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，是一种钙无关的酰基水解酶，可分解氧化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）。最近的许多研究将PAFAH活性与血管疾病和动脉粥样硬化的严重程度联系起来。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具有不容置疑的重要性，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的表达是由氧化磷脂（oxPL）诱导的，氧化磷脂是1-棕榈酰-2-花生四烯酰基- asn -甘油-3-磷脂胆碱的衍生物，是oxLDL的一种成分。这为我们提供了一个独特的模型，我们将用它来描述控制动脉粥样硬化病变中PAFAH表达的分子机制。我们假设长链前列腺素样oxPL通过结合树突状细胞上的CD14/ TLR4复合物和EP2前列腺素受体诱导PAFAH。这些相互作用被认为可以诱导白细胞介素6 （IL6）和关键转录因子的磷酸化和激活，这是PAFAH基因最佳转录所必需的事件。为了验证这一假设，我们提出了以下具体目标：1)研究EP2、IL6和CD14/ TLR4在oxPL诱导PAFAH中的作用；2)研究oxPL诱导PAFAH的分子机制。在Aim 1中，我们将验证oxPL与EP2和/或TLR4-CD14受体复合物结合诱导IL6的假设，并且该细胞因子对于最佳的PAFAH诱导是必不可少的。在Aim 2中，我们将研究GC-box、Sp1和Sp3在oxPL诱导PAFAH中的作用，并确定PAFAH的诱导是否依赖于Sp1/ Sp3的磷酸化和/或Sp1与活化的STAT3的关联。为此，我们组建了一个独特的研究团队，包括Norbert Leitinger博士（弗吉尼亚大学），oxPL研究的先驱，Zendra Zehner博士（VCU），转录分析专家，DNAP和ChIP分析需要确定oxPL诱导PAFAH的分子机制，首席研究员Suzanne Barbour博士，PAFAH酶和树突细胞（DC）生物学的专家。总之，拟议的研究应提供对血管疾病中控制PAFAH表达的生理机制的见解。我们预计我们的研究可能会发现控制PAFAH的新机制，从而最终导致预防动脉粥样硬化和其他血管疾病的发生或减缓进展的治疗方法。项目描述：血小板活化因子乙酰水解酶（PAFAH）是一种与心血管疾病的发病率和严重程度相关的脂蛋白相关磷脂酶A2。这组研究者的初步研究表明，PAFAH是由氧化低密度脂蛋白（LDL）颗粒中的氧化磷脂（oxPL）诱导的。本研究的目的是描述由oxPL诱导PAFAH的分子机制，希望这将揭示调节酶的新机制，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.