We studied the regulatory effects of the estragen receptorβ(ERβ)gene on the downstream estrogen signal transfection pathway in colon cancer cells and the possible mechanisms involved.A human ERβ gene recombinant expression plasmid,pEGFP-C1-ERβ,was constructed and transfected into the Caco-2 colon cancer cell line,a line with low ERβ gene expression.The expression of ERβ mRNA and protein was detected 72h after transfection.RT-PCR was used to examine the expression levels of the progesterone recepror(PR)gene containing the classic estrogen response element(ERE),the C-fos oncogene containing the AP-1 site(a non-classical ER binding site),the epigenetic modifying genes,such as Dnmt1,Dnmt3a,Dnmt3b,and histone methyltransferase(HMT),and the human mismatch repair gene hMLH1.Methylation-specific PCR was used to detect the changes in the methylated sites of the CpG islands in the promoters of the ERβ,PR,and C-fos genes.The results indicated that the human ERβ gene recombinant expression plasmid pEGFP-C1-ERβ was successfully constructed and transfected into Caco-2 cells.As compared with the control group,the mRNA and protein expression of ERβ gene was increased significantly 72h after the transfection of pEGFP-C1-ERβ into the Caco-2 cells.As compared with the control group,the mRNA expression of the PR,C-fos,Dnmt3a and Dnmt3b genes was increased significantly 72h after the transfection of pEGFP-C1-ERβ into the Caco-2 cells,but the mRNA expression of the Dnmt1,HMT,and hMLH1 genes decreased significantly(P<0.05).As compared with the control group,different degrees of demethylation occurred in the promoters of the ERβ,progesterone receptor(PR),and C-fos oncogene 72h after the transfection of pEGFP-C1-ERβ into the Caco-2 cells.The methylation index of the estrogen signal transfection pathway in Caco-2 cells was decreased significantly following the expression restoration of ERβ gene(P<0.05).It is concluded that the restoration or up-regulation of the ERβ gene in Caco-2 cells may significantly activate the expre
