Signal transduction mechanisms that mediate normal and pathologic angiogenesis

补助金名称 介导正常和病理性血管生成的信号转导机制 摘要 肺动脉高压（PAH）是一种进行性疾病，其特征是血管收缩、细胞凋亡、肺动脉高压和肺动脉高压。 增生和纤维化，导致肺动脉压升高，并经常引起右心衰竭 与死这种病无药可治。因此，新的机制研究和新的治疗策略 是迫切需要的。PAH患者血浆细胞因子升高是炎症的标志。作为主要 作为效应细胞，单核细胞释放细胞因子并浸润在肺的血管周围区域。单核细胞耗竭 减轻PAH动物模型中的血管重塑和血液动力学变化。这一证据表明， 单核细胞和内皮细胞（EC）之间的通信。然而，根本的机制不是 很好理解。目前的建议的目的是确定切割的细胞外结构域的作用， δ样蛋白4（exDll 4）在肺动脉高压发病中的作用及其机制在体外，我们发现钙蛋白酶1可以 裂解Dll 4。我们还发现TNF α和IL-1 β增加人单核细胞中D114的表达，D114的切割， 和exD 114从单核细胞释放。此外，我们发现重组exDll 4显著增加了 细胞凋亡和屏障功能降低。在寻找作用机制时，我们发现exDll 4与 与完整的Dll 4相互作用，这种相互作用阻止Dll 4与Notch 1结合，从而抑制Notch 1活化。在体内， 我们发现PAH小鼠单核细胞中Dll 4表达和exDll 4释放显著升高， 在PAH患者中。然而，Notch 1信号在小鼠和大鼠PAH进展期间在肺中减少。 基于这些发现，我们假设来源于单核细胞的exD 114对于PAH进展是至关重要的， 诱导细胞凋亡和损害肺EC的屏障功能。从机制上讲，exD 114与 Dll 4以阻止Dll 4和Notch 1的结合，阻断Notch 1信号传导。为了验证我们的假设，我们建议 三个目标。目标1。明确单核细胞Dll 4表达和exDll 4释放的调控机制， 呸。目标2.明确exDll 4-Notch 1在肺EC中的生物学功能及其分子机制。目的 3.表征抑制单核细胞Dll 4对PAH进展的治疗作用。完成这些 目的是：1）填补关于血管重塑和EC功能障碍机制的知识空白， 由单核细胞衍生的exDll 4介导的PAH的发病机制; 2）揭示Dll 4切割的调节;和3） 发明了通过靶向单核细胞Dll 4治疗PAH新策略。

Title of the grant Signal transduction mechanisms that mediate normal and pathologic angiogenesis Abstract Pulmonary arterial hypertension (PAH) is a progressive disease, characterized by vasoconstriction, cell proliferation, and fibrosis, leading to elevated pulmonary arterial pressure and often causing right heart failure and death. There is no cure for this disease. Therefore, novel mechanistic studies and new therapeutic strategies are urgently needed. Elevation of plasma cytokines in PAH patients is a hallmark of inflammation. As the major effector, monocytes release cytokines and infiltrate in perivascular regions of the lung. Depletion of monocytes attenuates vascular remodeling and hemodynamic changes in PAH animal models. This evidence implies a communication between monocytes and endothelial cells (ECs). However, the underlying mechanisms are not well understood. The objective of the current proposal is to define the role of extracellular domain of cleaved delta like 4 (exDll4) in the pathogenesis of PAH and its mechanisms. In vitro, we discovered that calpain1 can cleave Dll4. We also found that TNF and IL-1 increases Dll4 expression, Dll4 cleavage in human monocyte, and exDll4 release from monocytes. Furthermore, we found that recombinant exDll4 significantly increased apoptosis and decreased barrier function in ECs. Seeking the mechanism of action, we found exDll4 associated with intact Dll4, and this interaction prevents Dll4 binding to Notch1 and thus inhibits Notch1 activation. In vivo, we found that Dll4 expression and exDll4 release in monocyte are significantly elevated in PAH mice as well as in PAH patients. However, Notch1 signaling is decreased in lungs during PAH progression in mice and rats. Based on these findings, we hypothesize that exDll4 derived from monocytes is crucial for PAH progression by inducing apoptosis and impairing barrier function in lung EC. Mechanistically, exDll4 forms a heterodimer with Dll4 to prevent the association of Dll4 and Notch1, blocking Notch1 signaling. To test our hypothesis, we propose three aims. Aim 1. Define the regulatory mechanisms of Dll4 expression and exDll4 release from monocyte in PAH. Aim 2. Determine the biological function of exDll4-Notch1 in lung EC and its molecular mechanisms. Aim 3. Characterize the therapeutic effects of inhibiting monocyte Dll4 on PAH progression. Accomplishing these aims will 1) fill the knowledge gap regarding the mechanisms of vascular remodeling and EC dysfunction in the pathogenesis of PAH mediated by monocyte derived exDll4; 2) reveal the regulation of Dll4 cleavage; and 3) invent a novel therapeutic strategy for PAH by targeting monocyte Dll4.