Chen Chiao-nan, Ferrington Deborah A, Thompson LaDora V
Department of Physical Medicine and Rehabilitation, University of Minnesota, Minneapolis, MN 55455, USA.
J Appl Physiol (1985). 2008 Nov;105(5):1554-61. doi: 10.1152/japplphysiol.90680.2008. Epub 2008 Aug 28.
The identities of proteins that show disuse-related changes in the content of oxidative modification are unknown. Furthermore, it is unknown whether the global accumulation of oxidized proteins is greater in aged animals with muscle disuse. The purposes of this study are 1) to identify the exact proteins that show disuse-related changes in oxidation levels and 2) to test the hypothesis that the global accumulation of oxidized proteins with muscle disuse would be greater in aged animals. Adult and old rats were randomized into four groups: weight bearing and 3, 7, or 14 days of hindlimb unloading. Soleus muscles were harvested to investigate the protein oxidation with unloading. Slot blot, SDS-PAGE, and Western blot analyses were used to detect the accumulation of 4-hydroxy-2-nonenol (HNE)- and nitrotyrosine (NT)-modified proteins. Matrix-assisted laser desorption ionization-time of flight and tandem mass spectroscopy were used to identify modified proteins. We found that global HNE- and NT-modified proteins accumulated significantly with aging but not with muscle unloading. Two HNE and NT target proteins, four-and-a-half LIM protein 1 (FHL1) and carbonic anhydrase III (CAIII), showed changes in the oxidation levels with muscle unloading. The changes in the oxidation levels happened to adult rats but not old rats. However, old rats had higher baseline levels of HNE-modified FHL1. In summary, the data suggest that the muscle unloading-related changes of protein oxidation are more significant in specific proteins and that the changes are age related.
氧化修饰含量呈现与废用相关变化的蛋白质的身份尚不清楚。此外，肌肉废用的老年动物中氧化蛋白质的整体积累是否更多也不清楚。本研究的目的是：1）确定氧化水平呈现与废用相关变化的具体蛋白质；2）检验肌肉废用的老年动物中氧化蛋白质的整体积累会更多这一假设。成年和老年大鼠被随机分为四组：负重组以及后肢卸载3天、7天或14天组。采集比目鱼肌以研究卸载情况下的蛋白质氧化。采用狭缝印迹、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和蛋白质免疫印迹分析来检测4-羟基-2-壬烯醛（HNE）和硝基酪氨酸（NT）修饰的蛋白质的积累情况。利用基质辅助激光解吸电离飞行时间质谱和串联质谱来鉴定修饰的蛋白质。我们发现，HNE和NT修饰的蛋白质整体上随衰老而显著积累，但不随肌肉卸载而积累。两种HNE和NT靶蛋白，即四又二分之一LIM结构域蛋白1（FHL1）和碳酸酐酶III（CAIII），其氧化水平随肌肉卸载而发生变化。氧化水平的变化发生在成年大鼠而非老年大鼠中。然而，老年大鼠中HNE修饰的FHL1的基线水平更高。总之，数据表明蛋白质氧化与肌肉卸载相关的变化在特定蛋白质中更为显著，且这些变化与年龄有关。
