Lipolytic Mechanisms of PPAR Activation

描述(申请人提供)：过氧化物酶体增殖物激活受体(PPAR)是配体激活的核受体，对脂肪生成、血糖控制和脂肪代谢的转录调控起核心作用。最近的工作建立了PPAR在血管系统中的表达和作用，包括内皮细胞(EC)。尽管取得了这些进展，但重大悬而未决的问题依然存在。PPAR领域的一个中心悬而未决的问题是内源性PPAR配体的性质，它可能概括了合成的PPAR激动剂的作用，或者这些自然介质是如何在生理条件下产生的。同样，尽管广泛的研究表明PPAR与脂质生物学有关，但关于脂代谢途径和随后的PPAR激活之间的联系的数据很少。这种洞察力可能会对代谢紊乱和动脉粥样硬化产生重大影响。 有数据表明，脂蛋白脂酶(LPL)是富含甘油三酯的脂蛋白代谢的中心酶，是PPARpha激活的内源性机制。使用功能获得(LPL过表达)和功能丧失(PPARα缺乏)模型的体外和体内证据都支持这一说法。LPL介导的PPAR-α激活独立于LPL已知的非酶效应，并且对于PPAR(PPARpha；&gt；；PPARDelta&gt；PPARGamma)、脂蛋白底物(VLDL&gt；&gt；LDL&gt；高密度脂蛋白)和不含脂肪酶的脂肪酶具有选择性，并检测到其他脂肪酸释放脂肪酶。此外，单酰甘油(MAG)是一种LPL特异性产物，被认为是一种新的PPARpha激活剂，有助于这些LPL反应。我们的中心假设是脂解是内源性PPAR激活的主要途径，其远端效应取决于脂蛋白底物、脂肪酶和靶向PPAR的不同性质。这项提案概述了通过这些相同的参数定义PPAR激活的脂解机制的实验。初步数据表明，LPL介导的PPARpha激活的独特贡献者将被研究，包括MAG作为一个鲜为人知的信号分子和配体传递机制。脂蛋白底物和其他甘油三酯脂肪酶的关键脂解变量将根据它们的PPARα、-增量和-伽马效应进行检查。这些研究包括对现有的、特征良好的极低密度脂蛋白和来自小鼠和人类的血浆样本的脂解PPAR反应的测量。在小鼠模型和人类中存在具有一系列LPL活性的遗传LPL变体，这些变体将在体外和体内用于确定LPL功能分级如何改变已建立的PPAR反应。这些LPL模型包括通过瞬时LPL表达拯救的其他致命的LPL缺陷小鼠。通过这些程序性的努力，将深入了解脂解作为选择性内源性PPAR激活的机制及其远端转录效应。

DESCRIPTION (provided by applicant): Peroxisome Proliferator-Activated Receptors (PPARs) are ligand-activated nuclear receptors central to the transcriptional regulation of adipogenesis, glucose control and lipid metabolism. More recent work establishes PPAR expression and effects in the vasculature, including endothelial cells (EC). Despite these advances, major unresolved issues persist. One central unanswered question in the PPAR field is the nature of endogenous PPAR ligands, which might recapitulate the effects of synthetic PPAR agonists, or how such natural mediators are produced under physiologic conditions. Similarly, despite extensive studies implicating PPARs in lipid biology, little data exists regarding the links between pathways of lipid metabolism and subsequent PPAR activation. Such insight could have major implications for metabolic disorders and atherosclerosis. Data is presented establishing lipoprotein lipase (LPL), the central enzyme in triglyceride-rich lipoprotein metabolism, as an endogenous mechanism for PPARalpha activation. In vitro and in vivo evidence, using both gain of function (LPL overexpression) and loss of function (PPARalpha-deficiency) models, support this claim. LPL-mediated PPAR-alpha activation is independent of LPL's known non-enzymatic effects and is selective as to PPAR (PPARalpha>>PPARdelta> PPARgamma), lipoprotein substrate (VLDL>>LDL>HDL), and lipase- absent with other fatty acid-releasing lipases tested. Furthermore, monoacylglycerol (MAG), a LPL-specific product, is identified as a novel PPARalpha activator contributing to these LPL responses. Our central hypothesis is that lipolysis is a major pathway for endogenous PPAR activation, with distal effects determined by the varying nature of lipoprotein substrate, lipase, and targeted PPAR. This proposal outlines experiments to define lipolytic mechanisms for PPAR activation across these same parameters. Unique contributors to LPL-mediated PPARalpha activation, as suggested by preliminary data, will be studied, including MAG as a little studied signaling molecule and mechanisms of ligand delivery. Key lipolytic variables of lipoprotein substrate and other tiglyceride lipases will be examined in terms of their PPARalpha, -delta, and -gamma effects. These studies include measures of lipolytic PPAR responses on available well-characterized VLDL and plasma samples from both mice and humans. The existence of genetic LPL variants in mouse models and in humans, which have a range of LPL activity, will be utilized in vitro and in vivo to determine how graded LPL function alters well-established PPAR responses. These LPL models include the otherwise lethal LPL-deficient mice rescued by transient LPL expression. Through these programmatic efforts, insight will be gained into lipolysis as a mechanism for selective endogenous PPAR activation, and its distal transcriptional effects.