School of Food Science and Nutrition, University of Leeds, Leeds LS2 9JT, UK.
School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
Biochem Pharmacol. 2018 Jun;152:11-20. doi: 10.1016/j.bcp.2018.03.011. Epub 2018 Mar 14.
Only limited data are available on the inhibition of the sugar transporter GLUT5 by flavonoids or other classes of bioactives. Intestinal GLUT7 is poorly characterised and no information exists concerning its inhibition. We aimed to study the expression of GLUT7 in Caco-2/TC7 intestinal cells, and evaluate inhibition of glucose transport by GLUT2 and GLUT7, and of fructose transport by GLUT2, GLUT5 and GLUT7, by flavonoids. Differentiated Caco-2/TC7 cell monolayers were used to investigate GLUT7 expression, as well as biotinylation and immunofluorescence to assess GLUT7 location. For mechanistic sugar transport studies, X. laevis oocytes were injected with individual mRNA, and GLUT protein expression on oocyte membranes was confirmed. Oocytes were incubated with D-[C(U)]-glucose or D-[C(U)]-fructose in the presence of flavonoids, and uptake was estimated by liquid scintilation counting. In differentiated Caco-2/TC7 cell monolayers, GLUT7 was mostly expressed apically. When applied apically, or to both compartments, sorbitol, galactose, L-glucose or sucrose did not affect GLUT7 mRNA expression. Fructose applied to both sides increased GLUT7 mRNA (13%, p ≤ 0.001) and total GLUT7 protein (2.7-fold, p ≤ 0.05), while the ratio between apical, basolateral and total GLUT7 protein was unchanged. In the X. laevis oocyte model, GLUT2-mediated glucose and fructose transport were inhibited by quercetin, (-)-epigallocatechin gallate (EGCG) and apigenin, GLUT5-mediated fructose transport was inhibited by apigenin and EGCG, but not by quercetin, and GLUT7-mediated uptake of both glucose and fructose was inhibited by apigenin, but not by quercetin nor EGCG. Expression of GLUT7 was increased by fructose, but only when applied to Caco-2/TC7 cells both apically and basolaterally. Since GLUT2, GLUT5 and GLUT7 show different patterns of inhibition by the tested flavonoids, we suggest that they have the potential to be used as investigational tools to distinguish sugar transporter activity in different biological settings.
关于黄酮类化合物或其他类生物活性物质对糖转运蛋白 GLUT5 的抑制作用，仅有有限的数据。肠 GLUT7 的特征描述较差，并且不存在关于其抑制作用的信息。我们旨在研究 Caco-2/TC7 肠细胞中 GLUT7 的表达，并评估 GLUT2 和 GLUT7 对葡萄糖转运的抑制作用，以及 GLUT2、GLUT5 和 GLUT7 对果糖转运的抑制作用，黄酮类化合物。使用分化的 Caco-2/TC7 细胞单层来研究 GLUT7 的表达，并通过生物素化和免疫荧光评估 GLUT7 的位置。对于机制性糖转运研究，用个体 mRNA 注射非洲爪蟾卵母细胞，并确认卵母细胞膜上 GLUT 蛋白的表达。将卵母细胞与黄酮类化合物一起孵育 D-[C(U)]-葡萄糖或 D-[C(U)]-果糖，并通过液体闪烁计数估计摄取量。在分化的 Caco-2/TC7 细胞单层中，GLUT7 主要在上皮表达。当在上皮侧或两侧施加时，山梨醇、半乳糖、L-葡萄糖或蔗糖不会影响 GLUT7 mRNA 的表达。果糖施加于两侧增加 GLUT7 mRNA（13%，p≤0.001）和总 GLUT7 蛋白（2.7 倍，p≤0.05），而上皮、基底外侧和总 GLUT7 蛋白的比例保持不变。在非洲爪蟾卵母细胞模型中，GLUT2 介导的葡萄糖和果糖转运被槲皮素、(-)-表没食子儿茶素没食子酸酯（EGCG）和芹菜素抑制，GLUT5 介导的果糖转运被芹菜素和 EGCG 抑制，但不受槲皮素抑制，而 GLUT7 介导的葡萄糖和果糖摄取均被芹菜素抑制，但不受槲皮素或 EGCG 抑制。果糖增加 GLUT7 的表达，但仅当果糖施加于 Caco-2/TC7 细胞的上皮侧和基底外侧时才会增加。由于测试的黄酮类化合物对 GLUT2、GLUT5 和 GLUT7 的抑制作用表现出不同的模式，因此我们认为它们有可能作为研究工具，用于区分不同生物环境中糖转运体的活性。
