耐药乳腺癌MCF-7/Adr细胞NF-κB的激活与耐药的关系

Relationship between NF-κB activity and drug resistance in MCF-7/Adr human breast cancer cell

目的 :观察耐药MCF 7/Adr和其药敏亲本系MCF 7乳腺癌细胞中核因子κB(nuclearfactorkappaB ,NF κB)的激活水平 ,以及其抑制剂二硫代氨基甲酸吡咯烷 (pyrrolidinedithiocarbamate,PDTC)对耐药的逆转效应 ,探讨NF κB在耐药调控中的作用 .方法 :凝胶电泳迁移率法 (electrophoreticmobilityshiftassay ,EMSA)测定NF κB的DNA结合活性 .免疫印迹杂交法 (Westernblot)检测多药耐药基因 (MDR1 )产物P 糖蛋白 (P pg)的表达 .MTT法测定阿霉素 (Doxorubicin ,DOX)作用上述细胞的药物半数抑制浓度值 (IC50 ) .结果 :耐药MCF 7/Adr细胞中NF κB的DNA结合活性 (积分吸光度 :2 6 7± 9)显著高于MCF 7细胞中NF κB的DNA结合活性 (积分吸光度 :34± 2 ,P <0 .0 1 ) .DOX单独作用于MCF 7/Adr,IC50 为 (37.4± 2 .1 ) μmol/L ,与PDTC共同孵育后DOX的IC50值则显著下降为 (6 .8± 0 .8) μmol/L(P <0 .0 5 ) .抑制NF κB的激活不影响P pg的表达 (P >0 .0 5 ) .结论 :与其亲本药敏细胞相比较 ,MCF 7/Adr中的NF κB呈高水平的激活状态 .抑制NF κB的激活可以部分逆转MCF 7/Adr的DOX抵抗 ,提示NF

AIM: To investigate the activation level of nuclear transcription factor NF κB in MCF 7/Adr and MCF 7 human breast cancer cells and to explore the effects of the activity of NF κB in resistance against doxorubicin. METHODS: Electrophoretic motility shift assay (EMSA) was performed to examine the NF κB DNA binding activity of multi drug resistant MCF 7/Adr cell line and its parental cell line MCF 7. The protein expression of P gp was measured by Western blot and the IC 50 value of doxorubicin was evaluated by MTT assay. RESULTS: MCF 7/Adr cell line was shown to have a significantly higher activation level of NF κB (267±9) than its parent cell line MCF 7 (34±2, P < 0.01). The IC50 value of doxorubicin was (37.4±2.1) μmol/L in MCF 7/Adr cells, which decreased to (6.8± 0.8) μmol/L ( P <0.05) after coincubation with PDTC (50 μmol/L). When NF κB activation was inhibited with PDTC, the protein expression of P gp had no marked change. CONCLUSION: The inhibition of NF κB activation may partly reverse the resistance of MCF 7/Adr against doxorubicin, suggesting that NF κB may be involved in the mechanism of drug resistance in breast cancer cells.