Glutathione conjugation and DNA-binding of (±)-<i>trans</i>-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene and (±)-7β,8α-dihydroxy-9α,10α-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene in isolated rat hepatocytes

Abstract

Isolated rat liver hepatocytes, previously depleted of glutathione (GSH) by treatment with diethylmaleate, were allowed to incorporate [3H]glycine into their GSH. Incubation of 3H-labelled cells with 14C-labelled (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene ((+/-)-BP-7,8-dihydrodiol) or (+/-)-7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene ((+/-)-BPDE) revealed the formation of double labelled products. This together with evidence from amino acid analysis indicates formation of GSH-conjugates of the highly carcinogenic BP-derivatives. Incubation of hepatocytes isolated from 3-methylcholanthrene (MC) treated rats with 3H-labelled (+/-)-BP-7,8-dihydrodiol or (+/-)-BPDE resulted in binding of radioactivity to DNA. Reduction of the intracellular level of GSH to approximately 40% of the normal level resulted in an approximate 2-fold increase in the DNA-binding of either substrate. In addition there was a concurrent decrease in the amount of GSH-conjugates formed. These data clearly demonstrate that GSH participates in conjugation reactions with carcinogenic (+/-)-BP-7,8-dihydrodiol and (+/-)-BPDE and that the intracellular level of GSH is important in preventing reactive intermediates from reacting with the DNA in intact cells.