3D-bioprinting of self-healing hydrogels

Self-healing hydrogels are the most promising hydrogel-ink materials, especially for extrusion-based 3D-bioprinting, because, unlike traditional hydrogels, the bonds as well as their initial structure, properties and functionality can be recovered after extrusion, which together with shear-thinning property enables safe printing for cells, but also shape stability of the construct after printing. In addition to tunable viscoelastic properties given by these inks, they can also respond to cell forces by rearranging the network, while maintaining bulk physiological properties. Currently, mainly extrusion-based bioprinting has been used for these types of dynamic inks. Some basic 3D structures, such as letters, grids and patterns, have been printed with high shape fidelity and high cell viability using traditional 3D-bioprinting. More complex spiral, pyramidal, or vascular tree structures have also been printed using so-called gel-in-gel printing technique, and even some overhang geometries without the need for additional support bath. The current limitation of self-healing hydrogel inks has been their poor mechanical stability, which has been improved, for example, using additional crosslinking. However, the opposing characteristics of self-healing hydrogels, like toughness and fast self-healability, remain a challenge. Therefore, more studies are needed in the future to improve the self-healing hydrogel inks. This review collects some of the most relevant studies related to self-healing 3D-bioprintable hydrogels. It also discusses the importance of self-healing and shear-thinning properties for bioinks and bioprinted constructs, the effects of self-healing hydrogel bioinks and bioprinting on cells (and vice versa), as well as the current status and future prospects of self-healing hydrogel bioinks.Peer reviewe

Characterization of self-healing hydrogels for biomedical applications

Self-healing hydrogels have become attractive biomaterials due to their ability to repair their initial structure and properties in response to damage. When designing ideal self-healing hydrogels the understanding of their properties but also the actual healing process is required. Even though there currently are different characterization methods used, the lack of standardization makes comparison of different hydrogels difficult. The challenges in standardization arise, for example, from the use of different healing methods (i.e. healing environments) or different testing equipments used. In order to help the comparison of hydrogels, a group of characterization methods should be chosen and the measuring parameters and results in the literature should be presented more consistently. The characterization should include methods suitable to determine the presence of reversible interactions and their reversibility study, to investigate the self-healability of hydrogels and to determine the healing efficiencies of hydrogels, not forgetting time dependence and dynamics of self-healing. More quantitative, as well as theoretical studies are recommended. In this review different general characterization methods, including different measuring parameters and environments, used for self-healing hydrogels are charted, but also additional methods suitable for injectable/3D-bioprintable and conductive self-healing hydrogels are discussed. Some challenges of each method and future aspects for self-healing hydrogels and their characterization are also given.publishedVersionPeer reviewe

Design aspects and characterization of hydrogel-based bioinks for extrusion-based bioprinting

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Injectable and self-healing biobased composite hydrogels as future anticancer therapeutic biomaterials

Abstract Self-healing composite hydrogels are prepared from sustainable biopolymers by a green chemistry approach and analyzed by physicochemical and mechanical characterization techniques for future injectable anticancer biomaterials. Water-soluble chitosan (WSC) was prepared by grafting polyethylene glycol (PEG), glutamic acid and gallic acid onto the chitosan chain by carbodiimide chemistry. This WSC showed fast gelation (t ≈ < 60 seconds) with benzaldehyde-terminated 4-arm-PEG as a crosslinker through an amine/aldehyde Schiff base reaction. The compression modulus of these gels can be controlled between 6 and 67 kPa, which was dependent on both the crosslinker content as well as the total solid content (T%). It showed injectability and complete self-healing ability at the lower solid content (T = 2%). The hydrogel nanocomposites (HNCs) were synthesized together with gold (Au) and silver (Ag) nanoparticles (NPs) and tested for cytotoxicity using fibroblast cells (WI-38) for 48 hours, which showed good biocompatibility. The in-vitro assay against cancer cells (U87MG) for 48 hours indicated that only the HNCs with incorporated AuNPs were effective agents for cancer cell apoptosis in contrast to pristine gel, pure NPs (Ag and AuNPs) and HCNs with AgNPs. Therefore, these HNCs could be effective chemotherapeutic materials for designing anticancer nanomedicines in the future.publishedVersionPeer reviewe

Sequential Cross-linking of Gallic Acid-Functionalized GelMA-Based Bioinks with Enhanced Printability for Extrusion-Based 3D Bioprinting

The printability of a photocross-linkable methacrylated gelatin (GelMA) bioink with an extrusion-based 3D bioprinter is highly affected by the polymer concentration and printing temperature. In this work, we developed a gallic acid (GA)-functionalized GelMA ink to improve the printability at room and physiological temperatures and to enable tissue adhesion and antioxidant properties. We introduced a sequential cross-linking approach using catechol–Fe3+ chelation, followed by photocross-linking. The results show that the ink formulation with 0.5% (w/v) Fe3+ in GelMA (30% modification) with 10% GA (GelMA30GA-5Fe) provided the optimum printability, shape fidelity, and structural integrity. The dual network inside the printed constructs significantly enhanced the viscoelastic properties. Printed cylinders were evaluated for their printing accuracy. The printed structures of GelMA30GA-5Fe provided high stability in physiological conditions over a month. In addition, the optimized ink also offered good tissue adhesion and antioxidant property. This catechol-based sequential cross-linking method could be adopted for the fabrication of other single-polymer bioinks.publishedVersionPeer reviewe

Monitoring the gelation of gellan gum with torsion rheometry and brightness-mode ultrasound

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Bioactivated gellan gum hydrogels affect cellular rearrangement and cell response in vascular co-culture and subcutaneous implant models

Hydrogels are suitable soft tissue mimics and capable of creating pre-vascularized tissues, that are useful for in vitro tissue engineering and in vivo regenerative medicine. The polysaccharide gellan gum (GG) offers an intriguing matrix material but requires bioactivation in order to support cell attachment and transfer of biomechanical cues. Here, four versatile modifications were investigated: Purified NaGG; avidin-modified NaGG combined with biotinylated fibronectin (NaGG-avd); oxidized GG (GGox) covalently modified with carbohydrazide-modified gelatin (gelaCDH) or adipic hydrazide-modified gelatin (gelaADH). All materials were subjected to rheological analysis to assess their viscoelastic properties, using a time sweep for gelation analysis, and subsequent amplitude sweep of the formed hydrogels. The sweeps show that NaGG and NaGG-avd are rather brittle, while gelatin-based hydrogels are more elastic. The degradation of preformed hydrogels in cell culture medium was analyzed with an amplitude sweep and show that gelatin-containing hydrogels degrade more dramatically. A co-culture of GFP-tagged HUVEC and hASC was performed to induce vascular network formation in 3D for up to 14 days. Immunofluorescence staining of the αSMA+ network showed increased cell response to gelatin-GG networks, while the NaGG-based hydrogels did not allow for the elongation of cells. Preformed, 3D hydrogels disks were implanted to subcutaneous rat skin pockets to evaluate biological in vivo response. As visible from the hematoxylin and eosin-stained tissue slices, all materials are biocompatible, however gelatin-GG hydrogels produced a stronger host response. This work indicates, that besides the biochemical cues added to the GG hydrogels, also their viscoelasticity greatly influences the biological response.publishedVersionPeer reviewe

Mechanically robust, transparent, and UV-shielding composite of Na-Alginate and maleic acid-functionalized boron nitride nanosheets with improved antioxidant property

Maleic acid functionalized boron nitride nanosheets (BNNS-MA)/Na-Alginate composite with enhanced mechanical, UV-shielding and antioxidation properties have been fabricated for the first time by solvent evaporation from a homogeneous aqueous dispersion of BNNS-MA/Na-Alginate composite solution. The composite fabrication was driven by homogenous nano-integrations and chemistry of compatibilization of BNNS-MA with Na-Alginate through H-bonding interactions between -COOH functional group of BNNS-MA and -OH, -COONa groups of Na-Alginate. The BNNS-MA/Na-Alginate composites show significant enhancement of mechanical, UV-blocking and antioxidant properties compared to the Na-Alginate. Integrating only 1 wt% BNNS-MA improved the UV-blocking, tensile strength, and antioxidant properties of Na-Alginate film by 99.1%, 73% and 60.3%, respectively. Overall, our findings of BNNS-MA integrated Na-Alginate composite films with improved physical, mechanical, UV shielding, and antioxidant functionalities is very promising to open new insight in the field of transparent UV-protected biopolymer film for consumer products, packaging, cosmetics, and engineering applications.Peer reviewe