Viglino Christelle, Montessuit Christophe
Division of Cardiology, Department of Medical Specialties, University of Geneva School of Medecine, Geneva, Switzerland.
J Cell Biochem. 2017 Apr;118(4):670-677. doi: 10.1002/jcb.25655. Epub 2016 Dec 20.
Stimulation of glucose transport is markedly impaired in cardiomyocytes exposed to free fatty acids (FFA), despite relative preservation of canonical insulin- or metabolic stress signaling. We determined whether Focal Adhesion Kinase (FAK) activity is required for stimulation of glucose transport in cardiomyocytes, and whether FAK downregulation participates in FFA-induced impairment of glucose transport stimulation. Glucose transport, measured in isolated cultured cardiomyocytes, was acutely stimulated either by insulin treatment, or by metabolic inhibition with oligomycin resulting in AMP-activated kinase (AMPK) activation. FAK activity was inhibited pharmacologically by preincubation with PF-573,228 (PF). FAK activity was assessed from its autophosphorylation on residue Y397, and from the phosphorylation of its target paxillin on Y118. Y397 FAK phosphorylation was reduced in cultured cardiomyocytes chronically exposed to FFA. Preincubation with PF prior to determination of glucose transport resulted in a significant reduction of oligomycin-stimulated glucose transport, with a lesser reduction in insulin-stimulated glucose transport. Insulin and AMPK signaling was unaffected by PF preincubation. siRNA-mediated FAK knockdown also resulted in reduced oligomycin-stimulated glucose transport. Chronic treatment of FFA-exposed cardiomyocytes with phenylephrine or a phorbol ester restored FAK activity and improved glucose transport. In conclusion, stimulation of glucose transport in cardiomyocytes requires FAK activity prior to stimulation. The chronic reduction of FAK activity in cardiomyocytes exposed to FFA contributes to the loss of glucose transport responsiveness to insulin or metabolic inhibition. J. Cell. Biochem. 118: 670-677, 2017. © 2016 Wiley Periodicals, Inc.
尽管经典的胰岛素或代谢应激信号相对保持完整，但暴露于游离脂肪酸（FFA）的心肌细胞中葡萄糖转运的刺激明显受损。我们确定了粘着斑激酶（FAK）活性对于心肌细胞中葡萄糖转运刺激是否必需，以及FAK下调是否参与FFA诱导的葡萄糖转运刺激损伤。在分离培养的心肌细胞中测量的葡萄糖转运，可通过胰岛素处理或用寡霉素进行代谢抑制（导致AMP激活的蛋白激酶（AMPK）激活）来急性刺激。通过与PF-573,228（PF）预孵育以药理学方式抑制FAK活性。从其在Y397残基上的自磷酸化以及其靶标桩蛋白在Y118上的磷酸化来评估FAK活性。长期暴露于FFA的培养心肌细胞中Y397 FAK磷酸化减少。在测定葡萄糖转运之前用PF预孵育导致寡霉素刺激的葡萄糖转运显著减少，而胰岛素刺激的葡萄糖转运减少较少。胰岛素和AMPK信号不受PF预孵育的影响。siRNA介导的FAK敲低也导致寡霉素刺激的葡萄糖转运减少。用去氧肾上腺素或佛波酯对暴露于FFA的心肌细胞进行慢性处理可恢复FAK活性并改善葡萄糖转运。总之，心肌细胞中葡萄糖转运的刺激在刺激之前需要FAK活性。暴露于FFA的心肌细胞中FAK活性的慢性降低导致对胰岛素或代谢抑制的葡萄糖转运反应性丧失。《细胞生物化学杂志》118: 670 - 677, 2017。© 2016威利期刊公司
