Cleavage Products of Dietary Carotenoids: Occurrence & Nutritional Function

描述（由申请人提供）：食用膳食类胡萝卜素与降低心血管疾病和癌症的风险有关。人类通过摄入水果和蔬菜获得类胡萝卜素。历史上，人们认为某些类胡萝卜素最重要的作用是它们通过在中心双键上裂解成必需微量营养素维生素A（视黄醇）的代谢转化。最近，人们对膳食类胡萝卜素的“偏心分裂”产生伪胡萝卜素及其可能的氧化产生伪胡萝卜素酸的现象非常感兴趣。这些类胡萝卜素的代谢物可能在类胡萝卜素的非维生素A活性中发挥重要作用。目前还没有对这些化合物在人类食物和组织中的含量进行定量分析。完整的类胡萝卜素对这些化合物的代谢程度的证明及其作用机制的研究尚缺乏。我们将填补这些知识空白。(1)我们确定了底物特异性，并对两种已知的人类类胡萝卜素代谢酶（BCO1和BCO2）的产物进行了表征。这些研究也将建立细胞视黄醇结合蛋白和类胡萝卜素裂解酶在这些酶的代谢功能中的关系。(2)我们将利用分析化学来表征人类血浆、组织和食物中载胡萝卜素和伪胡萝卜素酸的存在并量化它们的含量，从而证明类胡萝卜素在人类中的相关性。这将通过在受试者食用一份含有营养相关量的-胡萝卜素或番茄红素的番茄汁后立即测定餐后血浆中亲本类胡萝卜素和代谢物的浓度，以及在每天食用4周后测定血浆中的稳态浓度来完成。(3)我们将使用LNCaP细胞，一种表现出番茄红素依赖性生长抑制并表达BCO2的人类前列腺癌细胞，在番茄红素处理的细胞中寻找载红素和载红素酸，以确定番茄红素在这些细胞中是否代谢以及在多大程度上代谢。我们将使用人类单核/巨噬细胞U937，这种细胞在类胡萝卜素处理下被抑制生长并诱导分化，来寻找母体类胡萝卜素的代谢。(4)我们将以多种方式探索类胡萝卜素代谢物的生物学（营养）功能：(a)我们将使用我们已经证明对膳食类胡萝卜素处理具有特征性功能反应的细胞系，并询问这些反应是否通过直接使用裂解产物获得；(b)我们将询问这些裂解产物和代谢物是否直接激活或拮抗配体依赖性转录因子。(c)我们将给BCO1敲除小鼠喂食β -胡萝卜素和番茄红素，以评估BCO2对这些类胡萝卜素的代谢程度以及类胡萝卜素代谢物在整个动物中的代谢作用。该结果将增强我们对富含水果和蔬菜的饮食促进健康作用的分子基础的理解。

DESCRIPTION (provided by applicant): Consumption of dietary carotenoids is associated with a decreased risk of cardiovascular disease and cancer. Humans obtain carotenoids in the diet by ingesting fruits and vegetables. Historically, it was thought that the most important role of some carotenoids was their metabolic conversion via cleavage at the central double bond to the essential micronutrient, vitamin A (retinol). There is intense recent interest in the "eccentric cleavage" of dietary carotenoids to apocarotenals and their possible oxidation to apocarotenoic acids. These metabolites of carotenoids may play important roles in the non-vitamin A activities of carotenoids. There are no quantitative analyses of the levels of these compounds in human foods and tissues. Demonstrations of the extent of metabolism of intact carotenoids to these compounds and studies of their mechanisms of action are lacking. We will address these gaps in knowledge. (1) We determine the substrate specificity and characterize the products of the two known human carotenoid-metabolizing enzymes (BCO1 & BCO2). These studies will also establish the relationship of cellular retinol-binding proteins and carotenoid cleavage enzymes in the metabolic function of these enzymes. (2) We will demonstrate the relevance of apocarotenoids in humans using analytical chemistry to characterize the presence and quantify the amounts of apo-carotenals and apocarotenoic acids in human plasma and tissues and in foods. This will be accomplished by quantitation of the immediate post- prandial plasma concentrations of parent carotenoids and metabolites after subjects consume a single serving of tomato juice containing nutritionally relevant amounts of beta-carotene or lycopene as well as by quantitation of steady state concentrations in plasma after 4 weeks of daily consumption. (3) We will use the LNCaP cell, a human prostate cancer cell that demonstrates lycopene-dependent growth inhibition and expresses BCO2 to look for apolycopenals and apolycopenoic acids in lycopene-treated cells to address whether and to what extent lycopene is metabolized in these cells. We will use the human monocyte/macrophage cell, U937, which is growth inhibited and induced to differentiate with carotenoid treatment, to look for metabolism of the parent carotenoids. (4) We will probe the biological (nutritional) function of carotenoid metabolites in multiple ways: (a) we will use cell lines that we have shown have characteristic functional responses to treatment with dietary carotenoids and ask whether these responses are obtained by direct treatment with the cleavage products and (b) we will ask whether these cleavage products and metabolites directly activate or antagonize ligand-dependent transcription factors. (c) We will feed the BCO1 knockout mouse both beta-carotene and lycopene to assess the extent of the metabolism of these carotenoids by BCO2 and the metabolic effects of carotenoid metabolites in the whole animal. The results will enhance our understanding of the molecular basis for the health-promoting effects of diets rich in fruits and vegetables. PUBLIC HEALTH RELEVANCE: Increased consumption of dietary carotenoids is associated with decreased risk of both cardiovascular disease and certain cancers. The proposed research will document the occurrence of new pathways of metabolism of dietary carotenoids (the health-promoting, colored pigments in fruits and vegetables) in humans. The research will also explore possible mechanisms that may explain the basis of the health-promoting effects of these components of the diet.