Biomaterial-mediated control over macrophage behavior for tissue regeneration

Claire Elizabeth Witherel

doi:10.17918/kfds-na77

Macrophages, the primary cell of the immune response, and fibroblasts, the major producers of extracellular matrix (ECM), have significant roles in wound healing and the foreign body response to implanted biomaterials. Macrophages are known to exhibit a spectrum of unique phenotypes in response to their environment; M1 macrophages, are associated with increased inflammation and initiating angiogenesis, while M2a macrophages have been associated with anti-inflammatory behavior and extracellular matrix deposition. Temporal control of macrophage phenotype from M1 to M2a has been shown to be critical in normal wound healing. On the other hand, dysregulated macrophage behavior has been associated with detrimental pathologies including chronic wounds and fibrosis. Therefore, the development of immunomodulatory biomaterials that can harness the natural immune response for repair and healing, hold significant promise for future of regenerative medicine. While it's appreciated that macrophages may hold the key to aberrant healing outcomes, it is not well understood how they direct pro-healing outcomes, especially within the context of a biomaterial implant. Therefore, the overall goals of this work were to 1. thoroughly characterize macrophage-biomaterial interactions and macrophage-fibroblast interactions further our understanding of how these cells may influence functional tissue regeneration, and 2. Utilize a model drug-eluting biomaterial to directly assess the role of M2a macrophages in tissue regeneration. To accomplish these goals, I investigated how macrophages change their behavior in response to 'successful', commercially- available wound matrices in vitro. Collectively, these studies showed that four commercially-available wound matrices evaluated pro-inflammatory macrophage behavior (more M1-like) and that direct contact (as opposed to soluble factors) of macrophages with a bioactive (i.e. containing cells and proteins) wound matrix was critical for reducing inflammation in pro-inflammatory macrophages. Next, I investigated the effects of macrophage-secreted signals on fibroblast behavior and matrix formation in vitro and employed model drug-eluting biomaterials to develop an M2a-promoting hydrogel to directly assess the role of M2a macrophages in the foreign body response in vivo. Together, these studies illustrated that M1 and M2a macrophage-derived signals significantly reduced the fiber diameter of fibroblast-derived deposition in vitro relative to other macrophage phenotypes. While clinically relevant macrophage-derived signals (M1+M2a and M1[right arrow]M2a) returned fibroblast matrix fiber diameter to the in vitro baseline. IL4+IL13-releasing hydrogel resulted in a significant reduction in leukocyte recruitment 21 days relative to the Blank hydrogel control, suggesting an increased presence of other key ECM-producing cells. Alternatively, these results may also imply that IL4+IL13 hydrogels may change the quality and composition of ECM surrounding a biomaterial or at the tissue-biomaterial interface. Interestingly and in contrast to pilot data, the IL4+IL13 hydrogel yielded minimal differences in gene expression and histological outcomes compared the Blank control. Collectively, these preliminary and somewhat inconclusive in vivo results are likely due to technical challenges there were discovered in the in vivo experimental design. A final validation study showed that the IL4+IL13 hydrogels that were originally implanted within the same mouse as the Blank controls were likely releasing enough drug to have an impact on the outcomes of the Blank controls. Preliminary data from the validation study yielded a robust response of the IL4+IL13 hydrogel relative to the Blank control at the Day 3 time point. Future work will focus on elucidating the long-term effects of these IL4+IL13 hydrogels on ECM deposition. This work is critical for understanding how transient control of macrophage behavior is directly linked to tissue repair and regeneration outcomes. Due to the importance of macrophage behavior in all tissues, this project has the potential to translate into a myriad of other fields, especially those in which abnormal inflammation prevent tissue repair or regeneration.

Biomaterial-mediated control over macrophage behavior for tissue regeneration

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Biomaterial-mediated control over macrophage behavior for tissue regeneration

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