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Gellan gum (GG) has been proposed for use in tissue engineering (TE) due to its structural and functional similarities with alginate. The most traditional crosslinking methods of GG, ionical and photocrosslinking, have downsides such as loss of stability or phototoxicity, which can limit their use in certain applications. In this study, an alternative hydrazone crosslinking method is introduced. Hydrazone crosslinking is a simple method that produces no toxic reagents or side-products. The method enables the fabrication of injectable hydrogels. GG was combined with hyaluronan (HA) to improve some properties such as cell attachment. The mechanical and physical properties of GG-HA hydrogels were controlled by changing the molecular weight, the degree of modification, and the ratio of polymer components. GG-HA hydrogels showed ionic nature of deswelling in the presence of cations enabling the control of physical properties in different solution environments. Due to the non-linear elastic behavior of hydrogels and tissues, the stiffness as a function of strain was represented instead of solely giving the second-order elastic constants. The stiffness of GG-HA hydrogels was similar to that of soft tissues at small strains. [ABSTRACT FROM AUTHOR]

Abstract Microstructure plays an essential role in the control of hydrogel properties. It is also an important factor when cells or drugs are encapsulated inside the hydrogel. In this work, the microstructures of hydrazone crosslinked hyaluronan-, alginate- and gellan gum-based hydrogels were evaluated thoroughly for the first time by using rheology- and diffusion (fluorescence recovery after photobleaching, FRAP)-based methods. The effect of gel parameters on the viscoelastic and diffusion properties of hydrogels, and further on their structural parameters (mesh size, average molecular weight of the polymer chain between neighboring crosslinks, crosslinking density) are shown. Results further show that diffusivity decreased when larger dextran sizes were used, which were equivalent to the mesh sizes of hydrogels (15 nm to 47 nm) evaluated by the rheological method. This mesh size range allows the transportation of smaller molecules, but also peptides and most of the proteins. A correlation between the storage modulus and the structural parameters was also shown. Overall, hydrazone crosslinking offers an easy way to produce polysaccharide-based hydrogels with variable microstructures by altering the gel parameters. Graphical Abstract Highlights • Microstructure of hydrogels can be determined using rheology- and FRAP-based methods. • The effect of material parameters on viscoelastic and diffusion properties is shown. • The higher the G', the lower the mesh size and M c , and the higher the crosslinking density. • The diffusivity decreases when larger dextran sizes (500 kDa and 2000 kDa) are used. • There is an equivalency between the FRAP results and mesh sizes evaluated by rheology. [ABSTRACT FROM AUTHOR]

Corneal blindness is a worldwide problem, plagued by insufficient amount of high-quality donor tissue. Cell therapy using human adipose stem cells (hASCs) has risen as an alternative to regenerate damaged corneal stromal tissue, the main structural and refractive layer of the cornea. Herein we propose a method to deliver hASCs into corneal defects in hyaluronan (HA)-based hydrogels, which form rapidly in situ by hydrazone crosslinking. We fabricated two different HA-based hydrazone-crosslinked hydrogels (HALD1-HACDH and HALD2-HAADH), and characterized their swelling, degradation, mechanical, rheological and optical properties and their ability to support hASC survival. To promote hASC attachment and survival, we incorporated collagen I (col I) to the more stable HALD1-HACDH hydrogel, since the HALD2-HAADH hydrogel suffered swift degradation in culture conditions. We then used an organ culture model with excised porcine corneas to study the delivery of hASCs in these three hydrogels for stromal defect repair. Although all hydrogels showed good hASC survival directly after encapsulation, only the collagen-containing HALD1-HACDH-col I hydrogel showed cells with elongated morphology, and significantly higher cell metabolic activity than the HALD1-HACDH gel. The addition of col I also increased the stiffness and reduced the swelling ratio of the resulting hydrogel. Most importantly, the corneal organ culture model demonstrated these hydrogels as clinically feasible cell delivery vehicles to corneal defects, allowing efficient hASC integration to the corneal stroma and overgrowth of corneal epithelial cells. [ABSTRACT FROM AUTHOR]