Oxidative stress (glutathionylation) and Na,K-ATPase activity in rat skeletal muscle.
Oxidative stress (glutathionylation) and Na,K-ATPase activity in rat skeletal muscle.
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DOI：
10.1371/journal.pone.0110514
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Juel C
中科院分区：
文献类型：
--
作者：
Juel C
肌肉活动期间骨骼肌膜上离子分布的变化会影响兴奋性并可能损害力量的发展。这些变化被 Na,K-ATP 酶抵消。因此，Na,K-ATP 酶的调节对于骨骼肌功能非常重要。本研究调查了大鼠骨骼肌膜中 Na,K-ATP 酶上氧化应激（谷胱甘肽化）的存在。使用抗谷胱甘肽抗体进行免疫沉淀，随后对 Na,K-ATP 酶蛋白亚基进行免疫检测，结果表明氧化和糖酵解骨骼肌中 α 亚型的谷胱甘肽化率分别为 9.0±1.3% 和 4.1±1.0%。在氧化肌中，20.0±6.1% 的 β1 单位被谷胱甘肽化，而在糖酵解肌中 14.8±2.8% 的 β2 亚基似乎被谷胱甘肽化。用还原剂二硫苏糖醇（DTT，1 mM）处理使糖酵解肌膜的体外最大 Na,K-ATP 酶活性增加 19%（P<0.05）。氧化型谷胱甘肽（GSSG，0-10 mM）可增加抗体检测到的体外谷胱甘肽水平，并以剂量依赖性方式降低体外最大 Na,K-ATP 酶活性，与糖酵解骨骼肌相比，氧化作用更大。本研究证明大鼠骨骼肌 Na,K-ATP 酶的 α 和 β 单位均存在基础谷胱甘肽化。此外，该研究表明谷胱甘肽水平与最大 Na,K-ATP 酶活性之间存在负相关。谷胱甘肽可能有助于骨骼肌中 Na,K-ATP 酶功能的复杂调节。特别是，氧化应激诱导的谷胱甘肽化可能在长时间肌肉活动期间的 Na,K-ATP 酶调节中发挥作用。
Changes in ion distribution across skeletal muscle membranes during muscle activity affect excitability and may impair force development. These changes are counteracted by the Na,K-ATPase. Regulation of the Na,K-ATPase is therefore important for skeletal muscle function. The present study investigated the presence of oxidative stress (glutathionylation) on the Na,K-ATPase in rat skeletal muscle membranes. Immunoprecipitation with an anti-glutathione antibody and subsequent immunodetection of Na,K-ATPase protein subunits demonstrated 9.0±1.3% and 4.1±1.0% glutathionylation of the α isoforms in oxidative and glycolytic skeletal muscle, respectively. In oxidative muscle, 20.0±6.1% of the β1 units were glutathionylated, whereas 14.8±2.8% of the β2-subunits appear to be glutathionylated in glycolytic muscle. Treatment with the reducing agent dithiothreitol (DTT, 1 mM) increased the in vitro maximal Na,K-ATPase activity by 19% (P<0.05) in membranes from glycolytic muscle. Oxidized glutathione (GSSG, 0–10 mM) increased the in vitro glutathionylation level detected with antibodies, and decreased the in vitro maximal Na,K-ATPase activity in a dose-dependent manner, and with a larger effect in oxidative compared to glycolytic skeletal muscle. This study demonstrates the existence of basal glutathionylation of both the α and the β units of rat skeletal muscle Na,K-ATPase. In addition, the study suggests a negative correlation between glutathionylation levels and maximal Na,K-ATPase activity. Glutathionylation likely contributes to the complex regulation of Na,K-ATPase function in skeletal muscle. Especially, glutathionylation induced by oxidative stress may have a role in Na,K-ATPase regulation during prolonged muscle activity.
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