PPARgamma, Endothelial Inflammation, and Atherosclerosis

PPARgamma、内皮炎症和动脉粥样硬化

• 批准号: 7142765
• 负责人: Jun-Ichi Abe
• 金额: $ 46.21万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7142765
• 关键词: JUN kinase; RNA interference; atherosclerosis; confocal scanning microscopy; disease /disorder etiology; disease /disorder model; enzyme activity; gene expression; genetic transcription; genetically modified animals; hemodynamics; human tissue; inflammation; laboratory mouse; laboratory rabbit; mitogen activated protein kinase; molecular pathology; nuclear factor kappa beta; organ culture; pathologic process; peroxisome proliferator activated receptor; protein protein interaction; shear stress; tissue /cell culture; tumor necrosis factor alpha; vascular cell adhesion molecule; vascular endothelium

There is increasing support for the idea that inflammation plays a major role in the development of atherosclerosis. Peroxisome proliferator-activated receptors (PPARgamma) are ligand-activated transcription factors that form a subfamily of the nuclear gene family. PPARgamma activators inhibit adhesion molecules expression in activated endothelial cells and significantly reduce monocyte/macrophage homing to atherosclerotic plaques. We found that steady laminar flow activates ERK5, and increases PPARgamma transcriptional activation. In addition, to utilize the newly identified Hinge-helix 1 region of PPARgamma fragment (Hin-H1 fragment), which specifically disrupts ERK5/PPARgamma interaction and inhibits activated ERK5-mediated PPARgamma activation, will provide a great tool to explore the specific role of ERK5 and PPARgamma interaction in laminar flow-mediated anti-inflammatory effect. Based on our exciting preliminary data that ERK5 activation increases PPARgamma activity and that this may inhibit the expression of adhesion molecules, we hypothesize that laminar flow acts as an anti-inflammatory modulator by increasing PPARgamma transcriptional activity, and decreasing VCAM-1 expression in endothelial cell. To prove the contribution of PPARgamma transcriptional activity (which is regulated by ERK5) to the atheroprotective effect of flow in endothelial cells we propose three aims. Aim 1 : Using cultured endothelial cells, define the role of ERK5/PPARgamma on TNF-alpha induced NF-kappaB, VCAM-1 promoter activation and subsequent VCAM-1 expression. Aim 2: Using an ex vivo organ culture system of intact aorta, determine the role of PPARgamma and ERK5 in inflammation-related expression of adhesion molecules Aim 3: Using an atherosclerosis model, determine endothelial ERK5, JNK, NF-kappaB activation and VCAM-1 expression in vivo by using en face confocal microscopy, and detect the role of endothelial ERK5 in atherosclerosis formation using gain and loss of function mice. In aim 1 we will show the importance of ERK5/PPARgamma interaction to regulate laminar flow-mediated anti-inflammatory effect using Hin-H1 fragment and ERK5 siRNA in adenovirus vector. In aim 2 we will utilize the organ culture system, and determine the role of ERK5-PPARgamma in flow in intact vessels. In aim 3 we will use endothelial specific constitutively active(CA)-MEK5 or ERK5' knock out in LDLR-/- knock out(KO) mice, which has been characterized by atherosclerosis formation. We anticipate that CA-MEK5 will inhibit and ERK5 KO will promote atherosclerosis formation. These experiments will provide a new molecular mechanism for the anti-inflammatory effect of PPARgamma and flow associated with ERK5 activity.

人们越来越多地支持炎症在动脉粥样硬化的发展中发挥重要作用的观点。 过氧化物酶体增殖物激活受体 (PPARgamma) 是配体激活的转录因子，形成核基因家族的一个亚家族。 PPARγ激活剂抑制活化内皮细胞中粘附分子的表达，并显着减少单核细胞/巨噬细胞归巢至动脉粥样硬化斑块。我们发现稳定的层流会激活 ERK5，并增加 PPARgamma 转录激活。此外，利用新鉴定的 PPARgamma 片段（Hin-H1 片段）的铰链螺旋 1 区域，特异性破坏 ERK5/PPARgamma 相互作用并抑制激活的 ERK5 介导的 PPARgamma 激活，将为探索 PPARgamma 的具体作用提供一个很好的工具。 ERK5 和 PPARgamma 在层流介导的抗炎作用中相互作用。基于我们令人兴奋的初步数据，即 ERK5 激活会增加 PPARgamma 活性，并且这可能会抑制粘附分子的表达，我们假设层流通过增加 PPARgamma 转录活性并减少内皮细胞中 VCAM-1 的表达来充当抗炎调节剂。为了证明 PPARgamma 转录活性（受 ERK5 调节）对内皮细胞血流的动脉粥样硬化保护作用的贡献，我们提出了三个目标。目标 1：使用培养的内皮细胞，确定 ERK5/PPARgamma 对 TNF-α 诱导的 NF-κB、VCAM-1 启动子激活和随后的 VCAM-1 表达的作用。目标 2：使用完整主动脉的离体器官培养系统，确定 PPARgamma 和 ERK5 在炎症相关粘附分子表达中的作用 目标 3：使用动脉粥样硬化模型，确定内皮 ERK5、JNK、NF-kappaB 激活和 VCAM- 1.使用enface共聚焦显微镜体内表达，并检测作用 使用功能获得和丧失的小鼠观察内皮 ERK5 在动脉粥样硬化形成中的作用。在目标 1 中，我们将展示 ERK5/PPARgamma 相互作用对于使用 Hin-H1 调节层流介导的抗炎作用的重要性 腺病毒载体中的片段和ERK5 siRNA。在目标 2 中，我们将利用器官培养系统，并确定其作用 完整血管中流动的 ERK5-PPARgamma。在目标 3 中，我们将在 LDLR-/- 敲除 (KO) 小鼠中使用内皮特异性组成型活性 (CA)-MEK5 或 ERK5' 敲除，其特征是动脉粥样硬化形成。我们 预计CA-MEK5会抑制动脉粥样硬化的形成，ERK5 KO会促进动脉粥样硬化的形成。这些实验将 为 PPARgamma 的抗炎作用和与 ERK5 活性相关的血流提供了新的分子机制。