Methacrylated Chitosan-gelatin Hydrogel Composites For 3D Bioprinting And Tissue Engineering

The advent and progress of tissue engineering(TE)holds a great promise as a technique to alleviate crisis associated with organ shortage and transplantation.Biomimetic constructs imitating the functions,structures and compositions of normal tissues are of great importance for tissue repair and regeneration.Hence,there is an increase in the number of manufacturing technologies such as 3D bioprinting for rapid fabrication of biomimetic complex structured scaffolds.Chitosan methacryloyl(Ch MA)is a polysaccharide with well-documented history for its potential in tissue engineering.In fact,by virtue of its favorable intrinsic biological properties and structural features,Ch MA has been validated through wide applications in the biomedical,food,pharmaceutical and cosmetics industries.Thanks to its biodegradability,biocompatibility,anti-inflammatory and anti-oxidation effects,it promises exceptional prospects in tissue-specific applications.Additionally,Ch MA as 3D polymeric hydrogels displays high water retention ability,which not only enables aqueous liquid absorption,but maintains their mechanical stability while creating the molecular environment desired for tissue regeneration.To be suitable for tissue engineering applications,Ch MA was blended with gelatin methacryloyl(Gel MA)which has shown excellent and tunable mechanical properties along with biochemical characteristics that could imitate the requirements for scaffolds.Therefore,this thesis describes a new protocol to develop printable and strong biomaterial inks for 3D printing by blending Ch MA with methacrylate gelatin(Gel MA)and hydroxyapatite(Hap).Furthermore,to offset the problem of inferior mechanical nature of hydrogels,two-step sequential crosslinking was employed to generate a mechanically enhanced photocrosslinkable double network(DN)hydrogel-based platform.The main contents are as follow:1.3D-Printable Thermo/Photo-Crosslinked Methacrylated Chitosan-Gelatin Hydrogel Composites for Tissue RegenerationThree-dimensional(3D)printing is an innovative method which can create patient-specific scaffolds through an automated and scalable process.Therefore,intricate biomimetic 3D tissue scaffolds with intrinsic architectural organization that can accelerate tissue regeneration is highly demanded.Here we present a printable hydrogel ink based on methylacrylate-modified chitosan(Ch MA)and gelatin(Gel MA)embedding nanohydroxyapatite(Hap).This polymer composite is first physically crosslinked by thermal gelation for post-printing structural stability,followed by covalent photo-crossslinking of Ch MA and Gel MA to form a long-term stable structure.The rheological behavior of the hydrogels and the mechanical strengths of the printed constructs are tuned by adjusting the content of Gel MA,which in turn enhances the shape retention after printing and enables the precise deposition of multilayered 3D scaffolds.Moreover,the formulated biomaterial inks exhibits biological characteristics that effectively support the spreading and proliferation of stem cells seeded on the scaffolds after 7 days of in vitro culture.Adding Hap has minor influences on the mechanical rigidity and cytocompatibility of the hydrogels compared with the group free of Hap.Together,the printable biomaterial inks with shear thinning and good structural integrity,along with biological cues,are promising for tissue engineering application.2.Double network hydrogels based on photo-crosslinkable chitosan and gelatin support cell growth and functionsAs another component of this work,double network hydrogels were created by a two-step photo-crosslinking using Ch MA and Gel MA.Hydrogel polymers chains have low density and small friction between their chains,causing them to exhibit sollike behavior,which result in inferior mechanical stiffness.Inspired by the unique contrasting double network(DN)structure,a DN hydrogel based on photocrosslinkable chitosan and gelatin is proposed to tackle the existing concerns.In this study,DN hydrogels was designed by interspersing a Gel MA network into a 3D polymeric hydrogel consisting of Ch MA via a two-step sequential photo polymerization.The objective was to generate a hydrogel framework with enhanced stiffness,in the perspective of obtaining mimetic functional tissues able to reconstruct lost or impaired tissues in vivo.Diffusion of Gel MA into the Ch MA network was observed to confirm the formation of DN.This inherent combination induces homogenous distribution of crosslinks and network structure which provides extra energy under loadings,resulting in the turning of Ch MA’s brittle nature.In fact,the mechanical properties of the DN hydrogels significantly increased with immersion time,indicating that the stiffness of the DN hydrogels could be controlled by adjusting Gel MA infiltration time.Moreover,the mechanical strengths of the sequentially cross-linked hydrogel could be manipulated with different Gel MA content,indicating that such DN hydrogels have widespread adaptability for in vitro cell studies.Results from the culture revealed biocompatible features with no cytotoxicity for the designed DN hydrogels.