Regulation of Intestinal Na Absorption

描述(申请人提供)：在哺乳动物的小肠中，钠主要通过绒毛细胞刷状缘膜上的氯化钠吸收和钠-葡萄糖共转运耦合(SGLT-1)来吸收。通过Na：H(特别是NHE3)和Cl：HCO3交换的双重作用，发生了耦合的氯化钠吸收。一氧化氮(NO)是最具生物活性的分子之一，其对正常肠道中NHE3和SGLT-1的调节尚不清楚。我们证明，无论是在体内和/或在体外抑制大鼠肠上皮细胞，抑制NO对绒毛细胞BBM SGLT-1和刺激NHE3均有抑制作用。SGLT-1的抑制机制是通过改变蛋白质的糖基化来降低其对葡萄糖的亲和力。相反，BBM NHE3的转录和合成增加导致BBM转运体数量增加，继而刺激NHE3。在证明了抑制NO调节NHE3和SGLT-1之后，下一个合乎逻辑的问题是在NO生理性增加的过程中发生了什么。事实上，到目前为止的研究结果表明，BBM NHE3和SGLT-1可能受到NO的代偿调节，以维持细胞内钠的稳态。这些观察结果让我们提出了一个最新的问题--这两个BBM转运蛋白能否直接相互调节？这是以前在肠道运输生理学中没有描述的现象。在这一背景下，本研究的总体假设是，肠细胞BBM中两条主要的钠吸收途径通过和/或直接调节彼此，以维持细胞内钠的稳态。因此，本方案的总体目标是确定NO和/或直接调节肠上皮细胞BBM NHE3和SGLT-1的代偿相互作用机制。具体地说，我们将采用体内和体外模型相结合的方法，结合合适的药理药物和特定的分子试剂，利用互补的生理和分子技术：1.阐明NO对BBM NHE3和SGLT-1调节的增强机制；2.确定相互直接调节BBM NHE3和SGLT-1的机制。这项研究将为了解肠上皮细胞BBM中两条主要的钠吸收途径是否可以直接调节彼此的功能以及NO如何介导NHE3和SGLT-1的代偿性相互调节提供新的见解。NA同化对保持健康至关重要，对腹泻和高血压等一系列常见疾病至关重要。同样，葡萄糖的吸收也很重要，不仅因为碳水化合物是饮食中的主要营养物质，还因为它在从糖尿病到肥胖症的各种常见疾病中都很重要。总之，这项建议中假设的对两条主要的钠吸收途径的独特和直接的调节可以成为制定策略的基础，以促进缺乏电解质和养分的吸收，并在有利的疾病状态下抑制电解质和养分的吸收。

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.