The influence of chemically modified Gellan Gum on macrophage polarization, fibroblast differentiation, and collagen orientation

Abstract

The wound healing process is a cascade of cell-cell signalling events that can be disturbed by the presence of a biomaterial. Macrophages and fibroblasts play critical roles in determining the host response to implanted biomaterials. As a consequence of the dysregulation of the macrophage or/and fibroblast response, increased pro-fibrotic/pro-inflammatory cytokines can induce chronic inflammation and/or fibrosis, which may negatively impact the function of the implant. The macrophage phenotype is dynamic throughout the host response. The plasticity of macrophage also strongly influences the proliferation and remodelling phase. Inflammatory mediators secreted from the polarized macrophages have a direct impact on fibroblast response. The contractility of activated fibroblasts (myofibroblasts) is largely responsible for wound closure and generation of collagen-rich (fibrotic) extracellular matrix (ECM). Both macrophage and fibroblasts respond rapidly to microenvironmental signals, such as: local paracrine signals from neighbouring cells, the secretion pattern of cytokines and chemokines, and physical and chemical cues arising from implantation. Thus, a balance of phenotypes of macrophage and controlled fibroblast transition is needed for timely progression from injury to proper wound healing. It is essential to predict how materials will modulate the response of phenotypic change of cells and macrophage-fibroblast interactions. Gellan gum, a polysaccharide derived from the bacterium Sphingomonas elodea, has been chemically modified with methacrylate and crosslinked through three polymerization methods: step-growth, chain-growth, and mixed model crosslinking. Macrophage phenotype shifts from M1 to M2 is controlled by the different crosslinking mechanisms, physical properties, and the chemistry of methacrylated gellan gum hydrogels. In tissue regeneration, the goal is to mimic the chemical and physical properties of living tissue and regenerate tissue similar to what was damaged, missing, or replaced. Fibronectin (FN)-coated methacrylated gellan gum crosslinked through the step-growth mechanism with varying stiffness can influence the function of macrophages. Untreated stiffer gels induced higher production of pro-inflammatory cytokines while FN surface coating shifts the macrophage phenotype on stiffer gels from an M1-like to an M2-like state. Using mechanical gradients and surface modification to control macrophage polarization can be a useful tool in ensuring a proper healing response and for tissue engineering. Fibrotic scar formation can induce dissimilar tissue characteristics compared to native tissue. Due to the strong dependence of fibroblasts on the macrophage phenotype in the wound healing process, a co-culture system was studied to better mimic the complexity of local implant-associated inflammation. Also, collagen orientation plays a critical role in wound contraction and scar formation and it is modulated by myofibroblasts. Macrophage soluble paracrine signals (secreted cytokines) have been shown have a direct influence on fibroblast proliferation, morphology, differentiation, migration, and deposition of collagen via focal adhesion kinase (FAK), Rho-associated protein kinase (ROCK), and myosin II pathways. Altering collagen orientation during the healing process by exerting physical/chemical properties of biomaterial is necessary for promoting tissue regeneration or/and perverting the functional and esthetic consequences. Macrophage phenotype and function have been shown to respond directly to ECM/substrate stiffness. The motility and phagocytic activity of macrophages are mediated by signaling involving Rac GTPase, Rho-associated protein kinase (ROCK), and myosin-II. It is essential for us to better understand macrophage functions on chemically modified gellan gum, which has the potential for repairing soft tissue. Inhibiting the RhoA pathway is thought to decrease the phagocytosis ability of macrophages and prevent macrophages from migrating into the graft. A carefully balanced inhibition of ROCK pathway and regulate material stiffness exerted on macrophage polarization, phagocytosis, and migration can be both beneficial and efficient for transplant survival.