食管鳞癌RASSF1A基因启动子区甲基化研究  

Hypermethylation Status of RASSF1A Gene Promoter Region in Esophageal Squamous Cell Carcinoma

作　　者：丁士刚[1] 吴笛[2] 邹检平[3] 邢秀娟[4] 鲁凤民[4] 陈香梅[4] 

机构地区：[1]北京大学第三医院消化科,北京100083 [2]广州市红十字会医院中医科,广州510220 [3]沈阳药科大学神经病理学系,沈阳110016 [4]北京大学医学部病原生物学系,北京100083

出　　处：《中国微创外科杂志》2007年第12期1213-1216,共4页Chinese Journal of Minimally Invasive Surgery

基　　金：国家自然科学基金主任基金(No.30540081)

摘　　要：目的研究肿瘤抑制基因RASSF1A启动子区甲基化所致该基因表达抑制在我国食管鳞癌发生中的作用及可能机制。方法用甲基化特异PCR技术(MSP)检测43例原发性食管鳞癌标本及6例对照食管上皮组织标本、食管鳞癌细胞系TE11和TE12中RASSF1A启动子甲基化状态;逆转录(RT)-PCR法检测5-氮-2′-脱氧胞苷(5-Aza-CdR)处理前后食管鳞癌细胞系中RASSF1A mRNA表达水平;Western blot方法检测食管鳞癌细胞系中细胞微管蛋白的表达。结果20.9%(9/43)原发性食管鳞癌标本有RASSF1A启动子超甲基化,正常食管上皮组织未发现该基因超甲基化改变。RASSF1A基因甲基化与食管癌患者的性别和TNM分期无关(P>0.05),但在不同年龄组间的差异有统计学意义(P<0.05)。TE12细胞RASSF1A启动子发生甲基化,RT-PCR检测不到RASSF1A mRNA。TE11细胞系启动子未发生甲基化,RT-PCR检测其有RASSF1A mRNA表达。5-Aza-CdR处理可使TE12重新表达RASSF1A mRNA。TE12细胞的β-微管蛋白水平比TE11细胞低87.8%。结论原发性食管鳞癌存在RASSF1A启动子超甲基化,该基因甲基化与年龄相关。RASSF1A在食管鳞癌细胞系表达抑制与其甲基化状态有关。RASSF1A表达缺失可能导致β-微管蛋白减少。Objective To study the involvement of promoter region hypermethylation induced gene silencing of tumor suppressor gene RASSF1A in the tumorigenesis of esophageal squamous cell carcinoma(ESCC).Methods Methylation specific PCR(MSP) was employed to test the methylation status of RASSF1A promoter in 43 primary ESCC specimens,6 samples of normal esophageal epithelial cells,and 2 ESCC cell lines,TE11 and TE12.The expression of RASSF1A was tested by RT-PCR before and after the cells being treated with de-methylation reagent 5-Aza-CdR.Western blot was carried out to detect the expression of β-tubulin in TE11 and TE12 cell lines.Results Hypermethylation of the RASSF1A promoter was detected in 9 of the 43 ESCC specimens(20.9%),while it is found in none of the normal esophageal epithelial cells.The methylation status of RASSF1A promoter showed no correlation with gender and TNM stage(P>0.05),but was significantly different among different age groups(P<0.05).RT-PCR demonstrated the expression of RASSF1A in un-methylated TE11,but failed to detect it in TE12,which was hypermethylated.After being treated with 5-Aza-CdR in vitro for 96 hours,RASSF1A mRNA was re-expressed in TE12.The level of β-tubulin in TE12 was lower than that in TE11 by 87.8%. Conclusions Hypermethylation of the RASSF1A promoter,which is correlated with age,can be found in ESCC tissues.The gene silencing of RASSF1A is related to the hypermethylation,and may lead to a decreased β-tubulin level.

关 键 词：食管鳞癌 甲基化 RASSF1A Β-微管蛋白 抑癌基因 

分 类 号：R735.1[医药卫生—肿瘤] R-33[医药卫生—临床医学]