Abstract
Macrophages are versatile cells of the innate immune system that can adopt a variety of functional phenotypes depending on signals in their environment. In previous work, we found that culture of macrophages on fibrin, the provisional extracellular matrix protein, inhibits their inflammatory activation when compared to cells cultured on polystyrene surfaces. Here, we sought to investigate the role of matrix stiffness in the regulation of macrophage activity by manipulating the mechanical properties of fibrin. We utilize a photo-initiated crosslinking method to introduce dityrosine crosslinks to a fibrin gel and confirm an increase in gel stiffness through active microrheology. We observe that matrix crosslinking elicits distinct changes in macrophage morphology, integrin expression, migration, and inflammatory activation. Macrophages cultured on a stiffer substrate exhibit greater cell spreading and expression of αM integrin. Furthermore, macrophages cultured on crosslinked fibrin exhibit increased motility. Finally, culture of macrophages on photo-crosslinked fibrin enhances their inflammatory activation compared to unmodified fibrin, suggesting that matrix crosslinking regulates the functional activation of macrophages. These findings provide insight into how the physical properties of the extracellular matrix might control macrophage behavior during inflammation and wound healing.
Highlights
Macrophages are innate immune cells that are central to many biological processes including development, metabolism, and tissue homeostasis.[1]
We found that bone marrow derived macrophages (BMDMs) displaced farther from their starting positions and exhibited higher velocities when cultured on fibrin gels, when compared to cells on glass (Fig. 3 and supplementary material, Figs. 2–4), suggesting that perhaps motility is enhanced on a extracellular matrix (ECM) matrix compared to a very stiff 2D surface
We showed scitation.org/journal/apb that culture of BMDMs on soft fibrin gels abrogated tumor necrosis factor alpha (TNF-a) secretion in response to LPS and IFN-c when compared to cells cultured on a stiff polystyrene surface.[36]
Summary
Macrophages are innate immune cells that are central to many biological processes including development, metabolism, and tissue homeostasis.[1] These cells are dynamic regulators of the wound healing process, advancing and resolving inflammation in response to cues in their microenvironment.[2] Macrophages are recognized for their remarkable plasticity and can assume a diverse range of phenotypes depending on cues from their microenvironment.[3] In response to pathogens or damaged cells, macrophages adopt a classically activated, pro-inflammatory phenotype to promote inflammation. In the presence of wound healing cytokines such as interleukin-4 (IL-4) and interleukin-13 (IL-13), macrophages polarize toward a pro-regenerative phenotype critical for tissue repair. Macrophage phenotypes are clearly regulated by soluble cues in their environment, but the role of adhesive and physical cues from the extracellular matrix (ECM) remains less well defined
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