A tuned gelatin methacryloyl (GelMA) hydrogel facilitates myelination of dorsal root ganglia neurons in vitro

Abstract

Investigating axonal myelination by Schwann cells (SCs) is crucial for understanding mechanisms underlying demyelination and remyelination, which may help gain insights into incurable disorders like neurodegenerative diseases. In this study, a gelatin-based hydrogel, gelatin methacryloyl (GelMA), was optimized to achieve the biocompatibility, porosity, mechanical stability, and degradability needed to provide high cell viability for dorsal root ganglia (DRG) neurons and SCs, and to enable their long-term coculture needed for myelination studies. The results of cell viability, neurite elongation, SC function and maturation, SC-axon interaction, and myelination were compared with two other commonly used substrates, namely collagen and Poly-d Lysine (PDL). The tuned GelMA constructs (Young's modulus of 32.6 ± 1.9 kPa and the median value of pore size of 10.3 μm) enhanced single axon generation (unlike collagen) and promoted the interaction of DRG neurons and SCs (unlike PDL). While DRG cells exhibited relatively higher viability on PDL after 48 h, i.e., 83.8%, the cells had similar survival rate on GelMA and collagen substrates, 66.7% and 61.5%, respectively. Further adjusting the hydrogel properties to achieve two distinct ranges of relatively small and large pores supported SCs to extend their processes freely and enabled physical contact with and wrapping around their corresponding axons. Staining the cells with myelin basic protein (MBA) and myelin-associated glycoprotein (MAG) revealed enhanced myelination on GelMA hydrogel compared to PDL and collagen. Moreover, the engineered porosity enhanced DRGs and SCs attachments and flexibility of movement across the substrate. This engineered hydrogel structure can now be further explored to model demyelination in neurodegenerative diseases, as well as to study the effects of various compounds on myelin regeneration.