Fabrication of GelMA - Agarose Based 3D Bioprinted Photocurable Hydrogel with In Vitro Cytocompatibility and Cells Mirroring Natural Keratocytes for Corneal Stromal Regeneration.

The complex anatomy of the cornea and the subsequent keratocyte-fibroblast transition have always made corneal stromal regeneration difficult. Recently, 3D printing has received considerable attention in terms of fabrication of scaffolds with precise dimension and pattern. In the current work, 3D printable polymer hydrogels made of GelMA/agarose are formulated and its rheological properties are evaluated. Despite the variation in agarose content, both the hydrogels exhibited G'>G'' modulus. A prototype for 3D stromal model is created using Solid Works software, mimicking the anatomy of an adult cornea. The fabrication of 3D-printed hydrogels is performed using pneumatic extrusion. The FTIR analysis speculated that the hydrogel is well crosslinked and established strong hydrogen bonding with each other, thus contributing to improved thermal and structural stability. The MTT analysis revealed a higher rate of cell proliferation on the hydrogels. The optical analysis carried out on the 14th day of incubation revealed that the hydrogels exhibit transparency matching with natural corneal stromal tissue. Specific protein marker expression confirmed the keratocyte phenotype and showed that the cells do not undergo terminal differentiation into stromal fibroblasts. The findings of this work point to the potential of GelMA/A hydrogels as a novel biomaterial for corneal stromal tissue engineering.

3D bioprinted photo crosslinkable GelMA/methylcellulose hydrogel mimicking native corneal model with enhanced in vitro cytocompatibility and sustained keratocyte phenotype for stromal regeneration.

Corneal transplantation serves as the standard clinical therapy for serious corneal disorders. However, rejection of grafts, significant expenditures, and most crucially, the global donor shortage, may affect the outcome. Recently, 3D bioprinting using biodegradable polymeric materials has become a suitable method for creating tissue replicas with identical architecture. One such most renowned material is GelMA, for its scaffold's three-dimensional structure, biocompatibility, robust mechanics, and favourable optical transmittance. However, GelMA's inadequate viscosity to print at body temperature with better form integrity remains an obstacle. The aim of this work is to create 3D printed GelMA/MC hydrogels for corneal stroma tissue engineering using MC's printability at room temperature and GelMA's irreversible photo cross-linking with UV irradiation. The print speed and pressure conditions for 3D GelMA/MC hydrogels were tuned. Thermal, morphological and physicochemical characteristics were studied for two distinct concentrations of GelMA/MC hydrogels. The hydrogels achieved a transparency of ~78 % (at 700 nm), which was on par with that of the normal cornea (80 %). The in vitro studies conducted using goat corneal stromal cells demonstrated the ability of both hydrogels to promote cell adhesion and proliferation. Expression of Vimentin and keratan sulphate validated the phenotype of keratocytes in the hydrogel. This 3D printed GelMA/MC hydrogel model mimics biophysical characteristics of the native corneal stroma, which may hold promise for clinical corneal stromal tissue engineering.