Molecular mechanisms in Abdominal Aortic Aneuysm

描述（由申请人提供）：腹主动脉瘤是一种常见的与年龄相关的潜在致命疾病，是慢性炎症、细胞外基质（ECM）降解和与内侧平滑肌细胞（SMCs）消耗相关的ECM生物合成受损之间复杂相互作用的结果。然而，在这错综复杂的神经网络中，中间的SMCs的确切作用尚未被阐明。我们的初步研究结果表明，信号分子蛋白激酶c - δ (PKC4)在人和动物动脉瘤组织的SMCs中上调。使用动脉瘤的CaCl2模型，我们证明PKC4基因缺陷小鼠免受动脉瘤的发展。初步的组织学分析表明，PKC4基因缺失会损害巨噬细胞的浸润，阻止内侧SMCs的消耗和ECM的降解。这些初步发现是新颖的，首次暗示PKC4参与动脉瘤的发病机制。我们推测PKC4可能是导致动脉瘤相关SMCs凋亡和促炎行为的一个重要因素。在本研究中，我们提出了三个具体目标来验证PKC4是导致内侧SMC耗竭和血管炎症的重要分子因子的假设。PKCd的促炎功能是通过单核细胞趋化蛋白-1 （MCP-1）介导的。在Specific Aim I中，我们将通过使用CaCl2模型和动脉瘤发展过程中的一系列组织学分析来确认PKC4敲除小鼠的动脉瘤抗性表型。此外，我们将使用通过不同机制诱导动脉瘤的弹性蛋白酶灌注模型来测试PKC4的作用。在Specific Aim II中，我们将描述PKC4基因缺失损害动脉瘤形成的机制。首先，我们将测试涉及Rho GTPase Cdc42和MAP激酶ERK的MCP-1基因表达的新范式。其次，我们将测试在PKC4基因敲除小鼠的主动脉壁上恢复MCP-1是否能挽救受损的动脉瘤。第三，我们将评估PKC4在巨噬细胞中的作用，以及炎症细胞对PKC4缺失小鼠抗动脉瘤表型的潜在贡献。在Specific Aim III中，我们将描述PKC4的两种肽抑制剂，其中一种是在我们的初步研究中开发的，以便开发一种分子试剂来在体内拮抗PKC4。所选抑制剂减弱动脉瘤发展的潜在功效将通过CaCl2模型来确定。我们认为，动脉PKC4上调例证了SMCs内的细胞内信号网络，该网络驱动促炎信号和细胞凋亡。通过提出的体内和体外研究，我们将更好地了解腹主动脉瘤的内侧SMCs发病机制，这将使我们能够为开发这种破坏性血管疾病的新疗法做出贡献。

DESCRIPTION (provided by applicant): Abdominal aortic aneurysm is a common age related and potentially lethal disease that develops as the result of a complex interplay among chronic inflammation, extracelluar matrix (ECM) degradation, and impaired ECM biosynthesis associated with depletion of medial smooth muscle cells (SMCs). However, the precise role of medial SMCs in this intricate network has yet to be elucidated. Results from our preliminary studies suggest that the signaling molecule protein kinase C-delta (PKC4) is upregulated in SMCs of human and animal aneurismal tissues. Using the CaCl2 model of aneurysm, we demonstrated that PKC4 gene deficient mice are protected from the development of aneurysms. Preliminary histological analyses indicate that PKC4 gene deficiency impairs infiltration of macrophages and prevents depletion of medial SMCs and degradation of ECM. These preliminary findings are novel and for the first time implicate PKC4 in the pathogenesis of aneurysms. We speculate that PKC4 might be an integral factor responsible for the apoptotic and pro-inflammatory behavior of SMCs associated with aneurysm. In this study, we propose three specific aims to test the hypothesis that PKC4 is an important molecular factor contributing to medial SMC depletion and vascular inflammation. The pro-inflammatory function of PKCd is mediated through monocyte chemoattactant protein-1 (MCP-1). In Specific Aim I, we will confirm the aneurysm-resistant phenotype of PKC4 knockout mice through the use of the CaCl2 model paired with a serial of histological analyses over the course of aneurysm development. In addition, we will test the role of PKC4 using the elastase perfusion model that induces aneurysm through a different mechanism. In Specific Aim II, we will delineate mechanisms through which PKC4 gene deficiency impairs aneurysm formation. First, we will test a new paradigm underlying MCP-1 gene expression involving Rho GTPase Cdc42 and MAP kinase ERK. Second, we will test whether restoring MCP-1 in the aortic wall rescues the impaired aneurysm in PKC4 knockout mice. Third, we will assess the role of PKC4 in macrophages and the potential contribution of inflammatory cells to the aneurysm resistant phenotype of PKC4 null mice. In Specific Aim III, we will characterize two peptide inhibitors of PKC4, one of which was developed during our preliminary studies, in order to develop a molecular reagent to antagonize PKC4 in vivo. The potential efficacy of the selected inhibitor to attenuate aneurysm development will be determined using the CaCl2 model. We believe that arterial PKC4 upregulation exemplifies an intracellular signaling network within SMCs that drives pro-inflammatory signaling and apoptosis. Through the proposed in vivo and in vitro studies, we will gain a better understanding of medial SMCs in the pathogenesis of abdominal aortic aneurysm, which will allow us to make contributions toward the development of novel therapies for this devastating vascular disease. PUBLIC HEALTH RELEVANCE: Abdominal aortic aneurysm (AAA) is a common degenerative disease associated with high mortality. Each year thousands of aneurysms, the majority of which are small, are identified through screening programs. Without treatment, these small aneurysms will enlarge and eventually rupture. Surgical repair is an effective approach to prevent deaths or complications resulting from a ruptured aneurysm, however, many patients with ruptured AAAs die before reaching a hospital. Therefore, there is a great need for alternative therapeutic strategies. By providing new information on regulatory mechanisms underlying smooth muscle cell apoptosis and inflammatory signaling, the proposed studies will further our understanding of the molecular mechanisms underlying the pathophysiology of aneurysms and facilitate development of novel therapeutic strategies aimed at slowing or reversing the growth of small aneurysms.