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AIM:To investigate the effect of toosendanin(TSN)on invasion and migration abilities of human ovarian cancer cells and the related mechanism.METHODS:The human ovarian cancer cell lines CAVO-3 and SKVO-3 were treated with TSN at different concentrations.The cell viabilty at 12,24,48,72 and 96 h after TSN treatment was measured by CCK-8 assay.Scratch wound healing assay and Transwell assay were employed to measure the invasion and migration abilities of CAVO-3 cells.The protein expression of nuclear factor-κB(NF-κB)p65,E-cadherin,N-cadherin,vimentin and Snail was determined by Western blot.RESULTS:TSN significantly inhibited the viability of CAVO-3 and SKVO-3 cells(P<0.05 ).Compared with control group ,the migration and invasion abilities of CAVO-3 cells in TSN group decreased significantly(P<0.05).In addition,the expression of NF-κB p65 and E-cadherin protein increased no-tably,followed with N-cadherin,vimentin and Snail protein decreased significantly(P<0.05).However,the inhibitor of NF-κB BAY11-7082 reversed the impact above.Compared with TSN group ,the migration and invasion abilities in TSN +BAY11-7082 group increased significantly(P<0.05).The protein expression of E-cadherin also decreased notably ,fol-lowed with the protein expression of N-cadherin,vimentin and Snail increased significantly(P<0.05).CONCLUSION:TSN inhibits the invasion and migration abilities of human ovarian cancer cells ,which is related to the inhibition of epitheli-al-mesenchymal transition process mediated by NF-κB/Snail signaling pathway.
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<p><b>OBJECTIVE</b>To construct eukaryotic expression plasmids containing green fluorescent protein gene and CYP19 wild-type or its variants (W39R, R264C, W39R-R264C) and to observe its expression in MCF-7 and Bcap-37 cells.</p><p><b>METHODS</b>The aromatase WT cDNA sequence was obtained by RT-PCR amplification and cloned into the eukaryotic expression vector pcDNA3.1(+). pcDNA3.1(+)-CYP19-GFP plasmid was then used as the template for site-directed mutation to create variant constructs (W39R, R264C, W39R-R264C). pcDNA3.1(+)-CYP19-GFP was transfected and expressed in MCF-7 and Bcap-37 cells.</p><p><b>RESULT</b>The construction of pcDNA3.1(+)-CYP19-GFP plasmid was confirmed by enzyme digestion and DNA sequencing. pcDNA3.1(+)-CYP19(W39R)-GFP, pcDNA3.1(+)-CYP19(R264C)-GFP, pcDNA3.1(+)- CYP19(W39R-R264C)-GFP plasmids were confirmed by DNA sequencing. The MCF-7 and Bcap-37 cells transfected with the pcDNA3.1(+)-CYP19-GFP plasmid expressed reporter gene of GFP.</p><p><b>CONCLUSION</b>The eukaryotic expression plasmids have been constructed and expressed in MCF-7 and Bcap-37 cells successfully, which lays the foundation for the research of biological activities of CYP19 variant allozymes.</p>