Metabolism Core

新陈代谢核心

• 批准号: 10747722
• 负责人: Leslie S Gewin
• 金额: $ 27.38万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10747722
• 关键词: Address; Affect; Biological Assay; Cell Count; Cells; Chronic; Chronic Kidney Failure; Communities; Consult; Consultations; Data; Disease Progression; Disease model; Doctor of Medicine; Doctor of Philosophy; Education; Electron Transport; Equipment; Faculty; Functional disorder; Genes; Genus Hippocampus; Glucose; Human; In Vitro; Injury; Injury to Kidney; Isotopes; Kidney; Kidney Diseases; Measures; Metabolic; Metabolism; Methods; Mitochondria; Obesity; Palmitates; Pathway interactions; Pharmaceutical Preparations; Play; Pre-Clinical Model; Process; Proteins; Protocols documentation; Pyruvate; Radioactive; Reagent; Recommendation; Renal function; Renal tubule structure; Reproducibility; Reproducibility of Results; Research; Research Personnel; Resolution; Resources; Respiration; Role; Services; Spirometry; Standardization; Tissues; Tracer; Training; Tubular formation; Universities; Update; Validation; Washington; Writing; biomedical resource; design; fatty acid oxidation; in vivo; instrumentation; interest; liquid chromatography mass spectrometry; long chain fatty acid; metabolomics; nutrition; oxidation; protein expression; response; stable isotope; transcriptomics

Project Summary/Abstract: Metabolism Core The Metabolism Core (Meta-Core) is a Biomedical Resource Core within the Washington University Chronic Kidney Disease National Resource Center. Chronically injured kidneys have altered metabolism, and recent data suggests that these metabolic changes play a causative role in progressive decline in kidney function. The ability to interrogate metabolism is crucial to understanding chronic kidney disease pathophysiology, but there are a number of barriers for kidney researchers to do so. First, many investigators interested in chronic kidney disease do not have expertise in metabolism and metabolic assays. Second, many of these assays require expensive equipment not readily available to many labs. To address these issues, the Meta-Core faculty consisting of Leslie Gewin, M.D., Core Director, Brian Finck, Ph.D., Associate Core Director, Gary Patti, Ph.D., Core Collaborator, and Leah Shriver, Ph.D., Core Collaborator, will perform consultations with Meta- Core users to guide and design the best metabolic assay to answer the research question. Several assays will be available to the Meta-Core users to interrogate metabolism at the cellular or whole kidney level. These assays include Seahorse bioflux analyses with the use of primary cells or freshly isolated tubules ex vivo, both more representative of the highly oxidative proximal tubules than commercially available primary human tubule cells or conditionally immortalized tubule cells. We can measure oxidation of fatty acids or glucose/pyruvate in kidney tissue ex vivo using radioactive substrate oxidation assays. The 3H-palmitate assay, not often performed on kidney tissue, has advantages over the more commonly used 14C-palmitate assay as it detects complete oxidation of long chain fatty acids through the electron transport chain and is easier to perform (collect 3H2O rather than 14CO2). We can also refer users to the Nutrition Obesity Research Center’s high- resolution respirometer (Oroboros Oxygraph 2k) to detect respiration in isolated mitochondria. In addition, untargeted metabolomics and stable isotope tracer studies will be performed on primary cells in vitro and in vivo. The Meta-Core faculty are all committed to sharing validated protocols and methods with the O’Brien Consortium as well as larger scientific community. In addition, we will help train the kidney scientific community how to generate primary proximal tubule cells and tubules and use the Seahorse bioflux analysis in a way that yields consistent, reproducible results. In particular, the importance of validating cell number to use, post- analysis correction for number of cells, and proper concentrations of reagents will be emphasized. The Meta- Core will leverage the expertise and equipment of other Cores at Washington University to produce complementary services for a reduced rate.

项目摘要/摘要：代谢核心 代谢核心（元核）是华盛顿大学慢性的生物医学资源核心 肾脏疾病国家资源中心。长期受伤的孩子改变了新陈代谢，最近 数据表明，这些代谢变化在肾脏功能的逐步下降中起着致病作用。 审问代谢的能力对于理解慢性肾脏病病理生理学至关重要，但是 肾脏研究人员有许多障碍。首先，许多对慢性感兴​​趣的调查员 肾脏疾病在代谢和代谢测定方面没有专业知识。其次，其中许多测定法 需要许多实验室不容易获得昂贵的设备。为了解决这些问题，元核 由M.D. Leslie Gewin的教师，Brian Finck博士核心主任，Gary Patti副总监，Gary Patti， 核心合作者和核心合作者Leah Shriver博士博士将与Meta-进行咨询 指导和设计最佳代谢测定法以回答研究问题的核心用户。会有几种测定 可以向元核用户使用在细胞或整个肾脏水平上询问代谢。这些 测定包括使用原代细胞或新鲜分离的管子的海马生物滤光管分析，均为体内 高度氧化近端管的代表性比市售的原代人体管更具代表性 细胞或有条件永生的小管细胞。我们可以测量脂肪酸或葡萄糖/丙酮酸的氧化 肾脏组织使用放射性底物氧化测定法。 3H-正确的测定法，不经常 在肾脏组织上进行的，比更常用的14c-贴木酸盐分析具有优势 通过电子传输链完全氧化长链脂肪酸，并且更容易执行 （收集3H2O而不是14CO2）。我们还可以将用户推荐给营养肥胖研究中心的高级 分辨率呼吸仪（Oroboros oxygraph 2k）检测分离的线粒体中的呼吸。此外， 未靶向的代谢组学和稳定的同位素示踪剂研究将在体外和在原代细胞上进行 体内。元核教师都致力于与O'Brien共享经过验证的协议和方法 财团以及更大的科学界。此外，我们将帮助培训肾脏科学界 如何生成原代近端管细胞和管子，并以一种方式使用Seahorse Bioflux分析 产生一致的，可再现的结果。特别是，验证使用单元格的重要性， 将强调细胞数量的分析校正和适当的试剂浓度。元 - 核心将利用华盛顿大学其他核心的专业知识和设备来生产 完全服务降低的速度。