重症肌无力外周血调节性T细胞线粒体自噬异常的研究  

The abnormal mitophagy of regulatory T cells in peripheral blood of patients with myasthenia gravis

作　　者：王娜[1] 邹利[1] 袁江[1] 章红妍[1] 唐莉[1] 王云甫[1] 卢祖能[2] 

机构地区：[1]湖北医药学院附属十堰市太和医院神经内科,442000 [2]武汉大学人民医院神经内科,430000

出　　处：《中国神经免疫学和神经病学杂志》2017年第4期270-275,共6页Chinese Journal of Neuroimmunology and Neurology

基　　金：国家自然科学基金项目(编号:81671238)

摘　　要：目的探讨线粒体自噬对重症肌无力(myasthenia Gravis,MG)患者外周血调节性T细胞(CD4^+CD25^+regulatory T cell,Treg)功能的影响及关系。方法选择首次确诊、尚未接受治疗的15例MG患者,以15名健康人作为对照,分别采集外周血,以磁珠分选法获得Treg细胞,经流式细胞仪鉴定纯度后,分别通过透射电镜检测Treg细胞线粒体自噬情况、Western Blot法检测微管相关蛋白轻链3-Ⅱ(microtubule associated protein light chain 3-Ⅱ,LC3-Ⅱ)的表达水平、JC-10荧光探针检测线粒体膜电位(以红色荧光细胞数/绿色荧光细胞数比值表示)变化、羧基荧光素琥珀酰亚胺酯(CFDA-SE,CFSE)检测Treg对正常CD4^+T细胞增殖抑制能力(荧光强度越强,增殖抑制能力越强)。结果与对照组比较,MG组电镜下线粒体自噬明显减少[(25.60±7.81)个、(19.20±5.49)个,t=-2.596,P<0.05],且变形线粒体较多;MG组LC3-Ⅱ表达明显减少(0.450±0.137、0.334±0.124,t=-2.413,P<0.05);MG组Treg细胞线粒体膜电位下降(2.153±0.537,1.726±0.494,t=2.126,P<0.05);MG组CFSE检测平均荧光强度减弱(34.82±10.64、26.49±5.94,t=2.646,P<0.05)。结论 MG患者有Treg细胞的线粒体自噬功能降低,这可能与其增殖抑制功能降低相关。Objective To explore the impact of mitophagy on the function of regulatory T cells （CD4＋CD25＋ regulatory T cell, Treg） in peripheral blood of patients with myasthenia gravis （MG） and the relationship between them.Methods Treg cells were isolated from 15 patients with MG who had not been treated and 15 healthy persons as the normal control, Treg cells were isolated from human PBMCs by using the magnetic separation and were identified by flow cytometry.Mitophagy was detected by transmission electron microscope and the expression of microtubule associated protein light chain 3-Ⅱ（LC3-Ⅱ）was measured by Western Blot, JC-10 fluorescent probe was used to detect the change of mitochondria transmembrane potential, the inhibition of Treg on the proliferation of normal CD4＋T cells was detected by using flow cytometry and carboxyfluorescein diacetate succinimidyl ester（CFDA-SE, CFSE）.Results Compared with the control group,less mitophagy occured in the MG group detected by transmission electron microscope（25.60＆#177;7.81,19.20＆#177;5.49,t=-2.596,P〈0.05）.The expression of LC3-Ⅱ was reduced （0.450＆#177;0.137,0.334＆#177;0.124,t=-2.413,P〈0.05） in the MG group.The mitochondria transmembrane potential of Treg cells（the ration of red fluorescent cells to green fluorescent）was lower（2.153＆#177;0.537,1.726＆#177;0.494,t=2.126,P〈0.05）in the MG group compared with the control group.The average fluorescence intensity of CFSE was lower in MG group（34.82＆#177;10.64,26.49＆#177;5.94,t=2.646,P〈0.05）.Conclusions The mitophagy of Treg cells is reduced in MG patients.This may be related with the decreased proliferation inhibiting function of Treg.

关 键 词：重症肌无力 线粒体自噬 TREG细胞 

分 类 号：R746.1[医药卫生—神经病学与精神病学]