Acute Effects of Sugars and Artificial Sweeteners on Small Intestinal Sugar Transport: A Study Using CaCo-2 Cells As an In Vitro Model of the Human Enterocyte

BackgroundThe gastrointestinal tract is responsible for the assimilation of nutrients and plays a key role in the regulation of nutrient metabolism and energy balance. The molecular mechanisms by which intestinal sugar transport are regulated are controversial. Based on rodent studies, two models currently exist that involve activation of the sweet-taste receptor, T1R2/3: an indirect model, whereby luminal carbohydrates activate T1R2/3 expressed on enteroendocrine cells, resulting in the release of gut peptides which in turn regulate enterocyte sugar transport capacity; and a direct model, whereby T1R2/3 expressed on the enterocyte regulates enterocyte function.AimsTo study the direct model of intestinal sugar transport using CaCo-2 cells, a well-established in vitro model of the human enterocyte.MethodsUptake of 10mM 14C D-Glucose and D-Fructose into confluent CaCo-2/TC7 cells was assessed following 3hr preincubation with sugars and artificial sweeteners in the presence and absence of the sweet taste receptor inhibitor, lactisole. Expression of the intestinal sugar transporters and sweet-taste receptors were also determined by RT-PCR.ResultsIn response to short term changes in extracellular glucose and glucose/fructose concentrations (2.5mM to 75mM) carrier-mediated sugar uptake mediated by SGLT1 and/or the facilitative hexose transporters (GLUT1,2,3 and 5) was increased. Lactisole and artificial sweeteners had no effect on sugar transport regulated by glucose alone; however, lactisole increased glucose transport in cells exposed to glucose/fructose. RT-PCR revealed Tas1r3 and SGLT3 gene expression in CaCo-2/TC7 cells, but not Tas1r2.ConclusionsIn the short term, enterocyte sugar transport activities respond directly to extracellular glucose levels, but not fructose or artificial sweeteners. We found no evidence of a functional heterodimeric sweet taste receptor, T1R2/3 in CaCo-2 cells. However, when glucose/fructose is administered together there is an inhibitory effect on glucose transport possibly mediated by T1R3.研究背景 胃肠道负责营养物质的同化吸收，并在营养代谢与能量平衡的调控中发挥关键作用。肠道糖转运的调控分子机制目前仍存在争议。基于啮齿类动物研究，当前存在两种涉及甜味受体（sweet-taste receptor）T1R2/3激活的调控模型：其一为间接模型，即肠腔碳水化合物激活肠内分泌细胞上表达的T1R2/3，进而触发肠道肽类激素释放，最终调控肠上皮细胞的糖转运能力；其二为直接模型，即肠上皮细胞自身表达的T1R2/3直接调控肠上皮细胞功能。 研究目的 本研究采用已被广泛验证的人肠上皮细胞体外模型CaCo-2细胞，探究肠道糖转运的直接调控模型。 实验方法 分别在添加与不添加甜味受体抑制剂拉克素（lactisole）的条件下，将汇合态CaCo-2/TC7细胞与糖类、人工甜味剂共孵育3小时，随后检测细胞对10mM ¹⁴C标记D-葡萄糖与D-果糖的摄取量。同时采用逆转录聚合酶链式反应（RT-PCR）检测肠道糖转运蛋白与甜味受体的基因表达水平。 实验结果 当细胞外葡萄糖及葡萄糖/果糖浓度在2.5mM至75mM范围内发生短期变化时，由SGLT1以及/或易化己糖转运蛋白（GLUT1、2、3、5）介导的载体依赖性糖摄取能力显著提升。仅由葡萄糖单独调控的糖转运过程不受拉克素与人工甜味剂的影响；但当细胞暴露于葡萄糖/果糖混合体系时，拉克素可增强葡萄糖转运能力。RT-PCR检测结果显示，CaCo-2/TC7细胞中存在Tas1r3与SGLT3基因的表达，但未检测到Tas1r2的转录产物。 研究结论 短期来看，肠上皮细胞的糖转运活性可直接响应细胞外葡萄糖水平，但不受果糖或人工甜味剂的调控。本研究未在CaCo-2细胞中检测到具有功能活性的异二聚体甜味受体T1R2/3。但当葡萄糖与果糖共同存在时，可对葡萄糖转运产生抑制作用，该调控过程可能由T1R3介导。