Knock-out SST Gene Enhancing Invasion And Migration Of Gastric Cancer Cell BGC823 And The Expression Of Potential Gastric Cancer Driver Gene SEM5A And KLF2

Background and Objective Somatostatin is a peptide hormone that is secreted by D cells of the gastrointestinal mucosa and is widely present in the gastrointestinal mucosa,especially in the antrum and stomach.Studies have shown that somatostatin has a certain inhibitory effect on the occurrence and development of gastric cancer,some studies have also found that different concentrations of SST can stimulate the growth of gastric cancer cells.Therefore,the relationship between SST and the occurrence and development of gastric cancer as well as its regulation mechanisms have not yet been clarified.SST,as a polypeptide hormone widely distributed on the gastric mucosa,can inhibit the secretion of gastrin and induce the occurrence of chronic atrophic gastritis.Chronic atrophic gastritis is a precancerous disease and pathological basis of gastric cancer.SEM5 A and KLF2,as potential driver genes of gastric cancer,may play important roles in the process of regulation mechanism of SST in gastric cancer.In the past,the study of SST in gastric cancer was limited to gene interference or gene silencing,while it did not knock out the SST gene actually.It has not reported whether SST plays a role in gastric cancer through driver genes.Therefore,we used CRISPR/Cas9 gene editing technology to knockout SST genes in gastric cancer cells to investigate the role of SST in gastric carcinogenesis.To detect the influnce in the proliferation,migration and invasion of gastric cancer cells as well as the expression of gastric driver genes,SEM5 A and KLF2,exploring whether SST exerts inhibitory effects through the regulation of gastric cancer-driver genes SEM5 A and KLF2.Methods 1.Two pairs of sg RNA as well as reporter were designed according to targeting sequence of SST gene for double-nicking.Vectors px330-SST-sg RNA and pmcherry EGFP-SST-reporter were constructed.Transfecting into HEK293 T for selecting sg RNA with higher cutting efficiency.2.After transfecting into BGC823,sorting the single selected cell by FACS.Getting the target cell strain which has knocked-out SST gene and verified by sequencing.The expression of SST was detected by immunocytochemistry and the cellular morphology was observed.3.Transwell was applied for testing migration and invasion ability,as well as MTT for proliferation ability.4.The protein expression of SEM5 A and KLF2 were observed by Western blotting and LSCM.Results 1.The plasmids px330-SST-sg RNA and pmcherry EGFP-SST-reporter was constructed successfully and the cell strain,1E9,which had knocked out SST gene,was established.2.The migration and invasion ability of 1E9 has increased significantly.The morphology of 1E9 turns less smooth and appears more antennae.3.The expression of SEM5 A and KLF2 turns higher in 1E9.Conclusions SST plays an inhibitory role in the proliferation,migration and invasion of gastric cancer cell BGC823.SST may inhibit the development of gastric cancer cells by regulating the gastric cancer-driver genes SEM5 A and KLF2.