Effect Of MEG3 Methylation On Cell Proliferation In Bladder Cancer T24 Cells

Objective: bladder cancer is one of the most common tumors in urinary system.Its high recurrence rate and high mortality have a great impact on the body and mind of patients.At present,the gold standard of bladder cancer detection is still cystoscopy,but it is still possible to miss 10% of papillary tumors.Therefore,it has become an urgent problem to seek effective early detection methods and new treatment methods for bladder cancer.The purpose of this study was to investigate the effect of 5-Aza-2’deoxycytidine(5-Aza-CdR),a methyltransferase inhibitor,on the proliferation of bladder cancer T24 cells and the methylation status of meg3 gene in T24 cells,so as to further explore the mechanism of meg3 gene in the occurrence and development of bladder cancer,and to seek a new scheme for the detection and treatment of bladder cancer.Methods: bladder cancer T24 cells were treated with different concentrations of 5-Aza-2’deoxycytidine(5-Aza-CdR)for 24h,48h and 72h respectively,and then the proliferation of T 24 cells was detected by MTT assay.There were 10 groups including zero adjustment group,blank control group,5μmol / l group,10 μmol / l group,15 μmol / l group,20 μmol / l group,25 μmol / l group,30 μmol / l group,35 μmol / l group and 40 μmol / l group.Then using the inhibition rate formula inhibition rate = [(control group od zero group OD)/(experimental group od zero group OD)] * 100% to calculate the inhibition rate of bladder cancer T24 cell proliferation in each drug group,in order to explore the effect of 5-Aza-CdR on the proliferation of bladder cancer T24 cells.The methylation status of MEG 3 gene in bladder cancer T24 cells was detected by bisulfite treated sequencing(BSP)method after 48h of culture in control group(without 5-Aza-Cd R treatment)and 30 μmol/L Group(with 30 μmol/L 5-Aza-CdR treatment).Results:(1)the proliferation of bladder cancer T 24 cells treated with different concentrations of 5-Aza-CdR was inhibited to varying degrees,and the inhibition rates calculated by each concentration group were > 0.(2)It was found that with the increase of 5-Aza-CdR concentration(5 μmol/L,10 μmol/L,15 μmol/L,20 μmol/L,25 μmol/L,30 μmol/L,35 μmol/L,40 μmol/L)at the same time,the proliferation inhibition rate of bladder cancer T24 cells was also gradually increased,and reached the highest value when the concentration of 5-Aza-CdR was 40 μmol / L,but there was no significant difference between some adjacent groups(P>0.05)5)At the same concentration of 5-Aza-CdR,the proliferation inhibition rate of T24 cells increased with the increase of time.Therefore,the proliferation inhibition of bladder cancer T24 cells was positively correlated with the concentration and time of 5-Aza-CdR.(3)The methylation status of MEG3 gene in bladder cancer T24 cells cultured under normal conditions and treated with 30μmol / L5-Aza-CdR for 48 h was detected by BSP method.The results showed that the methylation rate of MEG3 gene in bladder cancer T24 cells treated with 30μmol/L5-Aza-Cd R was lower than that in normal conditions(P<0.01).These results suggest that 5-Aza-CdR can reverse the methylation of MEG3 gene promoter.Conclusion:(1)5-Aza-CdR can effectively inhibit the proliferation of bladder cancer T24 cells.(2)5-Aza-CdR inhibited the proliferation of bladder cancer T24 cells in a concentration and time-dependent manner.(3)5-Aza-Cd R can reverse the methylation of MEG3 gene promoter.(4)Demethylation of meg3 gene promoter can reduce the proliferation of bladder cancer T24 cells.