MITOCHONDRIAL NADH-QUINONE OXIDE REDUCTASE OF THE OUTER-MEMBRANE IS RESPONSIBLE FOR PARAQUAT CYTOTOXICITY IN RAT LIVERS

We investigated the existence of an NADH-dependent paraquat (PQ) reduc tion system in rat liver mitochondria (Mt) in respect to the cytotoxic mechanisms of PQ. The outer membrane fractions, free from the contami nation of inner membranes but with a few microsomes, catalyzed rotenon e-insensitive NADH, but not NADPH, oxidation by menadione or PQ, Anti- NADH-cytochrome b(5) reductase antibody and its inhibitor p-hydroxymer curibenzonate did not inhibit the NADH-PQ reduction activity, Therefor e, the respiratory systems of the inner membranes and microsomal cytoc hrome P450 systems could not have been responsible for the reaction. D icoumarol, an inhibitor of NAD(P)H-quinone oxidoreductase (NQO), dose dependently suppressed the NADH oxidation in the outer membrane via PQ as well as menadione, with I-50 values of 190 (for menadione) and 150 mu M (for PQ). Because of a lower sensitivity to NADPH and the higher doses of dicoumarol required for its inhibition, the activity in the outer membrane may be an ''NADH-quinone oxidoreductase'' which partly differs from the NQO previously reported. This outer membrane enzyme p roduced superoxide anions in the presence of both NADH and PQ and was too tightly membrane-bound to be extracted by Triton X-100 and deoxych olate. From these results, we concluded that the free radical-producin g mitochondrial NADH-quinone oxidoreductase is a novel oxidation-reduc tion system participating in PQ toxicity, This is in good agreement wi th our previous results showing that PQ selectively damaged Mt in vivo and in vitro, resulting in cell death (K.-I. Hirai ct al., 1992, Toxi cology 72, 1-16). (C) 1998 Academic Press.