Effects Of Rad51 Gene Silencing On Cell Survival Of A549 Cells In Vivo And In Vitro

Objective: To silence gene Rad51 in A549 cells by Rad51 si RNA, and to observe variation of the colony-forming ability, the cell viability, the cell cycle progression and the early growth status of transplanted tumor in nude mouse after transfected by Rad51 si RNA. Then investigate the role of Rad51 in the survival of lung cancer with the aim to provide experimental data and theory data for targeted therapy of NSCLC.Methods: A549 cells were cultured as normal, Rad51 si RNA was designed and constructed to Lentivirus. The experiment was divided into three groups:①Normal group:the human A549 cells were cultured with DMEM medium supplemented with 10% Fetal Bovine Serum; ②Lenti-NC group:the A549 cells were infected with lentivirus of empty vector;③Lenti-Rad51 si group:infected with lentivirus of Rad51 si RNA; Real-time quantitative PCR analysis and Western blot were used to check the m RNA and protein level of Rad51 in A549 cells; cell proliferation ability was decided by Colony-forming assay; cell cycle were analyzed by fluorescence-activated cell sorter; the early growth status of transplanted tumor in nude mouse were observed.Result: 1. Lenti-Rad51 si and Lenti-NC virus were manufactured by Shanghai Genechem Company. The virus Lenti-Rad51 titer was 2×108 TU/ml and Lenti-NC titer was 8×108 TU/ml packaging; 2. A high transfection efficiency could be obtained after infection of Lenti-Rad51 si RNA and Lenti-NC in A549 cells; 3. After 48 h of the Lenti-Rad51 si infection, the m RNA and protein expression of Rad51 were downregulated detected by Quantitative RT-PCR and Western Blot; 4. Silencing gene Rad51 resulted in a reduction of the colony formation of A549 cells;5. The reusult of MTT assay indicated that infection of Lenti-Rad51 si can significantly inhibit the proliferation of A549 cells; 6. Cell cycle analysis revealed: the proportion of G1 phase increase obviously after the infection of Lenti-Rad51 si in A549 cells; 7. The results of nude mice transplant tumor show: compared with Normal group and Lenti-NC group, the growth ability of tumor cells was significantly reduced in Lenti-Rad51 si group.Conclusions: In vitro studies revealed: after inhibition of Rad51 by Rad51 si RNA, the colony formation ability and the proliferation of A549 cells were reduced obviously, numerous cells were arrested in G1 phase; in vivo, the growth ability of tumor cells was significantly reduced in Lenti-Rad51 si group.