Oxysterols, Estrogen Receptor Antagonism, and Vascular Disease

说明(申请人提供)：雌激素有可能提供有效的心血管保护。然而，有一些机制可以改变雌激素的血管作用，特别是在疾病已经存在的情况下。在目前的资助期间，我们确定27-羟基胆固醇(27HC)是第一个内源性选择性雌激素受体调节剂(SERM)，它随着高胆固醇血症而升高。我们发现，27HC拮抗血管内皮细胞和血管平滑肌上的雌激素受体(ER)功能，并导致E_2诱导的小鼠再内皮化受损。最近在小鼠身上的初步研究表明，代谢27HC的CYP7B1基因缺失也可能促进动脉粥样硬化的形成，减弱E2诱导的对新生内膜的保护，并损害E2对体重的调节。后一种发现也可能预示着葡萄糖动态平衡受损。我们的总体目标是现在确定27HC如何直接和间接地参与血管疾病的发病机制，以及如何改变27HC的水平和作用以减少血管疾病。目的1是确定应用27HC如何影响血管疾病的发病机制。27HC诱导的动脉粥样硬化病变丰度和特征的变化将在apoE+/+和apoE-/-雄性小鼠中进行评估，并在去卵巢的安慰剂和E2处理的雌性小鼠中进行评估。我们还将确定27HC如何影响E2介导的防止内膜增生的保护作用，并将在培养条件下研究27HC对E2调节巨噬细胞和VSM细胞功能的影响。目的2是确定27HC如何影响体重调节和血糖动态平衡，从而继而影响心血管健康。食物摄入量、能量消耗和肥胖度将在对照组、去卵巢安慰剂组、雌激素组C57BL/6雌性小鼠和西方饮食组中进行量化。将评估葡萄糖耐量和胰岛素敏感性以及决定它们的机制。27HC在内质网代谢调节中的中枢和外周作用将通过27HC的脑室递送来区分。目的3是确定预防高胆固醇血症所致的27HC升高如何影响雌二醇对动脉粥样硬化和代谢的保护作用。通过静脉注射编码该酶的腺病毒，以及建立肝脏特异性的CYP7B1转基因小鼠，肝脏过表达CYP7B1将降低血清27HC。目的4是开发新的策略，以减少血清27HC水平和27HC在靶细胞中的作用。利用我们开发的一种新的在小鼠体内使用的核外SERM，将在体内描述细胞培养研究中提出的非核ER信号在E2上调肝细胞色素P7B1中的作用。利用噬菌体展示识别的多肽与27HC连接的内质网唯一结合，将在培养细胞中探索选择性阻断由核和非核内质网介导的27HC作用的方法。通过实现这些目标，我们将增加我们对血管健康和疾病背景下唯一已知的内源性SERM生物学的基本理解。 与公共健康相关：27-羟基胆固醇是胆固醇的代谢物，当循环中的胆固醇水平较高时，它会在血液中升高，并与胆固醇一起在病变的动脉中积聚。我们发现，27-羟基胆固醇可以在许多细胞类型和器官中改变雌激素的作用。这项拟议的研究计划将确定27-羟基胆固醇如何通过其对雌激素作用的影响，促进血管疾病的发展以及体重和血糖调节受损。

DESCRIPTION (provided by applicant): Estrogen has the potential to provide potent cardiovascular protection. However, there are mechanisms that modify the vascular actions of estrogen, particularly if disease is already present. In the current funding period we identified 27-hydroxycholesterol (27HC), which is elevated with hypercholesterolemia, as the first endogenous selective estrogen receptor modulator (SERM). We showed that 27HC antagonizes estrogen receptor (ER) function in endothelium and vascular smooth muscle (VSM), and that it causes impaired E2- induced reendothelialization in mice. Recent preliminary studies in mice null for Cyp7b1, which metabolizes 27HC, suggest that 27HC may also promote atherogenesis, attenuate E2-induced protection from neointima formation, and impair E2 regulation of body weight. The latter finding is also likely indicative of impaired glucose homeostasis. Our Overall Objective is to now determine how 27HC contributes directly and indirectly to vascular disease pathogenesis, and how 27HC levels and actions can be modified to lessen vascular disease. Aim 1 is to determine how administered 27HC impacts vascular disease pathogenesis. 27HC-induced changes in atherosclerotic lesion abundance and characteristics will be evaluated in apoE+/+ vs. apoE-/- male and ovariectomized placebo vs. E2-treated female mice. We will also determine how 27HC impacts E2-mediated protection from intimal hyperplasia, and effects of 27HC on E2 regulation of macrophage and VSM cell function will be investigated in culture. Aim 2 is to determine how 27HC impacts body weight regulation and glucose homeostasis, which secondarily influence cardiovascular health. Food intake, energy expenditure and adiposity will be quantified in control vs. 27HC-treated ovariectomized placebo vs. E2-treated female C57BL/6 mice placed on control vs. Western diet. Glucose tolerance and insulin sensitivity and the mechanisms that determine them will be evaluated. CNS vs. peripheral actions of 27HC on ER regulation of metabolism will be distinguished by intraventricular delivery of 27HC. Aim 3 is to determine how prevention of the elevation in 27HC that occurs with hypercholesterolemia impacts the protective actions of E2 on atherosclerosis and metabolism. Serum 27HC will be lowered by hepatic overexpression of Cyp7b1 by IV injection of adenovirus encoding the enzyme and also by creation of a liver-specific Cyp7b1 transgenic mouse. Aim 4 is to develop new strategies to attenuate serum 27HC levels and 27HC actions in target cells. Using a novel extranuclear SERM we developed for in vivo use in mice, the role of non-nuclear ER signaling in hepatic Cyp7b1 upregulation by E2 suggested in cell culture studies will be delineated in vivo. Using peptides identified by phage display to uniquely bind to 27HC-liganded ER, approaches for the selective blockade of 27HC actions mediated by nuclear as well as non-nuclear ER will be pursued in cultured cells. By accomplishing these aims, we will increase our basic understanding of the biology of the only known endogenous SERM in the context of vascular health and disease. PUBLIC HEALTH RELEVANCE: 27-hydroxycholesterol is a metabolite of cholesterol that is elevated in the blood when circulating cholesterol levels are high, and it accumulates with cholesterol in diseased arteries. We have discovered that 27-hydroxycholesterol modifies the actions of estrogen in numerous cell types and organs. The proposed research program will determine how 27-hydroxycholesterol, through its impact on estrogen action, contributes to the development of blood vessel disease and to impaired body weight and glucose regulation.