Blood-brain barrier regulates leptin transport in obesity

血脑屏障调节肥胖中的瘦素转运

• 批准号: 7468501
• 负责人: ABBA J KASTIN
• 金额: $ 34.15万
• 依托单位: LSU PENNINGTON BIOMEDICAL RESEARCH CTR
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7468501
• 关键词: Affect; Antibodies; Appetite Depressants; Binding; Biological Assay; Blocking Antibodies; Blood; Blood - brain barrier anatomy; Blood Circulation; Brain; Cell Fractionation; Cells; Co-Immunoprecipitations; Condition; Eating; Endocytic Vesicle; Endocytosis; Endothelial Cells; Energy Transfer; Food; Food deprivation (experimental); Genetic Transcription; Heterodimerization; Immunoprecipitation; Leptin; Ligand Binding; Localized; Measures; Mediating; Microscopic; Microscopy; Mus; Obesity; Peptides; Perception; Phosphorylation; Physiological; Play; Positioning Attribute; Process; Protein Kinase; Proteins; Recruitment Activity; Regulation; Research Personnel; Role; Satiation; Signal Transduction; Signal Transduction Pathway; System; TNF gene; Testing; Therapeutic Uses; Time; Transfection; Tumor Necrosis Factor Receptor; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; Western Blotting; antisauvagine 30; cytokine; feeding; genetic regulatory protein; human MPP1 protein; human TNF protein; leptin receptor; mRNA Expression; novel; polypeptide; programs; protein protein interaction; radioligand; receptor; receptor mediated endocytosis; transcriptional coactivator p75; uptake; urocortin

DESCRIPTION (provided by applicant): The interactions of leptin with the blood-brain barrier (BBB) play an important role in some forms of obesity. In this proposal, we will elucidate the novel aspects of how leptin regulates the permeation of some feeding-related peptides and cytokines across the BBB. (a) to test the hypothesis that leptin recruits receptors of selective satiety peptides and activates transport, we will examine how leptin increases the blood-to-brain permeation of urocortin, another potent satiety peptide. We will first determine whether receptors for both leptin and urocortin are involved in this process. We will then determine whether there is heterodimerization of these receptors by use of co-immunoprecipitation and ligand binding assays, after the receptors are expressed by co-transfection on mouse TM-BBB4 brain endothelial cells. Further, we will determine by immunofluorescent microscopy whether there is co-localization of these receptors on endocytotic vesicles after binding to leptin. (b) To test the hypothesis that leptin can enhance an existing transport system for a cytokine that also reduces feeding, we will examine how leptin upregulates the transport of tumor necrosis factor alpha (TNFalpha). We predict that leptin will crease TNFalpha influx by modulating phosphorylation of the p55-receptors, p75-receptors, and proteins related to TNFalpha transport, as shown by transport assays, immunoprecipitation, and Western blot. (c) To test the physiological relevance of leptin-mediated regulation of TNFalpha and urocortin transport, we will measure the transport of TNFalpha, urocortin, as well as leptin itself in mice that are fed normally and in mice that are food-deprived. We predict that leptin will increase urocortin and TNFalpha transport only in mice with free access to food, thereby providing additional satiety signals to the brain when leptin is already relatively high in the circulation. In contrast, we predict that in food-deprived mice, the transport systems for both leptin and TNFalpha will be down-regulated and that for urocortin will not be activated, as expected for anorectic agents under the severe condition in which no food is available. By completing the proposed studies, we will demonstrate the presence of novel protein-protein interactions at the BBB, the mechanisms involved in these interactions, and the additional functional role of the BBB in regulating feeding. This information will not only add to a better understanding of the mechanisms of BBB transport in general but also to the potential therapeutic use of ingestive peptides.

描述(由申请人提供)： 瘦素与血脑屏障(BBB)的相互作用在某些形式的肥胖中起着重要作用。在这项提案中，我们将阐明瘦素如何调节一些与摄食相关的多肽和细胞因子通过血脑屏障的新方面。(A)为了验证瘦素招募选择性饱腹肽的受体并激活转运的假设，我们将研究瘦素如何增加另一种有效饱腹肽--尿皮质素--的血到脑的渗透。我们将首先确定瘦素和尿皮质素的受体是否都参与了这一过程。然后，我们将通过免疫共沉淀和配体结合分析来确定这些受体是否存在异二聚化，然后将这些受体通过共转染小鼠TM-BBB4脑内皮细胞来表达。此外，我们将通过免疫荧光显微镜来确定这些受体在与瘦素结合后是否在内吞囊泡上共定位。(B)为了验证瘦素可以增强现有细胞因子运输系统的假设，同时减少摄食，我们将研究瘦素如何上调肿瘤坏死因子α(TNFpha)的运输。我们预测，瘦素将通过调节p55受体、p75受体和与TNFpha转运相关的蛋白的磷酸化来增加TNFpha的内流，如运输分析、免疫沉淀和Western印迹所示。(C)为了测试瘦素介导的TNFpha和urocortin转运的生理学相关性，我们将测量正常喂养和缺乏食物的小鼠的TNFpha、urocortin以及瘦素本身的转运。我们预测，只有在自由进食的小鼠中，瘦素才会增加尿皮质素和肿瘤坏死因子α的转运，从而在瘦素在循环中已经相对较高的情况下向大脑提供额外的饱腹感信号。相反，我们预测，在食物匮乏的小鼠中，瘦素和肿瘤坏死因子α的运输系统都将下调，而尿皮质素的运输系统将不会被激活，这与厌食剂在没有食物的严重条件下所预期的一样。通过完成拟议的研究，我们将证明在血脑屏障存在新的蛋白质-蛋白质相互作用，这些相互作用涉及的机制，以及血脑屏障在调节摄食方面的额外功能作用。这些信息不仅将有助于更好地了解血脑屏障转运的一般机制，而且还将有助于消化肽的潜在治疗用途。