Effect of RNAi targetingBMI-1 gene on radiosensitivity of esophageal carcinoma cells
10.3969/j.issn.1007-3969.2016.09.007
- VernacularTitle:干扰基因BMI-1表达对食管癌细胞放射敏感性的影响
- Author:
Xingxiao YANG
;
Youmei LI
;
Heng SONG
;
Zhikun LIU
;
Ming MA
;
Shuchai ZHU
- Publication Type:Journal Article
- Keywords:
BMI-1 gene;
RNA interference;
Irradiation;
Radiosensitivity;
p16 gene;
CDK4 gene
- From:China Oncology
2016;26(9):763-769
- CountryChina
- Language:Chinese
-
Abstract:
Background and purpose:B cell-specific MLV integration site 1 (BMI-1) gene plays an important role in DNA damage after exposure to irradiation. The present study aimed to investigate the effect ofBMI-1 on radio-sensitivity of esophageal carcinoma cell after down-regulation of BMI-1 expression by silencing siRNA.Methods:Three pairs of siRNA based on the sequences of the BMI-1 mRNA were synthesized (siRNA1, siRNA2 and siRNA3) by compa-ny, and transfected into cultured TE13 cells as the BMI-1 siRNA groups, and a negative one was synthesized to be used as the negative control (NC) group. The untransfected group was named as the control group. BMI-1 mRNA and protein expression in esophageal cancer TE13 cells were detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blot in different groups. This study used flow cytometry assay to analyze cell cycle of transfected cells, and examined cellular growth and radiosensitivityin vitro by MTT and clone formation assay. mRNA and protein expression of p16 and CDK4 in esophageal cancer TE13 cells were detected by RT-PCR and Western blot.Results:The results of RT-PCR and Western blot showed that the expressions of BMI-1 at gene and protein levels were inhibited after silencing the BMI-1 gene. The mRNA and protein expression of BMI-1 in BMI-1 siRNA3 group were both significantly lower than that in BMI-1 siRNA1 and 2 groups. There was no significant difference in the cell proliferation among control, NC and BMI-1 siRNA3 groups. The values ofD0,Dq, and SF2 in BMI-1 siRNA3 group were 1.761, 2.122 and 0.6255, respectively, obvi-ously lower than those in control group (2.514, 2.694 and 0.8268) and those in NC group (2.506, 2.664 and 0.8231), while the value of N in BMI-1 siRNA3 group (3.336) was higher than that in control group (2.92) and that in NC group (2.895), which showed higher radiosensitivity in BMI-1 siRNA3 group. In addition, the cell cycle was arrested at G2/M phase after irradiation in control and NC groups. The percentage of G0/G1 phase in BMI-1 siRNA3 group was higher than that of control group and NC group, while the percentage of G2/M phase was lower than those in the latter. The up-regulation of p16 and down-regulation of CDK4 at gene and protein levels were detected after knockdown of BMI-1 expression by siRNA (P<0.01).Conclusion:siRNA could inhibitBMI-1 gene expression in esophageal cancer TE13 cells and enhance radiosensitivity, followed by eliminating the cell cycle arrest at G2/M stage after irradiationin vitro, which is related to the regulation of the protein expression ofp16 andCDK4.
