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898~903.ATG-F培养体系制备CIK细胞的表型特点及其对K562细胞的杀伤作用[J].张宇尘,沈燕,张萍,白凤霞,娄世锋.中国肿瘤生物治疗杂志,2018,(9)
ATG-F培养体系制备CIK细胞的表型特点及其对K562细胞的杀伤作用    点此下载全文    点此浏览HTML全文
张宇尘  沈燕  张萍  白凤霞  娄世锋
重庆医科大学第二附属医院血液内科,重庆400010
基金项目:重庆市自然科学基金资助项目(No. cstc2017jcyjAX0239)
DOI:10.3872/j.issn.1007-385X.2018.09.009
摘要:
      目的:探讨抗人T细胞兔免疫球蛋白-费森尤斯(anti-human T lymphocyte rabbit immunoglobulin-Fresenius,ATG-F)和IFN-γ、IL-2 组成的培养体系诱导细胞因子诱导的杀伤(cytokine induced killer,CIK)细胞的性能并用于临床细胞治疗的可行性。方法:采集分离健康供者外周血单个核细胞(peripheral blood mononuclear cell,PBMC),依照激活抗体的不同分组分别培养CIK细胞,在第7 和14 天计数总细胞数量。流式细胞术检测ATG-F组、CD3 组和TG(抗人胸腺细胞兔免疫球蛋白,Thymoglobulin)组CIK细胞组成及细胞表面活化性、抑制性受体分子的比例,还检测第14 天时ATG-F高剂量组、CD3 组和TG组CIK细胞对K562 靶细胞的杀伤性能。结果:使用ATG-F、IFN-γ、IL-2 培养体系成功诱导出CIK细胞。第14 天时高剂量ATG-F(F-H)组增殖倍数明显高于TG组(27.25±1.25 vs 16.60±1.72,P<0.01);其CD3+CD56+细胞比例与CD3 组无统计学差异(P<0.05),但CD3-CD56+的NK细胞显著高于TG 组及CD3 组[ (11.19±2.60)% vs(5.66±1.00)%、(1.42±0.51),P<0.01]、CD4+T 细胞比例显著低于CD3、TG 组[(4.35±1.47)% vs(26.88±5.01)%、(14.52±6.22)%,P<0.01]、CD56+CD94+ 、CD56+CD158a+ 、CD56+CD158b 细胞比例均明显高于CD3 组( 均P<0.01);F-H 组在靶效比为1∶10 时对K562 的杀伤效力显著高于CD3 组[ (60.52±2.05)% vs(30.02±6.67)%,P<0.01]。结论:ATG-F培养体系制备的CIK 细胞比常规培养体系所得CIK 细胞具有更高的NK细胞比例,其活化受体对K562细胞更强的细胞毒性。
关键词:细胞因子诱导的杀伤细胞  抗人T细胞兔免疫球蛋白-费森尤斯  生物治疗  CD3
Phenotypes of CIK cells prepared by ATG-F culture system and its killing effect against K562 cells    Download Fulltext
ZHANG Yuchen  SHEN Yan  ZHANG Ping  BAI Fengxia  LOU Shifeng
Department of Hematology, the Second Affiliated Hospital of Medical University of Chongqing, Chongqing 400010,China
Fund Project:Project supported by the Natural Science Foundation of Chongqing City(No.cstc2017jcyjAX0239)
Abstract:
      Objective: To investigate the function of CIK (cytokine induced killer) cells cultured using ATG-F (anti-human T lymphocyte rabbit immunoglobulin-Fresenius) and IFN - γ , IL-2 system and its feasibility in clinical practice. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from healthy donors and were used to culture CIK cells by different activating antibodies;the total cell count was calculated on Day 7 and 14. The CIK cell composition, cell surface activation and proportion of inhibitory receptor molecular in ATG-F group, CD3 group and TG (Thymoglobulin) group were analyzed by Flow cytometry, and the cytotoxicity of CIK cells against K562 cells were also determined by flow cytometry at day 14 in ATG-F high-dose group, CD3 group and TG group. Results: CIK cells were successfully cultured by ATG-F, IFN- γ , IL-2 system. The proliferation rate of ATGF high-dose group was significantly higher than that in TG group (27.25±1.25 vs 16.60±1.72, P<0.01), but the proportion of CD3+CD56+ cells showed no statistical difference compare with the CD3 group (P>0.05). The percentage of CD3-CD56+ NK cells in ATG-F high-dose group was significantly higher than that in TG group and CD3 group [(11.19±2.60)% vs(5.66±1.00)%,(1.42±0.51)% ,P<0.01], while the proportion of CD4+T cells was significantly lower than that in CD3, TG group [(4.35±1.47)% vs (26.88±5.01)%,(14.52±6.22)%, P<0.01]; the proportion of CD56+CD94+, CD56+CD158a+, CD56+CD158b cells was significantly higher than those in CD3 group (all P<0.01). The ATG-F high does group showed significantly higher cytotoxicity against K562 cells than that of CD3 group at the target/effect ratio of 1∶10. Conclusion: CIK cells cultured by ATG-F culture system has higher NK cell proportion than other ordinary culture system, and its activated receptor has more stronger cytotoxicity against K562 cells.
Keywords:cytokine induced killer(CIK)cell  anti-human T lymphocyte rabbit immunoglobulin-Fresenius(ATG-F)  Biological treatment  CD3
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