Role of NLRP3 signals in ischemia/reperfusion-induced organ injury

项目概述：该提案评估了细胞质固有的细胞内信号通路如何产生， 免疫受体NLRP 3在肾小管上皮细胞中受到调节。肾小管上皮细胞在肾脏疾病中起着重要作用。 缺血性肾损伤，我们的实验室和其他实验室已经表明，整体NLRP 3阻断可以防止实验性肾损伤， 缺血/再灌注损伤。NLRP 3信号传导经典地导致细胞死亡（焦亡），然而替代性NLRP 3 最近已经鉴定了导致IL 1b/IL 18分泌而没有细胞死亡的信号传导途径。因此，选择性地 靶向NLRP 3介导的细胞死亡信号可能会防止肾小管损伤，同时仍然保留宿主的能力， 分泌IL 1b/IL 18并产生所需的宿主防御反应（例如，抗微生物反应）。该项目高度 对于缺血性肾损伤的靶向治疗的发展具有重要意义，缺血性肾损伤在住院患者中经常发生。 患者和所有用于移植的器官。 广泛/长期目标：拟议研究的长期目标是确定NLRP 3如何有助于 肾脏中的损伤组织反应以及如何有效和选择性地靶向其信号传导。 具体目的：本提案的具体目的是检验细胞质模式识别 受体NLRP 3可以特异性靶向肾小管上皮细胞，以选择性地阻断NLRP 3介导的肾 肾小管细胞死亡，同时保留IL 1b/IL 18应答。目的1定义了驱动NLRP 3信号传导的机制 近端肾小管细胞中的通路。目的2确定NLRP 3信号传导如何可以选择性地靶向于细胞中。 近端肾小管细胞目的3确定NLRP 3信号传导及其选择性阻断如何影响肾脏 肾小管损伤/体内IL 1b/IL 18分泌，当NLRP 3表达被分离到肾小管上皮时。 研究设计和实现既定目标的方法：目标1比较和对比NLRP 3的激活 在人类和小鼠近端肾小管细胞中，并将确定NLRP 3激活如何促进细胞凋亡 和/或IL 1b/IL 18的产生。目的2将确定如何在细胞中靶向特定的NLRP 3激活途径。 这两个物种，以防止细胞死亡和保护细胞因子分泌。目标3将侧重于了解NLRP 3 在体内仅限于肾小管上皮细胞，可靶向预防肾小管损伤而不损害IL 1b/IL 18。 在第三个目标中，将使用两种不同的NLRP 3表达模型;一种是肾小管表达条件性的NLRP 3。 NLRP 3的功能丧失和另一种存在NLRP 3的超活化形式。这里的研究将决定 经典NLRP 3介导的细胞死亡信号是否可以与非经典信号解偶联以防止细胞死亡。 死亡，同时保持IL 1b/IL 18的分泌。 项目的健康相关性：如果本提案的目标得到满足，我们将了解NLRP 3（被认为是一种 中枢细胞应激检测器）可以被调节以靶向常见疾病，肾缺血/再灌注损伤。这 知识对于开发合理的靶向治疗以预防或改善肾损害至关重要。 在预期缺氧的临床情况下的缺血/再灌注损伤。

Project Summary: This proposal evaluates how intracellular signaling pathways generated by the cytoplasmic innate immune receptor NLRP3 are regulated in renal tubular epithelial cells. Renal tubule epithelial cells play a central role in ischemic kidney injury, and our lab and others have shown that global NLRP3 blockade can prevent experimental renal ischemia/reperfusion injury. NLRP3 signaling classically results in cell death (pyroptosis), however alternative NLRP3 signaling pathways have recently been identified that lead to secretion of IL1b/IL18 without cell death. Thus, selectively targeting NLRP3-mediated cell death signaling might prevent renal tubule injury while still preserving the host’s ability to secrete IL1b/IL18 and mount needed host defense responses (e.g., antimicrobial responses). The project is highly significant for the development of targeted therapeutics for ischemic kidney injury, which occurs frequently in hospitalized patients and in all organs procured for transplantation. Broad/long-term objectives: The long-term goals of the proposed research are to define how NLRP3 contributes to injurious tissue responses in the kidney and how its signaling can be effectively and selectively targeted. Specific Aims: The specific objective of this proposal is to test the hypothesis that the cytoplasmic pattern recognition receptor NLRP3 can be specifically targeted in renal tubular epithelial cells to selectively block NLRP3-mediated renal tubule cell death, while preserving IL1b/IL18 responses. Aim 1 defines the mechanisms that drive NLRP3 signaling pathways in proximal renal tubule cells. Aim 2 determines how NLRP3 signaling can be selectively targeted in the proximal renal tubule cells. Aim 3 determines how NLRP3 signaling, and its selective blockade, influences renal tubular injury/IL1b/IL18 secretion in vivo, when NLRP3 expression is isolated to the renal tubule epithelium. Research Design and Methods for Achieving the Stated Goals: Aim 1 compares and contrasts activation of NLRP3 in human and murine proximal renal tubular cells and will determine how NLRP3 activation contributes to pyroptosis and/or production of IL1b/IL18. Aim 2 will determine how specific NLRP3 activation pathways can be targeted in the two species to prevent cell death and preserve cytokine secretion. Aim 3 will focus on understanding how NLRP3 restricted to the renal tubule epithelium in vivo can be targeted to prevent renal tubular injury without impairing IL1b/IL18. In the third aim, two different models of NLRP3 expression will be used; one where the renal tubules express a conditional loss-of-function of NLRP3 and another where there is a hyper-activatable form of NLRP3. The studies here will determine whether canonical NLRP3-mediated cell death signaling can be uncoupled from noncanonical signaling to prevent cell death while preserving secretion of IL1b/IL18. Health Relatedness of Project: If the aims of this proposal are met, we will learn how activation of NLRP3 (deemed a central cellular stress detector) can be regulated to target a common disease, renal ischemia/reperfusion injury. This knowledge is crucial for the development of rational targeted therapies for prevention or amelioration of renal ischemia/reperfusion injury in clinical situations where hypoxia is anticipated.