抗人胸腺免疫球蛋白联合干扰素γ和白介素2诱导培养细胞因子诱导的杀伤细胞的效果*
Function and activity of cytokine induced killer cells induced by anti-human thymocyte immunoglobulin combined with interferon γ and IL-2

目的:探讨抗人胸腺免疫球蛋白(ATG)诱导培养的细胞因子诱导的杀伤细胞(CIK)的活性和功能,为CIK培养体系优化提供依据。方法:采集9例肿瘤患者外周血10 mL,分离单个核细胞,用CD3单抗或ATG(50、250、500 μg/L)联合干扰素γ、白介素2诱导培养。培养至第6～10天,采用流式细胞术检测细胞增殖状况,培养至第13天,采用流式细胞术检测CIK免疫表型(CD3、CD4、CD8、CD56)、激活性表面标志(CD28、CD27、CD69、NKG2D)、抑制性表面标志(PD1、CD152)及Granzyme-B、干扰素γ分泌量。结果:4种培养方式下,细胞增殖状况无统计学意义(P>0.05)。与CD3单抗相比,ATG培养的CIK中CD3+CD4+和CD3+CD8+细胞比例较低(P<0.001),CD3-CD56+与CD8+CD69+细胞比例较高,CD56+细胞干扰素γ和Granzyme-B分泌水平较高。结论:ATG诱导的CIK免疫活性优于用CD3单抗经典方案培养的CIK,抗肿瘤能力较强。
Aim: To investigate the activity and function of the cytokine induced killer cells(CIK)cultured by anti-human thymocyte immunoglobulin(ATG)combined with recombinant human interferon γ(IFN-γ)and recombinant human interleukin-2(IL-2).Methods: The peripheral blood(10 mL)was collected from 9 cancer patients. The peripheral blood mononuclear cells(PBMCs)were isolated by Ficoll density gradient centrifugation, and then were cultured with anti CD3 monoclonal antibody or different concentration(50,250,500 μg/L)of ATG combined with IFN-γ and IL-2. Cell proliferation was assessed using flow cytometry from D6 to D10. At the 13th day,the phenotype(CD3,CD4,CD8,CD56), the expressions of activated markers(CD28,CD27,CD69 and NKG2D)and inhibitory markers(PD1 and CD152), as well as the levels of IFN-γ and Granzyme-B secretion were analyzed by flow cytometry.Results: Compared with anti-CD3 group, the proportions of CD3+CD4+ cells and CD3+CD8+ cells were lower in ATG groups(P<0.001), while the percentages of CD3-CD56+ natural killer cells and CD8+CD69+ cells were elevated significantly in ATG groups(P<0.05). Furthermore, the levels of IFN-γ and Granzyme B secretion of CD56+ cells in all ATG groups were higher than that of anti-CD3 group(P<0.05).Conclusion: The CIK induced by ATG instead of CD3 monoclonal antibody combined with IFN-γ and IL-2 exhibit better immune-competence and enhanced cytotoxic activity