Colonic Cytokinetics and Cell Signaling: Dietary Effects

描述（由申请人提供）： 有令人信服的数据表明n-3多不饱和脂肪酸（PUFA）的保护作用，二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）对结肠癌的作用。相反，富含n-6 PUFA的膳食脂质，例如，亚油酸（LA）和花生四烯酸（AA），促进结肠肿瘤的发展。这是很重要的，因为典型的西方饮食中含有的n-6比n-3多不饱和脂肪酸多10到20倍。不幸的是，到目前为止，还缺乏一个统一的机制假说来解释为什么n-3 PUFA与n-6 PUFA（美国饮食中PUFA的主要饮食形式）相比选择性地抑制结肠癌。在当前的资助期内，我们证明了（i）饮食n-3 PUFA诱导结肠细胞膜微区的扰动（小窝）脂质组合物，抑制致癌信号传导和Ras蛋白运输;（ii）n-3 PUFA取代了结肠细胞膜磷脂中的花生四烯酸（AA），特别是EPA被代谢成新的3系列E-前列腺素（PGE 3），推定的抗肿瘤-环氧合酶（考克斯）代谢物;和（iii）当将高度可发酵的纤维、果胶（或其发酵产物-丁酸盐）添加到饮食中时，n-3 PUFA的促凋亡-化学保护作用增强。我们推测，未能解决膳食脂肪和纤维之间的相互作用可能解释为什么纤维的化学保护作用在前瞻性队列研究中被掩盖。因此，该提案的总体目标是进一步阐明膳食n-3 PUFA和可发酵纤维如何上调结肠细胞中的凋亡效应机制，从而降低结肠癌风险。将使用一系列实验模型，包括体内（注射氧化偶氮甲烷的氧化应激SOD 2 +/-、Gpx 4 +/-敲除、Gpx 4 Tg和fat-1转基因小鼠模型）和体外（正常和恶性转化的小鼠和人结肠细胞系），以检验我们的主要假设，即n-3 PUFA和丁酸盐以协调的方式发挥作用，从而增强膀胱介导的细胞凋亡。作为一个互补的假设，我们提出，DHA和可能EPA改变结肠细胞膜脂质微区组成，从而有利地调节类花生酸代谢和细胞外信号从表面受体到下游信号网络的中继。提出了以下具体目标：目标1将确定n-3 PUFA和丁酸相互作用调节内源性目的#2将确定n-3 PUFA改变Ras依赖性信号的时空分离的机制;和目标#3将表征新的EPA衍生的环氧合酶产物PGE 3的推定化学保护性质。EPA和DHA的消耗可能被证明是结肠癌的有效辅助治疗。因此，研究n-3多不饱和脂肪酸（n-3 polyunsaturated fatty acids，PUFA）对细胞信号通路的调控作用，降低结肠癌发生的风险，是一个非常重要的课题。与PA-07-100的目标和范围一致，通过营养组合优先考虑癌症预防的分子靶点，本提案的总体目标是阐明膳食n-3 PUFA和可发酵纤维如何上调结肠细胞中的凋亡效应机制，从而降低结肠癌风险。

DESCRIPTION (provided by applicant): There are cogent data indicating a protective effect of n-3 polyunsaturated fatty acids (PUFA) e.g., eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on colon cancer. In contrast, dietary lipids rich in n-6 PUFA, e.g., linoleic acid (LA) and arachidonic acid (AA), enhance the development of colon tumors. This is significant because the typical Western diet contains 10 to 20 times more n-6 than n-3 PUFA. Unfortunately, to date, a unifying mechanistic hypothesis addressing why n-3 PUFA selectively suppress colon cancer compared to n-6 PUFA (the major dietary form of PUFA in the U.S. diet) is lacking. In the current funding period, we demonstrated that (i) dietary n-3 PUFA induce perturbations in colonocyte membrane microdomain (caveolae) lipid composition, suppressing oncogenic signaling and Ras protein trafficking; (ii) n-3 PUFA supplant arachidonic acid (AA) in colonocyte membrane phospholipids, with EPA in particular being metabolized into a novel 3-series E-prostaglandin (PGE3), a putative antitumorigenic-cyclooxygenase (COX) metabolite; and (iii) the proapoptotic- chemoprotective effect of n-3 PUFA is enhanced when a highly fermentable fiber, pectin (or its fermentation product-butyrate) is added to the diet. We postulate that the failure to address an interaction between dietary fat and fiber may explain why the chemoprotective effects of fiber are obscured in prospective cohort studies. Therefore, the overall goal of this proposal is to further elucidate how dietary n-3 PUFA and fermentable fiber up-regulate apoptosis effector mechanisms in colonocytes, thereby reducing colon cancer risk. An array of experimental models will be used, including in vivo (azoxymethane-injected oxidatively stressed SOD2+/-, Gpx4+/- knock outs, Gpx4Tg and fat-1 transgenic mouse models) and in vitro (normal and malignant transformed mouse and human colonocyte cell lines) in order to test our primary hypothesis that n-3 PUFA and butyrate work in a coordinated manner to potentiate mitochondrial-mediated apoptosis. As a complementary hypothesis, we propose that DHA and possibly EPA alter colonocyte membrane lipid microdomain composition, thereby favorably modulating eicosanoid metabolism and the relay of extracellular signals from surface receptors to downstream signaling networks. The following specific aims are proposed: Aim #1 will determine the mechanisms by which n-3 PUFA and butyrate interaction modulate intrinsic (mitochondria-mediated) cell death signaling in the colon; Aim #2 will determine the mechanisms by which n-3 PUFA alter the spatio-temporal segregation of Ras-dependent signals; and Aim #3 will characterize the putative chemoprotective properties of a novel EPA-derived cyclooxygenase product, PGE3. The consumption of EPA and DHA may prove an effective adjuvant therapy in colon cancer. Therefore, it is both appropriate and timely to determine precisely how n-3 PUFA modulate cell signaling networks and reduce colon cancer risk.A growing body of literature supports the contention that bioactive food components containing n-3 polyunsaturated fatty acids (PUFA) are important in suppressing colon cancer. Consistent with the objectives and scope of PA-07-100, Prioritizing molecular targets for cancer prevention with nutritional combinations , the overall goal of this proposal is to elucidate how dietary n-3 PUFA and fermentable fiber up-regulate apoptosis effector mechanisms in colonocytes, thereby reducing colon cancer risk.