COLD PRESERVATION OF ISOLATED RABBIT PROXIMAL TUBULES INDUCES RADICAL-MEDIATED CELL INJURY

Background. Reactive oxygen species (ROS) are involved in reperfusion injury after preservation, Recent studies in isolated endothelial cell s and hepatocytes suggested the occurrence of ROS-mediated injury duri ng the period of cold incubation, In the present study, formation of R OS and subsequent cell injury were studied in freshly isolated rabbit proximal tubules (PTs). Methods, PTs were incubated in University of W isconsin (UW) solution, Euro-Collins solution, or a modified Krebs-Hen seleit buffer under aerobic conditions for up to 94 hr at 4 degrees, R OS formation and cell death were assessed as lipid peroxidation (forma tion of thiobarbituric acid-reactive substances [TBARS]) and release o f lactate dehydrogenase, respectively, The involvement of ROS was furt her investigated in UW solution using compounds that might interfere w ith ROS formation, In addition, tubules were studied under anaerobic c onditions (gassing with 95% N-2/5% CO2). Results. Cold preservation of rabbit PTs in any of the solutions under aerobic conditions caused pr ogressive lipid peroxidation and concomitant cell injury, Addition to UW solution of inhibitors of ROS formation, in particular 2,2'-dipyrid yl, or removal of oxygen by gassing with 95% N-2/5% CO2, prevented lip id peroxidation and protected rabbit PTs against cold injury, Both the nitric oxide (NO) synthase inhibitor L-NAME and dexamethasone, which blocks the inducible NO synthase, were ineffective, The cytoprotectant glycine affected neither TEARS formation nor lactate dehydrogenase re lease, Conclusions, Cold preservation of renal PTs under aerobic condi tions caused cell injury even in the specially designed preservation s olution UW. Cell injury is caused by iron-dependent, NO synthase-indep endent ROS formation.