ACID BASE PERMEABILITY AND TRANSPORT IN COLONIC CRYPTS

这一临床研究人员奖的候选人萨蒂什·辛格博士 完成了胃肠病学的临床培训，并将在 跟随亨利博士学习肠道运输生理学三年 粘合剂和沃尔特·博伦博士对细胞内pH(Phi)的调节。 在他多年的研究中，这位候选人掌握了荧光成像技术 监测PHI的技术。适用于肾脏灌流的技术 除了小管，他还成功地灌流了单个结肠隐窝。Dr。 辛格完全致力于学术医学的职业生涯。他的长期 目标是成为一名独立的临床医生/科学家， 在与胃肠道疾病相关的领域进行研究。 提出的研究重点是结肠的正常PHI生理学。 隐窝细胞，以及它们如何适应独特的酸碱环境 由细菌发酵产生的产物。结肠腔内含有 高水平的氨、短链脂肪酸和二氧化碳。这些是中性的 缓冲液有望很容易地渗透到大多数细胞膜上，并且 对Phi有很大影响。因为保持稳定的PHI对于正常是至关重要的 细胞功能和增殖，推测结肠隐窝 在独特的结肠中，细胞具有维持PHI的独特防御 环境。这一假设将通过微灌流的方式进行验证。 从兔近端结肠中分离出单个隐窝。PHI初步数据在 兔子的墓穴表明，根尖屏障不受NH3的影响， 而基侧膜具有正常的NH3通透性。初步 人近端结肠灌注隐窝的PHI数据表明 存在Na-H交换器。第一个目标是(I)确定地窖 细胞对亲脂性、中性酸碱产物具有顶端屏障 细菌发酵，以及(Ii)试图突破障碍 提供对其本质的基本洞察。第二个目标，是(I) 描述调节隐窝细胞pH和酸碱转运体的特征 (Ii)探讨隐窝分泌HCO-3进入管腔的机制。 第三个目标是将分析扩展到人类墓穴，最终 从结肠镜活检中获得这些。目标是确定关键 转运体的动力学性质，在未来可能是 与结肠病理患者的隐窝相比。因此， 拟议的工作对细胞具有基本的重要性。 生理学、胃肠生理学和医学。 辛格博士将继续其职业发展的环境 在细胞酸碱生理学方面有独特的优势(Boron博士)和 结肠生理学(宾德博士)两家赞助商都成功地进行了培训 伙计们。辛格博士的实验活动基于Boron博士的 细胞和分子生理学系实验室， 在离子通道/转运体和上皮细胞领域有很深的研究 生理学。Binder博士将提供关于细胞的总体指导 结肠生理学和病理生理学，关于临床方面的 人类研究，以及如何最好地发展职业生涯 临床医生/研究员。

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.