3D bioprinted neural construct for the in vitro modeling of spinal cord

INTRODUCTION: Spinal cord injury is a debilitating neurological condition that leads to permanent neural damage with consequent impairment of sensory and locomotor functions. Unfortunately, effective treatments are not yet available for a successful bridging of the injured segment and complete recovery of nervous functionality. Indeed, most of the experimental drugs are proven to be unsatisfactory during clinical trials, partially due to the weakness of actual preclinical models in replicating the pathophysiology of the central nervous system (CNS). In this context, 3D bioprinting technology has gained increasing attention in neural research, as it allows the creation of complex engineered structures which mimic the neural microenvironment. This work aims to develop a three‐dimensional neural construct for the in vitro modeling of the spinal cord. Specifically, the optimization of biomimetic photo‐crosslinkable hydrogels has been reported, intended for the bioprinting of the most represented cell phenotypes found in the spinal cord and the establishment of a multicellular 3D culture system. METHODS: Gelatin Methacryloyl and Thiolated Hyaluronic Acid were selected as the main constituents of the hydrogels because of their high flexibility and suitability for the printing process. Different concentrations of the two polymers were investigated to obtain a fast gelation rate and improve hydrogel extrudability in cell‐friendly conditions. Hydrogel characterization was conducted in terms of rheological properties and printability. In addition, preliminary in vitro studies were performed by encapsulating a motor neuron cell line (NSC‐34) and a neural stem cell line (NE‐4C) into specific hydrogel compositions. Lastly, the cell‐loaded hydrogels were printed, and the obtained constructs were cultivated for up to seven days. RESULTS: Rheological studies showed that the formulated hydrogels display shear‐thinning properties and the presence of a rapid sol‐gel transition after visible light exposure. According to morphological analysis, the printed constructs exhibited regular and well‐defined geometry and macroporosity, thus confirming hydrogel extrudability and shape fidelity. Moreover, in vitro assays demonstrated bioink cytocompatibility and revealed that the printing process does not affect cell viability and behavior. DISCUSSION & CONCLUSIONS: These results suggest that the optimized bioinks may be potential candidates for the bioprinting of a neural‐like construct with high‐resolution and good structural stability. Future investigations will be focused on the inclusion of topographical and biochemical cues to directly induce the differentiation of encapsulated cells into CNS cell subtypes.