Basic and clinical studies in immune function and metabolism

免疫功能和代谢的基础和临床研究

• 批准号: 8565575
• 负责人: Peter McGuire
• 金额: $ 75.19万
• 依托单位: NATIONAL HUMAN GENOME RESEARCH INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8565575
• 关键词: Acidosis; Acute; Address; Affect; Age; Amino Acids; Ammonia; Animals; Area; Arginine; Argininosuccinate lyase deficiency; Biological Models; CD3 Antigens; Catabolism; Cell Differentiation process; Cell Maintenance; Cell physiology; Cell surface; Cells; Cellular biology; Citrulline; Citrullinemia; Clinical; Clinical Data; Clinical Research; Communicable Diseases; Coupled; Cut protein; Cytokine Signaling; Defect; Development; Diet; Disease; Energy Intake; Enzymes; Event; Functional disorder; Future; Genes; Genetic; Goals; Hepatic; Hepatitis A Vaccines; Hepatocyte; Human; Hyperammonemia; Hypoglycemia; Immune; Immune System Diseases; Immune system; Immunity; Immunization; In Vitro; Inborn Errors of Metabolism; Individual; Infection; Infectious Agent; Inflammatory; Influenza; Intoxication; Laboratories; Lead; Link; Liver; Liver diseases; Longitudinal Studies; Lymphopenia; Malnutrition; Manuscripts; Measures; Mediating; Medical; Memory; Mesentery; Metabolic; Metabolic Diseases; Metabolic Pathway; Metabolism; Modeling; Molecular; Molecular Biology; Morbidity - disease rate; Mus; Nature; Nutrition Disorders; Ornithine carbamoyltransferase deficiency; Pathologic; Pathway interactions; Patients; Perception; Phenotype; Plasma; Polyamines; Predisposition; Preparation; Process; Proteins; Protocols documentation; Publishing; Rare Diseases; Recruitment Activity; Research; Role; Running; Spleen; T cell differentiation; T-Cell Activation; T-Cell Development; T-Cell Proliferation; T-Cell Receptor; T-Lymphocyte; Time; Translational Research; United States National Institutes of Health; Viremia; Virus Diseases; Work; adaptive immunity; argininosuccinate lyase; argininosuccinate synthase; clinically significant; cytokine; gene replacement therapy; immune activation; immune function; in vivo; insight; interest; lymph nodes; metabolomics; mouse model; novel; programs; prospective; protein intake; respiratory; response; stable isotope; treatment strategy; urea cycle; vaccine efficacy

Summary: Our translational research program studies intermediary metabolism and the immune system with a focus on inborn errors of metabolism. Intermediary metabolism is critical for the activation and differentiation of T-cells. Abnormalities in these pathways may lead to defects in effector and memory T-cell functions. Using a combination of mouse genetics, metabolomics, fluxomics, molecular and cell biology, and mouse models of inborn errors of metabolism, our goals are to understand how intermediary metabolism contributes to the normal function of cells in the immune system and their dysfunction in inborn errors of metabolism. This immune dysfunction may be primary or secondary. Genes involved in intermediary metabolism may be expressed in T-cells. In addition, intoxication phenotypes (e.g. acidosis or hyperammonemia) may also precipitate immune dysfunction. We are particularly interested in how these pathways affect responses to infectious diseases and immunization.
In addition, we also focus on disruptions of hepatic metabolism due to activation of the immune system systemically and locally. Hepatic metabolic decompensations in inborn errors of metabolism (e.g. hyperammonemia, acidosis, hypoglycemia) are most often precipitated by infection. Specifically our aims are: 1) To understand how intermediary metabolism affects T cell development, differentiation and function. A special focus is placed in understanding how metabolic enzyme defects and intoxication phenotypes in inborn errors of metabolism affect T-cell effector and memory cell differentiation and function in response to immunization and infection. This work is being pursued in the laboratory with mouse models as well as affected patients in the NIH clinical center. 2) To understand how immune activation systemically or locally can precipitate hepatic metabolic dysfunction in inborn errors of metabolism. Our current work focuses on modeling respiratory viral infection-induced hepatopathy and resultant hyperammonemia in mouse models of urea cycle disorders. We are working on the mechanisms of cytokine and viremia induced urea cycle dysfunction. Research: Intermediary metabolism and T-cells: Over the past year we have spent our time characterizing the role of the amino acids citrulline and arginine in T-cell function. For T-cells, arginine is essential for T-cell proliferation and maintenance of the T-cell receptor. In T-cells deprived of arginine, the CD3 zeta chain and T-cell receptor are downregulated from the cell surface reducing proliferation. Using stable isotopes, the fate ultimate fate of arginine so far seems to be protein and polyamine synthesis. These pathways are currently being dissected out further. During arginine deficiency states, such as during an infection, cells may utilize citrulline by upregulating argininosuccinate synthetase (ASS1) and argininosuccinate lyase (ASL) to make arginine. ASS1 and ASL are also enzymes of the hepatic urea cycle. This implies that individuals with urea cycle disorders due to ASS1 (citrullinemia) or ASL (argininosuccinic aciduria) deficiencies may have a conditional immune defect under low arginine conditions. To answer these questions, we used a mouse model of citrullinemia. These animals are unable to convert citrulline to arginine and have hypercitrullinemia and hyperammonemia. Pathologic studies revealed that these mice had T and B lymphopenia, small spleens, and absent mesenteric lymph nodes. Since most of the animals die by 3 weeks of age, in conjunction with the Venditti laboratory, we developed a liver-targeted gene replacement therapy approach. This approach would also allow us to study cell autonomous ASS1 defects. To date, we have rescued several animals, which still retain the ASS1 defect in their T-cells. We are currently in the process of evaluating T-cell function using influenza A immunization and natural infection approaches. To address the effects of systemic intoxication with ammonia on immune function, we are working with another model of a urea cycle disorder, the spf-ash mouse (ornithine transcarbamylase deficiency (OTC)). These mice are hyperammonemic at baseline. Immunization studies against influenza thus far have demonstrated reduced vaccine efficacy against natural infection. We are continuing to characterize the nature of this defect, concentrating on aspects of ammonia intoxication of T-cells. To accomplish this, we are characterizing a new model of OTC deficiency, the spf-j, which will be published by our lab shortly. The advantage of this model over previous models is that the mouse is also hyperammonemic but on a clean B6 background unlike its predecessor. These basic studies have extended to the NIH Clinical Center. Our protocol titled the NIH UNI Study: Urea Cycle Disorders, Nutrition and Immunity is up and running and actively recruiting patients. In this protocol we evaluate the development of adaptive immunity to influenza and Hepatitis A vaccines. Immune activation and hepatic intermediary metabolic dysfunction: Our second main area of work involves exacerbation of hepatic metabolic diseases by activation of the immune system. One of the most serious consequences of Urea Cycle Disorder (UCD) is acute hyperammonemia (HA), which may be caused by dietary indiscretion (i.e. high protein intake), dietary deficiency resulting in enhanced catabolism, and acute infection. Acute HA is caused most often by infectious precipitants. There is also a perception that the acute HA that develops is clinically different from other precipitants: higher plasma ammonias of longer duration. Using clinical data from the Rare Disease Clinical Research Network sponsored prospective longitudinal study of the Urea Cycle Disorders Consortium, we were able to show that acute HA due to infection is a distinct clinical entity and displays markers of increased morbidity. This manuscript is in preparation. Although enhanced catabolism is present during dietary deficiency as well as acute infection, we hypothesized that immune activation with inflammatory cytokine release may have more direct effects on hepatic urea cycle function. In support of this idea, we performed in vitro studies in primary human hepatocytes from controls and UCD patients, which demonstrated deleterious effects of inflammatory cytokines on ammonia metabolism. To examine this mechanism in vivo, we developed a model system of acute HA due to infection using two well-defined models, mouse adapted influenza A/PR/8 and the spf-ash mouse, a model of OTC deficiency. Via fluxomic and targeted metabolomic studies, we demonstrated that influenza infection results in unique perturbations in the disposal of ammonia and the availability of anaplerotic intermediates of the urea cycle. The clinical significance and translational importance of these findings are highlighted in the current management of acute HA in UCD. Regardless of the acute HA precipitant, the medical management strategy is the same: cut protein intake for 1-2 days, and provide high caloric intake. These measures are not always successful. Our findings provide novel insight into the metabolic pathophysiology behind acute infection, provides evidence for the use of alternative pathways of ammonia disposal in the spf-ash mouse and the possibility of applying anaplerotic intermediates or immune modifiers. This suggests that the ubiquitously applied treatment strategy of reversing catabolism during acute HA due to infection could be augmented. This work has resulted in a manuscript that is at present submitted for review. Future directions for this work include dissecting the molecular and metabolic events behind urea cycle inhibition. We will focus on cytokine signaling in hepatocytes and links to metabolism as well as viremia induced hepatopathy.

摘要：我们的转化研究计划研究中介代谢和免疫系统，重点是天生的新陈代谢错误。中间代谢对于T细胞的激活和分化至关重要。这些途径中的异常可能会导致效应子和记忆T细胞函数的缺陷。使用小鼠遗传学，代谢组学，通量学，分子和细胞生物学以及代谢的先天误差的小鼠模型的组合，我们的目标是了解中介代谢如何促进免疫系统中细胞的正常功能及其在生物误差中的异常功能。这种免疫功能障碍可能是初级或次要的。参与中间代谢的基因可以在T细胞中表达。另外，醉酒表型（例如酸中毒或高症血症）也可能导致免疫功能障碍。我们对这些途径如何影响对传染病和免疫的反应特别感兴趣。
此外，由于免疫系统在系统和局部激活，我们还专注于肝代谢的破坏。天生代谢错误（例如高症，酸中毒，低血糖）的肝代谢失代偿作用通常是由于感染而导致的。 特别是我们的目标是： 1）了解中介代谢如何影响T细胞的发展，分化和功能。特殊的重点是理解代谢酶缺损和代谢中的中毒表型如何影响T细胞效应子以及记忆细胞分化以及对免疫和感染的作用。这项工作是在实验室中使用小鼠模型以及NIH临床中心影响的患者进行的。 2）了解免疫激活如何系统地或局部如何在代谢的天生误差中沉淀肝代谢功能障碍。我们目前的工作着重于在尿素周期疾病的小鼠模型中对呼吸道病毒感染引起的肝病和由此产生的高氨血症进行建模。我们正在研究细胞因子和病毒血症诱导的尿素周期功能障碍的机制。 研究： 中间代谢和T细胞：在过去的一年中，我们花了时间来表征氨基酸瓜氨酸和精氨酸在T细胞功能中的作用。对于T细胞，精氨酸对于T细胞受体的T细胞增殖和维持至关重要。在被剥夺精氨酸的T细胞中，CD3 Zeta链和T细胞受体从细胞表面下调，减少了增殖。使用稳定的同位素，到目前为止，精氨酸的命运最终命运似乎是蛋白质和多胺合成。这些途径目前正在进一步剖析。在精氨酸缺乏状态期间，例如在感染期间，细胞可以通过上调Argininoscinate合成酶（ASS1）和Argininoscicatine裂解酶（ASL）来利用瓜氨酸来制造精氨酸。 ASS1和ASL也是肝尿素周期的酶。这意味着由于ASS1（瓜氨酸血症）或ASL（Argininosonoccinic酸性）缺陷引起的尿素周期疾病的个体可能在低精氨酸条件下可能有条件免疫缺陷。为了回答这些问题，我们使用了柑橘类血症的小鼠模型。这些动物无法将瓜氨酸转化为精氨酸，并且患有高胸链素血症和高症血症。病理学研究表明，这些小鼠患有T和B淋巴细胞减少症，小脾和肠系膜淋巴结。由于大多数动物在3周龄时死亡，并与venditti实验室一起死亡，因此我们开发了一种肝靶向的基因置换疗法方法。这种方法还可以使我们能够研究细胞自主的ASS1缺陷。迄今为止，我们已经救出了几只动物，这些动物仍然保留了其T细胞中的Ass1缺陷。我们目前正在使用流感A免疫和自然感染方法评估T细胞功能。为了解决氨中毒对免疫功能的影响，我们正在使用尿素周期疾病的另一种模型SPF-ASH小鼠（Ornithine transcarbamylase缺乏症（OTC））。这些小鼠在基线时是高氨血症。到目前为止，针对流感的免疫研究表明，针对自然感染的疫苗功效降低了。我们继续表征这种缺陷的性质，集中在T细胞中氨中毒的各个方面。 为了实现这一目标，我们正在表征一个新的OTC缺陷模型SPF-J，该模型将不久由我们的实验室发布。该模型的优点比以前的模型的优点是，小鼠也是高症，但在干净的B6背景上与其前身不同。这些基础研究已扩展到NIH临床中心。我们的协议标题为NIH UNI研究：尿素周期障碍，营养和免疫力正在启动并运行并积极招募患者。在此方案中，我们评估了对流感和肝炎A疫苗的适应性免疫的发展。 免疫激活和肝中介代谢功能障碍： 我们的第二个主要工作领域涉及通过激活免疫系统加剧肝代谢疾病。尿素循环障碍（UCD）最严重的后果之一是急性高症血症（HA），这可能是由饮食中的不加差异（即高蛋白质摄入量）引起的，饮食缺乏症会导致分解代谢增强和急性感染。急性HA最常由传染性降水剂引起。人们还认为，出现的急性HA在临床上与其他沉淀剂不同：持续时间较长的血浆氨氨基。利用稀有疾病临床研究网络的临床数据赞助了尿素周期疾病联盟的前瞻性纵向研究，我们能够证明由于感染而引起的急性HA是一个独特的临床实体，并且显示出发病率增加的标志物。此手稿正在准备。 尽管在饮食缺乏症和急性感染过程中存在增强的分解代谢，但我们假设炎性细胞因子释放的免疫激活可能会对肝尿素周期功能产生更直接的影响。为了支持这一想法，我们在对照组和UCD患者的原代人肝细胞中进行了体外研究，这表明炎性细胞因子对氨代谢的有害影响。为了在体内检查这种机制，我们使用了两个定义明确的模型，即适应的流感A/PR/8和SPF-ASH小鼠，这是OTC缺陷模型，我们开发了急性HA的模型系统。通过通量和靶向代谢组学研究，我们证明了流感感染会导致氨处置的独特扰动以及尿素周期的无毒中间体的可用性。这些发现的临床意义和翻译重要性在UCD中急性HA的当前管理中得到了强调。无论急性HA降水剂如何，医疗管理策略都是相同的：切割蛋白质摄入1-2天，并提供高热量摄入量。这些措施并不总是成功的。我们的发现为急性感染背后的代谢病理生理学提供了新的见解，为在SPF-ASH小鼠中使用氨处置的替代途径提供了证据，以及应用交换性中间体或免疫修饰的可能性。这表明可以增加由于感染引起的急性HA期间分解代谢的普遍应用治疗策略。 这项工作导致了目前已提交审查的手稿。这项工作的未来方向包括剖析尿素周期抑制背后的分子和代谢事件。我们将重点关注肝细胞中的细胞因子信号传导，并与代谢以及病毒血症诱导的肝病的联系。