Inhibition of gastric cancer BGC-823 cell proliferation by small interfering RNA specific to survivin gene in vitro

Objective To study the of proliferation and apoptosis of gastric cancer BGC-823 cells by using RNA interference technique to silence survivin gene in vitro and explore whether surviving siRNAs could become a promising tool in the treatment of gastric cancer.Method According to siRNA design principle,we designed three siRNAs targeting survivin gene,and constructed three expression plasmids (shRNA-survivin-1,shRNA-survivin-2 and shRNA-survivin-3) to evaluate their abilities in inhibition of survivin gene expression. SiRNAs expression plasmids (shRNAs) were transfected into BGC-823 cells via LipofectamineTM2000. The cells transfected with nonsense shRNA (shRNA-negative) served as a negative control group and those only treated with LipofectamineTM2000 as a blank control group. To detect their inhibition effect in BGC-823 cell proliferation,in vitro we applied real-time PCR,western blot,MTT assay,and flow cytometry assay.Result Among the three designed and constructed shRNAs targeting survivin gene,shRNA-survivin-1 showed a significant and effective inhibition of surviving gene expression. Real-time PCR results showed shRNA-survivin-1 could effectively decrease survivin mRNA levels (surviving/GAPDH to 1.05),and comparing with blank control group (surviving/GAPDH to 15.73),the difference was statistically significant(P0.05). Western blot showed comparing with blank control group and shRNA-negative group,that shRNA-survivin-1 significantly inhibited survivin protein expression; MTT assay results manifested 48 h growth inhibition rate in BGC-823 cells treated by shRNA-survivin-1 was 59.19%,and comparing with liposome group(32.15%)and shRNA-negative group (32.08%),the difference was statistically significant(P0.05). 72 h growth inhibition rate of shRNA-survivin-1 group (74.92%) was higher than those of liposome group and shRNA-negative group (P0.05). Flow cytometry assay showed cells treated by shRNA-survivin-1 had a significant increase in G2 /M phase,apoptosis rate significantly increased (P0.05).Conclusion shRNA-survivin-1 can effectively trigger a post-transcriptional survivin gene silencing,significantly suppress proliferation of BGC-823 cells and induce apoptosis to a certain degree. The findings about the designed effective siRNAs targeting survivin gene in our study provide a rational support for the development of siRNAs as a therapy for stomach cancer in humans.