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.
Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, National Institute of Health, Research Triangle Park, NC 27709, USA.
Mari
M
Iida
Colleen H
CH
Anna
Jennifer
J
Hartis
Maribel
M
Bruno
Barbara
B
Wetmore
Joshua R
JR
Dubin
Stella
S
Sieber
Lee
L
Bennett
Michael L
ML
Cunningham
Richard S
RS
Paules
Kenneth B
KB
Tomer
Christopher D
CD
Houle
Alex B
AB
Merrick
Robert C
RC
Sills
Theodora R
TR
Devereux
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