白细胞介素-27促进CIK细胞的增殖及其对淋巴瘤细胞的杀伤能力

Effect of interleukin-27 on the proliferation and cytotoxicity of lymphadenoma cells of cytokine induced killer cells

目的:初步探讨重组人白介素-27(interleukin-27,IL-27)体外对人外周血来源的细胞因子诱导的杀伤(cytokine induced killer,CIK)细胞增殖及其对肿瘤细胞的杀伤能力的影响。方法:无菌条件下分离健康志愿者外周血单个核细胞(peripheral blood mononuclear cell,PBMC),第0天加入IFN-γ、CD3单抗,之后根据加入不同细胞因子剂量随机分为六组进行CIK细胞培养:A组(IL-2:1 000 U/ml,正常对照组),B组(IL-2:1 000 U/ml,IL-27:20 ng/ml),C组(IL-2:1 000 U/ml,IL-27:10 ng/ml),D组(IL-2:500 U/ml,IL-27:10 ng/ml),E组(IL-2:1 000 U/ml,IL-27:5 ng/ml),F组(IL-2:1 000 U/ml,IL-27:40 ng/ml)。倒置相差显微镜下观察各组CIK细胞生长情况,流式细胞术检测各组CIK细胞CD3+CD56+T、CD8+T细胞的表达,自动细胞计数仪计数各组CIK细胞增殖情况,MTS方法测定各组CIK细胞对淋巴瘤K562细胞的杀伤活性。结果:培养第11天,D组与A、B、C组比较,CIK细胞中CD3+CD56+T细胞的表达[(66.57±2.44)%vs(60.03±1.75)%,(55.51±0.03)%,(56.07±0.83)%;均P<0.05]、CD8+T细胞的表达[(81.67±1.97)%vs(70.30±2.67)%,(74.92±2.47)%,(74.43±1.90)%;均P<0.05]都明显增强;D组CIK细胞培养的扩增倍数明显高于A、B、C组[(4 811.87±23.07)vs(3 257.73±91.97),(3790.92±64.49),(4 009.85±43.08)倍;均P<0.05];效靶比40∶1时;D组CIK细胞培养第11天时杀伤力为(76.71±2.21)%,显著高于其他各组(均P<0.05)。结论:细胞因子IL-27体外可显著提高CIK细胞的增殖能力和杀伤能力,最佳培养周期为11 d。

Objective：To investigate the effect of IL-27 on the proliferation and cytotoxicity of cytokine induced killer （CIK） cells derived from human peripheral blood mononuclear cells （PBMC）. Methods： Monocytes were purified from PBMCs obtained from healthy adults. After being challenged with interferon-γ （IFN-γ） and anti-human CD3, the cells were radomly divided into six groups by different stimulating factors： A group （IL-2：1 000 U/ml）, B group （IL-2： 1 000 U/ml, IL-27：20 ng/ml）, C group （IL-2：1 000 U/ml, IL-27：10 ng/ml）, D group （IL-2：500 U/ml, IL-27： 10 ng/ml）, E group （IL-2：1 000 U/ml, IL-27：5 ng/ml）, and F group （IL-2：1000 U/ml, IL-27：40 ng/ml）. The mor- phology of the CIK cells in different groups were evaluated by inverted microscopy. The proportion of CIK cells respective- ly expressing CD3 ＋ CD56 ＋ and CD8 ＋ were analyzed by fluorescence activating cell sorter （FACS） on days 7 , 9, 11 and 13 after treatments. The number of attached cells was counted by a computer-based cell counter. The cytotoxicity of CIKceils was determined by MTS test. Results： Compared with A, B and C groups, D group had significantly higher propor- tions of CD3 ＋CD56＋ CIK cells （[66.57 ±2.441% vs [60.03 ±1.751%, [55.51 ±0.031%, and [56.07 ± 0.831%; P〈0.05） and CD8 ＋ CIK cells （[81.67 ±1.971% vs [70.30±2.671%, [74.92 ±2.471%, and [ 74.43 ± 1.90 ] % ; P 〈 0.05 ）, and significantly higher index of cell expansion to culture median （4 811.87 ± 23.07 vs 3 257.73 ± 91.97, 3 790.92 ± 64.49, 4 009.85 ± 43.08 ; P 〈 0.05） on day 11 after treatments. The highest effector to target cell ratio on day 11 in D group was 40：1 with a cytotoxicity rate of （76.71 ± 2. 21 ）% which was significantly higher than that in groups A, B, and C. Conclusion： In vitro, IL-27 is capable of significantly enhancing the proliferation and cytotoxicity of CIK cells in a time-dependent manner and the optimal time of incubation seems to be 11 days.