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目的:探究IL-15、4-1BBL基因修饰的人白血病K562细胞(modi-K562细胞)联合IL-2体外高效扩增肾细胞癌患者自体自然杀伤(natural killer,NK)细胞的方法,研究扩增前后NK细胞对肾癌细胞株786-O的杀伤作用。方法:10例肾癌患者的外周血单个核细胞(peripheral blood mononuclear cell,PBMC)与modi-K562细胞在含不同浓度IL-2培养液中共育14 d,采用流式细胞术、Calcein-AM释放实验检测NK细胞的扩增情况、免疫表型及对肾癌786-O细胞的杀伤作用。结果:modi-K562细胞联合IL-2可有效扩增NK细胞,300 U/ml IL-2培养14 d时,NK细胞扩增(202.4±12.8)倍。在效靶比为20∶1时,扩增后NK细胞对786-O细胞的杀伤率为(72.0±4.3)%,显著高于扩增前NK细胞的杀伤率(34.2±3.6)%(P<0.01)。结论:IL-15、4-1BBL基因修饰的K562细胞联合IL-2在体外能有效扩增肾细胞癌患者NK细胞,扩增后NK细胞对肾癌786-O细胞的杀伤作用显著增强。
OBJECTIVE: To explore a method to efficiently expand natural killer (NK) cells in patients with renal cell carcinoma by IL-15,4-1BBL gene-modified human leukemia K562 cells (modi-K562 cells) in combination with IL-2 NK cell proliferation before and after the killing of renal cell carcinoma cell line 786-O. Methods: Peripheral blood mononuclear cells (PBMCs) and modi-K562 cells from 10 patients with renal cell carcinoma were co-cultured for 14 days in different concentration of IL-2 medium. Flow cytometry and Calcein-AM release The detection of NK cell proliferation, immunophenotype and cytotoxicity of 786-O cells in renal cell carcinoma. Results: NK cells were effectively expanded by modi-K562 cells combined with IL-2, and NK cells were expanded (202.4 ± 12.8) times after cultured with 300 U / ml IL-2 for 14 days. When the target ratio was 20:1, the killing rate of NK cells to 786-O cells after expansion was (72.0 ± 4.3)%, which was significantly higher than that of NK cells (34.2 ± 3.6)% (P <0.01). Conclusion: IL-15 and 4-1BBL gene-modified K562 cells combined with IL-2 can effectively expand NK cells in renal cell carcinoma patients in vitro, and the cytotoxicity of NK cells on renal carcinoma 786-O cells after the expansion is significantly enhanced.