| Objective:Recently,verteporfin(VP)has been discovered to display anti-tumor function,which makes it a promising adjuvant drug in cancer therapy.However,whether VP has anti-cervical cancer effect and its mechanism is unclear.In this study,we are designed to investigate the effect of VP on proliferation,invasion and migration of cervical cancer cell lines SiHa and HeLa,and to further explore the mechanism behind these effects.Methods:1 The inhibitory effect of different concentrations(0,0.5,1.0,2.0,4.0 ?M)of VP treated for different times(24,48,72 hours)on the proliferation of SiHa and HeLa cells were detected by CCK-8 method.In order to decide a proper time and concentration.2 The cell cloning assay was used to detect the clonality of control goup,1.5?M and 3.0?M VP groups for SiHa and HeLa.3 Transwell migration and invasion assay was used to detecte changes of migration and invasion ability of SiHa and HeLa cells after treated by 1.0?M and 2.5?M VP,respectively.4 The effect of 1.0?M and 2.5?M VP on the cell cycle of HeLa and SiHa cell linses were detected by flow cytometry(FCM).5 The mRNA and protein expression of Yes-associated protein(YAP)and connective tissue growth factor(CTGF)in 2.5?M VP-treated SiHa and HeLa cells were detected by RT-PCR and Western blot.Results:1 The inhibitory effect of VP on proliferation of HeLa and SiHa cells were in a time and dose dependent manner.After treated with VP for 48 hours,the IC50 concentration for SiHa and HeLa were about 2.0?M and 2.5 ?M,respectively.VP has a stronger proliferation inhibition effect on SiHa than HeLa.2 The cell cloning assay showed,the average clonogenic rates of Ctrl,1.5?M and 3.0?M VP goups were 16.4175%,9.8325%,4.9175 for HeLa cells and 18.0000%,7.0825%,3.2500%for SiHa cells.Compared with control group,the clonogenic rates of HeLa and SiHa cells were decreased after treated by 1.5?M or 3.0?M VP(p<0.001),and the 3.0?M group showed a lower clone formation rate than the 1.5?M group among HeLa cells(p<0.05).3 The results of migration assay showed that the number of perforated cells in 2.5?M,1.0?M VP treated groups and control goup were 44.33±3.786,64.33±5.859,169.67±11.676 for SiHa cells and 66.33±6.028,82.00±10.583,159.67±10.504 for HeLa cells(VP vs.Ctrl were p<0.001,2.5?M vs.1.0?M was p=0.047 for SiHa and p=0.177 for HeLa).As to invasion assay,the number of perforated cells in 2.5?M,1.5?M VP treated groups and control goup were 25.33±6.429,58.00±2.000,103.33±7.638 for SiHa cells and 28.33±5.859,77.67±8.505,125.00±5.568 for HeLa cells,respectively(VP vs.Ctrl were p<0.001,2.5pM vs.1.0?M was p=0.001 for SiHa and p<0.001 for HeLa).4 FCM results showed that the G1 phase cell proportion of Ctrl,1.0?M,2.5pM VP groups were 68.83%,75.76%,81.85%for HeLa and 68.31%,76.09%,83.33%for SiHa,which indicated that VP treated cells were G1 phase arrested compared with the control group(p<0.05).5 The expression of YAP and CTGF mRNA and protein in 2.5pM VP-treated cells was significantly decreased(Western blot:HeLa p<0.001,SiHa p<0.05;RT-PCR:p<0.001).Conclusion:VP can effectively suppress proliferation of HeLa and SiHa cells,decrease the migration and invasion ability of cervical cancer cell lines,which indicate that VP has potential anti-cervical cancer effect.The mechanism may be related to down-regulation of Hippo signaling pathway core protein YAP and its target CTGF. |
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