Disruptive cell cycle regulation involving epigenetic downregulation of Cdkn2a (p16(Ink4a)) in early-stage liver tumor-promotion facilitating liver cell regeneration in rats.
Cell cycle aberration was immunohistochemically examined in relation to preneoplastic liver cell foci expressing glutathione S-transferase placental form (GST-P) at early stages of tumor-promotion in rats with thioacetamide (TAA), a hepatocarcinogen facilitating liver cell regeneration. Immunoexpression of p16(Ink4a) following exposure to other hepatocarcinogens/promoters and its DNA methylation status were also analyzed during early and late tumor-promotion stages. GST-P(+) liver cell foci increased cell proliferation and decreased apoptosis when compared with surrounding liver cells. In concordance with GST-P(+) foci, checkpoint proteins at G(1)/S (p21(Cip1), p27(Kip1) and p16(Ink4a)) and G(2)/M (phospho-checkpoint kinase 1, Cdc25c and phospho-Wee1) were either up- or downregulated. Cellular distribution within GST-P(+) foci was either increased or decreased with proteins related to G(2)-M phase or DNA damage (topoisomerase IIα, phospho-histone H2AX, phospho-histone H3 and Cdc2). In particular, p16(Ink4a) typically downregulated in GST-P(+) foci and regenerative nodules at early tumor-promotion stage with hepatocarcinogens facilitating liver cell regeneration and in neoplastic lesions at late tumor-promotion stage with hepatocarcinogens/promoters irrespective of regenerating potential. Hypermethylation at exon 2 of Cdkn2a was detected at both early- and late-stages. Thus, diverse disruptive expression of G(1)/S and G(2)/M proteins, which allows for clonal selection of GST-P(+) foci, results in the acquisition of multiple aberrant phenotypes to disrupt checkpoint function. Moreover, increased DNA-damage responses within GST-P(+) foci may be the signature of genetic alterations. Intraexonic hypermethylation may be responsible for p16(Ink4a)-downregulation, which facilitates cell cycle progression in early preneoplastic lesions produced by repeated cell regeneration and late-stage neoplastic lesions irrespective of the carcinogenic mechanism.
Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan. k1_ gemini@nifty.com
Takuma
T
Tsuchiya
Liyun
L
Wang
Atsunori
A
Yafune
Masayuki
M
Kimura
Takumi
T
Ohishi
Kazuhiko
K
Suzuki
Kunitoshi
K
Mitsumori
Makoto
M
Shibutani
D000818:Animals; D002453:Cell Cycle / Q000502:physiology*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D044127:Epigenesis, Genetic; D020869:Gene Expression Profiling; D051549:Glutathione S-Transferase pi / Q000378:metabolism*; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology*; D008115:Liver Regeneration / Q000502:physiology*; D008297:Male; D009367:Neoplasm Staging; D012313:RNA / Q000737:chemistry / Q000235:genetics; D051381:Rats; D011916:Rats, Inbred F344; D060888:Real-Time Polymerase Chain Reaction; D013853:Thioacetamide / Q000633:toxicity
D019941:Cyclin-Dependent Kinase Inhibitor p16; D013853:Thioacetamide; D012313:RNA; D051549:Glutathione S-Transferase pi; C496558:Gstp1 protein, rat