HEMODYNAMICS AND MOLECULAR GENETIC RISK FACTORS IN ATHEROSCLEROSIS

动脉粥样硬化的早期损害，在人类受试者和 实验动物，始终以非随机的模式发展 在动脉血管系统中，以及这些容易受损的几何结构 面积~与分支点、曲率和其他区域相关 血液流动的改变。最近的研究表明， 生物力学力对骨形成蛋白表达的影响 血管内皮细胞(EC)的病理生理相关基因。 这让我们假设，血液动力学力量，特别是 层流和扰动层流产生的壁面剪应力 心流，既可以起到积极的刺激作用，也可以起到消极的刺激作用 动脉粥样硬化的发生，通过影响EC基因的表达。为了测试这一点 假设，我们提出了一系列相互关联的体外和体内 实验，利用细胞生物学、分子生物学的组合 生物学和实验病理学技术，三岁以下 具体目标：在第一个具体目标中，以RT-PCR为基础 将使用差异显示来定义分子遗传 稳定层流暴露在培养的EC中诱发的程序 “病变保护”中特有的剪应力 区域，与“病变倾向”中发现的改变的剪应力相比较 区域。这些不同的基因表达模式将是 在一个特别设计的模型系统中进行体外验证 在培养的EC单层上产生扰动的流场，以及 “动脉粥样硬化保护”和“动脉粥样硬化致病”候选基因将是 根据结构和功能数据选择。在第二个 特定的目的，已知的潜在的协同或拮抗 生化危险因素(例如，氧化脂蛋白的成分， 如溶血磷脂酰胆碱和高水平的 同型半胱氨酸)和抗动脉粥样硬化物质(如抗氧化剂) 随着剪应力的改变，刺激将在体外模型中进行检验。 在第三个具体目标中，候选人的表达方式 EC中的动脉粥样硬化保护和致病基因将 在病变保护和病变易发区域进行活体检查 动脉粥样硬化易感性(载脂蛋白E缺陷和低密度脂蛋白)的主动脉 受体缺陷)小鼠品系和人类动脉标本。一个 病理生理学相关性的最终测试将由 检测外源基因过表达对转基因植物生长的影响 病变定位/进展/消退的候选基因。这个 这些研究的结果应该会增加我们对 生物力学和生化“危险因素”的相互作用 动脉粥样硬化的形成，并可能导致临界EC的定义 基因参与其中。

The early lesions of atherosclerosis, in human subjects and experimental animals, consistently develop in a non-random pattern in the arterial vasculature, and the geometry of these ~lesion-prone areas~ correlates with branch points, curvatures and other regions of altered blood flow. Recent studies have demonstrated direct effects of biomechanical forces on the expression of pathophysiologically relevant genes by the endothelial cell (EC). This leads us to hypothesize that hemodynamic forces, in particular wall shear stresses generated by laminar and disturbed laminar flow, can function as both positive and negative stimuli in atherogenesis, via effects on EC gene expression. To test this hypothesis, we propose a series of interrelated in vitro and in vivo experiments, utilizing a combination of cell biological, molecular biological and experimental pathological techniques, under three Specific Aims: In the First Specific Aim, RT-PCR-based differential display will be used to define the molecular genetic programs elicited in cultured EC by exposure to the steady laminar shear stresses that characteristically occur in 'lesion-protected' areas, versus the altered shear stresses found in 'lesion-prone' areas. These differential patterns of gene expression then will be validated in vitro in a specially designed model system that generates a disturbed flow field over a cultured EC monolayer, and 'athero-protective' and 'athero-pathogenic' candidate genes will be selected, based on structural and functional data. In the Second Specific Aim, the potential synergism or antagonism of known biochemical risk factors (e.g., constituents of oxidized lipoproteins, such as lysophosphatidylcholine and elevated levels of homocysteine), and anti-atherogenic substances (e.g., antioxidants) with altered shear stress stimuli will be examined in vitro models. In the Third Specific Aim, the patterns of expression of candidate athero-protective and athero-pathogenic genes in EC will be examined in vivo in the lesion-protected and lesion-prone areas of the aorta of atherosclerosis-susceptible (ApoE-deficient and LDL receptor-deficient) mouse strains, and human arterial specimens. A final test of pathophysiological relevance will be attempted by examining the effects of transgenic overexpression of selected candidate genes on lesion localization/progression/regression. The results of these studies should increase our understanding of the interplay of biomechanical and biochemical 'risk factors' in atherogenesis, and may lead to the definition of the critical EC genes involved.