Impact of dyslipidemia on endothelial biomechanics

血脂异常对内皮生物力学的影响

• 批准号: 8321203
• 负责人: MICHAEL CHO
• 金额: $ 54.89万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-04 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8321203
• 关键词: Accounting; Actins; Address; Antiatherogenic; Aorta; Apolipoprotein E; Atherosclerosis; Atomic Force Microscopy; Belief; Biological Assay; Biomechanics; Blood Vessels; Cardiovascular Diseases; Caveolins; Cell membrane; Cell physiology; Cessation of life; Cholesterol; Complex; Deposition; Development; Disease; Dyslipidemias; Endothelial Cells; Endothelium; Environment; Event; Exposure to; Functional disorder; Funding; Goals; Grant; Guanosine Triphosphate Phosphohydrolases; Harvest; Image; Impairment; In Vitro; Individual; Inflammatory; Link; Lipids; Low-Density Lipoproteins; Mass Spectrum Analysis; Measures; Mechanics; Mediating; Membrane; Microscopy; Modeling; Modification; Morbidity - disease rate; Mus; Myosin ATPase; Myosin Light Chains; Pathway interactions; Permeability; Phosphorylation; Plasma; Play; Property; Regulation; Resistance; Rho-associated kinase; Risk Factors; Role; Scheme; Signal Pathway; Signal Transduction; Staging; Techniques; Testing; Tissues; Woman; athero susceptible; base; caveolin 1; cell injury; disorder prevention; gain of function; hemodynamics; in vivo; insight; loss of function; low density lipoprotein inhibitor; men; mutant; myosin phosphatase; novel; oxidized lipid; oxidized low density lipoprotein; response; rho; rho GTP-Binding Proteins; shear stress; two-photon; uptake

DESCRIPTION (provided by applicant): Biomechanical properties of endothelial cells (ECs) are crucially important in regulation of multiple EC functions, such as mechanotransduction and the integrity of the EC barrier. We have recently discovered that oxidized modifications of LDL (oxLDL) induce significant EC stiffening indicating that dyslipidemia plays a major role in the regulation of EC mechanics. Our long term goal is to elucidate the mechanisms responsible for dyslipidemia-induced changes in EC biomechanics and to determine the contribution of these mechanisms to endothelial dysfunction. During the first funding period of this grant, we have provided the first mechanistic insights into oxLDL-induced EC stiffening and demonstrated that it may facilitate the sensitivity of endothelial cells to flow. In the current proposal, we extend thee studies to address three new goals: In Aim 1, we will identify specific oxidized lipids that induce EC stiffening and address the hypothesis that EC stiffening is mediated by the insertion of oxidized lipids into the plasma membranes of endothelial cells and disruption of lipid packing of membrane domains. To achieve this goal, we will perform Mass Spectrometry analysis of oxidized lipids found in both oxLDL complex and in the vascular walls of aortas isolated from dyslipidemic ApoE-/- mice. EC stiffness will be measured using a combination of two biophysical techniques, Microaspiration and Atomic Force Microscopy and lipid packing will be assayed by two-photon microscopy. In Aim 2, we will elucidate the downstream signaling pathways that are responsible for oxLDL-induced EC stiffening focusing on the roles of caveolin and Rho-GTPases. Specifically, we will address a hypothesis that oxLDL/oxidized lipids-induced disruption of cholesterol-rich membrane domains activate a signaling pathway that includes phosphorylation of caveolin-1, activation of Rho-GTPase and its major downstream target, ROCK, with subsequent changes in actin/myosin organization. This hypothesis will be addressed using an array of gain-of-function and loss-of-function mutants of caveolin, Rho and Rac- GTPases and ROCK. In Aim 3, we will determine the impact dyslipidemia-induced EC stiffening on endothelial permeability under different hemodynamic environments in vitro and in vivo. More specifically, first we will test the hypothesis that oxLDL-induced EC stiffening impairs EC barrier and augments an increase in EC permeability under disturbed pro-atherogenic flow environment in vitro. Finally, we will determine whether an increase in EC stiffness correlates with an increase in endothelial permeability in vivo in ApoE-/- mice and determine whether disruption of the endothelial barrier in ApoE-/- mice can be rescued by caveolin-1 deficiency and/or ROCK inhibition. PUBLIC HEALTH RELEVANCE: Atherosclerosis, a disease of narrowing and blocking of major blood vessels, is a major cause for the development of the cardiovascular disease (CVD), which is responsible for 40% of all deaths and results in serious morbidity in both men and women. The crucial factor for atherosclerosis development is dyslipidemia, an increase in pro-atherogenic low-density lipoproteins (LDL), particularly when LDL undergoes oxidative modifications leading to formation of highly pro-inflammatory oxidized LDL (oxLDL). It is also known that the early key step in the development of atherosclerosis is dysfunction of endothelial cells, the inner lining of the blood vessels. Our studies suggest a novel paradigm for oxLDL-induced endothelial damage, an increase in endothelial stiffness, and we suggest that the stiffening of the endothelium plays a major role in the initiation of the disease. Furthermore, our studies suggest that treatments that rescue endothelial cells from oxLDL-induced stiffening and restore normal endothelial stiffness may have strong beneficial effects for the disease prevention.

描述（由申请人提供）：内皮细胞（ECS）的生物力学特性在调节多个EC功能（例如机械传导和EC屏障的完整性）方面至关重要。我们最近发现，LDL（OXLDL）的氧化修饰引起了显着的EC僵硬，表明血脂异常在EC机械的调节中起主要作用。我们的长期目标是阐明负责血脂异常引起的EC生物力学变化的机制，并确定这些机制对内皮功能障碍的贡献。在该赠款的第一个资金期间，我们为OXLDL诱导的EC加强提供了第一个机械洞察力，并证明它可能促进内皮细胞对流动的敏感性。在当前的建议中，我们扩展了您的研究以解决三个新目标：在AIM 1中，我们将确定诱导的特定氧化脂质 EC僵硬并解决了以下假设：EC僵硬是由将氧化脂质插入内皮细胞的质膜和膜结构域脂质堆积的破坏所介导的。为了实现这一目标，我们将对在OxLDL复合物中发现的氧化脂质和从血脂性APOE-/ - 小鼠分离的主动脉壁的血管壁中发现的氧化脂质分析。将通过两种生物物理技术，微型抽血和原子力显微镜和脂质堆积的组合测量EC刚度，通过两光子显微镜测定。在AIM 2中，我们将阐明负责OXLDL诱导的EC僵硬的下游信号通路，重点是小窝蛋白和Rho-GTPase的作用。具体而言，我们将解决一个假设，即OXLDL/氧化脂质诱导的富含胆固醇的膜结构域的破坏激活了一种信号传导途径，其中包括小窝蛋白-1的磷酸化，Rho-GTPase的激活，其主要下游靶标及其主要的下游靶标，随后在ACTIN/肌球蛋白组织中变化。该假设将使用一系列可爱蛋白，Rho和Rac-GTPases和Rock的功能障碍和功能丧失突变体来解决。在AIM 3中，我们将确定血脂异常诱导的EC僵硬对体外和体内不同血液动力学环境下内皮渗透性的影响。更具体地说，首先，我们将测试OXLDL诱导的EC僵硬损害的假设 在体外侵入的促动源性流动环境下，EC屏障并增加了EC渗透性的提高。最后，我们将确定EC刚度的增加是否与ApoE-/ - 小鼠体内内皮渗透性的增加相关，并确定是否可以通过Caveolin-1侵犯和/或岩石抑制作用来挽救APOE-/ - 小鼠中内皮屏障的破坏。 公共卫生相关性：动脉粥样硬化是一种狭窄和阻塞主要血管的疾病，是发展心血管疾病（CVD）的主要原因，该病因造成了所有死亡的40％，并导致男性和女性严重发病。动脉粥样硬化发育的关键因素是血脂异常，促动脉粥样硬化的低密度脂蛋白（LDL）的增加，尤其是当LDL经过氧化修饰导致高度促炎性氧化的LDL（OXLDL）形成氧化修饰时。还知道，动脉粥样硬化发展的早期关键步骤是内皮细胞的功能障碍，即血管内衬。我们的研究表明，OXLDL诱导的内皮损伤的新型范式，内皮刚度的增加，我们建议内皮的僵硬在疾病的启动中起主要作用。此外，我们的 研究表明，从OXLDL诱导的僵硬并恢复正常内皮僵硬的治疗方法可能对预防疾病具有强大的有益作用。