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Tuning hydrogel properties through minor modifications of the crosslinker structure is a beneficial approach for hydrogel design that could result in hydrogels with wide range of properties to match a desired application.

Liver extracellular matrix (ECM) composition, topography and biomechanical properties influence cell-matrix interactions. The ECM presents guiding cues for hepatocyte phenotype maintenance, differentiation and proliferation both in vitro and in vivo. Current understanding of such cell-guiding cues along with advancement of techniques for scaffold fabrication has led to evolution of matrices for liver tissue culture from simple porous scaffolds to more complex 3D matrices with microarchitecture similar to in vivo. Natural and synthetic polymeric biomaterials fabricated in different topographies and porous matrices have been used for hepatocyte culture. Heterotypic and homotypic cell interactions are necessary for developing an adult liver as well as an artificial liver. A high oxygen demand of hepatocytes as well as graded oxygen distribution in liver is another challenging attribute of the normal liver architecture that further adds to the complexity of engineered substrate design. A balanced interplay of cell-matrix interactions along with cell-cell interactions and adequate supply of oxygen and nutrient determines the success of an engineered substrate for liver cells. Techniques devised to incorporate these features of hepatic function and mimic liver architecture range from maintaining liver cells in mm-sized tailor-made scaffolds to a more bottoms up approach that starts from building the microscopic subunit of the whole tissue. In this review, we discuss briefly various biomaterials used for liver tissue engineering with respect to design parameters such as scaffold composition and chemistry, biomechanical properties, topography, cell-cell interactions and oxygenation.

To create an ideal graft substitute for regenerating bone, the scaffold should possess osteoconductive, osteoinductive, and osteogenic properties. Hydrogels are a very common scaffold, but the mechanical integrity and nanoporous structure are not advantageous for bone regeneration. Cryogelation is a technique in which the controlled freezing and thawing of a polymer creates a spongy, macroporous structure with ideal structural characteristics and promising mechanical stability. Hydrogels and cryogels of three different materials (chitosan-gelatin, N-vinyl-2-pyrrolidone, and silk fibroin (SF)) were compared to assess the optimal material and form of scaffold for this application. Cryogel and hydrogel structures were tested in parallel to evaluate porosity, swelling, mechanical integrity, cellular infiltration, and mineralization potential. Cryogels proved superior to hydrogels based on swelling potential and mechanical properties. Among the cryogels, SF demonstrated high pore diameter and area, mineralization upon cellular infiltration, and the largest presence of osteocalcin, a marker of bone formation. These results demonstrate the practicality of cryogels for a bone regeneration application and identify SF as a potential material choice.

Electrohydrodynamic spraying (EHS) has recently gained popularity for microencapsulation of cells for applications in cell delivery and tissue engineering. Some of the polymers compatible with EHS are alginate, chitosan, and other similar natural polymers, which are subject to ionotropic or physical gelation. It is desirable to further extend the use of the EHS technique beyond such polymers for wider biofabrication applications. Here, building upon our previous work of making PEG microspheres via EHS, we utilized the principles of EHS to fabricate cell-laden polyethylene glycol (PEG) hydrogel microspheres. The gelation of PEG hydrogel microspheres was achieved by forming covalent crosslinks between multiarm PEG acrylate and dithiol crosslinkers via Michael-type addition. We conducted a detailed investigation of the critical parameters of EHS, such as the applied voltage, inner needle diameter (i.d. needle), and flow rate, to obtain PEG microspheres with high cell viability and tightly-controlled diameters in the range of 70-300 μm. The polydispersity of cell-laden PEG hydrogel microspheres as measured by % coefficient of variation was between 6% and 23% for all conditions tested. We established that our method was compatible with different cell types and that all tested cell types could be encapsulated at high densities of 10-10 and ≥90% encapsulation efficiency. We observed cell aggregation within the hydrogel microspheres at applied voltage >5 kV. Since PEG is a synthetic polymer devoid of cell attachment sites, we could overcome this limitation by tethering Arg-Gly-Asp-Ser (RGDS) peptide to the PEG hydrogel microspheres; upon RGDS tethering, we observed uniform cell dispersion. The microencapsulated cells could be cultured in the PEG hydrogel microspheres of different sizes for up to one week without significant loss in cell viability. In conclusion, the EHS technique developed here could be used to generate cell-laden PEG hydrogel microspheres of controlled sizes for potential applications in cell delivery and organoid cultures.

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In this study, the potential of cryogel bilayer wound dressing and skin regenerating graft for the treatment of surgically created full thickness wounds was evaluated. The top layer was composed of polyvinylpyrrolidone-iodine (PVP-I) cryogel and served as the antiseptic layer, while the bottom regenerative layer was made using gelatin cryogel. Both components of the bilayer showed typical features of a cryogel interconnected macropore network, rapid swelling, high water uptake capacity of about 90%. Both PVP and gelatin cryogel showed high tensile strength of 45 and 10 kPa, respectively. Gelatin cryogel sheets were essentially elastic and could be stretched without any visible deformation. The antiseptic PVP-I layer cryogel sheet showed sustained iodine release and suppressed microbial growth when tested with skin pathogens (zone of inhibition ∼2 cm for sheet of 0.9 cm diameter). The gelatin cryogel sheet degraded in vitro in weeks. The gelatin cryogel sheet supported cell infiltration, attachment, and proliferation of fibroblasts and keratinocytes. Microparticles loaded with bioactive molecules (mannose-6-phosphate and human fibrinogen) were also incorporated in the gelatin cryogel sheets for their role in enhancing skin regeneration and scar free wound healing. In vivo evaluation of healing capacity of the bilayer cryogel was checked in rabbits by creating full thickness wound defect (diameter 2 cm). Macroscopic and microscopic observation at regular time intervals for 4 weeks demonstrated better and faster skin regeneration in the wound treated with cryogel bilayer as compared to untreated defect and the repair was comparable to commercial skin regeneration scaffold Neuskin-F. Complete skin regeneration was observed after 4 weeks of implantation with no sign of inflammatory response. Defects implanted with cryogel having mannose-6-phosphate showed no scar formation, while the wound treated with bilayer incorporated with human fibrinogen microparticles showed early signs of skin regeneration; epidermis formation occurred at 2 weeks after implantation.