Regulation of 15N Urea Isotopomers Production

15N尿素同位素生产监管

• 批准号: 8068083
• 负责人: ITZHAK NISSIM
• 金额: $ 10万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-06-18 至 2010-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8068083
• 关键词: 5' -AMP-activated protein kinase; Acetyl Coenzyme A; Acetyl-CoA Carboxylase; Acute; Address; Adult; Affect; Agmatine; Amination; Arginine; Arginine decarboxylase; Aspartate; Attenuated; Biochemical; Carbamoyl-Phosphate Synthase (Ammonia); Carbamyl Phosphate; Child; Chronic; Citric Acid Cycle; Citrulline; Congenital Abnormality; Cyclic AMP; Data; Development; Disease; Down-Regulation; Face; Failure; Fatty Liver; Funding; Glutamate Dehydrogenase; Glutamates; Glutaminase; Glutamine; Guidelines; Hepatic; Hepatocyte; Hyperammonemia; Hyperinsulinism; Intervention; Kinetics; Knowledge; Label; Lead; Lipids; Liver; Liver diseases; MALDI-TOF Mass Spectrometry; Malonyl Coenzyme A; Mass Fragmentography; Mediating; Medical; Metabolic; Metabolic syndrome; Metabolism; Methodology; Mitochondria; Modeling; Molecular Biology; N acetyl L glutamate; NADH; Nuclear Magnetic Resonance; Outcome; Oxaloacetates; Patients; Perfusion; Production; Progress Reports; Protein Isoforms; Publications; Pyruvate Carboxylase; Rattus; Regulation; Role; Secondary to; Signal Pathway; Supplementation; Synthase I; System; Testing; Up-Regulation; Urea; Zucker Rats; base; diagnosis evaluation; effective therapy; enzyme activity; fatty acid oxidation; hepatic ureagenesis; improved; in vivo; inorganic phosphate; lipid biosynthesis; mitochondrial dysfunction; nitrogen metabolism; non-alcoholic fatty liver; novel; oxidation; protein degradation; public health relevance; research study; tool; uptake; urea cycle

DESCRIPTION (provided by applicant): Impaired urea synthesis and consequent hyperammonemia (HA) are common occurrences in disorders of congenital defects of the urea cycle and of fatty acid oxidation (FAO), nonalcoholic fatty liver disease (NAFLD) and/or "Metabolic Syndrome" (MS). Still unknown are the biochemical and metabolic mechanisms by which defective FAO and fatty liver impair ureagenesis. Nor is an effective treatment available. During the current funding period we found that agmatine (AGM), the product of arginine decarboxylase, elevates hepatic [cAMP] and stimulates both ureagenesis and FAO. Our preliminary data demonstrate that AGM or 5-aminoimidazole-4-carboxamide-1-¿-D-ribofuranoside (AICAR), an activator of AMP-activated protein kinase (AMPK), enhances ureagenesis in a rat model of NAFLD or MS. Together, the observations strongly suggest that AGM has many of the effects expected for an activator of AMPK. In this renewal proposal our overall aim is to elucidate the mechanisms by which AGM or AICAR regulates hepatic glutamine metabolism and urea synthesis in NAFLD or MS. A long-term objective is to develop a clinically applicable pharmacotherapeutic intervention to improve ureagenesis in patients with NAFLD and/or MS. We propose to explore two related Specific Aims/Hypotheses: (i) Impaired ureagenesis in NAFLD is a consequence of mitochondrial dysfunction and a resultant decrease in synthesis of N-acetylglutamate (NAG), an obligatory activator of carbamoyl phosphate synthetase-I (CPS-I), the initial and the rate-limiting step of ureagenesis. AGM and AICAR augment FAO, thereby triggering a metabolic cascade that attenuates the metabolic derangements associated with NAFLD and MS. The result is an augmentation of ureagenesis; and (ii) NAFLD decreases hepatic uptake and metabolism of glutamine. This would limit mitochondrial [glutamate] and NAG synthesis. The net result is a failure of activation of CPS-I. AGM and AICAR stimulate FAO, improve glutamine uptake and permit more glutamate to be available for NAG synthesis, and thus, greater CPS-I activity. Based on these hypotheses, questions to be addressed include: (1) Is the action of AGM on FAO and ureagenesis mediated via activation of AMPK and/or cAMP-PKA? (2) How does acute or chronic treatment with AGM or AICAR affect metabolic coordination between hepatic FAO, the TCA cycle and ureagenesis in NAFLD or MS?; and (3) How does treatment with AGM or AICAR and subsequent activation of AMPK and/or cAMP-PKA affect hepatic glutamine uptake and metabolism, whole-body protein turnover and ureagenesis in NAFLD or MS. Experiments will be performed using a rat model of fatty liver and/or MS and various systems, including (a) isolated hepatocytes; (b) a liver perfusion system; (c) isolated mitochondria; and (d) in vivo study. We will use 15N and/or 13C labeled precursors and gas chromatography-mass spectrometry (GC-MS), MALDI-TOF-mass spectrometry, nuclear magnetic resonance (NMR) and molecular biology. Combining these methodologies provides a superb tool to pinpoint the primary mechanism(s) of AGM or AICAR action. Data obtained will provide new and pivotal information to elucidate the role of AGM or AICAR in the regulation of hepatic glutamine metabolism and ureagenesis. This knowledge may lead to development of a novel pharmacotherapeutic intervention to improve urea synthesis in NAFLD or MS. PUBLIC HEALTH RELEVANCE: Defective urea synthesis and consequent toxic hyperammonemia (HA) are common occurrences in case of nonalcoholic fatty liver disease (NAFLD) or Metabolic Syndrome (MS), an increasingly recognized medical problem in adults and children. The overall aims of this proposal are: (i) to elucidate the mechanism(s) regulating hepatic urea synthesis in NAFLD or MS; and (ii) to develop a clinically applicable pharmacotherapeutic intervention to improve ureagenesis in patients with NAFLD and/or MS. Our preliminary results suggest that such treatment is feasible. The outcome of the propose study may provide new guidelines for evaluation, diagnosis and treatment of defective urea synthesis in NAFLD or MS.

描述（由申请人提供）：尿素合成受损和随之而来的高氨血症 (HA) 是先天性尿素循环缺陷和脂肪酸氧化 (FAO)、非酒精性脂肪肝 (NAFLD) 和/或“代谢综合征”疾病中常见的情况缺陷性FAO和脂肪肝损害尿素生成的生化和代谢机制仍不清楚。在目前的资助期间，我们发现精氨酸脱羧酶的产物胍丁胺 (AGM) 会升高肝脏 [cAMP] 并刺激尿素生成和FAO。我们的初步数据表明，AGM 或 5-氨基咪唑-4-甲酰胺-。 1-¿ -D-呋喃核苷 (AICAR) 是 AMP 激活蛋白激酶 (AMPK) 的激活剂，可增强 NAFLD 或 MS 大鼠模型中的尿素生成，所有观察结果表明 AGM 具有 AMPK 激活剂所预期的许多作用。这项更新提案的总体目标是阐明 AGM 或 AICAR 调节 NAFLD 或 MS A 中肝脏谷氨酰胺代谢和尿素合成的机制。长期目标是开发一种临床适用的药物治疗干预措施，以改善 NAFLD 和/或 MS 患者的尿素生成。我们建议探索两个相关的具体目标/假设：(i) NAFLD 尿素生成受损是线粒体功能障碍和线粒体功能障碍的结果。 N-乙酰谷氨酸（NAG）的合成减少，NAG是氨基甲酰磷酸合成酶-I（CPS-I）的必需激活剂， AGM 和 AICAR 增强了尿素生成的初始和限速步骤，从而引发了代谢级联，从而减弱了与 NAFLD 和 MS 相关的代谢紊乱，并且 (ii) NAFLD 减少了肝脏的摄取和代谢。这会限制线粒体 [谷氨酸] 和 NAG 的合成，最终结果是 AGM 和 AICAR 刺激失败。改善谷氨酰胺吸收并允许更多谷氨酸用于 NAG 合成，从而提高 CPS-I 活性。基于这些假设，需要解决的问题包括：（1）AGM 对FAO 和尿素生成的作用是否通过介导。 AMPK 和/或 cAMP-PKA 的激活？ (2) AGM 或 AICAR 的急性或慢性治疗如何影响 NAFLD 或 NAFLD 中肝脏 FAO、TCA 循环和尿素生成之间的代谢协调？ MS？；以及（3）AGM 或 AICAR 治疗以及随后的 AMPK 和/或 cAMP-PKA 激活如何影响 NAFLD 或 MS 中的肝脏谷氨酰胺摄取和代谢、全身蛋白质周转和尿素生成。脂肪肝和/或 MS 的大鼠模型和各种系统，包括 (a) 分离的肝细胞；(b) 肝脏灌注系统；(c) 分离的线粒体；以及 (d) 体内研究。和/或 13C 标记前体和气相色谱-质谱 (GC-MS)、MALDI-TOF-质谱、核磁共振 (NMR) 和分子生物学结合这些方法提供了一个极好的工具来查明主要机制。获得的数据将为阐明 AGM 或 AICAR 在肝脏谷氨酰胺代谢和尿素生成的调节中的作用提供新的关键信息。这些知识可能会导致开发一种新的药物治疗干预措施，以改善 NAFLD 或 MS 中的尿素合成。 公共健康相关性：尿素合成缺陷和随之而来的中毒性高氨血症 (HA) 在非酒精性脂肪肝病 (NAFLD) 或代谢综合征 (MS) 中很常见，这是成人和儿童日益认识到的医疗问题。本提案的总体目标。是：(i) 阐明 NAFLD 或 MS 中肝脏尿素合成的调节机制；以及 (ii) 开发临床适用的方法；我们的初步结果表明，这种治疗方法是可行的，可以为 NAFLD 或 MS 患者尿素合成缺陷的评估、诊断和治疗提供新的指导。