The Expression Of MGMT And Survivin On Radiosensitivity And Chemosensitivity Of Esophageal Squamous Cancer Cell Lines

Objective To compare the sensitivity of KYSE-450,KYSE-70 and TE-1 esophageal squamous cancer cell lines to radiation and chemotherapeutic drugs which were5-fluorouracil and cisplatin.And compare the MGMT and Survivin protein expression.Methods The survival fraction of KYSE-450,KYSE-70 and TE-1 esophageal squamous cancer cell lines in different concentrations of cisplatin and 5-fluorouracil was measured using thiazolyl blue tetrazolium blue-base(MTT).Using IC50 value to evaluate the tolerance of celllines to chemotherapeutic drugs.The survival fraction of 3cell lines irradiated at different doses was detected by the clone formation assay.The radiation biological parameters of D0、Dq、N、SF2 were calculated by using the multiple target model to fit the dose survival curve to evaluate the sensitivity of the 3 cell lines to the radiation.The expression of MGMT and Survivin in 3 cell lines were detected by blot Western and q RT-PCR.Results Firstly,the survival curves of irradiated dose and their biological parameters of KYSE-450,KYSE-70 and TE-1 esophageal squamous cancer cell lines.The cell survival curve was fitted by Graph Pad Prism 5.0.The cell survival fractions of three esophageal squamous cancer cell lines decreased with the increase of irradiation dose.These results indicate that KYSE-70 was more sensitive to radiation while TE-1 and KYSE-450 were more resistant to radiation.Secondly,dose survival curves of KYSE-450,KYSE-70 and TE-1 cells in different concentrations of DDP and 5-FU and their IC50 values.The cell survival rate of esophageal squamous cancer cell lines of KYSE-450,KYSE-70 and TE-1 were significantly different in different concentrations of chemotherapeutic drugs.It showed that the tolerance to different concentrations of chemotherapeutic drugs was significantly different.According to the IC50 calculation software to calculate the IC50 relative to the different kinds of chemotherapeutic drugs in different cell lines,also confirmed that esophageal squamous cancer cell line of TE-1was more resistant to chemotherapeutic drugs DDP and 5-FU.Because IC50 TE-1 cells to DDP value was about 9 times the KYSE-70 and TE-1 cells on the value of 5-FU was about 133 times the KYSE-70.Compared with the IC50 value of TE-1 cells,the difference was statistically significant(F=199.9,157.9,P < 0.05).Thirdly,the expression of MGMT and Survivin protein in KYSE-450,KYSE-70 and TE-1 cell lines.The expression of MGMT protein and Survivin protein in TE-1 cells was the highest,followed by KYSE-70 and KYSE-450.Compared with TE-1 cells MGMT protein and Survivin protein expression level,the difference was statistically significant(F =367.7,213.5,P <0.05).Fourthly,FQ-PCR analysis.The expression of MGMT and Survivin in TE-1 which was tolerant cell line and KYSE-70 which was susceptible cell line were analyzed.Beta-actin was used as a control and Real-time PCR was used to verify the different expression.MGMT in TE-1 cells,compared with KYSE-70 high expression(5.47±0.32)times;Survivin in TE-1 cells,compared with KYSE-70 high expression(2.73±0.26)times,the difference was statistically significant(t=24.633,6.839,P <0.05).The expression of MGMT and Survivin in TE-1 were higher than KYSE-70,the difference was statistically significant(P <0.05).Conclusion In the experiment using MTT to detect TE-1 and KYSE-70 chemotherapy sensitivity experiment,TE-1 in cisplatin and fluorouracil in IC50 was higher than that of KYSE-70(P < 0.05),showed that in the chemotherapy of cisplatin and fluorouracil KYSE-70 is more sensitive to it and TE-1 is more tolerant.In the clone formation experiment of TE-1 and KYSE-70,the cell survival curve and the parameters showed that TE-1 was better than KYSE-70 in radiation tolerance(P <0.05).The expression of MGMT and Survivin in TE-1 and KYSE-70 was detected by Western blot.The results showed that the expression of MGMT protein and Survivin protein in the TE-1cells was the highest,followed by KYSE-70 and KYSE-450.The expression of MGMT and Survivin in TE-1 and KYSE-70 was detected by real-time fluorescence quantitative PCR.The results showed that there were high expression of both MGMT and Survivin in TE-1 cells compared with KYSE-70;Survivin in TE-1 cells,compared with KYSE-70 high expression.MGMT and Survivin expression levels were higher in the cell lines resistant to chemotherapy and radiotherapy.It was lower in the cell lines sensitive tochemotherapy and radiotherapy.It provided experimental basis for the study of the mechanism of esophageal squamous cell carcinoma with radiotherapy and chemotherapy.