Effects of hydroxyl radicals on ATPase and protein structure of myofibrils from rat heart.

Free oxygenated radicals frequently are involved in cardiac arrhythmias and contractility disorders during postischemic reperfusion. The aim of this study was to determine the effects of hydroxyl radicals (.OH) in vitro on myofibrillar Ca-adenosinetriphosphatase (ATPase), on the redox state of thiol groups and the electrophoretic pattern of myofibrillar proteins from rat heart. Myofibrils were treated up to 60 min by .OH generated with 0.3 mM H2O2 and 0.1 mM Fe2+. After a 60-min treatment with .OH, the measurement of thiol groups failed to show any oxidation. On the contrary, ATPase activity and electrophoretic pattern were affected dramatically by treatment with .OH. For all Ca2+ concentrations, ATPase was increased after treatment with .OH, but ATPase activation when Ca2+ rose from pCa 8 to pCa 4.5 was only 92% after 30 min of incubation rather than 226% for untreated myofibrils. The electrophoretic analysis of myofibrillar proteins showed a decrease in myosin heavy chain and formation of aggregates in treated myofibrils. All of these effects were reduced when incubation was performed in the presence of mannitol, a specific scavenger of .OH. No effect was observed with 0.1 mM Fe2+ alone or with 0.3 mM H2O2. The action of .OH was very fast to the extent that the effects were observed after only 15 s of incubation. The results reported in the present study may be related to the impaired relaxation and contracture described in vivo within the first minutes of a postischemic reperfusion and before any change in calcium homeostasis.