Differential Gene Expression in Estradiol-3-Benzoate-Treated Liver and Chemically-Induced Hepatocellular Carcinoma

In a previous study by the current authors, hepatocellular carcinoma (HCC) was determined to be epidemiologically sex-dependent, and the incidence and multiplicity of HCC found to decrease in estradiol-3 benzoate (EB)-treated F344 rats. Therefore, to ascertain the anticancer mechanism of EB, a commercially available cDNA microarray, with a total of 14,815 cDNA rat gene clones, was used to determine the differentially expressed genes in nontreated livers, EB-treated livers, and diethynitrosolamine (DEN)-induced HCC. In the sequenced experiment, a total of 85 genes were differentially expressed at either two or more times the rate of the normal expression, where 33 genes were downregulated by EB, and 52 genes upregulated. Candidate genes were selected according to significant changes observed in the mRNA expression in the EB-treated livers compared with the nontreated livers, then these genes were filtered according to their different expression patterns in the DEN-induced tumors compared to the estrogen-treated livers. To confirm the microarray data, a real-time PCR analysis was performed for ten selected genes: the H-ras revertant protein 107 (H­rev107), insulin-like growth factor binding protein (lOFBP), parathyroid hormone receptor (PI'HR), SH3 domain binding protein (SH3BP), metallothionein, src-suppressed C-kinase substrate (SSeCK) gene, phosphodiesterase I, CD44, epithelial membrane protein 3 (EMP3), and estrogen receptor a (ERa). The SSeCK and phosphodiesterase I genes were both upregulated in the DEN-induced hepatocarcinomas, yet their possible carcinogenic functions remain unknown. Meanwhile, the other genes were downregulated, including the genes related to growth regulation (IOFBP, H-revI07, ER$\alpha$), adipogenesis inhibition (PTHR), and tumor suppression (metallothionein).