MiR-326调节Th17细胞参与自身免疫甲状腺炎发病机制研究

Micro-RNA (miRNA) plays an important role in the pathogenesis of autoimmune diseases and the regulation of immune cells differentiation.Significantly elevated miRNA-326 were found in the spleen CD4+ T cells and thyroid tissues in NOD.H-2h4 mice spontaneous autoimmune thyroiditis (AIT) in our previous study, and miRNA-326 level was positively correlated with CD4+IL-17+ cells expression, which is the effective cell in the pathogenesis of AIT. Therefore we mean to study the expression of miR-326 in the peripheral blood and thyroid lymphocytes of patients with AIT, and to investigate the influence of these miR-326 on differentiation and function of T cells by using miRNA mimics and antagomir to make the two miRNAs overexpression or silencing in vitro culture system. Overexpression or silencing of miR-326 by injecting lentiviral vector in NOD.H-2h4 mouse model to observe the changes of the disease onset; By fluorescence in situ hybridization (FISH), we investigated whether the two miRNAs take part in the developing of AIT by regulating the differentiation and function of Th17,confirming possible targets Ets-1 for regulating Th17. This study will elucidate the function target and mechanisms of miR-326 in the pathogenesis of AIT, providing a theoretical basis for future target therapeutic strategies of autoimmune disease which based on miRNA.

已知微小RNA（miRNA）可调控免疫细胞分化。前期研究发现NOD.H-2h4小鼠自身免疫甲状腺炎（AIT）模型脾CD4+细胞及甲状腺组织中miRNA-326显著升高且与AIT的致病性细胞CD4+IL-17+ Th17细胞表达正相关。拟研究AIT患者外周血及甲状腺内淋巴细胞miR-326表达变化和临床意义；淋巴细胞体外培养，以miRNA mimic和antagomir过表达和沉默miR-326，观察T细胞分化及功能变化。过表达或抑制miR-326表达的慢病毒载体在体干预NOD.H-2h4小鼠，明确其对AIT发生发展的影响。FISH等方法找到miR-326作用的靶细胞，明确其是否通过调节Th17细胞分化和功能参与AIT发病，证实Ets-1是调控Th17的功能靶点。本研究将阐明miR-326在AIT发病中的作用靶点和机制，为开展以miRNA为靶点的自身免疫病治疗提供理论依据。

已知微小RNA（miRNA）可调控免疫细胞分化。前期研究发现NOD.H-2h4小鼠自身免疫甲状腺炎（AIT）模型脾CD4+细胞及甲状腺组织中miR-326显著升高且与AIT的致病性细胞CD4+IL-17+ Th17细胞表达正相关。拟研究AIT患者外周血及甲状腺内淋巴细胞miR-326表达变化和临床意义；淋巴细胞体外培养，过表达和沉默miR-326，观察T细胞分化及功能变化。抑制miR-326表达的慢病毒载体在体干预NOD.H-2h4小鼠，明确其是否通过调节Th17细胞分化和功能参与AIT发病，探究miR-326调控Th17的功能靶点。本研究发现HT 患者甲状腺组织及 PBMC 中 miR-326水平均显著高于对照组，Ets-1蛋白表达和ADAM17 mRNA水平及蛋白表达均显著低于对照组，且miR-326与IL-17A 的mRNA水平呈显著正相关，Ets-1蛋白表达和ADAM17 mRNA水平与miR-326均呈显著负相关。人群PBMC体外干预实验提示miR-326对Th17细胞具有显著正向调节作用，这种正向调节作用通过靶向调节ADAM17的表达，减少ADAM17对细胞膜IL-23R的剪切，间接上调HT患者中mIL-23R的水平，增强IL-23/IL-23R的信号传导来实现的。通过小鼠体外淋巴细胞Th17细胞极化培养后siRNA干预实验验证miR-326在转录后水平调节Ets-1蛋白水平而调控Th17细胞参与AIT，同时通过小鼠慢病毒体内干预实验，进一步证实了miR-326于翻译水平调节Ets-1蛋白表达，于转录水平调节ADAM17mRNA水平而调控Th17细胞参与自身免疫性甲状腺炎。上述研究阐明了miR-326在AIT发病中的作用靶点和机制，为开展以miRNA为靶点的自身免疫病治疗提供理论依据。另外，通过芯片筛查结果提示在桥本组甲状腺组织中IL-34的mRNA水平显著低于非桥本组。并进一步实验提示IL-34可以抑制甲状腺细胞凋亡，可能是评估甲状腺细胞损伤的潜在指标。