抗白细胞介素8单克隆抗体对子宫颈癌的抑制作用

Inhibitory effect of anti interleukin-8 monoclonal antibodies on cervical cancer

目的研究抗白细胞介素8（IL-8）单克隆抗体对子宫颈癌生长和转移的抑制作用。方法选取高表达IL-8的子宫颈鳞状细胞癌细胞株CaSki和已转染IL-8的SiHa细胞株（pcDNA3.1-IL-8-SiHa），体外实验采用Boyden法观察抗IL-8抗体对高表达IL-8的子宫颈癌细胞趋化能力的影响。体内实验通过建立裸鼠子宫颈癌移植瘤模型，采用反转录聚合酶链反应（RT-PCR）、酶联免疫吸附试验（ELISA）、TUNEL法，观察抗IL-8抗体对子宫颈癌细胞和裸鼠移植瘤生长的影响。实验用裸鼠随机分为10组（CaSki和pcDNA3.1-IL-8-SiHa细胞各5组），每组各5只，空白对照组（Ⅰ组）注射等体积的磷酸盐缓冲液（PBS）；阴性对照组（Ⅱ组）注射等体积的IgG；治疗组（Ⅲ组）注射抗IL-8抗体，100 μg/次，每2 d 1次；治疗组（Ⅳ组）注射抗IL-8抗体，500 μg/次，每3 d 1次；治疗组（Ⅴ组）注射抗IL-8抗体，1 000 μg/次，每周1次。结果体外趋化实验结果显示，CaSki细胞和pcDNA3.1-IL-8-SiHa细胞中不同浓度抗IL-8抗体治疗组的细胞趋化能力均低于空白对照组，差异均有统计学意义（CaSki细胞：F=289.6，P=0.000；pcDNA3.1-IL-8-SiHa细胞：F=79.0，P=0.005）。体内实验中，以抗瘤效果最好的Ⅳ组为例，Ⅳ组肿瘤重量小于Ⅰ组，差异具有统计学意义[接种CaSki细胞：（0.172±0.031）g比（0.735±0.015）g，P〈0.05；接种pcDNA3.1-IL-8-SiHa细胞：（0.400±0.029）g比（1.430±0.199）g，P〈0.05]。Ⅳ组移植瘤体积小于Ⅰ组，差异具有统计学意义[接种CaSki细胞：（0.049±0.028）cm3比（0.214±0.016）cm3，P〈0.05；接种pcDNA3.1-IL-8-SiHa细胞：（0.063±0.022）cm3比（0.600±0.072）cm3，P〈0.05]。肿瘤生长曲线提示抗IL-8抗体治疗组肿瘤生长缓慢，成瘤时间和裸鼠存活时间也相应延长。Ⅳ组IL-8 mRNA表达低于Ⅰ组，差异具有统计学意义（接种CaSki细胞：0.58±0.06比1.15±0.13，P〈0.05；接种pcDNA3.1-IL-8-SiHa细胞：0.69±0.08比1.16±0.13，P〈0.05）。Ⅳ组中IL-8蛋白的表达低于Ⅰ组，差异具有统计学意义（接种CaSki细胞：126±29比411±112，P〈0.05；接种pcDNA3.1-IL-8-SiHa细胞：134±47比327±69，P〈0.05）。Ⅳ组中凋亡指数高于Ⅰ组，差异具有统计学意义（接种CaSki细胞：81.8±3.0比26.0±5.6，P〈0.05；接种pcDNA3.1-IL-8-SiHa细胞：84.4±3.6比32.0±4.9，P〈0.05）。结论抗IL-8抗体可以在体外抑制人子宫颈癌细胞的迁移，在体内抑制移植瘤的生长和转移，促进细胞凋亡和坏死，并呈剂量依赖性。

ObjectiveTo investigate the inhibitory effect of anti interleukin （IL）-8 monoclonal antibodies on the growth and metastasis of cervical cancer.MethodsInvolved cervical cells included CaSki cells with high expression of IL-8 and SiHa cell lines with IL-8 plasmid transfected （pcDNA3.1-IL-8-SiHa）. Cervical cancer animal model was established on nude mice. Boyden method was used in vitro study to observe the effects of anti IL-8 antibodies on the chemotaxis of high-expressed IL-8 cervical cancer cells. The effect of antiIL-8 antibodies on the growth of cervical cancer cells and nude mice transplantation tumor was observed by the experiment in vivo through reverse transcription-polymerase chain reaction （RT-PCR）, enzyme linked immunosorbent assay （ELISA）, TUNEL method. Cell line （CaSki and pcDNA3.1-IL-8-SiHa） modeled nude mice were divided into 5 groups with 5 animals in each group. The blank control group （group Ⅰ） was given the equal volume of phosphate buffer solution （PBS）. Negative control group （group Ⅱ） was injected with IgG at the same volume of IgG. Treatment group （group Ⅲ） was injected with anti IL-8 antibodies at dose of 100 μg for once and intervals for once 2 days. Treatment group （group Ⅳ） was injected with anti IL-8 antibodies at dose of 500 μg for once and intervals for once 3 days. Treatment group （group V） was injected with anti IL-8 antibodies at dose of 1 000 μg for once and intervals for once 1 week.ResultsExperiments in vitro showed that the cell chemotaxis ability of anti IL-8 antibody in CaSki cells and pcDNA3.1-IL-8-SiHa cells was lower than that in the blank control group （CaSki cells： F = 289.6, P = 0.000; pcDNA3.1-IL-8-SiHa cells： F = 79.0, P = 0.005）. Group Ⅳ was taken as the example for its best anti-tumor effect in experiments in vivo. The tumor weight in group Ⅳ was lower than that in group Ⅰ [CaSki cells： （0.172±0.031） g vs. （0.735±0.015） g, P 〈 0.05; pcDNA3.1-IL-8-SiHa cells： （0.400±0.029） g vs. （1.430±0.199） g, P 〈 0.05]. The tumor volume in group Ⅳ was less than that in group Ⅰ [CaSki cells： （0.049±0.028） cm3 vs. （0.214±0.016） cm3, P 〈 0.05; pcDNA3.1-IL-8-SiHa cells： （0.063±0.022） cm3 vs. （0.600±0.072） cm3, P 〈 0.05]. The tumor growth curve also showed that tumor growth was slow, and the time of tumor formation as well as survival time was prolonged in anti IL-8 antibody treated group. The expression of mRNA in IL-8 in group IV was lower than that in group Ⅰ （CaSki cells： 0.58±0.06 vs. 1.15±0.13, P 〈 0.05; pcDNA3.1-IL-8-SiHa cells： 0.69±0.08 vs. 1.16±0.13, P 〈 0.05）. The protein expression of IL-8 in group Ⅳ was lower than that in group Ⅰ （CaSki cells： 126±29 vs. 411±112, P 〈 0.05; pcDNA3.1-IL-8-SiHa cells： 134±47 vs. 327±69, P 〈 0.05）. Apoptotic index in group Ⅳ was higher than that in group Ⅰ （CaSki cells： 81.8±3.0 vs. 26.0±5.6, P 〈 0.05; pcDNA3.1-IL-8-SiHa cells： 84.4±3.6 vs. 32.0±4.9, P 〈 0.05）.ConclusionAnti IL-8 antibody can inhibit cell migration of human cervical cancer in vitro, inhibit growth and metastasis of transplantation tumor in vivo, and promote apoptosis and necrosis with a dose-dependent way in vivo.