Heme trafficking and its impact on systemic iron homeostasis

描述(由申请者提供)：本项目的目标是研究血红素贩运如何影响全身铁稳态。由于每天需要25毫克的铁来维持红细胞的产生，每天只有1-2毫克的铁从胃肠道吸收，1-2毫克的铁每天通过萎靡不振的胃肠道、皮肤和肾脏粘膜细胞流失，因此大多数铁是循环利用的。传统的观点认为，当巨噬细胞吞噬衰老的红细胞时，它们的血红蛋白被降解为血红素，然后铁通过铁蛋白输出到转铁蛋白，然后输送到肝脏(用于储存)或骨髓(用于产生新的红细胞)。在以前的研究中，我们确定猫白血病病毒-C受体FLVCR特异性地从细胞中输出血红素(Cell 118：757-66,2004)。根据Western分析，FLVCR在十二指肠、肝脏和巨噬细胞中高表达，这些组织对铁的吸收和运输至关重要；Flvcr基因缺失的小鼠除了红细胞再生障碍性贫血外，还在这些部位存在过量的铁(Science 319：825-828,2008)。这些观察，加上对细胞系和体外原代巨噬细胞的研究，使我们假设血红素不仅是铁，而且是被贩卖的。这笔赠款将通过突变分析、非洲爪哇卵母细胞研究和电子结晶学，定义FLVCR的结构，并识别它将血红素输出到血红素结合蛋白(如血凝素和白蛋白)的机制。我们将确定铁如何调节FLVCR的细胞间定位，以帮助全身铁平衡。此外，我们将通过研究Flvcrflx/FLOX的巨噬细胞来确定红细胞中的血红素的命运；通过研究Flvcrflx/Flox的巨噬细胞在体外和体内的实验研究LysM-cre小鼠；通过研究Flvcrflx/Flox来确定一旦血红素进入肝脏后的命运；通过研究Alb-cre小鼠；确定肝细胞是否可以通过FLVCR将血红素分泌到胆汁中并允许铁排出体外；并探索这些发现在溶血、慢性炎症和血色沉着模型中的意义。总而言之，这些研究应该证明，铁的生理调节比之前所认识的更加复杂和复杂。

DESCRIPTION (provided by applicant): The goal of this project is to study how heme trafficking might impact systemic iron homeostasis. As 25 mg of iron is required daily to maintain red cell production, only 1-2 mg/d is absorbed from the GI tract, and 1-2 mg/d is lost through sloughed gastrointestinal, skin and kidney mucosal cells, most iron is recycled. The traditional view is that when macrophages engulf senescent red cells, their hemoglobin is degraded to heme then iron, which is exported via ferroportin to transferrin for delivery to the liver (for storage) or to the marrow (for new red cell production). In previous studies, we determined that the feline leukemia virus-C receptor, FLVCR, specifically exports heme from cells (Cell 118:757-66, 2004). On western analysis, FLVCR is highly expressed in the duodenum, liver and macrophage, tissues critical for iron absorption and trafficking; and Flvcr-deleted mice have excess iron at these sites, in addition to red cell aplasia (Science 319:825-828, 2008). These observations, plus studies of cell lines and primary macrophages in vitro, led us to hypothesize that heme, not only iron, is trafficked. This grant will define FLVCR structure and discern the mechanism by which it exports heme to heme-binding proteins, such as hemopexin and albumin, using mutagenesis analyses, Xenopus oocytes studies, and electron crystallography. We will determine how iron regulates the intercellular localization of FLVCR to aid systemic iron balance. In addition, we will determine the fate of heme from red cells after the cells are ingested by macrophages by studying macrophages from Flvcrflox/flox;LysM-cre mice in vitro and in vivo; determine the fate of heme once delivered to liver by studying Flvcrflox/flox;alb-cre mice; determine if hepatocytes can secrete heme via FLVCR into the bile permitting iron to exit the body; and explore the implications of these findings in models of hemolysis, chronic inflammation, and hemochromatosis. Together these studies should demonstrate that the physiologic regulation of iron is more intricate and complex than previously appreciated.