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: 7636746
• 负责人: James F. Collins
• 金额: $ 26.85万
• 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 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7636746
• 关键词: ATP phosphohydrolase; Address; Age; Anemia; Anemia due to Chronic Disorder; Animal Model; Apical; Cell Culture Techniques; Cells; Ceruloplasmin; Copper; Data; Development; Dietary Copper; Dietary Iron; Disease; Duodenum; Enterocytes; Epithelial; Epithelial Cells; Epithelium; Genes; Genetic; Goals; Hemochromatosis; Homeostasis; Human; Human Pathology; In Transferrin; In Vitro; Intestinal Absorption; Intestines; Investigation; Ion Transport; Ions; Iron; Knockout Mice; Learning; Link; Literature; Liver; Malnutrition; Mediating; Membrane; Menkes Kinky Hair Syndrome; Metals; Modeling; Molecular; Mucous Membrane; Mutation; Patients; Physiological; Physiological Processes; Play; Post-Transcriptional Regulation; Process; Proteins; Rattus; Regulation; Reporting; Rodent Model; Role; SLC11A2 gene; Serum; Small Interfering RNA; Small Intestines; Staging; Suggestion; Techniques; Testing; Trace Elements; Transcriptional Regulation; Vesicle; absorption; apical membrane; base; basolateral membrane; brush border membrane; deprivation; design; divalent metal; hypocupremia; in vitro Model; in vivo; in vivo Model; intestinal epithelium; jejunum; metal transporting protein 1; novel; postnatal; response

DESCRIPTION (provided by applicant): The overall control of iron homeostasis occurs at the transport step in the epithelium of the proximal small bowel, where absorption is precisely regulated to match body iron losses. Importantly, perturbations in intestinal iron transport are associated with several important disease states in humans, including anemia of chronic disease and hemochromatosis. Intestinal copper transport is enhanced in rats during iron-deficiency, and this is likely a physiological response related to the role of dietary copper in various aspects of overall body iron homeostasis. Interestingly, the Menkes copper ATPase (Atpya) is strongly induced in the duodenal mucosa of iron-deprived rats at different postnatal ages along with Divalent Metal Transporter i (Dmti), which can transport iron and copper. Thus, the overall goals of this proposal are i) to determine the roles that Dmti and Atpya play in the induction of copper transport during iron-deprivation, 2) to decipher the molecular mechanisms of induction of Dmti and Atpya during iron-deprivation and 3) to determine the effect that copper has on molecular mechanisms of trans-epithelial iron transport in the intestine. This will be accomplished by utilizing cell culture and rodent models of intestinal iron transport. Specific AIM i will test the hypothesis that induction of Dmti and Atpya is responsible for increased transepithelial copper transport seen during iron-deprivation. Iron and copper transport studies and siRNA knockdowns will be performed in our in vitro model of the intestinal epithelium, the IEC-6 cells. Once the transporter(s) involved in the induction of copper transport during iron-deficiency have been identified, we will perform complementary studies in in vivo models of iron-deficiency, including wild-type, iron-deficient rats, Belgrade (i.e. Dmti-deficient) rats and Atpya knockout mice. Specific AIM 2, will test the hypothesis that Dmti and Atpya are regulated by distinct molecular mechanisms during iron-deficiency. Iron-dependent, post- transcriptional regulation of Dmti will be examined (mediated by the 3' IRE) and transcriptional regulation of Atpya in IEC-6 cells will be studied. Finally, specific AIM 3 will test the hypothesis that increased copper transport during iron deficiency functions to enhance copper-dependent aspects of intestinal iron absorption. To accomplish this goal, iron transport studies will be performed in membrane vesicles isolated from iron-deficient rats deprived of dietary copper, and hephaestin activity in enterocytes and ceruloplasmin activity in serum will be determined. Overall, these studies will allow further definition of the copper- dependent processes that are involved in enhancing intestinal iron absorption during states of iron- deficiency. A detailed understanding of these relationships is critical, as intestinal iron transport controls overall body iron homeostasis. Moreover, this proposed investigation is novel, as studies addressing the impact of increased enterocyte and liver copper levels during iron-deficiency have not been reported to date.

描述（由申请人提供）：铁稳态的总体控制发生在近端小肠上皮的运输步骤中，其中吸收被精确调节以匹配体内铁损失。重要的是，肠道铁转运的扰动与人类的几种重要疾病状态有关，包括慢性病贫血和血色素沉着症。缺铁期间，大鼠的肠道铜转运增强，这可能是与膳食铜在全身铁稳态各个方面的作用相关的生理反应。有趣的是，门克斯铜 ATP 酶 (Atpya) 在不同出生后年龄的缺铁大鼠的十二指肠粘膜中与可转运铁和铜的二价金属转运蛋白 i (Dmti) 一起被强烈诱导。因此，该提案的总体目标是 i) 确定 Dmti 和 Atpya 在缺铁期间诱导铜转运中发挥的作用，2) 破译缺铁期间诱导 Dmti 和 Atpya 的分子机制，3) 确定铜对肠道跨上皮铁转运分子机制的影响。这将通过利用肠道铁转运的细胞培养和啮齿动物模型来实现。具体的 AIM i 将检验以下假设：Dmti 和 Atpya 的诱导导致缺铁期间观察到的跨上皮铜转运增加。铁和铜转运研究和 siRNA 敲低将在我们的肠上皮细胞体外模型（IEC-6 细胞）中进行。一旦确定了缺铁期间诱导铜转运的转运蛋白，我们将在缺铁体内模型中进行补充研究，包括野生型缺铁大鼠、贝尔格莱德（即 Dmti 缺乏）大鼠和 Atpya 基因敲除小鼠。具体的 AIM 2 将检验 Dmti 和 Atpya 在缺铁期间受到不同分子机制调节的假设。将检查 Dmti 的铁依赖性转录后调节（由 3' IRE 介导），并研究 IEC-6 细胞中 Atpya 的转录调节。最后，特定的 AIM 3 将检验以下假设：缺铁期间铜转运的增加可增强肠道铁吸收的铜依赖性。为了实现这一目标，将在从缺乏膳食铜的缺铁大鼠中分离的膜囊泡中进行铁转运研究，并测定肠细胞中的铁黄蛋白活性和血清中的铜蓝蛋白活性。总体而言，这些研究将进一步定义铜依赖性过程，这些过程涉及在缺铁状态下增强肠道铁吸收。详细了解这些关系至关重要，因为肠道铁转运控制着全身铁稳态。此外，这项拟议的研究是新颖的，因为迄今为止尚未报道铁缺乏期间肠细胞和肝铜水平增加的影响的研究。