, mediated by autoantibodies against NMDAR was proposed. Because paraneoplastic anti-NMDAR encephalitis has a better prognosis after tumor resection and immunotherapy, rapid quantitative systems for detecting functional autoantibodies against extracellular epitopes of NMDAR are necessary. To detect autoantibodies recognizing extracellular epitopes of NMDAR, we stably expressed mutant NMDAR that decreases Ca2+ permeability on a heterologous cell surface without any antagonist. Serum and CSF samples from patients were analysed using the cells expressing mutant NMDAR subunits by immunocytochemistry and on-cell Western analysis using live cells stably expressing mutant NMDAR. Furthermore, we were able to express mutant GluR zeta 1(NR1. GluN1) subunit of NMDAR alone on the cell surface and obtained direct evidence of the presence of autoantibodies recognizing extracellular epitopes of GluR zeta 1 and the induction of internalization by autoantibodies in serum and CSF from patients. The specificity of on-cell Western analysis was improved at 37 degrees C. The combination of this rapid quantitative assay using our on-cell Western analysis, detailed analysis of extracellular epitopes of NMDAR, and internalization assay of NMDAR will be valuable for the diagnosis, evaluation of clinical treatments, and follow-up of anti-NMDAR encephalitis. (C) 2011 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.