Toyoda Taro, Hayashi Tatsuya, Miyamoto Licht, Yonemitsu Shin, Nakano Masako, Tanaka Satsuki, Ebihara Ken, Masuzaki Hiroaki, Hosoda Kiminori, Inoue Gen, Otaka Akira, Sato Kenji, Fushiki Tohru, Nakao Kazuwa
Dept. of Medicine and Clinical Science, Kyoto Univ. Graduate School of Medicine, 54 Shogoin-Kawaharacho, Sakyo-ku, Kyoto, 606-8507, Japan.
Am J Physiol Endocrinol Metab. 2004 Jul;287(1):E166-73. doi: 10.1152/ajpendo.00487.2003. Epub 2004 Mar 16.
Recent studies have suggested that 5'AMP-activated protein kinase (AMPK) is activated in response to metabolic stresses, such as contraction, hypoxia, and the inhibition of oxidative phosphorylation, which leads to insulin-independent glucose transport in skeletal muscle. In the present study, we hypothesized that acute oxidative stress increases the rate of glucose transport via an AMPK-mediated mechanism. When rat epitrochlearis muscles were isolated and incubated in vitro in Krebs buffer containing the oxidative agent H(2)O(2), AMPKalpha1 activity increased in a time- and dose-dependent manner, whereas AMPKalpha2 activity remained unchanged. The activation of AMPKalpha1 was associated with phosphorylation of AMPK Thr(172), suggesting that an upstream kinase is involved in the activation process. H(2)O(2)-induced AMPKalpha1 activation was blocked in the presence of the antioxidant N-acetyl-l-cysteine (NAC), and H(2)O(2) significantly increased the ratio of oxidized glutathione to glutathione (GSSG/GSH) concentrations, a sensitive marker of oxidative stress. H(2)O(2) did not cause an increase in the conventional parameters of AMPK activation, such as AMP and AMP/ATP. H(2)O(2) increased 3-O-methyl-d-glucose transport, and this increase was partially, but significantly, blocked in the presence of NAC. Results were similar when the muscles were incubated in a superoxide-generating system using hypoxanthine and xanthine oxidase. Taken together, our data suggest that acute oxidative stress activates AMPKalpha1 in skeletal muscle via an AMP-independent mechanism and leads to an increase in the rate of glucose transport, at least in part, via an AMPKalpha1-mediated mechanism.
最近的研究表明，5'-腺苷酸激活蛋白激酶（AMPK）会在诸如收缩、缺氧以及氧化磷酸化抑制等代谢应激反应中被激活，这会导致骨骼肌中出现不依赖胰岛素的葡萄糖转运。在本研究中，我们假设急性氧化应激会通过一种AMPK介导的机制提高葡萄糖转运速率。当分离出大鼠肱三头肌并在含有氧化剂过氧化氢（H₂O₂）的Krebs缓冲液中进行体外孵育时，AMPKα1活性呈时间和剂量依赖性增加，而AMPKα2活性保持不变。AMPKα1的激活与AMPK苏氨酸（Thr）172位点的磷酸化有关，这表明上游激酶参与了激活过程。在抗氧化剂N-乙酰-L-半胱氨酸（NAC）存在的情况下，H₂O₂诱导的AMPKα1激活被阻断，并且H₂O₂显著增加了氧化型谷胱甘肽与谷胱甘肽（GSSG/GSH）浓度的比值，这是氧化应激的一个敏感标志物。H₂O₂并未导致AMPK激活的传统参数如AMP和AMP/ATP增加。H₂O₂增加了3-O-甲基-D-葡萄糖的转运，并且在NAC存在时，这种增加被部分但显著地阻断。当肌肉在使用次黄嘌呤和黄嘌呤氧化酶的超氧化物生成系统中孵育时，结果相似。综上所述，我们的数据表明急性氧化应激通过一种不依赖AMP的机制激活骨骼肌中的AMPKα1，并至少部分地通过一种AMPKα1介导的机制导致葡萄糖转运速率增加。
