Journal Article

Expression of stably transfected murine glutathione <i>S</i>-transferase A3-3 protects against nucleic acid alkylation and cytotoxicity by aflatoxin B<sub>1</sub> in hamster V79 cells expressing rat cytochrome P450-2B1

Wanda R. Fields, Charles S. Morrow, Johannnes Doehmer and Alan J. Townsend

in Carcinogenesis

Volume 20, issue 6, pages 1121-1125
Published in print June 1999 | ISSN: 0143-3334
Published online June 1999 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/20.6.1121
Expression of stably transfected murine glutathione S-transferase A3-3 protects against nucleic acid alkylation and cytotoxicity by aflatoxin B1 in hamster V79 cells expressing rat cytochrome P450-2B1

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Aflatoxin B1 (AFB1) is activated to AFB1-8,9-oxide (AFBO), a potent mutagenic and carcinogenic metabolite of AFB1. In the mouse, AFBO has been shown to be most efficiently detoxified by a specific isozyme of α-class glutathione S-transferase (GST), mGSTA3-3 (mGST-Yc). A hamster V79 cell line (V79MZr2B1, originally designated V79/SD1) previously transfected with the rat cytochrome P450-2B1 was stably transfected with an mGSTA3-3 expression vector, to study the chemopreventive role of GST in protecting against cytotoxicity or genotoxicity of AFBO. Immunoblotting demonstrated strong expression of an α-class GST in the mGSTA3-3 transfected cell line, whereas no detectable α-class GST protein was observed in the control (empty vector-transfected) cells. Previous studies with the V79MZr2B1 cell line indicated that it can activate AFB1 to a mutagenic metabolite via a transfected rat P450-2B1 stably expressed in the cells. We examined the ability of the expressed mGSTA3-3 to protect against AFB1-induced cytotoxicity or [3H]-covalent adduct formation in cellular nucleic acids. Exposure of empty vector-transfected control cells and mGSTA3-3 expressing cells to up to 600 nM [3H]-AFB1 indicated that a 70–80% reduction in DNA and RNA adducts was afforded by the expression of mGSTA3-3 in the transfected cells. Clonogenic survival assays showed that the mGSTA3-3 cell line was 4.6-fold resistant to AFB1 cytotoxicity as compared with the empty vector-transfected control SD1 cells, with IC50 values of 69 and 15 μM, respectively. The results of these studies demonstrate that mGSTA3-3 confers substantial protection against nucleic acid covalent modification and cytotoxicity by AFB1 in this transgenic cell model system.

Keywords: AFB1, aflatoxin B1; [3H]-AFB1, 3H-labelled AFB1; AFBO, aflatoxin B1-8,9-epoxide; CDNB, 1-chloro-2,4-dinitrobenzene; GST, glutathione S-transferase; hGSTM1, human glutathione S-transferase μ-1; hGSTT1, human glutathione S-transferase 𝛉-1; mGSTA3-3, murine glutathione S-transferase α-3 homodimer; rCYP2B1, rat cytochrome P450-2B1.

Journal Article.  3657 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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