Modulation of the arachidonic acid cascade by food polyphenols - A comprehensive analysis of their effects on 5-Lipoxygenase, Cytochrome-P450 and autoxidation pathways

类花生酸和其他多不饱和脂肪酸（PUFA）的氧化代谢产物参与细胞内信号传导以及疼痛调节，并代表最有效的内源性炎症介质类别之一。由于其重要的生理作用，世界上一半以上的药物直接影响这些氧化脂质的形成。虽然一些研究表明，营养改变了内源性氧化脂的模式，大多数食物成分，如多酚，对氧化脂的形成的影响尚未调查。此外，证明多酚对花生四烯酸（AA）级联和炎症的显著影响的研究集中于分析单个酶的表达水平或单个类花生酸的水平。这些方法忽视了生物过程的调节依赖于大量的氧脂素。因此，监测单个氧化脂质绘制了一个不完整的图片。因此，在这个项目中，我们将机械地研究多酚对AA级联的影响。为此，我们将采用液相色谱质谱法（LC-MS）作为靶向代谢组学方法，对AA级联的所有分支的一组全面的氧脂素进行定量。我们将分析多酚对环氧化酶，脂氧合酶，细胞色素P450和环氧化物水解酶途径的AA级联在不同的细胞培养试验，以及在无细胞的方法的活性的影响。此外，我们将利用氧化脂质模式来监测炎症过程中发生的氧化应激及其通过食物产生的抗氧化剂的改善。有了这些高含量的筛选工具，我们不仅可以发现潜在的活性多酚，还可以获得有关其作用模式的信息，有关其效力的定量数据以及有关其影响范围的定性数据。随后，将在脂多糖诱导的脓毒症和葡聚糖硫酸钠诱导的急性和慢性结肠炎的啮齿动物模型中研究最有效的化合物。氧化脂质模式的变化将与炎症的生理学、组织病理学和分子生物学参数相关。总的来说，这个项目将导致更深入地了解营养诱导的氧化脂质模式的改变及其对炎症的影响。我们将确定这些多酚，有力地改变调节脂质水平，阐明其作用机制模式，并评估其在体内的生物学效应。所获得的数据将有助于了解多酚对炎症的一般影响，并评估其在调节急性和慢性炎症过程中的潜在治疗价值。

Eicosanoids and oxidative metabolites of other poly-unsaturated fatty acids (PUFAs) are involved in intracellular signaling as well as the regulation of pain, and represent one of the most potent classes of endogenous mediators of inflammation. Due to their crucial physiological role more than half of the world pharmaceuticals directly influence the formation of these oxylipins. Although several studies demonstrate that nutrition alters the endogenous oxylipin pattern, the influence of most food ingredients, such as polyphenols, on the formation of oxylipins has not been investigated yet. Moreover, the studies demonstrating a significant impact of polyphenols on the arachidonic acid (AA) cascade and inflammation focused on the analysis of expression levels of single enzymes or the levels of single eicosanoids. These approaches disregard that the regulation of biological processes relies on a multitude of oxylipins. Thus, monitoring of single individual oxylipins draws an incomplete picture.Therefore, in this project, we will mechanistically investigate the impact of polyphenols on the AA cascade. For this purpose we will employ liquid chromatography mass spectrometry (LC-MS) for the quantification of a comprehensive set of oxylipins of all branches of the AA cascade as targeted metabolomics approach. We will analyze the effect of polyphenols on the activity of cyclooxgenase, lipoxygenase, cytochrome P450 and epoxide hydrolase pathways of the AA cascade in different cell culture assays as well as in cell free approaches. Moreover, we will utilize the oxylipin pattern to monitor oxidative stress occurring during inflammation and its amelioration by food-born antioxidants. With these high-content screening tools, we will not only discover potentially active polyphenols, but also gain information on their mode of action, quantitative data on their potency and qualitative data on the breadth of their effects. Subsequently, the most potent compounds will be investigated in rodent models of lipopolysaccharid induced sepsis and dextran sulfate sodium induced acute and chronic colitis. Changes in the oxylipin pattern will be correlated to physiological, histopathological and molecular biological parameters of inflammation. Overall, this project will lead to a deeper understanding of the nutritionally induced alterations in oxylipin patterns and their impact on inflammation. We will identify those polyphenols that potently alter regulatory lipid levels, elucidate their mechanistic mode of action and evaluate their biological effects in vivo. The obtained data will help to understand the general influence of polyphenols on inflammation and to evaluate their potential therapeutic value in modulating acute and chronic inflammatory processes.