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Abstract

Background: Tumor-associated macrophages (TAMs) are the predominant immune cells in the tumor microenvironment and portend poor prognosis. However, the molecular mechanisms underlying the tumor promotion of TAMs have not been fully elucidated.

Methods: Coculture of gastric cancer cells with U937 cells was performed to investigate the impact of TAMs on cancer cell behavior. MicroRNA (miRNA) microarray and bioinformatics were applied to identify the involved miRNAs and the functional target genes. The regulation of the miRNA on its target gene was studied using anti-miRNA and miRNA mimic.

Results: Coculture with CD204⁺ M2-like TAMs increased proliferation, migration, and epithelial-mesenchymal transition of gastric cancer cells. MiR-210 was the most upregulated miRNA in cancer cells identified by miRNA microarray after coculture. In gastric cancer tissues, miR-210 expression was positively correlated with CD204⁺ M2-like TAM infiltration. Inactivation of miR-210 by antimir attenuated CD204⁺ M2-like TAMs-induced cancer cell migration. Using pharmacological inhibitors and neutralizing antibodies, CD204⁺ M2-like TAMs-secreted TNFα was found to upregulate miR-210 through NF-κB/HIF-1α signaling. Bioinformatics analysis showed netrin-4 (NTN4) as a potential target of miR-210 to suppress gastric cancer cell migration. We also found an inverse expression between miR-210 and NTN4 in cancer cells after coculture or in tumor xenografts. Anti-miR-210 increased NTN4 expression, while miR-210 mimics downregulated NTN4 in cancer cells. Reporter luciferase assays showed that MiR-210 mimics suppressed NTN4 3' untranslated region-driven luciferase activity in cancer cells, but this effect was blocked after mutating miR-210 binding site.

Conclusions: CD204⁺ M2-like TAMs can utilize the TNF-α/NF-κB/HIF-1α/miR-210/NTN4 pathway to facilitate gastric cancer progression.

Keywords: Gastric cancer; Migration; NTN4; Tumor microenvironment; Tumor-associated macrophage; miR-210.