Effect Of MACC1 Overexpression On Radiotherapy Sensitivity Of Nasopharyngeal Carcinoma CNE-2 Cells And Its Mechanism

Objective: To investigate the effect of MACC1 overexpression on radiotherapy sensitivity of nasopharyngeal carcinoma CNE-2 cells and its mechanism.Methods :(1)The protein expression levels of MACC1 in 5-8F,HNE-1,CNE-1 and CNE-2 cell lines of nasopharyngeal carcinoma were detected by Western blot.(2)CNE-2 cell lines overexpressing MACC1 were established by lentivirus transfection technology,and stable transfected cells were screened.(3)QPCR and WB assays were used to detect the expressions of MACC1 mRNA and MACC1 protein,and subsequent cellular functional experiments were conducted after confirming that the upregulation efficiency was stable and effective.(3)The experiment was divided into three groups: experimental group(CNE-2 MACC1 overexpression stable strain group);Negative control group(CNE-2negative stable strain group);Blank control group(CNE-2 empty cell group).The radiosensitivity of CNE-2 cells after overexpression of MACC1 was detected by clonal formation assay.(4)Flow cytometry(FCM)was used to detect the effect of MACC1 overexpression on cell cycle and apoptosis of CNE-2 cells.Results :(1)The instantaneous transfection efficiency of CNE-2 cells was about 80-90%.Inverted fluorescence microscopy showed GFP fluorescence in the transfected cells.(2)The MACC1 gene transfection CNE-2 cells after 48 h,by QPCR detection,through the 2-? ? Ct analysis experimental MACC1 Ct value is 17.1,prompt MACC1 mRNA expression.Western blot assay showed obvious cell bands and overexpression of MACC1 protein in the experimental group,suggesting that the CNE-2 cell line of nasopharyngeal carcinoma with overexpression of MACC1 was successfully constructed.(3)Cloning formation experiment showed that:at 0,2,4,6Gy radiation dose,the survival scores(SF)of the experimental group were higher than those of the blank control group and the negative control group(p<0.05).The survival score(SF)of the blank control group was not significantly different from that of the negative control group(p>0.05).The single-click multi-target model was constructed to fit the cell survival curve.D0,Dq and N of the experimental group,blank control group and negative control group were(3.48,1.61,1.59),(2.37,1.26,1.70)and(2.68,1.20,1.57),respectively.The radiosensitization ratios(SER)between the experimental group and the blank control group and the negative control group were 0.68 and 0.77,respectively.(4)theresults of flow cytometry showed that compared with the blank control group and the negative control group,in the experimental group cell cycle G0-G1 phase distribution was decreased,the S phase distribution was increased,and the apoptosis rate was decreased(p<0.05).Conclusion: overexpression of MACC1 can reduce the radiotherapy sensitivity of nasopharyngeal carcinoma CNE-2 cells,which may be related to the decreased distribution of G0-G1 phase,increased distribution of S phase and decreased apoptosis of MACC1 overexpressed CNE-2 cells in nasopharyngeal carcinoma.