FILTERED, TAR-FREE AND AEROSOL-FREE CIGARETTE-SMOKE CAUSES ACCUMULATION OF THE TUMOR-SUPPRESSOR PROTEIN P53 IN RODENT CELLS

Cigarette smoke was filtered with a Cambridge glass fiber filter retai ning 99.9% of the tar and aerosol fraction and diluted 1:5 with air. T he murine cell line L929 was exposed to this smoke preparation for per iods of up to 10 min. Thereafter the following parameters were determi ned at different times: Nuclear accumulation of the tumor suppressor p rotein p53 indicating chromatin injury (by immunostaining); apoptotic DNA fragmentation (by DNA end labelling with biotin-16-dUTP in the pre sence of terminal deoxyribonucleotidyl transferase); the intracellular level of reactive oxygen intermediates (ROI) (by cytofluorimetry with the fluorigenic stain 2',7'-dichlorofluorescin diacetate). After 1 mi n exposure to 1:5 air-diluted filtered cigarette smoke maximal p53 acc umulation occured about 20 h later, whereas maximal DNA fragmentation and apoptosis and maximal ROI levels were found after 10 min of exposu re. Obviously, even the diluted, tar- and aerosol-free fraction of cig arette smoke has the potency, after 1 min of exposure only, to exert s evere DNA damage, a potential transformation risk for the surviving ce ll fraction, in murine cell cultures as indicated by stabilization and accumulation of the tumor suppressor protein p53.