Molecular Mechanisms of Intestinal Metal Ion Transport During Iron Deficiency

缺铁期间肠道金属离子转运的分子机制

• 批准号: 8813554
• 负责人: James F. Collins
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2016-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8813554
• 关键词: ATP phosphohydrolase; Anabolism; Anemia; Attenuated; Binding; Biology; Blood; Blood Circulation; Cells; Ceruloplasmin; Chronic; Complement; Copper; Coupled; Diet; Dietary Iron; Endosomes; Enterocytes; Enzymes; Epithelial Cells; Goals; Health; HepG2; Hepatic; Hepatocyte; Hereditary hemochromatosis; Human; Human Pathology; In Transferrin; Inflammation; Intercellular Fluid; Intestines; Ion Transport; Iron; Iron Overload; Knowledge; Liver; Location; Mediating; Membrane; Metallothionein; Metals; Molecular; Movement; Mus; Mutation; Nature; Pathway interactions; Pharmaceutical Preparations; Physiological; Play; Process; Production; Protein Biosynthesis; Proteins; Rattus; Regulation; Rodent; Role; System; Testing; Tissues; Vesicle; absorption; adverse outcome; base; copper-binding protein; copper-transporting ATPase; deprivation; in vitro Model; in vivo; insight; iron deficiency; metal transporting protein 1; mutant; novel; oxidation; research study; transcytosis

DESCRIPTION (provided by applicant): Absorption of dietary iron in the doudenum determines overall body iron levels as no active excretory systems exist. As such, this process must be tightly controlled to avoid the adverse consequences of tissue iron accumulation (e.g. in hereditary hemochromatosis) or deficiency (e.g. in anemia of chronic inflammation). We have been investigating molecular aspects of iron transport across intestinal epithelial cells (IECs) fo the past decade, with a long-term goal of developing drugs or dietary treatments to modulate iron absorption in humans. Although iron importers and exporters have been identified, a paucity of knowledge exists regarding the specific details of iron movement across IECs and export into the circulation (which is the rate-limiting step). We made the novel observation that copper-related processes are activated by iron deprivation of rodents. In enterocytes, a copper transporting ATPase (Atp7a) and a copper-binding protein (metallothionein) were upregulated in the setting of increased intracellular copper levels. These observations provided mechanistic insight into the relationship between body copper levels (which increase during iron deficiency & decrease in iron overload) and control of intestinal iron transport. Based upon these findings, identification of copper-specific mechanisms involved in control of iron flux was an imperative. Ferrous iron (Fe2+) export from enterocytes is functionally coupled to an oxidation step which is required for iron (Fe3+) binding to transferrin (Tf) in the interstitial fluid. A membrane-bound, multi-copper ferroxidase (FOX), Hephaestin (Heph), may mediate this step. However, Heph KO mice are viable and intestinal iron transport is only partially attenuated, suggesting that other FOXs exist. We recently discovered that enterocytes have two, distinct novel cyosolic ferroxidases (FOXs), one being an undiscovered, soluble form of Heph (sHeph) and the other termed cytoFOX. We postulate that cytosolic FOXs participate in transcytosis of iron across IECs. Studies in Heph KO mice demonstrated that both proteins contribute to cytosolic FOX activity (sHeph ~35-40%; cytoFOX ~60-65%). Another, circulating FOX, ceruloplasmin (Cp), may also participate in Fe export from IECs. The current proposal will elucidate specific, mechanistic details of intestinal Fe transport by testing the central hypothesis that novel multi-copper FOXs (sHeph and cytoFOX) in enterocytes and Cp in blood play integral roles in control of iron export from IECs. We will also define the mechanism(s) of Cu delivery for the biosynthesis of these proteins, likely involving intestinal Atp7a. The integrative approach outlined in this application, using unique in vivo and in vitro models of mammalian iron transport, will first decipher (in Aim 1) the role of Atp7a in delivering copper to sHeph to mediate iron efflx and in potentiating hepatic copper loading. Aim 2 will define the role(s) of novel soluble FOXs in the transcytotic iron pathway, while Aim 3 will clarify the role of Cp in iron absorption and will determine the mechanism of enhanced production of holo-Cp during iron deficiency. Overall, this project will advance the field of iron biology by revealing new mechanistic details of iron transport and may provide opportunities to develop molecular approaches to modulate iron absorption.

描述（由申请人提供）：饮食中的饮食铁的吸收决定了整体铁水平，因为不存在主动排泄系统。因此，必须严格控制此过程，以避免组织铁积累的不利后果（例如，在遗传性血色素沉着症中）或缺乏症（例如，在慢性炎症的贫血中）。在过去的十年中，我们一直在研究跨肠上皮细胞（IEC）的铁运输的分子方面，其长期目标是开发药物或饮食治疗以调节人类的铁吸收。尽管已经确定了铁的进口商和出口商，但是关于跨IEC的铁运动的具体细节并将其导出到循环中（这是限制速率的步骤）。我们做出了新的观察，即通过啮齿动物的铁剥夺与铜相关的过程被激活。在肠上皮细胞中，在升高的细胞内铜水平的情况下，将运输ATPase（ATP7A）的铜（ATP7A）和铜结合蛋白（金属硫硫蛋白）上调。这些观察结果提供了对体铜水平（铁缺乏症和铁超载减少的增加）和肠道铁转运的控制的机械洞察力。基于这些发现，必须鉴定参与铁通量控制的铜特异性机制。肠肠细胞的亚铁（Fe2+）导出在功能上耦合到氧化步骤，该步骤是在间质流体中与铁（Fe3+）结合与转铁蛋白（TF）所必需的。膜结合的多碳酸铁氧化酶（FOX），赫菲斯汀（Heph）可能会介导此步骤。但是，Heph KO小鼠是可行的，肠道铁的运输仅部分减弱，这表明存在其他狐狸。我们最近发现，肠细胞有两个独特的新型氰基铁氧化酶（狐狸），一种是未被发现的Heph（Sheph），另一种被称为Cytofox。我们假设胞质狐狸参与了IEC跨IEC的铁脑胞一次。在HEPH KO小鼠中的研究表明，两种蛋白质都有助于胞质FOX活性（Sheph〜35-40％；细胞毒素〜60-65％）。另一个循环的狐狸ceruloplasmin（CP）也可能参与IEC的FE出口。当前的提案将通过测试肠肠上皮细胞中新型多杆狐（Sheph和Cytofox）的中心假设来阐明肠道FE转运的特定机械细节，而在血液中CP中的CP在控制IEC中的铁中的积分作用中。我们还将定义这些蛋白质生物合成的Cu递送机制，可能涉及肠道ATP7A。在本应用程序中概述的综合方法，使用哺乳动物铁运输的独特体内和体外模型， 将首先破译（在AIM 1中）ATP7A在将铜运送到Sheph中的作用，以介导铁污水和增强肝铜负荷。 AIM 2将定义新型可溶性狐狸在跨环铁途径中的作用，而AIM 3将阐明CP在铁吸收中的作用，并将确定在铁缺乏症中增强Holo-CP产生的机制。总体而言，该项目将通过揭示铁运输的新机械细节来推进铁生物学领域，并可能提供开发分子方法调节铁吸收的机会。