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目的探索具有肿瘤杀伤活性(CKA)粒细胞的筛选方法,并对其体外抑瘤效应进行初步研究。方法 (1)收集健康志愿者外周抗凝全血21份,分离粒细胞,与人肺癌A549细胞共培养,通过检测肿瘤细胞增殖活性筛选出CKA粒细胞。(2)将筛选得到的CKA粒细胞作用于不同种类的肿瘤细胞,采用CCK-8法检测肿瘤细胞活性,双染法检测肿瘤细胞凋亡发生情况。(3)分别采用直接接触、Transwell共培养、条件培养液等共培养细胞方式,观察不同处理方式对CKA粒细胞抑制肿瘤细胞活性的影响。结果 (1)有6株粒细胞克隆对A549细胞活性具有明显的抑制作用,将其中作用最显著的克隆3标记为CKA粒细胞。(2)CKA粒细胞不仅对人肺腺癌细胞A549、人肝癌细胞HepG2和人宫颈癌细胞HeLa生长活性有明显抑制作用,且能促进这3种细胞的凋亡。(3)3种处理方式的抑制效率比较:直接接触>Transwell>条件培养。结论 CKA粒细胞对某些类型的肿瘤细胞活性具有明显的抑制作用;CKA粒细胞对肿瘤的杀伤作用可能与其与肿瘤的接触方式有关。
Objective To explore a screening method for neutrophils with tumor-killing activity (CKA) and to study its anti-tumor effect in vitro. Methods (1) 21 healthy volunteers were collected peripheral blood anticoagulant whole blood, isolated granulocytes, co-cultured with human lung cancer A549 cells, CKA granulocytes were screened by detecting tumor cell proliferation activity. (2) The CKA granulocytes were screened for different types of tumor cells, the activity of tumor cells was detected by CCK-8 assay, and the apoptosis of tumor cells was detected by double staining. (3) Using direct contact, Transwell co-culture, conditioned medium and other co-cultured cells, to observe the different treatment of CKA granulocytes inhibit the activity of tumor cells. Results (1) There were 6 strains of granulocyte clones on A549 cells significantly inhibited the activity, the most significant role in which clone 3 labeled as CKA granulocytes. (2) CKA granulocytes not only significantly inhibited the growth activity of human lung adenocarcinoma A549, HepG2 and HeLa, but also promoted the apoptosis of these three kinds of cells. (3) Comparison of inhibition efficiency of three treatments: direct contact> Transwell> conditioned culture. Conclusions CKA granulocytes have obvious inhibitory effect on the activity of certain types of tumor cells. The killing effect of CKA granulocytes on tumor may be related to their contact with tumor.