Ahmed Ali Hassan Abdellah, Di Jun, Mei Wu, Zhang Yu-Cheng, Li Yi, Du Zhen-Wu, Zhang Gui-Zhen
Department of Central Research, Second Hospital, Jilin University, Changchun, China E-mail :
Asian Pac J Cancer Prev. 2014;15(2):611-6. doi: 10.7314/apjcp.2014.15.2.611.
Dendritic cell (DC)-based tumor immunotherapy needs an immunogenic tumor associated antigen (TAA) and an effective approach for its presentation to lymphocytes. In this study we explored whether transduction of DCs with lentiviruses (LVs) expressing the human interleukin-12 gene could stimulate antigen- specific cytotoxic T cells (CTLs) against human lung cancer cells in vitro.
Peripheral blood monocyte- derived DCs were transduced with a lentiviral vector encoding human IL-12 gene (LV-12). The anticipated target of the human IL-12 gene was detected by RT-PCR. The concentration of IL-12 in the culture supernatant of DCs was measured by ELISA.Transduction efficiencies and CD83 phenotypes of DCs were assessed by flow cytometry. DCs were pulsed with tumor antigen of lung cancer cells (DC+Ag) and transduced with LV-12 (DC-LV-12+Ag). Stimulation of T lymphocyte proliferation by DCs and activation of cytotoxic T-lymphocytes (CTL) stimulated by LV-12 transduced DCs pulsed with tumor antigen against A549 lung cancer cells were assessed with methyl thiazolyltetrazolium (MTT).
A recombinant lentivirus expressing the IL-12 gene was successfully constructed. DC transduced with LV-12 produced higher levels of IL-12 and expressed higher levels of CD83 than non-transduced. The DC modified by interleukin -12 gene and pulsed with tumor antigen demonstrated good stimulation of lymphocyte proliferation, induction of antigen-specific cytotoxic T lymphocytes and anti- tumor effects.
Dendritic cells transduced with a lentivirus-mediated interleukin-12 gene have an enhanced ability to kill lung cancer cells through promoting T lymphocyte proliferation and cytotoxicity.
基于树突状细胞（DC）的肿瘤免疫疗法需要一种具有免疫原性的肿瘤相关抗原（TAA）以及将其呈递给淋巴细胞的有效方法。在本研究中，我们探讨了用表达人白细胞介素-12基因的慢病毒（LV）转导DC是否能在体外刺激针对人肺癌细胞的抗原特异性细胞毒性T细胞（CTL）。
用编码人IL-12基因的慢病毒载体（LV-12）转导外周血单核细胞来源的DC。通过RT-PCR检测人IL-12基因的预期靶点。用ELISA法检测DC培养上清中IL-12的浓度。通过流式细胞术评估DC的转导效率和CD83表型。用肺癌细胞的肿瘤抗原对DC进行脉冲处理（DC+Ag），并用LV-12进行转导（DC-LV-12+Ag）。用甲基噻唑基四氮唑（MTT）评估DC对T淋巴细胞增殖的刺激作用以及用LV-12转导并用肿瘤抗原脉冲处理的DC对A549肺癌细胞刺激的细胞毒性T淋巴细胞（CTL）的激活作用。
成功构建了表达IL-12基因的重组慢病毒。与未转导的DC相比，用LV-12转导的DC产生更高水平的IL-12，并表达更高水平的CD83。经白细胞介素-12基因修饰并用肿瘤抗原脉冲处理的DC表现出对淋巴细胞增殖的良好刺激作用、诱导抗原特异性细胞毒性T淋巴细胞以及抗肿瘤作用。
用慢病毒介导的白细胞介素-12基因转导的树突状细胞通过促进T淋巴细胞增殖和细胞毒性而具有增强的杀伤肺癌细胞的能力。
