MiR-204-3p overexpression inhibits gastric carcinoma cell proliferation by inhibiting the MAPK pathway and RIP1/MLK1 necroptosis pathway to promote apoptosis

Figure 1 Expression of miR-204-3p in gastric carcinoma. A: The expression level of miR-204-3p in gastric carcinoma tissues and para-carcinoma tissues was analyzed by the GEO database; B: The expression level of miR-204-3p in normal gastric mucosal epithelial cells and gastric carcinoma cells, ^cP < 0.001.

Figure 2 Effect of miR-204-3p on AGS and HGC-27 cell viability. A: Transfection efficiency of lentiviral vector. Scale bar: 1000 μm, with representative images shown, fluorescence distribution of green fluorescent protein in cells indicating transfection efficiency; B and C: The expression level of miR-204-3p in lentivirus transfected AGS and HGC-27 cells; D and E: Effect of miR-204-3p on AGS and HGC-27 cell viability. ^aP < 0.05, ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 3 Effect of miR-204-3p on colony forming ability in AGS and HGC-27 cells. A and B: Crystal violet staining diagram of AGS and HGC-27 cells; C and D: Statistical results of colony forming ability in AGS and HGC-27 cells. ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 4 Effects of miR-204-3p on apoptosis in AGS and HGC-27 cells. A and B: Flow cytometry was used to analyze apoptosis of AGS and HGC-27 cells; C and D: Statistical results of AGS and HGC-27 cell apoptosis. ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 5 Effects of miR-204-3p on the cell cycle in AGS and HGC-27 cells. A and B: Cell cycle analysis in AGS and HGC-27 cells by flow cytometry; C and D: Percentage of cell cycle distribution in AGS and HGC-27 cells. ^bP < 0.01, ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 6 Effect of miR-204-3p on tumor size in BABL/c nude mice. A: Comparison of gastric carcinoma tumor xenograft models in BABL/c nude mice; B: Comparison of subcutaneous tumors on the back in BABL/c nude mice; C: Statistical graph of tumor growth curve for gastric carcinoma tumor xenograft models in BABL/c nude mice; D: Statistical graph of body weight growth curve in BABL/c nude mice. ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 7 Effect of miR-204-3p on the MAPK signaling pathway in AGS and HGC-27 cells. A and B: Effects of miR-204-3p on mRNA levels of KRAS, ERK1 and ERK2 in AGS and HGC-27 cells; C and D: Effects of miR-204-3p on protein levels of KRAS, ERK1/2 and p-ERK1/2 in AGS cells; E and F: Effects of miR-204-3p on protein levels of KRAS, ERK1/2 and p-ERK1/2 in HGC-27 cells. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 8 Effects of miR-204-3p on apoptosis-related proteins and necroptosis-related proteins in AGS and HGC-27 cells. A and B: Effect of miR-204-3p on apoptosis-related proteins BCL-2, BAX and Caspase-3 in AGS cells; C and D: Effect of miR-204-3p on apoptosis-related proteins BCL-2, BAX and Caspase-3 in HGC-27 cells; E and F: Effects of miR-204-3p on necroptosis-related proteins RIP1, p-RIP1, MLK1, and p-MLK1 in AGS cells; G and H: Effects of miR-204-3p on necroptosis-related proteins RIP1, p-RIP1, MLK1, and p-MLK1 in HGC-27 cells. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. NC: Empty lentiviral vector; KD: miR-204-3p knockdown lentiviral vector; OE: miR-204-3p overexpression lentiviral vector.

Figure 9 Mechanism of the effect of miR-204-3p on the MAPK signaling pathway, apoptosis and necroptosis. TNF: Tumor necrosis factor; GF: Germ-free.