SALT TRANSPORT IN SURGICAL DIARRHEAL DISEASE

上皮氯（C1-）分泌调节缺陷是 一些重要疾病的临床表现， 从囊性纤维化到分泌性腹泻， 肠毒性感染或肠道炎症。虽然数量可观， 已经花费了大量的努力来确定顶膜C1的作用， 在这一过程中的渠道，相对知之甚少 基底外侧膜事件。 这背后的核心假设 研究表明，基底外侧膜Na +-K +-2C1-协同转运蛋白 （NKCC1）是一个重要的独立调控位点，控制着净C1- 分泌能力 越来越多的证据表明，NKCC 1是 调节在三个水平：1）磷酸化依赖和- 协同转运蛋白功能的独立变化，2）通过改变细胞 表面表达，和3）基因表达的长期变化。 该项目解决了各级监管的基本问题 并利用培养的人肠上皮细胞系作为模型， 系统. 细胞内C1-对NKCC1的抑制作用 活动将被定义，和F的潜在监管作用- 肌动蛋白细胞骨架和酪氨酸激酶信号传导的NKCC1将是 评估。 蛋白激酶C对F-肌动蛋白和基底外侧核的影响 膜再循环将被探索为改变的可能机制。 响应佛波醇酯的NKCC1表面表达。 最后，长- NKCC1基因表达和C1分泌的长期变化 将定义炎症、渗透压和代谢应激。 的 建议的研究应该提高对C1分泌的理解， 胃肠道，气道和外分泌器官，通过阐明基本 NKCC1的调节机制。NKCC1在大多数细胞中表达， 被认为在控制细胞中起着普遍管家作用 体积除了其在支持离子矢量传输中的作用之外 穿过极化的上皮细胞 长期目标是建立一个 设计新的治疗方法的合理基础， 电解质运输 此外，这些研究可能具有重要的 对这种普遍存在的膜转运调节的影响 在健康和疾病期间哺乳动物细胞中的蛋白质。

Defective regulation of epithelial chloride (C1-) secretion underlies the clinical manifestations of a number of important diseases ranging from cystic fibrosis to the secretory diarrhea associated with enterotoxic infection or intestinal inflammation. While considerable effort has been expended to define the role of apical membrane C1- channels in this process, comparatively little is known about basolateral membrane events. The central hypothesis underlying this research is that the basolateral membrane Na+-K+-2C1-cotransporter (NKCC1) is a critical independent regulatory site that controls net C1- secretory capacity. Accumulating evidence suggests that NKCC1 is regulated at three levels: 1) by phosphorylation-dependent and - independent changes in cotransporter function, 2) by altered cell surface expression, and 3) by long-term changes in gene expression. This project addresses fundamental questions at each level of regulation and utilizes cultured human intestinal epithelial cell lines as model systems. The inhibitory influence of intracellular C1- on NKCC1 activity will be defined, and the potential regulatory roles of the F- actin cytoskeleton and tyrosine kinase signaling on NKCC1 will be assessed. Effects of protein kinase C on F-actin and basolateral membrane recycling will be explored as a possible mechanism for altered NKCC1 surface expression in response to phorbol ester. Finally, long- term changes in NKCC1 gene expression and C1- secretion in response to inflammatory, osmotic, and metabolic stress will be defined. The proposed studies should improve understanding of C1- secretion in the gastrointestinal tract, airway, and exocrine organs by elucidating basic mechanisms of regulation of NKCC1. NKCC1 is expressed in most cells and is thought to play a generalized housekeeping role in controlling cell volume in addition to its role in supporting vectorial transport of ions across polarized epithelia. The long-term objective is to establish a rational basis for designing new therapeutic approaches to disorders of electrolyte transport. Moreover, these studies may have important implications for the regulation of this ubiquitous membrane transport protein in mammalian cells during health and disease.