Optic nerve sheath meningiomas are the second common optic nerve tumor that affect the optic nerve sheath of primarily middle-aged individuals. In general, invasive measures are avoided, as in most situations, surgery yields no good outcome and is often only considered for blind individuals or those with severe proptosis. Regeneration of the optic nerve fibres bearing newly functional sheaths with the aid of tissue engineering to replace the worn out nerve fibres, could be a long-term solution. Therefore, to regenerate these nerve fibres, it is important to first produce essential tools such as the cell-laden core filaments in vitro. The current research was carried out to produce cell-laden core filaments by coaxially bioprinting a hyaluronic acid-tyramine (HA-TA) bioink using enzymatic crosslinking.
In this study, bovine primary chondrocytes were isolated and cultured. The confluent cell pellets were combined with tyramine substituted hyaluronic acid of high molecular weight (HMW HA-TA) to form the bioink. This cell-laden bioink was extruded through a coaxial nozzle of 22/16G in a core/shell printing set-up to print the cell-laden core filaments. To improve the gelation mechanism used during the coaxial bioprinting process, biocompatible near-instantaneous enzymatic crosslinking, based on the enzyme horseradish peroxidase (HRP), was explored to yield cell-laden core filaments. The extruded cell-laden core filaments were cultured for up to 7 days in vitro. In addition, the physical properties such as rheological properties, swelling ratio and degradation of the biomaterial (HMW HA-TA) and corresponding hydrogels were examined.
HMW HA-TA was synthesized successfully through DMTMM-mediated amidation. HA-TA with 3 different tyramine substitution degrees (DS) were prepared and analysed. The functional biomaterial was assessed and proven through the formation of gels upon HRP-mediated crosslinking of the phenolic moieties. The gelation time for the enzymatic crosslinking of HMW HA-TA at 3 different percentages (1.3, 1.8 or 2.2 %w/v) with 5.5 U/ml HRP was instantaneous. These cell encapsulated core filaments were printed in the presence of a sacrificial ink sheet based on Pluronic F127. The physical properties of the resulting hydrogel were analysed; the swelling ratio decreased with increasing DS and increased with increasing polymer concentration. The enzymatic degradation rate of the HA hydrogel was inversely proportional to the concentration of polymer and DS.
We developed a near-instantaneous crosslinking HMW HA-TA bioink for the production of cell-laden filaments by core/shell bioprinting, which can be used for nerve fibres’ regeneration in the future.