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)，如二十碳五烯酸(EPA)和二十二碳六烯酸(DHA)，对结肠癌具有保护作用。相比之下，富含n-6多不饱和脂肪酸的膳食脂质，如亚油酸(LA)和花生四烯酸(AA)，促进了结肠癌的发展。这一点很重要，因为典型的西式饮食含有的n-6多出n-3多不饱和脂肪酸10到20倍。不幸的是，到目前为止，缺乏一个统一的机制假说来解释为什么n-3多不饱和脂肪酸比n-6多不饱和脂肪酸(美国饮食中多不饱和脂肪酸的主要饮食形式)选择性地抑制结肠癌。在目前的资助期间，我们证明了(I)饮食n-3PUFA引起结肠细胞膜微区(小窝)脂组成的扰动，抑制致癌信号和RAS蛋白的运输；(Ii)n-3PUFA替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(PGE3)，可能是抗肿瘤发生环氧合酶(COX)的代谢物；以及(Iii)当在饮食中添加高度可发酵的纤维、果胶(或其发酵产物丁酸)时，n-3PUFA的促凋亡-化学保护作用得到增强。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了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，这项建议的总体目标是进一步阐明饮食中n-3多不饱和脂肪酸和可发酵纤维如何上调结肠细胞的凋亡效应机制，从而降低结肠癌风险。我们将使用一系列实验模型，包括体内(偶氮甲烷注射氧化应激SOD2/-、Gpx4/-敲除、Gpx4Tg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类结肠癌细胞系)，以验证我们的初步假设，即n-3多不饱和脂肪酸和丁酸盐协同作用加强线粒体介导的细胞凋亡。作为补充假设，我们认为DHA和可能的EPA改变了结肠细胞膜脂微域的组成，从而有利于调节二十烷类代谢和细胞外信号从表面受体到下游信号网络的传递。提出了以下具体目标：目标1将确定n-3多不饱和脂肪酸和丁酸相互作用调节结肠固有(线粒体介导的)细胞死亡信号的机制；目标2将确定n-3多不饱和脂肪酸改变RAS依赖信号的时空分离的机制；目标3将表征一种新的EPA衍生的环氧合酶产物PGE3的化学保护特性。EPA和DHA的消耗可能被证明是结肠癌的一种有效的辅助治疗。因此，准确地确定n-3多不饱和脂肪酸如何调节细胞信号网络和降低结肠癌风险是合适和及时的。越来越多的文献支持含有n-3多不饱和脂肪酸(PUFA)的生物活性食品成分在抑制结肠癌方面的重要性。与PA-07-100的目标和范围一致，本建议的总体目标是阐明饮食n-3多不饱和脂肪酸和可发酵纤维如何上调结肠细胞的凋亡效应机制，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.