SLC15A4介导Nod1依赖的NF-κB信号通路对SLE自身反应性炎症的调控作用机制

The final stage of the pathogenesis in systemic lupus erythematosus(SLE) is the development of the autoimmunity inflammation. It is the key to improve the therapeutic effect of SLE to specifically inhibit some pathways of the autoimmune inflammatory response. Studies have shown that NF-κB pathway play an important role in promoting autoimmune inflammation of SLE, but the exact regulatory mechanism in SLE is not clear. SLC15A4 modulates Nod1 signaling pathway and starts the proinflammatory response,which was found as susceptiblity gene in SLE with genome-wide association study.In our previous work,we found that differential expression of SLC15A4 in T lymphocyte of SLE.Based on it, we presume that SLC15A4 regulates autoreactive inflammation by Nod1-dependent NF-κB molecular pathway in SLE. In present project,we will study the single nucleotide polymorphism of SLC15A4 in the peripheral blood lymphocyte subsets from SLE patients. Then we analyze the relationship between SLC15A4 expression, its gene polymorphism and the disease activity,clinical phenotypes, expression of Nod1 and NF-κB-related inflammatory factors. In transfected Jurkat cells, to investigate the activation of the Nod1 pathway with up-regulation or down-regulation of SLC15A4 expression. In vivo, SLE disease activity will be evaluated after SLC15A4 expression is down-regulated in lupus mice and the mechanism of SLC15A4 modulating Nod1-dependent NF-κB pathway activation is further confirmed. The study will provide a new experimental basis to clarify the pathogenesis of autoimmune inflammation formation and to may reveal novel therapeutic targets in SLE.

自身免疫性炎症形成是SLE发病过程的最终环节，特异性阻断炎症反应是提高SLE疗效的关键。NF-κB通路在SLE自身免疫性炎症形成中发挥重要作用，但确切调控机制尚不明确。全基因扫描发现SLC15A4为SLE易感基因，在某些炎症性疾病中SLC15A4通过Nod1通路启动促炎症应答。我们前期研究发现SLC15A4在SLE患者T细胞中差异表达，由此提出SLC15A4通过Nod1依赖NF-κB信号通路调控SLE炎症形成的假说。本课题拟先研究SLE患者SLC15A4基因多态性与疾病活动性、Nod1依赖NF-κB通路相关炎性因子变化的相关性；体外试验观察干预SLC15A4表达后Nod1通路分子活化情况；进一步通过下调狼疮鼠体内SLC15A4表达，验证其在炎症反应形成中的作用，最终明确SLE发病中SLC15A4调控Nod1依赖NF-κB活化的作用机制，为阐明SLE的发病机制和发现新的治疗靶点提供实验依据。

系统性红斑狼疮（SLE）是一种累及多器官系统的自身免疫性疾病，其病因尚不十分清楚，已知的因素包括基因水平的改变、表观遗传学的调控以及环境和激素等相互影响。研究发现SLC15A4的2个SNPs（rs10847697，rs1385374）位点在SLE患者与对照组之间有显著性的差异，推测其可能通过Nod1通路启动促炎症应答，在SLE的自身免疫性炎症形成过程中起着重要作用。我们收集中国西南部地区汉族人口中的355例SLE患者及375例健康者的外周血，对SLC15A4的基因多态性与SLE的关系进行相关性分析。结果表明：1. SLC15A4基因多态性的等位基因频率分布：在18个位点中，其中5个在SLE患者和健康对照之间有显著性差异。2. SLC15A4基因多态性的基因型分布频率：在共显性模型分析中，rs3765108中的基因型AG显著高于基因型AA(P=0.019，OR=1.447，95%CI=1.063-1.970)，rs7308691中的基因型AT分布频率明显高于基因型AA分布频率(P=0.049，OR=1.645，95%CI=1.000-2.705)。在显性模型中，rs1385374与SLE的发病显著相关（TT+CTvsCC，P=0.042，OR=1.1363，95%CI =1.010-1.840）。3. SLC15A4的不同SNP位点与SLE的不同疾病表型具有相关性，其中盘状红斑及狼疮肾炎与5个SNP位点均存在显著相关性。4. 连锁不平衡分析：对18个SNP位点进行连锁不平衡分析发现存在2个单体型块。单体型块1长度为1kb并包括3个SNPs，其中频率最高的单体型是AAT；单体型块2长度为29kb，包含以下15个SNPs，其中频率最高的单体型是TTTGGCACAAGCTTC和CCCGAGTCAAATTTC，后者与SLE的发病相关(P = 0.033, OR 1.376, 95% CI 1.026-1.845)。5．SLC15A4基因mRNA及蛋白在SLE患者PBMC中的表达水平高于健康对照组。由此可见，SLC15A4的5个SNP位点（rs959989、rs1385374、rs983492、rs12298615和rs10847697）在SLE中存在显著差异，可能是SLE的发病危险因子之一， SLC15A4基因的单核苷酸多态性与中国汉族人群SLE的发病密切相关。