Polyunsaturated fatty acid (PUFA), inflammation and antioxidants

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THE PROBLEM Over the last century, Canadian diets has been inundated with n-6 polyunsaturated fatty acids (n-6 PUFA), like linoleic acid (LA). Along with such changes, various chronic diseases like diabetes and atherosclerosis, characterized by aberrant inflammation and oxidative stress have soared among Canadians. Fundamental knowledge gaps exist in our understanding of PUFA metabolism and inflammation/oxidative stress. BACKGROUND Most of our  knowledge of n-6 PUFA driven inflammation is centered around, arachidonic acid (ARA) which is actually lower in chronic diseases. However, recent research indicates that other n-6 PUFAs like linoleic (LA) and gamma linolenic acid (GLA) are major players during inflammation. In this regard, oxidative modifications of LA and elongase/desaturase mediated biotransformations of PUFA remain understudied. However, it is difficult to understand specific roles of LA as it is rapidly bioconverted sequentially to GLA, DGLA and finally to ARA in vivo. Thus overall, the LONG-TERM GOAL of this program is to elucidate the role of n-6 PUFA bio-transformation on inflammation and oxidative stress. My overall HYPOTHESIS is that under high LA availability, biological transformations of LA is vital in regulating inflammation/oxidative stress. THE PROGRAM The overarching aim of this program is to understand the roles of oxidative and metabolic modifications of LA and GLA towards inflammation and oxidative stress in cardiomyocytes and macrophages. We aim to target the impact of oxidized LA metabolites and 4-HNE in a high n-6 PUFA milieu both in vitro (using H9c2 and RAW 264.7 cell lines) and in vivo following various fatty acid diets (Objective 1). Next, we aim to identify the role of LA and GLA in causing inflammation/ oxidative stress independent of ARA (Objective 2). In this regard, we do possess a unique knockout mice model of Elovl5 gene deletion, which leads to in vivo accumulation of LA and ALA but reduces ARA. Thus, we will isolate cardiomyocytes and macrophages from wildtype littermates and Elovl5KO following monounsaturated (MUFA, as control) and n-6 PUFA-rich diet regimens. Mass Spectrometry and gas chromatography-will be used to analyze eicosanoids, fatty acids and oxidized lipid metabolites. Cell signaling, inflammation, oxidative stress will also be evaluated in these cells as shown in earlier studies. SIGNIFICANCE This application is to understand the fundamental role of enzymatic and non-enzymatic bio-transformations of individual PUFA in mammalian cells like macrophages and cardiomyocytes that are spatially related but functionally distinct along with their responses during inflammation and oxidative stress. I believe under the onslaught of high doses of dietary PUFAs, to which every Canadian is subjected to, this program if funded can lead to fundamental knowledge synthesis in PUFA metabolism and identification of true causes of n-6 PUFA mediated inflammation in mammalian cells.