Bio-fabrication of peptide-modified alginate scaffolds: Printability, mechanical stability and neurite outgrowth assessments

Peripheral nerve tissue requires appropriate biochemical and physical cues to guide the regeneration process after injury. Bioprinted peptide-conjugated sodium alginate (PCSA) scaffolds have the potential to provide physical and biochemical cues simultaneously. Such scaffolds need characterisation in terms of printability, mechanical stability, and biological performance to refine and improve application in nerve tissue regeneration. In this study, it was hypothesized that 3D scaffold printed with low concentrated multiple PCSA precursor would be supportive for axon outgrowth. Therefore, a 2% (w/v) alginate precursor was conjugated with either arginine-glycine-aspartate (RGD) or tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptides, or a mixture of RGD and YIGSR (1:2) and was bioprinted in this study. The printability of the composite PCSA scaffolds was tested in three different concentrations of crosslinker (i.e. 50, 100, and 150 mM of CaCl2), and was evaluated by measuring strand width, pore geometry, and angle-formation accuracy. Swelling, degradation, and compression experiments were conducted over a 3 week period to evaluate the mechanical stability of the composite PCSA scaffolds. Scanning electron microscopic (SEM) images were taken to study the surface morphology of the degraded scaffolds. Biological performance was assessed both for single and composite PCSA scaffolds by quantifying the viability and morphology of seeded or incorporated Schwann cells (SCs), amount of secreted brain derived neurotrophic factor (BDNF) by incorporated SCs, and directional neurite outgrowth of neuron cells in a 2D culture. Experimental results suggest that 30 kPa extrusion pressure and 18 mm/s needle speed are suitable to fabricate composite PCSA scaffolds with reasonable strand or pore printability (∼0.95–1.0), and 50 mM CaCl2 facilitated better strand and pore printability than the two other concentrations. Captured SEM images demonstrate that all the composite PCSA scaffolds preserved the initial biofabricated porous structure over 3 weeks, but they lost ∼70% of the initial elastic modulus. In terms of biological performance, composite PCSA scaffolds facilitated better viability and morphology of SCs, as well as supported superior directional neurite outgrowth compared to that of a single PCSA scaffold.