树突状细胞的体外培养及其诱导的抗肺癌免疫作用

In Vitro Culture of Dendritic Cell and Its Induced Anti-tumor Immonotherapy

目的建立体外培养扩增树突状细胞(DC)的方法,并观察其诱导的抗肿瘤免疫作用.方法分离外周血单个核细胞,应用GM-CSF及IL-4诱导DC产生,用TNF-A和PGE2促进其成熟,并用肺癌细胞系GLC-82可溶性抗原致敏.将成熟DC与自体LAK细胞混合培养(DC-LAK),用MTT法检测DC-LAK细胞及LAK细胞对多种肿瘤细胞系的杀伤作用.结果经GM-CSF、IL-4、TNF-A、PGE2作用后,细胞表达CD1a阳性率为71.0%,CD83 50.0%,CD14阳性率低于10.0%,高表达HLA-Ⅰ、HLA-Ⅱ和CD45,为典型的DC表型特征.DC-LAK和LAK细胞对4种肿瘤细胞(GLC-82,A549,moser和MCF-7)的杀伤率,前者为84.7%、66.9%、49.3%和56.0%,后者为43.5%、31.3%,45.0%和32.4%.结论成功的诱导出具有典型形态特征及增强LAK细胞杀伤活性的DC.

Objective To establish a method to culture dendritic cells （DC） in vitro and to observe the anti -tumor immunological effect induced by DC. Methods Dendritic cells were derived from PBMC stimulated with GM -CSF and IL-4 and DC were pulsed with tumor soluble antigen abstracted from lung cancer cell line GLC - 82. TNF - A and PGE2 were used to enhance, antigen presenting ability of DC. Phenotype of DC were analyzed through FACS. Mature DC were co - cultured with LAK cells and the mixed cells were named DC - LAK. Cytotoxicity of DC - LAK and LAK was measured by MTT assay. Results The cultured DC expressed HLA - Ⅰ, HLA -Ⅱ and CD45 and positive rates of CDla, CD83 and CD14 were 71.0%, 50. 0% and less than 10. 0% respectively. Kill rates of DC - LAK and LAK to the four cell lines （ GLC - 82, A549, moser and MCF -7） were 84. 7%, 66. 9%, 49.3%, 56. 0% and 43.5%, 31.3%, 45.0%, 32. 4% respectively. Conclusion DC that has typical phenotype and could be enhance cytotoxicity of LAK could be induced from PBMC successfully.