Changes in global gene and protein expression during early mouse liver carcinogenesis induced by non-genotoxic model carcinogens oxazepam and Wyeth-14,643.
We hypothesized that the mouse liver tumor response to non-genotoxic carcinogens would involve some common early gene and protein expression changes that could ultimately be used to predict chemical hepatocarcinogenesis. In order to identify a panel of genes to test, we analyzed global differences in gene and protein expression in livers from B6C3F1 mice following dietary treatment with two rodent carcinogens, the benzodiazepine anti-anxiety drug oxazepam (2500 p.p.m.) and the hypolipidemic agent Wyeth (Wy)-14,643 (500 p.p.m.) compared with livers from untreated mice. Male mice were exposed for 2 weeks and 1, 3 or 6 months to oxazepam or Wy-14,643 in an age-matched study design. By histopathological evaluation, no liver preneoplastic foci or tumors were detected at 6 months in treated or control groups. By cDNA microarray analysis [NIEHS Mouse Chip (8700 genes); n = 3 individual livers/group, four hybridizations/sample], expression of 36 genes or 220 genes were changed relative to control livers following 6 months of oxazepam or Wy-14,643 treatment, respectively. To obtain a more comprehensive picture of gene/protein expression changes, we also conducted a proteomics study by 2D-gel electrophoresis followed by matrix assisted laser desorption/ionization-mass spectrometry on cytoplasmic, nuclear, and microsomal subcellular fractions of the same liver samples utilized for the cDNA microarray analysis. Real-time PCR, western blot analysis and immunohistochemistry were utilized for validation and to expand the results to other time points. Cyp2b20, growth arrest- and damage-inducible gene beta (Gadd45beta), tumor necrosis factor alpha-induced protein 2 and insulin-like growth factor binding protein 1 (Igfbp5) genes and proteins were upregulated by oxazepam, and Cyp2b20, Cyclin D1, proliferating cell nuclear antigen, Igfbp5, Gadd45beta and cell death-inducing DNA fragmentation factor alpha subunit-like effector A exhibited higher expression after Wy-14,643 treatment. Most of these genes/proteins were also deregulated at 2 weeks. There appeared to be more distinct than common changes in the expression of carcinogenesis-related genes/proteins between the two compounds, suggesting that the major carcinogenic pathways are different for these compounds and may be distinct for different chemical classes.
Iida
Mari
M
Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, National Institute of Health, Research Triangle Park, NC 27709, USA.
Anna
Colleen H
CH
Hartis
Jennifer
J
Bruno
Maribel
M
Wetmore
Barbara
B
Dubin
Joshua R
JR
Sieber
Stella
S
Bennett
Lee
L
Cunningham
Michael L
ML
Paules
Richard S
RS
Tomer
Kenneth B
KB
Houle
Christopher D
CD
Merrick
Alex B
AB
Sills
Robert C
RC
Devereux
Theodora R
TR
D000818:Animals; D001483:Base Sequence; D002273:Carcinogens; D017931:DNA Primers; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D008114:Liver Neoplasms, Experimental / Q000139:chemically induced* / Q000235:genetics; D008297:Male; D051379:Mice; D010076:Oxazepam / Q000633:toxicity*; D016133:Polymerase Chain Reaction; D011743:Pyrimidines / Q000633:toxicity*
D002273:Carcinogens; D017931:DNA Primers; D011743:Pyrimidines; D010076:Oxazepam; C006253:pirinixic acid