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（CF）是白人中最常见的致命遗传性疾病 人口。随着CF患者寿命的延长，肝病已成为第二大疾病 死亡的主要原因。 CF 肝病的发展被认为是 胆管分泌缺陷导致阻塞的结果 顽强的胆汁分泌物会破坏胆管，从而导致局部 门静脉周围胆道纤维化/肝硬化。这个解释除了 最近发现CFTR仅在胆管细胞上表达，而不在胆管细胞上表达 肝细胞，表明研究胆汁分泌对于 了解 CF 的病理生理学并制定治疗策略 肝。通过应用从大鼠身上开发的分离方法，最近，新的 极化隔离胆管装置 (IBDU) 已与正常和 CF 隔离 老鼠。因此，本研究的目的是进一步表征 来自正常和 CF 敲除小鼠的胆管细胞和 IBDU，以表征离子 这些细胞中的转运蛋白，并研究各种转运蛋白的作用和机制 胆汁分泌中包括神经内分泌肽在内的促分泌剂 寻找激活替代的、独立于营地的胆汁分泌的方法 CF 小鼠中的通路。最近成功从正常小鼠中分离出 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 小鼠，但需要进一步表征。最近的实验研究 容量激活的氯离子通道作为胆汁分泌的可能替代方案 CF 胆管细胞中的通路表明，通过低渗挑战后调节体积减少（RVD）评估的细胞体积调节在 CF 中受损 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/底物运输，此 R03 资助 提议检查介导细胞的潜在离子传输机制 音量调节及其改变，并找到可以纠正的代理 CF 胆管细胞的细胞体积调节受损。定量 视频显微镜将用于检查特定抑制剂的效果 参与 RVD 的离子通道和转运蛋白，并筛选潜在药物 纠正 CF 小鼠胆管细胞受损的 RVD。这些有前途的 因此离子通道和转运蛋白的抑制剂和刺激剂 确定的将通过同位素流出研究进一步调查 它们对 Cl- 和 K+ 离子传输的影响，以及膜片钳技术 表征 RVD 中涉及的离子通道和转运蛋白。 了解胆道运输系统及其基本机制 正常小鼠和 CF 小鼠的分泌和细胞体积调节将有助于 制定治疗方法来克服 CFTR 缺陷。这个项目，在 反过来，将为候选人提供一个极好的机会来拓宽和 发展研究和认知技能，成为独立研究员 开展拟议研究项目所需的研究资源，例如 并帮助成功竞争未来的研究资助。