Ultra-low content physio-chemically crosslinked gelatin hydrogel improves encapsulated 3D cell culture.

Gelatin-based hydrogels are extensively used for 3D cell culture, bioprinting, and tissue engineering due to their cell-adhesive nature and tunable physio-chemical properties. Gelatin hydrogels for 3D cell culture are often developed using high-gelatin content (frequently 10-15 % w/v) to ensure fast gelation and improved stability. While highly stable, such matrices restrict the growth of encapsulated cells due to creating a dense, restrictive environment around the encapsulated cells. Hydrogels with lower polymer content are known to improve 3D cell growth, yet fabrication of ultra-low concentration gelatin hydrogels is challenging while ensuring fast gelation and stability. Here, we demonstrate that physical gelation and photo-crosslinking in gelatin results in a fast-gelling hydrogel at a remarkably low gelatin concentration of 1 % w/v (GelPhy/Photo). The GelPhy/Photo hydrogel was highly stable, allowed uniform 3D distribution of cells, and significantly improved the spreading of encapsulated 3T3 fibroblast cells. Moreover, human cholangiocarcinoma (HuCCT-1) cells encapsulated in 1 % GelPhy/Photo matrix grew and self-assembled into epithelial cysts with lumen, which could not be achieved in a traditional high-concentration gelatin hydrogel. These findings pave the way to significantly improve existing gelatin hydrogels for 3D cell culture applications.

Pair of Functional Polyesters That Are Photo-Cross-Linkable and Electrospinnable to Engineer Elastomeric Scaffolds with Tunable Structure and Properties.

A pair of alkyne- and thiol-functionalized polyesters are designed to engineer elastomeric scaffolds with a wide range of tunable material properties (e.g., thermal, degradation, and mechanical properties) for different tissues, given their different host responses, mechanics, and regenerative capacities. The two prepolymers are quickly photo-cross-linkable through thiol-yne click chemistry to form robust elastomers with small permanent deformations. The elastic moduli can be easily tuned between 0.96 ± 0.18 and 7.5 ± 2.0 MPa, and in vitro degradation is mediated from hours up to days by adjusting the prepolymer weight ratios. These elastomers bear free hydroxyl and thiol groups with a water contact angle of less than 85.6 ± 3.58 degrees, indicating a hydrophilic nature. The elastomer is compatible with NIH/3T3 fibroblast cells with cell viability reaching 88 ± 8.7% relative to the TCPS control at 48 h incubation. Differing from prior soft elastomers, a mixture of the two prepolymers without a carrying polymer is electrospinnable and UV-cross-linkable to fabricate elastic fibrous scaffolds for soft tissues. The designed prepolymer pair can thus ease the fabrication of elastic fibrous conduits, leading to potential use as a resorbable synthetic graft. The elastomers could find use in other tissue engineering applications as well.

Advances in Gelatin Bioinks to Optimize Bioprinted Cell Functions.

Gelatin is a widely utilized bioprinting biomaterial due to its cell-adhesive and enzymatically cleavable properties, which improve cell adhesion and growth. Gelatin is often covalently cross-linked to stabilize bioprinted structures, yet the covalently cross-linked matrix is unable to recapitulate the dynamic microenvironment of the natural extracellular matrix (ECM), thereby limiting the functions of bioprinted cells. To some extent, a double network bioink can provide a more ECM-mimetic, bioprinted niche for cell growth. More recently, gelatin matrices are being designed using reversible cross-linking methods that can emulate the dynamic mechanical properties of the ECM. This review analyzes the progress in developing gelatin bioink formulations for 3D cell culture, and critically analyzes the bioprinting and cross-linking techniques, with a focus on strategies to optimize the functions of bioprinted cells. This review discusses new cross-linking chemistries that recapitulate the viscoelastic, stress-relaxing microenvironment of the ECM, and enable advanced cell functions, yet are less explored in engineering the gelatin bioink. Finally, this work presents the perspective on the areas of future research and argues that the next generation of gelatin bioinks should be designed by considering cell-matrix interactions, and bioprinted constructs should be validated against currently established 3D cell culture standards to achieve improved therapeutic outcomes.