【摘要】 目的探讨恶性肿瘤对于细胞因子诱导的杀伤(cytokine induced killer,CIK)细胞的增殖抑制以及诱导凋亡等方面的作用,并探讨肿瘤细胞影响CIK的作用机制。方法在CIK细胞培养体系中分别加入胃癌细胞系BGC-823、结肠癌细胞系HCT-8细胞的培养上清,观察对于CIK细胞的增殖倍数、主要效应细胞(CD3~+ CD56~+细胞)的含量以及杀伤活性的影响。为探讨肿瘤细胞来源的胞外体在影响CIK细胞活性中的作用,采用超速离心技术分离肿瘤细胞培养上清中的胞外体结构,观察其对于CIK细胞的抑制作用。结果培养体系中肿瘤培养上清的加入降低了CIK细胞的增殖倍数和CD3~+ CD56~+细胞的比例,并且肿瘤培养上清的加入抑制了CIK细胞对于同类肿瘤细胞的细胞毒活性。肿瘤细胞和CIK细胞的共同培养增加了CIK细胞中凋亡细胞的比例,同时降低了增殖细胞比例。通过超速离心方法从BGC-823胃癌细胞培养上清液中分离出胞外体结构,电镜显示为直径介于40～100 nm之间的膜性微小囊泡,并且BGC-823细胞培养上清中胞外体结构的去除可以减轻其对于CIK细胞的增殖抑制。结论肿瘤细胞对于CIK细胞的增殖和生物学活性具有抑制作用,免疫抑制性细胞因子和胞外体都是可能的作用机制。更多还原

【Abstract】 Objective To analyze the influence of malignant cancer cells on the proliferation and cyto- toxicity of cytokine induced killer (CIK) cells. Methods Supernatant from gastric cancer cell line BGC-823 and colon cancer cell line HCT-8 was added into the culture system of CIK cells. The proliferation of CIK was analyzed by BrdU and MTT method. The percentage of the major effect cells (CD3~+ CD56~+ cells) were enumerated by flow cytometry. The cytotoxicity was evaluated using LDH method. To analyze the mechanisms of the tumor cells affect- ing CIK, the exosome of BGC-823 cells was isolated and tested by electron microscopy. Results The addition of the supernatant from cancer cell culture system decreases the proliferating rate of CIK cells and the prevalence of CD3~+ CD56~+ cells. The cytotoxicity of CIK cells against other similar cancer cell lines was also decreased. Exo- somes, small (40-90 nm in diameter) single-membrane bound vesicles were extracted by ultra-speed centrifugation from the supernatant of BGC-823 cells, and were identified by electron microscopy. Depletion of the exosomes from the supematants of BGC-823 cells decreases their suppressive capability to the proliferation rate of CIK cells. Con- clusion Cancer cells can inhibit the biological activities of CIK cells. The exosomes secreted from cancer cells may act as an important factor in this course.更多还原