细胞因子诱导的杀伤细胞体外逆转K562/ADR细胞株多药耐药性观察  

Experimental research of CIK cells reverse multi-drug resistance in vitro

作　　者：蒋东霞[1] 何琳[1] 徐虹[1] 胡杰英[1] 买玲[1] 宋永平[1] 杨萍[2] 

机构地区：[1]河南省肿瘤研究所医学生物实验室,郑州450003 [2]河南省职工医学院生物化学教研室,郑州451191

出　　处：《郑州大学学报（医学版）》2008年第5期905-908,共4页Journal of Zhengzhou University(Medical Sciences)

基　　金：河南省科技厅专项基金资助项目341700900

摘　　要：目的:研究细胞因子诱导的杀伤细胞(CIK)体外逆转K562/ADR细胞株多药耐药(MDR)的可行性。方法:在含细胞因子(人重组粒细胞-巨噬细胞集落刺激因子、人重组白细胞介素-1、人重组白细胞介素-4和人重组肿瘤坏死因子-α)的RPMI1640完全培养液中诱导K562细胞株、K562/ADR耐药细胞株分化成树突状细胞(DC)。正常人外周血单个核细胞(PBMC)在上述细胞因子诱导下培养成CIK,与成熟的DC共培养成CIK+K562/ADR-DC。应用流式细胞术检测CIK及CIK+K562/ADR-DC的细胞表型。MTT法测定CIK和K562/ADR-DC对K562/ADR的细胞毒作用,流式细胞仪检测CIK组和CIK+K562/ADR-DC组细胞中糖蛋白(P-gp)、阿霉素(ADR)的含量,RT-PCR检测mdr-1基因表达。结果:CIK、CIK+K562/ADR-DC细胞经流式细胞仪检测,均具有自己独特的表型。PBMC诱导培养4d后,CIK开始增殖,第7～9天细胞数量明显增多。K562/ADR来源的DC和CIK共培养后,细胞增殖速度明显加快,9d以后与CIK相比2组细胞的增殖速度呈现明显差异。CIK+K562/ADR-DC细胞对K562/ADR的细胞毒作用在效靶比20:1范围内明显高于CIK细胞(P<0.05)。CIK组和CIK+K562/ADR-DC组细胞的P-gp和Mdr-1与对照组相比显著降低(P<0.05),ADR表达明显升高。结论:CIK+K562/ADR-DC对高表达P-gp的K562/ADR耐药细胞株表现出了特异性细胞毒作用,有效提高了细胞内ADR的含量,下调了P-gp蛋白和mdr-1基因的表达,从而使免疫效应细胞体外逆转MDR的效果得以显现。Aim ： To explore the feasibility of CIK cells reverse multi-drug resistance in vitro. Methods： Resistant K562/ADR cell and sensitive K562 cell were induced from differentiating to DC in complete RPMI 1640 culture medium supplemented with GM-CSF, IL-4 and TNF-α. PBMC were induced into CIK, and then made the application of the MDR reverse experiment with mature DC,co-cultured with mature DC,in vitro. The expression of P-gp and intracellular concentration of ADR in target cells were detected by flow cytometry, while the cytotoxicity was measured by MTT, and the gene expressions of mdr-1 were detected by RT-PCR. Results. When tumor antigen of K562/ADR cell line loaded on DC cocultured with CIK, the cell proliferation was obviously accelerated and the multiplications show obvious differences after 9 days. The growth inhibition ratio of effector cells on target cells showed higher and more specific growth, while the cytotoxicity of the CIK cell to target cell was on upward. P-gp of target cell was declined in the comparison with the control group, in the E/T scope of 20 ： 1. The effect of cytotoxicity of CIK ＋ K562/ADR-DC was much higher than that of the CIK cell （P 〈 0.05）. The expression of mdr-1 gene also showed a downward trend. Conclusion： CIK ＋ K562/ADR-DC has a specificity cytotoxicity on K562/ADR drug-resistant cell lines with high P-gp expression,notably increased the content of ADR while the expression of P-gp and mdr-1 were declined. The vitro MDR reversal of immune cells was shown significantly.

关 键 词：细胞因子 杀伤细胞 DC细胞 K562/ADR细胞株 多药耐药 

分 类 号：R733.7[医药卫生—肿瘤]