Regulation of Intestinal Na Absorption

描述（由申请人提供）：在哺乳动物小肠中，钠主要通过绒毛细胞刷状边缘膜上的NaCl吸收和Na-葡萄糖共转运（SGLT-1）耦合吸收。耦合NaCl吸收是通过Na:H（特别是NHE3）和Cl:HCO3交换的双重作用发生的。作为最具生物活性的分子之一，一氧化氮（NO）在正常肠道中对NHE3和SGLT-1的调控尚不清楚。我们证明，无论是在兔体内还是在体外对大鼠肠上皮细胞的抑制，NO的抑制都能抑制绒毛细胞BBM SGLT-1并刺激NHE3。抑制SGLT-1的机制是通过改变该蛋白的糖基化来降低其对葡萄糖的亲和力。相比之下，NHE3通过BBM NHE3转录和合成的增加而继发于BBM转运体数量的增加。已经证明抑制NO可以调节NHE3和SGLT-1，下一个合乎逻辑的问题是在NO的生理增加过程中会发生什么。事实上，迄今为止的结果表明，BBM NHE3和SGLT-1可能受到NO的代偿调节，以维持细胞Na稳态。这些观察结果让我们提出了一个最新奇的问题——这两种BBM转运蛋白能直接相互调节吗？这种现象在肠运输生理学中还没有描述。在此背景下，本研究的总体假设是肠细胞BBM中的两种主要钠吸收途径通过NO和/或直接相互调节以维持细胞内钠稳态。因此，本提案的总体目的是确定NO和/或直接调节肠上皮细胞BBM NHE3和SGLT-1的代偿互惠机制。具体地说，使用体内和体外模型的结合，采用互补的生理和分子技术，适当的药理学药物和特定的分子试剂，我们将：阐明增强NO介导的BBM NHE3和SGLT-1和2的调控机制。确定BBM NHE3和SGLT-1相互直接调控的机制。本研究将对肠上皮细胞BBM中的两种主要Na吸收途径是否可以直接相互调节功能以及NO如何介导NHE3和SGLT-1的代偿性相互调节提供新的见解。钠的吸收对维持健康至关重要，它对腹泻和高血压等多种常见疾病至关重要。同样，葡萄糖的吸收是必不可少的，不仅因为碳水化合物是饮食中的主要营养物质，而且因为它在从糖尿病到肥胖的各种常见疾病中都很重要。综上所述，本研究假设的两种主要钠吸收途径的独特而直接的调控可以为制定促进电解质和营养物质吸收的策略奠定基础，在缺乏电解质和营养物质吸收的情况下，在有利的疾病状态下抑制电解质和营养物质吸收。公共卫生相关性：肠道内钠和葡萄糖的同化对保持身体健康至关重要。它们的吸收对许多常见疾病，包括腹泻、糖尿病、肥胖和高血压更为关键。一氧化氮是最具生物活性的分子之一，可能介导钠和葡萄糖的吸收调节。事实上，我们的研究表明，主要负责钠和葡萄糖吸收的转运蛋白（SLGT-1和NHE3）可能直接相互调节。更好地理解这两种主要钠吸收途径的独特和直接调节，可以形成制定策略的基础，在缺乏时促进电解质和营养物质的吸收，在疾病状态下抑制它们，这将是有利的。

DESCRIPTION (provided by applicant): In the mammalian small intestine Na is primarily absorbed by coupled NaCl absorption and Na-glucose co-transport (SGLT-1) on the brush border membrane of villus cells. Coupled NaCl absorption occurs via the dual operation of Na:H (specifically NHE3) and Cl:HCO3 exchange. The regulation of NHE3 and SGLT-1 in the normal intestine by nitric oxide (NO), one of the most biologically active molecules, was unclear. We demonstrated that whether in vivo in rabbits and/or in vitro inhibition in rat intestinal epithelial cells, inhibition of NO inhibited villus cell BBM SGLT-1 and stimulated NHE3. The mechanism of inhibition of SGLT-1 was by reducing its affinity for glucose specifically by altering the glycosylation of the protein. In contrast, NHE3 was stimulated secondary to an increase in BBM transporter numbers by an increase in the transcription and synthesis of BBM NHE3. Having shown that inhibition of NO regulates NHE3 and SGLT-1, the next logical question is what happens during physiological increases in NO. In fact, results to date indicate that BBM NHE3 and SGLT-1 may be compensatorily regulated by NO to maintain cellular Na homeostasis. These observations led us to ask a most novel question -- can these two BBM transport proteins directly regulate one another? A phenomenon here to fore not described in intestinal transport physiology. Given this background, the overall hypothesis of this proposal is that the two primary Na absorptive pathways in the enterocyte BBM regulate one another via NO and/or directly to maintain cellular Na homeostasis. Thus, the overall aim of this proposal is to determine the compensatory reciprocal mechanism of regulation of intestinal epithelial cell BBM NHE3 and SGLT-1 by NO and/or directly. Specifically using a combination of in vivo and in vitro models, employing complementary physiological and molecular techniques with appropriate pharmacological agents and specific molecular reagents we will: 1. Elucidate the enhanced NO mediated mechanism of regulation of BBM NHE3 and SGLT-1 and 2. Determine the mechanism of direct regulation of BBM NHE3 and SGLT-1 by each other. This study will provide novel insight into whether the two primary Na absorptive pathways in the BBM of intestinal epithelial cells can directly regulate the functioning of each other and how NO mediates the compensatory reciprocal regulation of NHE3 and SGLT-1. Na assimilation is essential to maintain health and it is critical for a wide range of common diseases such as diarrhea and hypertension. Similarly, glucose absorption is not only essential because carbohydrates represent the predominant nutrient in the diet, but also because it is important in a variety of common diseases from diabetes to obesity. In conclusion, the unique and direct regulation of the two primary Na absorptive pathways as hypothesized in this proposal could form the basis for developing strategies to promote electrolyte and nutrient absorption where deficient and inhibit the same in disease states where it would be advantageous. PUBLIC HEALTH RELEVANCE: Sodium and glucose assimilation in the intestine is essential to maintain good health. Their absorption is even more critical in numerous common diseases including diarrhea, diabetes, obesity and hypertension. Nitric oxide, one of the most biologically active molecules, may mediate the regulation of the absorption of sodium and glucose. In fact, our studies show that transporters primarily responsible for sodium and glucose absorption (SLGT-1 and NHE3) may directly regulate one another. Better understanding of the unique and direct regulation of these two primary Na absorptive pathways as hypothesized in this proposal could form the basis for developing strategies to promote electrolyte and nutrient absorption where deficient and inhibit the same in disease states where it would be advantageous.