Hsa-miR-16 Inhibited The Proliferation Of Human Gastric Cancer MGC-803 Cell Line And Regulated The Expression Of WWP1

Objective: to investigate whether hsa-mir-16 can target the binding of WWP1 mRNA and regulate its protein expression in gastric cancer cells,as well as its effect on the proliferation of gastric cancer cells.Methods:1.The bioinformatics method was used to predict the target genes of mir-16,focusing on whether the target genes could be bound to WWP1 mRNA 3 'UTR,and the GO analysis and KEGG analysis were performed on mir-16.2.Use lip2000 miR-16 mimic,miR-16 inhibit transfection into the MGC-803 gastric cancer cells and extract its total RNA,real-time fluorescent quantitative PCR,verify the transfection,and verify WWP1 mRNA expression level with or without changes.3.Western blot was used to verify whether there were differences in the expression of WWP1 protein after overexpression or inhibition of mir-16 at the protein level.4.The expression vectors of the dual luciferase reporting systemwere constructed and transfected into 293 T cells to quantitatively evaluate the targeting relationship between mir-16 and WWP1 through the relative luciferase activity.5.CCK8 assay explored the effects of mir-16 and silencing target gene WWP1 on proliferation of gastric cancer cells mgc-803 at the cellular level.Results:1.Multiple databases predicted the existence of targeted binding sites between mir-16 and WWP1 mRNA 3' utr,and GO analysis and KEGG analysis confirmed that mir-16 was involved in regulating tumor development.2.Real-time fluorescent quantitative PCR results showed that after transfection with a mir-16 mimic in human poorly differentiated gastric cancer MGC803 cells,the expression of mir-16 was increased compared with that of the control group(p<0.01),while after transfection with mir-16 inhibitor,the expression of mir-16 was decreased compared with that of the control group(p<0.001),suggesting that the transfection was effective.However,compared with the control group,there was no significant change in WWP1 mRNA levels in the mir-16 mimic group and the mir-16 inhibitor group.3.Western blot experiments showed that WWP1 protein expression was decreased in the mir-16 mimic group compared with the controlgroup(p<0.01),while WWP1 protein expression was increased in the mir-16 inhibitor group(p<0.05).The differences were statistically significant.4.The results of the dual luciferase reporting system showed that after the wild-type WWP1 reporter plasmid was co-transfected with mir-16 or mir-nc,the relative fluorescence value in the mir-16 group was0.45 ± 0.032,significantly lower than that in the mir-nc transfection group,and the difference was statistically significant(p<0.0001),indicating that mir-16 could target and bind to WWP1 mRNA 3 'UTR.5.CCK8 results showed that transfection with mir-16 inhibitor could enhance the proliferation of mgc-803 in human poorly differentiated gastric cancer cells,while co-transfection with mir-16 inhibitor and siRNA of WWP1 could eliminate this proliferation.Conclusions: combined with the above results,mir-16 can target and bind WWP1 mRNA 3'utr to regulate the expression of WWP1 protein in mgc-803 cells of human gastric cancer.After inhibiting the level of mir-16 in human gastric cancer mgc-803 cells,the proliferation capacity of human gastric cancer mgc-803 cells was enhanced.