Bone marrow-derived, alternatively activated macrophages enhance solid tumor growth and lung metastasis of mammary carcinoma cells in a Balb/C mouse orthotopic model

Abstract

Introduction: Tumor-associated macrophages, which are derived from the infiltration of circulating bone marrow-derived monocytes, consist primarily of a polarized M2 macrophage (M2-M phi) population and are associated with poor prognosis in various cancers. In the present study, we attempted to assess whether M2-M phi s derived from bone marrow stimulate the promotion and progression of mammary tumors. Methods: 4T1 murine mammary carcinoma cells were injected either alone or coupled with M2-M phi s into the mammary fat pads of syngeneic female Balb/C mice. M2-M phi s were prepared by treating monocytes isolated from female Balb/C mouse bone marrow with IL-4. Tumor cell growth was determined using an in vivo imaging system and the expression of cell proliferation-related, angiogenesis-related, and lymphangiogenesis-related proteins in tumor tissues was immunohistochemically analyzed. To evaluate the effects of the crosstalk between 4T1 cells and M2-M phi s on the secretion and mRNA expression of cytokines and the migration of monocytes, 4T1 cells and M2-M phi s were co-cultured and cytokine antibody array, real-time RT-PCR, and trans-well migration assays were conducted. Results: The co-injection of M2-M phi s into the mammary fat pads of mice increased solid tumor growth and lung metastasis of 4T1 cells as well as the infiltration of CD45(+) leukocytes into tumor tissues. The proportions of Ki-67(+) proliferating cells and the expression of hypoxia inducible factor-1 alpha, vascular endothelial cell growth factor A, CD31, vascular endothelial cell growth factor C, and lymphatic vessel endothelial receptor-1 were increased significantly in the tumor tissues of mice co-injected with 4T1 cells and M2-M phi s. The in vitro results revealed that the proliferation of 4T1 cells, the migration of monocytes, and the secretion of granulocyte colony-stimulating factor, IFN gamma, IL-1 alpha, IL-2, IL-16, IFN gamma-induced protein-10, keratinocyte-derived chemokine, macrophage colony-stimulating factor, monocyte chemotactic protein-1, macrophage inflammatory protein-1 alpha, and RANTES were increased when 4T1 cells were co-cultured with M2-M phi s, as compared with when the 4T1 cells were cultured alone. Conclusion: The crosstalk between 4T1 cells and M2-M phi s increased the production of cytokines, which may have induced immune cell infiltration into tumor tissues, tumor cell proliferation, angiogenesis, and lymph angiogenesis, thereby increasing solid tumor growth and lung metastasis.