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RELATIVE SENSITIVITY OF THE 32P-POSTLABELING OF DNA AND THE AUTORADIOGRAPHIC UDS ASSAY IN THE LIVER OF RATS EXPOSED TO 2-ACETYLAMINOFLUORENE (2AAF)
Citation:
Gallagher, J., T. Shank, J. Lewtas, P. Lefevre, AND J. Ashby. RELATIVE SENSITIVITY OF THE 32P-POSTLABELING OF DNA AND THE AUTORADIOGRAPHIC UDS ASSAY IN THE LIVER OF RATS EXPOSED TO 2-ACETYLAMINOFLUORENE (2AAF). U.S. Environmental Protection Agency, Washington, D.C., EPA/600/J-91/163 (NTIS PB91231316), 1991.
Description:
Groups of male rats were dosed concomitantly with 2-AAF by gavage at doses between 0.01 mg/kg and 40 mg/kg, and livers sampled 2-72h later. he liver of one group of animals was perfused to yield hepatocytes which were assayed in vitro for unscheduled DNA synthesis (UDS) via incorporation of tritiated thymidine and autoradiography. NA was extracted from the livers of the other group and DNA adduct levels determined using the 32P-postlabeling technique. he major C-8 2-aminofluorene/guanosine and three minor adducts were quantitated enabling the relative sensitivity of the two techniques to be compared. ose and time related UDS response was observed, which at the most sensitive time point enabled DNA repair to be discerned at a dose level of 0.1 mg/kg of 2 AAF, a response classified formally at 5 mg/kg 2AAF. he C-8 adduct as determined by 32P-postlabeling analysis was discernible at doses as low as 0.01 mg/kg of 2AAF. t is concluded that as part of a well-defined dose response, UDS can be discerned with confidence for doses of 2 AAF between 0.1-5 mg/kg and DNA adducts for doses of 2AAF between 0.01-1.0 mg/kg. iscernible UDS for 2AAF in the rat liver is at 13 DNA (total adducts/10 8 nucleotides or 8 DNA adducts (c-8)adducts/10 8 nucleotides). he presumed C-8 2-acetylaminofluorene/guanosine adduct prepared by reaction of 2-acetoxy-2-acetylamino fluorene (2AAAF) with DNA was a significant but unreliable marKer of 2AAF/DNA adducts in the liver in vivo. NA repair did not appear to remove DNA adducts selectively and adducts remained in DNA when discernible DNA repair had ceased.