Function and Regulation of Phosphodiesterase in Atherogenesis

描述(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诱因，在工业化世界中仍是主要的死亡原因。对新型抗动脉粥样硬化药物干预的医学需求尚未得到满足。这一努力中的一个关键障碍是我们目前无法完全了解动脉粥样硬化形成中的致病和调控途径。动脉粥样硬化形成的重要病理特征之一是异常聚集的平滑肌样细胞，即所谓的合成平滑肌细胞(SMC)。人工合成的SMC具有增殖、迁移、分泌、炎症和凋亡等功能，因此在SMC的发生、发展和破裂过程中起关键作用。 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)在病变血管的合成的SMC中特异表达，而在正常血管中不表达，而且促动脉粥样硬化刺激血管紧张素II(Ang II)和反应性氧化应激(ROS)可以通过核糖体S6激酶(P90RSK)激活PDE1C。PDE1C的激活对于介导Ang II诱导的合成SMC中溶酶体胆固醇积聚和趋化因子的表达是必不可少的。我们还发现，PDE1C存在于溶酶体中，可能参与了溶酶体的失稳和胆固醇的积聚，从而诱导ROS的产生和因溶酶体功能障碍而导致的应激诱导细胞凋亡。因此，我们将探索一种假设，即PDE1C在动脉粥样硬化过程中作为一种新的关键正向调节因子，调节合成的SMC的各种促动脉粥样硬化特性。通过详细研究PDE1C调控的SMC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生发展的新的分子机制，并开发新的治疗策略。为了实现我们的目标并解决我们的假设，我们提出了以下具体目标。在目标1中，我们将使用一系列生物化学和细胞生物学方法来确定PDE1C如何调节合成的SMC中的溶酶体功能和胆固醇积累以及炎症反应。在目标2中，我们将了解p90RSK是如何激活PDE1C的，并建立p90RSK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，我们将使用全局和SMC特异性的PDE1C基因敲除小鼠，确定PDE1C缺乏在多大程度上减轻动脉粥样硬化小鼠动脉粥样硬化病变的形成和血管病理。我们还将研究它们的结合部位肽对p90RSK介导的PDE1C激活的干扰在血管动脉粥样硬化重建中的作用。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中合成SMC病理的新的分子调控机制，识别新的治疗靶点，并设计新的治疗方法来抑制合成的SMC病理，因为PDE超家族代表着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.