Marin-Corral J, Minguella J, Ramírez-Sarmiento A L, Hussain S N A, Gea J, Barreiro E
Respiratory Medicine and Thoracic Surgery Depts, Muscle and Respiratory System Research Unit, IMIM Hospital del Mar, PRBB, Barcelona, Spain.
Eur Respir J. 2009 Jun;33(6):1309-19. doi: 10.1183/09031936.00072008. Epub 2009 Feb 5.
In the diaphragms of chronic obstructive pulmonary disease (COPD) patients, the nature of oxidatively modified proteins and superoxide anion production were explored. Diaphragm specimens were obtained through thoracotomy because of localised lung lesions in COPD patients (16 severe and eight moderate) and 10 control subjects. Lung and respiratory muscle functions were evaluated. Oxidised proteins were identified using immunoblotting and mass spectrometry. Protein and activity levels of the identified proteins were determined using immunoblotting and activity assays. Lucigenin-derived chemiluminescence signals in a luminometer were used to determine superoxide anion levels in muscle compartments (mitochondria, membrane and cytosol) using selective inhibitors. In severe COPD patients compared with controls, respiratory muscle function was impaired; creatine kinase, carbonic anhydrase III, actin and myosin were oxidised; myosin carbonylation levels were increased five-fold; creatine kinase content and activity and myosin protein were reduced; superoxide anion levels were increased in both mitochondria and membrane compartments; and the percentage of superoxide anion inhibition achieved by rotenone was significantly greater. In severe COPD patients, oxidation of diaphragm proteins involved in energy production and contractile performance is likely to partially contribute to the documented respiratory muscle dysfunction. Furthermore, generation of the superoxide anion was increased in the diaphragms of these patients.
在慢性阻塞性肺疾病（COPD）患者的膈肌中，对氧化修饰蛋白的性质和超氧阴离子的产生进行了研究。由于COPD患者（16例重度和8例中度）及10名对照受试者存在局部肺部病变，通过开胸手术获取膈肌标本。评估肺和呼吸肌功能。使用免疫印迹和质谱法鉴定氧化蛋白。使用免疫印迹和活性测定法测定所鉴定蛋白的蛋白质和活性水平。使用发光计中基于光泽精的化学发光信号，通过选择性抑制剂测定肌肉区室（线粒体、膜和细胞质）中的超氧阴离子水平。与对照组相比，重度COPD患者的呼吸肌功能受损；肌酸激酶、碳酸酐酶III、肌动蛋白和肌球蛋白被氧化；肌球蛋白羰基化水平增加了五倍；肌酸激酶含量、活性和肌球蛋白蛋白减少；线粒体和膜区室中的超氧阴离子水平均升高；并且鱼藤酮实现的超氧阴离子抑制百分比显著更高。在重度COPD患者中,参与能量产生和收缩性能的膈肌蛋白氧化可能部分导致了已记录的呼吸肌功能障碍。此外，这些患者膈肌中超氧阴离子的生成增加。
