雷帕霉素改善Graves眼病小鼠CD4^(+)T细胞亚群失衡的研究  

Rapamycin restores CD4^(+)T cell subsets imbalance in Graves′ophthalmopathy mice model

作　　者：张萌[1] 王玲[1] 丁习 何明倩 陈子怡 吴晓燕[1] 伍丽萍[1] 施秉银[1] 王悦[1] Zhang Meng;Wang Ling;Ding Xi;He Mingqian;Chen Ziyi;Wu Xiaoyan;Wu Liping;Shi Bingyin;Wang Yue(Department of Endocrinology,First Affiliated Hospital of Xi′an Jiaotong University,Xi′an 710061,China)

机构地区：[1]西安交通大学第一附属医院内分泌科,西安710061

出　　处：《中华内分泌代谢杂志》2023年第11期964-973,共10页Chinese Journal of Endocrinology and Metabolism

基　　金：国家自然科学基金项目(82201238、82270827、82170805、81970679);陕西省自然科学基础研究计划项目(2020JM-392)。

摘　　要：目的本文从Graves眼病(Graves′ophthalmopathy,GO)的发病机制出发,旨在研究雷帕霉素对于GO小鼠CD4^(+)T细胞亚群紊乱的调节作用,进而探索其对于GO小鼠眼病及甲状腺功能亢进症(甲亢)表型的影响,为GO的病因治疗提供新的思路和可能。方法利用表达TSHR A亚单位的重组腺病毒肌肉重复9次注射Balb/c小鼠制备GO小鼠。通过在饲料中添加雷帕霉素(14 ppm)给予小鼠干预,末次免疫后4周安乐死小鼠,收集外周血、甲状腺、眼眶、脾脏等组织检测TT4、促甲状腺素受体抗体(TRAb)、甲状腺病理改变和眼眶病理改变,并通过流式细胞学分析、免疫组化等评估CD4^(+)T细胞亚群等免疫学相关指标。结果GO小鼠造模后不仅表现以球后纤维化和球后脂肪新生的眼部病变(80%~90%),还表现为自身抗体升高和血清总甲状腺素增高的甲亢病变(80%)。球后进一步的脾脏细胞流式细胞学、甲状腺免疫组化和眶后组织的免疫组化显示GO小鼠CD4^(+)T细胞亚群的偏移:Th1/Th2细胞平衡向Th1倾斜,并伴随有Treg细胞比率降低。雷帕霉素干预后的小鼠脾脏细胞流式细胞学显示Th1、Th17细胞比率回落和Th2、Treg细胞比率回升。甲状腺和眼眶组织免疫组化也显示Th1/Th2细胞失衡和Treg细胞降低得到了改善。结论GO小鼠模型表现了显著的CD4^(+)T细胞亚群失衡,而雷帕霉素不仅可以调节GO小鼠的CD4^(+)T细胞亚群紊乱,还能有效改善GO小鼠眼病及甲亢表型。因此,CD4^(+)T细胞亚群失衡是GO的病因干预环节之一,雷帕霉素是潜在的GO干预方式,未来可以进行随机临床研究进一步探索研究。Objective To study the effect of rapamycin on the disorder of CD4^(+)T cell subsets in Graves′ophthalmopathy(GO)mice,as well as the ophthalmopathy and hyperthyroidism phenotype,providing new possibilities for the treatment of GO.Methods 6-8 weeks old female Balb/c mice were injected intramuscularly with adenovirus expressing the A-subunit of TSHR(A-sub-Ad)9 times.Rapamycin was given by embedding in the feed(14 ppm).The animals were euthanized 4 weeks after the final injection to obtain blood,spleen cells,thyroid glands,and orbital tissue.TT4,thyrotropin receptor antibody(TRAb),thyroid,and orbital pathologic changes were detected,and the CD4^(+)T cell subgroup was evaluated by flow cytometry and immunohistochemistry.Results After final immunization,the mice showed characteristics of GO:increased retrobulbar fibrosis and retrobulbar adipogenesis that indicated ophthalmopathy,increased autoantibodies,and serum total thyroxin that indicated hyperthyroidism.After the intervention of rapamycin,retrobulbar fibrosis and retrobulbar adipogenesis were significantly improved,and the incidence of ophthalmopathy was reduced from 80%-90%to 20%.Moreover,the increase of total thyroxin was reduced from 80%to 20%,and the metabolic condition and thyroid pathology were also improved.Flow cytometry of the spleen,immunohistochemistry of the thyroid and orbital tissue revealed that GO mice exhibited Th1 dominance in Th1/Th2 balance and reduction of Treg cells.After the intervention of rapamycin,flow cytometry showed that the ratio of Th1 and Th17 cells decreased and the ratio of Th2 and Treg cells increased.Immunohistochemistry of thyroid and orbital tissues also confirmed improvement of Th1/Th2 cell imbalance and Treg cell reduction.Conclusion GO mouse model showed a significant imbalance of CD4^(+)T cell subsets,and rapamycin could not only regulate the disorder of CD4^(+)T cell subsets in GO mice,but also effectively improve the phenotype of ophthalmopathy and hyperthyroidism.Therefore,the imbalance of CD4^(+)T cell subsets is one

关 键 词：GRAVES眼病 雷帕霉素 CD4^(+)T细胞亚群 BALB/C小鼠 动物模型 

分 类 号：R581[医药卫生—内分泌] R771.3[医药卫生—内科学]