靶向抑制基因DENN-SV诱导人乳腺癌细胞系MCF-7凋亡与NF-κB的关系  

Relationship between NF-kappa B and human breast cancer cell line MCF-7 apoptosis induced by targeting inhibiting DENN-SV gene

作　　者：李锦成[1] 骆振华[1] 

机构地区：[1]辽宁医学院附属第一医院乳腺外科,辽宁锦州121000

出　　处：《中国妇幼保健》2014年第6期953-957,共5页Maternal and Child Health Care of China

基　　金：辽宁省科技厅社会发展攻关计划资助课题〔2010225034〕

摘　　要：目的:研究DENN-SV基因在人乳腺癌细胞系MCF-7中抗凋亡作用的可能机制。方法:将前期试验筛选出的含最佳靶点序列的菌株扩增并抽提质粒,将实验分为空白组、空白载体转染组及最佳靶点组。将实验分为两组,每组分为空白组、空白载体转染组及最佳靶点组,一组转染后48h给予TRAIL 25 ng/ml处理4 h,另一组不予TRAIL处理。收集细胞,采用流式细胞技术检测细胞凋亡率,采用Elisa法测定细胞裂解液中NF-κB含量,采用EMSA法检测核蛋白样品中NF-κB活性。结果:流式细胞仪检测TRAIL诱导凋亡效果增强,与空白组、阴性对照组相比较差异具有统计学意义(P<0.05);Elisa检测NF-κB结果示实验组的NF-κB表达量显著降低差异有统计学意义(P<0.05),EMSA检测NF-κB结果示实验组的NF-κB活性减弱差异有统计学意义(P<0.05)。结论:干扰DENN-SV基因的表达后,给予TRAIL处理可引起MCF-7细胞的凋亡率增高。这种现象的机制可能与NF-κB活性有关,且该现象与NF-κB的关系与TRAIL的作用没有直接联系。Objective: To research the possible mechanism of anti- apoptotic effect of DENN- SV gene in human breast cancer cell line MCF- 7. Methods: The strains containing the best targeting sequence screened out were amplified,and then plasmid was extracted. The experiment was divided into blank group,empty vector transfection group and the best target group. The experiment was divided into two groups,each group was divided into blank group,empty vector transfection group and best target group,at 48 hours after transfection, one group was given TRAIL( 25 ng / ml) for 4 hours,and the other group was not given TRAIL treatment,and the cells were collected. The apoptotic rate was detected by flow cytometry; ELISA assay was used to detect the content of NF- κB in cell lysis solution; EMSA assay was used to detect NF- κB activity in nucleoprotein samples. Results: Flow cytometry revealed that TRAIL- induced apoptosis effect enhanced,there was statistically significant difference compared with blank group and negative control group( P < 0. 05); ELISA assay indicated that NF- κB expression level in experimental group decreased significantly( P < 0. 05),EMSA detection showed that NF- κB activity in experimental group decreased significantly( P < 0. 05). Conclusion: In this study,after intervening the expression of DENN- SV gene, TRAIL treatment can lead to increase of apoptotic rate of MCF- 7 cells. The mechanism of this phenomenon may be related to NF- κB activity,and the phenomenon is not directly linked with the roles of NF- κB and TRAIL.

关 键 词：DENN—SV 短发夹RNA RNA干扰 人MCF-7细胞系 NF—κB 

分 类 号：R737.9[医药卫生—肿瘤]