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目的:研究负载膀胱癌酸洗抗原肽对树突状细胞(DCs)分泌IL-12的影响和意义。方法:用弱酸洗脱法获得膀胱癌细胞株BIU-87肿瘤抗原肽;联合应用重组人GM-CSF、IL-4和TNF-α体外诱导健康志愿者DCs并负载膀胱癌肿瘤抗原肽;DC诱导膀胱癌抗原特异性CTLs;用MTT法检测其对BIU-87体外杀伤效应;用ELISA法检测第3、6、9天培养细胞的上清液,评价DCs分泌IL-12 p70的能力。结果:联合应用重组人GMCSF、IL-4和TNF-α可在体外诱导出DCs;负载膀胱癌抗原肽的DC诱导的特异性CTLs可杀伤高达(78.5±7.0)%的BIU-87细胞,显著高于各对照组(P<0.01);经ELISA方法检测不同时期的DCs分泌IL-12的量不同,而且负载抗原肽DCs较未负载抗原DCs有更强的分泌能力(P<0.05)。结论:IL-12 p70在DCs刺激特异性CTLs的过程中发挥重要作用,其分泌量受DCs的成熟状态及某些刺激信号的影响,膀胱癌酸洗抗原肽是其刺激信号之一。
Objective: To investigate the effect and significance of bladder cancer acid-washed antigen peptide on the secretion of IL-12 by dendritic cells (DCs). Methods: Human bladder cancer cell line BIU-87 tumor antigen peptide was obtained by weak acid-eluting method. DCs were induced in vitro in combination with recombinant human GM-CSF, IL-4 and TNF-α and loaded with tumor antigen peptide of bladder cancer. Bladder cancer antigen-specific CTLs were detected by MTT assay. The cytotoxicity of BIU-87 in vitro was detected by MTT assay. The supernatant of cultured cells on day 3, day 6, and day 9 was assayed by ELISA to evaluate the ability of DCs to secrete IL-12 p70. Results: DCs were induced by recombinant human GMCSF, IL-4 and TNF-α in combination with DCs. DC-specific CTLs loaded with bladder cancer antigen peptides could kill up to (78.5 ± 7.0)% of BIU-87 cells, (P <0.01). The amount of IL-12 secreted by DCs in different periods detected by ELISA was different, and the DCs loaded with antigenic peptides had stronger secretion ability than DCs without load (P <0.05). CONCLUSION: IL-12 p70 plays an important role in the stimulation of specific CTLs by DCs. The secretion of IL-12 is influenced by the maturation status of DCs and some stimulation signals. Pickling antigen peptide of bladder cancer is one of the stimulatory signals.