Gut mucosal mast cells are activated by fat absorption: physiology and mechanism

描述（由申请人提供）：在糖尿病、肥胖或心血管疾病患者中，循环促炎介质经常升高。我们观察到肠粘膜肥大细胞（MMC）被脂肪吸收激活。在我们建立的有意识淋巴瘘大鼠中，我们发现脂肪吸收激活肠粘膜肥大细胞（MMC）并使其脱颗粒，肠道淋巴中MMC标记蛋白酶II明显增加（约20倍），相对于空腹淋巴（预摄入脂肪）。从肠道生理学的角度来看，这一观察结果非常令人兴奋；它可能具有深远的临床意义。我们每天消耗脂肪几次，所以MMC在白天被频繁激活。随着MMC的激活和脱颗粒，肠道可能是循环促炎介质的重要贡献者。我们假设脂质的吸收和乳糜微粒的形成和分泌激活了MMCs。这种作用是脂质特异性的，不为蛋白质或碳水化合物所共有。我们进一步假设，急性和慢性消耗脂肪，脂肪的类型，影响其激活MMCs的能力，效力的顺序是ω - 6（亚油酸和花生四烯酸）bb0 ω - 9（油酸）bb1 ω - 3脂肪酸（亚麻酸）。为了验证这些假设，我们提出了以下研究：我们将确定肠道脂肪吸收和MMC激活之间的关系。目的：1)通过测量脂肪吸收过程中淋巴和组织中肥大细胞介质的释放来进一步表征MMC的激活，我们将通过肥大细胞缺陷的Ws/Ws大鼠来确定脂质介质、细胞因子、趋化因子的释放是否主要来自MMC；2)确定MMC稳定剂的存在或不存在是否会影响膳食脂质的肠道摄取和淋巴运输。我们还将研究Ws/Ws大鼠的脂肪吸收。具体目标2。我们假设，由饮食中的多不饱和长链脂肪酸（FA）亚油酸（18:2,n-6）产生的CMs是肥大细胞激活的一个强有力的决定因素，这种作用受到脂肪酸类型的不同影响。我们将确定急性和慢性暴露于不同程度的不饱和脂肪酸和类型（n-3， n-6和n-9）脂肪酸对MMC激活的脂肪吸收。具体目标3。我们将验证我们的假设，即脂肪酸或肠细胞对CMs的处理会导致导致MMCs激活的因子（例如NF-kB反应）的释放。为了验证我们的假设，我们提出了以下两个子目标。1)在体外研究不同类型FAs通过NF-kB信号通路对大鼠嗜碱性白血病细胞（RBL-2H3细胞，大鼠MMC的替代物）激活的影响；2)研究肠道上皮细胞系（Caco-2细胞）乳糜淋巴及其成分乳糜微粒（CMs）或含有乳糜微粒的培养基对红细胞- 2h3细胞NF-kB反应的相互作用。如果结果是肯定的，我们将隔离所涉及的因素。

DESCRIPTION (provided by applicant): In patients with diabetes, obesity, or cardiovascular disease, the circulating proinflammatory mediators are often elevated. We made the novel observation that intestinal mucosal mast cells (MMC) are activated by fat absorption. In our well established conscious lymph fistula rats, we showed that fat absorption activates the intestinal mucosal mast cells (MMC) and they degranulate as evidenced by the marked increase (~ 20 fold) of the MMC marker protease II in intestinal lymph relative to fasting lymph (pre-ingestion of fat). This observation is very exciting from the standpoint of gut physiology; it may have profound clinical implications. We consume fat a few times a day and so the MMC are activated frequently during the day. With the activation and degranulation of MMC, the gut may be an important contributor to the circulating proinflammatory mediators. We hypothesized that the absorption of lipids and the formation and secretion of chylomicrons (CMs) activate the MMCs. This action is lipid specific and is not shared by proteins or carbohydrates. We further hypothesize that acute and chronic consumption of fat, the type of fat, influence its ability to activate the MMCs and that the order of potency is omega 6 (linoleate & arachidonate) > omega 9 (oleate) > omega 3 fatty acids (linolenate). To test these hypotheses, we proposed the following studies: Specific Aim 1. We will determine the relationship between intestinal fat absorption and MMC activation. Subaims: 1) to further characterize the activation of MMC by measuring the release of mast cell mediators in lymph and tissue during fat absorption, we will determine if the release of lipid mediators, cytokines, chemokines is mostly from MMC by using the mast cell deficient Ws/Ws rats; 2) to determine if the presence or absence of MMC stabilizers affects intestinal uptake and lymphatic transport of dietary lipids. We will also study fat absorption in Ws/Ws rats. Specific Aim 2. We hypothesize that the production of CMs, resulting from dietary polyunsaturated long-chain fatty acids (FA) linoleic acid (18:2, n-6) is a potent determinant of the mast cell activation and this action is differently affected by the type of fatty acids. We will determine the acute and chronic exposure to fatty acids with different degree of unsaturation and the type (n-3, n-6, and n-9) fatty acids on MMC activation by fat absorption. Specific Aim 3. We will test our hypothesis that fatty acid or the processing of CMs by the enterocytes results in the release of factor/factors that cause the activation of the MMCs (e.g. NF-kB response). To test our hypotheses, we have proposed the following 2 subaims. 1) We will study the in vitro effects of different types of FAs on rat basophil leukemia cells (RBL-2H3 cells, a surrogate of the rat MMC) activation through NF-kB signaling pathway; 2) We will study the interaction of chylous lymph and its components e.g. chylomicrons (CMs) or the media containing CMs by the intestinal epithelial cell line (Caco-2 cells) on RBC-2H3 cell NF-kB response. If the outcome is positive, we will isolate the factor/factors involved. PUBLIC HEALTH RELEVANCE: By focusing on the relationship between fat absorption and mucosal mast cell degranulation, our proposed studies will identify the role of gut-derived inflammatory factors in the normal process of dietary fat absorption and transport, identify which inflammatory agents are generated by normal fat absorption and may enter the general circulation, and contribute to the etiology of various diseases such as diabetes, metabolic syndrome, and atherosclerosis and coronary heart disease. Our results will also greatly enhance our understanding of function of gastrointestinal lymphatics in health and disease by clarifying the changes which occur in GI lymph parameters during normal physiological states such as fasting and lipid absorption, laying the ground work for future studies comparing the differences in lymph between diseased states and normal physiological states.