中文摘要：

目的 探讨Tfh细胞在重症肌无力（MG）患者胸腺中乙酰胆碱受体抗体（AChR-Ab）产生中的作用及机制。方法 收集28例MG患者、9例无胸腺异常的先天性心脏病患者和9例未合并MG的胸腺增生患者。采用流式细胞术检测胸腺中滤泡辅助T（Tfh）细胞和B细胞的比例,分别通过RT-PCR和Western-Blotting技术检测胸腺中Th1、Th2和Tfh细胞特异性转录因子T-bet、GATA-3、Bcl-6 mRNA水平和细胞因子干扰素γ（IFN-γ）、白细胞介素4（IL-4）、IL-21表达;采用HE和多标免疫荧光染色观察胸腺中生发中心以及Tfh细胞和B细胞之间的组织结构关系;采用放射免疫沉淀技术（RIA）检测MG患者胸腺和血清中AChR-Ab滴度;免疫磁珠提取胸腺中Tfh细胞和B细胞共培养,检测培养上清中IL-21和AChR-Ab滴度水平。结果 MG患者胸腺中Tfh细胞和B细胞比例及Tfh细胞相关转录因子Bcl-6mRNA水平和细胞因子IL-21表达均较对照组增高（P〈0.01）;MG患者胸腺中抗AChR-Ab滴度较对照组增高（P〈0.01）;MG患者胸腺中存在异位生发中心,且在生发中心内Tfh细胞和B细胞存在共定位;MG患者胸腺Tfh和B细胞共培养上清中IL-21和AChR-Ab水平较对照组增多（P〈0.01）,且AChR-Ab产生可以被IL-21中和抗体阻断。结论 Tfh细胞通过在MG患者胸腺中辅助B细胞形成异位生发中心及促进抗AChR-Ab产生参与MG的发生和发展。

英文摘要：

Objective To study the role of T follicular helper （Tfh） cells in the pathogenesis of myasthenia gravis （MG）. Methods Twenty-eight MG patients were recruited in our study. Non-MG-associated thymic hyperplasia （NMG） and non-thymic abnormal congenital heart disease patients were used as controls. The frequencies of Tfh ceils and B cells in thymocytes were detected via flow cytometry. The transcription factor T- bet, GATA-3 and Bcl-6 mRNA expressions in thymocytes were detected with Reverse Transcription-Polymerase Chain Reaction （RT-PCR）. Cytokines interferon 7 （IFN-γ）, interleukin-4 （IL-4） and interleukin-21 （IL-21） in thymocytes were measured by Western-Blotting. Anti-human acetylcholine receptor （AChR） antibody in the serum or thymic tissue homogenate supernatant was assayed using the radioimmunoprecipitation method （RIA）. CD4, CD19, CD35, CXCR5 in the thymus were observed by hematoxylin-eosin （HE） and immunofluorescenee staining. CD19^＋ B cells and CD4^＋ CXCR5^＋ T cells were isolated from thymus using immunomagnetic sorting method and were co-cultured. The IL-21 and anti-AChR antibody secretions in co-culture supernatant were detected after 7 days. Results The frequency of Tfh cells and B cells increased significantly in thymocytes of MG patients compared with NMG and HC, and Tfh cells counts positively correlated with disease severity. Tfh cell-associated transcription factors Bcl-6 mRNA and cytokines IL-21 expression were enhanced in the thymocytes of MG patients. Germinal center structure was observed in MG thymus through HE and immunofluorescence staining, which was not observed in the control group. Tfh cells and B cells co-localized in the germinal center of the MG thymus. High level of IL-21 and anti-AChR antibody concentrations were detected in the co-culture of patients Tfh and B cells but the autoantibody titers decreased dramatically by the presence of anti-IL-21 antibodies. Conclusions The enhanced frequency of Tfh cell was involved in the pathological