Characterization of hepatocellular resistance and susceptibility to styrene toxicity in B6C3F1 mice

Short-term inhalation exposure of B6C3F1 mice to styrene causes necrosis of centrilobular (CL) hepatocytes. However, in spite of continued exposure, t he necrotic parenchyma is rapidly regenerated, indicating resistance by reg enerated cells to styrene toxicity. These studies were conducted to test th e hypothesis that resistance to repeated styrene exposure is due to sustain ed cell proliferation, with production of hepatocytes that have reduced met abolic capacity. Male mice were exposed to air or 500 ppm styrene (6 h/day) ; hepatotoxicity was evaluated by microscopic examination, serum liver enzy me levels, and bromodeoxyuridine (BrdU)-labeling index (LI). Metabolism was assessed by measurement of blood styrene and styrene oxide. Both single an d repeated exposures to styrene resulted in mortality by Day 2; in mice tha t survived, there was CL necrosis with elevated BrdU LI at Day 6, and compl ete restoration of the necrotic parenchyma by Day 15. The BrdU LI in mice g iven a single exposure had returned to control levels by Day 15. Re-exposur e of these mice on Day 15 resulted in additional mortality and hepatocellul ar necrosis, indicating that regenerated CL cells were again susceptible to the cytolethal effect of styrene following a 14-day recovery. However, in mice repeatedly exposed to styrene for 14 days, the BrdU LI remained signif icantly increased on Day 15, with preferential labeling of CL hepatocytes w ith enlarged nuclei (karyomegaly). If repeated exposures were followed by a 10-day recovery period, CL karyomegaly persisted, but the BrdU LI returned to control level and CL hepatocytes became susceptible again to styrene to xicity, as demonstrated by additional mortality and acute necrosis after a challenge exposure. These findings indicated a requirement for continued st yrene exposure and DNA synthesis in order to maintain this resistant phenot ype. Analyses of proliferating-cell nuclear-antigen (PCNA) labeling were co nducted to further characterize the cell cycle kinetics of these hepatocyte s. The proportion of cells in S-phase was increased by repeated exposure. H owever, PCNA analysis also revealed an even larger increase in the G1 cell compartment with repeated exposures, without a concurrent increase in G2 ph ase or in mitotic cell numbers. These data indicate that resistance to styr ene-induced necrosis under conditions of repeated exposure is not due to su stained cell turnover and production of new, metabolically inactive cells, but rather is due to some other, as yet unknown, protective phenotype of th e regenerated cells.