integrin-mediated regulation of enterocyte lipid homeostasis

摘要 肠道对膳食脂肪的吸收对肥胖及其并发症的发生具有重要意义。长的- 学期目标是了解细胞表面基质受体的整合素家族在调节肠道中的作用。 营养吸收。本申请的总体目标是阐明αvβ3和αvβ5的作用 整合素在调节小肠脂稳态中的作用。中心假设是，对摄取的反应 在饮食脂肪中，肠细胞分泌整合素配体Mfge8。Mfge8然后与肠细胞αvβ3和αvβ5结合 整合素激活一条信号通路，增加膳食脂肪的吸收与合成 内质网中的甘油三酯(TG)和细胞质脂滴中的TG的水解性。这 该假说基于的数据表明，缺乏Mfge8或αvβ3和αvβ5整合素的小鼠 出现脂肪泻，为了应对饮食脂肪的挑战，减少脂肪的吸收，积累过多的细胞质 脂滴和分泌较少的乳糜粒。这些研究的基本原理是，勾勒出 Mfge8-整合素轴在肠道脂质吸收中的作用是理解其机制所必需的。 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) 确定营养物质如何诱导肠细胞产生Mfge8转录本和蛋白以及生理学 Mfge8和αvβ3和αvβ5整合素在膳食脂肪吸收中的作用；2)确定 αvβ3和αvβ5整合素在调节肠细胞胞浆脂滴甘油三酯水解中的作用； 确定靶向αvβ3和αvβ5整合素治疗肥胖的治疗潜力。目标 1将研究Mfge8在分化的Caco-2细胞和原代肠道细胞中的转录调节， Mfge8分泌到肠腔对不同营养物质的响应，以及细胞的鉴定 利用肠道细胞特异性缺失的转基因小鼠获得脂肪吸收所需的Mfge8的来源 Mfge8或Mfge8缺陷小鼠具有肠细胞特异性表达Mfge8。Aim 2将研究 Mfge8和αvβ3和αvβ5整合素调节脂滴中甘油三酯水解的机制 使用整合素和Mfge8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Mfge8-整合素轴如何调节亚细胞定位和磷酸化 脂解中介物。目的3检测口服抑制αvβ3和αvβ5整合素的治疗效果。 诱导肥胖小鼠体重减轻，并开发具有优化效力和 选择性地阻断这条途径。申请人认为，拟议的研究是创新的，因为它 确定摄取膳食脂肪与细胞质脂中甘油三酯的降解有关的机制 用于乳胶管生产的液滴。拟议的研究具有重要意义，因为它将扩大 了解肠道如何吸收和处理供出口的膳食脂肪。这一知识具有 有可能为开发可减少肥胖和餐后血脂的疗法提供信息。

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.