Expression And CpG Island Methylation Of SOX7Gene In Esophageal Squamous Cell Carcinoma

Objective: There is a high incidence of esophageal squamous cellcarcinoma (ESCC) in China. Due to the dormant clinical symptoms, mostpatients are diagnosed at middle or advanced stages with poor prognosis. Dueto the high incidence and poor survival, ESCC is one of the major healthissues in China. Therefore, to explore the pathogenesis of esophagealcarcinoma and to find the effective target of gene therapy is an important wayto reduce the incidence and mortality of esophageal cancer.Esophageal cancer is a multi-factor, multi-phase, and multiple geneticvariation involved process. Many oncogenes, tumor suppressor genes andprotein changes are involved in the carcinogenesis of esophageal cancer. Geneepigenetics modification is one of the pathogenesis of cancer, and as a majorepigenetic changes, DNA methylation may play important roles in thepathogenesis of esophageal cancer. Studies have shown that abnormalexpressions of SOX gene family are present in many tumors, and closelylinked with Wnt/β-catenin signal pathway. Sequence search found multipleCpG islands in the promoter region and exon1area of SOX7; suggestingaberrant DNA methylation may be an important epigenetic change of this genein tumors. In the present study, we detected SOX7methylation status in theesophageal cancer cells, ESCC tumor tissues and corresponding paracancerousesophageal squamous epithelial tissueses, analyzed the correlation between thepresence of methylation and expression of SOX7gene, to explore the role ofSOX7gene in ESCC.Method:1Reverse transcription polymerase chain reaction (RT-PCR) method wasused to detect the mRNA level of SOX7gene in esophageal cancer celllines (TE1, EC109, T.TN, Yes-2) treated or untreated with5-aza-dC, ESCC tumor tissues and corresponding paracancerous esophagealsquamous epithelial tissueses.2Immunohistochemical method was used to detect the protein expression ofSOX7in ESCC tumor tissues and corresponding paracancerousesophageal squamous epithelial tissueses.3Methylation specific PCR (MSP) method was used to examine themethylation status of SOX7gene in esophageal cancer cell lines treated oruntreated with5-aza-dC,58ESCC tumor tissues and correspondingparacancerous esophageal squamous epithelial tissueses.4SPSS13.0was applied to analyze the results of experiments.Results:1The relationship between mRNA and protein expression of SOX7andclinical pathological data in ESCC.1.1The mRNA expression of SOX7in ESCCmRNA expression of SOX7in tumor tissues (0.36±0.09) wassignificantly lower than that in corresponding paracancerous esophagealsquamous epithelial tissueses (0.61±0.13)(t=12.772, P<0.05). The mRNAexpression of SOX7was correlated with status of lymph node metastasis,TNM stage and pathological differentiation of ESCC patients (P <0.05).1.2The protein expression of SOX7in ESCCThe protein expression of SOX7in tumor specimens (27.6%,16/58) wassignificantly lower than that in corresponding paracancerous esophagealsquamous epithelial tissueses (72.4%,42/58)(χ2=16.004, P<0.01). The proteinexpression of SOX7was significantly associated with lymph node metastasisof ESCC (P<0.05), however, it was not associated with pathologicaldifferentiation and TNM stage (P>0.05).2The mechanism of the low expression of SOX7in ESCC2.1The mRNA expression of SOX7in esophageal cancer cell lines treated oruntreated with5-aza-dCmRNA expression of SOX7gene was only detected in TE1cell line ofthe four cell lines. After the cell lines were treated with5-aza-dC, mRNA expression of SOX7can be detected in the four cell lines.2.2The methylation status of SOX7in esophageal cancer cell lines treated oruntreated with5-aza-dCSOX7gene was fully methylated in EC109, T.T N, and Yes-2cell lines,while semi-methylation of SOX7was dected in TE1cells. After treatmentwith5-aza-dC, SOX7gene was detected unmethylation status in the fourtreated cell lines.2.3Relationship between methylation status of SOX7and clinicalpathological data in ESCCThe promoter methylation frequency of SOX7in tumor specimens(53.4%,31/58) was significantly higher than that in correspondingparacancerous esophageal squamous epithelial tissueses (29.3%,17/58)(χ2=8.652, P<0.05). Methylation frequencies of SOX7gene in poordifferentiation group (71.2%,23/31) was much higher than that in moderateand high differentiation group (29.6%,8/27)(χ2=11.518, P<0.05). Methylationfrequencies of SOX7in lymph node metastasis group (72.4%,21/29) wassignificantly higher than that in non lymph node metastasis group (22.6%,10/29)(χ2=8.385, P<0.05). Methylation frequencies of SOX7in TNM stagesIII and IV group (82.4%,14/17) was higher than that in stages I and II group(41.5%,17/41)(χ2=8.074, P<0.05). Methylation status of SOX7gene was notassociated with TNM stage, gender, smoking, and drinking (P>0.05).2.4Relationship between methylation status of SOX7and its expressionThe mRNA expression of SOX7in tumor tissues with promoterhypermethylation of the gene (0.28±0.08) was significantly lower than that intumor tissues with unmethylation of the gene (0.41±0.08)(t=-6.249, P<0.05).The positive protein expression of SOX7in tumor tissues with promoterhypermethylation of the gene (12.9%,4/31) was significantly lower than thatin tumor tissues with unmethylation of the gene (44.4%,12/27)(χ2=7.187,P<0.05).Conclusions:1The mRNA and protein expression of SOX7gene in ESCC is lower than that in corresponding paracancerous esophageal squamous epithelialtissueses, suggesting that SOX7gene may be a candidate tumor suppressorgene in ESCC, and its decreased expression may be one of thepathogenesis of ESCC.2The decreased expression and hypermethylation of SOX7gene in most ofthe esophageal cancer cell lines, and the reverse of the expression after thecell lines were treated with5-aza-dC, indicating aberrant DNAmethylation may be one of the mechanisms that lead to the decreasedexpression of SOX7in esophageal cancer.3Methylation frequency of SOX7in ESCC tumor tissues was significantlyhigher than that in corresponding paracancerous esophageal squamousepithelial tissueses, suggesting hypermethylation may be one of themechanisms in the pathogenesis of ESCC.4The close correlation between SOX7promoter hypermethylation statusand lymph node metastasis, pathological differentiation and TNM stage,indicating promoter hypermethylation of SOX7may be associated withthe development of ESCC, and may be act as an effective target for theprognosis of ESCC patients.