Department of Molecular Genetics, Maastricht University, Maastricht, Netherlands.
Am J Physiol Cell Physiol. 2010 Jun;298(6):C1549-59. doi: 10.1152/ajpcell.00334.2009. Epub 2010 Apr 7.
Insulin and contraction stimulate both cardiac glucose and long-chain fatty acid (LCFA) uptake via translocation of the substrate transporters GLUT4 and CD36, respectively, from intracellular compartments to the sarcolemma. Little is known about the role of vesicular trafficking elements in insulin- and contraction-stimulated glucose and LCFA uptake in the heart, especially whether certain trafficking elements are specifically involved in GLUT4 versus CD36 translocation. Therefore, we studied the role of coat proteins, actin- and microtubule-filaments and endosomal pH on glucose and LCFA uptake into primary cardiomyocytes under basal conditions and during stimulation with insulin or oligomycin (contraction-like AMP-activated protein kinase activator). Inhibition of coat protein targeting to Golgi/endosomes decreased insulin/oligomycin-stimulated glucose (-42%/-51%) and LCFA (-39%/-68%) uptake. Actin disruption decreased insulin/oligomycin-stimulated glucose uptake (-41%/-75%), while not affecting LCFA uptake. Microtubule disruption did not affect substrate uptake under any condition. Endosomal alkalinization increased basal sarcolemmal CD36 (2-fold), but not GLUT4, content, and concomitantly decreased basal intracellular membrane GLUT4 and CD36 content (-60% and -62%, respectively), indicating successful CD36 translocation and incomplete GLUT4 translocation. Additionally, endosomal alkalinization elevated basal LCFA uptake (1.4-fold) in a nonadditive manner to insulin/oligomycin, and decreased insulin/oligomycin-stimulated glucose uptake (-32%/-68%). In conclusion, 1) CD36 translocation, just like GLUT4 translocation, is a vesicle-mediated process depending on coat proteins, and 2) GLUT4 and CD36 trafficking are differentially dependent on endosomal pH and actin filaments. The latter conclusion suggests novel strategies to alter cardiac substrate preference as part of metabolic modulation therapy.
胰岛素和收缩通过分别将底物转运体 GLUT4 和 CD36 从细胞内隔室易位到肌膜上来刺激心脏中的葡萄糖和长链脂肪酸 (LCFA) 摄取。关于囊泡转运元件在胰岛素和收缩刺激心脏中的葡萄糖和 LCFA 摄取中的作用知之甚少，特别是某些转运元件是否专门参与 GLUT4 与 CD36 的易位。因此，我们研究了在基础条件下以及在胰岛素或寡霉素（收缩样 AMP 激活蛋白激酶激活剂）刺激下，网格蛋白/内体靶向蛋白、肌动蛋白和微管丝以及内体 pH 对原代心肌细胞中葡萄糖和 LCFA 摄取的作用。抑制网格蛋白向高尔基体/内体的靶向作用可降低胰岛素/寡霉素刺激的葡萄糖（-42%/-51%）和 LCFA（-39%/-68%）摄取。肌动蛋白破坏降低了胰岛素/寡霉素刺激的葡萄糖摄取（-41%/-75%），而不影响 LCFA 摄取。微管丝破坏在任何情况下均不影响底物摄取。内体碱化增加了基础肌膜 CD36（2 倍），但不增加 GLUT4，同时还减少了基础细胞内膜 GLUT4 和 CD36 含量（分别为-60%和-62%），表明 CD36 易位成功，而 GLUT4 易位不完全。此外，内体碱化以非累加的方式升高了基础 LCFA 摄取（1.4 倍），超过了胰岛素/寡霉素的作用，并降低了胰岛素/寡霉素刺激的葡萄糖摄取（-32%/-68%）。总之，1）CD36 易位与 GLUT4 易位一样，是一种依赖网格蛋白的囊泡介导过程，2）GLUT4 和 CD36 的转运对内体 pH 和肌动蛋白丝的依赖性不同。后一结论提示了改变心脏底物偏好的新策略，作为代谢调节治疗的一部分。
