Résumé
Objective To study the inhibition effects of various gastrin-shNAs on gastrin expression in gastric cancer cell line BGC-823. Methods Four nucleotide sequences of shRNA were designed corresponding to various sites of gastrin gene.Four shRNAs were synthesized by in vitro transcription and transfected into gastric cancer cell line BGC-823 at the final concentration of 10nmol/L,20nmol/L,40nmol/L and 80nmol/L respectively.In situ hybridization and immunohistochemistry techniques were applied to investigate the inhibition of gastrin expression and screen the most effective shRNA.The inhibitory effect on gastrin mRNA of screened shRNA was further identified by RT-PCR.MTT assay was used to determine the inhibitory effect of 4 shRNAs at various final concentrations on the growth of BGC-823 cells. Results The gastrin mRNA and protein exression were suppressed distinctly 24,48,and 72hours after transfection,and exhibited time-and concentration-dependent tendency.The highest suppression efficiency on both mRNA(54.27?0.042)% and protein(41.69?0.038)% level occurred 72 hours later in the cells transfected with shRNAs.The RT-PCR result showed that the inhibitory ratio of shRNA3 on gastrin mRNA of BGC-823 was 48.1%.MTT displayed a proliferative inhibition of the BGC-823 cells after transfection of shRNAs with a concentration-denpendent tendency except the shRNA4 treated cells.Conclusion Four gastrin-shRNAs showed a significant inhibition effect on gastrin expression of gastric cancer cell BGC-823 on mRNA and protein level.shRNAs might be the most effective gastrin-shRNA.Inhibited gastrin expression by shRNAs resulted in a significant decrease of proliferative ability of BGC-823 cells.