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  • Title: Immunohistochemical localization of O6-ethyldeoxyguanosine and deoxyguanosin-8-yl-(acetyl)aminofluorene in liver sections of rats treated with diethylnitrosamine, ethylnitrosourea or N-acetylaminofluorene.
    Author: Menkveld GJ, Van der Laken CJ, Hermsen T, Kriek E, Scherer E, Den Engelse L.
    Journal: Carcinogenesis; 1985 Feb; 6(2):263-70. PubMed ID: 3882258.
    Abstract:
    Antibodies raised in rabbits against the bovine serum albumin conjugates of O6-ethylguanosine (O6-EtGuo) and N-(guanosin-8-yl)-N-acetyl-2-aminofluorene (Guo-8-AAF) have been used in a double peroxidase-antiperoxidase staining assay to visualize the localization of DNA-O6-ethyldeoxyguanosine (O6-EtdGuo) and some of the interaction products of N-acetyl-2-aminofluorene (AAF) with DNA guanine in liver sections of rats treated with diethylnitrosamine (DEN), ethylnitrosourea (ENU), or AAF respectively. O6-EtdGuo could be detected in nuclei of parenchymal cells after injection of DEN (12-50 mg/kg) or ENU (140 mg/kg). Clear time and dose dependencies were observed. The lowest dose of DEN which, at 5 h after injection, resulted in immunohistochemically detectable levels of O6-EtdGuo, was 12 mg/kg; 5 h after 6 mg/kg no consistent difference between treated and untreated rats could be observed. A striking heterogeneity in staining pattern was observed after DEN: centrilobular regions were stained much more than peripheral zones. At 7 days after a single DEN injection of 50 mg/kg small rims of significantly stained hepatocytes could still be observed around the central veins. No heterogeneity of staining pattern was observed 2 h after ENU. ENU, in contrast with DEN, also resulted in a significant staining of nonparenchymal cells. At 24 h after ENU no significant staining of hepatocytes could be detected, but positive staining was still present in bile duct cells, vascular endothelial cells and sinusoidal cells. The results indicate a detection level of approximately 5 mumol O6-EtdGuo/mol DNA-P, i.e., 5 X 10(4) O6-EtdGuo residues per diploid genome. Using the anti-Guo-8-AAF antiserum, positive results were obtained 6 days after a single AAF dose of 0.5-10 mg/kg, corresponding to a detection limit of less than or equal to 0.4 mumol dGuo-8-(A)AF/mol DNA-P. Staining was rather homogenously distributed over the liver lobules. Persistency of the AAF-DNA interaction products was investigated both after 10 and 2 mg/kg AAF. Increased nuclear staining could be observed up to 8 weeks after 10 mg/kg and 4 weeks after 2 mg/kg.
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