Volume Regulatory Pathways in Cystic Fibrosis Mice

囊性纤维化小鼠的容量调节途径

• 批准号: 6623078
• 负责人: WON KYOO CHO
• 金额: $ 6.3万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-01 至 2003-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6623078
• 关键词: cell morphology; chloride channels; cystic fibrosis; disease /disorder model; drug screening /evaluation; gallbladder; gallbladder disorder; gastrointestinal agents; genetically modified animals; ion transport; laboratory mouse; liver disorder; osmotic pressure; potassium channel; secretion; video microscopy; voltage /patch clamp

DESCRIPTION (provided by applicant): Cystic fibrosis (CF) is the most common lethal inherited diseases in white population. As CF patients live longer, liver disease has become the second leading cause of death. The development of CF liver disease is believed to result from the secretory defects in the bile ducts leading to the obstructions of bile ductules by tenacious bile secretions, thereby resulting in focal periportal biliary fibrosis/cirrhosis. This explanation in addition to the recent finding that CFTR is only expressed on bile duct cells, but not on hepatocytes, suggest that studying biliary secretion is crucial to understanding the pathophysiology and developing therapeutic strategies for CF liver. By applying the isolation methods developed from rat, recently, novel polarized isolated bile duct unit (IBDU)s have been isolated from normal and CF mice. Therefore, the aims of this research are to further characterize cholangiocytes and IBDU from normal and CF knockout mice, to characterize ion transporters in these cells, and to study the actions and mechanisms of various secretagogues including neuroendocrine peptides in biliary secretion in order to find ways to activate alternative, camp-independent biliary secretory pathways in CF mice. Recent success to isolate IBDU from normal mouse yielded intact polarized functional IBDU that responds to secretin, vasoactive intestinal peptide, and DBcAMP/IBMX. Similar IBDUs were also isolated from CF mice but need further characterization. Recent experiments to study the volume-activated chloride channel as a possible alternative biliary secretory pathway in CF cholangiocytes indicate that cell volume regulation, assessed by regulatory volume decrease (RVD) after hypotonic challenge, is impaired in CF cholangiocytes. Since cell volume regulation is thought to be vital for many cell functions including secretion and ion/substrate transport, this R03 grant is proposing to examine underlying ion transport mechanisms mediating cell volume regulation and their alterations, and to find agent(s) which can correct the impaired cell volume regulation in CF cholangiocytes. Quantitative videomicroscopy will be used to examine the effect of specific inhibitors to ion channels and transporters involved in RVD, and to screen potential agents to correct the impaired RVD of CF mouse cholangiocytes. These promising inhibitors and stimulating agents to ion channels and transporters thus identified will be further investigated by isotope efflux study to examine their effect on Cl-and K+ ion transport, and patch clamping techniques to characterize the ion channels and transporters involved in the RVD. Understanding transport systems and their underlying mechanisms of biliary secretion and cell volume regulation in normal and CF mice will help to formulate therapeutic approaches to overcome the CFTR defect. This project, in turn, will provide the candidate with an excellent opportunity to broaden and develop research and cognitive skills to become independent researcher, the necessary research resources to carry out the proposed research projects, as well as help successfully compete for future research grants.

描述(由申请人提供)： 囊性纤维化是白人中最常见的致死性遗传病。 人口。随着慢性肝病患者寿命的延长，肝病已成为第二大疾病 头号死因。CF肝病的发展被认为是 由导致梗阻的胆管分泌缺陷所致 通过顽固的胆汁分泌物破坏胆小管，从而导致局灶性 门静脉周围胆管纤维化/肝硬变。这一解释除了 最近发现CFTR只在胆管细胞上表达，而在胆管上不表达 肝细胞，表明研究胆汁分泌对 了解CF的病理生理机制并制定治疗策略 肝脏。通过应用从老鼠身上开发的分离方法，最近，新的 极化隔离胆管单位S从正常和CF中分离出来 老鼠。因此，这项研究的目的是进一步表征 正常和CF基因敲除小鼠的胆管细胞和IBDU的离子特性 这些细胞中的转运蛋白，并研究各种不同的作用和机制 胆汁分泌物中包括神经内分泌肽在内的促分泌剂 寻找激活替代的、不依赖于cAMP的胆汁分泌的方法 在CF小鼠中的通路。最近从正常小鼠中成功分离出IBDU 完整的极化功能性IBDU，对促胰液素、血管活性有反应 肠肽和DBcAMP/IBMX。类似的IBDU也从CF中分离出来 老鼠，但还需要进一步的定性。最新的实验研究 容量激活氯通道可能是一种替代的胆汁分泌 低张激发后的调节性体积减少(RVD)反映了CF胆管细胞的容量调节功能受损 胆管细胞。由于细胞体积调节被认为对许多人来说至关重要 细胞功能，包括分泌和离子/底物转运，此R03授权 正在提议研究介导细胞的潜在离子传输机制 音量调节及其变化，并寻找能纠正的代理人(S) CF型胆管细胞体积调节功能受损。量化 将使用视频显微镜来检查特定抑制剂对 参与RVD的离子通道和转运体，以及筛选潜在的药物 纠正CF型小鼠胆管细胞RVD受损。这些前景看好 离子通道和转运体的抑制剂和刺激剂 将通过同位素外流研究进行进一步调查以检查 它们对Cl-和K+离子转运的影响以及膜片钳技术 描述RVD中涉及的离子通道和转运体。 了解胆道转运系统及其潜在机制 正常和CF小鼠的分泌和细胞体积调节将有助于 制定克服CFTR缺陷的治疗方法。这个项目，在 反过来，将为候选人提供一个极好的机会来扩大和 发展研究和认知技能，成为独立的研究人员， 开展拟议研究项目所需的研究资源，如 以及帮助成功竞争未来的研究拨款。