Characterization and functional study of macrophages associated with pulmonary metastasis

Abstract

There is a growing appreciation of the importance of tumor-stroma interactions in the progression of tumors to malignancy. In the tumor stroma, macrophages are abundant and have been shown to enhance malignant processes and in particular to enhance metastasis. Experiments reported in this thesis have demonstrated that the two last stages of pulmonary metastasis, seeding and persistent growth are regulated by a population of macrophages newly recruited to the lung in response to these cells. This requirement was demonstrated using mice homozygous for a null mutation in the colony stimulating factor-1 (CSF-1) gene that is depleted in the recruited macrophages and by using liposome Clodronate depletion of macrophages in vivo. Fluorescent activated cell sorting analysis revealed recruitment of CSF-1R+CD11b+Gr1-VEGFR1 highCCRhigh macrophages in the metastasis-bearing lung. To test their function, the CD11b+ macrophages were deleted in vivo with diphtheria toxin (DT) treatment in mosaic animals generated by bone marrow transplant using a transgenic mouse expressing simian DT receptor driven by the CD11b promoter as a bone marrow donor that confers susceptibility to these cells to DT. The deletion of CD11b+ macrophages reduced the tumor cell seeding efficiency and growth rate in the lung. Importantly depletion of macrophages following initial tumor cell seeding blocked the growth of already established metastases. An intact lung 3D imaging system revealed an increased macrophage and tumor cell interaction over time and this is temporally correlated with tumor cell extravasation and survival in the lung. Depletion of macrophages using liposome Clodronate was shown using this method to inhibit tumor extravasation in vivo. Microarray analysis revealed a distinct gene expression signature of these metastasis-associated macrophages. Cell surface adhesion molecules and cytokine receptors expressed by these macrophages were studied both in vivo and in vitro to reveal the molecular mechanism for their recruitment and function in metastasis seeding and growth. Together these data clearly demonstrated the importance of a recruited macrophage population displaying a distinct phenotype in tumor cell extravasation, seeding and growth in a metastatic target organ. Furthermore, these studies reveal the potential of targeting specific pathways in these cells to block the malicious spread of tumor cells.