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）：由于不存在活性排泄系统，因此十二指肠对膳食铁的吸收决定了全身铁水平。因此，必须严格控制这一过程，以避免组织铁积累（例如遗传性血色病）或缺乏（例如慢性炎症贫血）的不良后果。在过去的十年里，我们一直在研究铁在肠上皮细胞（IEC）中转运的分子方面，长期目标是开发药物或饮食治疗来调节人类的铁吸收。虽然已经查明了铁的进口商和出口商，但对铁在国际电工委员会之间的流动和出口进入流通（这是限速步骤）的具体细节了解甚少。我们提出了新的观察，铜相关的过程被激活的铁剥夺的啮齿动物。在肠上皮细胞中，铜转运ATP酶（Atp 7a）和铜结合蛋白（金属硫蛋白）在细胞内铜水平增加的情况下上调。这些观察结果提供了对体内铜水平（在缺铁期间增加和铁过载减少）与肠道铁转运控制之间关系的机械见解。基于这些研究结果，鉴定参与控制铁通量的铜特异性机制是必要的。从肠上皮细胞输出的亚铁（Fe 2+）在功能上与铁（Fe 3+）结合至间质液中的转铁蛋白（Tf）所需的氧化步骤偶联。膜结合的多铜铁氧化酶（FOX），Hephaestin（Heph），可以介导这一步骤。然而，Heph KO小鼠是可行的，肠铁转运仅部分减弱，表明存在其他FOX。我们最近发现，肠细胞有两个不同的新的环索铁氧化酶（FOX），一个是未发现的，可溶性形式的Heph（sHeph）和其他称为cytoFOX。我们假设胞浆FOX参与铁跨IEC的转胞吞作用。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%; cytoFOX ~60-65%）。另一种是循环FOX，血浆铜蓝蛋白（Cp），也可能参与IEC的铁输出。目前的建议将阐明具体的，机制的细节，肠道铁转运的中心假设，即新的多铜FOX（sHeph和cytoFOX）在肠上皮细胞和Cp在血液中发挥不可或缺的作用，在控制铁输出IEC。我们还将定义这些蛋白质的生物合成的Cu递送机制，可能涉及肠Atp 7a。本申请中概述的整合方法，使用哺乳动物铁转运的独特体内和体外模型， 将首先破译（在目的1中）Atp 7a在将铜递送至sHeph以介导铁流出和增强肝铜负荷中的作用。目标2将定义新型可溶性FOX在转胞吞铁途径中的作用，而目标3将阐明Cp在铁吸收中的作用，并确定缺铁期间增强全Cp产生的机制。总的来说，该项目将通过揭示铁转运的新机制细节来推进铁生物学领域，并可能为开发调节铁吸收的分子方法提供机会。