摘要
目的探讨通过构建表达人白细胞抗原(HLA)HLA-A2的肺癌细胞系以诱生肿瘤特异性细胞毒性T淋巴细胞(CTL)。方法将编码HLA-A2基因的质粒pcDNA3.1D/V5转染人肺腺癌A549细胞,得到能够稳定表达HLA-A2的肺腺癌细胞系。将其用丝裂霉素(MMC)处理后,行混合淋巴细胞肿瘤细胞培养(mixed lymphocyte tumor cell culture,MLTC)以诱导HLA-A2+的健康人外周血淋巴细胞(peripheral blood lymphocyte,PBL)产生肿瘤特异性CTL,并通过特异性杀伤实验、单克隆抗体阻断实验、淋巴细胞增殖实验进行检验。结果HLA-A0201基因在A549细胞中的瞬时与稳定表达率分别为0.32、0.91。成功获得稳定表达HLA-A2的A549细胞株。MLTC诱导最终产生4组T淋巴细胞,且符合肿瘤特异性CTL的形态学特征与大量增殖的特点,但CTL杀伤实验和单克隆抗体阻断实验结果示肿瘤细胞未被有效杀伤,可能与肿瘤特异性CTL的数量较少及活性较低有关。结论本研究通过构建表达HLA-A2的肺癌细胞系,对建立肿瘤特异性CTL模型作了初步探讨,为进一步完善肺癌特异性CTL细胞模型并为研究肿瘤免疫逃避、免疫耐受及反杀伤淋巴细胞等生物学行为打下基础。
Objective To induce tumor specific cytotoxic T lymphocytes （CTLs） with construction of human lung adenocarcinoma cell line expressing HIA-A2 gene. methods Plasmid pcDNA3.1D/V5 encoding HIA-A2 gene was transfected into human lung adenocarcinoma A549 cells to obtain cell line expressing HIA-A2 gene. Then the cell line was treated with mitomycin （MMC）. Tumor specific CTLs were induced with mixed lymphocyte tumor cell culture （MLTC） in HIA-A2^＋ peripheral blood lymphocyte （PBL） from healthy donators and were verified by specific injury test, antibody blocking test, and lymphocyte proliferation test. Results Transient expression efficiency and stable expression efficiency of HIA-A2 gene in A549 cells were 0.32 and 0.91, respectively. A549 cell line exressing HIA-A2 gene had been constructed successfully. After MLTC, 4 groups of CI＇Ls were obtained, whose morphology and proliferation features were consistent with that of tumor-specific CI＇Ls. The results of CTL killing test and the mono antibody blocking test showed that the tumor cells had not be killed, which may due to the number and the activity of CTLs was very little. Conclusion Some experience and experimental data are obtained from the initial study on foundation of tumor specific CTL with HIA-A2^＋ adenocarcinoma, which is very helpful for the optimization of CTL model for studying immune evading and tolerance in tumor cells.
出处
《免疫学杂志》
CAS
CSCD
北大核心
2007年第1期40-45,共6页
Immunological Journal
基金
国家自然科学基金(30400440)
第三军医大学博士创新基金(20050030)资助
