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European journal of gynaecological oncology

Effect of lentivirus mediated cyclooxygenase-2 gene shorthairpinRNA on invasiveness of endometrial carcinoma.


PMID 25872333

Abstract

This study aims to explore the effects of lentivirus mediated cyclooxygenase-2 gene shorthairpinRNA (COX-2-shRNA) on invasiveness of endometrial carcinoma HEC-1B cells. Double-stranded DNA oligonucleotide of COX-2-shRNA was designed and synthesized, and the recombinant lentiviral vector COX-2-ShRNA (LV-COX-2-ShRNA) was constructed. LV-COX-2-ShRNA, pHelper 1, and pHelper 2 were transferred into 293T cells, followed by lentiviral packaging. The virus titer was tested according to expression level of GFP in 293T cells. HEC-1B cells were infected with recombinant lentivirus. The silencing of COX-2 gene was assessed by real-time PCR and western-blot, and the in vivo invasiveness of HEC-1B cells was analyzed by transwell invasion assay. Recombinant lentiviral vector expressing siRNA targeting COX-2 gene was successfully constructed to harvest the recombinant lentivirus with the concentrated virus suspension titer of 5 x 10(7)Tu/ml. Compared with control group, the inhibitory rate of COX-2 expression in HEC-1B cells in siRNA group were 61.87% and 67.48% at mRNA and protein level, respectively. The mean number of cells penetrating matrigel was 16.6, which was significantly less than the control group 50.2 and non-specific siRNA infection group 47.2, the invasion inhibition rate being 64.8% (p < 0.01). RNA interference can inhibit the invasiveness of HEC-in cells.