摘要
目的:研究HSP70-TKD诱导的自然杀伤细胞(NK)体外对细胞表面HSP70表达不同的胰腺癌细胞和结肠癌细胞的迁移作用差异。方法:用干细胞培养基富集人外周血单个核细胞(Peripheral blood mononuclear cell,PBMC)至第10天加入2μg/ml TKD,4 d后用流式细胞仪检测NK细胞表面杀伤性受体CD94/NKG2C的表达情况;应用流式分选将胰腺癌细胞系Colo357和结肠癌细胞系CW2分为HSP70高表达细胞亚系Colo+、CW2+和HSP70低表达细胞亚系Colo-、CW2-;Transwell小室实验检测HSP70-TKD诱导的NK细胞对这4种细胞的迁移能力,MTT法检测HSP70-TKD诱导的NK细胞对这4种细胞的杀伤活性。结果:流式结果显示NK细胞在干细胞培养基中培养14天可大量增殖,细胞纯度最高达(92.50±1.25)%;经HSP70-TKD诱导后,NK细胞表面杀伤性受体CD94/NKG2C的表达明显增加;经流式分选后,Colo+、Colo-细胞表面HSP70的表达分别为(78.2±2.2)%和(27.3±1.2)%,而CW2+、CW2-细胞表面HSP70的表达分别为(91.1±2.5)%和(18.2±1.0)%;HSP70-TKD诱导的NK细胞对Colo+迁移率为(68.6±2.8)%,明显高于对Colo-的迁移率(22.8±1.5)%,NK细胞对CW+迁移率为(73.5±2.7)%,明显高于对CW2-的迁移率(18.2±1.3)%;HSP70-TKD诱导的NK细胞对Colo+、Colo-的杀伤活性有显著性差异,当效靶比为20∶1时,杀伤活性分别为(61.2±3.0)%、(24.5±1.5)%,NK对CW2+、CW2-的杀伤活性有显著性差异,当效靶比为20∶1时,杀伤活性分别为(63.8±3.2)%、(22.4±1.8)%。结论:HSP70-TKD诱导的NK细胞是一种新型、高效的免疫活性细胞,在体外易向HSP70表达阳性的肿瘤细胞迁移。
Objective： To investigate the Migration ability toward human pancreatic carcinoma cell line and human colon carcinoma cell line with difference HSP70 plasma membrane expression. Methods： CD3^-CD56^＋NK cells were obtained from human peripheral blood mononuclear（ PBMC） in stem cell growth medium SCGM,2 μg / ml TKD was added to the medium on 10 th day,the activating receptor CD94 / NKG2 C expression levels on NK cells was detected with FAC after 4 days. The human pancreatic carcinoma cell line Colo357 and the human colon carcinoma cell line CW2 were separated into Colo^＋and CW2^＋with high HSP70 expression and Colo^-and CW2^-with low HSP70 expression; Migration assays of NK to the four difference cell lines were performed in a Transwell cell culture system. The cytolytic activity of TKD-activated NK cells against the four subline with HSP70 expression on their cell surface was analyzed by MTT assay. Results： Flow cytometry analysis showed that CD3^-CD56^＋NK cells could expanded after 2 weeks in SCGM medium,and the largest percentage of NK cell was（ 92. 50 ± 1. 25） %. CD94 expression levels on NK cells increased obviously after TKD inducement the cell surface HSP70 expression of Colo^＋,Colo^-were（ 78. 2 ± 2. 2） % and（ 27. 3 ± 1. 2） % separately,the cell surface HSP70 expression of CW2^＋,CW2^-were（ 91. 1 ± 2. 5） % and（ 18. 2 ± 1. 0） % separately after FACS; the Migration of NK cells toward Colo^＋was（ 68. 6 ± 2. 8） %,higher than the migration toward Colo^-with（ 22. 8 ± 1. 5） %; the Migration of NK cells toward CW2^＋was（ 73. 5 ± 2. 7） %,higher than the migration toward CW2^-with（ 18. 2 ± 1. 3） %; the cytolytic activity of NK against Colo^＋was（ 61. 2 ± 3. 0） % compared to（ 24. 5 ± 1. 5） % against Colo-when the ratio of effector cells and target cell was 20∶ 1,the cytolytic activity of NK against CW2^＋was（ 63. 8 ± 3. 2） % compared to（ 22. 4 ± 1. 8） % against CW2^-when the ratio of effector cells and target cell was 20∶ 1. Co
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2016年第8期1123-1127,共5页
Chinese Journal of Immunology
基金
福建省医学创新课题(2011-CX-17)
