Intervention of sulfur mustard toxicity by downregulation of cell proliferation and metabolic rates

Metabolically active and proliferating basal cells in the skin are most sen sitive to the potent skin blistering chemical warfare compound HD (bis-(2-c hloroethyl) sulfide). We previously described a Ca2+-dependent mechanism of HD (0.3-1 mM) toxicity that was inhibited by the cell-permeant Ca2+ chelat or BAPTA AM (1,2-bis(O-aminophenoxy)ethane-N,N,N',N' acid acetoxymethyl est er), We describe some cellular effects of BAPTA AM that suggest a mechanism for its protective action. Monolayer log-phase normal human epidermal kera tinocytes were incubated (37 degreesC) first in keratinocyte growth medium (KGM) containing BAPTA AM (10-40 muM) for 30 min and then in KGM alone over night prior to evaluation. The BAPTA AM inhibited cell growth in a concentr ation-dependent manner with same cellular degeneration above 30 muM (light microscopy). At 20-30 muM, BAPTA AM also inhibited cellular metabolic proce sses, as evidenced by a lower incorporation of [H-3]-thymidine (DNA synthes is, 54 +/- 5%), [H-3]-uridine (RNA synthesis, 29 +/- 6%) and [C-14]-valine (protein synthesis, 12 +/- 2%) as well as a lower protein content per cultu re (30 +/- 3%) compared with corresponding untreated controls. However, 20- 30 muM BAPTA AM did not cause any demonstrable cytopathology based on morph ological (electron microscopy) as well as biochemical (lactate dehydrogenas e release, an indicator of cell viability loss) criteria, indicating a lack of acute toxicity. These results suggest that a mechanism of protection by BAPTA AM against HD may be via decreasing some metabolic, and therefore pr oliferative, rates.