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目的:检测miR-6867及其宿主基因Rap型鸟苷酸交换因子1(rap guanine nucleotide exchange factor like 1,RAPGEFL1)在食管癌细胞系及食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)组织中的表达水平及甲基化状态,并探讨其在ESCC发生发展中的作用。方法:选取河北医科大学第四医院2014年1月至2016年1月收治的ESCC手术患者组织标本87例。应用实时荧光定量PCR(qRT-PCR)检测miR-6867及RAPGEFL1在食管癌细胞系和ESCC组织及其相应癌旁组织中的表达水平,分析miR-6867和RAPGEFL1基因表达之间的相关性。应用甲基化特异性PCR法(MSP)检测RAPGEFL1在食管癌细胞系和ESCC组织及其相应癌旁组织中的甲基化状态。结果:miR-6867在ESCC组织中的相对表达量显著低于其相应癌旁组织(P<0.05),并与淋巴结转移及TNM分期有关(P<0.05);RAPGEFL1基因在ESCC组织中的表达显著低于其相应癌旁组织(P<0.05),并与淋巴结转移、组织分化程度及TNM分期相关(P<0.05);miR-6867与RAPGEFL1在ESCC组织中的表达呈明显正相关(P<0.05)。5-氮杂-2’-脱氧胞苷(5-Aza-2’-deoxycytidine,5-Aza-dC)处理后,4种食管癌细胞系中miR-6867和RAPGEFL1基因的表达均增高,并且其甲基化程度明显降低。RAPGEFL1基因在ESCC组织中的甲基化率显著高于其相应癌旁组织(P<0.05),并与淋巴结转移、组织分化程度及TNM分期有关(P<0.05)。结论:ESCC的发生发展可能与miR-6867和RAPGEFL1的异常低表达及RAPGEFL1高甲基化状态有关,miR-6867与RAPGEFL1表达具有一致性,且RAPGEFL1基因启动子区甲基化可能是导致miR-6867与RAPGEFL1表达沉默的机制之一。
Objective: To detect the expression of miR-6867 and its host gene rap guanine nucleotide exchange factor 1 (RAPGEFL1) in esophageal squamous cell carcinoma (ESCC) and esophageal squamous cell carcinoma And its methylation status, and to explore its role in the development of ESCC. Methods: Totally 87 patients with ESCC from January 2014 to January 2016 were selected from the Fourth Hospital of Hebei Medical University. The expression of miR-6867 and RAPGEFL1 in esophageal cancer cell lines and ESCC tissues and corresponding paracancerous tissues was detected by real-time quantitative PCR (qRT-PCR), and the correlation between miR-6867 and RAPGEFL1 gene expression was analyzed. Methylation-specific PCR (MSP) was used to detect the methylation status of RAPGEFL1 in esophageal cancer cell lines and ESCC tissues and their corresponding paracancerous tissues. Results: The relative expression level of miR-6867 in ESCC tissues was significantly lower than that in paracancerous tissues (P <0.05), and correlated with lymph node metastasis and TNM stage (P <0.05). The expression of RAPGEFL1 gene in ESCC tissues was significantly (P <0.05), and was correlated with lymph node metastasis, histological differentiation and TNM stage (P <0.05). There was a significant positive correlation between the expression of miR-6867 and RAPGEFL1 in ESCC tissues (P <0.05) ). After treatment with 5-Aza-2’-deoxycytidine (5-Aza-dC), the expression of miR-6867 and RAPGEFL1 genes were all increased in the four esophageal cancer cell lines The degree of methylation was significantly reduced. The methylation rate of RAPGEFL1 gene in ESCC tissues was significantly higher than that in its corresponding paracancerous tissues (P <0.05), and was related to lymph node metastasis, histological differentiation and TNM stage (P <0.05). Conclusion: The occurrence and development of ESCC may be related to the abnormal low expression of miR-6867 and RAPGEFL1 and the hypermethylation state of RAPGEFL1. The expression of miR-6867 and RAPGEFL1 are consistent, and the promoter methylation of RAPGEFL1 gene may be the result of miR- One of the mechanisms by which RAPGEFL1 silences expression.