SERCA2/CaMKIIγ axis in osteocytes regulates RANKL-mediated osteoclastogenesis in response to mechanical loading

Osteocytes are the principal mechanosensory cells in bone and a main source of receptor activator of nuclear factor-κB ligand (RANKL), which promotes osteoclast formation during adult bone remodeling. However, the mechanisms regulating RANKL production in osteocytes, particularly in response to mechanical cues, remain incompletely understood. Sarco/endoplasmic reticulum Ca²⁺-ATPase 2 (SERCA2) and Ca²⁺/calmodulin-dependent protein kinase II (CAMKII) have both been implicated in osteocytic mechanotransduction, but their roles in regulating osteocytic RANKL relatively unknown. This study investigated the role of the osteocytic SERCA2/CAMKIIγ axis in RANKL-mediated osteoclastogenesis and its relationship with mechanical loading. In an ovariectomy (OVX)-induced osteoporosis model, bone loss and increased osteoclast activity were accompanied by reduced SERCA2 and elevated RANKL expression. In vitro, SERCA2 knockdown in MLO-Y4 cells reduced CAMKIIγ phosphorylation, and increased RANKL expression and the osteoclastogenic capacity of co-cultured bone marrow-derived macrophages (BMMs). KN93 inhibition and siRNA knockdown of CAMKIIγ produced similar effects, whereas SERCA2 re-expression restored CaMKIIγ phosphorylation and reduced RANKL expression. Fluid shear stress (FSS) increased SERCA2 expression and CAMKIIγ phosphorylation while decreasing RANKL expression, whereas these responses were attenuated by SERCA2 knockdown. Ca²⁺ chelation altered RANKL expression and diminished its further response to FSS. In vivo, local AAV9-mediated SERCA2 overexpression attenuated trabecular bone loss and osteoclast-related changes in OVX mice but showed limited effects in tail-suspended mice. Collectively, these findings support the involvement of SERCA2/CaMKIIγ signaling in the mechanically responsive regulation of RANKL in MLO-Y4 cells and indicate that local SERCA2 overexpression can alleviate OVX-associated bone loss. SERCA2 may therefore represent a potential target for limiting excessive bone resorption in osteoporosis.