Molecular pro filing of genes specifically up-regulated during

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In search of genes steadily up-regulated during the promotion stage in carcinogenesis, suppression PCR subtractive hybridization and following Northern blot screening were performed using a phenobarbital (PB)-promotion model based on a medium-term liver bioassay. Two weeks after an initiation with diethylnitrosamine (DEN), rats were given PB for up to 64 weeks. For comparison, animals given ethinylestradiol (EE) or butylated hydroxytoluene (BHT) in the diet at promotion stage were also included. Rats were subjected to partial hepatectomy (PH) at week 3.In addition, dose-dependence of PB at week 8 of promotion and responsiveness to representative non-genotoxic carcinogens without DEN-initiation were examined. Fragments of a total of 67 different genes were isolated from the up-regulated gene population in the liver at day 10 of PB-treatment by subtracting from basal expression of DEN+PH alone. By Northern blot screening for signal-detectable 48 genes, 16 genes showed up-regulation in th … More e livers at week 8 of promotion, common to the PB-and EE-treatments as compared to the basal expression of unpromoted liver. The majority of these genes were also up-regulated at week 8 by BHT-treatment, and were also constitutively expressed in the DEN(-), PH(-)-untreated rat livers. Among the up-regulated genes common to the PB- and EE-promotion, and not responding to the non-genotoxic carcinogens in uninitiated liver, the following 6 genes showed overexpression in PBpromoted hepatocellular carcinomas at week 64 ; ubiquitously expressed mammalian ABC half transporter, apolipoprotein A4 (APOA4), nuclear receptor binding factor-2, CD81, hypothetical protein (HSPC014), and one unidentified gene. These genes might be candidates for biomarkers in screening of non-genotoxic hepatocarcinogens by analysis in two-stage carcinogenesis models. In addition, in situ localization of APOA4mRNA-signal was observed specific to GST-P-positive foci in the thioacetamide-promoted livers. Although further study is required on the role in the hepatocarcinogenesis, APOA4 may link to the formation of neoplastic lesions. Less