title: An in vivo method for using 5-bromo-2'-deoxyuridine (BrdU) as a marker of chemically-induced hepatocellular proliferation in the Japanese medaka (Oryzias latipes).
abstract: Japanese medaka fish (Oryzias latipes) were used to develop an in vivo method to assess hepatocellular proliferation in a nonmammalian model. Proliferative responses were assessed in medaka at 7, 17, 24, and 94 days after a 48-hour exposure to 10 or 100 mg/L diethylnitrosamine (DEN). Subgroups of medaka were exposed to 50 or 75 mg/L of 5-bromo-2'-deoxyuridine (BrdU) in water for 72 hours, sacrificed, and then processed for immunohistochemical staining. Proliferative indices of BrdU-labeled hepatocytes were quantified and compared using both count and area measurements. There was a significant increase (p < 0.05) in hepatocellular proliferation in the 100 mg/L DEN-treated fish as compared to controls and 10 mg/L DEN-treated fish for the first 3 time points. Hepatocarcinogenicity was evaluated 26 weeks post-DEN exposure. There was a significant increase (p < 0.0001) in hepatocellular neoplasms in 100 mg/L DEN-treated fish compared to other fish. Effective BrdU-labeling of S-phase hepatocytes in medaka was achieved by adding BrdU to the aquarium water, and an increase in hepatocellular proliferation using this method was detected 7 days after exposure to a carcinogenic concentration of DEN. Additionally, the new method of area measurement indices of proliferation were as precise as count indices (R2 > or = 0.92).
authors.affiliation: GEO-CENTERS, INC, Fort Detrick, Maryland 21702-5010, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Brennan
authors.forename: E M
authors.initials: EM
authors.lastname: Boncavage-Hennessey
authors.forename: M J
authors.initials: MJ
authors.lastname: Wolfe
authors.forename: M W
authors.initials: MW
authors.lastname: Toussaint
authors.forename: W E
authors.initials: WE
authors.lastname: Dennis
authors.forename: A B
authors.initials: AB
authors.lastname: Rosencrance
authors.forename: H S
authors.initials: HS
authors.lastname: Gardner
mesh_terms: D018248:Adenoma, Liver Cell / Q000139:chemically induced / Q000473:pathology; D000818:Animals; D014408:Biomarkers, Tumor; D001973:Bromodeoxyuridine / Q000008:administration & dosage / Q000378:metabolism*; D015197:Carcinogenicity Tests; D002273:Carcinogens / Q000633:toxicity; D006528:Carcinoma, Hepatocellular / Q000139:chemically induced / Q000473:pathology; D002455:Cell Division / Q000187:drug effects; D004052:Diethylnitrosamine / Q000633:toxicity; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008113:Liver Neoplasms / Q000139:chemically induced / Q000473:pathology; D009990:Oryzias / Q000502:physiology*; D016196:S Phase; D013997:Time Factors
chemical_list: D014408:Biomarkers, Tumor; D002273:Carcinogens; D004052:Diethylnitrosamine; D001973:Bromodeoxyuridine