The response of infiltrating and resident macrophages to melanoma challenge in the mouse lung: Involvement of the CC-chemokine axis in the recruitment of pro-tumoral macrophages

In a pulmonary metastasis model using B16F10 melanoma, we found monocyte/macrophage infiltration gradually increasing in the course of metastatic progression. The gene expression pattern of the infiltrating macrophages revealed an early (d3) inflammatory response that shifted to a tumor-promoting late response (d21). The pro-tumoral expression pattern was characterized by Arg1 and elevated expression of Ccl2, Vcam1, Vegfa, Stab1, Lyve1 and Ptgs2. In contrast, while enhancing their anti-inflammatory phenotype, resident lung (alveolar) macrophages also expressed pro-inflammatory genes, notably Il12b.We found at least three chemokine receptors involved in the recruitment of macrophages into the metastatic lung: CCR1, CCR2 and CCR5. The B16F10 cells secreted large quantities of RANTES which in turn activates CCR1 and CCR5, and thus contribute to macrophage chemotaxis. Infiltrating monocytes/macrophages, as well as alveolar macrophages, revealed an increased chemokine expression level in response to tumor challenge. These chemokines included the ligands for CCR1 and CCR5, Mip1α and Mip1β, thus providing a constant means of further macrophage recruitment. Interestingly, the expression of the CCR2 ligand, CCL2, was found strikingly increased by infiltrating monocytes/ macrophages, thus ensuring a positive feed-back loop for macrophage recruitment.In order to explore the involvement of CC chemokine receptors in pulmonary metastasis, we treated the mice with antagonists against CCR1, CCR5 and CCR2 after i.v. B16F10 challenge. The CCR1 and CCR5 antagonist treatments resulted in a 20% decrease in the number of lung colonies each. FACS analysis of the B16F10 colony bearing lungs revealed a distribution of macrophage subsets quite different from untreated lungs, highlighting the importance of CC-chemokine receptors in the recruitment of pro-tumoral macrophages. The effect of a CCR2 antagonist on lung colony formation will be discussed, together with its effect on the phenotype of macrophage subpopulations.