Neuronal differentiation and inhibition of glial differentiation of murine neural stem cells by pHPMA hydrogel for the repair of injured spinal cord

Currently, several therapeutic methods of treating the effects of spinal cord injury (SCI) are being considered. On the one hand, transplantation of stem cells (SCs), in particular, neural stem/progenitor cells (NSPCs), is promising, as these cells have the potential to differentiate into nervous tissue cells, able to enhance endogenous regeneration and prevent the development of inflammatory processes. On the other hand, it is quite promising to replace the damaged nervous tissue with synthetic matrices, in particular hydrogels, which can create artificial conditions for the regenerative growth of injured nerve fibers through the spinal cord injury area, i.e. stimulate and support axonal regeneration and myelination. In this work, we combined both of these novel approaches by populating (injecting or rehydrating) a heteroporous pHPMA hydrogel (NeuroGel) with murine hippocampal NSPCs. Being inside the hydrogel (10 days of cultivation), NSPCs were more differentiated into neurons: 19.48% ± 1.71% (the NSPCs injection into the hydrogel) and 36.49% ± 4.20% (the hydrogel rehydration in the NSPCs suspension); in control cultures, the level of differentiation in neurons was only 2.40% ± 0.31%. Differentiation of NSPCs into glial cells, in particular into oligodendrocyte progenitor cells, was also observed – 8.89% ± 2.15% and 6.21% ± 0.80% for injection and rehydration variants, respectively; in control – 28.75% ± 2.08%. In the control NSPCs culture, there was a small number of astrocytes – 2.11% ± 0.43%. Inside the hydrogel, NSPCs differentiation in astrocytes was not observed. In vitro data showed that the hydrogel promotes the differentiation of NSPCs into neurons, and inhibits the differentiation into glial cells. And in vivo showed post-traumatic recovery of rat spinal cord tissue after injury followed by implantation of the hydrogel+NSPCs complex (approximately 7 months after SCI). The implant area was closely connected with the recipient tissue, and the recipient cells freely grew into the implant itself. Inside the implant, a formed dense neuronal network was visible. In summary, the results are primarily an experimental ground for further studies of implants based on pHPMA hydrogel with populated different origin SCs, and the data also indicate the feasibility and efficiency of using an integrated approach to reduce possible negative side effects and facilitate the rehabilitation process after a SCI. •Neural stem cells, NeuroGel, differentiation, implanta