CpG/TLR9信号转导对母胎界面免疫功能和妊娠结局影响的机制研究

Miscarriage and preterm birth are the most common complications of pregnancy. Intrauterine infection is one of the most common causes. The key to reduce miscarriage and preterm birth lies in the finding and blocking the signal transduction pathway which leads to intrauterine infection. Systemic or intrauterine bacterial infection results in excessive production of hypomethylated CpG DNA motifs that are recognized by TLR9. Based on our previous work, we found that CpG/TLR9 signaling influenced the immune activation on the maternal-fetal interface and led to miscarriage and preterm birth in immunodeficient mice, but not in widetype mice. With transcriptome analysis, flow cytometry and immunohistochemistry technology et al., this project is proposed to explore the abnormal mechanism in NOD mice during the CpG/TLR9-induced miscarriage and preterm birth, and to identify the defense mechanism against the intrauterine infection in wide-type mice. It ultimately seeks to explore the detailed mechanism of the effects of CpG/TLR9-induced inflammatory on pregnancy outcomes. The fruits of the research will make a deeper exploration in the fatal-maternal immune intolerance, thereby, helping lay the foundation of seeking more efficient targets for clinical intervention, treatment of spontaneous abortion and preterm birth.

流产和早产是最常见的妊娠并发症，而宫内炎症是导致妊娠中晚期流产和早产的最常见原因之一，因此发现并阻断导致宫内炎症的信号转导途径是减少流产和早产的关键。全身性或宫内细菌感染会产生大量低甲基化的CpG基序，被Toll样受体9（TLR9）识别。课题组前期研究发现CpG/TLR9信号转导可通过改变母胎界面免疫细胞的功能状态，诱导免疫缺陷孕小鼠发生流产和早产，而对野生型小鼠无影响，其作用机制尚不清楚。本项目拟以NOD和BALB/c小鼠为模型，应用RNA-Seq高通量测序、流式细胞、免疫组化等技术，探索：1）CpG/TLR9炎症反应引起NOD小鼠流产率和早产率增加的异常机制；2）野生型小鼠抵御CpG ODN模拟的宫内炎症，维持正常妊娠过程的防御机制。本课题的结果将进一步揭示母胎界面免疫耐受状态形成和维持的调控机制，为发展更有效的自然流产和早产临床防治技术提供理论基础和实践依据。

已知CpG/TLR9信号转导可通过改变母胎界面免疫细胞的功能状态，诱导免疫缺陷孕小鼠发生流产和早产，而对野生型小鼠无影响，其作用机制尚不清楚。本项目以NOD和BALB/c小鼠为模型，应用RNA-Seq高通量测序技术确定胎盘与蜕膜组织中的基因表达差异，当小鼠受到细菌感染时，首先激活的是抗微生物/抗菌体液免疫。WT小鼠体内由C1q和PPAR信号通路调节的巨噬细胞M2型极化，NOD小鼠巨噬细胞的M1型极化是两种小鼠在感染情况下出现不同妊娠结局的主要原因。利用中和抗体抑制的小鼠C1q功能发现，C1q在抵抗WT小鼠应对CpG ODN刺激，维持正常妊娠结局过程中发挥重要的促进作用。此外，收集人绒毛组织，通过免疫组化和Western blot实验验证STMN1和Prdx2的表达水平，利用细胞系通过基因敲除技术验证其对滋养层细胞增殖、凋亡和迁移的影响，并确定了TTP对STMN1的表达、c-Myc对Prdx2表达的调控。本课题的结果将进一步揭示母胎界面免疫耐受状态形成和维持的调控机制，为发展更有效的临床防治技术提供理论基础和实践依据。