Abnormal subchondral bone remodeling plays a pivotal role in the progression of osteoarthritis (OA). Here, we analyzed subchondral bone samples from OA patients and observed a significant upregulation of ubiquitin carboxy-terminal hydrolase L1 (UCHL1) specifically in subchondral bone osteoclasts. Notably, we found a strong correlation between UCHL1 expression and osteoclast activity in the subchondral bone during OA progression in both human and murine models. Conditional UCHL1 deletion in osteoclast precursors exacerbated OA progression, while its overexpression, mediated by adeno-associated virus 9, alleviated this process in male mice. Mechanistically, RANKL stimulates UCHL1 expression in osteoclast precursors, subsequently stabilizing CD13, augmenting soluble CD13 (sCD13) release, and triggering an autocrine inhibitory effect on the MAPK pathway, thereby suppressing osteoclast formation. These findings unveil a previously unidentified negative feedback loop, RANKL-UCHL1-sCD13, that modulates osteoclast formation and presents a potential therapeutic target for OA.

The C-X-C motif chemokine ligand 2 (CXCL2), a member of the CXC receptor ligand family, is involved in various immune and inflammatory processes, but its effect(s) on bone formation have not yet been reported. We report here that CXCL2 is enriched in bone marrow and show abundant expression of CXCL2 in osteoblasts of osteoporotic mice. CXCL2 neutralization within the bone marrow by using antibody alleviated bone loss in mice, indicating a negative role of CXCL2 in bone formation. In line with this, CXCL2 overexpression attenuated proliferation, as well as differentiation, of osteoblasts in vitro. By contrast, CXCL2 downregulation promoted osteoblast expansion and differentiation. Mechanistically, CXCL2 inhibits the ERK1/2 (MAPK3/1) signaling pathway in osteoblasts. Activation of ERK1/2 abolishes the inhibitory effect of CXCL2 in osteoblasts, whereas inactivation of ERK1/2 reverses the osteogenic role of CXCL2 inhibition. These results show that CXCL2 attenuates osteoblast differentiation through inhibition of the ERK1/2 signaling pathway. We demonstrate here that CXCL2 is a negative regulator of bone formation and clarify the responsible mechanisms. Therefore, pharmaceutical coordination of CXCL2 and of the pathways through which it is regulated in osteoblasts might be beneficial regarding bone formation.

Senescence impairs preosteoblast expansion and differentiation into functional osteoblasts, blunts their responses to bone formation-stimulating factors and stimulates their secretion of osteoclast-activating factors. Due to these adverse effects, preosteoblast senescence is a crucial target for the treatment of age-related bone loss; however, the underlying mechanism remains unclear. We found that mTORC1 accelerated preosteoblast senescence in vitro and in a mouse model. Mechanistically, mTORC1 induced a change in the membrane potential from polarization to depolarization, thus promoting cell senescence by increasing Ca²⁺influx and activating downstream NFAT/ATF3/p53 signaling. We further identified the sodium channel Scn1a as a mediator of membrane depolarization in senescent preosteoblasts. Scn1a expression was found to be positively regulated by mTORC1 upstream of C/EBPα, whereas its permeability to Na⁺was found to be gated by protein kinase A (PKA)-induced phosphorylation. Prosenescent stresses increased the permeability of Scn1a to Na⁺by suppressing PKA activity and induced depolarization in preosteoblasts. Together, our findings identify a novel pathway involving mTORC1, Scn1a expression and gating, plasma membrane depolarization, increased Ca²⁺influx and NFAT/ATF3/p53 signaling in the regulation of preosteoblast senescence. Pharmaceutical studies of the related pathways and agents might lead to novel potential treatments for age-related bone loss.

Introduction: Estrogen deficiency leads to bone loss in postmenopausal osteoporosis, because bone formation, albeit enhanced, fails to keep pace with the stimulated osteoclastic bone resorption. The mechanism driving this uncoupling is central to the pathogenesis of postmenopausal osteoporosis, which, however, remains poorly understood. We previously found that Cxcl9 secreted by osteoblasts inhibited osteogenesis in bone, while the roles of Cxcl9 on osteoclastic bone resorption and osteoporosis are unclear. Materials and Methods: Postmenopausal osteoporosis mouse model was established by bilateral surgical ovariectomy (OVX). In situ hybridization was performed to detect Cxcl9 mRNA expression in bone. ELISA assay was conducted to assess Cxcl9 concentrations in bone and serum. Cxcl9 activity was blocked by its neutralizing antibody. Micro-CT was performed to determine the effects of Cxcl9 neutralization on bone structure. Cell Migration and adhesion assay were conducted to evaluate the effects of Cxcl9 on osteoclast activity. TRAP staining and Western blot were performed to assess osteoclast differentiation. CXCR3 antagonist NBI-74,330 or ERK antagonist SCH772984 was administered to osteoclast to study the effects of Cxcl9 on CXCR3/ERK signaling. Results: Cxcl9 was expressed and secreted increasingly in OVX mice bone. Neutralizing Cxcl9 in bone marrow prevented bone loss in the mice by facilitating bone formation as well as inhibiting bone resorption. In vitro, Cxcl9 secreted from osteoblasts facilitated osteoclast precursors adhesion, migration and their differentiation into mature osteoclasts. The positive role of osteoblastic Cxcl9 on osteoclasts was eliminated by blocking CXCR3/ERK signaling in osteoclasts. Estrogen negatively regulated Cxcl9 expression and secretion in osteoblasts, explaining the increased Cxcl9 concentration in OVX mice bone. Conclusion: Our study illustrates the roles of Cxcl9 in inhibiting bone formation and stimulating bone resorption in osteoporotic bone, therefore providing a possible therapeutic target to the treatment of postmenopausal osteoporosis.

Chemokine C-C motif ligand 7 (CCL7) is implicated in various immune and inflammatory processes; however, its role in rheumatoid arthritis (RA) remains unclear. In this study, we observed that CCL7 expression was upregulated in synovial M1-polarized macrophages and in the serum of RA mice and patients. CCL7 was found to promote macrophage polarization toward the M1 phenotype while inhibiting M2 differentiation in vitro. Furthermore, intra-articular injection of recombinant CCL7 protein in mice resulted in enhanced M1 polarization, increased inflammation, and fibrosis within synovial tissues, which exacerbated arthritis-associated pain. These effects were partially mitigated by treatment with a CCL7 neutralizing antibody. Mechanistically, we identified a CCL7 autocrine positive feedback loop that amplifies inflammation via the CCL7-CCR1-JAK2/STAT1 pathway. Collectively, our findings reveal a previously unrecognized CCL7-mediated autocrine inflammatory amplification loop that modulates macrophage polarization and exacerbates RA progression, positioning CCL7 as a potential therapeutic target for RA.