title: Disruption of spindle checkpoint function ahead of facilitation of cell proliferation by repeated administration of hepatocarcinogens in rats.
abstract: We aimed to clarify the hepatocarcinogen-specific disruption of cell cycle checkpoint functions and its time course after repeated administration of hepatocarcinogens. Thus, rats were repeatedly administered with hepatocarcinogens (methapyrilene, carbadox and thioacetamide), a marginal hepatocarcinogen (leucomalachite green), hepatocarcinogenic promoters (oxfendazole and β-naphthoflavone) or non-carcinogenic hepatotoxicants (promethazine and acetaminophen) for 7, 28 or 90 days, and the temporal changes in cell proliferation, expression of G1/S and spindle checkpoint-related molecules, and apoptosis were examined using immunohistochemistry and/or real-time RT-PCR analysis. Hepatocarcinogens facilitating cell proliferation at day 28 of administration also facilitated cell proliferation and apoptosis at day 90. Hepatocarcinogen- or hepatocarcinogenic promoter-specific cellular responses were not detected by immunohistochemical single molecule analysis even after 90 days. Expression of Cdkn1a, Mad2l1, Chek1 and Rbl2 mRNA also lacked specificity to hepatocarcinogens or hepatocarcinogenic promoters. In contrast, all hepatocarcinogens and the marginally hepatocarcinogenic leucomalachite green induced Mdm2 upregulation or increase in the number of phosphorylated MDM2(+) cells from day 28, irrespective of the lack of cell proliferation facilitation by some compounds. However, different Tp53 expression levels suggest different mechanisms of induction or activation of MDM2 among hepatocarcinogens. On the other hand, hepatocarcinogenic methapyrilene and carbadox downregulated the number of both ubiquitin D(+) cells and proliferating cells remaining in M phase at day 28 and/or day 90, irrespective of the lack of cell proliferation facilitation in the latter. These results suggest that hepatocarcinogens disrupt spindle checkpoint function after 28 or 90 days of administration, which may be induced ahead of cell proliferation facilitation.
authors.affiliation: Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology.
authors.forename: Masayuki
authors.initials: M
authors.lastname: Kimura
authors.forename: Sayaka
authors.initials: S
authors.lastname: Mizukami
authors.forename: Yousuke
authors.initials: Y
authors.lastname: Watanabe
authors.forename: Yasuko
authors.initials: Y
authors.lastname: Hasegawa-Baba
authors.forename: Nobuhiko
authors.initials: N
authors.lastname: Onda
authors.forename: Toshinori
authors.initials: T
authors.lastname: Yoshida
authors.forename: Makoto
authors.initials: M
authors.lastname: Shibutani
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D001562:Benzimidazoles / Q000008:administration & dosage / Q000633:toxicity; D002218:Carbadox / Q000008:administration & dosage / Q000633:toxicity*; D002273:Carcinogens / Q000008:administration & dosage* / Q000633:toxicity*; D049109:Cell Proliferation / Q000187:drug effects* / Q000235:genetics; D000071877:Checkpoint Kinase 1; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D008099:Liver / Q000166:cytology*; D059566:M Phase Cell Cycle Checkpoints / Q000187:drug effects*; D008297:Male; D008701:Methapyrilene / Q000008:administration & dosage / Q000633:toxicity*; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D011916:Rats, Inbred F344; D012394:Rosaniline Dyes / Q000633:toxicity; D013853:Thioacetamide / Q000633:toxicity*; D013997:Time Factors; D014452:Ubiquitins / Q000235:genetics / Q000378:metabolism
chemical_list: D001562:Benzimidazoles; D002273:Carcinogens; C493958:Cdkn1a protein, rat; D050759:Cyclin-Dependent Kinase Inhibitor p21; D012394:Rosaniline Dyes; C109184:UBD protein, human; D014452:Ubiquitins; D013853:Thioacetamide; C015320:leucomalachite green; D008701:Methapyrilene; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C000605871:Chek1 protein, rat; D002218:Carbadox; C011030:oxfendazole