3D culture of the spinal cord with roots as an ex vivo model for comparative studies of motor and sensory nerve regeneration

Abstract

Motor and sensory nerves exhibit tissue-specific structural and functional features. However, in vitro models designed to reflect tissue-specific differences between motor and sensory nerve regeneration have rarely been reported. Here, by embedding the spinal cord with roots (SCWR) in a 3D hydrogel environment, we compared the nerve regeneration processes between the ventral and dorsal roots. The 3D hydrogel environment induced an outward migration of neurons in the gray matter of the spinal cord, which allowed the long-term survival of motor neurons. Tuj1 immunofluorescence labeling confirmed the regeneration of neurites from both the ventral and dorsal roots. Next, we detected asymmetric ventral and dorsal root regeneration in response to nerve growth factor (NGF) and glial cell line–derived neurotrophic factor (GDNF), and we observed motor and sensory Schwann cell phenotypes in the regenerated ventral and dorsal roots, respectively. Moreover, based on the SCWR model, we identified a targeted effect of collagen VI on sensory nerve fasciculation and characterized the protein expression profiles correlating to motor/sensory-specific nerve regeneration. These results suggest that the SCWR model can serve as a valuable ex vivo model for comparative study of motor and sensory nerve regeneration and for pharmacodynamic evaluations.