CAJ | 학술논문

目的检测前列腺癌中N-myc下游调节基因-1(N-myc downstream regulated gene-1,NDRG1)启动子区的甲基化状态,探讨甲基化转移酶抑制剂5-氮杂胞苷对NDRG1基因在前列腺癌细胞mRNA的表达及对细胞增殖的影响.方法 2013年1月至2014年4月采用亚硫酸盐测序PCR法分别检测前列腺癌组织、良性前列腺增生(BPH)组织、前列腺癌细胞系(PC3、22RV1、LNCaP、DU145)和人正常前列腺细胞系RWPE-1中的NDRG1基因启动子区甲基化状态.用10 μmol/L 5-氮杂胞苷分别作用于LNCaP和DU145细胞72 h后,噻唑盐法分析5-氮杂胞苷对LNCaP和DU145细胞增殖的影响;RT-PCR法检测两种细胞系中NDRG1 mRNA的表达.结果 NDRG1基因在前列腺癌细胞系PC-3、22RV1、LNCaP和DU145中甲基化率分别为(24.8±3.3)％、(36.2±2.5)％、(48.6±2.8)％、(69.5±1.7)％,人正常前列腺细胞系RWPE-1甲基化率为(4.8±4.5)％;前列腺癌组织中为(48.6±5.3)％,BPH组织中为(4.3±2.1)％,组间比较差异均有统计学意义(P＜0.05).10 μmol/L5-氮杂胞苷处理LNCaP和DU145细胞72 h后,两种细胞中NDRG1基因发生了去甲基化,mRNA表达水平较处理前增强8～9倍,细胞生长受到抑制(P＜0.05).结论 NDRG1基因启动子区的高甲基化是其在前列腺癌中异常表达的原因之一,5-氮杂胞苷能逆转NDRG1基因的甲基化状态,调控该基因mRNA表达,并能抑制前列腺癌细胞的增殖.
목적 검측전렬선암중N-myc하유조절기인-1(N-myc downstream regulated gene-1,NDRG1)계동자구적갑기화상태,탐토갑기화전이매억제제5-담잡포감대NDRG1기인재전렬선암세포mRNA적표체급대세포증식적영향.방법 2013년1월지2014년4월채용아류산염측서PCR법분별검측전렬선암조직、량성전렬선증생(BPH)조직、전렬선암세포계(PC3、22RV1、LNCaP、DU145)화인정상전렬선세포계RWPE-1중적NDRG1기인계동자구갑기화상태.용10 μmol/L 5-담잡포감분별작용우LNCaP화DU145세포72 h후,새서염법분석5-담잡포감대LNCaP화DU145세포증식적영향;RT-PCR법검측량충세포계중NDRG1 mRNA적표체.결과 NDRG1기인재전렬선암세포계PC-3、22RV1、LNCaP화DU145중갑기화솔분별위(24.8±3.3)％、(36.2±2.5)％、(48.6±2.8)％、(69.5±1.7)％,인정상전렬선세포계RWPE-1갑기화솔위(4.8±4.5)％;전렬선암조직중위(48.6±5.3)％,BPH조직중위(4.3±2.1)％,조간비교차이균유통계학의의(P＜0.05).10 μmol/L5-담잡포감처리LNCaP화DU145세포72 h후,량충세포중NDRG1기인발생료거갑기화,mRNA표체수평교처리전증강8～9배,세포생장수도억제(P＜0.05).결론 NDRG1기인계동자구적고갑기화시기재전렬선암중이상표체적원인지일,5-담잡포감능역전NDRG1기인적갑기화상태,조공해기인mRNA표체,병능억제전렬선암세포적증식.
Objective To evaluate the methylation status of prostate cancer NDRG1 gene promoter region,and to explore the influence of methylation inhibitor 5-azacytidine on NDRG1 gene's mRNA expression in prostate cancer cells and its effects on cell proliferation.Methods Bisulfite-sequencing PCR (BSP) were used to detect the NDRG1 gene promoter methylation status in prostate cancer and BPH tissue,prostate cancer cell lines (PC3,22RV1,LNCaP and DU145) and human normal prostate cell line's RWPE-1.After 10 μmol/L 5-azacytidine were used on LNCaP and DU145 cells for 72 h,5-azacytidine's influence on cell proliferation was analyzed with MTT,two prostate cancer cell lines NDRG1 mRNA expressions were detected with RT-PCR.Results The methylation rates of NDRG1 gene in prostate cancer cell lines PC-3,22RV1,LNCaP and DU145 were (24.8 ± 3.3) ％,(36.2 ± 2.5) ％,(48.6 ± 2.8) ％ and (69.5 ± 1.7) ％,respectively.Methylation rate of Human normal prostate cell lines RWPE-1 was (4.8 ± 4.5) ％;prostate carcinoma was (48.6 ± 5.3) ％,BPH tissue was (4.3 ± 2.1) ％.The differences between groups were statistically significant.After 10 μmol/L 5-azacytidine added on LNCaP and DU145 cells for 72 h,NDRG1 gene demethylation occurred in both cells,its mRNA expression enhanced 8-9 times compared with previous and its cell growth was inhibited (P ＜ 0.05).Conclusions NDRG1 gene promoter region's hypermethylation is one of the reasons of its aberrant expression in prostate cancer.5-azacytidine can reverse NDRG1 gene promoter methylation status,regulate the expression of the gene and can inhibit prostate cancer cell proliferation.