Regulation of endothelial cell quiescence by Zeb1 as a regulator of angiogenesis

Zeb1 作为血管生成调节剂对内皮细胞静止的调节

• 批准号: 2274901
• 金额: --
• 依托单位: University of Nottingham
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2274901
• 关键词: Regulation; endothelial; cell; quiescence; Zeb1

The growth of new blood vessels from the pre-existing vasculature (angiogenesis) is an important physiological process driven by hypoxia. All blood vessels are lined by endothelial cells (EC) which under normal conditions form a smooth monolayer, which is non-proliferative, non-motile, tightly controlled junctions, non-adherent surface with no inflammation or leukocyte binding. Under these conditions the endothelium is in a state of 'quiescence' allowing efficient oxygen and other solute exchange into the interstitium and respiring tissues. The EC maintains this quiescent phenotype through the activity of a combination of epigenetic, surface receptor and junctional receptor activity, metabolic and of course transcriptional control of key genes. As tissues change their metabolic needs, the endothelium responds. A responding EC can be considered activated and depending on the specifics of the activation, can result in a phenotypic remodelling causing increased leukocyte adherence, increased inflammatory marker expression, proliferation, motility and significant changes in signalling pathways 1. Endothelial cell dysfunction occurs across a wide variety of diseases. During diabetes the altered microenvironment leads to EC dysfunction resulting in hypoxia and inflammation which results in excessive vascular leakage and growth. Diabetic retinopathy is a frequent complication of diabetes in which vessel growth in the retina results in loss of vision. In cardiovascular diseases (including coronary and peripheral vascular disease, stroke and diabetes) and vascular related diseases (neurodegeneration cancer, arthritis and sepsis) the EC is activated, and disease progression results in vascular remodelling, inducing the growth of new vessels (angiogenesis) or inflammatory conditions. Understanding how the quiescence-activation switch occurs will enable modification of this process for therapeutic benefit: if we can understand how to restore (or generate methods to maintain) EC quiescence we will be able to inhibit undesired EC activation. We have recently identified Zeb1 (a transcription factor with known roles in tumour progression and epithelial to mesenchymal transition) as being expressed in quiescent ECs. This was originally identified in vitro, and subsequently in vivo using the angiogenic mouse retina as an experimental model. Using inducible and endothelial specific Zeb1 knockout mice we have demonstrated preliminary evidence to show mice spontaneously undergo aberrant endothelial growth in adult retinae and increase solute leakage during choroidal neovascularisation. These preliminary data suggest altered Zeb1 signalling contributes to retinal angiogenesis and vessel leakage in adult mice, which could be important in the progression of DR. Therefore, this project will aim to investigate angiogenic remodelling in a series of developmental and pathological angiogenesis models. Aim 1: Does Zeb1 alter angiogenic phenotype in development? Zeb1 will be knocked out using a EC specific, conditional transgene. Retinal angiogenesis will be quantified in neonatal mice, and adult mice. Pathological angiogenesis will be induced using hindlimb ischaemia and choroidal neovascularisation models. Characterisation of EC phenotypes and interactions with angiogenesis modulating leukocyte populationsAim 2: Contribution of EC Zeb1 to the progression of diabetic retinopathy. Diabetes will be induced within the inducible EC specific KO mice and retinal permeability will be measured in vivo. Characterization of EC phenotype will be made by immunohistochemistry and molecular characterisation Aim 3: Is Zeb1 regulated angiogenesis VEGF dependent? Signalling pathways linking VEGFR2, hypoxia and glycolysis will be explored in vitro. This will also be explored in vivo with the use of VEGF antibodies to prevent VEGF stimulation within the EC Zeb1 KO diabetic model.

从预先存在的脉管系统生长新血管（血管生成）是由缺氧驱动的重要生理过程。所有血管都衬有内皮细胞（EC），其在正常条件下形成光滑的单层，其是非增殖的、非运动的、严格控制的连接、非粘附表面，没有炎症或白细胞结合。在这些条件下，内皮处于“静止”状态，允许有效的氧气和其他溶质交换到呼吸组织中。EC通过表观遗传、表面受体和连接受体活性、代谢以及当然关键基因的转录控制的组合的活性来维持这种静止表型。当组织改变其代谢需求时，内皮细胞会做出反应。响应的EC可以被认为是激活的，并且根据激活的具体情况，可能导致表型重塑，导致白细胞粘附增加、炎症标志物表达增加、增殖、运动性和信号通路的显着变化1。内皮细胞功能障碍发生在多种疾病中。在糖尿病期间，改变的微环境导致EC功能障碍，导致缺氧和炎症，这导致过度的血管渗漏和生长。糖尿病视网膜病变是糖尿病的常见并发症，其中视网膜中的血管生长导致视力丧失。在心血管疾病（包括冠状动脉和外周血管疾病、中风和糖尿病）和血管相关疾病（神经变性癌症、关节炎和败血症）中，EC被激活，并且疾病进展导致血管重塑，诱导新血管的生长（血管生成）或炎性病症。了解静止-激活开关是如何发生的，将能够修改这一过程以获得治疗益处：如果我们能够了解如何恢复（或产生维持）EC静止的方法，我们将能够抑制不希望的EC激活。我们最近发现Zeb 1（一种在肿瘤进展和上皮间质转化中具有已知作用的转录因子）在静止的EC中表达。这最初是在体外确定的，随后在体内使用血管生成小鼠视网膜作为实验模型。使用诱导型和内皮特异性Zeb 1基因敲除小鼠，我们已经证明了初步的证据，表明小鼠自发地经历异常的内皮生长在成年视网膜和增加溶质渗漏脉络膜新生血管形成。这些初步数据表明，改变Zeb 1信号有助于视网膜血管生成和血管渗漏在成年小鼠，这可能是重要的DR的进展，因此，该项目的目的是研究血管生成重塑在一系列的发展和病理血管生成模型。目的1：Zeb 1是否改变发育中的血管生成表型？将使用EC特异性条件转基因敲除Zeb 1。将在新生小鼠和成年小鼠中定量视网膜血管生成。将使用后肢缺血和脉络膜新生血管形成模型诱导病理性血管生成。EC表型的特征及其与血管生成调节白细胞群的相互作用目的2：EC Zeb 1在糖尿病视网膜病变进展中的作用将在诱导型EC特异性KO小鼠内诱导糖尿病，并将在体内测量视网膜渗透性。EC表型的表征将通过免疫组织化学和分子表征来进行目的3：Zeb 1调节的血管生成是否依赖于VEGF？将在体外探索连接VEGFR 2、缺氧和糖酵解的信号通路。这也将通过使用VEGF抗体在EC Zeb 1 KO糖尿病模型中预防VEGF刺激进行体内探索。