Analysis of the Angiopoietin/Tie2 signaling cascade with regard to vascular permeability and inflammation as well as study of the acute regulation of Tie2 expression in experimental sepsis

血管生成素/Tie2信号级联与血管通透性和炎症相关的分析以及实验性脓毒症中Tie2表达的急性调节研究

• 批准号: 248484453
• 负责人: Professor Dr. Sascha David
• 金额: --
• 依托单位: Klinik für Nieren- und Hochdruckerkrankungen
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/248484453?language=en
• 关键词: Analysis; Angiopoietin; Tie2; signaling; cascade

Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to infection. It is not the infection per se but rather the overwhelming response that leads to multiple organ failure and death. The endothelium pervades every organ and all its physiological processes can be affected in sepsis, thus systemic vascular changes have severe consequences for organ function. Ligands of the endothelial tyrosine kinase receptor Tie2 are markedly imbalanced in severe infections associated with vascular leakage, yet regulation of the receptor itself has been understudied in this context. We have observed that Tie2 expression dramatically drops within a few hours affecting down-stream signaling and vascular barrier function. Moreover, experimental suppression is sufficient to mimic the septic vascular phenotype. We hypothesize that Tie2 is a critical determinant of the hosts vascular response to sepsis and we will focus on 2 major aspects of its regulation: 1) The search for a common denominator of Tie2 suppression in various critical illnesses (hemorrhagic shock, sepsis, influenza, anthrax, etc.) led to the realization that all these conditions share a rather simple clinical symptom, i.e. hemodynamic shock and microvascular hypoperfusion. Translating this to the bench, we found that Tie2 transcription was highly depended on flow. Here, we will analyze the link between the flow-responsive GATA3 transcription factor and Tie2 expression upon flow in vitro and in vivo. Conditional endothelial specific knockouts and transgenics will be generated. 2) Tie2 expression is also affected by posttranslational modifications. We have observed extensive Tie2 shedding cross-species in an MMP14-dependent fashion. Besides pharmacological inhibition of MMP14 in murine experimental sepsis, we plan to identify the extracellular Tie2 cleavage site by mass spectrometry followed by experimental mutation and further functional analysis. If Tie2 shedding is indeed injurious or adaptive will be evaluated in a mouse model of overexpressed soluble (s)Tie2.

败血症是一种危及生命的器官功能障碍，由宿主对感染的反应失调引起。这不是感染本身，而是压倒性的反应，导致多器官衰竭和死亡。脓毒症患者血管内皮遍布于各个器官，其所有生理过程均受影响，因此全身血管的改变对器官功能有严重影响。内皮酪氨酸激酶受体Tie2的配体在与血管渗漏相关的严重感染中显着失衡，但在这种情况下，受体本身的调节尚未得到充分研究。我们观察到Tie2表达在几小时内急剧下降，影响下游信号传导和血管屏障功能。此外，实验抑制足以模拟脓毒性血管表型。我们假设Tie2是宿主血管对脓毒症反应的关键决定因素，我们将重点关注其调节的两个主要方面：1)寻找各种危重疾病（失血性休克、脓毒症、流感、炭疽等）中Tie2抑制的共同点，从而认识到所有这些疾病都有一个相当简单的临床症状，即血流动力学休克和微血管灌注不足。将其翻译到实验台上，我们发现Tie2转录高度依赖于流动。在这里，我们将分析流动响应GATA3转录因子与Tie2在体外和体内流动时的表达之间的联系。条件内皮特异性敲除和转基因将产生。2) Tie2的表达也受到翻译后修饰的影响。我们观察到Tie2以依赖mmp14的方式在物种间广泛脱落。除了MMP14在小鼠实验性脓毒症中的药理抑制作用外，我们计划通过质谱法确定细胞外Tie2切割位点，然后进行实验突变和进一步的功能分析。如果Tie2脱落确实是有害的或适应性的，将在可溶性Tie2过表达的小鼠模型中进行评估。