RÉSUMÉ
Objective:To study the expression of miR-496 in gastric cancer cells, and explore its role and mechanism in the proliferation and apoptosis of gastric cancer cells. Methods:Real-time fluorescence quantitative PCR (qPCR) was used to detect the expression of miR-496 in normal gastric epithelial cell lines and gastric cancer cell lines AGS and MKN45. miR-496 was knocked down in AGS cells with the lowest expression level, and a negative control group and a blank control group were set up. Cell proliferation and apoptosis were detected by CCK8 assay and flow cytometry. LYN, the target gene of miR-496, was screened using bioinformatics software, and the effect of transfection of miR-496 on LYN expression was detected by qPCR. Subsequently, rescure experiment was conducted to further study the mechanism of miR-496 on gastric cancer cells through regulation of LYN. Data were analyzed by GraphPad Prism 9 software. Measurement data were presented as mean ± standard deviation, and the comparison between the two groups was performed by t test. Results:The expression of miR-496 in AGS and MKN45 was significantly lower than that in normal gastric epithelial cells ( P<0.05). After overexpression of miR-496, the proliferation of AGS cells could be inhibited and the apoptosis ratio of AGS cells could be significantly increased ( P<0.05). QPCR results showed that miR-496 overexpression group could inhibit the expression of LYN ( P<0.05). Bioinformatics analysis showed that miR-496 binds to LYN kinase ( LYN) 3 ′UTR region, and overexpression of miR-496 can inhibit the expression of LYN in AGS cells, while CCK8 rescue experiment showed that overexpression of LYN could remove the inhibitory effect of miR-496 on cell proliferation. Flow cytometry showed that LYN expression could cancel the promoting effect of miR-496 on apoptosis ( P<0.05). Conclusion:miR-496 is low expressed in gastric cancer cells, and it inhibits the proliferation and promotes apoptosis of gastric cancer cells by targeting the expression of LYN in gastric cancer cells.
RÉSUMÉ
Objective:To study the effect and mechanism of Rab4A knockout expression on proliferation, migration and invasion of gastric cancer cells. Methods:The expression of Rab4A in four human gastric cancer cell lines MGC-803, SGC-790, MKN45 and AGS was detected by Western blot. Rab4A was knocked out in AGS cells with the highest expression level, and untransfected gastric cancer cells were used as control group. Cell proliferation, migration and invasion were detected by CCK8 and Transwell assay, respectively. Western blot analysis was used to investigate the expression changes of epidermal growth factor receptor (EGFR), downstream pathway proteins AKT and β-catenin induced by Rab4A knockout. The interaction between Rab4A and MiR- 496 was detected by dual luciferase reporter gene, and the effect of MiR- 496 transfection on Rab4A expression was detected by qPCR and Western blot. GraphPad Prism 9 software was used for data analysis, t-test was used for comparison between the two groups, and normal distribution measurement data were expressed as mean ± standard deviation ( ± s). Results:The expression of Rab4A was the highest in AGS cells, and the knockdown of Rab4A inhibited the proliferation, migration and invasion of AGS cells ( P<0.05). In Rab4A knockout gastric cancer cells, the surface expression of epidermal growth factor receptor (EGFR) was significantly decreased, and the expression of downstream pathway proteins p-AKT and β-catenin was also inhibited ( P<0.05). The luciferase reporter showed that MiR- 496 could bind the 3′UTR of Rab4A. In addition, MiR- 496 down-regulated the expression of Rab4A in AGS cells( P<0.05). Conclusion:The expression of Rab4A is inhibited by MiR- 496, and the proliferation, migration and invasion of gastric cancer cells can be inhibited by down-regulating the surface expression of EGFR after inhibiting Rab4A expression.