Hemodynamics: Heterogeneous Endothelial Gene Expression

内皮细胞的作用及其与血流动力学的关系是动脉粥样硬化易感性的一个突出的候选者 在复杂的扰动流区域。未患病动脉中敏感部位的内皮细胞 表达一组独特的促病理和保护途径增强的平衡， 易感内皮表型。在项目5中，我们提出（a）高胆固醇血症（HC）将改变 具有不同空间和时间足迹的不同动脉中易感位点的表型平衡， 和（B）在补充的体外研究中，血液动力学流动特征是以下的表观遗传调节剂： 表型平衡我们提出了公正的基因组分析和集中的机制调查， 主要扰动途径，内质网（ER）应激，作为表型的补充方法 在体内和体外的转变。在目标1中，将在空间上绘制11个动脉区域的内皮表型 在与FDA中心的持续合作中， 位于马里兰州月桂的设备和放射卫生。特别关注内皮细胞的内质网 内质网（ER）应激/未折叠蛋白反应（UPR）途径，我们最近确定为 在正常动物体内的动脉粥样硬化易感性的特征。少数内皮细胞的表型分析 细胞（EC），包括那些覆盖发展中损伤的细胞，将在激光捕获显微镜检查后进行， 线性mRNA扩增和微阵列，以及免疫染色中鉴定的靶蛋白。 成绩单研究。在目的2中，血流动力学力和脂质环境对内皮细胞的作用， 将在暴露于动脉血流波形的EC中体外测量表型转变。将研究受控流动特性对mRNA转录谱和miRNA差异表达的影响-特别关注miR 10a/10 b调控。在目标3中，重点将在体内和体外的蛋白质水平上针对内皮ER应激和UPR机制。该提案在发现和机制水平上解决了空间上定义为内皮细胞的功能如何从病理性的 作为其脂质和血液动力学环境的函数的病理表型的失衡。

The role ofthe endothelium and its relation to hemodynamics is a prominent candidate for atherosusceptibility in regions of complex disturbed flow. Endothelium at susceptible sites in undiseased arteries express a distinct set of balances of pro-pathological and protective pathway enhancement that define the susceptible endothelial phenotype. In Project 5 we propose (a) that hypercholesterolemia (HC) will change the phenotype balance of susceptible sites in different arteries with distinct spatial and temporal footprints, and (b) in complementary in vitro studies, that hemodynamic flow characteristics are epigenetic regulators of phenotype balance. We propose unbiased genomic analyses and focused mechanistic investigations of a primary perturbed pathway, endoplasmic reticulum (ER) stress, as complementary approaches to phenotype transition in vivo and in vitro. In Aim 1, endothelial phenotypes of 11 arterial regions will be mapped spatially and temporally during 0-6 months of HC in adult swine in an ongoing collaboration with the FDA Center for Devices and Radiological Health in Laurel, MD. Particular focus will be upon endothelial endoplasmic reticulum (ER) stress/unfolded protein response (UPR) pathways that we have very recently identified as characteristic of athero-susceptibility in normal animals in vivo. Phenotyping of small numbers of endothelial cells (EC) including those overlying developing lesions wili be conducted following laser capture microscopy, linear mRNA amplification and microarray, and immunostaining of targeted proteins identified in the transcript studies. In Aim 2, the contributions of hemodynamic forces and lipid environment to endothelial phenotype transition will be measured in vitro in EC exposed to arterial flow waveforms. The influence of controlled flow characteristics on mRNA transcript profiles and miRNA differential expression - with special interest in miR10a/10b regulation will be investigated. In Aim 3, focus will be directed to endothelial ERstress and UPR mechanisms at the protein level both in vivo and in vitro. The proposal addresses at the discovery and mechanistic levels how spatially-defined as a fnction of endothelium transitions from a propathological imbalance to a pathological phenotype as a function of its lipid and hemodynamic environment.