Carcinogenesis, Vol. 20, No. 10, 1971-1977,
October 1999
© 1999 Oxford University Press
Carcinogenesis |
Glutathione S-transferase-catalyzed conjugation of bioactivated aflatoxin B1 in human lung: differential cellular distribution and lack of significance of the GSTM1 genetic polymorphism
1 Department of Pharmacology and Toxicology,
2 Department of Medicine and
3 Department of Surgery, Queen's University, Kingston, Ontario K7L 3N6, Canada
Epidemiological studies suggest that aflatoxin B1 (AFB1), a mycotoxin produced by certain Aspergillus species, may play a role in human respiratory cancers in occupationally-exposed individuals. AFB1 requires bioactivation to the corresponding exo-8,9-epoxide for carcinogenicity, and glutathione S-transferase (GST)-catalyzed conjugation of the epoxide with glutathione (GSH) is a critical determinant of susceptibility to AFB1. Of the purified human GST enzymes studied, the polymorphic hGSTM1-1 has the highest activity towards AFB1 exo-epoxide. The influence of the GSTM1 polymorphism on AFB1–GSH formation, as well as the abilities of cytosols from preparations enriched in different isolated lung cell types to conjugate AFB1-epoxides, were examined. In whole-lung cytosols from patients undergoing clinically indicated lobectomy, GSTM1 genotype correlated with GSTM1 phenotype as determined by [3H]trans-stilbene oxide conjugation: GSTM1-positive = 295 ± 31 pmol/mg/h (n = 6); GSTM1-negative = 92.8 ± 23.3 pmol/mg/h (n = 4) (P < 0.05). In contrast, conjugation of microsome-generated [3H]AFB1-epoxides with GSH was low and variable between patients, and did not correlate with GSTM1 genotype: GSTM1-positive = 11.9 ± 8.1, 111 ± 66 and 510 ± 248 fmol/mg/h (n = 6); GSTM1-negative = 15.3 ± 16.7, 167 ± 225 and 540 ± 618 fmol/mg/h (n = 4) (for 1, 10 and 100 µM [3H]AFB1, respectively). GSH conjugates of AFB1 exo-epoxide and the much less mutagenic stereoisomer AFB1 endo-epoxide were produced in a ratio of ~1:1 in cytosols from both whole lung and isolated cells. Total cytosolic AFB1-epoxide conjugation was significantly higher in fractions enriched in alveolar type II cells (3.07 ± 1.61 pmol/mg/h) than in unseparated lung cells (0.143 ± 0.055 pmol/mg/h) or fractions enriched in alveolar macrophages (0.904 ± 0.319 pmol/mg/h; n = 4) (P < 0.05). Furthermore, AFB1–GSH formation and percentage of alveolar type II cells in different cell fractions were correlated (r = 0.78, P < 0.05). These results demonstrate that human lung GSTs exhibit very low conjugation activity for both AFB1-8,9-epoxide stereoisomers, and that this activity is heterogeneously distributed among cell types, with alveolar type II cells exhibiting relatively high activity. Of the GSTs present in human peripheral lung which contribute to AFB1 exo- and endo-epoxide detoxification, hGSTM1-1 appears to play at most only a minor role.
Abbreviations: AFB1, aflatoxin B1; CDNB, 1-chloro-2,4-dinitrobenzene; EDTA, ethylenediaminetetraacetic acid; GSH, glutathione; GST, glutathione S-transferase; HEPES, N-2-hydroxy-ethylpiperazine-N-2-ethanesulfonic acid; HPLC, high-performance liquid chromatography; PCR, polymerase chain reaction; tSBO, trans-stilbene oxide.
4 Present address: Department of Surgery, University of Massachusetts, Worchester, MA, USA
5 To whom correspondence should be addressed Email: masseyt{at}post.queensu.ca
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