Impact of dyslipidemia on endothelial biomechanics

摘要： 内皮生物力学在多种内皮功能中起着关键作用，已被越来越多地认识到。我们的 研究的重点是氧化脂质对血管内皮细胞生物力学的调节，我们发现它可以诱导血管内皮细胞 显著的内皮细胞硬化。我们的长期目标是阐明导致 血脂异常引起的内皮生物力学改变及其作用的研究 内皮功能障碍的机制。在这笔赠款的当前资助期内，我们提供了 关于血脂异常引起的主动脉内皮细胞(EC)僵硬的重要机制研究 这在很大程度上依赖于CD36清道夫受体和RhoA/ROCK信号通路的激活。我们也 发现oxLDL/血脂异常与促动脉粥样硬化紊乱的血流(DF)环境有协同作用 在体内外诱导内皮细胞僵硬的作用。在目前的建议中，我们将这些研究扩展到 解决三个新目标：在目标1中，我们专注于进一步阐明oxLDL诱导的EC的机制 变得僵硬。首先(目标1A)，我们将确定CD36在EC僵化中的作用是否提供了途径 氧化低密度脂蛋白内化和氧化脂质随后掺入膜中，或者 CD36是诱导导致EC僵硬的信号级联反应所必需的。我们还将确定 已知与CD36结合的脂肪酸对EC硬度的影响。在目标(1B)的第二部分，我们将 探讨CD36介导的oxLDL摄取和RhoA激活之间的机制联系 CASCADE，这是基于我们的初步数据，我们建议通过RhoA从 抑制调节剂GDI-1。在目标2中，我们重点研究oxLDL/DF诱导的内皮细胞僵硬在 破坏内皮屏障和内皮-单核细胞黏附(目标2B)。第一(目标2A)，我们是 建议深入研究oxLDL和DF对细胞活化的协同影响 RhoA/ROCK级联，并区分依赖于RhoA的EC硬化和 细胞凋亡对EC屏障的破坏作用。在目标(2B)的第二部分中，我们调查了 通过区分内皮细胞硬化的影响来区分oxLDL诱导的单核细胞黏附中的内皮细胞硬化 Vs.oxLDL诱导的炎性NFkB级联反应的激活和表达的增加 内皮细胞黏附分子。在目标3中，这些研究被扩展到探讨 血脂异常所致的体内内皮细胞硬化及其在动脉粥样硬化形成中的作用 损伤。这一目标将使用两种内皮特异性CD36缺陷小鼠模型来实现，一种是Ti2e- CD36天生缺陷的驱动模型和VEcad驱动的诱导型模型。这两款车型都将是 在两种主要的小鼠血脂异常模型ApoE-/-和LDLR-/-的背景下进行测试。加在一起， 这些新的研究有望为我们理解内皮细胞提供重要的新见解 血脂异常状态下的生物力学特性，特别是在促动脉粥样硬化的血流动力学中 环境。

Abstract: Endothelial biomechanics is increasingly recognized to play a key role in multiple endothelial functions. Our studies focus on regulation of endothelial biomechanics by oxidized lipids, which we showed to induce significant endothelial stiffening. Our long term goal is to elucidate the mechanisms responsible for dyslipidemia-induced changes in endothelial biomechanics and to determine the contribution of these mechanisms to endothelial dysfunction. During the current funding period of this grant, we provided significant mechanistic insights into dyslipidemia-induced stiffening of aortic endothelial cells (EC) showing that it critically depends on CD36 scavenger receptor and activation of RhoA/ROCK cascade. We also discovered that oxLDL/dyslipidemia and pro-atherogenic disturbed flow (DF) environment have a synergistic effect in inducing EC stiffening in vitro and in vivo. In the current proposal, we extend these studies to address three new goals: In Aim 1, we focus on elucidating further the mechanism of oxLDL-induced EC stiffening. First (aim 1A), we will determine whether the role of CD36 in EC stiffening is to provide the route for oxLDL internalization with subsequent incorporation of oxidized lipids into the membrane or whether CD36 is required to induce a signaling cascade that leads to EC stiffening. We will also determine the impact of fatty acids known to bind to CD36 on EC stiffness. In the second part of the aim (1B), we will investigate the mechanistic link between CD36 mediated oxLDL uptake and activation of the RhoA cascade, which based on our preliminary data, we propose to be mediated by the dissociation of RhoA from the inhibitory regulator GDI-1. In Aim 2, we focus on the role of oxLDL/DF-induced EC stiffening in the disruption of the endothelial barrier and endothelial-monocyte adhesion (aim 2B). First (aim 2A), we are proposing to investigate in depth the synergistic impact of oxLDL and DF on the activation of the RhoA/ROCK cascade and to discriminate between the contributions of RhoA-dependent EC stiffening vs. apoptosis in the disruption of the EC barrier. In the second part of the aim (2B), we investigate the role of oxLDL-induced EC stiffening in monocyte adhesion by discriminating between the impacts of EC stiffening vs. oxLDL-induced activation of the inflammatory NFkB cascade and increase in the expression of endothelial adhesion molecules. In Aim 3, these studies are extended to investigate the mechanism of dyslipidemia-induced endothelial stiffening in vivo and its contribution to the formation of atherosclerotic lesions. This goal will be achieved using two models of endothelial-specific CD36-deficient mice, a Ti2e- driven model, which is CD36-deficient from birth and VEcad-driven inducible model. Both models will be tested on the backgrounds of two major models of mouse dyslipidemia, ApoE-/- and LDLR-/-. Taken together, these new studies are expected to provide significant new insights into our understanding of endothelial biomechanical properties under dyslipidemic conditions particularly in pro-atherogenic hemodynamic environment.