抗原冲击树突状细胞介导细胞毒性T淋巴细胞特异性抗白血病作用的体外研究

Specific anti-leukemic cell effect mediated by dendritic cells pulsed with acute myelogenous leukemia antigen in vitro

目的研究白血病来源树突状细胞体外诱导的有效方法；观察不同抗原诱导的特异性细胞毒性T淋巴细胞（CTL）的抗白血病效应。方法分离白血病患者骨髓单个核细胞，经钙离子载体A23187诱导分化，将弱酸洗脱抗原、低渗抗原分别冲击树突状细胞，96h后树突状细胞与T细胞共同培养，采用MTT法比较CTL细胞对白血病细胞的杀伤活性。结果骨髓来源的单个核细胞经过100ng／ml的GM—CSF、500ng／ml钙离子载体A23187成功诱导为树突状细胞，倒置显微镜下具有树突状细胞的典型形态，流式细胞术测定其CD1a、CD83表达较诱导前明显升高，差异有统计学意义（P〈0．01）。弱酸洗脱法获得抗原冲击树突状细胞致敏T细胞后杀伤白血病的能力最高，未负载抗原的树突状细胞致敏T细胞杀伤白血病细胞的能力最低，两者差异具有统计学意义（P〈0．01）。结论白血病来源的骨髓单个核细胞经GM—CSF、钙离子载体A23187能够成功诱导为树突状细胞；弱酸洗脱后获得的抗原冲击树突状细胞致敏T细胞能够获得更强的杀伤白血病细胞的能力。

Objective To explore the effective method for in vitro culture of the dendritic cells（DCs） and the specific anti-leukemic cell effect mediated by dendritic cells pulsed with acute myelogenous leukemia antigen. Methods Bone marrow mononucleas cells （BMMNCs） isolated from AML patients were induced to undergo differentiation with 500 ng/ml A23187 and pulsed with AML antigen. After 96 h, DCs and T cells were co-cultured for 5 to 7 days. The cytotoxic activity of cytotoxic T lymphocyte（CTL） to AML were detected with MTY colorimetry. Results BMMNCs isolated from AML patients treated with 100 ng/ml rhGM-CSF in combination with 500 ng/ml A23187 for 96 h exhibited typical morphology of DCs with rapidly increased expression of CD1a and CD83 （P 〈0.01）. Dendritic cells pulsed with acid eluted tumor antigen peptides group displayed the strongest cytotoxic activity of CTL to AML. There was a significant difference between two groups（P 〈0.01）. Conclusion BMMNCs isolated from AML patients can be successfully induced to DCs with rhGM-CSF and A23187 and dendritic cells pulsed with acid eluted tumor antigen peptides group had the strongest cytotoxic activity of CTL to AML.