integrin-mediated regulation of enterocyte lipid homeostasis

总结 肠道对膳食脂肪的吸收对于肥胖及其并发症的发展是重要的。很长的- 我们的目标是了解细胞表面基质受体整合素家族在调节肠道免疫反应中的作用。 养分吸收本申请的总体目标是阐明αvβ3和αvβ5在 整合素在调节小肠脂质稳态中的作用核心假设是，在对摄入的反应中， 肠细胞分泌整合素配体Mfge 8。然后Mfge 8结合肠上皮细胞αvβ3和αvβ5 整合素激活信号通路，增加膳食脂肪的吸收， 甘油三酯（TG）在内质网中的作用和TG在细胞质脂滴中的水解。这 该假设基于证明Mfge 8或αvβ3和αvβ5整联蛋白缺陷的小鼠 发展成脂肪代谢障碍，并对膳食脂肪挑战作出反应，吸收较少的脂肪，积累过量的细胞质 脂滴和分泌较少的乳糜微粒。这些研究的基本原理是， Mfge 8-整合素轴在肠脂质吸收中的作用对于理解其机制是必要的， 肠细胞调节脂蛋白的产生。这一假设将通过3个具体目标进行检验：1） 确定营养素如何诱导肠上皮细胞产生Mfge 8转录物和蛋白质，以及营养素的生理作用。 Mfge 8和αvβ3和αvβ5整联蛋白在膳食脂肪吸收中的作用; 2）确定Mfge 8和αvβ3和αvβ5整联蛋白在膳食脂肪吸收中的作用。 αvβ3和αvβ5整合素调节肠上皮细胞胞质脂滴中TG的水解;和3） 确定靶向αvβ3和αvβ5整联蛋白治疗肥胖症的治疗潜力。目的 1将检测Mfge 8在分化的Caco-2细胞和原代肠上皮细胞中的转录调控， Mfge 8分泌到肠腔中以响应不同的营养物，并鉴定Mfge 8的细胞表达。 通过使用肠上皮细胞特异性缺失的转基因小鼠获得脂肪吸收所需的Mfge 8来源 具有Mfge 8的肠细胞特异性表达的Mfge 8或Mfge 8缺陷型小鼠。目标2将审查 Mfge 8和αvβ3和αvβ5整联蛋白调节脂滴中TG水解的机制 使用整合素和Mfge 8缺陷小鼠的体内研究和分化的Caco-2细胞的体外研究。 研究将确定Mfge 8-整联蛋白轴如何调节亚细胞定位和磷酸化， 脂解介质。目的3：检测口服αvβ3和αvβ5整合素抑制剂对胃癌的治疗效果。 在肥胖小鼠中诱导体重减轻，并开发具有优化效力的新化合物， 选择性阻断该途径。申请人认为，拟议的研究具有创新性，因为它 确定了一种机制，将摄入的膳食脂肪与细胞质脂质中TG的水解联系起来 用于乳糜微粒产生的液滴。这项研究意义重大，因为它将扩大 了解肠道如何吸收和加工膳食脂肪以供输出。这种知识具有 潜在的信息开发的治疗，可以减少肥胖和餐后脂血。

SUMMARY Intestinal absorption of dietary fats is important for the development of obesity and its complications. The long- term goal is to understand the role of the integrin family of cell surface matrix receptors in regulating intestinal nutrient absorption. The overall objective of this application is to elucidate the role of the αvβ3 and αvβ5 integrins in regulating small intestinal lipid homeostasis. The central hypothesis is that in response to ingestion of dietary fat, enterocytes secrete the integrin ligand Mfge8. Mfge8 then binds enterocyte αvβ3 and αvβ5 integrins activating a signaling pathway that increases the absorption of dietary fat coupled with synthesis of triglyceride (TG) in the endoplasmic reticulum and hydrolysis of TG in cytoplasmic lipid droplets. This hypothesis is based on data demonstrating that mice deficient in Mfge8 or the αvβ3 and αvβ5 integrins develop steatorrhea and in response to a dietary fat challenge absorb less fat, accumulate excess cytoplasmic lipid droplets and secrete less chylomicrons. The rationale for these studies is that delineating the role of the Mfge8-integrin axis in intestinal lipid absorption is necessary to understand the mechanisms by which enterocytes regulate lipoprotein production. This hypothesis will be tested through 3 specific aims: 1) Determining how nutrients induce enterocyte production of Mfge8 transcript and protein and the physiological role of the Mfge8 and the αvβ3 and αvβ5 integrins in absorption of dietary fat; 2) Determining the role of the αvβ3 and αvβ5 integrins in regulating hydrolysis of TG in enterocyte cytoplasmic lipid droplets; and 3) Determining the therapeutic potential of targeting the αvβ3 and αvβ5 integrins for the treatment of obesity. Aim 1 will examine transcriptional regulation of Mfge8 in differentiated Caco-2 cells and primary enterocytes, secretion of Mfge8 into the intestinal lumen in response to different nutrients, and identification of the cellular source of Mfge8 required for fat absorption through use of transgenic mice with enterocyte specific deletion of Mfge8 or Mfge8 deficient mice with enterocyte specific expression of Mfge8. Aim 2 will examine the mechanisms by which Mfge8 and the αvβ3 and αvβ5 integrins regulate hydrolysis of TG from lipid droplets using in vivo studies with integrin and Mfge8 deficient mice and in vitro studies with differentiated Caco-2 cells. Studies will determine how the Mfge8-integrin axis regulates subcellular localization and phosphorylation of lipolytic mediators. Aim 3 will test the therapeutic efficacy of oral inhibition of the αvβ3 and αvβ5 integrins in inducing weight loss in obese mice coupled with development of novel compounds with optimized potency and selectivity for blocking this pathway. The proposed research is innovative, in the applicant's opinion, because it identifies a mechanism for linking uptake of ingested dietary fats with hydrolysis of TG from cytoplasmic lipid droplets for chylomicron production. The proposed research is significant because it will expand the understanding of how the intestine absorbs and processes dietary fat for export. This knowledge has the potential to inform the development of therapeutics that can reduce obesity and postprandial lipemia.