Helicobacter pylori-induced miR-136 is a potential predictor of early-stage gastric cancer

Figure 1 miR-136 upregulation in Helicobacter pylori-positive human precursor lesions of gastric cancer and gastric cancer tissues. A-D: Overexpressed miRNA-136 and under expressed PDCD11 in Helicobacter pylori (H. pylori)-positive gastric cancer (GC) tissues compared with H. pylori-negative GC tissues; E and F: The expression of miR-136 was negatively related to the expression of PDCD11 in GC tissues (r1 = -0.782; r2 = -0.812); G-K: MiR-136 expression was upregulated and PDCD11 expression was downregulated significantly in precursor lesions of GC compared with non-active gastritis, chronic gastritis, and GC compared with non-active gastritis, chronic gastritis, precursor lesions of GC. ^aP < 0.01. HP+: With Helicobacter pylori infection; HP-: Without Helicobacter pylori; NAG: Non-active gastritis without Helicobacter pylori infection; CAG: Chronic gastritis with Helicobacter pylori-infection; PLGC: Precursor lesions of gastric cancer with Helicobacter pylori-infection; GC: Gastric cancer.

Figure 2 miR-136 upregulation in Helicobacter pylori-positive gastric cancer cells in vitro. A-E: Overexpressed miRNA-136 and underexpressed PDCD11 in SGC-7901 and BGC-823 gastric cancer cells with Helicobacter pylori (H. pylori) infection compared with cells without H. pylori infection; F and G: Overexpressed miRNA-136 and underexpressed PDCD11 in H. pylori-positive SGC-7901 and BGC-823 gastric cancer cells, respectively, compared with H. pylori-positive GES-1. ^aP < 0.01. HP+: With Helicobacter pylori-infection, HP-: Without Helicobacter pylori-infection.

Figure 3 Nuclear factor kappa-B regulated miR-136 expression after Helicobacter pylori infection. A and B: Helicobacter pylori induced a significant increase in the relative luciferase activity of wild-type reporter vector; C-F: Nuclear factor kappa-B inhibitor could reduce the upregulated expression of miRNA-136 induced by Helicobacter pylori. ^aP < 0.01; NS: Not significant; Wt: Wild-type reporter vector; Mut: Mutant reporter vector; Bay: Nuclear factor kappa-B inhibitor; HP: Helicobacter pylori; HP+: With Helicobacter pylori infection; HP-: Without Helicobacter pylori.

Figure 4 miR-136 promoted the proliferation and migration of Helicobacter pylori-positive gastric cancer cells in vitro. A and B: The expression of miRNA-136 was increased in SGC-7901 and BGC-823 gastric cancer cells transfected with miR-136 mimic, and the expression of miRNA-136 was decreased in SGC7901 and BGC-823 gastric cancer cells transfected with miR-136 inhibitor; C-I: MiR-136 mimic significantly promoted the proliferation and migration of SGC-7901 and BGC-823 gastric cancer cells. Conversely, the miR-136 inhibitor had the opposite effect on SGC-7901 and BGC-823 gastric cancer cells via cell counting kit-8 assay, clone formation, scratch repair assays, and Transwell assay. ^aP < 0.01.

Figure 5 miR-136 promoted tumorigenesis of Helicobacter pylori-positive gastric cancer cells by targeting PDCD11 in vitro. A and B: MiR-136 mimic significantly attenuated luciferase activity of the wild-type reporter vector. Rather, miR-136 mimic had no effect on the Mut reporter vector via the luciferase assay; C and D: MiR-136 inhibitor significantly increased the luciferase activity of the wild-type reporter vector. Rather, miR-136 inhibitor had no effect on the Mut reporter vector via the luciferase assay; E-I: The expression of PDCD11 mRNA and protein was reduced in SGC-7901 and BGC-823 cells transfected with miR-136 mimic via reverse transcription quantitative PCR and western blot assay. When SGC-7901 and BGC-823 cells were transfected with miR-136 inhibitor, the opposite occurred; J: PDCD11 plasmid cannot be inhibited by miR-136; K-P: The proliferation and migration of gastric cancer cells induced by miR-136 were abrogated by transfecting PDCD11 plasmid. ^aP < 0.01. wt: Wild-type reporter vector; mut: Mutant reporter vector.

Figure 6 miR-136 promoted tumorigenesis of Helicobacter pylori-positive gastric cancer cells by targeting PDCD11 in vivo. A: The promoted tumor growth in miR-136 mimic group and the reduced tumor growth in miR-136 inhibitor group compared with that in the control group; B: The tumor growth induced by miR-136 were abrogated by PDCD11. ^aP < 0.01.