IL-21基因对宫颈癌HeLa细胞抑制作用的体外试验研究

The inhibitory effect of IL-21 gene on cervical carcinoma HeLa cells in vitro

目的探讨白介素-21(interleukin-21,IL-21)基因对宫颈癌细胞(HeLa细胞)的作用。方法构建人源性IL-21质粒并电转染至体外培养的宫颈癌细胞(HeLa细胞),选择性培养获得表达IL-21的HeLa细胞(HeLa-IL-21)。将96只重症联合免疫缺陷(severe combined immune deficiency,SCID)小鼠随机分为免疫重建组与非免疫重建组,每组各48只。采用人外周血淋巴细胞重建免疫重建组小鼠的免疫功能。再分别将两组小鼠随机均分为A、B、C 3个亚组,每组各16只。其中A组小鼠接种HeLa细胞,B组小鼠接种HeLa-空载体细胞,C组小鼠接种HeLa-IL-21细胞。比较各组小鼠肿瘤生长情况、小鼠脾细胞与不同靶细胞作用时干扰素-γ(IFN-γ)表达水平及脾细胞毒性,以探究IL-21基因抗肿瘤的作用机制。结果接种后第20、30、40天,非免疫重建A组小鼠肿瘤体积分别为(28.4±3.7)mm^3、(95.7±13.0)mm^3、(232.4±21.6)mm^3,均明显大于免疫重建A组小鼠[(19.7±2.9)mm^3、(65.7±12.7)mm^3、(154.7±16.9)mm^3](P<0.05),免疫重建C组小鼠肿瘤体积分别为(16.4±2.7)mm^3、(48.5±11.0)mm^3、(53.4±10.7)mm^3,均明显大于免疫重建C组小鼠[(12.4±2.1)mm^3、(22.7±5.6)mm^3、(37.4±10.3)mm^3,](P<0.05),免疫重建C组小鼠肿瘤体积均明显小于免疫重建A组小鼠(P<0.05),非免疫重建C组小鼠肿瘤体积均明显小于非免疫重建A组小鼠(P<0.05)。检测不同来源小鼠脾细胞与不同靶细胞作用时IFN-γ表达水平发现,HeLa细胞作为靶细胞时,各组小鼠IFN-γ高表达(P<0.05);较非免疫重建组小鼠,免疫重建组小鼠对不同靶细胞IFN-γ高表达(P<0.05);较免疫重建A、B组小鼠,免疫重建C组IFN-γ高表达(P<0.05)。小鼠脾细胞毒性实验发现,转染IL-21质粒能增加小鼠脾细胞对HeLa细胞的杀伤毒性。结论细胞因子IL-21有一定的抗肿瘤作用,可能与其增强T细胞对肿瘤细胞的杀伤作用有关,为宫颈癌患者的免疫治疗奠定了一定的研究基础。

Objective To investigate the effect of IL-21 gene on HeLa cells of cervical cancer. Method Constructed anthropogenic IL-21 plasmid and electric transfected to cervical cancer ceils （HeLa） in vitro culture to selective gain HeLa cell expressed IL-21 （HeLa-IL-21）. Immunized 48 severe combined immune deficiency （SCID） mice （immune reconstruction group） with human peripheral blood lymphocyte to reconstruction their immune function and selected another 48 SCID mice as non-reconstruction group. Then divided both groups into equal 3 subgroups as A, B, C group. And inoculated HeLa cells, HeLaempty cells, HeLa cells-IL-21, respectively. Compared tumor growth, IFN-T expression level and spleen cell toxicity between groups so as to explore antiturnor mechanism oflL-21 gene. Result 20, 30, 40 days after inoculation, the tumor volume of non-immune reconstruction A group were （28.4 ± 3.7） mm3, （95.7 ± 13.0） mm3, （232.4 ± 21.6） mm3, which were significantly larger than immune reconstruction A group [（19.7±2.9） mm3, （65.7± 12.7） mm3, （154.7±16.9） mm3] （P 〈 0.05）; tumor volume of non-immune rebuild C group were （16.4± 2.7） mm3, （48.5 ± 11.0） mm3, （53.4±10.7） mm3, which were significantly larger than immune reconstruction C group [（12.4±2.1） mm3, （22.7±5.6） mm3, （37.4± 10.3） mm3] （P 〈 0.05） ; tumor volume of immune reconstruction C group were smaller size compared to immune reconstruction A group （P 〈 0.05）; tumor volume of non-immune reconstruction C group were significantly smaller than non-immune reconstruction （P 〈 0.05）, When HeLa ceils setted as target cells, IFN-γ expressed high level in immune reconstitution mice can express higher levels of IFN-γ, the transfected IL-21 plasmid can express higher levels of IFN-γ. Mice spleen cell toxicity test showed that the transfection of IL-21 plasmid can increase the cytotoxicity ofmurine spleen cells to HeLa cells. Conelusion IL-21 has a certain anti-tumor effect, that may be relate to IL-21 can reinforce the tumor cell killing effect ofT cells, in order to provide a certain basis for the clinical treatment of cervical cancer immunotherapy.