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: 10490324
• 负责人: Nicholas Ah Mew
• 金额: $ 22.31万
• 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 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10490324
• 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; Persons; 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.

抽象的 任何原因（包括多种可筛查疾病）引起的新生儿高氨血症 (HA) 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HA的治疗方法有 旨在降低血氨水平；虽然它们可以防止因脑水肿而死亡，但它们 无法有效减少或预防脑损伤。我们对氨毒性机制的了解 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、 星形胶质细胞中的 K+ 代谢导致渗透压和脑水肿。但目前尚不清楚如何 HA 影响发育中的大脑，因为动物模型适合研究分子、生物化学、 目前还不存在 HA 对新生儿大脑的病理生理学和神经学影响。我们有 创建了诱导型 HA 动物模型，即 N-乙酰谷氨酸合酶敲除 (NAGSko) 小鼠。 纯合基因敲除小鼠在接受 N-氨甲酰谷氨酸和 N-氨甲酰谷氨酸治疗后可存活至成年并繁殖 瓜氨酸，并在治疗停止时形成 HA。我们建议使用 NAGSko 小鼠作为模型 诱导型新生儿HA。我们的目标是建立可用于临床前和临床的 HA 生物标志物 神经保护药物的测试。我们的具体目标是： 1. 在新生 NAGSko 小鼠中诱导 HA 并确定 尿素生成正常化后，大脑中相关的 MRS 生化变化是否持续存在。 我们将在出生后第 13 天 (P13) 在 NAGSko 小鼠中诱导 HA 并测量大脑中代谢物的差异 HA发作期间和HA恢复后2周的HA NAGSko小鼠和非HA同窝小鼠的结果。脑 将使用质子磁共振波谱 (1H-MRS) 测量代谢物。 2. 确定 新生儿HA发作是否会导致与血液相关的持续异常星形胶质细胞功能 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 2 光子 显微镜监测新生儿 HA 发作期间 Ca2+ 信号传导的变化及其长期后果 关于清醒动物星形细胞 Ca2+ 信号传导，Ca2+ 信号传导的变化将与血清生物标志物相关 脑损伤 S100B 和 NSE（神经元特异性烯醇化酶）。 3. 确定新生儿HA发作是否引起 持续异常的脑电活动。我们将评估新生儿 HA 发作的频率是否会增加 以及 NAGSko 小鼠尿素生成正常化后癫痫发作的严重程度。验证后，我们计划使用 脑电图模式、1H-MRS 和脑损伤的血液生物标志物作为临床前和临床中 HA 的生物标志物 评估保护大脑免受氨中毒的药物和疗法。如果成功的话，药物 这些试验的结果将补充当前的治疗方法并改善治疗结果 患有HA的患者。