RNA解旋酶DDX46调控蜕膜巨噬细胞炎症反应介导母胎界面免疫紊乱的机制研究

The immune inflammatory disorder of maternal fetal interface is an important factor to induce pregnancy related diseases, but the precisely regulation mechanism of immune response in decidual macrophage (dMϕ) is poorly understood. We previously found that RNA helicase DDX46 play an important role in the negative regulation of antiviral innate immunity (Nat Immunol, 2017), we newly discovered that DDX46 can negatively regulate the inflammatory cytokines expression in dMϕ, and the embryo resorption rate of DDX46+/- mice was significantly higher than that in wild-type control mice，but the mechanism is not clear. In this project we proposed to analyze the effect of DDX46 on the biological function of dMϕ, trophoblast cells and other immune cells in DDX46+/- mice. We found the target genes of DDX46 through the iCLIP and transcriptome sequencing methods, then found the key interaction proteins of DDX46 identified by mass spectrometry, to uncover the molecular mechanism of how DDX46 regulates inflammatory cytokines expression of dMϕ by recruiting key proteins to regulate target genes, we hope to elucidate a new mechanism of the regulation of maternal fetal interface immune disorder induced by DDX46 from the inflammation perspective，in order to explore a new way for clinical diagnosis and treatment of pregnancy related diseases mediated by inflammation.

母胎界面微环境中的免疫炎症紊乱是诱导妊娠相关疾病的重要因素，而有关调控蜕膜巨噬细胞（dMϕ）免疫炎症反应的机制却知之甚少。申请人以前发现RNA解旋酶DDX46在负调控抗病毒天然免疫中发挥重要作用(Nat Immunol, 2017)，新近发现DDX46能负调控dMϕ炎症细胞因子表达，且DDX46+/-孕母鼠的胚胎吸收率明显高于对照野生型孕鼠，而确切机制尚不清楚。本项目拟在DDX46+/-小鼠体内分析DDX46对dMϕ、滋养细胞和其它免疫细胞生物学功能的影响。通过iCLIP和转录组测序寻找与DDX46结合并受其调控的靶标基因，质谱鉴定与DDX46互作的关键蛋白，从而研究DDX46如何通过招募关键蛋白调控靶标基因进而调控dMϕ炎症细胞因子表达的分子机制，以期从炎症调控的角度阐明dMϕ中DDX46调控母胎界面免疫紊乱的新机制，为临床炎症失衡介导复发性流产等妊娠相关疾病的诊治提供新思路。

母胎界面微环境中的免疫炎症紊乱是诱导妊娠相关疾病的重要因素，蜕膜巨噬细胞（dMϕ）在调控免疫炎症中发挥关键作用，而目前精确调控dMϕ免疫炎症反应的表观分子机制却知之甚少。我们发现RNA解旋酶DDX46+/-孕母鼠的胚胎吸收率明显高于对照野生型小鼠，去除蜕膜巨噬细胞后能逆转这种效应。进一步发现DDX46能负调控dMϕ炎症细胞因子的表达，且DDX46敲减后与滋养细胞共孵育能抑制滋养细胞的迁移和侵袭。通过iCLIP发现DDX46能结合一系列的靶标RNAs（包括NFκB1），质谱鉴定发现与DDX46互作的关键蛋白为ALKBH5，发现DDX46能招募ALKBH5使得靶标NFκB1mRNA发生去m6A甲基化修饰，使得NFκB1mRNA发生核滞留，从而抑制NFκB1表达及其下游炎症细胞因子的表达，从而揭示了DDX46调控dMϕ炎症细胞因子表达的分子机制。从临床样本中发现，相比于正常早孕期蜕膜组织，DDX46在自然流产组织中的表达显著降低。我们从炎症调控的角度阐明dMϕ中DDX46调控母胎界面免疫紊乱的新机制，为临床炎症介导复发性流产等妊娠相关疾病的诊治提供新思路。