Detection and quantification of ploidy, nuclearity, and DNA synthesis in rat hepatocytes after administration of a peroxisome proliferator.
The peroxisome proliferator methylclofenapate (MCP) induces species-specific liver growth and cancer in rats and mice. The acute hyperplastic effects of MCP were studied in rats given MCP (25 mg/kg by daily gavage) and injected IP (0.5 mL, 50 mM) with bromodeoxyuridine (BrdU) 6 hr before each sampling time. The animals were killed at 6-hr intervals and hepatocyte suspensions prepared from their livers by collagenase perfusion. The cells were stained with propidium iodide (PI) and BrdU antibody combined with fluorescein isothiocyanate (FITC). DNA content (PI-red fluorescence) and S phase (BrdU/FITC-green fluorescence) were analyzed simultaneously by two-parameter flow cytometry and the frequency of S phase in different ploidy classes determined. At the same time, S-phase cells from different ploidy groups were sorted onto slides by fluorescence-activated cell sorting and examined microscopically to determine the frequency of binucleated cells undergoing DNA synthesis. The results show that MCP-induced acute hyperplasia occurs mainly in a sensitive subpopulation of binucleated hepatocytes.
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JA
Imperial Chemical Industries, Central Toxicology Laboratory, Macclesfield, Cheshire, UK.
D000818:Animals; D001973:Bromodeoxyuridine / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000648:ultrastructure; D002992:Clofenapate / Q000633:toxicity; D004247:DNA / Q000096:biosynthesis*; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D008297:Male; D051379:Mice; D008830:Microbodies / Q000187:drug effects* / Q000648:ultrastructure; D011003:Ploidies; D051381:Rats; D016196:S Phase / Q000187:drug effects
D002992:Clofenapate; D004247:DNA; D001973:Bromodeoxyuridine