Physiology Core

生理学核心

• 批准号: 9983064
• 负责人: Marcelo Daniel Carattino
• 金额: $ 14.52万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 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/9983064
• 关键词: Address; Animal Model; Animals; Biochemical; Biological Assay; Biological Models; Biological Process; Carrier Proteins; Cell Line; Cell Separation; Cells; Characteristics; Collaborations; Complement; Data; Development; Disease; Electric Capacitance; Electrophysiology (science); Enzymes; Epithelial; Epithelial Cells; Epithelium; Equipment; Fluorescence; Generations; Genetic Transcription; Goals; Health; Imaging technology; Immunoblotting; In Vitro; Individual; Instruction; International; Ions; Kidney; Libraries; Light; Measurement; Mediating; Membrane; Modeling; Molecular; Mutate; Nephrons; Organ; Phenotype; Phosphorylation; Physiology; Post Translational Modification Analysis; Post-Translational Protein Processing; Proteins; RNA analysis; Regulation; Regulatory Pathway; Renal function; Research; Research Personnel; Resources; Reverse Transcriptase Polymerase Chain Reaction; Series; System; Technical Expertise; Techniques; Technology; Therapeutic; Tissues; Training; Transcript; Ubiquitination; design; genetic regulatory protein; glycosylation; imaging system; kidney imaging; mutant; palmitoylation; protein expression; single molecule; solute; tool; transcriptome sequencing

Abstract The Physiology Core provides a series of graded in vitro technologies for studying the function and regulation of transport and other membrane resident proteins with progressive degrees of complexity from single molecules to model systems to native epithelia. To gain understanding of biological processes that mediate kidney function in health and disease states, in light of the phenotypic diversity and functional complexity of this organ, the use of strategies that allow specific segment/cell/protein to be studied in isolation under defined conditions is required. The overall goal of this core is to elucidate at a molecular and cellular level the function and regulation of key proteins involved in kidney health (including development) and disease. To accomplish this goal, the Core will provide investigators with: a) microdissected tubules for analysis of RNA expression and/or abundance, protein expression, immunolocalization, and enzyme/transporter microassays, b) functional fluorescence assays of channel/transporter function in individually identified cells in isolated tubules microperfused in vitro, c) measurements of transepithelial ion/solute fluxes across isolated tubules microperfused in vitro, d) electrophysiological assays for functional analysis of transport proteins in heterologous expression systems, either wild-type or mutated, alone or in combination with putative regulatory proteins, e) analyses of transcellular and paracellular transport in model and native epithelia in Ussing chambers, f) technologies for analysis of post-translational modification of proteins including phosphorylation, ubiquitination, palmitoylation and glycosylation, and g) technologies to study composition, dynamics, and regulatory mechanisms through the assessment of transcript abundance in isolated single cells. A central function of the Core is to provide instruction in all the techniques performed by the Core and the use of the Core equipment. The Physiology Core will interact synergistically with the other Cores to: a) extend the studies conducted in either heterologous expression systems or isolated tubules to animals models, b) identify potential therapeutic compounds that target transporters and associated regulatory pathways, and c) perform detailed and quantitative analysis using imaging technologies of the expression of transport and regulatory proteins in heterologous expression systems, model and native epithelia, and isolated tubules.

抽象的 生理核心提供了一系列分级的体外技术，用于研究功能和调节 从单一的运输和其他具有复杂程度的膜驻留蛋白 分子以建模到天然上皮的系统。了解介导的生物过程 肾脏在健康和疾病状态中的功能，鉴于表型多样性和功能复杂性 器官，使用允许在定义下隔离研究特定段/细胞/蛋白质的策略 需要条件。该核心的总体目标是在分子和细胞水平上阐明功能 以及对肾脏健康（包括发育）和疾病的关键蛋白质的调节。完成 该目标，核心将为研究人员提供：a）微解析小管以分析RNA表达 和/或丰度，蛋白质表达，免疫定位和酶/转运蛋白微粒，b）功能 在分离小管中单独鉴定的细胞中通道/转运蛋白功能的荧光测定 在体外微填充，c）测量跨小管的跨度离子/溶质通量的测量 在体外微培养，d）用于运输蛋白功能分析的电生理测定 单独的或突变的异源表达系统，单独或与推定的调节性结合 蛋白质，e）模型中的跨细胞和细胞细胞转运的分析和ussing中的天然上皮 钱伯斯，f）用于​​分析蛋白质翻译后修饰的技术，包括磷酸化， 泛素化，棕榈酰化和糖基化以及G）研究组成，动力学和 通过评估孤立的单个细胞中转录物丰度的评估来调节机制。中央 核心的功能是在核心执行的所有技术和使用中提供指令 核心设备。生理核心将与其他核心协同相互作用至：a）扩展研究 在异源表达系统或针对动物模型的孤立小管中进行的 靶向转运蛋白和相关调节途径的潜在治疗化合物，c）执行 使用传输表达和调节的成像技术的详细和定量分析 异源表达系统，模型和天然上皮和孤立小管中的蛋白质。