Temperature-dependent quinone cytotoxicity in platelets involves arylation

Abstract

Menadione (MEN), a representative quinone compound, produces cytotoxicity in many cells by arylation with protein thiols and oxidative stress due to redox cycling. Previously it was demonstrated that protein arylation appears to be a primary mechanism for MEN-induced toxicity in platelets. To test the hypothesis that temperature conditions may be important in MEN-induced cytotoxicity in noncancer cells, platelets were incubated with menadione at 25, 37, or 42°C. As temperature was increased, MEN significantly enhanced lactate dehydrogenase (LDH) leakage. MEN-induced depletion of protein thiol levels also increased as temperature was elevated. To investigate the mechanism of temperature-dependent MEN cytotoxicity, MEN-induced platelet toxicity was compared to two other quinone substances. Benzoquinone (BQ), which acts via arylation, produced cytotoxic effects similar to those of MEN. Dimethoxy-1,4-naphthoquinone (DMNQ), which exerts toxicity via oxidative radical generation, failed to produce cytotoxicity at all three temperatures. While MEN and DMNQ enhanced O 2 consumption in a temperature-dependent manner, BQ did not affect this parameter. MEN, which possesses an electrophilic 3-position, was found to react with thiols to form a thioether linkage, a direct indicator of arylation. In the case of MEN uptake kinetics, the amount of cellular uptake was not different at various temperatures, but concentration of MEN in extracellular medium decreased temperature dependently. This might be due to increased arylation capacity binding to cellular proteins as temperature rises. These data suggest that MEN-induced platelet cytotoxicity involves arylation that is temperature related.