The Primary and Secondary Metabolism of Aflatoxin B1 by Rat Hepatocytes Cultured on Matrigel

Abstract

Rat hepatocytes on culturing rapidly lose the ability to metabolize many xenobiotics including the mycotoxin aflatoxin B1 (AFB1). Rat hepatocytes cultured for 5 days on plastic retain <10% of the initial cytochrome P450 content compared with 34% in cells grown on matrigel alone and 74% in cells cultured on matrigel plus phenobarbital (PB). Although microsomal activation of AFB1 was preserved in matrigel cultured hepatocytes and increased by PB in all the culturing conditions examined, the AFB1 macromolecular binding capacity was reduced to <10% of that present in the initial cultures. This lack of correlation between cytochrome P450 levels, microsomal activation, and AFB1 binding involved cytosolic glutathione-S-transferase activity (GST). This increased in matrigel cultured cells and was further enhanced by PB. Western blotting of the cytosols showed that the expression of the alpha class GST subunit Yc2 correlated with AFB1-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 suppressed 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.