METHYLENE-CHLORIDE - AN INHALATION STUDY TO INVESTIGATE TOXICITY IN THE MOUSE LUNG USING MORPHOLOGICAL, BIOCHEMICAL AND CLARA CELL-CULTURE TECHNIQUES

Single exposures of mice to methylene chloride (MC) cause vacuolation and necrosis of the bronchiolar Clara cells which subsequently recover normal morphology on continued exposure. Both cytochrome P-450 (CYP)- and glutathione S-transferase (GST)-dependent metabolism of MC are kn own to occur. The current studies have investigated the metabolism of MC in mouse lung using inhibitors of both GST and CYP-dependent routes of metabolism, the consequences of metabolic inhibition on the Clara cell vacuolation, and any changes in cell proliferation, assessed in v itro, in Clara cells cultured from exposed individuals. Vacuolated bro nchiolar cells were seen in mice exposed to 2000 and 4000 ppm MC but w ere not seen at lower concentrations, while addition of the CYP inhibi tor, piperonyl butoxide, significantly reduced the bronchiolar cell va cuolation seen following exposure to 2000 ppm MC. Treatment of mice wi th the glutathione depletor, buthionine sulphoximine, had no effect on the number of vacuolated bronchiolar cells following MC. Exposure of mice to 1000 ppm MC and above for 6 h caused a burst of DNA synthesis in bronchiolar Clara cells cultured in vitro from the lungs of exposed animals. The results suggest that the Clara cell vacuolation followin g MC exposure is mediated via CYP metabolism, that depression of the C YP metabolic pathway occurs following exposure, and that Clara cell va cuolation may have a priming role in stimulating cell proliferation in the unaffected cell population.