Molecular Mechanisms Regulating Ammonia Metabolism

调节氨代谢的分子机制

基本信息

批准号：
10491245
负责人：
I. David Weiner
金额：
$ 33.55万
依托单位：
UNIVERSITY OF FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-12-15 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10491245
关键词：
Acids Acute Address Alkalosis Ammonia Anions Apical Atrophic Bicarbonates CXCL12 gene Cardiovascular system Cell membrane Cell physiology Characteristics Chronic Kidney Failure Clinical Medicine Coupled DNA Sequence Alteration Disease Disease Progression Duct (organ) structure Electron Microscopy Enzymes Event Excretory function GDF15 gene GPR4 gene Gene Combinations Gene Deletion Generations Genes Glycoproteins Health Homeostasis Hypokalemia Impairment In Vitro Intercalated Cell Intercalated Duct Intervention Trial Kidney Knockout Mice Lead Limb structure Maintenance Mediating Metabolic acidosis Metabolism Modeling Molecular Mus Pathway interactions Pattern Persons Phenotype Physical Function Play Potassium Procedures Process Proteins Proton-Translocating ATPases Protons Publishing RNA Splicing Regulation Renal Replacement Therapy Role Series Serum Signal Pathway Signal Transduction Site Techniques Thick Thyroid Function Tests Urine Variant Vesicle Work base bone metabolism collecting tubule structure extracellular glucose metabolism hensin hyperkalemia improved insulin sensitivity kidney interstitial tissue mortality muscle form muscle strength response transcription activator-like effector nucleases

项目摘要

Project Summary/Abstract The renal maintenance of acid-base homeostasis is critical for optimal health. Collecting duct intercalated cells play a central role in this process through adaptive changes in proton secretion, bicarbonate secretion, and Rh glycoproteins-mediated ammonia transport. Our current paradigm is that the primary determinant of this response involves direct effects of extracellular pH on intercalated cells. We suggest a new paradigm. Deletion of the proximal tubule-specific basolateral bicarbonate transporter, NBCe1-A, causes severe metabolic acidosis, yet inhibits intercalated cell phenotypic characteristics of acid secretion and inhibits the intercalated cell plasticity response to acid-loading. This is not an off-target effect of the TALEN gene-editing procedure; identical effects were seen with NBCe1-A/B deletion generated using Cre-lox techniques. Published work shows that the K+ disorders, hypokalemia and hyperkalemia, alter intercalated cell phenotype and plasticity in a pattern which cannot be explained by extracellular pH. The effects of NBCe1-A or NBCe1-A/B deletion, or of K+ disorders likely involve ammonia. Each alters proximal tubule- derived ammonia generation in a pattern which parallels the observed intercalated cell responses. Isolated perfused collecting duct studies show ammonia acutely and directly regulates intercalated cell H⁺ and bicarbonate transport. Thus, we propose a new paradigm, that proximal tubule-derived ammonia, which is concentrated in the renal interstitium by the TAL, is a primary determinant of intercalated cell phenotypic characteristics and plasticity response. Our proposed studies investigate this new paradigm in detail. Specific Aim 1 will determine the effect of gene deletion maneuvers which directly and specifically alter proximal tubule and thick ascending limb ammonia metabolism on intercalated cell phenotype and plasticity. We will use a combination of gene deletion approaches, including proximal tubule-specific deletion of PDG, the initial and the rate-limiting enzyme in ammoniagenesis, and TAL NHE4 deletion, which directly impacts ammonia concentration into the renal interstitium where we propose it regulates intercalated cells. We will study the effect of these gene deletion so in a variety of conditions, including basal state, acid-loading, and abnormal potassium homeostasis, both hypokalemia and hyperkalemia. Specific Aim 2 will determine the signaling mechanisms through which proximal tubule regulates intercalated cell phenotypic characteristics and plasticity. We will identify whether this is direct stimulator of pathways or acts parallel to and independent of signaling pathways known to alter intercalated cells, including GDF15, hensin, GPR4, and SDF1. These studies will substantially advance our understanding of the molecular mechanisms regulating thereby acid-base homeostasis.

项目摘要/摘要酸碱稳态的肾脏维持对于最佳健康至关重要。收集管道插入细胞在此过程中通过质子分泌，碳酸氢盐分泌和 RH糖蛋白介导的氨转运。我们目前的范式是这是主要的决定者反应涉及细胞外pH对插入细胞的直接影响。我们建议一个新的范式。删除近端管特异性基底外侧碳酸氢盐的删除转运蛋白NBCE1-A会导致严重的代谢性酸中毒，但抑制了插层的细胞表型特征酸的分泌并抑制对酸负荷的插入细胞可塑性反应。这不是一个脱离目标塔伦基因编辑程序的影响；通过产生的NBCE1-A/B缺失可见相同的效果使用Cre-Lox技术。已发表的工作表明，K+疾病，低钾血症和高钾血症已改变插入的细胞表型和可塑性，该模式无法通过细胞外pH来解释。效果 NBCE1-A或NBCE1-A/B缺失或K+疾病的含量可能涉及氨。每个都改变了代理管 - 衍生的氨产生的模式与观察到的插入细胞反应相似。孤立完善的收集管道研究表明，氨敏感，直接调节插入的细胞H⁺和碳酸氢盐运输。那就是我们提出了一个新的范式，那是近端管衍生的氨 TAL中浓缩肾脏间质，是插入式细胞表型的主要决定剂特性和可塑性响应。我们提出的研究详细研究了这一新范式。具体目标1将确定基因删除操作的效果，该操作直接，专门改变近端小管和较厚的肢体氨代谢在插入的细胞表型和可塑性上。我们将使用基因缺失方法的组合，包括PDG的近端管特异性缺失以及氨化中的限速酶和TAL NHE4缺失，直接影响氨集中于肾脏间质，我们建议它调节插入的细胞。我们将研究效果这些基因删除中的多种条件中的缺失，包括碱性状态，酸负荷和异常钾稳态，低钾血症和高钾血症。特定的目标2将确定近端细胞调节的信号传导机制插入的细胞表型特征和可塑性。我们将确定这是否是直接的刺激器途径或行为平行于并独立于已知会改变插入细胞的信号通路，包括 GDF15，Hensin，GPR4和SDF1。这些研究将大大提高我们对调节分子机制的理解因此酸碱稳态。