Reduced 4-aminobiphenyl-induced liver tumorigenicity but not DNA damage in arylamine N-acetyltransferase null mice

Abstract

The aromatic amine 4-aminobiphenyl (ABP) is a liver procarcinogen in mice, requiring enzymatic bioactivation to exert its tumorigenic effect. To assess the role of arylamine N-acetyltransferase (NAT)-dependent acetylation capacity in the risk for ABP-induced liver tumors, we compared 1-year liver tumor incidence following the postnatal exposure of wild-type and NAT-deficient Nat1/2(-/-) mice to ABP. At an ABP exposure of 1200 nmol, male Nat1/2(-/-) mice had a liver tumor incidence of 36% compared to 69% in wild-type males, and at 600 nmol there was a complete absence of tumors compared to 60% in wild-type mice. Only one female wild-type mouse had a tumor using this exposure protocol. However, levels of N-deoxyguanosin-8-yl-ABP-DNA adducts did not correlate with either the strain or sex differences in tumor incidence. These results suggest that female sex and NAT deficiency reduce risk for ABP-induced liver tumors, but by mechanisms unrelated to differences in DNA-damaging events.

Fig. 1

Representative H&E liver sections from wild-type or Nat1/2(−/−) mice sacrificed at one year of age after neonatal ABP (dose as indicated) or DMSO vehicle administration. (A) Liver from a representative male wild-type mouse dosed with 1200 nmol of ABP showing compression of hepatocytes at the border of a nodule (x40). (B) Mitotic body (arrowhead) present in the adenoma from a male wild-type dosed with 1200 nmol ABP (x200). (C) Degenerative changes indicated by the presence of intracytoplasmic inclusions and macrovacuoles in an adenoma with inflammatory cells demarcated by an arrow from a representative male wild-type mouse dosed with 1200 nmol of ABP (x200). (D) Nuclear pleomorphism and absence of sinusoidal radial pattern in a hepatocellular carcinoma from a male Nat1/2(−/−) mouse dosed with 1200 nmol ABP (x200). (E) Region of necrosis in a hepatocellular carcinoma from a male Nat1/2(−/−) mouse treated with 1200 nmol of ABP (x200). (F) Presence of erythrocytes in sinsusoidal spaces and disorganized architecture in a female wild-type dosed with 600 nmol ABP (x200). (G) Region of focal periportal inflammation in a liver displaying overall normal architecture from a female wild-type mouse dosed with 1200 nmol of ABP (x100). (H) Steatosis in a male wild-type mouse dosed with 600 nmol ABP (x200). (I) Liver from a representative male Nat1/2(−/−) mouse dosed with 600 nmol of ABP displaying an overall normal architecture (x200). (J) Liver from a representative female Nat1/2(−/−) dosed with 1200 nmol ABP, displaying an overall normal morphology (x200).

Fig. 2

Comparison of NAT activities in wild-type neonatal and adult male and female mice. Liver cytosols prepared from either postnatal day 8 (PND8), postnatal day 15 (PND15) or 8 week old wild-type male and female mice were assayed for NAT activity using 0.1 mM ABP in the presence of 0.1 mM AcCoA, as described in Materials and Methods. N-acetylated ABP (AABP) was separated and quantified by HPLC. Product formation rates represent the means ± SD from n=3 animals per sex and age. There was no statistically significant difference in enzyme activities between males and females at any age.

Fig. 3

Levels of dG-C8-ABP adducts in adult (A) and neonatal (B) male and female wild-type and Nat1/2(−/−) mice dosed with ABP. Adult animals were dosed i.p. with 20 mg/kg ABP and sacrificed 24 hours later. Neonatal animals were dosed using the 1200 nmol ABP dosing regimen on postnatal days 8 and 15, and sacrificed 24 hours later. Liver genomic DNA was isolated and levels of dG-C8-ABP were measured by HPLC/MS. *, significantly different from wild-type (p< 0.05).