Regulation of PAF Acetylhydrolase Expression by Oxidized Phospholipids

描述（由申请人提供）：血小板活化因子乙酰水解酶（PAFAH）也称为脂蛋白相关磷脂酶A2，是一种钙非依赖性酰基水解酶，可分解代谢氧化LDL（oxLDL）颗粒中的氧化磷脂（oxPL）以及脂质介质血小板活化因子（PAF）。最近的许多研究表明PAFAH活性与血管疾病和动脉粥样硬化的严重程度相关。尽管这种酶在动脉粥样硬化和其他血管疾病的发病机制中的重要性毋庸置疑，但对其表达和活性的调节机制知之甚少。我们的初步研究表明，PAFAH的表达诱导氧化磷脂（oxPL），衍生物1-棕榈酰-2-花生四烯酸-sn-甘油-3-磷酸胆碱，oxLDL的一个组成部分。这为我们提供了一个独特的模型，我们将使用它来描绘动脉粥样硬化病变中PAFAH表达的分子机制。我们假设长链、类胡萝卜素oxPL通过与发展中动脉粥样硬化病变中树突状细胞上的CD 14/TLR 4复合物和EP 2前列腺素受体结合来诱导PAFAH。这些相互作用被认为是诱导白细胞介素6（IL 6）和磷酸化和激活的关键转录因子，事件必不可少的最佳转录PAFAH基因。为了验证这一假设，我们提出了以下具体目标：1）研究EP 2、IL 6和CD 14/TLR 4在oxPL诱导PAFAH中的作用; 2）研究oxPL诱导PAFAH的分子机制。在目的1中，我们将检验oxPL与EP 2和/或TLR 4-CD 14受体复合物的结合诱导IL 6以及该细胞因子对于最佳PAFAH诱导是必需的这一假设。在目的2中，我们将研究GC盒，Sp1和Sp3在oxPL诱导PAFAH中的作用，并确定PAFAH诱导是否依赖于Sp1/ Sp3的磷酸化和/或Sp1与活化的STAT 3的关联。为此，我们组建了一支独特的调查团队，其中包括Norbert Leitinger博士。（弗吉尼亚大学），oxPL研究的先驱，Zendra Zehner博士（VCU），以及确定oxPL诱导PAFAH的分子机制所需的转录测定，DNAP和ChIP分析的专家，以及首席研究员Suzanne Barnard博士，PAFAH酶和树突细胞（DC）生物学方面的专家。总之，拟议的研究应该提供深入了解的生理机制，控制PAFAH表达血管疾病。我们预计，我们的研究可能会发现控制PAFAH的新机制，最终可能导致治疗，以防止动脉粥样硬化和其他血管疾病的开始或缓慢进展。项目叙述：血小板活化因子乙酰水解酶（PAFAH）是脂蛋白相关的磷脂酶A2，与心血管疾病的发病率和严重程度相关。这组研究人员的初步研究表明，PAFAH是由氧化低密度脂蛋白（LDL）颗粒中的氧化磷脂（oxPL）诱导的。本研究的目的是描绘PAFAH诱导oxPL的分子机制，希望这将揭示新的机制，调节酶，从而控制心血管疾病。

DESCRIPTION (provided by applicant): Platelet-activating factor acetylhydrolase (PAFAH) also known as the lipoprotein-associated phospholipase A2 is a calcium-independent acylhydrolase that catabolizes oxidized phospholipids (oxPL) in the oxidized LDL (oxLDL) particle as well as the lipid mediator platelet-activating factor (PAF). Many recent studies have correlated PAFAH activity with severity of vascular disease and atherosclerosis. Despite the unquestioned importance of this enzyme in the pathogenesis of atherosclerosis and other vascular disease, very little is known about mechanisms regulating its expression and activity. Our preliminary studies indicate that PAFAH expression is induced by oxidized phospholipids (oxPL), derivatives of 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine, a component of oxLDL. This provides us with a unique model that we will use to delineate the molecular mechanisms controlling PAFAH expression in the atherosclerotic lesion. We hypothesize that long chain, prostaglandin-like oxPL induce PAFAH by binding to the CD14/ TLR4 complex and the EP2 prostaglandin receptor on dendritic cells in the developing atherosclerotic lesion. These interactions are proposed to induce interleukin 6 (IL6) and the phosphorylation and activation of key transcription factors, events essential for optimal transcription of the PAFAH gene. To test this hypothesis, we propose the following specific aims: 1) Investigate Roles of EP2, IL6, and CD14/ TLR4 in oxPL Induction of PAFAH and 2) Investigate Molecular Mechanisms of oxPL Induction of PAFAH. In Aim 1, we will test the hypothesis that oxPL binding to EP2 and/ or the TLR4-CD14 receptor complex induces IL6 and that this cytokine is essential for optimal PAFAH induction. In Aim 2, we will investigate role of a GC-box, Sp1, and Sp3 in induction of PAFAH by oxPL and determine whether PAFAH induction is dependent on phosphorylation of Sp1/ Sp3 and/ or the association of Sp1 with activated STAT3. We have assembled a unique team of investigators for this purpose, including Dr. Norbert Leitinger (University of Virginia), a pioneer in the study of oxPL, Dr. Zendra Zehner (VCU), and expert in the transcriptional assays, DNAP, and ChIP analyses required for determination of the molecular mechanism of PAFAH induction by oxPL, and the Principal Investigator, Dr. Suzanne Barbour, an expert in the PAFAH enzyme and dendritic cell (DC) biology. Together, the proposed studies should provide insights into the physiological mechanisms that control PAFAH expression in vascular disease. We anticipate that our studies may uncover novel mechanisms to control PAFAH that could eventually result in treatments to prevent initiation or slow progression of atherosclerosis and other vascular diseases. Project Narrative: Platelet-activating Factor Acetylhydrolase (PAFAH) is the lipoprotein-associated phospholipase A2 that has been correlated with the incidence and severity of cardiovascular disease. Preliminary studies from this group of investigators indicate that PAFAH is induced by oxidized phospholipids (oxPL) in the oxidized low density lipoprotein (LDL) particle. The objective of this study is to delineate the molecular mechanisms of PAFAH induction by oxPL in hopes that this will uncover novel mechanisms for regulating the enzyme and thereby controlling cardiovascular disease.