The role of MRP1 in Protection of Cardiac Injury

MRP1 在保护心脏损伤中的作用

• 批准号: 8115153
• 负责人: Mary Vore
• 金额: $ 26.19万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-25 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8115153
• 关键词: 4 hydroxynonenal; ABCC1 gene; ATP-Binding Cassette Transporters; Adriamycin PFS; Alkylation; Antibodies; Biochemical; Biological Assay; Cardiac; Cardiac Myocytes; Cardiotoxicity; Cell Line; Cell membrane; Cells; Collaborations; Coupled; Cysteine; Development; Dose; Esters; Fractionation; Genes; Genotype; Glutathione; Glutathione Disulfide; Goals; Heart; Histidine; Human; Immunohistochemistry; In Vitro; Injury; Knockout Mice; Lead; Lysine; Measures; Mediating; Messenger RNA; Mitochondria; Modality; Morphology; Mus; Normal tissue morphology; Oxidation-Reduction; Oxidative Stress; P-Glycoprotein; P-Glycoproteins; Production; Protein Isoforms; Proteins; Proteomics; Quantitative Microscopy; Regulation; Role; Sarcolemma; Schiff Bases; Site; Testing; Therapeutic; Time; Tissues; Toxic effect; Transgenic Mice; Transgenic Organisms; Tumor Necrosis Factor-alpha; Vesicle; Wild Type Mouse; adduct; base; buthionine; cancer therapy; chemotherapeutic agent; cytotoxic; functional group; in vivo; novel; overexpression; oxidant stress; oxidative damage; response; treatment effect; uptake

SPACE PROVIDED. ' The goal of the present application is to characterize the role of multidrug resistance protein 1 (Mrp1) in protecting the heart from oxidative stress. We postulate that cancer treatment with Adriamycin (ADR)leads to oxidative stress, which in turn leads to production of tumor necrosis factor-a (TNF) that amplifies oxidative stress and causes normal tissue injury. Mrp1 is an ATP-binding cassette (ABC) transporter that mediates the ATP-dependent efflux of glutathione (GSH) and its conjugates, including the GSH conjugate of the cytotoxic product of oxidative stress, 4-hydroxynonenal (HNE; GS-HNE). We will test the hypotheses that 1) Cardiac expression of Mrp1 protects the heart from oxidative stress and injury induced by ADR by mediating efflux of GS-HNE; 2) Mrp1 expression increases and is localized in plasma membrane and mitochondria in response to oxidative stress and/or TNF, and 3) excessive production of HNE and GS-HNE inactivates Mrp1, overwhelming its protective role, and exacerbating oxidative injury. Four Specific Aims will test these hypotheses; Aim 1will utilize Mrp1 null mice to assess its role in protecting the heart from ADR-induced oxidative stress and injury, the ability of MnSODto compensate for loss of Mrp1, and the function of TNF in regulating Mrp1 expression. Aim 2 will characterize the subcellular localization and function of Mrp1 following ADR and TNF treatment. Aim 3 will assessthe role of oxidative stress in the regulation of Mrp1 expression and localization. Finally, Aim 4 will characterize the ability of HNEand GS-HNE to inactivate Mrp1 by alkylation of key cysteine, histidine or lysine residues. We will utilize mice of various genotypes (Mrp1 null mice, MnSOD transgenic and heterozygous (+/-) mice) to assessthe roles of these genes in protection against cardiac injury, confocal immunofluorescent immunohistochemistry and quantitative immunogold analysis for localization of Mrp1 expression in the cardiomyocyte, and HEK293 cells for expression of Mrp1 to characterize its function; proteomic analyses will be used to identify potential structural isoforms of Mrp1 and the sites of HNE alkylation of Mrp1. Understanding the roles of Mrp1, TNF and GSH in protecting the heart from ADR-induced tissue injury will lead to the development of ancillary therapeutic modalities to protect against such injury, and thus permit utilization of higher doses of this highly effective chemotherapeutic agent.

空间 假如。 ' 本应用的目的是表征多药抗性蛋白1（MRP1）在 保护心脏免受氧化应激。我们假设用adrimycin（ADR）铅癌治疗 氧化应激又导致产生肿瘤坏死因子A（TNF），从而扩增了氧化性氧化。 压力并导致正常的组织损伤。 MRP1是ATP结合盒（ABC）转运蛋白，可介导 谷胱甘肽（GSH）及其偶联物的ATP依赖性外排，包括细胞毒性的GSH共轭物 氧化应激的产物，4-羟基烯烯（HNE; GS-HNE）。我们将测试假设1）心脏 MRP1的表达可保护心脏免受ADR诱导的氧化应激和损伤的影响 gs-hne; 2）MRP1表达增加并定位在质膜和线粒体中 氧化应激和/或TNF，以及3）HNE和GS-HNE的过量产生MRP1，MRP1， 压倒了其保护作用，并加剧了氧化损伤。四个具体目标将测试这些 假设； AIM 1WILL利用MRP1 NULL小鼠评估其在保护心脏免受ADR诱导的角色中的作用 氧化应激和损伤，MNSODTO补偿MRP1的损失的能力以及TNF的功能 调节MRP1表达。 AIM 2将表征MRP1之后MRP1的亚细胞定位和功能 ADR和TNF处理。 AIM 3将评估氧化应激在MRP1表达调节中的作用 和本地化。最后，AIM 4将表征Hneand GS-HNE灭活MRP1的能力 关键半胱氨酸，组氨酸或赖氨酸残基的烷基化。我们将利用各种基因型的小鼠（MRP1 NULL 小鼠，mnsod转基因和杂合（+/-）小鼠评估这些基因在保护中的作用 针对心脏损伤，共聚焦免疫荧光免疫组织化学和定量免疫金 分析MRP1​​表达在心肌细胞和HEK293细胞中的定位用于MRP1的表达 表征其功能；蛋白质组学分析将用于鉴定MRP1的潜在结构同工型 以及MRP1 HNE烷基化的位置。了解MRP1，TNF和GSH在保护它中的作用 ADR引起的组织损伤的心脏将导致辅助治疗方式发展为 防止这种伤害，从而允许使用较高剂量的高剂量 化学治疗剂。