H2O2-induced lipid peroxidation in mitochondria of pulmonary vascular smooth muscle tissue and its modification by DFO, DMTU and DIDS.

Treatment of bovine pulmonary artery smooth muscle tissue mitochondria with H2O2 stimulated iron release, hydroxyl radical (OH) production and lipid peroxidation. Pretreatment of mitochondria with deferoxamine (DFO) and dimethyl thiourea (DMTU) prevented OH production and markedly reduced lipid peroxidation without appreciably altering iron release caused by H2O2. Simultaneous treatment of either DFO or DMTU with H2O2 significantly reduced lipid peroxidation and also prevented OH production without causing marked decrease in iron release. In contrast, addition of DFO or DMTU even 2 min after treatment of the mitochondria with H2O2 did not significantly altered iron release, OH production and lipid peroxidation. Pretreatment of the mitochondria with 4,4'-dithiocyano-2,2'-disulfonic acid stilbene (DIDS) markedly reduced lipid peroxidation without appreciably altering the increase in OH production and iron release caused by H2O2.

Involvement of a protease in tert-butylhydroperoxide-mediated activation of Ca2+ ATPase in microsomes of pulmonary smooth muscle.

Microsomes isolated from bovine pulmonary artery smooth muscle tissue treated with the oxidant t-buOOH stimulated Ca2+ ATPase activity dose-dependently as also protease activity when tested with a synthetic substrate N-benzoyl-DL-arginine p-nitroanilide. At 300 microM, t-buOOH optimally stimulated these activities. Treatment of the microsomes with t-buOOH stimulated ATP dependent Ca2+ uptake while Na+ dependent Ca2+ uptake was inhibited by t-buOOH. Pretreatment of the microsomes with vitamin E (1 mM) and aprotinin (1 mg/ml) prevented t-buOOH caused stimulation of protease activity and Ca2+ ATPase activity, and also stimulation of ATP dependent Ca2+ uptake while t-buOOH caused inhibition of Na+ dependent Ca2+ uptake was reversed by vitamin E and aprotinin. Treatment of the microsomes with trypsin (1 microgram/ml) stimulated Ca2+ ATPase and ATP dependent Ca2+ uptake while Na+ dependent Ca2+ uptake was inhibited. Pretreatment of the microsomes with aprotinin prevented trypsin caused stimulation of Ca2+ ATPase and ATP dependent Ca2+ uptake, while trypsin caused inhibition of Na+ dependent Ca2+ uptake was reversed by aprotinin.

Redox state of pyridine nucleotides, but not glutathione, regulate Ca2+ release by H2O2 from mitochondria of pulmonary smooth muscle.

Treatment of bovine pulmonary artery smooth muscle mitochondria with H2O2 stimulated oxidation of GSH and NAD(P)H along with an increase in Ca2+ release. Addition of oxaloacetate to mitochondrial suspension stimulated Ca2+ release and oxidation of NAD(P)H while GSH level remained unchanged. Subsequently, addition of beta-hydroxybutyrate which reduced mitochondrial pyridine nucleotides caused reuptake of the released Ca2+ without causing appreciable alteration of GSH level. Treatment of the mitochondria with 1,3-bis(2-dichloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase, significantly decreased GSH level without producing discernible change in Ca2+ release and NAD(P)H oxidation.

Protective role of anion channel blocker in lipid peroxidation caused by H2O2 in microsomes of bovine pulmonary arterial smooth muscle tissue.

The role of hydroxyl radical (OH.) in H2O2-mediated stimulation of lipid peroxidation in microsomes of bovine pulmonary arterial smooth muscle tissue and the protective effects of DIDS, the anion channel blocker have been studied. Treatment of microsomes with H2O2 (1 mM) stimulate iron release, OH. production and lipid peroxidation. Pretreatment with DFO (an iron chelator) or DMTU (a hydroxyl radical scavenger) prevents OH. production and thereby reduces lipid peroxidation without any appreciable reduction of iron release. Simultaneous treatment of either DFO or DMTU with H2O2 significantly reduces lipid peroxidation and prevents OH. production without any significant reduction of iron release. However, addition of DFO or DMTU 2 min after treatment of the microsome with H2O2 does not produce any significant reduction of lipid peroxidation, OH production and iron release. Pretreatment of microsomes with DIDS markedly reduces the stimulation of lipid peroxidation without appreciably altering the increase in OH. production and iron release caused by H2O2.