Inflammation and Infection in Atherosclerosis

描述（由申请人提供）：冠状动脉疾病（CAD）是全球发病率和死亡率的主要原因。最近的兴趣集中在慢性感染性疾病，如牙周感染作为CAD的潜在贡献者，因为传统的风险因素，包括高胆固醇血症，吸烟和高血压不能完全解释CAD的发病率。流行病学研究表明，牙周感染的个体患CAD的可能性增加30%至100%。在过去的资助周期中，我们产生了大量的数据，这些数据与牙龈卟啉单胞菌（P.g）相关动脉粥样硬化形成的小鼠模型中感染传感电路的作用和随后的炎症细胞因子表达有关。针对这一竞争性应用，我们评估了P.g侵袭在动脉粥样硬化形成中的作用。我们发现，当侵袭缺陷型P.g.菌株DPG 3取代野生型P.g，或者当包括用侵入干扰抗生素甲硝唑处理时，随后的动脉粥样硬化形成减少约50%，支持存在显著促进P. g驱动的动脉粥样硬化形成的侵入介导的胞质过程。我们也开始研究感染和动脉粥样硬化形成的机制。核苷酸结合寡聚化结构域样受体（NOD 1和NOD 2）代表了检测细胞内微生物分子存在的免疫监视系统。最近，肽聚糖（PGN; TLR 2配体）被细胞表面TLR 2独立于其胞壁酰二肽（MDP）组分（NOD 2配体）识别，并且还通过涉及MyD 88的独特TLR信号传导途径激活NF-：B。这引入了TLR 2和NOD 2信号传导相关联的可能性，并且NOD 2的一个功能是调节TLR 2。然而，NOD 2在TLR介导的细胞因子应答中的作用仍然存在争议。我们的初步数据显示，将NOD 2 siRNA引入到P. g刺激的巨噬细胞中增强了促炎反应。然而，NOD 2激活TLR 2介导的细胞因子反应被发现依赖于MDP剂量：低配体刺激似乎是协同的，而高配体水平出现抑制。 这种调节导致NF：B调节以及IL-1和TNF转录调节。我们的假设是NOD 2的激活负性调节TLR 2反应，从而减少动脉粥样硬化;缺乏这种调节导致免疫反应增强和动脉粥样硬化加重。为了检验我们的假设，我们在目的1中提出使用功能丧失研究来确定NOD 2在P.g驱动的动脉粥样硬化中的作用;在目的2中提出确定参与NOD 2调节先天性炎症反应和侵袭介导的胞质过程的分子机制;以及在目的3中，使用功能获得研究来确定胞壁酰二肽（MDP）或NOD 2阻断剂的施用是否保护小鼠免于P. g诱导的动脉粥样硬化。这些实验将清楚地定义NOD 2在P.g动脉粥样硬化形成中的作用，并为旨在减少或预防动脉粥样硬化形成的药理学干预铺平道路。 公共卫生相关性：冠状动脉疾病（CAD）是全球人类发病和死亡的主要原因。拟议实验的结果将确定免疫传感在感染相关动脉粥样硬化形成中的作用，并为旨在减少或预防动脉粥样硬化形成的药理学干预铺平道路。

DESCRIPTION (provided by applicant): Coronary artery disease (CAD) is a major cause of morbidity and mortality worldwide. Recent interest has focused on chronic infectious diseases such as periodontal infection as potential contributors to CAD since traditional risk factors, including hypercholesterolemia, smoking, and hypertension fail to fully explain the incidence of CAD. Epidemiological studies indicate that individuals with periodontal infection are 30 to 100% more likely to have CAD. During the past funding cycle, we generated substantial data related to the role of infection-sensing circuits and ensuing inflammatory cytokine expression in a mouse model of Porphyromonas gingivalis (P.g)-associated atherogenesis. Pertinent to this competing application, we evaluated the role of P.g invasion in atherogenesis. We found that when invasion-deficient P.g. strain DPG3 was substituted for wild-type P.g, or when treatment with the invasion interfering antibiotic metronidazole was included, subsequent atherogenesis was reduced by about 50%, supporting the existence of an invasion-mediated cytosolic process that contributes significantly to P.g-driven atherogenesis. We have also begun to characterize the mechanisms linking infection and atherogenesis. Nucleotide binding oligomerization domain- like receptors (NOD1 and NOD2) represent an immune surveillance system that detects the presence of microbial molecules inside the cell. Recently peptidoglycan (PGN; a TLR2 ligand) was shown to be recognized independently of its muramyl dipeptide (MDP) components (NOD2 ligands) by cell-surface TLR2, and also to activate NF-:B through a distinct TLR signaling pathway involving MyD88. This introduced the possibility that TLR2 and NOD2 signaling are linked, and that one function of NOD2 is the regulation of TLR2. However, the role of NOD2 in TLR-mediated cytokine responses remains controversial. Our preliminary data show that introducing NOD2 siRNA into P.g-stimulated macrophages heightens the pro-inflammatory response. However, NOD2 activation of TLR2-mediated cytokine response was found dependent on MDP dose: low ligand stimulation appears to be synergistic while high ligand levels appear inhibitory. This regulation leads to NF:B modulation and IL-1 and TNF transcriptional regulation. Our hypothesis is that activation of NOD2 negatively regulates TLR2 responses, which in turn reduce artherosclerosis; the absence of such regulation leads to heightened immune responses and aggravated atherosclerosis. To test our hypothesis we are proposing in Aim 1 to use loss of function studies to determine the role of NOD2 in P.g- driven atherosclerosis; in Aim 2 to determine the molecular mechanisms involved in NOD2 regulation of the innate inflammatory response and invasion-mediated cytosolic process; and in Aim 3 to determine whether administration of muramyl dipeptide (MDP) or NOD2 blockers protect mice from P. g induced atherosclerosis using gain of function studies. These experiments will clearly define the role of NOD2 in P.g atherogenesis and pave the way for pharmacological interventions aimed at reducing or preventing atherogenesis. PUBLIC HEALTH RELEVANCE: Coronary artery disease (CAD) is a major cause of morbidity and mortality in humans worldwide. The results of the proposed experiments will define the role of immune sensing in infection-associated atherogenesis and pave the way for pharmacological interventions aimed reducing or preventing atherogenesis.