3D printable hyaluronic acid-based hydrogel for its potential application as a bioink in tissue engineering

Abstract

Background: After recognition of 3D printing and injectable hydrogel as a critical issue in tissue/organ engineering and regenerative medicine society, many hydrogels as bioinks have been developed worldwide by using polymeric biomaterials such as gelatin, alginate, hyaluronic acid and others. Even though some gels have shown good performances in 3D bioprinting, still their performances do not meet the requirements enough to be used as a bioink in tissue engineering.

Method: In this study, a hydrogel consisting of three biocompatible biomaterials such as hyaluronic acid (HA), hydroxyethyl acrylate (HEA) and gelatin-methacryloyl, i.e. HA-g-pHEA-gelatin gel, has been evaluated for its possibility as a bioprinting gel, a bioink. Hydrogel synthesis was obtained by graft polymerization of HEA to HA and then grafting of gelatin- methacryloyl via radical polymerization mechanism. Physical and biological properties of the HA-based hydrogels fabricated with different concentrations of methacrylic anhydride (6 and 8%) for gelatin-methacryloylation have been evaluated such as swelling, rheology, morphology, cell compatibility, and delivery of small molecular dimethyloxalylglycine. Printings of HA-g-pHEA-Gelatin gel and its bioink with bone cell loaded in lattice forms were also evaluated by using home-built multi-material (3D bio-) printing system.

Conclusion: The experimental results demonstrated that the HA-g-pHEA-gelatin hydrogel showed both stable rheology properties and excellent biocompatibility, and the gel showed printability in good shape. The bone cells in bioinks of the lattice-printed scaffolds were viable. This study showed HA-g-pHEA-Gelatin gel's potential as a bioink or its tissue engineering applications in injectable and 3D bioprinting forms.

Keywords: 3D bioprinting; Biocompatible; Gelatin; Hyaluronic acid; Tissue engineering.

Fig. 1

Swelling behaviors of HA-g-pHEA-Gelatin hydrogel at pH 7.0 and pH 7.4 and 37 °C, where Gelatin-MA were fabricated with 6% (a) and 8% (b) methacrylic anhydride, respectively

Fig. 2

Digital (a, d) and surface (b, e) and cross-section (c, f) of scanning electron microscopy (b, c, e and f) morphologies of HA-g-pHEA-Gelatin hydrogel (a, b and c: 6% methacrylic anhydride and D, E and F: 8% methacrylic anhydride). The gel composition is 0.25 g HA, 3 mL HEA, 0.25 g Gelatin-MA, and the scale bars of (b and c) and (e and f) are 5 μm and 10 μm, respectively

Fig. 3

Rheological behaviors of HA-g-pHEA-Gelatin (6% methacrylic anhydride) hydrogel, where the relations of (a) viscosity change over shear rate; storage-loss modulus and complex viscosity over oscillation stress (b) and frequency (c)

Fig. 4

Release of DMOG drug in different amount from the HA-HEA-Gelatin hydrogel over time

Fig. 5

Optical images of printed HA-g-pHEA-Gelatin gel scaffolds with/without cells. (a, b, c, d) Non-cell loaded hydrogel and (e, f, g, h) cell-loaded bioinks were extruded from the nozzle by applying different air pressures of 450, 500, 550 and 600 kPa respectively

Fig. 6

Live and dead assay results of the bone cells s in HA-g-pHEA-Gelatin (a, b) and 1 day in vitro cell culture after bioprinting of the HA-g-pHEA-Gelatin gel as a bioink (c, d)