Excessive Copper Levels Disrupt Hepatic Nuclear Receptor Function

过量的铜水平会破坏肝核受体功能

• 批准号: 8059216
• 负责人: Clavia Ruth Wooton-Kee
• 金额: $ 4.76万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-25 至 2012-09-24
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8059216
• 关键词: Excessive; Copper; Levels; Disrupt; Hepatic

DESCRIPTION (provided by applicant): Our long-term objective is to determine the effect of copper on the DNA-binding activity and structure of hepatic nuclear receptors and expression of hepatic nuclear receptor target genes in a mouse animal model of Wilson's disease. Wilson's disease is an autosomal recessive disease that results in excessive hepatic copper accumulation due to mutations in the Cu-transporting P-type ATPase, ATP7b, and is associated with a variety of symptoms including steatosis, cholestasis, cirrhosis, and liver failure, as well as neurological dysfunction. Wilson's disease is a chronic and severe liver disorder that is fatal if not treated. Chelation and/or zinc therapy started before the onset of severe liver dysfunction has been shown to manage the symptoms of Wilson's disease. The Wilson's disease animal models (Long Evans Cinnamon (LEC) rat and Atp7b-/- mouse) have decreased expression of genes involved in several metabolic pathways, including bile acid synthesis (cholesterol 71-hydroxylase (Cyp7a1)), cholesterol synthesis (3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA reductase)), biliary bile acid (bile salt export pump (Bsep, Abcb11) and glutathione conjugated- compound transport (multidrug resistance-associated protein 2 (Mrp2, Abcc2)). The expression of these genes is regulated by nuclear receptors, including the farnesoid X receptor (FXR) and liver X receptor (LXR). Our hypothesis is that excess copper inactivates nuclear receptors and disrupts signaling pathways critical for maintaining metabolic homeostasis in the liver. We will utilize the Atp7b-/- mouse to determine nuclear receptor DNA-binding activity via chromatin immunoprecipitation (ChIP) assays and to measure parameters of liver function, such as bile flow, biliary secretion of copper, bile acids, and bilirubin. We will test the following Specific Aims: (1) determine the effects of excess copper on ligand-mediated induction of target gene expression and nuclear receptor and coactivator recruitment to chromatin (2) determine the effect of copper on the structure of nuclear receptors (3) determine the effect of zinc and/or n-acetyl cysteine on nuclear receptor DNA-binding activity and liver function in wild-type and Atp7b-/- mice. This Specific Aim will address if zinc restores the DNA-binding activity of nuclear receptors and target gene mRNA expression and improves liver function. Additionally, n-acetyl cysteine and zinc will be administered to Atp7b-/- mice to determine if increasing hepatic glutathione levels enhance Mrp2-mediated biliary copper excretion. These studies will provide insight into the molecular mechanisms involved in liver dysfunction associated with Wilson's disease. Hence, our findings will directly advance the research of liver disease by elucidating molecular mechanisms involved in the progression of Wilson's disease and therapeutic interventions with available pharmacological agents, zinc and n-acetyl cysteine. PUBLIC HEALTH RELEVANCE: We will measure the DNA-binding activity of nuclear receptors in a mouse animal model for Wilson's disease, a chronic disease that is characterized by excessive hepatic copper accumulation and severe liver dysfunction that results in death if not treated. Our studies will determine molecular mechanisms that are disrupted by excessive copper levels and therapeutic regimens to restore nuclear receptor regulation of hepatic liver gene expression and hence improve liver function. This research specifically addresses the biological mechanisms relevant to human liver pathology and to Wilson's disease.

描述（由申请人提供）：我们的长期目标是确定铜对肝核受体的DNA结合活性和结构的影响，以及在威尔逊氏病小鼠动物模型中肝核受体靶基因的表达。威尔逊氏病是一种常染色体隐性疾病，由于Cu传播P型ATPase，ATP7B的突变导致过度肝铜的积累，并且与多种症状有关，包括静脉炎，胆汁淤积，胆汁淤积，Cirrhosis，cirrhosis和Fliver衰竭以及伴随神经系统疾病。威尔逊氏病是一种慢性且严重的肝脏疾病，即使没有治疗，这是致命的。在严重的肝功能障碍发作之前，螯合和/或锌疗法已开始证明可以控制威尔逊氏病的症状。威尔逊疾病动物模型（Long Evans肉桂（LEC）大鼠和ATP7B - / - 小鼠）的基因表达降低了几种代谢途径的基因表达，包括胆汁酸合成（胆固醇71-羟化酶（CYP7A1）），胆固醇合成（3-Hydroxy-3-甲基-glutartaryl-coa hydroxy-grutaryl-coAA， ），胆汁胆酸（胆汁盐导出泵（BSEP，ABCB11）和谷胱甘肽共轭 - 化合物转运（多抗耐药性相关蛋白2（MRP2，ABCC2，ABCC2））。这些基因的表达受核受体的调节，包括核受体，包括Farnes X受体（Farnes X受体）和coperor（fxr）和coperor（lx）。灭活核受体并破坏对肝脏中代谢稳态至关重要的信号通路。 We will test the following Specific Aims: (1) determine the effects of excess copper on ligand-mediated induction of target gene expression and nuclear receptor and coactivator recruitment to chromatin (2) determine the effect of copper on the structure of nuclear receptors (3) determine the effect of zinc and/or n-acetyl cysteine on nuclear receptor DNA-binding activity and liver function in wild-type and Atp7b-/- mice.如果锌恢复核受体的DNA结合活性并靶向基因mRNA表达并改善肝功能，则该特定目的将解决。另外，将N-乙酰基半胱氨酸和锌将用于ATP7B - / - 小鼠，以确定增加肝谷胱甘肽水平是否会增强MRP2介导的胆道铜排泄。这些研究将洞悉与威尔逊氏病有关的肝功能障碍所涉及的分子机制。因此，我们的发现将直接通过阐明参与威尔逊疾病和可用药理剂，锌和N-乙酰基半胱氨酸的分子机制和治疗干预措施的分子机制来直接推进对肝病的研究。 公共卫生相关性：我们将在威尔逊氏病小鼠动物模型中测量核受体的DNA结合活性，这是一种慢性疾病，其特征是过度肝铜的积累和严重的肝功能障碍，如果不治疗，导致死亡。我们的研究将确定分子机制受到过度铜水平和治疗方案破坏的分子机制，以恢复肝肝基因表达的核受体调节，从而改善肝功能。这项研究专门涉及与人肝脏病理和威尔逊氏病有关的生物学机制。