THE PRIMARY AND SECONDARY METABOLISM OF AFLATOXIN B-1 BY RAT HEPATOCYTES CULTURED ON MATRIGEL

Rat hepatocytes on culturing rapidly lose the ability to metabolize ma ny xenobiotics including the mycotoxin aflatoxin B-1 (AFB(1)). Rat hep atocytes cultured for 5 days on plastic retain <10% of the initial cyt ochrome P450 content compared with 34% in cells grown on matrigel alon e and 74% in cells cultured on matrigel plus phenobarbital (PB). Altho ugh microsomal activation of AFB(1) was preserved in matrigel cultured hepatocytes and increased by PB in all the culturing conditions exami ned, the AFB(1) macromolecular binding capacity was reduced to <10% of that present in the initial cultures. This lack of correlation betwee n cytochrome P450 levels, microsomal activation, and AFB(1) binding in volved cytosolic glutathione-S-transferase activity (GST). This increa sed in matrigel cultured cells and was further enhanced by PB. Western blotting of the cytosols showed that the expression of the alpha clas s GST subunit Yc(2) correlated with AFB(1)-GSH conjugate formation. By contrast, expression of the alpha class GST Ya-type subunit showed no similar close correlation. The expression of pi class GST Yf subunit, which is associated with dedifferentiation in hepatocytes, was suppre ssed by culturing on matrigel. The results obtained using hepatocytes cultured on matrigel indicate that despite limitations, this comprises a potentially useful model for the in vivo situation. (C) 1994 Academ ic Press, Inc.