High‐fat diet‐induced obesity increases lymphangiogenesis and lymph node metastasis in the B16F10 melanoma allograft model: Roles of adipocytes and M2‐macrophages

To examine the effects of high-fat diet (HFD) on melanoma progression, HFD-fed C57BL/6N mice were subcutaneously injected with syngeneic B16F10 melanoma cells. At 3 weeks post-injection, the tumors were resected; the mice were then sacrificed at 2 weeks post-resection. HFD stimulated melanoma growth and lymph node (LN) metastasis as well as tumor and LN lymphangiogenesis. Lipid vacuoles in the tumor and M2-macrophage (MΦ)s in the adipose and tumor tissues were increased in HFD-fed mice. CCL19 and CCL21 contents were higher in LNs than in tumors. HFD increased both CCL19 and CCL21 levels in LNs and CCR7 in tumors. Adipose tissue-conditioned media (CM) from HFD-fed mice enhanced lymphangiogenesis, and mature adipocyte (MA)/M2-MΦ co-culture CM markedly stimulated the tube formation of lymphatic endothelial cell (LEC)s and B16F10 migration. Monocyte migration was moderately stimulated by B16F10 or MA CM, but tremendously stimulated by B16F10/M2-MΦ co-culture CM, which was enhanced by MA/B16F10/M2-MΦ co-culture CM. The co-culture results revealed that MAs increased CCL2, M-CSF and CCR7 mRNAs in B16F10s; vascular endothelial growth factor (VEGF)-D mRNA in M2-MΦs; and CCL19, CCL21 and VEGF receptor (VEGFR)3 mRNA in LECs. M2-MΦs increased CCL2, M-CSF and VEGF-A mRNAs in B16F10s, whereas B16F10s increased VEGF-C mRNAs in M2-MΦs and VEGFR3 mRNA in LECs. These results indicate that in HFD-fed mice, MA-induced CCL2 and M-CSF in tumor cells increase M2-MΦs in tumor; the crosstalk between tumor cells and M2-MΦs further increases cytokines and angiogenic and lymphangiogenic factors. Additionally, MA-stimulated CCL19, CCL21/CCR7 axis contributes to increased LN metastasis in HFD-fed mice.