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Copper Transport in Lactation

哺乳期的铜转运

基本信息

批准号：
7846308
负责人：
MARIA C LINDER
金额：
$ 1.51万
依托单位：
CALIFORNIA STATE UNIVERSITY FULLERTON
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-06-01 至 2010-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7846308
关键词：
ATP phosphohydrolase ATP7A protein Address Affect Affinity Animal Model Animals Antibodies Apical Appearance Back Binding Binding Sites Blood Blood Cells Breast Cell Culture Techniques Cell membrane Cell model Cell surface Cells Ceruloplasmin Complex Confocal Microscopy Copper Diet Drug or chemical Tissue Distribution Endocytosis Enterocytes Epithelial Epithelial Cells Epithelium Exocytosis Gastrointestinal tract structure Goals Hepatocyte Hepatolenticular Degeneration Hereditary Disease Heterozygote Homeostasis Hormones Human Human Milk Immunoassay Immunofluorescence Immunologic Immunoprecipitation Infant Intestines Kidney Knock-out Knowledge Laboratories Lactation Ligands Liquid substance Liver Location Macroglobulins Mammary gland Measures Mediating Membrane Membrane Transport Proteins Messenger RNA Metabolism Milk Modeling Molecular Chaperones Mothers Mus Mutant Strains Mice Mutation Pattern Peptide Hydrolases Physiological Plasma Pregnancy Principal Investigator Process Production Proteins Pump Radioactive Radioactive Tracers Rattus Relative (related person)Rest Rodent Rodent Model Role Route SLC11A2 gene Side Small Interfering RNA Testing Time Tissues Transfection Transgenic Mice Translations Vesicle Wild Type Mouse Wilson disease protein cell type hypocupremia inhibitor/antagonist mammary epithelium monolayer mutant neonate offspring overexpression polarized cell prevent pup receptor receptor expression response trafficking trans-Golgi Network uptake

项目摘要

DESCRIPTION (provided by applicant): Many aspects of the mechanisms by which Cu is taken up from the blood plasma by different kinds of cells (including the transporters and delivery proteins involved), and just how (after cell entry) Cu finds its way to specific secretions, remain to be defined. We have obtained evidence that lactation profoundly alters the tissue distribution of newly absorbed Cu, diverting most of it from the liver and kidney to the mammary gland, where it rapidly appears in the milk and in milk ceruloplasmin. Cu in milk ceruloplasmin may be more bio-available to the infant than the other forms of Cu in the milk, which have not been well defined. A polarized cell culture model responsive to lactational hormones has been developed for studying milk production by mammary epithelial cells. Mutant rodent models lacking specific proteins that could be involved in the steps required for uptake of Cu by mammary gland and its delivery to developing milk are available. The long term objective is to understand how Cu distribution and transport are regulated under normal conditions and in gestation and lactation. The immediate objectives are to determine how Cu from the exchangeable plasma pool enters mammary (and hepatic) cells, under normal conditions and in lactation, to explain the marked changes in distribution observed in the latter condition (what potential membrane transporters are involved, and how their expression and/or disposition is changed by lactation). The objectives are also to determine how, after cell entry, copper is routed to the milk and milk cemloplasmin; the role(s) of ATOX1, WND and/or MNK in this process and in regulating the copper content of the milk; and to further define the copper components of mammalian milk. Expression of potential transporters will be measured at the mRNA and protein levels, by Real Time PCR and immunoassays, respectively; and localization will be followed with immunofluorescence and confocal microscopy. Alterations in expression will be induced in the cell culture model with lactational hormones, transfection, and antisense oligos. Effects of natural and knockout mutations of specific transporters in whole animal models on copper distribution and transport will be determined. Cu uptake and its emergence in milk and milk ceruloplasmin will be followed with 67Cu/64Cu and immunoprecipitation. Effects on Cu uptake of specific antibodies against membrane transporters will be measured. Cu binding components in milk secretions will be characterized. It is expected that the proposed studies will greatly advance our knowledge of how Cu is distributed to cells from the blood and particularly how mammary epithelial cells put Cu into components of the milk that may have specific effects on the infant.

描述（由申请人提供）：不同种类的细胞（包括所涉及的转运蛋白和递送蛋白）从血浆中摄取Cu的机制的许多方面，以及Cu（进入细胞后）如何进入特定分泌物，仍有待确定。我们已经获得的证据表明，哺乳期深刻地改变了新吸收的铜的组织分布，将大部分从肝脏和肾脏转移到乳腺，在那里它迅速出现在牛奶和牛奶铜蓝蛋白。牛奶铜蓝蛋白中的铜可能比牛奶中其他形式的铜更能被婴儿生物利用，而这些形式还没有很好的定义。一种对泌乳激素敏感的极化细胞培养模型已被开发用于研究乳腺上皮细胞的产奶量。目前已有缺乏特定蛋白质的突变啮齿动物模型，这些蛋白质可能参与乳腺吸收铜并将其输送到发育中的乳汁所需的步骤。长期目标是了解铜的分布和运输是如何在正常条件下，在妊娠和哺乳期调节。直接的目标是确定如何铜从可交换血浆池进入乳腺（和肝）细胞，在正常条件下，在哺乳期，以解释显着的变化，在后一种条件下观察到的分布（涉及哪些潜在的膜转运蛋白，以及它们的表达和/或处置是如何改变哺乳期）。目的还在于确定在细胞进入后，铜如何被路由至乳汁和乳汁cemloplasmin; ATOX 1、WND和/或MNK在该过程中以及在调节乳汁铜含量中的作用;以及进一步确定哺乳动物乳汁的铜组分。将分别通过真实的时间PCR和免疫测定法在mRNA和蛋白质水平上测量潜在转运蛋白的表达;并将使用免疫荧光和共聚焦显微镜进行定位。在细胞培养模型中，用哺乳激素、转染和反义寡核苷酸诱导表达的改变。将确定整个动物模型中特定转运蛋白的天然和敲除突变对铜分布和转运的影响。铜的吸收和它的出现在牛奶和牛奶铜蓝蛋白将遵循与67 Cu/64 Cu和免疫沉淀。将测量针对膜转运蛋白的特异性抗体对Cu摄取的影响。乳汁分泌物中的铜结合组分将被表征。预计拟议的研究将大大推进我们对铜如何从血液中分布到细胞的知识，特别是乳腺上皮细胞如何将铜放入可能对婴儿产生特定影响的牛奶成分中。