Mechanisms of toxicity of 2‐ and 5‐hydroxy‐1,4‐naphthoquinone; absence of a role for redox cycling in the toxicity of 2‐hydroxy‐1,4‐naphthoquinone to isolated hepatocytes

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Abstract
The mechanisms of toxicity to isolated rat hepatocytes of two structurally related naphthoquinones have been studied. Both 5-OH-1,4-naphthoquinone (5-OH-1,4-NQ; juglone) and 2-OH-1,4-naphthoquinone (2-OH-1,4-NQ; lawsone) caused a concentration-dependent cytotoxicity to hepatocytes which was preceded by a depletion of intracellular glutathione. 5-OH-1,4-NQ caused a depletion of intracellular glutathione when incubated either at 4°C or 37°C whereas 2-OH-1,4-NQ caused a depletion of intracellular glutathione when the hepatocytes were incubated at 37°C but not at 4°C. 5-OH-1,4-NQ but not 2-OH-1,4-NQ reacted with glutathione in buffered solution. These results suggested that the depletion of intracellular glutathione by 2-OH-1,4-NQ is enzyme mediated whereas in the case of 5-OH-1,4-NQ the direct chemical reaction with glutathione may be largely responsible for the depletion. A critical role for depletion of protein thiols in menadione-induced cytotoxicity has been proposed. In agreement with earlier work, menadione caused a decrease in protein sulphydryls prior to cell death, however, at cytotoxic concentrations of both 2-OH-1,4-NQ and 5-OH-1,4-NQ this decrease only accompanied rather than preceeded cell death. The mechanism of toxicity of 5-OH-1,4-NQ is similar to that of other naphthoquinones and involves formation of its corresponding naphthosemiquinone, active oxygen species and redox cycling as it stimulated a disproportionate increase in both microsomal NADPH oxidation and oxygen consumption. No evidence for redox cycling of 2-OH-1,4-NQ was obtained either with microsomes or hepatocytes and thus its cytotoxicity must involve a different mechanism to that currently proposed for other naphthoquinones. This inability of 2-OH-1,4-NQ to redox cycle may be due to its very low one-electron reduction potential (E1/7-415 mV). Thus, it is a very poor substrate for reduction to a semiquinone by cellular reductases.