紫外线诱导系统性红斑狼疮DNA甲基化的分子机制研究

Systemic lupus erythematosus (SLE) is the prototype of autoimmune disease. At present, the etiology and pathogenesis is still unknown. It is believed that the genetic factor, environmental factors and their interaction cause abnormal immune, eventually leading to multiple systemic organ damage. The level of DNA methylation in epigenetic plays an important role in SLE pathogenesis, and is a research hotspot in recent years. SLE exists abnormal methylation. And high expression of many T cell surface molecules is relevant to the low promoter methylation. Some studies have shown that environmental factors can affect the level of DNA methylation, especially the ultraviolet. On the basis of our previous experiments, in the present study, we aim to explore the molecular mechanisms of DNA hypomethylation of ultraviolet-induced SLE, by test the level of DNA methylation in CD4 + T cells, the methyltransferase enzyme mRNA and protein levels, the methylation transferase activity and the MAPK signal transduction pathway before and after different doses of ultraviolet radiation in SLE and healthy control groups; compare the changes of gene expression profiles before and after ultraviolet irradiation using the human genome chip, and test the gene promoter methylation regional CpG island, to provide a new target and way in the treatment of SLE.

系统性红斑狼疮（SLE）是自身免疫病的原型，目前其病因和发病机制仍然未明，一般认为是遗传因素、环境因素及其交互作用引起免疫异常，最终导致多器官、系统损害。表观遗传学中DNA甲基化水平的改变在SLE发病机制中的起重要作用，且是近年来研究的热点，SLE存在甲基化异常，多种T细胞表面分子高表达与启动子低甲基化有关。已有研究表明，环境因素能够影响DNA甲基化水平，环境因素中紫外线对SLE的影响尤为重要。本课题拟在近期研究基础上，通过检测经不同剂量紫外线照射前后SLE和健康对照CD4+T细胞的DNA甲基化水平，甲基化转移酶mRNA水平和蛋白质水平,甲基化转移酶的活性以及MAPK信号转导的检测，探讨紫外线诱导系统性红斑狼疮DNA低甲基化的分子机制；采用人类全基因组芯片比较紫外线照射前后基因表达谱的变化，检测基因启动子区域CpG岛甲基化情况，为SLE的治疗提供新的靶点和途径。

研究证实表观遗传调控参与系统性红斑狼疮（SLE）的发病机制，而DNA甲基化是一种主要表观遗传修饰形式。本研究检测紫外线照射前后SLE患者CD4+T细胞DNA甲基化水平，并采用real-time PCR和western blot检测CD4+T细胞DNA甲基化转移酶DNMT1和DNMT3a的表达情况。结果表明SLE活动期患者DNA甲基化水平显著降低，经50mJ/cm2紫外线照射后，SLE活动组及稳定组的DNA甲基化水平与对照组相比均显著降低。甲基化转移酶DNMT1 mRNA表达水平在SLE活动、SLE稳定和对照组中没有显著差异，50 mJ/cm2 紫外线照射可显著降低SLE活动组和SLE稳定组CD4+T细胞DNMT1 mRNA的表达，经100 mJ/cm2紫外线照射后稳定组和健康对照组DNMT1蛋白表达下降。不同剂量紫外线照射后，CD4+T细胞DNMT3A mRNA的表达变化在三组中无统计学差异，DNMT3A蛋白表达量在健康对照和SLE活动组中均有所下降，但都没有达到统计学差异。适当剂量紫外线照射可降低CD4+T细胞DNMT1的表达，但对DNMT3A的表达无显著影响。以上结果提示CD4+T细胞甲基化水平可能参与SLE的发病。