Mechanisms for Human TLR8 induced pregnancy loss in a mouse model of SLE

系统性红斑狼疮(SLE)是一种多系统的自身免疫性疾病，可导致 生活质量和早期死亡率显著下降。遗传学研究已经确定 激活先天免疫系统，过量产生I型干扰素是主要原因 致因途径。RNA敏感的Toll样受体是先天免疫的重要保护性受体 免疫系统，但也与自身免疫有关，因为它们可以识别核酸 从受损细胞和含有核酸的免疫复合体中释放出来。功能性 内体识别RNA的固有受体TLR7和TLR8基因多态性易患 对系统性红斑狼疮和过量的小鼠TLR7和人TLR8可加速狼疮小鼠模型。人类 TLR8在功能上与TLR7不同，因为它识别不同的配体并表达 不同的免疫细胞亚群。 TLR8在小鼠中不是RNA传感器，因为5个氨基酸的缺失会减弱其 RNA结合位点。因此，我们将人TLR8 BAC转基因(HuTLR8)培育成小鼠，在其中 易感基因Sle1是一种轻度狼疮表型。转基因的表达 是否受到生理调节，使得huTLR8在髓系树突状细胞和 单核细胞，但在淋巴细胞中的水平低50-100倍。我们发现huTLR8诱导了一种抗- 导致自发性胎盘炎症的Sle1小鼠的磷脂样综合征，胎儿 高达60%的女性大坝出现吸收和晚期妊娠丢失。在本提案中，我们将 通过研究huTLR8作为一种信号通路的作用，测试huTLR8介导的妊娠丢失的机制 胎盘滋养层细胞和免疫细胞中的RNA传感器。 在目标1中，我们将研究自身抗体在胎盘损伤中的作用。 并确定在B细胞中huTLR8的表达是否会导致 产生具有增强致病性的自身抗体。在目标2中，我们将确定时间 胎盘损伤的特征，母体细胞和胎儿细胞中huTLR8对 损伤与抗磷脂抗体激活的胎盘滋养层细胞是否释放 增强局部炎症的huTLR8配体。在目标3中，我们将确定髓系细胞 来自huTLR8tg小鼠的细胞本质上比它们的野生型细胞更容易发炎 从而放大抗体介导的胎盘损伤的效应器反应。 这些实验应该可以让我们区分huTLR8在母体细胞中的作用， 滋养层细胞和免疫细胞亚群在诱导胎盘损伤中的作用 妊娠丢失。我们的研究将告诉我们，特定的huTLR8拮抗剂或 应该测试huTLR8配体预防或治疗胎盘损伤的能力。

Systemic lupus erythematosus (SLE) is a multisystem, autoimmune disease that causes a significant decrease in quality of life and early mortality. Genetic studies have identified activation of the innate immune system, with excess production of Type I interferons as a major causative pathway. RNA-sensing Toll-like receptors are crucial protective receptors of the innate immune system but are also implicated in autoimmunity because they can recognize nucleic acids released from damaged cells and within nucleic acid containing immune complexes. Functional polymorphisms of the endosomal RNA-recognizing innate receptors TLR7 and TLR8 predispose to SLE and excess mouse TLR7 and human TLR8 accelerate lupus in mouse models. Human TLR8 differs in its function from TLR7 because it recognizes different ligands and is expressed on different immune cell subsets. TLR8 is not an RNA sensor in mice because of a 5 amino acid deletion that attenuates its RNA-binding site. We therefore bred a human TLR8 BAC transgene (huTLR8) into mice in which a mild lupus phenotype is conferred by the susceptibility locus Sle1. Expression of the transgene is physiologically regulated such that huTLR8 is expressed at high levels in myeloid DCs and monocytes but at 50-100-fold lower levels in lymphocytes. We found that huTLR8 induces an anti- phospholipid like syndrome in Sle1 mice that leads to spontaneous placental inflammation, fetal resorptions and late-term pregnancy loss in up to 60% of female dams. In this proposal we will test the mechanism for huTLR8 mediated pregnancy loss by addressing the role of huTLR8 as a sensor of RNA in placental trophoblasts and immune cells. In Aim 1 we will examine the role of autoantibodies in placental injury in mice with or without huTLR8 expression and determine whether huTLR8 expression in B cells results in the production of autoantibodies with enhanced pathogenicity. In Aim 2 we will determine the timing and characteristics of placental injury, the contribution of huTLR8 in maternal vs. fetal cells to injury and whether placental trophoblasts activated by anti-phospholipid antibodies release huTLR8 ligands that enhance local inflammation. In Aim 3 we will determine whether myeloid cells from huTLR8tg mice are intrinsically more pro-inflammatory than those of their wild-type counterparts and therefore amplify the effector response to antibody mediated placental injury. These experiments should allow us to distinguish the role of huTLR8 in maternal cells, trophoblasts and immune cell subsets in the induction of placental damage that leads to pregnancy loss. Our studies will inform us whether specific huTLR8 antagonists or antagonists of huTLR8 ligands should be tested for their ability to prevent or treat placental injury.