Grefner N M, Gromova L V, Gruzdkov A A, Snigirevskaia E S, Komissarchik Ia Iu
Tsitologiia. 2006;48(4):355-63.
To elucidate mechanisms providing transport of sugars across intestinal epithelium, on taking into account the current hypotheses (active transport, participation of paracellular transport and passive component of transcellular transport), it was important to reveal structural changes of tight junctions and distribution of the carriers of facilitated diffusion of GLUT2 and protein kinase C during absorption of glucose. On using confocal and electron microscopy, ultrastructural and immunocytochemical studies of enterocytes after perfusion of isolated rat small intestine fragment with 75 mM glucose (chronic experiment) have shown: 1) fluorescent labels of transporter GLUT2 and PKCbetaII are located in the apical area of enterocytes situated at the upper half of the villus. Antibodies against GLUT2, conjugated with gold, are revealed at the microvilli or apical membrane and in the area of terminal network; 2) no ultrastructural changes of the tight junction are detected on ultrathin sections and freeze--fracture replics. At the same time, fluorescent and gold labels against actin are concentrated in the vicinity of the lateral membrane in the tight junction area. The results obtained can serve a confirmation of a hypothesis that at high glucose concentrations GLUT2 participates in its transfer across the apical membrane.
为阐明糖跨肠上皮细胞转运的机制,考虑到当前的假说(主动转运、细胞旁转运的参与以及跨细胞转运的被动成分),揭示紧密连接的结构变化以及葡萄糖吸收过程中GLUT2易化扩散载体和蛋白激酶C的分布情况至关重要。运用共聚焦显微镜和电子显微镜,对分离的大鼠小肠片段灌注75 mM葡萄糖后(慢性实验)肠上皮细胞进行超微结构和免疫细胞化学研究,结果显示:1)转运蛋白GLUT2和PKCβII的荧光标记位于绒毛上半部分肠上皮细胞的顶端区域。与金结合的抗GLUT2抗体在微绒毛或顶端膜以及终末网区域被发现;2)超薄切片和冷冻断裂复制品上未检测到紧密连接的超微结构变化。同时,针对肌动蛋白的荧光和金标记集中在紧密连接区域侧膜附近。所得结果可证实一种假说,即在高葡萄糖浓度下GLUT2参与其跨顶端膜的转运。