Piezo2⁺ Mechanosensory Neurons Orchestrate Heterotopic Ossification in Achilles Tendon

Heterotopic ossification (HO) of the Achilles tendon induces pain and impairs tendon mechanical function, yet its pathogenic mechanisms remain largely unknown. Given that neurotransmitters, inflammatory cytokines, and mechanical loading are highly associated with HO incidence, we raised a hypothesis that activation of sensory neurons within the injured Achilles tendon may be one critical cause driving HO progression. This present work demonstrates that Netrin-1, predominantly secreted by macrophages at the injured site, profoundly promotes the regeneration of Piezo2-expressing sensory nerves in the Achilles tendon of mice after puncture. This is accompanied by an aberrant rise in the pro-osteogenic neuropeptide calcitonin gene-related peptide (CGRP) in both sensory nerves at the injury site and the dorsal root ganglion (DRG), particularly under the synergistic influence of mechanical stimulation and prostaglandin E2 (PGE2), a pro-inflammatory cytokine initially produced by macrophages. Consequently, osteogenic differentiation of tendon-derived stem cells (TDSCs) is enhanced, which in turn further stimulates PGE2 secretion from osteoblasts. Of note, such an effect and subsequent HO formation can be significantly attenuated by tail suspension treatment or by blocking signaling pathways mediated by CGRP, Netrin-1, PGE2, or Piezo2. Together, these findings elucidate the crosstalk among macrophages, osteoblasts, and Piezo2⁺ sensory neurons in the injured Achilles tendon, highlighting the critical role of sensory hyperinnervation in promoting HO progression.