High-Affinity Low-Capacity and Low-Affinity High-Capacity N-Acetyl-2-Aminofluorene (AAF) Macromolecular Binding Sites Are Revealed During the Growth Cycle of Adult Rat Hepatocytes in Primary Culture
| Autor: | T. Moran, Katherine S. Koch, Hyam L. Leffert, W. Thomas Shier |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
DNA Replication Metabolite Cells Primary Cell Culture Biology Toxicology DNA Synthesis Inhibition Rats Sprague-Dawley 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine medicine polycyclic compounds Animals Binding site primary hepatocytes Cells Cultured 030304 developmental biology Cell Proliferation 0303 health sciences Inbred F344 Cultured Binding Sites procarcinogen binding DNA synthesis hepatocarcinogenesis DNA replication Pharmacology and Pharmaceutical Sciences DNA 2-Acetylaminofluorene Molecular biology Rats Inbred F344 Rats N-Acetyl-2-Aminofluorene Binding Sites in Cultured Rat Hepatocytes genomic DNA 030104 developmental biology medicine.anatomical_structure chemistry Biochemistry 030220 oncology & carcinogenesis Hepatocyte Carcinogens Hepatocytes Autoradiography Sprague-Dawley N-acetyl-2-aminofluorene |
| Zdroj: | Koch, KS; Moran, T; Shier, WT; & Leffert, HL. (2018). High-Affinity Low-Capacity and Low-Affinity High-Capacity N-Acetyl-2-Aminofluorene (AAF) Macromolecular Binding Sites Are Revealed During the Growth Cycle of Adult Rat Hepatocytes in Primary Culture. TOXICOLOGICAL SCIENCES, 163(1), 26-35. doi: 10.1093/toxsci/kfy007. UC San Diego: Retrieved from: http://www.escholarship.org/uc/item/16j0s31d Toxicological sciences : an official journal of the Society of Toxicology, vol 163, iss 1 |
| Popis: | Long-term cultures of primary adult rat hepatocytes were used to study the effects of N-acetyl-2-aminofluorene (AAF) on hepatocyte proliferation during the growth cycle; on the initiation of hepatocyte DNA synthesis in quiescent cultures; and, on hepatocyte DNA replication following the initiation of DNA synthesis. Scatchard analyses were used to identify the pharmacologic properties of radiolabeled AAF metabolite binding to hepatocyte macromolecules. Two classes of growth cycle-dependent AAF metabolite binding sites – a high-affinity low-capacity site (designated Site I) and a low-affinity high-capacity site (designated Site II) – associated with two spatially distinct classes of macromolecular targets, were revealed. Based upon radiolabeled AAF metabolite binding to purified hepatocyte genomic DNA or to DNA, RNA, proteins and lipids from isolated nuclei, Site IDAY 4targets (KD[APPARENT]≈ 2-4 x 10−6M and BMAX[APPARENT]≈ 6 pmols/106cells/24 h) were consistent with genomic DNA; and with AAF metabolized by a nuclear cytochrome P450. Based upon radiolabeled AAF binding to total cellular lysates, Site IIDAY 4targets (KD[APPARENT]≈ 1.5 x 10−3M and BMAX[APPARENT]≈ 350 pmols/106cells/24 h) were consistent with cytoplasmic proteins; and with AAF metabolized by cytoplasmic cytochrome P450s. DNA synthesis was not inhibited by concentrations of AAF that saturated DNA binding in the neighborhood of the Site I KD. Instead, hepatocyte DNA synthesis inhibition required higher concentrations of AAF approaching the Site II KD. These observations raise the possibility that carcinogenic DNA adducts derived from AAF metabolites form below concentrations of AAF that inhibit replicative and repair DNA synthesis. |
| Databáze: | OpenAIRE |
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