ACID BASE PERMEABILITY AND TRANSPORT IN COLONIC CRYPTS

该临床研究者奖的候选人Satish Singh博士 完成了胃肠病学的临床培训， 我跟随亨利博士研究了三年的肠道运输生理学 结合剂和细胞内pH值（pHi）的调节与博士沃尔特硼。 在他的研究年里，候选人已经掌握了荧光成像 监测pHi的技术。肾脏灌注技术的改进 他还成功地灌注了单个结肠隐窝。博士 辛格完全致力于学术医学事业。他的长期 目标是成为一名独立的临床医生/科学家，执行基本的 研究与胃肠道疾病相关的领域。 这项研究的重点是结肠的正常pHi生理学， 以及它们如何适应独特的酸碱环境 由细菌发酵的产物产生。结肠腔包含 高浓度的氨短链脂肪酸和二氧化碳这些中性 预期缓冲液容易渗透大多数细胞膜， 对pHi影响很大。因为维持稳定的pH值对正常的 细胞功能和增殖，假设结肠隐窝 细胞具有独特的防御，以维持独特的结肠中的pHi， 环境这一假设将通过微灌注的管腔进行测试， 从兔近端结肠分离的单个隐窝。初步pHi数据 兔子的隐窝表明顶端屏障对NH3是不渗透的， 而基底外侧膜具有正常的NH3渗透性。初步 来自人近端结肠的灌注隐窝的pHi数据表明， 存在Na-H交换剂。第一个目的是（i）确定是否地穴 细胞对亲脂性中性酸碱产物具有顶端屏障 （二）试图突破细菌发酵的障碍， 提供了对其本质的基本见解。第二个目标是（一） 表征调节隐窝细胞pH的酸碱转运蛋白， (ii)探讨隐窝分泌HCO-3进入管腔的机制。 第三个目标是最终将分析扩展到人类隐窝 从结肠镜活检中获得这些。我们的目标是确定 运输工具的动力学特性，在未来， 与结肠病变患者的隐窝相比。因此，在本发明中， 所提议的工作对细胞具有根本重要性 生理学、胃肠道生理学和医学。 辛格博士将继续其职业发展的环境 在细胞酸碱生理学（Boron博士）和 结肠生理学（宾德博士）。两个赞助商都训练成功 伙计们辛格博士的实验活动是基于博士硼的 细胞和分子生理学系的实验室， 在离子通道/转运蛋白和上皮细胞的区域具有很大的深度 physiology.宾德博士将提供细胞的总体指导 生理学和病理生理学的结肠，在临床方面， 人类研究，以及如何最好地发展职业生涯的一个 临床医生/研究者。

The candidate for this Clinical Investigator Award, Dr. Satish Singh, has completed his clinical training in gastroenterology and will have spent three years studying the physiology of intestinal transport with Dr. Henry Binder and the regulation of intracellular pH (pHi) with Dr. Walter Boron. During his research years, the candidate has mastered fluorescence imaging techniques for monitoring pHi. Adapting techniques for perfusing renal tubules, he also has succeeded in perfusing single colonic crypts. Dr. Singh is fully committed to a career in academic medicine. His long-term goal is to become an independent clinician/scientist, performing basic research in areas relevant to gastrointestinal disease. The research proposed focuses on the normal pHi physiology of colonic crypt cells, and how they have adapted to the unique acid-base environment created by products of bacterial fermentation. The colonic lumen contains high levels of ammonia, short-chain fatty acids, and CO2. These neutral buffers would be expected to readily permeate most cell membranes, and greatly affect pHi. Because maintaining a stable pHi is critical to normal cell function and proliferation, it is hypothesized that colonic crypt cells possess unique defenses for maintaining pHi in the unique colonic environment. This hypothesis will be tested by microperfusing the lumen of single crypts isolated from rabbit proximal colon. Preliminary pHi data on rabbit crypts suggest that the apical barrier is impervious to NH3, whereas the basolateral membrane has normal NH3 permeability. Preliminary pHi data on perfused crypts from human proximal colon indicate the presence of a Na-H exchanger. The first aim is to (i) determine if crypt cells possess an apical barrier to lipophilic, neutral acid-base products of bacterial fermentation, and (ii) attempt to breach the barrier to provide fundamental insight into its nature. The second aim, is to (i) characterize the acid-base transporters that regulate crypt cell pH and (ii) explore the mechanism by which crypts secrete HCO-3 into the lumen. The third aim, is to extend the analysis to human crypts, eventually obtaining these from colonoscopic biopsies. The goal is to determine key kinetic properties of the transporters which, in the future, might be compared to those of crypts from patients with colonic pathology. Thus, the proposed work has aspects of fundamental importance to cell physiology, GI physiology, and medicine. The environment in which Dr. Singh will continue his career development has unique strengths in both cellular acid-base physiology (Dr. Boron) and colonic physiology (Dr. Binder). Both sponsors have trained successful fellows. Dr. Singh's experimental activities are based in Dr. Boron's laboratory in the Department of Cellular and Molecular Physiology, which has great depth in the areas of ion channels/transporters and epithelial physiology. Dr. Binder will provide overall guidance on the cell physiology and pathophysiology of the colon, on the clinical aspects of the human studies, and on how to best develop the career of a clinician/investigator.