Tumor-associated macrophages induce capillary morphogenesis of lymphatic endothelial cells derived from human gastric cancer

Abstract

Tumor lymphangiogenesis is a major prognostic indicator of gastric cancer. Tumor-induced inflammation has been shown to attract tumor-associated macrophages (TAMs) that affect lymphangiogenesis. However, detailed mechanisms of macrophage-induced lymphangiogenesis have not been elucidated. Here, we evaluated the interaction between TAMs and lymphatic endothelial cells (LECs) derived from lymph nodes (LNs) of human gastric cancer. LECs were directly or indirectly co-cultured with macrophages from healthy human blood, with or without the supernatant of the gastric cancer cell line, OCUM-12. We analyzed the effect of cancer pretreated macrophages and of macrophages from metastatic LNs of gastric cancer on LECs. We observed morphological changes of LECs in co-culture and assessed the gene expression of possible lymphangiogenic molecules of macrophages and LECs after contact co-culture, and of cancer pretreated macrophages, by qRT-PCR. Specimens of metastatic LN of gastric cancer were immunofluorescently stained. We found that tubulogenesis of LECs was observed only in the contact co-culture model. OCUM-12 cells promoted macrophage-induced tubulogenesis of LECs. Relative gene expression of matrix metalloproteinases (MMP) and adhesion molecules was significantly up-regulated in both capillary-forming LECs and co-cultured macrophages. Cancer pretreated macrophages up-regulated lymphangiogenic factors including inflammatory cytokines, MMPs, adhesion molecules and VEGF-C. Blocking of ICAM-1 and macrophage activation suppressed tubulogenesis of LECs. Immunohistochemistry showed macrophages localized around lymphatic vessels. Our results suggested that interaction between LECs and macrophages may be an important initial step of tumor lymphangiogenesis developing LN metastasis. Understanding of its mechanisms could be useful for future therapeutics of gastric cancer.

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