A possible involvement of plasma membrane NAD[P]H oxidase in the switch mechanism of the cell death mode from apoptosis to necrosis in menadione-induced cell injury

The effects of inhibitors of plasma membrane NADPH oxidase on menadione-in­duced cell injury processes were studied using human osteosarcoma 143B cells. The intracellular level of superoxide in the cells treated with menadione for 6 h reached a maximum followed by an abrupt decrease. The population of apoptotic cells detected by Annexin V and propidium iodide double staining also reached its maximum at 6 h of menadione-treatment while that of necrotic cells increased continuously reaching 90% of the total population at 9 h of the treatment. Pretreatment of the cells with in­hibitors of NADPH oxidase, including diphenyliodonium chloride, apocynin, N-vani- llylnonanamide and staurosporine was effective in lowering the menadione-induced elevations of superoxide, and also in the suppression of the switch of the cell death mode from apoptosis to necrosis in menadione-treated cells except for the case of staurosporine. These results strongly suggest that superoxide generated by NADPH oxidase, besides that generated by the mitochondria, may contribute to the remark­able increase in the intracellular level of superoxide in the cells treated with menadione for 6 h resulting in the switch from apoptosis to necrosis, although a di­rect evidence of the presence of active and inactive forms of NADPH oxidase in con­trol and menadione-treated 143B cells is lacking at present.