Molecular Determinants of Arterial Remodeling in Atherogenesis

描述(申请人提供)：人类冠状动脉动脉瘤中的激活细胞表达高水平的基质降解酶，例如胶原酶和弹性蛋白酶。我们以前的研究表明，这种胶原酶和弹性蛋白的溶解活动调节病变动脉的完整性。最近由NIH R01支持的一系列研究为基质金属蛋白酶(MMP)家族的胶原酶在动脉重构中的重要作用提供了直接的体内证据。这一新的应用建议进一步检查胶原酶的遗传和药物操作对斑块胶原代谢的影响，并深入研究弹性分解酶在动脉疾病中的作用。我们还将探索使用生物标记物检测基质降解的可能性。具体目标1将完成我们对各种间质胶原酶在调节小鼠动脉粥样硬化斑块结构中的作用的研究。我们将研究另一种胶原酶-8的缺失对斑块结构的影响。我们将进一步分析缺乏两种主要胶原酶MMP13和MMP8的复合突变小鼠动脉粥样硬化中的胶原结构。使用一种新的成像技术和一种基质金属蛋白酶-13选择性抑制剂的研究将检验药物抑制胶原酶将改变斑块结构的假设。特定目标2将验证这样的假设，即apoE和Niemann Pick病，C1型(NPC1)蛋白联合缺陷的小鼠将显示出对动脉扩张的高度易感性。由于复合突变体apoE和NPC1缺陷小鼠过表达组织蛋白酶K，这是一种有效的弹性蛋白酶，我们将通过分子成像来跟踪这些动物体内的组织蛋白酶活性。我们还将对这些被操纵的突变小鼠的动脉组织中弹性蛋白的结构进行成像和表征。具体目标3将开发和验证细胞外基质蛋白降解的生物标记物，以提供新的工具来探测体内弹性蛋白和胶原的降解。这些标记物将包括MSGC对翻译后修饰的氨基酸及其凝聚物的测量，作为弹性蛋白和胶原分解的标志。探索弹性蛋白溶解的新标记物在体内的用途将使用来自各种弹性蛋白酶突变的小鼠的生物标本，包括组织蛋白酶K和S、基质金属蛋白酶-9和12以及中性粒细胞弹性蛋白酶，以及来自遗传诱导的心肌病小鼠的生物标本。如果我们成功地开发和验证了新的生物标记物，我们将把它们应用于我们机构进行的各种大型临床试验中患者的血液样本库中，以检验干预措施(如矿物皮质激素受体阻滞剂或他汀类药物注射)影响人体体内基质代谢的假设。 公共卫生相关性：人类冠状动脉粥样斑块中的胶原酶和弹性蛋白溶解活性可能调节动脉粥样硬化斑块的基质结构，这是急性血栓并发症的关键决定因素。我们最近的研究在NIH R01的支持下，为胶原酶的重要作用提供了直接的体内证据。这一新的应用建议进一步检查胶原酶的药理和遗传操作对斑块胶原代谢的影响。我们还建议深入研究弹性酶在动脉疾病发病机制中的作用。此外，我们还将探索使用生物标记物检测基质降解的可能性。该项目有助于了解动脉粥样硬化形成过程中动脉重塑的机制。这些互补性研究将进一步将临床前的发现转化为临床心血管预防医学。

DESCRIPTION (provided by applicant): Activated cells in human coronary atheromata express high levels of matrix-degrading enzymes, e.g., collagenases and elastases. Our previous studies suggested that such collagenolytic and elastolytic activities regulate the integrity of diseased arteries. A series of recent studies supported by this NIH R01 provided the direct in vivo evidence for the important role of collagenolytic enzymes of the matrix metalloproteinase (MMP) family in arterial remodeling. This renewal application proposes to examine further the effects of genetic and pharmacologic manipulations of collagenases on plaque collagen metabolism, and to study in depth the role of elastolytic enzymes in arterial disease. We will also explore the possibility of detecting matrix degradation using biomarkers. Specific Aim 1 will complete our studies of the roles of various interstitial collagenases in the regulation of the structure of atherosclerotic plaques in mice. We will examine the effects of deficiency of another collagenase MMP-8 on plaque structure. We will further analyze collagen structure in atheromata of the compound mutant mice lacking two major collagenases MMP-13 and MMP-8. Studies using a novel imaging technology and an MMP-13 selective inhibitor will test the hypothesis that pharmacologic inhibition of this collagenase will alter the plaque structure. Specific Aim 2 will test the hypothesis that mice with combined deficiency of apoE and Niemann Pick disease, type C1 (NPC1) protein will show heightened susceptibility to arterial ectasia. As compound mutant apoE- and NPC1-deficient mice overexpress cathepsin K, a potent elastase, we will track cathepsin activity in vivo in these animals by molecular imaging. We will also image and characterize the structure of elastin in arterial tissue from these manipulated mutant mice. Specific Aim 3 will develop and validate biomarkers of extracellular matrix protein degradation to provide novel tools to probe elastin and collagen degradation in vivo. These markers will include MSGC measurement of post-translationally modified amino acids and their condensates that serve as signatures for elastin and collagen breakdown. Exploration of the in vivo utility of novel markers of elastinolysis will use biological specimens derived from mice with mutations in various elastases including cathepsins K and S, MMP-9 and -12, and neutrophil elastase, as well as from mice with genetically-induced cardiomyopathies. If we succeed in developing and validating novel biomarkers, we will apply them to banks of blood specimens on patients in various large clinical trials conducted at our institution to test the hypothesis that the interventions (e.g. mineralocorticid receptor blocking agent or statin administration) affect matrix metabolism in humans in vivo. PUBLIC HEALTH RELEVANCE: Collagenolytic and elastolytic activities in human coronary atheromata likely regulate the matrix structure of atherosclerotic plaques, a key determinant of the acute thrombotic complications. Our recent studies supported by this NIH R01 provided the direct in vivo evidence for the important role of collagenolytic enzymes. This renewal application proposes to examine further the effects of pharmacologic and genetic manipulations of collagenases on plaque collagen metabolism. We also propose to study in depth the role of elastolytic enzymes in the pathogenesis of arterial disease. In addition, we will explore the possibility of detecting matrix degradation using biomarkers. This project should help in understanding the mechanisms of arterial remodeling during atherogenesis. These complementary studies will translate further preclinical findings into clinical preventive cardiovascular medicine.