AChR-Fc融合蛋白靶向BCR选择性识别、清除AChR反应性B细胞  

Selective recognition and elimination of acetylcholine receptor-reactive B cells with a fusion protein AChR-Fc by targeting B cell antigen receptor

作　　者：常婷[1] 林宏[1] 刘煜[1] 李柱一[1] 

机构地区：[1]第四军医大学唐都医院神经内科,710038

出　　处：《中国神经免疫学和神经病学杂志》2012年第1期17-21,共5页Chinese Journal of Neuroimmunology and Neurology

基　　金：国家自然科学基金面上资助项目(30870841)

摘　　要：目的研究乙酰胆碱受体(AChR)-Fc融合蛋白选择性识别、清除AChR反应性B细胞的作用。方法克隆人AChRα1亚单位胞外段基因(Hα1-210),插入含有CMV启动子、小鼠Kappa链先导序列以及人IgG1从铰链区到CH3基因片段的真核表达载体pAN1782中,构建重组表达载体pAN-Hα1-210,表达AChR-Fc融合蛋白;同分泌乙酰胆碱受体抗体(AChRAb)的杂交瘤细胞TIB175共孵育,采用流式细胞仪观察、分析AChR-Fc融合蛋白同杂交瘤细胞的亲和力以及对杂交瘤细胞增殖和凋亡的影响。结果成功构建真核表达载体pAN-Hα1-210,在CHO-K1细胞中稳定表达具有一定生物活性的AChR-Fc融合蛋白;AChR-Fc融合蛋白对TIB175细胞具有较高的亲和力,可特异性抑制该细胞的增殖并促进其凋亡。结论 AChR-Fc融合蛋白可特异性识别、清除AChR反应性B细胞,为进一步靶向B细胞治疗重症肌无力奠定基础。Objective To study the effects of acetylcholine receptor（AChR）-Fc fusion protein on the recognization and elimination of AChR reactive B cells.Methods The extracellular domain of human AChRα1-subunit（Hα1-210） was cloned and then inserted into a pAN1782 mammalian expression vector with a cytomegalovirus promoter,a murine immunoglobulin κ-chain leader sequence and the human genomic IgG γ1,containing regions from the hinge to the end of CH3.Recombinant the AChR-Fc fusion protein expressing plasmid,pAN-Hα1-210,was constructed.After incubation with hybridoma cells TIB175,which secret AChRAb,the binding capacity,the proliferation and apoptosis effects of AChR-Fc fusion protein on TIB175 were analyzed by flow cytometry.Results Eukaryotic expression vector pAN-Hα1-210 was constructed successfully and the AChR-Fc fusion protein with biological activity was expressed by CHO-K1 cells.AChR-Fc showed a higher binding capacity to hybridoma cells.In addition,AChR-Fc inhibited the proliferation and induced apoptosis of hybridoma cells.Conclusions AChR-Fc fusion protein can specifically recognize and eliminate AChR reactive B cells,which may lay a foundation for further works on the myasthenia gravis treatment by targeting B cells.

关 键 词：重症肌无力 融合蛋白 靶向治疗 自身反应性B细胞 FcγRⅡB 

分 类 号：R746.1[医药卫生—神经病学与精神病学] Q782[医药卫生—临床医学]