ABSTRACT
Objective To discuss the role of DNA-dependent protein kinase catalytic subunit (DNA-DPKCS) in human lung adenocarcinoma cell line (A549) by using small interfering RNA (siRNA) to specifically knockdown DNA-DPKCS expression and its effects on cell proliferation, cell cycle and radio-sensitivity. Methods The DNA-DPKCS-siRNA expression vector was constructed and transfected into A549 cell line. The transformed clones were randomly selected and isolated. The cell cycle distribution and apop-tesis were analyzed by flowcytometry analysis. Cell survival was detected by using clonogenic formation as-say. Results With specific inhibition of DNA-DPKCS expression, stable transfected cell line 549pRNA-DNA-DPKCS was constructed by RNA interference technique. The 549pRNA-C and 549pSUPER cell lines were the control cell lines tansfected with control and blank plasmids, respectively. Compared with A549 cells, the expression levels of DNA-DPKCS mRNA (0.110: 1. 000), protein (0. 870: 2.967) and activity of DNA-DPKCS (0.004: 0.266) in 549pRNA-DNA-DPKCS cells were significantly lower (F = 80.55 ,P < 0.01;F=63.96, P<0.01;F=51.62,P<0.01, respectively). The analysis of SF_2(0.25:0.76), D_0 (1.42:1.62) and D_q (0.06: 1. 00) showed significant difference between 549pRNA-DNA-DPKCS and A549 cells (F = 996.86, P < 0.01 ; F = 17.41, P < 0.05 ; F = 68.92, P < 0.01). The number of 549pRNA-DNA-DPKCS cells in S (24.5%: 35.5%) and G_2 (10.7%: 11.0%) phases was significantly decreased (F = 4.83, P<0.05 and F=32.04, P <0.01, respectively). Conclusions In A549 cells, inhibit of DNA-DPKCS gene expression can enhance the radiosensitivity and affect cell cycle distribution.
ABSTRACT
Objective To observe the effect of DNA-PKCS antisense oligodeoxynucleotides (ASODN) on the radiosensitivity of lung cancer cell line A549 through transfecting DNA-PKcs ASODN into A549 cells.Methods DNA-PKCS ASODN and unrelated ODN were transfected into A549 cells (testing group and control group).These cells were irradiated with 0,0.5,1.0,2.0,4.0,6.0 or 8.0 Gy X-ray.Clo nogenic assay was performed to determine the survival fraction.The parameters Do,Dq and N for the multi target single-hit model,as well as the parameters a,13 and SF2 for the linear-quadratic model,were calculated to evaluate cell radiesensitivity.ResultsIn the control group and testing group,the α value was 0.14 and O.31 ,The βvalue was 0.030 and 0.018 ,the SF2 was 0.63 and 0.41 ,the Do was 2.38 and 2.09 ,the Dq was 1.43 and 0.60 ,respectively.In the testing group,the α value of A549 cells was increased,but the β value, SF2,DO and Dq were decreased.Conclusions DNA-PKCS ASODN can enhance the radiosensitivity of A549 cells and is a potential target in treating lung cancer.