IL-2和IL-15刺激脐血NK细胞对K562/Jurkat细胞的杀伤活性研究

Cytotoxic Effect of IL-2/IL-15 Stimulated Cord Blood Derived NK Cells on K562/Jurkat Cell Lines

本研究旨在探讨脐血(cord blood,CB)NK细胞的杀伤功能及细胞因子IL-2和IL-15对NK细胞杀伤K562/Jurkat细胞活性的影响。采用autoMACS及CD3阴选后CD56阳选两步免疫磁珠分选法从脐血单个核细胞纯化NK细胞,分选后NK细胞用流式细胞仪测定纯度,通过IL-2及IL-2和IL-15两种细胞因子组合培养3天,用LDH释放法检测培养前后脐血NK细胞对白血病细胞株K562和Jurkat的细胞毒活性(效靶比10∶1),并记录不同细胞因子培养体系条件下NK细胞的生长状况。结果表明,CB-MNC中CD3-CD56+细胞含量为(14.88±9.2)%,分选后CD3-CD56+细胞含量为(92.39±0.8)%;培养3天后,IL-2组NK细胞形成集落数为148.60±13.0,IL-2复合IL-15组NK细胞形成集落数为831.80±23.0,两者比较,有显著性差异;以K562为靶细胞时,新鲜纯化CB-NK的杀伤活性为(27.76±8.8)%,IL-2组杀伤活性为(61.9±9.1)%,IL-2复合IL-15组杀伤活性为(87.62±3.7)%,差别有统计学意义;以Jurkat为靶细胞时,新鲜纯化CB-NK细胞的杀伤活性为(29.32±2.5)%,IL-2组杀伤活性为(69.43±4.4)%,IL-2复合IL-15组杀伤活性为(92.95±3.2)%,差别有统计学意义。结论:脐血新鲜纯化NK细胞对K562、Jurkat细胞有一定的杀伤作用,但自发细胞毒活性较低,IL-2/IL-15刺激后杀伤活性显著提高,IL-15复合IL-2对促进脐血NK细胞的生长和提高其细胞毒活性明显优于单用IL-2。

The aim of this study was to explore the cytotoxicity of fresh cord blood（CB） NK cells and the influence of IL-12 and IL-15 on activity of the NK cells killing K562 and Jurkat cells lines.The NK cells were isolated from cord blood by depleting CD3＋ cells and then enriching CD56＋ cells using sorting with immunomagnetic beads.The experiment was diveded into 3 groups： group A（fresh CB-NK cells without cytokines）,group B（CB-NK cells cultured by IL-2） and group C（CB-NK cells cultured by IL-2 and IL-15）.The purity of NK cells was determined by flow cytometry;the cytotoxity of fresh and different cytokine-treated CB-NK cells on K562 and Jurkat cell lines was detected by LDH release test.The results showed that the purity of NK cells before and after sorting was 14.88±9.2% and 92.39±0.8% respectively.After culture for 3 days,NK-forming colony amounts in group B and group C were 148.60±13.0 and 831.80±23.0 respectively,the comparison between group B and group C showed the significant difference（p〈0.05）.The cytotoxicities of NK cells in group A,B and C on K562 and Jurkat cell lines were 27.76±8.8%,61.90±9.1% and 87.62±3.7%;29.32±2.5%,69.43±4.4% and 92.95±3.2% respectively,the difference was significant（p〈0.05）.It is comcluded that the fresh isolated CB-NK cells show low cytotoxic activity.After stimulated with IL-2 or IL-2 plus IL15,cytotoxicity of CB-NK cells increases obviously,the effect of IL-2 plus IL-15 is much better than IL-2 alone for promoting the growth and enhancing the cytotoxicity of CB-NK cells.