DNA Adduct Formation and Removal in N-Acetoxy-2-Acetylaminofluorene-Exposed Cultured Cells and in Organs from Rats Fed 2-Acetylaminofluorene
The development of carcinogen DNA-adduct antibodies has made possible a new approach to investigate carcinogen-DNA interactions (1). To quantitate adducts of a particular carcinogen, highly-avid rabbit antibodies have been employed to allow detection by radioimmunoassay (RIA) of one adduct in 10 5DNA bases (1). The studies described here employed the antiserum anti-guanosin-(8-yl)-acetylaminofluorene (anti-G-8-AAF) elicited against the nucleoside-adduct coupled covalently to bovine serum albumin and injected into rabbits (2). The antiserum is specific for the acetylated and deacetylated C-8 adducts of 2-acetylaminofluorene (2-AAF) with DNA (dG-8-AAF and dG-8-AF) (see Figure 1). It does not cross-react with the minor adduct, 3-deoxyguanison(N2-yl)-acetylaminofluorene (dG-N2-AAF) (see Figure 1), the carcinogen alone, or DNA (2,3). Since the C-8 adducts comprise the major proportion (80 to 90%) of adducts formed upon interaction of 2-AAF, or its activated derivative N-acetoxy-2-acetylaminofluorene (N-Ac-AAF) with DNA in vivo (3,4,5), the anti-G-8-AAF was considered appropriate for initial studies. The antibody has been utilized to distinguish between the acetylated and deacetylated C-8 adducts of 2-AM in DNA, and to quantitate the proportions of each in DNA extracted from either cultured cells exposed to N-Ac-AAF or from livers and kidneys of male rats fed 2-AAF.
KeywordsHigh Pressure Liquid Chromatography Adduct Level Normal Human Fibroblast Cesium Chloride Retinyl Acetate
Unable to display preview. Download preview PDF.
- 6.Poirier, M.C., G.M. Williams, and S.H. Yuspa. 1980. Effect of culture conditions, cell type, and species of origin on the distribution of acetylated and deacetylated deoxyguanosine C-8 adducts of N-acetoxy-2-acetylaminofluorene. Molec. Pharmacol. 18: 234–240.Google Scholar
- 7.Poirier, M.C. 1981. Measurement of the formation and removal of DNA adducts of N-acetoxy-2-acetylaminofluorene. In: DNA Repair: A Laboratory Manual of Research Procedures, Volume 1A. E.C. Friedberg and P.C. Hanawalt, eds. Marcel Dekker, Inc.: New York. pp. 143–153.Google Scholar
- 9.Poirier, M.C., and R.J. Connor. (in press). A radioimmunoassay for 2-acetylaminofluorene-DNA adducts. In: Immunochemical Techniques, Volume 2. H. Van Vunakis and J. Langone, eds. Academic Press: New York.Google Scholar
- 14.Sakai, S., C.E. Reinhold, P.J. Wirth, and S.S. Thorgiersson. 1978. Mechanism of in vitro mutagenic activation and covalent binding of N–hydroxy–2–acetylaminofluorene in isolated liver cell nuclei from rat and mouse. Cancer Res. 38–2058–2067.Google Scholar
- 15.Schut, H.A., P.J. Wirth, and S.S. Thorgiersson. 1978. Mutagenic activation of N-hydroxy-2-acetylaminofluorene in the salmonella test system: The role of deacetylation by liver and kidney fractions from mouse and rat. Molec. Pharmacol. 14: 682–692.Google Scholar
- 16.Howard, P.C., F.A. Beland, and D.A. Casciano. 1981. Quantitation of N-hydroxy-2-acetylaminofluorene-mediated DNA adduct formation and the subsequent repair in primary rat hepatocyte cultures. J. Supra Struct. Cell. Biochem. Suppl. 5, Abstract 534, p. 197.Google Scholar
- 18.Cerutti, P.A. 1978. Repairable damage in DNA. In: DNA repair mechanisms: ICN-UCLA Symposia on Molecular and Cellular Biology, IA. P.C. Hanawalt, E.C. Friedberg, and C.F. Fox, eds. Academic Press: New York. pp. 1–14.Google Scholar
- 19.Levinson, J.W., B. Konze-Thomas, V.M. Maher, and J.J. McCormick. 1979. Evidence for a common rate-limiting step in the repair process of ultraviolet light and N-acetoxyacetylaminofluorene-induced damage in the DNA of human fibroblasts. Proc. Am. Assoc. Cancer Res. 20: 105.Google Scholar