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目的:检测人食管鳞癌(esophageal squamous cell carcinoma,ESCC)组织中Ras相关区域家族7(Ras-association domain family 7,RASSF7)基因的mRNA、蛋白表达情况及其甲基化状态,探究RASSF7在ESCC发生发展中的作用。方法:组织标本取自河北医科大学第四医院2011—2012年间手术切除的69例ESCC原发灶组织及癌旁组织。分别应用RT-PCR及甲基化特异性PCR(methylation specific polymerase chain reaction,MSP)方法检测DNA甲基转移酶抑制剂5-氮杂-2’-脱氧胞苷(5-aza-2’-deoxycitydine,5-Aza-dC)处理前后的4株食管癌细胞系(TE13、T.Tn、YES-2、Ec109)和69例病灶组织及其癌旁组织中RASSF7 mRNA表达水平及甲基化状态,应用免疫组织化学方法检测69例ESCC组织及相应癌旁组织中RASSF7的蛋白表达。结果:RASSF7基因在TE13、T.Tn、YES-2细胞系中表达阳性,在Ec109细胞系中表达缺失;经5-Aza-dC处理后,RASSF7在TE13、T.Tn、YES-2细胞中表达下调,在Ec109细胞中表达阳性。5-Aza-dC处理前后4株食管癌细胞系中均未检测到RASSF7的甲基化。人ESCC组织中RASSF7的mRNA相对表达量(0.63±0.08 vs 0.42±0.20,P<0.01)与蛋白表达阳性率[81.2%(56/69)vs 53.6%(37/69),P<0.01]均显著高于相应癌旁组织,且均与患者的淋巴结转移情况及分化程度有关(P<0.05或P<0.01),与TNM分期、年龄和性别无关(均P>0.05)。ESCC组织和相应癌旁组织中均未检测到RASSF7的甲基化。结论:4株食管癌细胞系、人ESCC组织和癌旁组织中RASSF7基因的表达差异与RASSF7本身甲基化状态无关,ESCC组织中RASSF7的高表达可能参与了ESCC的发生及转移。
OBJECTIVE: To detect the mRNA and protein expression of Ras-associated domain family 7 (RASSF7) gene and its methylation status in esophageal squamous cell carcinoma (ESCC) tissues. To explore the role of RASSF7 in ESCC The development of the role. Methods: Tissue samples were obtained from 69 ESCC primary tumor tissues and adjacent tissues excised from 2011 to 2012 in the Fourth Hospital of Hebei Medical University. RT-PCR and methylation specific polymerase chain reaction (MSP) were used to detect the DNA methyltransferase inhibitor 5-aza-2’-deoxycitydine The expression of RASSF7 mRNA and methylation status in four esophageal cancer cell lines (TE13, T.Tn, YES-2, Ec109) and 69 lesions and adjacent normal tissues before and after treatment with 5-Aza- Immunohistochemistry was used to detect the protein expression of RASSF7 in 69 ESCC tissues and corresponding paracancerous tissues. Results: The expression of RASSF7 gene was positive in TE13, T.Tn and YES-2 cell lines and was absent in Ec109 cell line. After treatment with 5-Aza-dC, RASSF7 was expressed in TE13, T.Tn and YES-2 cells The expression was down-regulated and was positive in Ec109 cells. No methylation of RASSF7 was detected in the four esophageal cancer cell lines before and after 5-Aza-dC treatment. The positive expression rate of RASSF7 mRNA in human ESCC tissues was significantly higher than that in normal human ESCC tissues (0.63 ± 0.08 vs 0.42 ± 0.20, P <0.01) and positive rate of protein expression (81.2% (56/69) vs 53.6% (37/69), P <0.01] (P <0.05 or P <0.01), which were not related to TNM staging, age and gender (all P> 0.05). No methylation of RASSF7 was detected in ESCC tissues and corresponding paracancerous tissues. CONCLUSION: The expression of RASSF7 gene in four esophageal cancer cell lines, human ESCC tissues and paracancerous tissues is not related to the methylation status of RASSF7 itself. The high expression of RASSF7 may be involved in the occurrence and metastasis of ESCC.