3H2脱氧D葡萄糖摄取测定方法检测心肌细胞对葡萄糖摄取量.docx

生理学报 Acta Physiologica Sinica, April 25, 2004, 56(2): 224-229 HYPERLINK Received 2003-07-29 Accepted 2003-09-19 This work was supported by the National Basic Research Priorities Programme of Peoples Republic of China (No. G2000056906) and the National Natural Science Foundation of China (No. *Corresponding author. Tel: 010 Fax: 010 E-mail: zhangyy@ 　 ?-肾上腺素受体激动对新生大鼠心肌细胞代谢的影响 闫 洁, 陈 锴, 徐 明, 吕志珍, 韩启德, 张幼怡* 北京大学第三医院血管医学研究所, 分子心血管学教育部重点实验室, 北京 100083 摘 要: 为了观察?-肾上腺素受体 (?-AR) 持续激动对心肌细胞代谢的影响, 本工作以培养的新生大鼠心肌细胞为研究对象, 采用[3H]-亮氨酸 ([3H]-leucine）掺入法和BCA蛋白分析法检测心肌细胞的蛋白合成代谢与蛋白含量, [3H]-2-脱氧-D-葡萄糖摄取测定方法检测心肌细胞对葡萄糖的摄取量, 并采用蛋白免疫印迹杂交方法检测腺苷酸活化蛋白激酶 (adenosine monophosphate activated protein kinase, AMPK) 磷酸化程度。结果显示, ?-AR激动剂异丙肾上腺素 (isoproterenol, ISO) 持续刺激心肌细胞48 h, 心肌细胞[3H]-leucine掺入量和蛋白质含量与对照组相比均无显著差异。用ISO或去甲肾上腺素(?1-AR特异性拮抗剂哌唑嗪存在下)持续激动?-AR 48 h, 心肌细胞的葡萄糖摄取量和AMPK磷酸化均显著高于对照组。上述结果表明, ?-AR持续激动对培养的新生大鼠心肌细胞蛋白质合成及蛋白质含量没有明显的作用, 但可引起葡萄糖摄取量明显增加和AMPK的激活, 提示?-AR可能与心肌肥厚过程中能量代谢状态的变化有关。 关键词: ?-肾上腺素受体; 代谢; 心肌细胞; 腺苷酸活化蛋白激酶 中图分类号: Q463 Effects of ?-adrenoceptor activation on metabolism in neonatal rat cardiomyocytes YAN Jie, CHEN Kai, XU Ming, LU Zhi-Zhen, HAN Qi-De, ZHANG You-Yi* Institute of Vascular medicine, peking university third hospital and Key Laboratory of Molecular Cardiology, Education Ministry, beijing 100083, China Abstract: The aim of the present study was to investigate the effects of ?-adrenergic receptor (?-AR) activation on metabolism in cultured neonatal rat cardiomyocytes. The protein synthesis and total protein content of cardiomyocytes were determined by [3H]-leucine incorporation and BCA protein content assay. Cardiomyocyte glucose uptake was measured by [3H]-2-deoxy-D-glucose uptake analysis. Adenosine monophosphate activated protein kinase (AMPK) phosphorylation was detected by Western blot. The results showed that sustained stimulation with isoproterenol (ISO), a ?-adrenoceptor agonist, had no effect on [3H]-leucine incorporation and