宫颈癌细胞促进血管新生斑马鱼模型的建立

Pro-angiogenic Effects of Cervical Cancer Cells in a Zebrafish Xenotransplantation Model

[目的]建立宫颈癌细胞异种移植斑马鱼模型,研究其对血管新生的影响.[方法]以斑马鱼作为研究对象,使用线性化的红色荧光质粒pcDNA3.0＋DsRed转染人宫颈癌SiHa细胞,通过显微注射法将稳定表达红色荧光的SiHa细胞移植入48 hpf（发育后时数）的FLK转基因斑马鱼胚胎内,建立人宫颈癌斑马鱼模型,观察人宫颈癌细胞在斑马鱼胚胎体内存活、增值和诱导血管增生情况,探讨宫颈癌细胞对斑马鱼模型血管新生的影响.[结果]注射不同数量人宫颈癌SiHa细胞对斑马鱼卵有不同的影响,其中注射50～ 100个细胞产生最佳的促进血管新生作用.注射Matrigel的对照组斑马鱼在注射后的第一天和第二天突出到肠下静脉血管篮外到卵黄的垂直分枝平均血管数频率分别为2.8％±1.4％和5.4％±2.2％.而注射人宫颈癌细胞系SiHa的斑马鱼的新生血管数比对照组明显增加,在注射后第一天和第二天突出到肠下静脉血管篮外到卵黄的垂直分枝平均血管数频率分别为8.0％±2.7％和27.4％±3.2％（P＜0.01）.[结论]本研究成功建立宫颈癌细胞促进血管新生的斑马鱼模型,这为研究宫颈癌的发病机制、筛选新的抗肿瘤血管生成化合物和基因靶点研究提供了新的技术平台.

[Objective] To establish a zebrafish xenotransplantation model of cervical cancer and investigate the pro-angiogenic effects of cervical cancer.[Methods] A linearized plasmid pcDNA3.0 vector expressing Ds-Red fluorescent protein were used to fluorescently label human cervical cancer SiHa cells.The fluorescent-labeled SiHa cells were then transplanted into the FLK 48 hpf transgenic zebrafish embryos using microinjection.The survival,proliferation and induction of angiogenesis were evaluated in zebrafish xenotransplantation embryos and larvae.[Results] Different numbers of xenograft human cervical cancer SiHa cells had different effects on zebrafish and transplantation of 50-100 c SiHa cells achieved an excellent angiogenesis.Xenograft SiHa cells significantly increased new blood vessels compared with the control group.The numbers of the vertical branch of subintestinal vein outside yolk in matrigel control group and xenograft group were 2.8 ± 1.4％ and 8.0 ± 2.7％ in day 1,5.4 ± 2.2％ and 7.4 ± 3.2％ in day 2 after injection （all P ＜ 0.01）.[Conclusion] This study has successfully established zebrafish xenotransplantation model of cervical cancer and demonstrated the pro-angiogenesis of cervical cancer,which provides a new technology platform for the study of the pathogenesis of cervical cancer development of new compounds and gene targets for anti-tumor angiogenesis.