Hundal H S, Ramlal T, Reyes R, Leiter L A, Klip A
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Endocrinology. 1992 Sep;131(3):1165-73. doi: 10.1210/endo.131.3.1505458.
The effects of the oral hypoglycemic drug metformin on glucose and amino acid transporter activity and subcellular localization of GLUT1 and GLUT4 glucose transporters were tested in cultured L6 myotubes. In muscle cells preexposed to maximal doses of metformin (2 mM, for 16 h), 2-deoxyglucose uptake was stimulated by over 2-fold from 5.9 +/- 0.3 to 13.3 +/- 0.5 pmol/min.mg protein. Uptake of the nonmetabolizable amino acid analog methylaminoisobutyrate was unaffected by treatment with the drug under identical conditions. Extracellular calcium was required to preserve the full response to the biguanide. Exposure of muscle cells to insulin in the presence of metformin resulted in further activation of 2-deoxyglucose transport. The latter effect was additive to the maximum effect of metformin, suggesting that the biguanide stimulates hexose uptake into muscle cells by an insulin-independent mechanism. Glucose transporter number quantified by performing studies of D-glucose-protectable binding of cytochalasin-B in plasma membranes (PM) and internal membranes (IM) prepared from L6 myotubes revealed that a 16-h treatment with 800 microM metformin significantly elevated glucose transporter number in the PM (by 47%), with an equivalent decrement in glucose transporter number (47%) in the IM. Western blot analysis using antisera reactive with the GLUT1 and GLUT4 isoforms of glucose transporters showed that metformin caused a reduction in GLUT1 content in the IM fraction and a concomitant increase in the PM. Unlike insulin, metformin treatment had no effect on the subcellular distribution of GLUT4. We propose that the molecular basis of metformin action in skeletal muscle involves the subcellular redistribution of GLUT1 proteins from an intracellular compartment to the plasma membrane. Such a recruitment process may form an integral part of the mechanism by which the drug stimulates glucose uptake (and utilization) in skeletal muscle and facilitates lowering of blood glucose in the management of type II diabetes.
在培养的L6肌管中测试了口服降糖药二甲双胍对葡萄糖和氨基酸转运体活性以及葡萄糖转运蛋白1（GLUT1）和葡萄糖转运蛋白4（GLUT4）亚细胞定位的影响。在预先暴露于最大剂量二甲双胍（2 mM，16小时）的肌肉细胞中，2-脱氧葡萄糖摄取量从5.9±0.3增加到13.3±0.5 pmol/分钟·毫克蛋白，刺激倍数超过2倍。在相同条件下，该药物处理对不可代谢氨基酸类似物甲基氨基异丁酸的摄取没有影响。需要细胞外钙来维持对双胍类药物的完全反应。在二甲双胍存在下，肌肉细胞暴露于胰岛素会导致2-脱氧葡萄糖转运进一步激活。后一种作用与二甲双胍的最大作用相加，表明双胍类药物通过胰岛素非依赖机制刺激己糖摄取到肌肉细胞中。通过对L6肌管制备的质膜（PM）和内膜（IM）中细胞松弛素B的D-葡萄糖保护结合进行研究来量化葡萄糖转运体数量，结果显示，用800 microM二甲双胍处理16小时可使PM中的葡萄糖转运体数量显著增加（47%），而IM中的葡萄糖转运体数量相应减少（47%）。使用与葡萄糖转运体的GLUT1和GLUT4亚型反应的抗血清进行的蛋白质印迹分析表明，二甲双胍导致IM部分中GLUT1含量降低，同时PM中GLUT1含量增加。与胰岛素不同，二甲双胍处理对GLUT4的亚细胞分布没有影响。我们提出，二甲双胍在骨骼肌中作用的分子基础涉及GLUT1蛋白从细胞内区室向质膜的亚细胞重新分布。这样的募集过程可能是该药物刺激骨骼肌中葡萄糖摄取（和利用）并促进II型糖尿病管理中血糖降低机制的一个组成部分。
