Enhancing Radiosensitivity Of A549 Cell Lines By Transfection Of Bcl-2 SiRNA

Objective:To investigate the effect on apoptosis of A549 cell by transfection of Bcl-2 SiRNA,and to explore the relationship between Bcl-2 SiRNA and radio -sensitivity of A549 cell.We combine the SiRNA and Radiotherpy,and the aim is to seek a new method to enhance radiosensitivity of tumor cell.Methods:1.Vectors containing SiRNA targeting Bcl-2 gene were constructed and trans -fected into A549.The Real time RT-PCR and Western blot were used to compare the efficiency of gene silencing.2.Flow cytometry was used to determine the effects of pBcl-2-siRNA,and we have compared the induced-apoptotic effect between A549 cell transfected with pBcl-2-siRNA or A549 cell transfected with pNeg-siRNA or A549 cell transfected with pRNA-U6.3/Hygro and Normal A549 cells.Later,we compare the induced -apoptotic effect of the four cell lines with 6Gy irradiation.3.The radiosensitivity and survival fraction of A549 cell transfected with pBcl-2 -siRNA and A549 cell transfected with pNeg-siRNA and A549 cell transfected with pRNA-U6.3/Hygro and Normal A549 cells were investigated by cellular clone formation experimenResults:1.Vectors containing SiRNA targeting Bcl-2 gene were constructed succeedly and transfected into A549 cell.Three stable transfected cell lines A549/Bcl2-Ins,A549/ Bcl2-Neg and A549/SD were screened from the transfected A549 cells.Western blot analysis and Real time RT-PCR indicated that the expression of Bcl-2 was suppressed by the pBcl-2-siRNA.2.Flow cytometry was used,The apoptosis rate of A549/Bcl2-Ins was higher than that or controls.The apoptosis rate of A549/Bcl2-Ins at 6Gy irradiation was higher than that of controls and unirradiated groups.It indicated that the pBcl-2-siRNA had good effect on the apoptosis rate and the pBcl-2-siRNA got better effect with irra -diation..3.The radiosensitivity of A549/Bcl2-Ins was evaluated by Clone formation array. The D₀ and D_q of A549/Bcl2-Ins were 1.399 and 1.298,respectively,lower than those of A549/Bcl2-Neg(1.695 and 2.480) or those of A549/SD(1.706 and 2.227) or those of A549(1.816 and 2.777).The radiosensitivity of A549 cell was enhanced by the transfection. Conclusions:1.Vectors containing SiRNA targeting Bcl-2 gene were constructed succeedly, and the expression of Bcl-2 was suppressed.2.The apoptosis rate of A549 cell was enhanced strongly by the pBcl-2-siRNA. And The apoptosis rate will get higher with irradiation.3.The radiosensitivity of A549 cell can be enhanced by the transfection.