Role of Inflammation in the Pathobiology of Abdominal Aortic Aneurysm

炎症在腹主动脉瘤病理学中的作用

• 批准号: 8598786
• 负责人: Kathleen Raman
• 金额: --
• 依托单位: ST. LOUIS VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-10-01 至 2017-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8598786
• 关键词: Abdominal Aortic Aneurysm; Acute; Advanced Glycosylation End Products; Affect; Aorta; Apoptosis; Apoptotic; Atherosclerosis; Biology; Blood Vessels; Bone Marrow; Caliber; Cell Culture Techniques; Cells; Chronic; Cytoplasmic Tail; Data; Development; Dominant Negative Receptor; Elastases; Elastic Fiber; Elastin; Environmental Risk Factor; Family; Functional disorder; Gelatinase B; Genetic; HMGB1 Protein; Human; Immunohistochemistry; In Situ Nick-End Labeling; In Vitro; Incidence; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Knockout Mice; Lead; Leukocyte L1 Antigen Complex; Leukocytes; Life; Ligand Binding; Ligands; Link; Macrophage Activation; Matrix Metalloproteinases; Measures; Medial; Mediating; Mediator of activation protein; Molecular; Mus; Pathogenesis; Pattern; Peptide Hydrolases; Perfusion; Phenotype; Population; Prevention strategy; Primary Cell Cultures; Protein Family; RNA; Receptor Activation; Receptor Inhibition; Research Project Grants; Role; Signal Pathway; Signal Transduction; Smooth Muscle Myocytes; Staining method; Stains; Stimulus; Technology; Testing; Time; Tissue Inhibitor of Metalloproteinases; Tissues; Transgenic Mice; Vascular Diseases; Veterans; Wild Type Mouse; cell behavior; clinically relevant; cytokine; in vitro Model; in vivo; indexing; innovation; macrophage; member; mouse model; novel diagnostics; novel therapeutics; promoter; public health relevance; receptor; reconstitution; tool

DESCRIPTION (provided by applicant): Abdominal aortic aneurysms (AAAs) are a common and potentially life-threatening vascular condition prevalent among US veterans. Aortic wall inflammation results in two fundamental structural changes which underlie AAA; elastin degradation and loss of smooth muscle cells (SMC) within the aortic media. Aneurysmal degeneration is related to increased elaboration and activity of elastin-specific proteinases, particularly matrix metalloproteinases (MMPs). While macrophages and SMC are key cellular mediators in the pathogenesis of AAAs; the upstream stimulus which triggers the inflammatory cascade responsible for matrix changes remains unknown. Activation of the receptor for advanced glycation end products (RAGE) leads to both an acute inflammatory response and perpetuation of a chronic, sustained inflammatory state leading to tissue damage. Additionally, along with endogenous tissue inhibitors of metalloproteinases (TIMPs), RAGE upregulates the expression and activity of MMPs including MMP-9, a protease expressed by macrophages that is critical to AAA formation. SMC apoptosis has been linked to the interaction of RAGE with diaphanous 1 (mDia-1). RAGE blockade may be effected using soluble RAGE (sRAGE), a ligand-binding decoy of RAGE, as well as RAGE null mice. Transgenic mice in which SMCs express dominant negative (DN) RAGE driven by the SM22¿ promoter allow assessment of cell-specific contributions of RAGE activation. We hypothesize that RAGE alters macrophage polarization and SMC apoptosis, leading to matrix degeneration fundamental to AAA development. Thus, the specific aims of this proposal are: Aim #1: Determine if RAGE-mediated signaling alters macrophage activation and polarization underlying the proinflammatory pathophysiology of AAA. We hypothesize that RAGE mediates the polarization of macrophages from an M2 (reparative) to an M2 (inflammatory) phenotype that transmits a damage signal and contributes to AAA. We will delineate the M1 and M2 subtypes in the aortic tissues of RAGE null, sRAGE-treated, and untreated mice following elastase perfusion. We will confirm our findings with in vitro studies using primary bone marrow derived macrophages stimulated with RAGE ligands to determine the effects of RAGE on MMP and TIMP activity and macrophage polarization. We will reconstitute RAGE null mice with RAGE-sufficient bone marrow to determine whether the presence of RAGE on bone marrow derived leukocytes is sufficient to polarize macrophages into an M1 subtype in the aorta of elastase-perfused mice. Aim #2: Characterize if RAGE-mediated SMC apoptosis within the aortic wall leads to the pathogenesis of AAA. We hypothesize that RAGE, through its interaction with mDia-1, mediates SMC apoptosis and proliferation within the aortic wall. We will utilize SMC DN RAGE mice to assess SMC-specific contributions of RAGE activation in elastase-induced AAA. Using Ki67 and TUNEL staining, we will quantify proliferative and apoptotic indices in SMCs of elastase-perfused aorta of wild-type, RAGE null, and sRAGE- treated wild-type mice. In vitro, we will stimulate primary aortic SMC with RAGE ligands and measure effects of RAGE on SMC proliferative and apoptotic indices. We will determine if mDia-1 is necessary for RAGE's effects using inhibitory RNA technology. This proposal is innovative in that we will use in vivo and in vitro models to understand how RAGE-mediated alterations in macrophage polarization and SMC apoptosis result in AAA development. The studies described in this proposal may eventually lead to novel diagnostic and therapeutic strategies for prevention and treatment. The clinical relevance of this proposal is underscored by the increasing incidence of AAAs among the VA population and the limited treatment options that are currently available.

描述（由申请人提供）： 腹主动脉瘤（AAA）是美国退伍军人中常见的一种可能危及生命的血管疾病。主动脉壁炎症导致AAA基础的两种基本结构变化;主动脉中膜内弹性蛋白降解和平滑肌细胞（SMC）损失。动脉瘤变性与弹性蛋白特异性蛋白酶，特别是基质金属蛋白酶（MMPs）的加工和活性增加有关。虽然巨噬细胞和SMC是AAAs发病机制中的关键细胞介质;但触发负责基质变化的炎症级联反应的上游刺激仍然未知。晚期糖基化终末产物受体（AGEs）的激活导致急性炎症反应和慢性持续炎症状态的持续，从而导致组织损伤。此外，沿着内源性金属蛋白酶组织抑制剂（TIMP），MMP-9（一种由巨噬细胞表达的蛋白酶，对AAA形成至关重要）等MMP的表达和活性上调。平滑肌细胞凋亡与mDia-1相互作用有关。可使用可溶性β-内酰胺酶（sNAR）（一种β-内酰胺酶的配体结合诱饵）以及β-内酰胺酶缺失小鼠实现β-内酰胺酶阻断。在转基因小鼠中，SMC表达由SM 22启动子驱动的显性阴性（DN）β 2受体，可以评估β 2受体活化的细胞特异性贡献。我们推测，腹主动脉瘤改变巨噬细胞极化和平滑肌细胞凋亡，导致基质变性的AAA发展的基础。因此，本提案的具体目标是：目标#1：确定RAGE介导的信号传导是否改变AAA的促炎病理生理学基础的巨噬细胞活化和极化。我们假设，AAA介导巨噬细胞从M2（修复性）到M2（炎症性）表型的极化，M2表型传递损伤信号并促成AAA。我们将描述弹性蛋白酶灌注后，在EST-null、sRAGE治疗和未治疗小鼠的主动脉组织中的M1和M2亚型。我们将通过体外研究证实我们的发现，该研究使用经MMP配体刺激的原代骨髓来源的巨噬细胞来确定MMP和TIMP活性以及巨噬细胞极化的影响。我们将用RAGE充足的骨髓重建RAGE缺失小鼠，以确定骨髓源性白细胞上RAGE的存在是否足以使弹性蛋白酶灌注小鼠主动脉中的巨噬细胞极化为M1亚型。目的#2：表征主动脉壁内RAGE介导的SMC凋亡是否导致AAA的发病机制。我们假设，通过与mDia-1的相互作用，介导主动脉壁内SMC的凋亡和增殖。我们将利用SMC DN AAA小鼠来评估SMC特异性在弹性蛋白酶诱导的AAA中的活化作用。使用Ki 67和TUNEL染色，我们将量化弹性蛋白酶灌注的野生型、β-淀粉样蛋白缺失和β-淀粉样蛋白治疗的野生型小鼠主动脉SMC的增殖和凋亡指数。在体外，我们将刺激主动脉平滑肌细胞的配体和测量的影响，平滑肌细胞增殖和凋亡指数。我们将使用抑制性RNA技术来确定mDia-1是否是Escherichia's效应所必需的。这个建议是创新的，我们将使用在体内和体外模型，以了解如何RAGE介导的改变巨噬细胞极化和SMC凋亡导致AAA的发展。本提案中描述的研究可能最终导致新的预防和治疗诊断和治疗策略。VA人群中AAA的发生率增加以及目前可用的治疗选择有限，强调了该提议的临床相关性。