树突状细胞与细胞因子诱导的杀伤细胞共培养对白血病耐药细胞杀伤作用的实验研究

Effect of dendritic cells pulsed with tumor lysate antigens co-cultured with cytokine induced killer cells on cytotoxicity against multidrug resistant cells.

目的探讨负载P-糖蛋白(P-gp)高表达的多药耐药(MDR)白血病K562/A02细胞冻融抗原的树突状细胞(DC)与同源细胞因子诱导的杀伤细胞(CIK)共培养对MDRK562/A02杀伤作用的影响。方法提取健康人骨髓单个核细胞,常规诱导出DC及CIK,将K562/A02细胞冻融物作为抗原冲击的DC,与CIK共培养作为实验组,抗原不冲击的DC与CIK共培养作为对照组,以CIK及DC单独培养分别作为空白对照组1和空白对照组2。光镜下观察细胞形态,流式细胞术分析细胞表型,MTT法检测杀伤活性。结果实验组、对照组细胞增殖活性均大于CIK组(P<0.05)。实验组对K562/A02、K562的杀伤活性在效靶比5∶1、10∶1、20∶1时分别为(42.90±0.67)%、(49.85±0.28)%、(63.36±0.46)%和(23.56±0.43)%、(26.11±0.34)%、(34.46±0.35)%,均高于对照组及空白对照组1(P<0.05);实验组对K562/A02的杀伤活性高于K562和MCF7(P<0.05)。结论DC与CIK共培养物是一种增殖活性和细胞毒活性高于CIK的免疫活性细胞,而经冻融抗原冲击的DC与CIK共培养能明显提高对MDRK562/A02的杀伤活性。

Objective To investigate the effect of dendritic cells （DC） pulsed with tumor lysate antigens of K562/A02 co- cultured with cytokine induced killer （CIK） cells on cytotoxicity against muhidrug resistant （MDR） leukemia K562/A02 cells with high expression of p - glycoprotein （P - gp）. Methods Bone marrow mononuclear cells isolated from healthy adult donors were induced to obtain CIK cells and DC respectively and then CIK were co - cultured with DCs which were pulsed with tumor lysate. The morphologic features were observed with optical microscopy and expressions of markers typical were measured using flow cytometry. The cytotoxieity of three effect cells was measured with MTT assay respectively. Results After pulsing DCs with tumor lysate antigens co - cultured CIK or DC - CIK led to a significant increase of lymphocyte proliferation of both populations. The cytotoxicity of CIK co - culturing with DCs pulsed with tumor lysate antigens to K562/A02 and K562 cells was （42.90 ±0. 67 ） % , （49. 85 ±0. 28 ） % （63.36 ±0.46） % and （23.56 ±0.43 ） % , （26. 11 ±0. 34） % , （ 34. 46 ±0. 35 ） % respectively at an effector - to - target ratio of （5 ： 1, 10： 1, 20：1 ）. Conclusion This study indicates that DC pulsed with tumor lysate antigens co - cultured with CIK can improve lymphocyte proliferation and enhance the cytotoxic activity. Especially, it can induce a P - gp specific anti - leukemic immune.