The Expression And Mechanism Of Invasion And Migration Of Matriptase Gene In Human Ovarian Cancer Cell Lines HO-8910and HO-8910PM

Objective：To detect the different expression of Matriptase in different metastaticpotential of human ovarian cancer cells. And disturb Matriptase gene expression inHO-8910PM cells by RNAi，In order to investigate the relationship in Matriptasegene on cell migration and invasion ability in ovarian cancer cell lines.Methods: High-metastatic human ovarian cancer line HO-8910PM and ovariancancer cell HO-8910were collected.The ability of metastatic of the former wasstronger than that of the latter. Compare the ability of invasion and migration inHO-8910PM and HO-8910by scratch assay and by millicell chamber artificialreconstituted basement membrane invasion assay. Detect Matriptase mRNAexpression levels in HO-8910PM and HO-8910through RT-PCR and Real-time PCR;Detect Matriptase protein expression levels in HO-8910PM and HO-8910throughimmunocytochemistry methods and Western blot. RNAi techenique was applied toconstruct three Matriptase gene lentivirus siRNA expression vector and transfectovarian cancer lines HO-8910PM. Real-time PCR and Western blot were used toobserve the inhibitory effect of RNAi on Matriptase mRNA and protein expression.Transwell migration assay was used to detect the effect of Matriptase geneknockdown on the invasion ability of HO-8910PM cells. Scratch assay was used todetect the effect of Matriptase gene knockdown on the migration ability ofHO-8910PM cells.Results:1.The24hour’ migration distance （347.23±8.41）μm of HO-8910PM cells weresignificantly higer than HO-8910group (153.95±9.56)μm (P<0.01).2.The number90.67±2.08of HO-8910PM cells that penetrated the Matrigel after24hour’incubation were significantly higer than in HO-8910group63.33±1.52(P<0.01). 3.The expression of Matripase mRNA0.724±0.026in HO-8910PM cells washigher than HO-8910group0.376±0.037(P<0.01); and the migration was positivelycorrelated with the Matripase mRNA expression levels(r=0.992, P<0.01);and theinvassiveness was also positively correlated with the Matripase mRNA expressionlevels(r=0.969, P<0.01).Real-time PCR results showed the expression of MatripasemRNA0.001330±0.000025in HO-8910PM cells was higher than HO-8910group0.000723±0.000004（P<0.01）.4.As far protein level, immunocytochemistry results showed that Matripaseexpression is maily in the cytoplasm and cell membrane，and HO-8910PM cells wasobviously lighter than that in HO-8910group,The expression of Matripase protein15.63±0.83in HO-8910PM cells was higher than HO-8910group7.65±1.30(P<0.01).and the migration was positively correlated with the Matripase protein expressionlevels(r=0.971，P＜0.05);and the invassiveness was also positively correlated withthe Matripase protein expression levels（r=0.958，P＜0.05）.Western blot resultsshowed that the expression of Matripase protein0.6692±0.0216in HO-8910PM cellswas higher than HO-8910group0.3327±0.0107(P<0.01).5.Three Matriptase lentivirus siRNA plasmid expression vectors wereconstructed successfully.Compared with Control cell group， the expression ofMatriptase mRNA was decreased by19.63%，32.43%and89.72%in HO-8910PMcells tranfected with siRNA8985-1，siRNA8988-1，siRNA8987-3respectively，andsiRNA8987-3was the most effective one among the three siRNAs. Meanwhile，Western blot results showed that siRNA8987-3inhibited Matriptase proteinexpression significantly (P<0.01).6.Transwell assay results showed that Matriptase gene silencing inhibited theinvasive ability of HO-8910PM cells significantly (P<0.01).7.Scratch assay results showed that Matriptase gene silencing inhibited themigration ability of HO-8910PM cells significantly (P<0.01).Conclusion:1.Matriptase highly expressed in HO-8910PM and lowly expressed in HO-8910 indicated that it may related with tumor metastasis．2. Knockdown of Matriptase suppressed invasive and migration capability ofovarian cancer cells HO-8910PM.Matriptase may played a critical role in themetastasis of human ovarian cancer.