Mechanisms regulating fetal membrane and neutrophil responses to infection

摘要/摘要 绒毛膜炎-胎膜（FM）的炎症-以中性粒细胞浸润为特征， 早产的主要危险因素。即使在没有早产的情况下，绒毛膜炎也可能对胎儿有害。 胎儿尽管细菌感染、绒毛膜炎和早产之间有很强的相关性， 所涉及的机制尚未完全了解。通过表达先天免疫模式识别 受体（PRR）、Toll样受体（TLR）和Nod样受体（NLR），FM具有逃避和 防止感染。然而，取决于信号传导和调节的性质，这些保护性的信号传导和调节可能会导致细胞凋亡。 免疫机制可产生可导致病理学的炎性环境。特别地，IL-8是一种免疫调节剂。 主要的中性粒细胞化学引诱物和炎性体介导的IL-1b是组织损伤的主要诱导物， 早产的中介。我们已经发现绒毛膜隔室是FM IL-8和IL-10的主要部位。 响应细菌脂多糖（LPS）、肽聚糖（PDG）和胞壁酰二肽的1b产生 (MDP)分别激活TLR 4、TLR 2和Nod 2。而TLR和NLR可以直接激活 信号通路导致炎性细胞因子/趋化因子的产生，有可能产生更多的 这些过程的复杂调制、调节和微调以及所产生的响应类型， 尤其是IL-1b。该补助金的重点是要求两个或更多的PRR依次启动 通过新型中间体暴露于细菌触发物的FM中。microRNA（miRs）的非经典家族 激活ssRNA传感器TLR 7和TLR 8，以引发炎症反应。这些miR也可以是 携带于外泌体中并递送至靶细胞中的TLR 7或TLR 8。我们的初步数据支持这个概念 TLR 8激活的miR-146 a-3 p可能作为一种新的中间信号，驱动FM趋化性， 对细菌TLR和NLR激动剂的炎症反应。我们也有初步数据表明， 含有TLR 8活化miR的FM衍生的外泌体触发中性粒细胞活化和中性粒细胞释放 细胞外陷阱最后，我们发现FM组织和循环外泌体TLR 8激活的miR-146 a-3 p是 在早产妇女中升高。基于此，我们的中心假设是TLR 8激活miR 介导FM趋化性IL-8和炎性小体介导的炎性IL-1b对细菌 触发器，并通过它们的释放和交付通过外泌体活跃的母体中性粒细胞。这导致 导致母胎界面炎症，增加绒毛膜羊水炎的风险。为了验证这一点，我们 具体目标是确定： 目标1。TLR 8激活miR在暴露于细菌触发物后介导FM趋化性IL-8应答。 目标二。TLR 8激活miR有助于FM炎性小体激活和炎性IL-1b产生。 目标3。含有TLR 8活化miR的FM外泌体诱导嗜中性粒细胞活化。

Summary/Abstract Chorioamnionitis - inflammation of the fetal membranes (FM) - is characterized by neutrophil infiltration and is a major risk factor for preterm birth. Even in the absence of prematurity, chorioamnionitis can be detrimental to the fetus. Despite a strong association between bacterial infection, chorioamnionitis, and preterm birth, the mechanisms involved are not fully understood. Through expression of the innate immune pattern recognition receptors (PRR), Toll-like receptors (TLRs) and Nod-like receptors (NLRs), FMs have strategies to evade and protect against infection. However, depending upon the nature of signaling and regulation, these protective immune mechanisms may create an inflammatory milieu that can contribute to pathology. In particular, IL-8 is a major neutrophil chemoattractant and inflammasome-mediated IL-1b is a major inducer of tissue injury and mediator of preterm birth. We have found that the chorionic compartment is the primary site of FM IL-8 and IL- 1b production in response to bacterial lipopolysaccharide (LPS), peptidoglycan (PDG), and muramyl dipeptide (MDP) which activate TLR4, TLR2, and Nod2, respectively. While TLRs and NLRs can directly activate signaling pathways leading to inflammatory cytokine/chemokine production, there is the potential for far more complex modulation, regulation, and fine-tuning of these processes and the type of responses generated, particularly for IL-1b. This grant focusses on the requirement of two or more PRRs to be activated sequentially in FMs exposed to bacterial triggers via novel intermediates. A non-classical family of microRNAs (miRs) activate the ssRNA sensors, TLR7 and TLR8, to elicit an inflammatory response. These miRs can also be carried in exosomes and delivered to TLR7 or TLR8 in target cells. Our preliminary data supports the concept that TLR8-activating miR-146a-3p may acts as a novel intermediate signal that drives FM chemotactic and inflammatory responses to bacterial TLR and NLR agonists. We also have preliminary data demonstrating that FM-derived exosomes containing TLR8-activating miRs trigger neutrophil activation and release of neutrophil extracellular traps. Finally, we found that FM tissue and circulating exosomal TLR8-activating miR-146a-3p is elevated in women with preterm birth. Based on this, our central hypothesis is that TLR8-activating miRs mediate FM chemotactic IL-8 and inflammasome-mediated inflammatory IL-1b in response to bacterial triggers, and through their release and delivery via exosomes active maternal neutrophils. This leads to inflammation at the maternal-fetal interface, increasing the risk for chorioamnionitis. To test this, our specific aims are to determine if: Aim 1. TLR8-activating miRs mediate a FM chemotactic IL-8 response after exposure to bacterial triggers. Aim 2. TLR8-activating miRs contribute to FM inflammasome activation and inflammatory IL-1b production. Aim 3. FM exosomes containing TLR8-activating miRs induce neutrophil activation.