M2-polarized macrophages promote metastatic behavior of Lewis lung carcinoma cells by inducing vascular endothelial growth factor-C expression

OBJECTIVES: Tumor-associated macrophages that generally exhibit an alternatively activated (M2) phenotype have been linked to tumor progression and metastasis. However, the role of M2-polarized macrophages in the growth and metastasis of lung adenocarcinoma remains enigmatic. The aim of this study was to explore the effect of M2 macrophages on the proliferation and migration of mouse Lewis lung carcinoma cells and tumor-induced lymphangiogenesis. METHODS: Trypan blue staining and the Transwell migration assay were performed to evaluate the effects of activated (M1 or M2) macrophages on the proliferation and migration of Lewis cells. Furthermore, vascular endothelial growth factor-C expression in Lewis cells and nitric oxide secretion from activated macrophages were detected during the co-culture assay. Following treatment with activated macrophages, lymphatic endothelial cells differentiated into capillary-like structures, and the induction of Lewis cell migration was assessed using a twodimensional Matrigel-based assay. RESULTS: In the co-culture Transwell system, the proliferation and migration of Lewis cells were promoted by M2 macrophages. Moreover, the co-culture significantly increased the expression of vascular endothelial growth factor-C by Lewis cells and reduced the secretion of nitric oxide from M2 macrophages, which subsequently led to the capillary morphogenesis of lymphatic endothelial cells. Interestingly, following co-culture with Lewis cells, the function of RAW264.7 cells was polarized toward that of the M2 macrophage phenotype. CONCLUSION: M2-polarized macrophages promoted the metastatic behavior of Lewis cells by inducing vascular endothelial growth factor-C expression. Thus, the interruption of signaling between M2 macrophages and Lewis cells may be considered to be a new therapeutic strategy.

M2-Polarized tumor-associated macrophages are associated with poor prognoses resulting from accelerated lymphangiogenesis in lung adenocarcinoma

OBJECTIVES: Tumor-associated macrophages have been implicated in promoting tumor growth, progression and metastasis. However, the activated phenotype (M1 or M2) of tumor-associated macrophages remains unknown in solid tumors. Therefore, this study examined the density and prognostic significance of M2-polarized tumor-associated macrophages in lung adenocarcinoma. METHODS: Tumor specimens from 65 lung adenocarcinoma patients were assessed by ELISA for Th1/Th2 cytokine concentrations. The activated phenotype (M1 or M2) of tumor-associated macrophages was determined utilizing immunofluorescence staining. Additionally, to evaluate lymphangiogenesis, peritumoral lymphatic microvessel density was measured using D2-40. The correlation between tumor-associated macrophage subtype and overall patient survival was analyzed using the Kaplan-Meier method and compared using the log-rank test. RESULTS: A shift toward Th2 cytokine expression was detected within lung adenocarcinoma microenvironments. Approximately 79.71±16.27 percent of tumor-associated macrophages were M2 polarized; the remaining 20.35±5.31 percent were M1 polarized. The infiltration of M2-polarized macrophages was significantly associated with P-TNM staging and lymph node metastasis. The peritumoral lymphatic microvessel density was significantly higher in the high M2-polarized tumor-associated macrophage group than in the low M2-polarized tumor-associated macrophage group. A significant difference in overall patient survival was detected not only between patients with tumors with high and low macrophage counts but also between patients with tumors with high and low counts of M2-polarized macrophages. CONCLUSION: Tumor-associated macrophages in lung adenocarcinoma have an M2-polarized subtype and are associated with poor prognoses, perhaps resulting from accelerated lymphangiogenesis and lymph node metastasis.