Understanding and Controlling Macrophage Behavior in Angiogenesis

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Project summary At the heart of angiogenesis and biomaterial vascularization lies the inflammatory response, orchestrated primarily by macrophages, which dramatically shift phenotype over time in response to microenvironmental cues. In the normal response to injury, macrophages are initially pro-inflammatory (aka M1), and at later stages they are replaced by a mixed population referred to collectively as M2 that upregulate factors associated with resolution of the wound healing process. Previously we showed that M1 macrophages are critical for the initiation of angiogenesis, but they must switch to M2 for stable angiogenesis and wound healing. The extent of the diversity of this M2 population, and how they regulate angiogenic processes, is still unknown. At later stages of angiogenesis and biomaterial vascularization, M2 macrophages are generated 1) via transition from M1 macrophages, or 2) from direct differentiation of newly arriving monocytes. The differences between the M2 macrophages arising from each population have not been investigated. Preliminary data suggest that M1-derived M2 macrophages possess enhanced angiogenic functionality, and that biomaterials that transiently stimulate the initial M1 phase may enhance the subsequent response to M2-promoting biomaterials to achieve enhanced vascularization and healing. The overarching hypothesis of this project is that biomaterials that promote sequential M1 and M2 activation of the same population of macrophages will enhance vascularization. We recently found that adoptively transferred macrophages and M2-promoting microparticles are synergistic in promoting tissue revascularization in a murine hindlimb ischemia model. However, the macrophages were rapidly cleared from the site of injury (within 2 days), making it difficult to study the phenotype changes in adoptively transferred macrophages. Therefore, the goal of this project is to adoptively transfer macrophages using biomaterial scaffolds as a cell carrier, so that the macrophages are retained at the site of injury for long enough to track their phenotype changes and to observe their interactions with other cell types involved in angiogenesis. We will use porous scaffolds (as opposed to hydrogels) because other work in our lab has shown using in vitro studies that macrophage-T cell crosstalk is important for the M1-to-M2 transition of macrophages, so it is important that T cells are able to inflitrate the biomaterial carrier. In the current project, we will administer pro-inflammatory M1 macrophages to the site of injury using porous scaffolds in order to test the hypothesis that M1 macrophage crosstalk with T cells promotes Th2 differentiation in T cells, the M1-to-M2 transition in macrophages, and enhanced angiogenesis. The hypothesis is that adoptively transferred M1 macrophages will transition into a distinct M2 phenotype with enhanced angiogenic functions (compared to M0 macrophages) via crosstalk with endogenous T cells.