Function and Regulation of Phosphodiesterase in Atherogenesis

描述（由申请人提供）：动脉粥样硬化是心肌梗死、中风和外周血管疾病的主要触发因素，在工业化国家，这些疾病仍然是主要的死亡原因。对于新型抗动脉粥样硬化药物干预存在未满足的医学需求。这一奋进的一个关键障碍是我们目前无法完全理解动脉粥样硬化形成中的致病和调节途径。动脉粥样硬化形成的重要病理特征之一是平滑肌样细胞（所谓的合成平滑肌细胞（SMC））的异常积聚。合成的平滑肌细胞具有增殖、迁移、分泌、炎性和凋亡等特性，在平滑肌细胞的发生、发展和破裂中起着重要作用 动脉粥样硬化斑块。我们最近发现环核苷酸磷酸二酯酶1C（PDE1C）在病变血管的合成SMC中特异性表达，而在正常血管中不表达，并且促动脉粥样硬化刺激物血管紧张素II（Ang II）和反应性氧化应激（ROS）可以通过核糖体S6激酶（p90 RSK）激活PDE1C。PDE1C激活对于介导合成SMC中Ang II诱导的溶酶体胆固醇积累和趋化因子表达至关重要。我们还发现PDE1C存在于溶酶体中，并且可能参与溶酶体不稳定和胆固醇积累，随后诱导ROS产生和由于溶酶体功能障碍引起的应激诱导的细胞凋亡。因此，我们将探讨的假设，PDE1C作为一种新的关键的积极调节剂的各种促动脉粥样硬化的合成平滑肌细胞在动脉粥样硬化的过程中的功能。通过详细描述PDE1C调控的SMC发病机制在动脉粥样硬化中的功能关系，我们旨在阐明动脉粥样硬化发展的新分子机制，并开发治疗这种疾病的新治疗策略。为了实现我们的目标和解决我们的假设，我们提出了以下具体目标。在目标1中，我们将采用一系列生物化学和细胞生物学方法来确定PDE1C如何调节溶酶体功能和胆固醇积累以及合成SMC中的炎症反应。在目标2中，我们将了解PDE1C是如何被p90 RSK激活的，并建立p90 RSK介导的PDE1C激活在合成SMC发病机制中的生物学联系。在目标3中，我们将确定PDE1C缺陷在多大程度上减弱动脉粥样硬化病变形成和血管病理学在一个完善的动脉粥样硬化小鼠模型中使用全球和SMC特异性PDE1C基因敲除小鼠。我们还将表征通过其结合位点肽破坏p90 RSK介导的PDE1C活化对血管致动脉粥样硬化重塑的影响。我们在这里提出的创新方法和技术将使我们能够揭示动脉粥样硬化形成中合成SMC病理学的新分子调控机制，识别新的治疗靶点，并设计新的治疗方法来抑制合成SMC病理学，因为PDE超家族代表了一类非常有吸引力的药物靶点，用于开发特定的治疗药物。

DESCRIPTION (provided by applicant): Atherosclerosis is the main trigger of myocardial infarction, stroke and peripheral vascular disease, which remains the leading cause of death in the industrialized world. There is an unmet medical need for novel anti- atherosclerotic drug interventions. A critical roadblock in this endeavor is our current inability to fully understand pathogenic and regulatory pathways in atherogenesis. One of the important pathological features of atherogenesis is abnormal accumulation of smooth muscle-like cells, so-called synthetic smooth muscle cells (SMCs). Synthetic SMCs are proliferatory, migratory, secretory, inflammatory and apoptotic, and thus play critical roles in the initiation, progression and rupture of atherosclerotic plaques. We recently discovered that cyclic nucleotide phosphodiesterase 1C (PDE1C) is specifically expressed in synthetic SMCs of diseased vessels but not in normal vasculature and that proatherogenic stimuli angiotensin II (Ang II) and reactive oxidative stress (ROS) can activate PDE1C through ribosome S6 kinase (p90RSK). PDE1C activation is essential for mediating Ang II-induced lysosomal cholesterol accumulation and chemokine expression in synthetic SMCs. We also found that PDE1C is present in lysosomes and is likely involved in lysosomal destabilization and cholesterol accumulation, which subsequently induce ROS production and stress-induced cell apoptosis due to lysosome dysfunction. We will therefore explore the hypothesis that PDE1C acts as a novel critical positive regulator of various pro-atherogenic features of synthetic SMCs in the atherosclerotic process. By characterizing the functional relationship of PDE1C-regulated SMC pathogenesis in atherosclerosis in detail, we aim to elucidate the novel molecular mechanism of atherosclerosis development, and to develop novel therapeutic strategies for treating this disease. To achieve our goals and address our hypotheses we propose the following Specific Aims. In Aim 1 we will employ an array of biochemistry and cell biology approaches to determine how PDE1C regulates lysosomal function and cholesterol accumulation as well as inflammatory response in synthetic SMCs. In Aim 2 we will understand how PDE1C is activated by p90RSK and establish the biological link of p90RSK-mediated PDE1C activation in pathogenesis of synthetic SMCs. In Aim 3 we will determine the extent to which PDE1C deficiency attenuates atherosclerosis lesion formation and vascular pathologies in a well-established mouse model of atherosclerosis using global and SMC-specific PDE1C-knockout mice. We will also characterize the effects of disrupting p90RSK-mediated PDE1C activation by their binding-site peptide on vascular atherogenic remodeling. The innovative approaches and technologies we propose here will enable us to unveil the novel molecular regulatory mechanisms underlying synthetic SMC pathology in atherogenesis, to identify novel therapeutic targets, and to design new therapies to inhibit synthetic SMC pathologies given that PDE superfamily represents a highly attractive class of drug targets for the development of specific therapeutic agents.