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目的:探讨不同接种密度对C57BL/6小鼠细胞因子诱导的杀伤细胞(CIK细胞)增殖分化及杀瘤作用的影响,进一步优化CIK细胞培养方法。方法:按照1×10~6/mL(A组)、4×10~6/mL(B组)、8×10~6/mL(C组)、12×10~6/mL(D组)4种接种密度培养细胞,加入必要的细胞因子,14 d后收获细胞,通过流式细胞术、CCK-8法对细胞增殖、分化、杀瘤作用进行分析。结果:培养过程中,C组细胞形态及增殖能力优于A、B组,在14 d时收获细胞并对其进行检测时发现,C组CD3~+/NK1.1~+细胞所占比例明显高于A、B组,杀瘤活性也优于A、B组;D组细胞密度过大,在7 d细胞进入快速增殖期后出现大面积死亡。结论:适当提高C57BL/6小鼠CIK细胞的接种密度利于细胞增殖、分化及杀瘤作用的形成,选择8×10~6/mL的接种密度是较为合适的。
Objective: To investigate the effects of different inoculation density on the proliferation and differentiation of cytokine-induced killer cells (CIK cells) in C57BL / 6 mice and the cytotoxicity of CIK cells, and to further optimize the CIK cell culture method. Methods: The mice were randomly divided into four groups according to the following criteria: 1 × 10 ~ 6 / mL group A, 4 × 10 ~ 6 / mL group B, 8 × 10 ~ 6 / mL group C and 12 × 10 ~ Four kinds of inoculation density cells were cultured, the necessary cytokines were added, and the cells were harvested after 14 days. The cell proliferation, differentiation and tumoricidal activity were analyzed by flow cytometry and CCK-8. Results: The cell morphology and proliferation ability of group C were better than group A and B during the culture process. When cells were harvested at 14 days and detected, the percentage of CD3 ~ + / NK1.1 ~ + cells in group C was significantly higher Higher than group A and group B, tumor killing activity was also superior to group A and group B. In group D, the cell density was too large, and large area of death occurred after 7 days of rapid cell proliferation. Conclusions: It is more appropriate to select the proper density of 8 × 10 ~ 6 / mL when the inoculation density of CIK cells in C57BL / 6 mice is favorable to the proliferation, differentiation and tumorigenesis.