Yonsei Med J.  2019 Jan;60(1):38-47. 10.3349/ymj.2019.60.1.38.

Up-Regulation of MiR-1915 Inhibits Proliferation, Invasion, and Migration of Helicobacter pylori-Infected Gastric Cancer Cells via Targeting RAGE

• Xu XC
• Zhang WB
• Li CX
• Gao H
• Pei Q
• Cao BW
• He TH

• KMID: 2428276
• DOI: http://doi.org/10.3349/ymj.2019.60.1.38

Abstract

PURPOSE
Helicobacter pylori (HP)-infected gastric cancer (GC) is known to be a fatal malignant tumor, but the molecular mechanisms underlying its proliferation, invasion, and migration remain far from being completely understood. Our aim in this study was to explore miR-1915 expression and its molecular mechanisms in regulating proliferation, invasion, and migration of HP-infected GC cells.
MATERIALS AND METHODS
Quantitative real-time PCR and western blot analysis were performed to determine miR-1915 and receptor for advanced glycation end product (RAGE) expression in HP-infected GC tissues and gastritis tissues, as well as human gastric mucosal cell line GES-1 and human GC cell lines SGC-7901 and MKN45. CCK8 assay and transwell assay were performed to detect the proliferation, invasion, and migration capabilities. MiR-1915 mimics and miR-1915 inhibitor were transfected into GC cells to determine the target relationship between miR-1915 and RAGE.
RESULTS
MiR-1915 was under-expressed, while RAGE was over-expressed in HP-infected GC tissues and GC cells. Over-expressed miR-1915 could attenuate cellular proliferation, invasion, and migration capacities. RAGE was confirmed to be the target gene of miR-1915 by bioinformatics analysis and luciferase reporter assay. Moreover, HP-infected GC cellular proliferation, invasion, and migration were inhibited after treatment with pcDNA-RAGE.
CONCLUSION
MiR-1915 exerted tumor-suppressive effects on cellular proliferation, invasion, and migration of HP-infected GC cells via targeting RAGE, which provided an innovative target candidate for treatment of HP-infected GC.

Keyword

Gastric cancer; microRNA-1915; MKN45; receptor for advanced glycation end product; SGC-7901

MeSH Terms

Blotting, Western
Cell Line
Cell Proliferation
Computational Biology
Gastritis
Helicobacter pylori
Helicobacter*
Humans
Luciferases
Rage*
Real-Time Polymerase Chain Reaction
Stomach Neoplasms*
Up-Regulation*
Luciferases

Figure

• 

  Fig. 1 Under-expressed miR-1915 and over-expressed RAGE in HP (+) GC tissues. (A) Relative miR-1915 level in gastritis or GC tissues with or without HP infection by qRT-PCR. (B) Expression of RAGE in gastritis or GC tissues with or without HP infection by western blotting. *p<0.05 vs. HP (−) Gastritis, †p<0.05 vs. HP (−) GC, ‡p<0.05 vs. HP (+) Gastritis. RAGE, receptor for advanced glycation end product; HP, Helicobacter pylori; GC, gastric cancer.

• 

  Fig. 2 Under-expressed miR-1915 and over-expressed RAGE in HP (+) GC cells. (A) HP infection was performed in human SGC-7901, MKN45 GC cell lines, and GES-1 human gastric epithelial cell line. Relative miR-1915 level was detected in each cell line with or without HP infection by qRT-PCR. (B) Expression of RAGE was detected in each cell line with or without HP infection by western blotting. *p<0.05 vs. HP (−) GES-1 or HP (−) 7901 or HP (−) MKN45 or HP (+) GES-1. RAGE, receptor for advanced glycation end product; HP, Helicobacter pylori.

• 

  Fig. 3 Over-expressed miR-1915 inhibits proliferation, invasion, and migration of HP (+) GC cells. (A) HP (+) 7901 and MKN45 cells were transfected by miR-1915 mimic. Relative miR-1915 level was detected by qRT-PCR. (B) Cell proliferation of HP (+) 7901 and MKN45 cells in 0, 24, 48, and 72h was detected by CCK8 assay. (C) Invasive/migratory capability of HP (+) 7901 and MKN45 cells was detected by transwell assay. *p<0.05 vs. pre-NC. GC, gastric cancer; HP, Helicobacter pylori.

• 

  Fig. 4 Target relationship between miR-1915 and RAGE. (A) Complementary sequence information of miR-1915 and RAGE. (B) 293T cells were transfected by miR-1915 inhibitor. Relative luciferase activity of RAGE 3′UTR (WT or Mut) was detected by luciferase reporter assay. Expression of RAGE was detected by qRT-PCR and western blotting. (C) 293T cells were transfected by miR-1915 mimic. Relative luciferase activity of RAGE 3′UTR (WT or Mut) was detected by luciferase reporter assay. Expression of RAGE was detected by qRT-PCR and western blotting. *p<0.05 vs. NC, †p<0.05 vs. pre-NC. RAGE, receptor for advanced glycation end product; HP, Helicobacter pylori.

• 

  Fig. 5 Proliferation, invasion, and migration of HP (+) GC cells induced by miR-1915 and RAGE. (A) HP (+) 7901 and MKN45 cells were transfected by miR-1915 mimic, miR-1915 mimic+pcDNA, or miR-1915 mimic+pcDNA-RAGE. Relative miR-1915 level was detected by qRT-PCR. (B) Expression of RAGE was detected by qRT-PCR and western blotting. (C) Cell proliferation in 0, 24, 48, and 72 h was detected by CCK-8 assay. (D) Cell invasive capability was detected by transwell assay. (E) Cell migratory capability was detected by transwell assay. *p<0.05 vs. pre-NC, †p<0.05 vs. pcDNA. RAGE, receptor for advanced glycation end product; HP, Helicobacter pylori; GC, gastric cancer.

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