Sculpting the blank slate: how fibrin's support of vascularization can inspire biomaterial design

Abstract

Fibrin is the primary extracellular constituent of blood clots, and plays an important role as a provisional matrix during wound healing and tissue remodeling. Fibrin-based biomaterials have proven their utility as hemostatic therapies, scaffolds for tissue engineering, vehicles for controlled release, and platforms for culturing and studying cells in three dimensions. Nevertheless, fibrin presents a complex milieu of signals to embedded cells, many of which are not well understood. Synthetic extracellular matrices (ECMs) provide a blank slate that can ostensibly be populated with specific bioactive cues, including growth factors, growth factor binding motifs, adhesive peptides and peptide crosslinks susceptible to proteases, thereby enabling a degree of customization for specific applications. However, the continued evolution and improvement of synthetic ECMs requires parallel efforts to deconstruct native ECMs and decipher the cues they provide to constituent cells. The objective of this review is to reintroduce fibrin, a protein with a well-characterized structure and biochemistry, and its ability to support angiogenesis specifically. Although fibrin's structure-function relationships have been studied for decades, opportunities to engineer new and improved synthetic hydrogels can be realized by further exploiting fibrin's inspiring design.

Keywords: Angiogenesis; Biomaterial; Extracellular matrix; Fibrin.

Figure 1. Schematic of fibrinogen and its assembly into fibrin

This cartoon representation shows the central E domain of fibrinogen flanked by the two globular D domains linked together by coiled-coil regions. Thrombin cleavage of fibrinopeptides A and B from the E domain leads to dimerization of soluble fibrin molecules into fibrin dimers, the first step of fibrin polymerization. Additionally, thrombin cleavage of factor XIII produces factor XIIIa, which crosslinks adjacent γ chains, linking the D domains in the growing fibrin polymer. Figure adapted from [6] with permission from John Wiley and Sons.

Figure 2. Fibrin hydrogels are useful as 3D model systems of angiogenic sprouting

High magnification confocal images of a sprouting capillary in a 3D fibrin matrix containing FITC-conjugated fibrinogen. HUVECs cultured in fibrin gels form capillaries with hollow lumens as demonstrated by confocal sections obtained at the bottom (left), center (middle), and top (right) z-planes of a capillary. HUVECs were stained with Oregon Green 488 phalloidin and nuclei were counterstained with DAPI. The HUVECs create a 3D channel in the fibrin matrix via proteolysis.

Figure 3. Engineering fibrin for controlled growth factor presentation

(A) The recombinant protein TG-ephrin B2, the full length ectodomain of ephrin B2 fused to a transglutaminase-sensitive (TG) motif, was conjugated to fibrin via factor XIIIa through the TG region of the protein. (B) Modifying fibrin in this way improved the angiogenic response of acellular fibrin implants in a chick chorioallantoic membrane (CAM) model, producing localized angiogenesis at the implant site. Pure fibrin implants showed little vessel ingrowth. (B) In vivo examination of microvascular growth on the growing CAM revealed strong and specific induction of new vessels by ephrin-B2 from fibrin grafts. Blood vessels were monitored by in vivo fluorescence microscopy performed after Intravenous injections of FITC-dextran, and showed larger diameters of vessels induced by ephrin-B2 indicative of arterioles or venules; by contrast, no changes of the regular vessel pattern of the CAM were observed in response to plain fibrin. Figure adapted from [88] with permission from Elsevier.