INTESTINAL SECRETION & INFLAMMATION - IMPACT OF AMMONIA

肠道分泌

• 批准号: 6381296
• 负责人: JEFFREY B. MATTHEWS
• 金额: $ 30.6万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6381296
• 关键词: actins; adenosinetriphosphatase; ammonia; ammonium chloride; calcium; chloride channels; chlorine; colitis; colon; cyclic AMP; cystic fibrosis; electrophysiology; gastrointestinal epithelium; human tissue; intestines; ion transport; membrane lipids; membrane transport proteins; omega 3 fatty acid; potassium; potassium channel; secretion; sodium; tissue /cell culture; voltage /patch clamp

Numerous diseases affecting the GI tract, ranging from secretory diarrhea to cystic fibrosis, are characterized by dysregulation of epithelial C1- secretion. This project originally identified that ammonium ion (NH4+, normally present at high concentrations in the colonic lumen) may be a novel endogenous regulator of C1- secretion via effects via effects on K+ channels and begins to define the interaction of NH4+ with the basolateral membrane K+ transporters also required for Cl-secretion. Based on work already accomplished, the current application considers how altered K+ channel regulation may influence various intestinal disease states. Preliminary data indicate that the ammonia-derived oxidant monochloramine (NH2Cl) may contribute to the diarrhea of colitis by potentiating Ca2+- dependent K+ channels. Experiments also suggest that docosahexaenoic acid (DHA, a component of fish oil) can augment Ca+2- dependent K+ channels, finding of particular interest as DHA begins clinical evaluation as therapy in CF. Preliminary findings suggest that the actin cytoskeleton an functionally alter Ca2+-dependent K+ channels, and conversely, that these K+ channels can modulate cell functions such as epithelial restitution that involve actin remodeling. Three sets of studies are proposed. First, the impact of ammonia on colonic epithelial transport will be further characterized in cultured epithelial ells and in human colonic mucosal preparations, with attention to the interaction of NH4+ with the basolateral Na+-k+-2Cl- co- transporter, Na+_K+ ATPase, and K+ channels. Second, potentiation of basolateral Ca+2-dependent K+ channels by cAMP and NH2Cl will be explored using cultured epithelial cells as model systems with the goal of defining a common mechanism for K+ channel potentiation by these seemingly diverse stimuli. The potential for therapeutic modulation of basolateral K + channels will be explored, specifically examining wheth4r docosahexaenoic acid (DHA) can augment Ca2+-dependent Cl- secretion in T84 cells and human colon, and, if so, to determine its mechanism of action. Finally, the studies will define the effect of chemical manipulation of F-actin on Ca2+-dependent K+ channel regulation and extend preliminary findings suggesting that K+ channel regulation affects the actin-regulated process of epithelial restitution. These studies highlight the importance of basolateral K+ channels in the regulation of secretion and other epithelial functions and reinforce their potential as targets for new drug design.

从分泌性腹泻到囊性纤维化的许多影响胃肠道的疾病的特征是上皮C1-分泌失调。该项目最初确定铵离子（通常以高浓度为limen的NH4+）可能是通过对K+通道的影响通过效应进行效应的新型内源性调节剂，并且开始定义NH4+与基底膜K+转运蛋白的相互作用，也需要进行cl-固定。基于已经完成的工作，当前的申请考虑了K+通道调节的改变可能如何影响各种肠道疾病状态。初步数据表明，氨衍生的氧化剂单氯胺（NH2CL）可能通过增强Ca2+依赖的K+通道来导致结肠炎的腹泻。实验还表明，二十六烯酸（DHA，鱼油的一个组成部分）可以增加Ca+ 2-依赖的K+通道，因为DHA开始临床评估作为CF的治疗。初步发现表明，肌动蛋白细胞骨架在功能上改变了Ca2+依赖性的K+通道，相反，这些K+通道可以调节涉及肌动蛋白重塑的上皮恢复等细胞功能。提出了三组研究。首先，将在培养的上皮ELL和人类结肠粘膜制剂中进一步表征氨对上皮转运的影响，并注意Nh4+与基底外侧Na+ -K+ -k+ -k+ -2cl- cl- cl-co- transporter，Na+ _K+ _K+ Atpase和K+ Channels的相互作用。其次，将使用培养的上皮细胞作为模型系统来探索基底外侧Ca+ 2依赖性的K+通道，以定义这些看似多样化的刺激的共同机制。将探索基底外侧K +通道的治疗性调节潜力，特别检查WHETH4R DOCOSAHEXAENOIC（DHA）可以增强T84细胞和人类结肠中Ca2 +依赖性的CL-分泌，如果是这样，则可以确定其作用机制。最后，研究将定义F-肌动蛋白化学操纵对Ca2+依赖性K+通道调节的影响，并扩展初步发现，表明K+通道调节会影响肌动蛋白调节的上皮恢复过程。这些研究强调了基底外侧K+通道对分泌和其他上皮功能的调节的重要性，并增强了它们作为新药设计目标的潜力。