脐血及外周血NKT细胞体外扩增及其表型比较.docVIP

脐血及外周血NKT细胞体外扩增及其表型比较 　 作者：刘嬿，范华骅，阮鸣，高跞，聂晓绚，杨懿铭，陆华中，高峰 【摘要】 为了建立体外有效扩增脐血与外周血天然杀伤T细胞（NKT细胞）的方法并研究其表型的差异，分别采用单加IL-2及同时加IL-2和α-半乳糖神经酰胺（α-Galcer）的方法，从人脐血单个核细胞（UCB-MNC）和人外周血单个核细胞（PB-MNC）中扩增NKT细胞；用流式细胞检测技术测定NKT细胞（TCR Vα24+ TCR Vβ11+）的比例及其它表型特征。结果显示： UCB-MNC在第7天时，其TCR Vαβ+ NKT细胞的扩增量占淋巴细胞的（24.48±419）%，是原来的（8.74±4.37）×102倍 （ Plt;0.01），而且大多不表达NK1.1（CD161）；TCR Vβ11+较TCR Vα24+高表达；第14天时，PB-MNC中的TCR Vαβ+NKT细胞扩增量占淋巴细胞的（11.1±3.61）%，是原来的（372±201）×102倍（Plt;0.01），且NK1.1高表达，TCR Vβ11+与 TCR Vα24+表达率基本一致。结论： α-Galcer对NKT细胞有特异的扩增功能，且脐血NKT的扩增效率较外周血高。以表型划分，脐血中的NKT细胞多为NK1.1-，而外周血多为NK1.1+，是一种新的NKT细胞亚型。 【关键词】 脐血 　　Proliferation of Natural Killer T Cells in Umbilical Cord Blood and Periperal Blood and Their Different Phenotypes 　　　Abstract Purpose of this study was to establish an effective method in vitro to proliferate natural killer T (NKT) cells from umbilical cord blood (UCB) and periperal blood (PB)，and to study their different phenotype. Mononuclear cells (MNC) from UCB and PB were cultured in the presence of IL-2 (100 U/ml)，with or without alpha-Galcer. TCR Vα24 Vβ11 double positive natural killer T-cells (NKT cells) and their other phenotypes were determined by flow cytometry. The results showed that after expansion for 7 days，TCRVαβ+ NKT cells from UCB-MNCs increased by （8.74±4.37）×102 times as much，but most of them did not express NK1.1 and its TCR Vβ11+ was higher than TCR Vα24+. After expansion for 14 dyas，TCR Vαβ+ NKT cells from PB-MNCs increased by （3.72±2.01）×102 times，the expression of NK1.1 was high and its TCR Vβ11+ was almost equal to TCR Vα24+. It is concluded that human TCR Vα24 Vβ11 double positive NKT cells can expand by addition of α-Galcer. The proliferation efficiency in UCB-MNCs is greater than that in PB-MNCs. Most of the UCB-NKT is NK1.1-，while the PB-NKT is NK1.1+，a new subset of NKT cells. 　　Key words umbilical cord blood；periperal blood；natural killer T cell；α- Galcer；phenotype 新近研究发现，构成机体免疫系统的淋巴细胞中，除T、B、NK细胞外，还存在一种新的细胞系——NKT细胞。这种细胞有严格的CD1d限制性，且具有大量释放IL-4和I