title: Inhibition by linoleic acid hydroperoxide of alveolar macrophage superoxide production: effects upon mitochondrial and plasma membrane potentials.
abstract: Linoleic acid hydroperoxide (LOOH) is a naturally occurring product of lipid peroxidation. Incubation of rat alveolar macrophages with LOOH produced alterations of membrane properties and function at concentrations of LOOH as low as 0.1 microM. These included phorbol myristate acetate (PMA)-stimulated superoxide production, mitochondrial membrane potential, and plasma membrane potentials. These effects were clearly separated from gross loss of structural integrity as measured by lactate dehydrogenase release, in terms of both time of incubation and concentration of LOOH. PMA-stimulated superoxide production measured 15 min after addition of 10 microM LOOH was inhibited approximately 50%; however, addition of this concentration of the hydroperoxide after PMA stimulation was without effect. Superoxide production was also measured in a cell-free system produced by incubation of alveolar macrophages with sodium dodecyl sulfate. Prior incubation of alveolar macrophages with LOOH, H2O2, or t-butyl hydroperoxide, under conditions that significantly inhibited superoxide production by the intact cells, did not produce inhibition of the NADPH-dependent superoxide generating system in the cell-free preparation. These results suggest that the effect of LOOH was upon signal transduction involved in the stimulation of superoxide production rather than on the NADPH oxidase itself. Measurements of membrane potential changes were made using the lipophilic ions, 3,3'-dipentyloxacarbocyanine (DiOC5(3] and bis(3-phenyl-5-oxoisoxazol-4-yl)pentamethineoxonol (oxonol V). On the basis of their charge, DiOC5(3) fluorescence primarily reports mitochondrial potential and oxonol V absorbance reports plasma membrane potential. With 10 microM LOOH, depolarization of the plasma and mitochondrial membranes appeared to occur within seconds. As prior depolarization depresses superoxide production, these hydroperoxide-induced changes in membrane potential may be responsible for decreased PMA-stimulated superoxide production.
authors.affiliation: Department of Pediatrics, Children's Hospital, Los Angeles, California 90027.
authors.forename: H J
authors.initials: HJ
authors.lastname: Forman
authors.forename: E
authors.initials: E
authors.lastname: Kim
mesh_terms: D000818:Animals; D002474:Cell-Free System; D005978:Glutathione / Q000502:physiology; D007425:Intracellular Membranes / Q000187:drug effects* / Q000502:physiology; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008041:Linoleic Acids / Q000494:pharmacology*; D008054:Lipid Peroxides / Q000494:pharmacology*; D008264:Macrophages / Q000201:enzymology / Q000378:metabolism* / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects*; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D011650:Pulmonary Alveoli; D051381:Rats; D011919:Rats, Inbred Strains; D013481:Superoxides / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D008041:Linoleic Acids; D008054:Lipid Peroxides; D013481:Superoxides; C008285:linoleic acid hydroperoxide; D007770:L-Lactate Dehydrogenase; D005978:Glutathione; D013755:Tetradecanoylphorbol Acetate