Induced pluripotent stem cell-derived neural stem cells promote bone formation in mice with calvarial defects

Nerve-derived factors have attracted attention in bone regeneration therapy due to their ability to promote bone regeneration and nerve innervation. Mesenchymal stem cells transported to target sites promote osteogenesis. However, there are few reports on the effects of neural stem cells on bone regeneration. Therefore, the aim of this study was to investigate the role of neural stem cells in osteogenesis. Here, embryoid bodies (EB) or primary neurospheres (1NS) were generated using mouse induced pluripotent stem cells (iPS cells), which were then seeded onto gelatin (Gel) sponges. The seeded Gel sponges were then transplanted into mouse calvarial bone defects. We noted that 1NS-seeded Gel promoted bone regeneration and the presence of tartrate-resistant acid phosphatase (TRAP)-positive cells, whereas the EB-seeded Gel did not. RNA-sequencing of the 1NS-seeded and EB seeded Gels showed an upregulation of the transforming growth factor (TGF)-β signaling pathway in the 1NS-seeded Gel group. Immunostaining confirmed the presence of Id3 positive cells in mice with bone defects treated with the 1NS-seeded Gel. These findings suggest that the transplantation of neural stem cells may contribute to the promotion of bone regeneration. STATEMENT OF SIGNIFICANCE: This study aimed to investigate whether neural stem cells, when seeded in Gel sponges, promoted bone regeneration. It has been well documented that bone is tightly linked with the nervous systems. Bioscaffolds comprising factors that promote innervation and bone regeneration have been investigated for use in bone therapy. However, there is limited research on the use of neural stem cells for promoting bone formation. To assess this relationship, we conducted both in vivo and in vitro assays to determine whether neural stem cells promoted bone formation. We noted that 1NS-seeded Gel sponges promoted bone formation significantly in mice with calvarial defects after 4 weeks. This study provides a novel approach of neural stem cells for bone therapy.