title: Hyaluronic acid-fibrin interpenetrating double network hydrogel prepared in situ by orthogonal disulfide cross-linking reaction for biomedical applications.
abstract: To strengthen the mechanical properties of a fibrin gel and improve its applicability as a scaffold for tissue engineering (TE) applications, a strategy for the in situ preparation of the interpenetrating network (IPN) of fibrin and hyaluronic acid (HA) was developed on the basis of simultaneous and orthogonal fibrinogenesis and disulfide cross-linking. The synthetic pathway included the preparation of mutually reactive HA derivatives bearing thiol and 2-dithiopyridyl groups. Combining thiol-derivatized HA with thrombin and 2-dithiopyridyl-modified HA with fibrinogen and then mixing the obtained liquid formulations afforded IPNs with fibrin-resembling fibrillar architectures at different ratios between fibrin and HA networks. The formation of two networks was confirmed by conducting reference experiments with the compositions lacking one of the four components. The composition of 2% (w/v) fibrin and 1% (w/v) HA showed the highest storage modulus (G'), as compared with the single network counterparts. The degradation of fibrin in IPN hydrogels was slower than that in pure fibrin gels both during incubation of the hydrogels in a fibrin-cleaving nattokinase solution and during the culturing of cells after their encapsulation in the hydrogels. Together with the persistence of HA network, it permitted longer cell culturing time in the IPN. Moreover, the proliferation and spreading of MG63 cells that express the hyaluronan receptor CD44 in IPN hydrogel was increased, as compared with its single network analogues. These results are promising for tunable ECM-based materials for TE and regenerative medicine.
  
  
  
  The present work is devoted to in situ fabrication of injectable extracellular matrix hydrogels through simultaneous generation of networks of fibrin and hyaluronic acid (HA) that interpenetrate each other. This is accomplished by combination of enzymatic fibrin cross-linking with orthogonal disulphide cross-linking of HA. High hydrophilicity of HA prevents compaction of the fibrin network, while fibrin provides an adhesive environment for in situ encapsulated cells. The interpenetrating network hydrogel shows an increased stiffness along with a lower degradation rate of fibrin in comparison to the single fibrin network. As a result, the cells have sufficient time for the remodelling of the scaffold. This new approach can be applied for modular construction of in vitro tissue models and tissue engineering scaffolds in vivo.
authors.affiliation: Science for Life Laboratory, Division of Polymer Chemistry, Department of Chemistry-Ångström, Uppsala University, SE 751 21 Uppsala, Sweden.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Austrian Cluster for Tissue Regeneration, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Donaueschingenstrasse 13, A-1200 Vienna, Austria; Trauma Care Consult GmbH, Gonzagagasse 11/25, A-1010 Vienna, Austria.
authors.forename: Philipp
authors.initials: P
authors.lastname: Heher
authors.affiliation: Science for Life Laboratory, Division of Polymer Chemistry, Department of Chemistry-Ångström, Uppsala University, SE 751 21 Uppsala, Sweden.
authors.forename: Jöns
authors.initials: J
authors.lastname: Hilborn
authors.affiliation: Austrian Cluster for Tissue Regeneration, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Donaueschingenstrasse 13, A-1200 Vienna, Austria; Trauma Care Consult GmbH, Gonzagagasse 11/25, A-1010 Vienna, Austria. Electronic address: office@trauma.lbg.ac.at.
authors.forename: Heinz
authors.initials: H
authors.lastname: Redl
authors.affiliation: Science for Life Laboratory, Division of Polymer Chemistry, Department of Chemistry-Ångström, Uppsala University, SE 751 21 Uppsala, Sweden. Electronic address: dmitri.ossipov@kemi.uu.se.
authors.forename: Dmitri A
authors.initials: DA
authors.lastname: Ossipov
mesh_terms: D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003432:Cross-Linking Reagents / Q000737:chemistry; D005109:Extracellular Matrix / Q000737:chemistry; D005337:Fibrin / Q000737:chemistry*; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis*; D006820:Hyaluronic Acid / Q000138:chemical synthesis / Q000737:chemistry*; D020100:Hydrogels / Q000138:chemical synthesis / Q000737:chemistry*; D008422:Materials Testing
chemical_list: C497483:CD44 protein, human; D003432:Cross-Linking Reagents; D018960:Hyaluronan Receptors; D020100:Hydrogels; D005337:Fibrin; D006820:Hyaluronic Acid
keywords: 3D cell culture; Biodegradation; Fibrin; Hyaluronic acid; Interpenetrating hydrogel