Deer antler polypeptides promote osteogenesis for fracture healing via activating neurosensory PGE2/EP4/p-CREB axis.

BACKGROUND

Deer antler polypeptides (DAPs), one of the most bioactive components of deer antler, have been reported to possess multiple biological functions. However, the role of DAPs in fracture healing remains unclear. This study aims to investigate the therapeutic effects of DAPs on fracture healing and elucidate their potential pharmacological mechanism.

METHODS

LC-MS/MS was employed to analyze the peptide compositions and serum metabolites of DAPs. Sensory nerve-specific EP4 receptor knockout (EP4 Avil ) mice, sensory nerve- and skeletal stem/progenitor cells-specific fluorescence reporter mice were subjected to a transverse tibial fracture followed by oral administration of DAPs at 50 mg/kg/day. In vitro, primary bone marrow cells (BMCs) were incubated with serum containing DAPs, and dorsal root ganglion (DRG) neurons were subsequently treated with the conditioned medium harvested from these BMCs. A series of experimental approaches including gait analysis, μCT, three-point bending test, calcein double-labeling staining, IHC, IF, ELISA, qRT-PCR and western blotting were performed on both in vivo and in vitro models.

RESULTS

LC-MS/MS analysis identified 303 functional peptides and 32 differential metabolites. DAPs substantially accelerated mouse fracture healing by enhancing callus formation. PGE2 production was significantly increased in the fracture callus of DAPs-treated mice and in BMCs stimulated with DAPs-containing serum, which activated the EP4 receptor in sensory nerves and DRG neurons in vivo and in vtiro. This activation consequently improved hypothalamic CREB phosphorylation and diminished sympathetic tone, thereby promoting the osteogenic differentiation of SSPCs. However, DAPs-mediated osteogenesis in the fracture callus was abolished in EP4 Avil mice due to the blockade of neurosensory PGE2/EP4/p-CREB signaling.

CONCLUSION

This study demonstrate that DAPs enhance fracture healing by promoting osteogenesis via neurosensory PGE2/EP4/p-CREB axis.

THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: Deer antler polypeptides (DAPs) are the primary bioactive components of deer antler, a self‑regenerative mammalian organ. This study reveals that DAPs can promote fracture healing in mice through activation of the neurosensory PGE2/EP4/p‑CREB axis. These findings provide a novel bioactive candidate for enhancing bone repair, which holds translational potential for developing therapeutics to treat fractures or other bone defects in clinical practice". at the end of the abstract as it should be included as part of the abstract.