The ability of
naphthalene 1,2-oxide to diffuse across intact cellular membranes, the subsequent biotransformation of this
epoxide and its potential to produce losses in cellular viability have been examined in incubations of isolated hepatocytes. Addition of 1R,2S- or 1S,2R-naphthalene
oxide enantiomers (15, 30 and 60 microM) to isolated hepatocytes resulted in a rapid depletion of intracellular
glutathione. Depletion of
glutathione was concentration dependent and maximal at 5-15 min. Addition of either of the enantiomeric
oxides at 60 microM resulted in the loss of more than 20 nmol
glutathione/10(6) cells (1 ml cells); thus more than a third of the added
epoxide was available for conjugation with intracellular
glutathione. The time course and concentration dependence of
glutathione depletion corresponded to the rapid, concentration-dependent formation of
naphthalene oxide glutathione conjugates. The levels of
glutathione adduct were highest 1 min after addition of
naphthalene oxide and declined to 25% of this level after 30 min. Loss of
glutathione conjugates from incubations correlated with the formation of
N-acetylcysteine adducts. In contrast, the levels of
glutathione adducts added exogenously to hepatocytes were relatively stable over a 120-min incubation suggesting that although further metabolism of
naphthalene oxide glutathione adducts formed intracellularly is possible, extracellular
glutathione adducts cannot penetrate the hepatocellular membrane. Small amounts of radiolabel from [3H]
naphthalene 1,2-oxide were bound covalently to macromolecules in hepatocytes; the rate of this binding slowed rapidly after the first minute of incubation. Severe blebbing of the surface of the hepatocytes was noted in cells incubated for 30 min with 480 microM
naphthalene oxide. Many of the cells were vacuolated at 60 min and progressed to frank
necrosis with pyknotic nuclei and inability to exclude
trypan blue. Cells incubated with
1-naphthol responded in a qualitatively similar fashion to those cells incubated with
epoxide; however, hepatocytes incubated with
1-naphthol progressed to frank cellular
necrosis at a slower rate. In hepatocytes partially depleted of
glutathione by pretreatment with
buthionine sulfoximine, addition of 1S,2R-naphthalene
oxide at a rate of 1 nmol/min/10(6) cells resulted in significant losses in cell viability. In contrast, no losses in cell viability were observed with the enantiomer, 1R,2S-naphthalene
oxide. Both
epoxides produced similar losses in cellular
glutathione levels.(ABSTRACT TRUNCATED AT 400 WORDS)