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Hyaluronic acid-fibrin interpenetrating double network hydrogel prepared in situ by orthogonal disulfide cross-linking reaction for biomedical applications
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Polymer Chemistry.
Ludwig Boltzmann Inst Expt & Clin Traumatol, Austrian Cluster Tissue Regenerat, Donaueschingenstr 13, A-1200 Vienna, Austria.;Trauma Care Consult GmbH, Gonzagagasse 11-25, A-1010 Vienna, Austria..
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Polymer Chemistry.
Ludwig Boltzmann Inst Expt & Clin Traumatol, Austrian Cluster Tissue Regenerat, Donaueschingenstr 13, A-1200 Vienna, Austria.;Trauma Care Consult GmbH, Gonzagagasse 11-25, A-1010 Vienna, Austria..
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2016 (English)In: Acta Biomaterialia, ISSN 1742-7061, E-ISSN 1878-7568, Vol. 38, 23-32 p.Article in journal (Refereed) PublishedText
Abstract [en]

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. Statement of Significance 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.

Place, publisher, year, edition, pages
2016. Vol. 38, 23-32 p.
Keyword [en]
Interpenetrating hydrogel, Fibrin, Hyaluronic acid, Biodegradation, 3D cell culture
National Category
Polymer Chemistry Bio Materials
URN: urn:nbn:se:uu:diva-300044DOI: 10.1016/j.actbio.2016.04.041ISI: 000378963800003PubMedID: 27134013OAI: oai:DiVA.org:uu-300044DiVA: diva2:950772
EU, European Research Council
Available from: 2016-08-02 Created: 2016-08-02 Last updated: 2016-09-02Bibliographically approved
In thesis
1. Hyaluronan Based Biomaterials with Imaging Capacity for Tissue Engineering
Open this publication in new window or tab >>Hyaluronan Based Biomaterials with Imaging Capacity for Tissue Engineering
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis presents the preparation of hyaluronan-based biomaterials with imaging capability and their application as scaffolds in tissue engineering. First, we have synthesized HA derivatives functionalized with different chemoselective groups. Then, functional ligands with capacities for hydrophobic drug loading, imaging, and metal ion coordination were chemically conjugated to HA by chemoselective reactions with these groups. An injectable in situ forming HA hydrogel was prepared by hydrazone cross-linking between hybrid iron-oxide nanogel and HA-aldehyde (paper-I). The degradation of this hydrogel could be monitored by MRI and UV-vis spectroscopy since it contained iron oxide as a contrast agent and pyrene as a fluorescent probe. Additionally, this hydrogel has a potential for a delivery of hydrophobic drugs due to its pyrene hydrophobic domains. The degradation study showed that degradability of the hydrogel was correlated with its structure. Based on the obtained results, disulfide cross-linked and fluorescently labeled hydrogels with different HA concentration were established as a model to study the relationship between the structure of the hydrogel and its degradability (paper-II). We demonstrated that disulfide cross-linked HA hydrogel could be tracked non-invasively by fluorescence imaging. It was proved that the in vivo degradation behavior of the hydrogel is predictable basing on its in vitro degradation study. In paper-III, we developed a disulfide cross-linked HA hydrogel for three-dimensional (3D) cell culture. In order to improve cell viability and adhesion to the matrix, HA derivatives were cross-linked in the presence of fibrinogen undergoing polymerization upon the action of thrombin. It led to the formation of an interpenetrating double network (IPN) of HA and fibrin. The results of 3D cell culture experiments revealed that the IPN hydrogel provides the cells with a more stable environment for proliferation. The results of the cellular studies were also supported by in vitro degradation of IPN monitored by fluorescence measurements of the degraded products. In paper-IV, the effect of biomineralization on hydrogel degradation was evaluated in a non-invasive manner in vitro. For this purpose, two types of fluorescently labeled hydrogels with the different ability for biomineralization were prepared. Fluorescence spectroscopy was applied to monitor degradation of the hydrogels in vitro under two different conditions in longitudinal studies. Under the supply of Ca2+ ions, the BP-modified hydrogel showed the tendency to bio-mineralization and reduction of the rate of degradation. Altogether, the performed studies showed the importance of imaging of hydrogel biomaterials in the design of optimized scaffolds for tissue engineering.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 60 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1402
hyaluronan, hydrogel, scaffold, tissue engineering, imaging, interpenetrating network, MRI, fluorescence imaging, scaffold degradation
National Category
Materials Chemistry Polymer Chemistry
urn:nbn:se:uu:diva-300799 (URN)978-91-554-9649-4 (ISBN)
External cooperation:
Public defence
2016-09-29, 80101, Lägerhyddsvägen 1, Uppsala, 09:00 (English)
Available from: 2016-09-06 Created: 2016-08-14 Last updated: 2016-09-13

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