Objective The aim of this study was to identify the role of Piezo1-mediated mechanotransduction in entheseal pathological new bone formation and to explore the underlying molecular mechanism.

Methods Spinal ligament tissues were collected from 14 patients with ankylosing spondylitis (AS) and 14 non-AS controls and bulk RNA sequencing was conducted. Collagen antibody-induced arthritis models were established to observe pathological new bone formation. Pharmacological inhibition and genetic ablation of Piezo1 was performed in animal models to identify the essential role of Piezo1. Entheseal osteo-chondral lineage cells were collected and in vitro cell culture system was established to study the role and underlying mechanism of Piezo1 in regulation of chondrogenesis, osteogenesis and its own expression.

Results Piezo1 was aberrantly upregulated in ligaments and entheseal tissues from patients with AS and animal models. Pharmaceutical and genetic inhibition of Piezo1 attenuated while activation of Piezo1 promoted pathological new bone formation. Mechanistically, activation of CaMKII (Calcium/calmodulin dependent protein kinase II) signalling was found essential for Piezo1-mediated mechanotransduction. In addition, Piezo1 was upregulated by AS-associated inflammatory cytokines.

Conclusion Piezo1-mediated mechanotransduction promotes entheseal pathological new bone formation through CaMKII signalling in AS.