Inhibitory Effect Of MiR-145 On Transplanted Tumor Of Ovarian Carcinoma In Nude Mice

Objective: To investigate the effect of miR-145 on invasion and migration of ovarian cancer SKOV3 cells and its possible mechanism,and to provide a new direction for the clinical treatment of ovarian carcinoma.Methods:(1)The establishment of stable expression of miR-145 cell lines(miR-145-CMV-RFP-SKOV3)and negative control cell lines(mock-CMW-RFPSKOV3):to lentivirus as a carrier for packaging,lentivirus was infected with SKOV3 cells,puromycin screened stable cell lines,picking monoclonal and expanding culture,multiple screening of strong secretion of the target protein cell lines,to obtain stable expression of miR-145 cell lines(miR-145-CMV-RFP-SKOV3)and negative control cell lines(mock-CMW-RFP-SKOV3);(2)The establishment of subcutaneous transplantation model of SKOV3 cells in nude mice and in vivo experiment of nude mice:establishment of subcutaneous transplantation model of SKOV3 cells in nude mice,observation of tumor growth,draw the growth curve,bioluminescence imaging technique for tracing the growth and spread of ovarian cancer in nude mice;Mice were sacrificed to obtain tumor tissue,Western blot was used to detect the expression of MUC1 and EMT marker protein(E-cadherin ?N-cadherin ?Fibronectin?Vimentin)in tumor tissue.Results: The cell lines expressing miR-145 and red fluorescent protein(miR-145-CMV-RFP-SKOV3)and negative control cell lines expressing nonsense sequences sequence and red fluorescent protein(mock-CMW-RFP-SKOV3)stably were successfully constructed;Successful establishment of three groups of SKOV3 cells subcutaneously transplanted in nude mice model;Draw a tumor growthcurve:Compared with the blank control group(SKOV3),the tumor volume of nude mice was significantly decreased in experimental group(miR-145-CMV-RFP-SKOV3),the difference was statistically significant(P <0.01).Compared with the blank control group(SKOV3),the negative control group(mock-CMW-RFP-SKOV3)had a reduced tumor volume in nude mice,but had no significant difference(P> 0.05);Tumor volume increased with time,The expression of luciferase was observed in the experimental group(miR-145-CMV-RFP-SKOV3)after 5 days of inoculation,and the expression of luciferase in the negative control group was much higher than that in the experimental group.With the increase in the number of days of inoculation,cell concentration increased,the expression of fluorescein increased rapidly;The expression of MUC1,N-cadherin,Fibronectin and Vimentin was significantly decreased,while the expression of E-cadherin was increased by Western Blot in the experimental group.Conclusion: In this study,we successfully constructed cell lines stably expressing miR-145 and red fluorescent protein(miR-145-CMV-RFP-SKOV3)and negative control cell lines expressing nonsense sequences and red fluorescent protein(mock-CMW-RFP-SKOV3).Experiments in nude mice showed that,the high expression of miR-145 inhibits the growth of SKOV3 cell transplanted tumors.Highly expressed miR-145 can reduce the expression of MUC1 protein in ovarian cancer cells.E-cadherin expression increased,the expression of N-cadherin,Fibronectin and Vimentin was significantly decreased,suggesting that miR-145 may inhibit the EMT process of tumor cells and inhibit the invasion and metastasis of ovarian cancer cells.