Sepsis increases contraction-related generation of reactive oxygen species in the diaphragm

Pulmonary Division, Department of Medicine, Case Western Reserve University, and MetroHealth Medical Center, Cleveland, Ohio 44109 Recent work indicates that free radicals mediate sepsis-induced diaphragmatic dysfunction. These previous experiments have not, however, established the source of the responsible free radical species. In theory, this phenomenon could be explained if one postulates that sepsis elicits an upregulation of contraction-related free radical formation in muscle. The purpose of the present study was to test this hypothesis by examination of the effect of sepsis on contraction-related free radical generation [i.e., formation of reactive oxygen species (ROS)] by the diaphragm. Rats were killed 18 h after injection with either saline or endotoxin. In vitro hemidiaphragms were then prepared, and ROS generation during electrically induced contractions (20-Hz trains delivered for 10 min) was assessed by measurement of the conversion of hydroethidine to ethidium. ROS generation was negligible in noncontracting diaphragms from both saline- and endotoxin-treated groups (2.0 ± 0.6 and 2.8 ± 1.0 ng ethidium/mg tissue, respectively), but it was marked in contracting diaphragms from saline-treated animals (19.0 ± 2.8 ng/mg tissue) and even more pronounced (30.0 ± 2.8 ng/mg tissue) in diaphragms from septic animals ( P