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.