Department of Otolaryngology Head and Neck Surgery, First Affiliated Hospital of Liaoning Medical Colledge, Jinzhou, Liaoning 121001, China.
Chin Med J (Engl). 2011 May;124(9):1357-61.
Objective evaluation of the antitumor effect of interleukin-12 (IL-12) gene-transfected dendritic cell (DC) vaccine on laryngeal carcinoma requires in vivo and in vitro tests. The aim of this study was to investigate the function of IL-12 gene transfected DC at initiating specific immune response to laryngeal carcinoma in vitro.
Recombinant adenovirus with IL-12 gene was constructed. DCs were isolated from the peripheral blood of patients with laryngeal carcinoma, pulsed with tumor lysate of laryngeal carcinoma cells (DC⁺Ag), and transfected with IL-12 (DC-IL-12⁺Ag). The cells pheotypes including CD83, CD86 and HLA-DR on surface of DCs were assayed by flow cytometry (FCM). The concentration of IL-12 in culture supernatant of DCs and interferon γ (IFN-γ) in culture supernatant of T cells cocultured with DCs were quantified by ELISA. Methyl thiazolys tetrazolium (MTT) was used to evaluate proliferation of autologous T lymphocytes and activation of cytotoxic T lymphocytes (CTL) stimulated by IL-12-transfected DCs pulsed with tumor lysate against laryngeal carcinoma cells.
The recombinant adenovirus expressing IL-12 gene was constructed successfully. Gene-transfected DC plused with tumor lysate with IL-12 (DC-IL-12⁺Ag) expressed higher level of CD83, CD86 and produced higher level of IL-12 than untransfected DCs (DC⁺Ag) (CD83: (60.2 ± 1.8)% vs. (50.7 ± 1.2)%, P < 0.05; CD86: (88.9 ± 2.1)% vs. (78.2 ± 3.9)%, P < 0.05; IL-12: (262.5 ± 3.0) ng/L vs. (103.8 ± 5.1) ng/L, P < 0.05). The proliferation of autologous T lymphocytes and production of IFN-γ stimulated by DC transfected with IL-12 were more obviously than untransfected DCs. Cytotoxicity of CTL stimulated by IL-12-transfected DC pulsed with tumor lysate against laryngeal carcinoma cells were significantly stronger than stimulated by untransfected DC.
It is a promising approach for IL-12-transfected DC pulsed with tumor lysate to increase the antitumoral effect.
为了客观评价白细胞介素 12（IL-12）基因转染树突状细胞（DC）疫苗对喉癌的抗肿瘤作用，需要进行体内和体外试验。本研究旨在探讨 IL-12 基因转染的 DC 在体外启动针对喉癌细胞的特异性免疫反应的功能。
构建携带 IL-12 基因的重组腺病毒。从喉癌患者外周血中分离出 DC，用喉癌细胞肿瘤裂解物（DC⁺Ag）冲击，用 IL-12 转染（DC-IL-12⁺Ag）。流式细胞术（FCM）检测 DC 表面 CD83、CD86 和 HLA-DR 等细胞表型。酶联免疫吸附试验（ELISA）检测 DC 培养上清液中 IL-12 的浓度和与 DC 共培养的 T 细胞上清液中干扰素 γ（IFN-γ）的浓度。噻唑蓝（MTT）比色法评估 IL-12 转染的 DC 冲击肿瘤裂解物刺激自体 T 淋巴细胞增殖和细胞毒性 T 淋巴细胞（CTL）的激活。
成功构建了表达 IL-12 基因的重组腺病毒。转染 IL-12 基因并加用肿瘤裂解物的 DC（DC-IL-12⁺Ag）表达更高水平的 CD83、CD86，产生更高水平的 IL-12，与未转染的 DC（DC⁺Ag）相比（CD83：（60.2 ± 1.8）%比（50.7 ± 1.2）%，P < 0.05；CD86：（88.9 ± 2.1）%比（78.2 ± 3.9）%，P < 0.05；IL-12：（262.5 ± 3.0）ng/L 比（103.8 ± 5.1）ng/L，P < 0.05）。IL-12 转染的 DC 刺激自体 T 淋巴细胞增殖和产生 IFN-γ的作用明显强于未转染的 DC。IL-12 转染的 DC 冲击肿瘤裂解物刺激的 CTL 对喉癌细胞的细胞毒性明显强于未转染的 DC。
IL-12 转染的 DC 冲击肿瘤裂解物可增强抗肿瘤作用，是一种很有前途的方法。
