脂多糖诱导Treg失衡对实验性自身免疫性重症肌无力的免疫调节作用  

Immunomodulatory effects of lipopolysaccharide-induced Treg imbalance on experimental autoimmune myasthenia gravis

作　　者：黎良康 司伟岳 吴兴原 周阳 魏舒丽 董晶 李晓丽[1,2] 段瑞生[1,2] LI Liangkang;SI Weiyue;WU Xingyuan;ZHOU Yang;WEI Shuli;DONG Jing;LI Xiaoli;DUAN Ruisheng(Department of Neurology,Shandong Provincial Qianfoshan Hospital of Shandong University,Jinan 250014,Shandong,China;Department of Neurology,The First Affiliated Hospital of Shandong First Medical University&Shandong Provincial Qianfoshan Hospital,Shandong Institute of Neuroimmunology,Shandong Provincial Medicine and Health Key Laboratory of Neuroimmunology,Jinan 250014,Shandong,China)

机构地区：[1]山东大学附属山东省千佛山医院神经内科,山东济南250014 [2]山东第一医科大学第一附属医院(山东省千佛山医院)神经内科,山东省神经免疫研究所,山东省医药卫生神经免疫重点实验室,山东济南250014

出　　处：《山东大学学报（医学版）》2023年第10期1-8,共8页Journal of Shandong University:Health Sciences

基　　金：山东第一医科大学学术提升计划(2019QL013);山东省千佛山医院国家自然科学基金培育基金(QYPY2020NSFC0610)。

摘　　要：目的探究实验性自身免疫性重症肌无力(EAMG)合并急性感染时免疫细胞的改变及其功能特征。方法利用大鼠乙酰胆碱受体(AChR)α亚基的97-116(R97-116)肽段免疫6~8周龄雌性Lewis大鼠,发病高峰期腹腔注射脂多糖(LPS)诱导急性感染模型为LPS组,腹腔注射PBS作为对照(PBS组)。记录临床评分及体质量。流式细胞术检测外周循环和淋巴器官中Th1、Th17、Treg细胞的比例以及脾脏滤泡辅助T细胞(Tfh)、树突状细胞(DC)、生发中心B细胞(GCB)的比例和功能。对脾脏分别进行PNA及CD4、Foxp3免疫荧光检测。采用酶联免疫吸附实验(ELISA)检测血清抗R97-116 IgG抗体及亚型水平。结果与PBS组相比,LPS组大鼠临床症状较重、体质量减轻,脾脏Treg细胞(CD4^(+)CD25^(+))比例(P=0.016)及Foxp3的表达均明显减少。另外,LPS组大鼠脾脏PNA的表达增多且呈簇状,且GCB细胞表达MHC-Ⅱ及分泌IL-6的比例明显增加(P=0.002、P=0.017)。LPS组大鼠DC亚群表达CD80的平均荧光素强度(MFI)增加,但无统计学差异(P=0.057)。LPS组大鼠外周血免疫球蛋白G(IgG)水平无变化,保护性抗体IgG1水平降低(P=0.005),致病性抗体IgG2b水平无变化,IgG1/IgG2b下调(P=0.018)。结论LPS可能通过减少脾脏Treg细胞的比例及Foxp3的表达,增强脾脏生发中心反应、GCB细胞的功能以及DC亚群共刺激的能力来下调保护性抗体水平,最终导致EAMG大鼠的临床症状加重。Objective To investigate the changes and functional characteristics of immune cells in experimental autoimmune myasthenia gravis(EAMG)complicated with acute infection.Methods Female Lewis rats aged 6-8 weeks were immunized with R97-116 peptide of rat acetylcholine receptor alpha subunit.Lipopolysaccharide(LPS)was injected intraperitoneally to induce acute infection(LPS group),and PBS was injected intraperitoneally to set controls(PBS group).Clinical scores and body weight were recorded.Flow cytometry was used to detect the frequencies of Th1,Th17 and Treg cells from lymphoid organs and peripheral circulation,and follicular helper T(Tfh)cells,dendritic cells(DCs)and germinal center B(GCB)cells in the spleen.Peanut agglutinin(PNA),CD4 and Foxp3 levels in the spleen were detected with immunofluorescence.The levels of anti-R97-116 IgG antibody and its subclasses in serum were analyzed with ELISA.Results Compared with the PBS group,the LPS group had aggravated clinical symptoms and lower body weight.The frequency of Treg(CD4^(+)CD25^(+))in the spleen was decreased(P=0.016),and the Foxp3 expression in the spleen was reduced.In addition,the PNA in the spleen of the LPS group was increased and clustered,and the expression of MHC-Ⅱ(P=0.002)and secretion of IL-6(P=0.017)of GCB cells were increased.The mean fluorescence intensity(MFI)of DC subclass expressing CD80 was increased but not statistically significant(P=0.057).The level of protective antibody IgG1 in peripheral blood was decreased in LPS group(P=0.005),the level of pathogenic antibody IgG2b did not change,and IgG1/IgG2b was down-regulated(P=0.018).Conclusion LPS may reduce the frequency of Treg cells and the expression of Foxp3 in the spleen,enhance the germinal center response,function of GCB cells and ability of DC co-stimulation,eventually leads to the aggravation of clinical symptoms of EAMG rats.

关 键 词：脂多糖 感染 实验性自身免疫性重症肌无力 TREG细胞 

分 类 号：R746.1[医药卫生—神经病学与精神病学]