The response of infiltrating and resident macrophages to melanoma challenge in the mouse lung reveals 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 inflammatory response that shifted to expression of Arg1, and elevated Ccl2, Vcam1, Vegfa, Stab1, Lyve1 and Ptgs2. In contrast, at the later times, resident lung (alveolar) macrophages expressed pro-inflammatory genes, notably Il12b. The macrophages expressed three chemokine receptors that could direct the recruitment of macrophages to metastatic colonies: CCR1, CCR2 and CCR5. B16F10 cells secrete large quantities of RANTES which engages CCR1 and CCR5 and thus can contribute to macrophage chemotaxis. Infiltrating monocytes/macrophages, as well as alveolar macrophages, showed increased chemokine expression in response to tumor challenge. These chemokines included the ligands for CCR1 and CCR5, Mip1a and Mip1b, thus providing a constant source of further macrophage recruitment. The expression of the CCR2 ligand, CCL2, was also strikingly increased by the infiltrating monocytes/ macrophages, and thus constituted 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 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 also be discussed, together with its effect on the phenotype of macrophage subpopulations.