shRNA靶向干扰COX-2促进人肝癌裸鼠皮下移植瘤肿瘤细胞凋亡

Promotive effect of cyclooxygenase-2 gene inhibited by shRNA on the tumor cells apoptosis of subcutaneous xenograft human liver cancers in nude mice

目的:探讨重组干扰质粒pshRNA-COX-2对人肝癌细胞Hep3B裸鼠皮下移植瘤肿瘤细胞凋亡的促进作用。方法:重组干扰质粒pshRNA-COX-2转染Hep3B细胞并筛选后,将被成功转染的Hep3B细胞种植于裸鼠皮下,测量肿瘤大小,成瘤4周后处死裸鼠,绘制肿瘤生长曲线,RT-PCR和Western blot检测肿瘤组织中COX-2 mRNA和蛋白表达,电镜观察和TUNEL法检测肿瘤细胞凋亡。结果:与空白组相比,干扰组肿瘤组织中COX-2 mRNA和蛋白表达抑制率分别为48.5%和61.4%。干扰组最终瘤体大小明显小于阴性组和空白组<P<0.01)。电镜观察显示干扰组凋亡细胞增多,TUNEL法显示干扰组肿瘤细胞凋亡指数明显高于阴性组和空白组(P<0.01)。结论:pshRNA-COX-2通过靶向抑制COX-2基因表达,可明显抑制人肝癌细胞Hep3B裸鼠皮下移植瘤生长,促进肿瘤细胞凋亡。

Objective：To investigate the promotive effect of recombinant interference plasmid pshRNA-COX-2 on the tumor cells apoptosis of human liver cancer cell Hep3B subcutaneous xenograft tumors in nude mice.Methods： Hep3B cells were transplanted into the subcutaneous tissue of nude mice after Hep3B cells were transfected with plasmid pshRNA-COX-2 and selected.Xenograft tumor volumes were measured every three days and growth curves of tumors were drawn.The mice were killed four weeks after tumors formation.The COX-2 mRNA and protein expressions in tumor tissue were measured by reverse transcription polymerase chain reaction（RT-PCR） and Western blot respectively.The tumor cells apoptosis was detected by electron microscope observation and TdT-mediated dUTP nick end labeling in situ assay（TUNEL）.Results： Compared to the untreated group,the inhibition rates of COX-2 mRNA and protein expressions in tumor tissue were 48.5% and 61.4% respectively in the pshRNA-COX-2 group.The ultimate tumor volume in the pshRNA-COX-2 group was significantly smaller than the pshRNA-HK group and the untreated group（P0.01）.The apoptosis cells increased in the pshRNA-COX-2 group observed with electron microscopy,and results of TUNEL revealed that the apoptosis index（AI） of tumor cells in the pshRNA-COX-2 group was much higher than the pshRNA-HK group and the untreated group（P0.01）.Conclusions： The recombinant interference plasmid pshRNA-COX-2 can significantly promote the tumor cells apoptosis of human liver cancer cell Hep3B subcutaneous xenografts in nude mice through targeted inhibiting COX-2 expression.