THE INTEGRATED ROLES OF IPLA2G IN OBESITY, INFLAMMATION AND HEPATIC DYSFUNCTION

描述（由申请人提供）：工业化社会中死亡率和发病率的主要原因是高脂饮食诱导的肥胖症（HFDIO）的多种下游后遗症。以前，我们证明了钙非依赖性磷脂酶A2的重要性？（iPLA2？）作为一个中央酶介质整合细胞信号和有机体生物能通过全球iPLA 2的生成和表征？KO小鼠。值得注意的是，iPLA 2？基因敲除小鼠对HFDIO完全耐受，并出现胰岛素抵抗。然而，由于在整个动物中多器官系统受到影响，iPLA 2？？敲除，iPLA 2的机械作用？？在每种组织中，导致该小鼠对HFDIO完全耐受的原因尚不清楚。所提出的研究的首要目标是组织的机械确定， iPLA 2？？的细胞器特异性作用促进炎症、适应不良的细胞信号传导和功能失调的生物能量学，导致胰岛素抵抗和HFDIO的病理性终末器官后遗症。因此，我们产生了含有iPLA 2？？floxed构建体的小鼠。活性位点，该位点已与组织特异性Cre小鼠杂交，导致产生肝细胞特异性和骨骼肌肌细胞特异性iPLA 2？？敲除小鼠拟议的研究将协同使用这些使能遗传模型，结合我们开发的综合脂质组学和代谢组学平台，以确定iPLA 2？参与HFDIO期间肥胖、炎症和胰岛素抵抗的发展。第一个具体的目标将确定肝细胞特异性iPLA 2？？在介导脂质第二信使的产生，脂质代谢，脂肪肝和生物能量学在HFDIO不受iPLA 2？？在生殖系敲除中存在的其他细胞类型中的功能丧失。在特定目标2中，我们将研究骨骼肌肌细胞特异性消融iPLA 2？？对骨骼肌代谢、胰岛素抵抗和线粒体功能障碍的影响，这些在WT小鼠的高脂肪喂养期间存在，但在种系敲除小鼠中得到拯救。最后，在具体目标3，我们将确定互动机制的作用，细胞器特异性亚型的iPLA 2？？通过转基因的重新引入无论是特定的或过氧化物酶异构体的iPLA 2？？进入肝细胞特异性iPLA 2？？敲除小鼠。此外，通过脂质第二信使产生的iPLA 2？？在肝细胞、骨骼肌肌细胞和脂肪细胞中，这些代谢交织组织之间介导的器官间通讯将通过我们开发/改进的协同高穿透性技术来鉴定。通过采用新型基因试剂、高质量准确度质谱技术和综合化学生理学方法的这种多学科方法，可以鉴定减轻HFDIO后遗症的新型药理学靶点。

DESCRIPTION (provided by applicant): The major causes of mortality and morbidity in industrialized societies result from the multiple downstream sequelae of high fat diet-induced obesity (HFDIO). Previously, we demonstrated the importance of calciumindependent phospholipase A2?(iPLA2?) as a central enzymic mediator integrating cellular signaling and organismal bioenergetics through the generation and characterization of a global iPLA2??KO mouse. Remarkably, the iPLA2??knockout mouse was completely resistant to HFDIO and the development of insulin resistance. However, due to the multiple organ systems affected in the whole animal iPLA2??knockout, the mechanistic roles of iPLA2??in each tissue contributing to the complete resistance of this mouse to HFDIO are unknown. The overarching goal of the proposed research is the mechanistic determination of the tissue- and Organelle-specific roles of iPLA2??in promoting inflammation, maladaptive cellular signaling and dysfunctional bioenergetics that result in insulin resistance and the pathologic end-organ sequelae of HFDIO. Accordingly, we generated mice containing a floxed construct of the iPLA2??active site which has been crossed with tissue specific Cre mice resulting in the generation of hepatocyte-specific and skeletal muscle myocyte-specific iPLA2??knockout mice. The proposed research will synergistically use these enabling genetic models in conjunction with the integrated lipidomics and metabolomics platforms we developed to identify the mechanisms through which iPLA2??participates in the development of obesity, inflammation, and insulin resistance during HFDIO. The first specific aim will identify the roles of hepatocyte-specific iPLA2??in mediating alterations in the generation of lipid 2nd messengers, lipid metabolism, hepatosteatosis and bioenergetics during HFDIO unencumbered by the effects of iPLA2??loss of function in other cell types that are present in the germline knockout. In Specific Aim 2, we will examine the effects of skeletal muscle myocyte-specific ablation of iPLA2??on skeletal muscle metabolism, insulin resistance and mitochondrial dysfunction that are present during high fat feeding in WT mice, but are rescued in the germline knockout mouse. Finally, in Specific Aim 3, we will determine the interactive mechanistic roles of the organelle-specific isoforms of iPLA2??through the transgenic reintroduction of either the mitochondrial-specific or peroxisomal-specific isoforms of iPLA2??into the hepatocyte-specific iPLA2??knockout mouse. Moreover, the mechanisms through which lipid 2nd messengers generated by iPLA2??in hepatocytes, skeletal muscle myocytes and adipocytes mediate inter-organ communication between these metabolically interwoven tissues will be identified through synergistic highly penetrating technologies we have developed/refined. Through this multidisciplinary approach employing novel genetic reagents, high mass accuracy mass spectrometry technologies and integrated chemophysiologic approaches, novel pharmacologic targets to attenuate the sequelae of HFDIO can be identified.