CORE--CELL PHYSIOLOGY RESOURCE

核心--细胞生理资源

• 批准号: 7069753
• 负责人: Phillip Darwin Bell
• 金额: $ 15.72万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-30 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7069753
• 关键词: autosomal recessive trait; biomedical facility; cell biology; cell line; confocal scanning microscopy; disease /disorder model; epithelium; fluorescence; immunofluorescence technique; isolation perfusion; kidney; laboratory mouse; molecular /cellular imaging; pathologic process; physiology; polycystic kidney; renal tubule; tissue /cell culture; tissue /cell preparation; voltage /patch clamp

Genetically engineered mouse models of autosomal recessive polycystic kidney disease (ARPKD) provide model systems to study cyst development in kidney, liver and pancreas. This work has led to the surprising observation that apical cilia play a pivotal role in the pathogenesis of ARPKD. However, there is a lack of information concerning the physiological mechanisms that result in cyst formation. This Cellular Physiology Core will provide a comprehensive facility to perform state-of-the-art physiological experiments in cells and tissues from mouse models of ARPKD. One unique aspect of this Core is the breadth of experimental tools and resources that have been amassed to attack specific problems and questions. The other unique aspect of this Core is the ability to isolate renal tubules and ducts, to produce primary and immortalized cell lines, and to apply experimental procedures and techniques that maximize the physiological information that can be obtained from freshly isolated tissues or cultured cells. The Physiology Core is capable of assessing alterations in the rate of electrolyte and water transport in isolated perfused renal tubules and ductal epithelium and in cultured cells grown on permeable supports. In addition, this Core utilizes advanced fluorescence imaging and multi-photon confocal microscopy to study transporter and channel function, transport rates, and regulation of cytosolic calcium, pH, sodium and chloride. Furthermore, using 2-photon microscopy, this Core now has the ability to visualize and resolve subcellular domains in living cells providing novel and detailed information on ion concentrations in cellular microdomains. For instance, recent work in the Core laboratory has revealed elevated subapical membrane calcium levels in an ARPKD collecting duct cell model. Other capabilities include patch clamp analysis, advanced immunofluorescence techniques, and evaluation of paracrine and autocrine signaling using a biosensor approach. Mice that are generated by the Engineered Mouse Core will be used to obtain fresh tissue and to establish primary and immortalized cell lines. The ability to compare the results obtained in freshly isolated tissues versus primary and immortalized cell lines is one of the major strengths of this Core facility. Overall the Cellular Physiology Core will provide the RPKDCC Investigators with a wide range of tools and expertise to define the physiological mechanisms that are involved in the pathogenesis of ARPKD in renal tubules and epithelial cells.

常染色体隐性多囊肾脏疾病（ARPKD）的基因工程小鼠模型提供了研究肾脏，肝脏和胰腺囊肿发育的模型系统。这项工作导致了令人惊讶的观察，即顶端纤毛在ARPKD的发病机理中起关键作用。但是，缺乏有关导致囊肿形成的生理机制的信息。该细胞生理核心将为ARPKD小鼠模型中的细胞和组织中进行最新的生理实验提供全面的设施。该核心的一个独特方面是实验工具和资源的广度，这些工具和资源已积累以攻击特定问题和问题。该核心的另一个独特方面是能够分离肾小管和导管，产生原发性和永生的细胞系，并采用实验程序和技术，从而最大程度地利用从新鲜分离的组织或培养细胞获得的生理信息。生理核心能够评估电解质和水运输速率的改变 分离的灌注肾小管和导管上皮以及在可渗透支撑物上生长的培养细胞中。此外，该核心利用高级荧光成像和多光子共聚焦显微镜研究转运蛋白和通道功能，传输速率以及胞质钙，pH，pH，钠和氯化物的调节。此外，使用2光子显微镜，该核心现在可以在活细胞中可视化和解析亚细胞结构域，从而提供有关细胞微区离子浓度的新颖和详细信息。例如，核心实验室的最新研究表明，在ARPKD收集管道细胞模型中，亚质膜钙水平升高。其他功能包括斑块夹分析，先进的免疫荧光技术以及使用生物传感器方法评估旁分泌和自分泌信号。由工程小鼠核产生的小鼠将用于获得新鲜组织并建立原发性和永生的细胞系。比较在新鲜分离的组织与原发性和永生细胞系中获得的结果的能力是该核心设施的主要优势之一。总体而言，细胞生理核心将为RPKDCC研究人员提供广泛的工具和专业知识，以定义肾小管和上皮细胞中ARPKD发病机理的生理机制。