重组质粒pIRES—GM—CSF—IL-21的构建及其治疗小鼠肝癌移植瘤的疗效  

Construction of recombinant plasmid plRES-GM-CSF-IL-21 and its therapeutic effect on orthotopically transplanted liver cancer in mice

作　　者：程明荣[1] 李清[2] 洪晓武[3] 叶桃[1] 何秉[1] 程志俭[1] 徐宏智[1] 查兵兵[1] 

机构地区：[1]复旦大学附属上海市第五人民医院普外科,200240 [2]复旦大学附属上海市第五人民医院浦江分院内科,200240 [3]复旦大学上海医学院免疫学系

出　　处：《中华肿瘤杂志》2011年第9期661-665,共5页Chinese Journal of Oncology

基　　金：上海市自然科学基金（09ZRl424700）

摘　　要：目的构建重组质粒plRES—GM—CSF—IL-21，观察其在小鼠体内的抑瘤作用。方法将体外培养的肝癌H22细胞接种于小鼠肝左叶，成瘤后，将小鼠分为5组，每组10只，分别尾静脉注射重组质粒plRES—GM—CSF—IL-21、plRES-GM—CSF、plRES—IL-21、plRES和PBS，观察各组荷瘤小鼠的肿瘤生长情况以及GM—CSF和IL-21的抗肿瘤效应。采用酶联免疫吸附法检测小鼠血清γ干扰素（IFN-γ）和白细胞介素2（IL-2）的含量。采用四甲基偶氮唑蓝法检测小鼠脾脏自然杀伤细胞（NK）和细胞毒T淋巴细胞（CTL）的活性。结果H22组、H22／Neo组、H22／GM—CSF组、H22／IL-21组和H22／GM-CSF—IL-21组小鼠的肿瘤重量分别为（1．591±0．280）g、（1．489±0．155）g、（0．603±0．223）g、（0．583±0．290）g和（0．303±0．323）g，H22／GM—CSF—IL-21组、H22／IL-21组和H22／GM-CSF组小鼠的肿瘤重量明显低于H22组和H22／Neo组（P〈0．01），H22／GM—CSF—IL-21组小鼠的肿瘤重量明显低于H22／GM—CSF组和H22／IL-21组（P〈0．05）。与H22／GM—CSF组和H22／IL-21组比较，H22／GM—CSF-IL-21组小鼠血清中IFN-γ和IL-2的浓度显著升高（P〈0．01），而H22组和H22／Neo组小鼠血清中IFN-γ和IL-2的浓度显著降低（P〈0．01）。与H22／GM—CSF组和H22／IL-21组比较，H22／GM-CSF—IL-21组小鼠脾脏CTL和NK细胞的杀伤活性显著增强（P〈0．01），而H22组和H22／Neo组小鼠脾脏CTL和NK细胞的杀伤活性显著降低（P〈0．01）。结论重组质粒pIRES-GM—CSF-IL-21对小鼠肝癌移植瘤具有明显的抗肿瘤效应，并能促进小鼠体内IFN-γ和IL-2的分泌，增强脾脏中NK和CTL细胞的活性，其效果优于单一GM—CSF或IL-21基因治疗。Objective To construct a recombinant plasmid pIRES-GM-CSF-IL-21, and to investigate its antitumor effect on tumors in the mice. Methods Fifty Bal b/c mice were included in this study. Cultured hepatoma H22 cells were inoculated in the left lobe of the liver to develop orthotopically transplanted liver tumor models. The mice with orthotopically transplanted liver tumor were randomly divided into 5 groups （ n = 10） ： （ 1 ） Each mouse received injection of recombinant plasmid pIRES-GM-CSF-IL-21 ; （2） Received injection of plasmid pIRES-GM-CSF ; （ 3 ） pIRES-IL-21 ; （4） Received injection of ampty plasmid pIRES （H22/neo group ）; （5） Received injection of PBS （H22 group ） via the tail vein, respectively. Therefore, the anti-tumor effect was induced by both GM-CSF and IL-21, or by either of them alone. The serum levels of IFN-γ/and IL-2 were detected by ELISA, and the cytotoxicity of spleen NK and CTL cells were tested by MTT colorimetry. Results Comparing with the H22 and H22/Neo groups, the tumor weight in the mice of H22/GM-CSF group was （0. 603 ±0. 223）g, H22/IL21-treated group （0. 583 ± 0.290） g and H22/GM-CSF-IL21-treated group （0.303 ± 0. 323 ）g, significantly lower than that in the H22 group [ （ 1.591 ± 0.280） g ] and H22/Nco group [ （ 1. 489 ± 0. 155 ） g]. Among them the tumor growth was most significantly inhibited in the H22/GM-CSF-IL-21 group （0.303± 0.323 ）g, compared with that of H22 and H22/neo groups （ P 〈 0.01 ）. But there was no significant difference between the tumor weights of the H22/GM-CSF group and H22/IL-21 group, and between the tumor weights of the H22 and H22/Neo groups （ P 〉 0. 05 ）. The levels of IFN-γ/and IL-2 in peripheral blood of mouse models treated with H22/GM-CSF- IL-21 were significantly increased than that in the H22/GM-CSF group and H22/IL-21 group （all P 〈 0.01）, but significantly decreased in the H22group and H22/Neo group （ P 〈 0.01 ）. The anti-tumor activity of splenic NK cells and CTLs i

关 键 词：免疫 细胞 粒细胞-巨噬细胞集落刺激因子 白细胞介素21 Γ干扰素 肝肿瘤 

分 类 号：R735.7[医药卫生—肿瘤]