| Objective: To investigate the effect of dichloroacetate(DCA)on the proliferation,invasion and migration of human bladder cancer T24 cell line and its possible mechanism,providing a theoretical basis of the Anti-tumor therapy with DCA.Methods: Human bladder cancer cell line T24 was selected as the research object and incubated under appropriate environmental conditions.Different concentrations of DCA were added to the culture medium of the logarithmic growth phase of human bladder cancer T24 cell line.The effect of DCA on the proliferation of bladder cancer T24 cells was detected by MTT assay.Colony formation assays were detected with Giemsa staining.Cell wound scratch assay and Transwell assay were applied to evaluate the ability of the T24 cell invasion and migration and the apoptosis was examined by Hoechst33258 staining.Western blot was used to detect the protein expression of Bcl-2,Bax,caspase-3,E-cadherin,Slug,Snail,vimentin and N-cadherin.Results: The MTT assay results showed that human bladder cancer cell line T24 were cultured with different concentrations of DCA(0 mM,1.25 mM,2.5 mM,5 mM,10 mM,20 mM,40 mM)for 24 hours,48 hours and 72 hours.It was found that DCA had no significant inhibitory effect on human bladder cancer cell line T24 at a concentration of 1.25 mM and 2.5mM,with no significant difference(P>0.05).However,in the experimental group,the survival rates of human bladder cancer T24 cells were lower than that of the control group at the concentrations of 5mM,10 mM,20mM and 40 mM.And with the increase of DCA concentration and administration time,the survival rate of bladder cancer T24 cells decreased significantly,the difference was statistically significant(P <0.05).Cell cloning experiments showed that compared with the control group,DCA could reduce the clonality of human bladder cancer T24 cell line in a concentration-dependent manner(P <0.05).Cell scratch test results show that at 0h,the scratch width of the control group and each experimental group was basically the same.However,after 24 h,48h and 72 h exposure to 5mM,10 mM and 20 mM DCA,the human bladder cancer T24 cells migration rate showed different degrees of reduction.Compared with the control group,the difference was statistically significant(P<0.05),and in a time and concentration-dependent manner.The results of Transwell assay showed that the number of invasive cells in the experimental group treated with 5mM,10 mM and 20 mM DCA for 24 h were(361.7 ± 20)/ field,(165.0 ± 17.1)/ field and(71.0 ± 16.1)/ field,respectively.Compared with the control group(536.7 ± 27.6)/ field,the difference was statistically significant(P <0.01),indicating that DCA can invade the invasiveness of human bladder cancer T24 cell lines in a concentration-dependent manner.Hoechst33258 fluorescence staining results showed that the apoptotic rate of T24 cells in human bladder cancer cells treated with 5mM,10 mM and 20 mM DCA for 48 h were(9 ± 1.83)/ HP,(26.5 ± 5.8)/ HP and(48.75 ± 2.99)/ HP,respectively.Compared with the control group(3.25 ± 0.96)/ HP,the difference was statistically significant(all P <0.05),indicating that DCA can promote the apoptosis of human bladder cancer T24 cell line in a concentration-dependent manner.Western blot results showed that after treated with different concentrations of DCA(0mM,5mM,10 mM and 20mM)for 48 h,the protein levels of E-cadherin,Bax and caspase-3 were up-regulated with the increase of DCA concentration.Compared with the control group,the difference was statistically significant(P <0.05).However,the protein levels of Bcl-2,Slug,Snail,vimentin and N-cadherin decreased with the increase of DCA concentration,which was significantly different from the control group(P <0.05).Conclusion: DCA can effectively inhibit the proliferation,clone formation,invasion and migration of human bladder cancer T24 cells and induce its apoptosis.The mechanism may be related to mitochondrial apoptosis pathway and epithelial-mesenchymal transition. |
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