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Caveolin-1与葡萄糖转运蛋白4共同参与PC12细胞缺糖应激调节

张骐麒, 黄亮, 韩朝, 关心, 王亚君, 刘晶, 万京华, 邹伟

辽宁师范大学生命科学学院，大连 116081；辽宁省生物技术与分子药物研发重点实验室，大连 116029；大连医科大学附属第一医院，中英再生医学中心，大连 116011；大连市第五人民医院神经内科，大连 116029

摘要

近年研究表明，作为构成胞膜窖(Caveolae)主要的组分之一，窖蛋白-1 (caveolin-1, Cav-1)除了在细胞胆固醇平衡、信号转导和整合以及细胞生长等过程中起重要作用外，还参与细胞营养改变的神经元代谢调节过程。本文旨在探讨 Cav-1和葡萄糖转运蛋白4 (glucose transporter 4, GLUT4)在神经细胞内营养环境改变时的功能变化和关系。采用Western blot和激光共聚焦法观察了两种蛋白在PC12细胞葡萄糖剥夺(glucose deprivation, GD)前后的表达水平与分布，发现GD 6 h后能诱导PC12细胞内Cav-1和GLUT4蛋白表达水平增加，CCK检测和流式细胞术结果显示细胞活力下降、细胞内钙离子浓度([Ca2+]i)升高、线粒体膜电位(mitochondrial membrane potential, MMP)下降。采用siRNA技术敲低Cav-1基因后，GD组PC12细胞死亡率和[Ca2+]i进一步升高，MMP进一步下降；Cav-1敲低细胞系和甲基化β环化糊精(methylated-β-Cyclodextrin, M-β-CD)法处理实验组中，Cav-1和GLUT4蛋白表达均下降。此外还发现，GD可促进GLUT4从细胞质转位到细胞膜。结果提示，Cav-1可能通过调节GLUT4在GD情况下发挥神经保护作用。

关键词： 窖蛋白1; 葡萄糖转运蛋白4; 葡萄糖营养剥夺; PC12细胞

Caveolin-1 and glucose transporter 4 involved in the regulation of glucose-deprivation stress in PC12 cells

ZHANG Qi-Qi, HUANG Liang, HAN Chao, GUAN Xin, WANG Ya-Jun, LIU Jing, WAN Jing-Hua, ZOU Wei

College of Life Science, Liaoning Normal University, Dalian 116081, China； Liaoning Key Laboratory of Biotechnology and Molecular Medicine Development, Dalian 116029, China； Regenerative Medicine Centre, First Affiliated Hospital of Dalian Medical University, Dalian 116011, China； Department of Neurology, the Fifth People’s Hospital of Dalian, Dalian 116029, China

Abstract

Recent evidence suggests that caveolin-1 (Cav-1), the major protein constituent of caveolae, plays a prominent role in neuronal nutritional availability with cellular fate regulation besides in several cellular processes such as cholesterol homeostasis, regulation of signal transduction, integrin signaling and cell growth. Here, we aimed to investigate the function of Cav-1 and glucose transporter 4 (GLUT4) upon glucose deprivation (GD) in PC12 cells. The results demonstrated firstly that both Cav-1 and GLUT4 were up-regulated by glucose withdrawal in PC12 cells by using Western blot and laser confocal technology. Also, we found that the cell death rate, mitochondrial membrane potential (MMP) and intracellular free Ca2+ concentration ([Ca2+]i) were also respectively changed followed the GD stress tested by CCK8 and flow cytometry. After knocking down of Cav-1 in the cells by siRNA, the level of [Ca2+]i was increased, and MMP was reduced further in GD-treated PC12 cells. Knockdown of Cav-1 or methylated-β-Cyclodextrin (M-β-CD) treatment inhibited the expression of GLUT4 protein upon GD. Additionally, we found that GLUT4 could translocate from cytoplasm to cell membrane upon GD. These findings might suggest a neuroprotective role for Cav-1, through coordination of GLUT4 in GD.

Key words: caveolin-1; glucose transporter 4; glucose deprivation; PC12 cell

收稿日期：2014-11-17　　录用日期：2015-02-04

通讯作者：邹伟　　E-mail: weizou60@126.com

引用本文：

张骐麒, 黄亮, 韩朝, 关心, 王亚君, 刘晶, 万京华, 邹伟. Caveolin-1与葡萄糖转运蛋白4共同参与PC12细胞缺糖应激调节[J]. 生理学报 2015; 67 (4): 349-356.

ZHANG Qi-Qi, HUANG Liang, HAN Chao, GUAN Xin, WANG Ya-Jun, LIU Jing, WAN Jing-Hua, ZOU Wei. Caveolin-1 and glucose transporter 4 involved in the regulation of glucose-deprivation stress in PC12 cells. Acta Physiol Sin 2015; 67 (4): 349-356 (in Chinese with English abstract).