miR-182及其启动子甲基化状态在葡萄膜炎发生中的作用和机制研究

Behçet diseases and VKH syndrome are the two most common uveitis entities leading to blindness in China. Our previous studies have shown that the abnormity of DNA methylation，the abnormity of dendritic cells (DC), aberrant activation of Th17 cells and diminished Treg cells are critical to the development of both diseases. Recent studies have identified that miR-182 and FoxO1 are responsible for regulating the differentiation and function of Th17/Treg cells. We have found that DNA methylation status in the promoter of miR-182 was decreased, whereas the mRNA expression of miR-182 was elevated， which showed negative correlation significantly. Therefore, we speculate that decreased DNA methylation status in the promoter of miR-182 may cause the increased the mRNA expression of miR-182, and then was involved in the pathogenesis of uveitis by regulating its target genes. This project is designed to investigate the effect on the expression of FoxO1 and the differentiation of DC and Th17/Treg cells by promoting or suppressing the expression of miR-182 from three levels: case-control, vitro study and animal experiment. Collectively the aims of these studies are to disclose the key mechanisms of miR-182 and its promoter methylation status in pathogenesis of uveitis by regulating its target genes, which may provide a new target for the prevention and treatment of uveitis.

Behçet病和VKH综合征是我国致盲率最高的葡萄膜炎类型，DNA甲基化异常、树突状细胞功能异常、Th17细胞过度激活以及Treg细胞数量和功能降低是其发病基础。最近的研究发现miR-182能够调控Th17/Treg细胞的分化和功能。我们的预实验也发现，相对于正常对照，Behçet病和VKH综合征患者miR-182启动子甲基化水平降低，而miR-182 mRNA表达升高，二者呈显著负相关。由此推测miR-182启动子低甲基化可能导致miR-182的表达升高，进而通过调控下游靶基因参与葡萄膜炎的发病。本项目拟在前期研究基础上，从病例对照、离体研究和动物实验三个层面，采用促进和抑制miR-182表达的不同手段、观察其对靶基因表达以及对DC、Th17和Treg细胞分化三条路径的影响，全方位多层次阐明miR-182及其启动子甲基化状态参与葡萄膜炎发病的关键作用机制，为葡萄膜炎的防治提供新靶点。

白塞病（BD）和VKH综合征是我国致盲率最高的葡萄膜炎类型，Usher综合症是一种以耳聋和视网膜色素变性为特征的常染色体隐性遗传性疾病，Graves病(GD)是一种最常见的自身免疫性甲状腺疾病。本项目围绕BD、VKH综合征、Usher综合症和GD，开展了系列研究，主要研究结果包括以下6个方面：1. 从全基因组水平鉴定了与BD相关联的4332个差异DNA甲基化位点，其中547个位点在BD中呈现高甲基化，3785个位点在BD中呈现低甲基化；发现4个差异的DNA甲基化位点（cg03546163, cg25114611, cg23261343 and cg14290576）可以作为BD疾病诊断的生物标记物；发现BD患者中FKBP5的低甲基化能促进FKBP5基因的表达。2. 从全基因组水平鉴定了与VKH综合征相关联的561个差异DNA甲基化位点，其中254个位点在VKH综合征中呈现高甲基化，307个位点在VKH综合征中呈现低甲基化。3. 鉴定了MYO7A基因c.6070C>T (p.R2024X) 和c.5168+1G>A复合杂合突变会导致Usher综合征的发病。4. 发现IL2RA/rs2104286位点多态性与中国西南地区汉族人群GD和Graves眼病（GO）的易感性显著关联，它可能通过影响IL2RA基因mRNA的表达和细胞因子（IL-17，IL-10）分泌水平参与GD和GO的发生。5. 发现VDR/Apa Ⅰ位点多态性与中国西南地区汉族人群GD和GO遗传易感性显著相关，可能通过影响VDR基因mRNA表达和细胞因子（IL-17，TGF-β1）分泌进而参与GD和GO的发生。6. 从全基因组水平鉴定了与GD相关联的548个差异DNA甲基化位点，其中355个位点GD中呈现高甲基化，193个位点在GD中呈现低甲基化。综上所述，这些研究为BD、VKH综合征、Usher综合症和GD的防治提供了新的靶点。