Piezo1 Mechanosensor Downregulates Osteoclastogenesis by Suppressing the AKT/GSK3β Pathway

Objectives: Dysregulated osteoclastogenesis is implicated in bone lytic diseases, such as periodontitis. It is well established that osteoclastogenesis is induced by RANKL-mediated activation of NFATc1 transcription factor which, in turn, elicits the expression of genes required for osteoclast functions. Although emerging evidence supports that mechanosensitive Ca²⁺-channel Piezo1 plays a pivotal role in bone regeneration, possible engagement of Piezo1 in osteoclastogenesis and bone resorption in periodontitis is largely unknown. Thus, we sought to establish the role of Piezo1 in osteoclast differentiation and function.
Methods: RAW 264.7 cells stimulated with or without RANKL (10ng/mL), in the presence or absence of Yoda1 (2.5μM, Piezo1 activator), were subject to Ca2+ influx assay. Fluorescent confocal microscopy and qPCR were employed to monitor the Piezo1 expression pattern in RAW 264.7 cell as well as to evaluate OC-STAMP, TRAP (ACP5) and MMP9 mRNA expression, respectively. To evaluate the formation of multinucleated mature osteoclasts and bone resorptive activity, TRAP staining and pit formation assays were performed. The phosphorylation status of AKT and GSK3β was determined by Western blot.
Results: In RANKL-primed RAW 264.7 cells (Day 2), Yoda1 significantly promoted Ca2+ influx (P<0.05) but significantly suppressed the expression of osteoclast-related genes, OC-STAMP, MMP9 and ACP5 (P<0.05). Moreover, the number and size of multi-nucleated TRAP-positive cells as well as the formation of pits were also reduced significantly by treatment with Yoda1 (P<0.05). RANKL upregulated the phosphorylation of AKT and GSK3β in RAW264.7 cells which was abrogated by the presence of Yoda1 (P<0.05).
Conclusions: The present study demonstrated that the activation of Piezo1 decreases osteoclast differentiation and activity by inhibiting OC-STAMP, TRAP and MMP9 mRNA expression, while suppressing the AKT/GSK3β signaling axis, suggesting that mechanosensitive Ca²⁺-channel Piezo1 may play a gatekeeper role in restraining over-activation of osteoclastogenesis and function to resorb bone in the context of periodontitis.