DIETHYLDITHIOCARBAMATE AND NITRIC-OXIDE SYNERGIZE WITH OXIDANTS AND WITH MEMBRANE-DAMAGING AGENTS TO INJURE MAMMALIAN-CELLS

The effect of diethyldithiocarbamate (DDC) and sodium nitroprusside (S NP) on the killing of endothelial cells and on the release of arachido nate by mixtures of oxidants and membrane-damaging agents was studied in a tissue culture model employing bovine aortic endothelial cells la beled either with (51)Chromium or (3)arachidonic acid. While exposure to low, subtoxic concentrations of oxidants (reagent H2O2, glucose-oxi dase generated peroxide, xanthine xanthine oxidase, AAPH-generated per oxyl radical, menadione-generated oxidants) did not result either in c ell death or in the loss of membrane-associated arachidonic acid, the addition of subtoxic amounts of a variety of membrane-damaging agents (streptolysin S, PLA(2), histone, taurocholate, wheatgerm agglutinin) resulted in a synergistic cell death. However, no significant amounts of arachidonate were released unless proteinases were also present. Th e addition to these reaction mixtures of subtoxic amounts of DDC (an S OD inhibitor and a copper chelator) not only very markedly enhanced ce ll death but also resulted in the release of large amounts of arachido nate (in the complete absence of added proteinases). Furthermore, the inclusion in DDC-containing reaction mixtures of subtoxic amounts of S NP, a generator of NO, further enhanced, in a synergistic manner, both cell killing and the release of arachidonate. Cell killing and the re lease of arachidonate induced by the DDC and SNP-containing mixtures o f agonists were strongly inhibited by catalase, glutathione, N-acetyl cysteine, vitamin A, and by a nonpenetrating PLA(2) inhibitor as well as by tetracyclines. A partial inhibition of cell killing was also obt ained by 1,10-phenanthroline and by antimycin. It is suggested that DD C might amplify cell damage by forming intracellular, loosely-bound co mplexes with copper and probably also by depleting antioxidant thiols. It is also suggested that ''cocktails'' containing oxidants, membrane -damaging agents, DDC, and SNP might be beneficial for killing of tumo r cells in vivo and for the assessment of the toxicity of xenobiotics in vitro.