慢病毒介导人乳头瘤病毒16型E6基因短发夹状干扰RNA对宫颈癌细胞感染效率和侵袭能力的影响

目的 研究以慢病毒为载体靶向人乳头瘤病毒16型(HPV16)E6基因的短发夹状干扰RNA(shRNA)对宫颈癌Caski细胞侵袭能力的影响.方法 将靶向HPV16型E6的shRNA表达序列(干扰组)和无义表达(无义干扰组)克隆到慢病毒工作质粒中包装成病毒颗粒,感染宫颈癌Caski细胞.观察绿色荧光阳性细胞的比例以了解慢病毒对Caski细胞的感染效率;用RT-PCR法检测shRNA作用后3组Caski细胞HPV16型E6 mRNA含量的变化;Western blot法检测E6蛋白表达的变化;Transwell小室法检测肿瘤细胞侵袭能力的变化.结果 慢病毒对Caski细胞的最佳每个细胞中的病毒颗粒数目(MOI)为2.5;与无义干扰组相比,干扰组细胞中HPV16E6的mRNA含量减少70%,蛋白表达水平降低63%,Caski细胞的侵袭能力显著下降.结论 慢病毒携带的shRNA能干扰宫颈癌Caski细胞中HPV16型E6的表达,并抑制癌细胞的侵袭能力.

Effect of human papilloma virus16 E6 shRNA mediated by lentivirus on infection ratio and invasive ability of cervical cancer cell

Objective To investigate the influence of lentivirus mediated short hairpin RNA(shRNA)target to human papilloma virus(HPV)16 E6 on invasive ability of cervical cancer Caski cells.Methods Lentivirus was produced after shRNA target to human papilloma virus(HPV)16 E6 and to nonsense was cloned to lentivirus work vector.Infection ratio was assessed by assay of EGFP positive cells of Caski.Total mRNA of E6 was determined by RT-PCR after Caski cells were infected by lentivirus.The change of E6 protein expression was analyzed by Western blot.The invasive ability of Caski cells was assayed employing Transwell.Results The optimal MOI (Multiplicity of infection)of lentivirus to Caski was 2.5.Total mRNA and protein of E6 were decreased (by 70%and 63%)in interfering group compared with control group.The invasive ability of Caski cells also reduced after infected by lentivirus.Conclusion shRNA mediated by lentivirus can inhibit expression of HPV16 E6 and invasive ability of cervical cancer cells.