Identification of biomarkers for biochemical, pathophysiological and neurological effects of high ammonia concentration on the central nervous system in a preclinical model of neonatal hyperammonemia

在新生儿高氨血症临床前模型中鉴定高氨浓度对中枢神经系统生化、病理生理学和神经学影响的生物标志物

• 批准号: 10302593
• 负责人: Nicholas Ah Mew
• 金额: $ 26.78万
• 依托单位: CHILDREN'S RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-18 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10302593
• 关键词: Acute; Adult; Affect; Ammonia; Animal Model; Animals; Argininosuccinate lyase deficiency; Astrocytes; Biochemical; Biological Markers; Biotinidase Deficiency; Blood; Brain; Brain Edema; Brain Injuries; Cells; Cessation of life; Chemicals; Chronology; Citrulline; Citrullinemia; Coma; Complement; Cultured Cells; Data; Development; Disease; Electroencephalography; Epilepsy; Exposure to; Frequencies; Functional disorder; Glutamates; Glutamine; Goals; Holocarboxylase Synthetase Deficiency; Human; Hyperammonemia; Hyperargininemia; Hypoxia; Impairment; Infant; Intellectual functioning disability; Knock-out; Knockout Mice; Lead; Ligase; Liver; Long-Term Effects; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Maple Syrup Urine Disease; Measures; Metabolism; Methods; Modeling; Molecular; Monitor; Mus; N acetyl L glutamate; N-carbamylglutamate; Neonatal; Neonatal Brain Injury; Neuraxis; Neurocognitive Deficit; Neurologic; Neurologic Effect; Neuron-Specific Enolase; Newborn Animals; Organ; Ornithine Carbamoyltransferase; Outcome; Patient-Focused Outcomes; Patients; Pattern; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Pre-Clinical Model; Preclinical Testing; Protons; Recovery; Resources; Risk; Seizures; Serum; Severities; Signal Transduction; Stress; Supplementation; Symptoms; Testing; Toxic effect; Transplant Recipients; United States Health Resources and Services Administration; Validation; Water; Withdrawal; Zebrafish; awake; base; biomarker identification; brain abnormalities; brain electrical activity; drug testing; experience; experimental study; fatty acid oxidation; imaging study; improved outcome; injury recovery; insight; liver transplantation; mature animal; neonatal brain; neonatal mice; neonate; neuroprotection; neurotoxicity; novel; novel therapeutics; postnatal; preclinical evaluation; prevent; research clinical testing; screening panel; therapeutically effective; two photon microscopy

Abstract Neonatal hyperammonemia (HA) from any cause, including several screenable disorders, results in brain injury leading to irreversible intellectual and developmental disabilities, and even death. Current therapies for HA are targeted at reducing blood ammonia levels; although they can prevent death from brain edema, they are inefficient at reducing or preventing brain damage. Our understanding of the mechanism of ammonia toxicity to the brain is based on experiments in cultured cells and adult animal models; HA disrupts glutamine, glutamate, and K+ metabolism in the astrocytes leading to osmotic stress and brain edema. However, it is not known how HA affects the developing brain because animal models suitable for studying molecular, biochemical, pathophysiological, and neurological effects of HA on the neonatal brain currently do not exist. We have created an animal model of inducible HA, the N-acetylglutamate synthase knockout (NAGSko) mouse. Homozygous knockout mice survive into adulthood and reproduces when treated with N-carbamylglutamate and citrulline, and develop HA when treatment is stopped. We propose to use the NAGSko mice as a model of inducible neonatal HA. Our goal is to establish biomarkers of HA that could be used in both preclinical and clinical testing of neuroprotection drugs. Our specific aims are: 1. To induce HA in neonatal NAGSko mice and determine whether associated MRS biochemical changes in their brains persist after ureagenesis has been normalized. We will induce HA in NAGSko mice at postnatal day 13 (P13) and measure metabolite differences in the brains of HA NAGSko mice and non-HA littermates during HA episode and 2 weeks post recovery from HA. Brain metabolites will be measured using proton magnetic resonance spectroscopy (1H-MRS). 2. To determine whether neonatal HA episode causes persistent abnormal astrocyte function that correlates with blood biomarkers of brain damage. We will use our novel NAGSko/ALDH1L1/GCamp5G-tdTm mice and 2-photon microscopy to monitor changes in Ca2+ signaling during neonatal HA episode and its long-term consequences on astrocytic Ca2+ signaling in awake animals, Changes in Ca2+ signaling will be correlated to serum biomarkers of brain injury S100B and NSE (neuron-specific enolase). 3. To determine whether neonatal HA episode causes persistent abnormal brain electrical activity. We will assess whether a neonatal HA episode increases frequency and severity of seizures after normalization of ureagenesis in the NAGSko mice. After validation, we plan to use EEG patterns, 1H-MRS and blood biomarkers of brain damage as biomarkers of HA in pre-clinical and clinical evaluation of drugs and therapies for the protection of the brain from ammonia toxicity. If successful, drugs that result from these trials will complement current treatment approaches and improve the outcome of patients with HA.

摘要 任何原因引起的新生儿高氨血症(HA)，包括几种可筛查的疾病，都会导致脑损伤 导致不可逆转的智力和发育障碍，甚至死亡。目前治疗HA的方法有 旨在降低血氨水平；尽管它们可以防止死于脑水肿，但它们 在减少或预防脑损伤方面效率低下。我们对氨中毒机制的认识 大脑基于培养细胞和成年动物模型的实验；HA破坏谷氨酰胺，谷氨酸， 星形胶质细胞的K+代谢导致渗透应激和脑水肿。然而，目前还不知道是如何 HA影响发育中的大脑，因为动物模型适合研究分子、生化、 目前尚不存在HA对新生儿大脑的病理生理和神经学影响。我们有 建立了诱导性HA的动物模型，N-乙酰谷氨酸合成酶基因敲除(NAGSko)小鼠。 纯合子基因敲除小鼠在N-氨基甲酰谷氨酸和N-氨基甲酰谷氨酸处理后存活并繁殖 瓜氨酸，并在治疗停止时产生HA。我们建议使用NAGSko小鼠作为一种 可诱导的新生儿HA。我们的目标是建立可用于临床前和临床的HA生物标记物 神经保护药物的测试。我们的具体目标是：1.诱导新生NAGSko小鼠HA并测定 在尿失禁正常化后，他们大脑中相关的MRS生化变化是否会持续下去。 我们将在出生后第13天(P13)在NAGSko小鼠体内诱导HA，并测量大脑中代谢物的差异 在HA发病期间和从HA中恢复2周后，观察HA NAGSko小鼠和非HA仔鼠的情况。脑区 代谢物将使用质子磁共振波谱(1H-MRS)进行测量。2.确定 新生儿HA事件是否导致持续性星形胶质细胞功能异常与血液相关 脑损伤的生物标志物。我们将使用我们的新型NAGSko/ALDH1L1/GCamp5G-TDTM小鼠和双光子 显微镜监测新生儿HA发病过程中钙信号的变化及其长期后果 清醒动物星形胶质细胞钙信号的变化将与血清生物标志物相关 脑损伤的S100B和NSE(神经元特异性烯醇化酶)。3.确定新生儿HA发病是否导致 持续异常的脑电活动。我们将评估新生儿HA发作是否会增加频率 以及NAGSko小鼠尿失禁正常化后癫痫发作的严重程度。经过验证后，我们计划使用 脑电图型、1H-MRS和脑损伤的血液生物标志物作为HA的临床前和临床生物标志物 保护大脑免受氨中毒影响的药物和治疗方法的评价。如果成功，药物 这些试验的结果将补充目前的治疗方法，并改善 患有HA的患者。