CD73 is a multi-functional protein that regulates liver injury

CD73是一种调节肝损伤的多功能蛋白

• 批准号: 9882993
• 负责人: Natasha T Snider
• 金额: $ 34.43万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-03-01 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9882993
• 关键词: 5' -Nucleotidase; ATF2 gene; Active Sites; Acute; Address; Adenosine; Adenosine Monophosphate; Albumins; Alcohols; Animals; Antibodies; Apoptosis; Binding; Biochemical; Biological; Biology; Cardiovascular Diseases; Catalytic Domain; Cell surface; Cells; Cessation of life; Chemicals; Chronic; Chronic Hepatitis C; Cirrhosis; Clinical; Couples; Data; Development; Diet; Disease; Disease Progression; Down-Regulation; Enzymes; Estrogen receptor positive; Ethanol; Etiology; Event; Fibrosis; Genes; Genetic Transcription; Goals; Hepatocyte; High Fat Diet; Homeostasis; Human; Image; In Vitro; Inflammation; Injury; Knockout Mice; Leukotrienes; Ligands; Lipids; Liver; Liver diseases; LoxP-flanked allele; Malignant Neoplasms; Messenger RNA; Metabolic; Metabolism; Mitochondria; Modeling; Molecular; Mus; Names; National Institute on Alcohol Abuse and Alcoholism; Pain; Pathology; Patients; Pharmaceutical Preparations; Phosphorylation; Play; Predisposition; Primary carcinoma of the liver cells; Protein Isoforms; Proteins; Proteomics; RNA Splicing; Recombinants; Regulation; Research Personnel; Resistance; Resources; Role; Sampling; Signal Transduction; Site; Slice; Solid; Stress; Suggestion; Thromboxane B2; Tissues; Transcript; Up-Regulation; Variant; Virus; Wild Type Mouse; Work; base; chronic liver disease; endoplasmic reticulum stress; experimental study; extracellular; feeding; foot; glycoproteomics; glycosylation; hepatocellular injury; hepatocyte injury; improved; in vivo; insight; liver function; liver injury; loss of function; mRNA Precursor; mutant; non-alcoholic fatty liver disease; nonalcoholic steatohepatitis; novel; problem drinker; protein function; receptor; resilience; screening; therapeutic target

Abstract. Their central importance to the synthetic, secretory and metabolic functions of the liver renders hepatocytes particularly vulnerable to injury. Reinforcing hepatocyte resilience to damaging insults holds potential to block liver disease progression at early stages, independent of etiology. To that end, ecto-5’-nucleotidase (CD73) is a promising novel liver disease target, as it plays critical roles in multiple mouse liver injury models, and is dysregulated in livers of patients with chronic liver diseases of multiple etiologies. The classical function of CD73 is metabolism of extracellular adenosine monophosphate (AMP) to adenosine. We found that global CD73-/- mice are resistant to chemically-induced hepatocellular injury, an unexpected finding given that liver injury induces near-complete loss of cell surface CD73 AMPase activity in wild-type mice. This points to alternative functions, which is also supported by our identification of a novel, catalytically-inactive CD73 isoform (CD73S) in advanced human liver disease. We observe differences in apoptosis-related signaling in primary hepatocytes expressing the two different human CD73 isoforms. CD73 binds two bioactive lipids in vitro, suggesting that it may regulate lipid homeostasis. Our central hypothesis is that non-enzymatic and alternative isoform-related mechanisms of hepatocyte CD73 promote liver injury and disease progression. The hypothesis will be addressed via two specific aims: (i) Define the non-enzymatic and isoform-specific functions of CD73 in hepatocytes and (ii) Examine the in vivo and tissue-level mechanisms of hepatoprotection in the absence of CD73. Our approach for Aim 1 will couple high throughput ligand screening, high content imaging, and proteomics with detailed biochemical, molecular and cell-biological assessments to determine if CD73 is a lipid-sensing receptor or a regulator of ER stress, and to determine the specific function of the non-catalytic CD73S variant in hepatocyte resistance to stress. For Aim 2, we will examine liver-specific contributions of CD73 in vivo using two models of alcohol and high fat diet-induced injury and our newly-generated liver-specific CD73-/- mice. We will utilize recombinant enzyme in vivo and precision-cut liver slices to perform mechanistic rescue and loss-of-function studies. This work will lead to a thorough integration of CD73 function in hepatocytes with the potential to yield novel mechanistic approaches for hepatoprotection.

抽象的。 它们对肝脏的合成、分泌和代谢功能至关重要，使得肝细胞 增强肝细胞对损伤性损伤的抵抗力具有阻断的潜力。 与病因无关的早期肝脏疾病进展，外切 5'-核苷酸酶 (CD73) 是一种药物。 有前景的新型肝病靶标，因为它在多种小鼠肝损伤模型中发挥着关键作用，并且 多种病因的慢性肝病患者的肝脏失调 CD73 的经典功能。 是细胞外单磷酸腺苷（AMP）代谢为腺苷我们发现全局CD73-/-小鼠。 对化学诱导的肝细胞损伤有抵抗力，考虑到肝损伤会诱导肝细胞损伤，这是一个意外的发现 野生型小鼠的细胞表面 CD73 AMPase 活性几乎完全丧失，这表明有替代功能， 我们在先进技术中鉴定出一种新型的、无催化活性的 CD73 亚型 (CD73S)，也支持了这一点。 我们观察到原代肝细胞表达中凋亡相关信号的差异。 两种不同的人类 CD73 亚型在体外结合两种生物活性脂质，表明它可能具有调节作用。 我们的中心假设是非酶促和替代亚型相关的机制。 肝细胞 CD73 会促进肝损伤和疾病进展，这一假设将通过两个具体的过程得到解决。 目标：(i) 定义肝细胞中 CD73 的非酶促和异构体特异性功能，以及 (ii) 检查 在缺乏 CD73 的情况下，我们的目标 1 的方法将是体内和组织水平的肝脏保护机制。 将高通量配体筛选、高内涵成像和蛋白质组学与详细的生物化学结合起来， 分子和细胞生物学评估以确定 CD73 是否是脂质敏感受体或 ER 调节剂 应激，并确定非催化 CD73S 变体在肝细胞抗性中的具体功能 对于目标 2，我们将使用酒精和压力两种模型来检查 CD73 在体内的肝脏特异性贡献。 高脂肪饮食引起的损伤和我们新产生的肝脏特异性 CD73-/- 小鼠我们将利用重组。 体内酶和精确切割的肝脏切片进行机械救援和功能丧失研究。 这项工作将导致肝细胞中 CD73 功能的彻底整合，并有可能产生新的 保肝的机制方法。