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John H.n. Meerman – One of the best experts on this subject based on the ideXlab platform.
Blockage of transcription as a trigger for p53 accumulation by 2-Acetylaminofluorene DNA-adductsLife sciences, 2003Co-Authors: Hilde E. Van Gijssel, Leon H.f. Mullenders, Michiel F Van Oosterwijk, John H.n. MeermanAbstract:
The hepatocarcinogen 2-Acetylaminofluorene is one of the most studied experimental carcinogens. We have shown previously that normal rat hepatocytes accumulate the tumour suppressor p53 after exposure to this compound while preneoplastic rat hepatocytes do not. We suggested that the lack of p53 response may confer a growth advantage on preneoplastic hepatocytes and may be an important factor in hepatic tumor promotion by 2-Acetylaminofluorene and other genotoxic compounds. Inhibition of RNA polymerase II driven transcription by DNA lesions may constitute one of the mechanisms leading to accumulation of the tumour suppressor p53. We have investigated the accumulation of p53 by structurally different DNA lesions of 2-Acetylaminofluorene for which the rate of nucleotide excision repair (NER) and inhibition of transcription are known. Experiments were performed with NER proficient human fibroblasts as well as repair deficient xeroderma pigmentosum group A (XPA) cells, XPC cells [only transcription coupled repair (TCR)] and Cockayne syndrome (CS)B cells [only global genome repair (GGR)]. The cells were exposed to N-acetoxy-acetylaminofluorene (NAAAF) in the presence or absence of paraoxon inducing dG-C8-AAF or dG-C8-AF adducts respectively. Both treatments led to accumulation of p53 in all cells. However, dG-C8-AAF adducts produced greater p53 induction than dG-C8-AF adducts. The percentage p53-positive cells was highest and the threshold for p53 accumulation was lowest in XPA and CSB cells. Our results further demonstrate that both the potency of a lesion to inhibit transcription as well as the restoration of RNA synthesis determines the magnitude of p53 induction.
Electrochemical Detection and Quantification of the Acetylated and Deacetylated C8-Deoxyguanosine DNA Adducts Induced by 2-AcetylaminofluoreneAnalytical biochemistry, 1997Co-Authors: Sandra A.m. Bol, Anton J.l. De Groot, Roeline B. Tijdens, John H.n. Meerman, Leon H.f. Mullenders, Albert A. Van ZeelandAbstract:
Abstract The genotoxic agent 2-Acetylaminofluorene induces, upon metabolic activation, two main types of DNA adducts in animal tissue, i.e., (deoxyguanine-8-yl)-aminofluorene (dG-C8-AF) andN-(deoxyguanine-8-yl)-acetylaminofluorene (dG-C8-AAF). Quantification of the frequency of these adducts usually relies on the use of radioactively labeled 2-Acetylaminofluorene. Here, we report the development of a sensitive, non-radioactive method for the quantification of dG-C8-AF and dG-C8-AAF. Essentially, the modified DNA bases are separated by high-performance liquid chromatography (HPLC) and quantified by electrochemical detection. We established that both modified bases guanine-C8-aminofluorene and guanine-C8-acetylaminofluorene are electrochemically active. Subsequently, a procedure was developed to quantify dG-C8-AF and dG-C8-AAF in genomic DNA. Following DNA hydrolysis the adducted bases were extracted by ethyl acetate, separated by HPLC, and detected electrochemically. This procedure has been applied in the analysis of dG-C8-AAF inN-acetoxy-2-Acetylaminofluorene-modified calf thymus DNA and in the detection of dG-C8-AAF and dG-C8-AF in liver DNA of mice injected intraperitoneally with 150–450 mgN-hydroxy-2-Acetylaminofluorene/kg. The quantification of relatively low dG-C8-AF and dG-C8-AAF adduct levels (i.e., 0.1–1 adduct/106nucleotides) in mouse liver DNA demonstrates the sensitivity of this electrochemical detection procedure. The detection limit of the method is 1 adduct per 106nucleotides for both adducts using 20 μg of DNA and 4 adducts per 108nucleotides using 500 μg DNA.
Bioactivation of the hepatocarcinogen N-hydroxy-2-Acetylaminofluorene by sulfation in the rat liver changes during the cell cycleLife sciences, 1992Co-Authors: Ron A.h.j. Gilissen, John H.n. MeermanAbstract:
Sulfation activity towards N-hydroxy-2-Acetylaminofluorene and 4-nitrophenol was determined in male rat liver cytosol at several time points after partial hepatectomy corresponding to G1-, S-, and M-phase. N-hydroxy-2-Acetylaminofluorene sulfation activity decreased by 80% when hepatocytes entered the G1-phase. This lower activity was maintained during the S-phase and M-phase, but was restored when hepatocytes entered the G0-phase again. Sulfation activity towards 4-nitrophenol did not alter after hepatectomy. Various other cytosolic enzyme activities were determined after hepatectomy to investigate the specificity of the decrease in sulfation activity. Lactate dehydrogenase and glucose-6-phosphate dehydrogenase activities were increased in the S- and M-phase by maximally 80% and 60%, respectively. Glutathione-S-transferase and glutamate-pyruvate transaminase activity did not alter during the cell cycle. These results indicate that sulfation of N-hydroxy-2-Acetylaminofluorene in hepatocytes may depend on the phase of the cell cycle. The relevance of the finding is discussed in relation to the resistance of proliferating (pre)neoplastic hepatocytes to the toxic and mitoinhibitory effects of N-hydroxy-2-Acetylaminofluorene.
Khalid Abdul Kadir – One of the best experts on this subject based on the ideXlab platform.
Effect of tocotrienols on hepatocarcinogenesis induced by 2-Acetylaminofluorene in rats.The American Journal of Clinical Nutrition, 1991Co-Authors: Wan Zurinah Wan Ngah, Zanariah Jarien, Myint Myint San, Alini Marzuki, Gapor Md Top, Nor Aripin Shamaan, Khalid Abdul KadirAbstract:
The effects of tocotrienols on hepatocarcinogenesis in rats fed with 2-Acetylaminofluorene (AAF) were followed morphologically and histologically for a period of 20 wk. NO differences between treated and control rats in the morphology and histology of their livers was observed. Cell damage was extensive in the livers of AAF-treated rats but less extensive in the AAF-tocotrienols-treated rats. 2-Acetylaminofluorene significantly increases the activities of both plasma and liver microsomal γ-glutamyltranspeptidase (GGT) and liver microsomal UDP-glucuronyltransferase (UDP-GT)
Robin E. Neft – One of the best experts on this subject based on the ideXlab platform.
Genetic toxicity of 2-Acetylaminofluorene, 2-aminofluorene and some of their metabolites and model metabolitesMutation research, 1994Co-Authors: Robert H. Heflich, Robin E. NeftAbstract:
2-Acetylaminofluorene and 2-aminofluorene are among the most intensively studied of all chemical mutagens and carcinogens. Fundamental research findings concerning the metabolism of 2-Acetylaminofluorene to electrophilic derivatives, the interaction of these derivatives with DNA, and the carcinogenic and mutagenic responses that are associated with the resulting DNA damage have formed the foundation upon which much of genetic toxicity testing is based. The parent compounds and their proximate and ultimate mutagenic and carcinogenic derivatives have been evaluated in a variety of prokaryotic and eukaryotic assays for mutagenesis and DNA damage. The reactive derivatives are active in virtually all systems, while 2-Acetylaminofluorene and 2-aminofluorene are active in most systems that provide adequate metabolic activation. Knowledge of the structures of the DNA adducts formed by 2-Acetylaminofluorene and 2-aminofluorene, the effects of the adducts on DNA conformation and synthesis, adduct distribution in tissues, cells and DNA, and adduct repair have been used to develop hypotheses to understand the genotoxic and carcinogenic effects of these compounds. Molecular analysis of mutations produced in cell-free, bacterial, in vitro mammalian, and intact animal systems have recently been used to extend these hypotheses.