REGULATION OF 15N UREA ISOTOPOMERS PRODUCTION

描述（由申请人提供）：正常和病理状态下肝性尿潴留的急性调控一直是广泛研究和众多争议的主题。最近，我们发现精氨酸代谢通过精氨酸脱羧酶（ADC）反应产生的谷氨酸刺激脂肪酸氧化（FAO），从而合成n -乙酰谷氨酸（NAG），这是磷酸氨基甲酰合成酶- i （CPS-I）的活化剂。此外，从婴儿高胰岛素/高氨血症（HI/HA）综合征转基因小鼠模型中获得的结果显示，胍丁氨酸可提高肝脏中NAG水平和尿素合成。因此，胍丁氨酸可能被证明是一种有价值的治疗HI/HA的辅助药物。因此，目前更新建议的总目标是：(i)阐明agmatine调节FAO、NAG和尿素合成的机制；（ii）仔细研究先天性HI损害肝脏尿潴积并导致HA的机制。在这些目的中所寻求的信息可能会推进agmatine在治疗受损性尿原中的潜在应用。根据我们的研究结果，我们建议探索两个主要假设：(i) agmatine刺激FAO，触发代谢级联反应，导致乙酰辅酶a和谷氨酸的可获得性增加，用于NAG的合成，从而导致CPS-I激活；（ii） HI/HA患者β细胞不受控制的胰岛素释放增加导致FAO下降。由于gdh相关的功能获得突变导致FAO的减少，加上肝脏谷氨酸氧化增加，导致乙酰辅酶A和谷氨酸的消耗，从而减少NAG和尿素合成。然而，胍丁氨酸将通过刺激FAO和增加NAG合成来逆转这种代谢级联反应。

DESCRIPTION (provided by applicant): The acute regulation of hepatic ureagenesis under normal and pathologic states has been the subject of extensive investigation and numerous controversies. Recently, we have discovered that agmatine, the product of arginine metabolism via the arginine decarboxylase (ADC) reaction, stimulates fatty acid oxidation (FAO), and thus synthesis of N-acetylglutamate (NAG), an activator of carbamoyl phosphate synthetase-I (CPS-I). In addition, agmatine elevates NAG levels and urea synthesis in livers obtained from the transgenic mouse model of infant hyperinsulinism/hyperammonemia (HI/HA) syndrome. Thus, agmatine might prove a valuable therapeutic adjunct in the case of HI/HA. Therefore, the overall aims of the current renewal proposal are: (i) To elucidate the mechanism(s) by which agmatine regulates FAO, NAG and urea synthesis; and (ii) To scrutinize the mechanism(s) by which congenital HI impairs hepatic ureagenesis and leads to HA. The information sought in these aims may advance the potential application of agmatine in the treatment of impaired ureagenesis. Based on our findings we propose to explore two main hypotheses: (i) The stimulation of FAO by agmatine, triggers a metabolic cascade that leads to an increased availability of acetyl-CoA and glutamate for the synthesis of NAG, thus resulting in the activation of CPS-I; and (ii) The increased uncontrolled release of insulin by beta-cells in HI/HA patients leads to decreased FAO. A decrease in FAO, together with increased hepatic glutamate oxidation, resulting from the GDH-linked gain-of-function mutation, leads to the depletion of acetyI-CoA and glutamate and thus, diminished NAG and urea synthesis. Agmatine, however, will reverse this metabolic cascade by stimulating FAO and increasing NAG synthesis. We will use wild type and transgenic mice expressing the glutamate dehydrogenase (GDH) gain-of-function mutation in pancreatic beta-cells or the liver as a model of human HI/HA. In conjunction, we will use state-of-the-art methodologies, including 15N and/or 13C labeled precursors, Gas Chromatography-Mass Spectrometry (GC-MS) and Nuclear Magnetic Resonance (NMR), to determine the beneficial effect of agmatine on hepatic NAG and ureagenesis in this mouse model of HI/HA. The proposed studies are of clinical as well as scientific significance. The data to be generated may have tremendous clinical impact by advancing the development of a protocol to ameliorate impaired urea synthesis.