5-氮杂-2’-脱氧胞苷对肺癌SPC—A1细胞和非小细胞肺癌组织中抑癌基因甲基化的作用  

Effects of 5-Aza-2-deoxycytidine on DNA methylation of anti-oncogenes in non-small cell lung cancer cells

作　　者：方汉林[1] 于在诚[1] 祝会斌 金永堂[3] 

机构地区：[1]安徽医科大学第一附属医院胸外科,合肥230022 [2]合肥第二人民医院胸外科 [3]浙江大学医学院公共卫生学院

出　　处：《中华肿瘤杂志》2012年第9期658-663,共6页Chinese Journal of Oncology

基　　金：国家自然科学基金（30471427）

摘　　要：目的观察分泌型卷曲相关篮白1（SFRP1）基凶在非小细胞肺癌（NSCLC）组织中的甲基化状态，研究抑甲基化制剂5-氮杂-2’-脱氧胞苷（5-Aza—CdR）对肺癌SPC—A1细胞中SFRP1、p16和O^6-甲基鸟嘌呤-DNA甲基转移酶（MGMT）基因甲基化的影响。方法采用甲基化特异性PCR（MSP）法和免疫组化sP法，检测60例NSCLC组织中SFRPl基因甲基化的状态和蛋白的表达，以21例肺良性病变组织作为对照。以5-Aza—CdR处理肺癌SPC—A1细胞，采用MSP法和RT—PCR法检测处理前后细胞中SFRP1、p16和MGMT基因的甲基化状态及相应mRNA的表达。结果SFRP1基因在60例NSCLC组织中的甲基化率为58．3％，明显高于肺良性病变组织（14．3％；X2=12．118，P=0．001）；SFRP1基因的甲基化与NSCLC的分化程度和淋巴结转移情况有关（均P〈0．05）。SFRP1蛋白在35例SFRPI基因甲基化的NSCLC组织中的阴性表达率为68．6％，明显高于非甲基化的NSCLC组织（24．0％；x2=9．613，P=0．002）；SFRPl蛋白的阴性表达与NSCLC的分化程度、临床分期以及淋巴结转移情况有关（均P〈0．05）。末用5-Aza—CdR处理时，SPC—A1细胞中SFRPI、p16和MGMT基因甲基化和mRNA的表达甚很低；应用不同浓度的5-Aza—dCR处理后，SPC—A1细胞中SFRP1、p16和MGMT基凶甲基化和mRNA的表达均显著上调（均P〈0．05）。结论SFRPl基因的甲基化与NSCI．C的发生发展密切相关；5-Aza—CdR能逆转SFRPI、p16和MGMT基凶的甲基化状态，促进其重新表达。Objective To observe the expression of SFRP1 gene methylation in non-small cell lung cancer （NSCLC）, and study the effect of 5-Aza-2 -deoxycytidine （5-Aza-CdR） on DNA methylation and expression of SFRP1, p16 and MGMT genes in the human lung cancer cell line SPC-A-1 cells. Methods SP immunohistochemistry and methylation-specific PCR were used to detect the SFRP1 methylation in 60 NSCLC cases, and 21 cases of benign lung diseases were used as control group. SPC-A-1 cells were cultured and treated with 5-Aza-CdR. The promoter methylation status of SFRP1, p16 and MGMT genes were detected by methylation-specific polymerase （MSP） chain reaction, and mRNAs were detected by real-time PCR. Results The positive rate of SFRP1 gene methylation in NSCLC was signifcantly higher than that in normal X2 lung tissue （58.3% vs. 14.3% ; = 12.118 ,P=0.001 ）. SFRP1 gene methylation was closely correlated with lymph node metastasis and degree of differentiation in NSCLC （ P 〈 0.05 ）. SFRP1 protein expression was correlated with clinical stage, degree of differentiation and lymph node metastasis in NSCLC （P 〈 0.05 ）. The positive expression of SFRP1 protein in 30 cases of NSCLC tissue containing SFRP1 gene methylation was significantly higher than that in non-methylated NSCLC （68.6% vs. 24.0% ; X2=9. 613, P = 0.002）. SFRPI gene methylation was closely correlated with SFRP1 gene protein expression in NSCLC （P 〈 0.05 ）. Negative expression of SFRP1 protein was correlated with the differentiation, clinical stage, and lymph node metastasis in NSCLC （ all P 〈 0.05 ）. Without 5-Aza-CdR treatment, the expressions of methylation of SFRP1, p16 and MGMT genes and their mRNA were low. After 5-Aza-CdR treatment at different concentrations, their expressions were significantly elevated （ all P 〈 0.05 ）. Conclusions SFRP1 gene methylation is closely associated with carcinogenesis and development of NSCLC. 5-Aza-CdR may reverse the methylation of SFRP1, p16 and MGMT genes, and facilitate the re-expres

关 键 词：5-氮杂-2’-脱氧胞苷 癌 非小细胞肺 SPC—A1细胞 分泌型卷曲相关蛋白1 DNA甲基化 

分 类 号：R734.2[医药卫生—肿瘤]