Role of endothelial JAK/STAT signaling in the regulation of vascular leakage

内皮 JAK/STAT 信号在血管渗漏调节中的作用

基本信息

批准号：
10451718
负责人：
Alejandro Pablo Adam
金额：
$ 36.45万
依托单位：
ALBANY MEDICAL COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-20 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10451718
关键词：
ADAMTS Actins Acute Address Affect Anti-Inflammatory Agents Blood Vessels Breeding Candidate Disease Gene Cell Communication Cessation of life Clinical Complex Conflict (Psychology)Cytokine Signaling Cytoskeleton Data Endothelial Cells Endothelium Enzymes Extravasation Feedback Functional disorder Future Gene Expression Gene Expression Regulation Genes Genetic Transcription Glycocalyx Grant Hyaluronan Hyaluronic Acid Hyaluronidase IL6 gene Immune response In Vitro Inflammation Inflammatory Inflammatory Response Interleukin-6 Janus kinase Knock-in Knock-out Knockout Mice Knowledge Lead Link Literature Lung Mediating Mediator of activation protein Metalloproteases Modeling Monomeric GTP-Binding Proteins Mus Mutation Organ Outcome Pathway interactions Permeability Pharmacology Phosphoric Monoester Hydrolases Phosphorylation Post-Translational Protein Processing Regulation Research Project Grants Role STAT3 gene Sepsis Septic Shock Signal Pathway Signal Transduction Stress Fibers Testing Therapeutic Tissues Transcriptional Regulation Vascular Permeabilities arm base cecal ligation puncture cytokine design healing improved in vivo long-term sequelae loss of function mortality new therapeutic target novel novel therapeutic intervention pathogen prevent response septic tissue injury tissue repair transcription factor translational potential

项目摘要

Sustained vascular leakage due to prolonged or exacerbated inflammation leads to organ damage, lasting sequelae and increased mortality. The mechanisms that mediate long-term loss of endothelial barrier function are poorly understood. We propose that transcriptional changes in the endothelium, and in particular STAT3- dependent gene regulation, may explain these long-term changes. STAT3 can promote opposing effects on the endothelium. While inhibition of its phosphorylation (mediated by JAK kinases) is anti-inflammatory and reduces endothelial permeability, endothelial STAT3 knockout actually increases vascular leakage. The central hypothesis of this grant is that prolonged STAT3 phosphorylation promotes a pro-inflammatory STAT3 transcriptional response that involves changes in the glycocalyx, including degradation of hyaluronic acid, and drastic alterations in the actin cytoskeleton. This hypothesis challenges the current understanding of the role of STAT3 in the endothelial inflammatory response by linking cytokine signaling to sustained vascular leakage via a novel mechanism. A better understanding of this pathway not only can explain multiple clinical observations, but also is expected to have a significant positive impact on future anti- inflammatory treatments by identifying novel pharmacological targets to prevent and/or revert the systemic vascular leakage without affecting other arms of the immune response that may be required for pathogen clearing or tissue healing. We will combine in vitro and in vivo experimentation in the three separate aims to: Identify the specific role of STAT3-dependent transcriptional mediators in endothelial barrier loss. We identified novel STAT3-dependent genes downstream of IL-6, including endothelial glycocalyx catabolic enzymes and modulators of the actin cytoskeleton. Loss of hyaluronan and actin stress fibers temporally correlate with IL-6- induced loss of barrier function in HUVEC. We will determine any causal roles for these candidate genes. Determine the requirement of endothelial STAT3 phosphorylation in promoting barrier function loss in vivo. Phosphorylated and non-phosphorylated STAT3 may have different and even opposing roles. We will use established models of sepsis to assess mice survival, vascular leakage and endothelial glycocalyx and cytoskeletal regulation of STAT3 (STAT3iEKO) and gp130iEKO knockout mice, as well as mice harboring an inducible, endothelial-specific knockin Y705F mutation (STAT3iE-Y705F). Determine the role of feedback loops leading to STAT3 sustained activation in barrier function. We will determine how an IL-6-induced sustained activation of STAT3 can lead to prolonged vascular leakage by interfering with negative feedback loops involving SOCS3 and TC45. We will identify potential post- translational modifications of the negative regulator SOCS3 that could lead to sustained STAT3 activation. To assess the role of these loops in vivo, we will create endothelial-specific SOCS3 knockouts by breeding SOCS3fl mice to cdh5-CreERT2 mice.

由于长期或加剧的炎症而导致的持续血管渗漏会导致器官损伤，后遗症和死亡率增加。介导内皮屏障功能长期丧失的机制人们了解甚少。我们认为内皮细胞的转录变化，特别是 STAT3- 依赖基因调控，可以解释这些长期变化。 STAT3可以促进相反的作用内皮细胞。虽然抑制其磷酸化（由 JAK 激酶介导）具有抗炎作用降低内皮通透性，内皮 STAT3 敲除实际上会增加血管渗漏。这该资助的中心假设是 STAT3 磷酸化延长会促进促炎 STAT3 转录反应涉及糖萼的变化，包括透明质酸和肌动蛋白细胞骨架的剧烈改变。这一假设挑战了当前的通过将细胞因子信号传导与 STAT3 联系起来，了解 STAT3 在内皮炎症反应中的作用通过一种新的机制持续血管渗漏。更好地理解这条途径不仅可以解释了多项临床观察结果，而且预计会对未来的抗- 通过识别新的药理学靶点来预防和/或恢复全身性炎症治疗血管渗漏而不影响病原体可能所需的其他免疫反应清除或组织愈合。我们将结合体外和体内实验来实现三个不同的目标：确定 STAT3 依赖性转录介质在内皮屏障丧失中的具体作用。我们确定了 IL-6 下游的新型 STAT3 依赖性基因，包括内皮糖萼分解代谢酶和肌动蛋白细胞骨架的调节剂。透明质酸和肌动蛋白应激纤维的损失与 IL-6- 暂时相关导致 HUVEC 屏障功能丧失。我们将确定这些候选基因的因果作用。确定内皮 STAT3 磷酸化在促进体内屏障功能丧失中的要求。磷酸化和非磷酸化的 STAT3 可能具有不同甚至相反的作用。我们将使用建立脓毒症模型来评估小鼠存活率、血管渗漏和内皮糖萼 STAT3 (STAT3iEKO) 和 gp130iEKO 敲除小鼠以及携带可诱导的内皮特异性敲入 Y705F 突变 (STAT3iE-Y705F)。确定反馈环路在屏障功能中导致 STAT3 持续激活的作用。我们将确定 IL-6 诱导的 STAT3 持续激活如何导致血管渗漏延长干扰涉及 SOCS3 和 TC45 的负反馈回路。我们将确定潜在的后负调节因子 SOCS3 的翻译修饰可能导致 STAT3 持续激活。到评估这些环在体内的作用，我们将通过育种创建内皮特异性 SOCS3 敲除 SOCS3fl 小鼠至 cdh5-CreERT2 小鼠。