Postmenopausal osteoporosis is caused by the deficiency of estrogen, which breaks bone homeostasis and induces levels of pro-inflammatory cytokines. Muscone is a potent anti-inflammatory agent and is used to treat bone fracture in traditional Chinese medicine. However, its anti-osteoclastogenic effects remain unclear. For in vitro study, morphology tests of osteoclastogenesis were firstly performed. And then, factors in RANK-induced NF-kB and MAPK pathways were examined by RT-PCR and Western blot, and the binding of TNF receptor-associated factor (TRAF)6 to RANK was inspected by coimmunoprecipitation and immunofluorescence staining. For in vivo experiments, C57BL/6 ovariectomized (OVX) mice were used for detection, including H&E staining, TRAP staining, and micro CT. As a result, muscone reduced OVX-induced bone loss in mice and osteoclast differentiation in vitro, by inhibiting TRAF6 binding to RANK, and then suppressed NF-kB and MAPK signaling pathways. The expression of the downstream biomarkers was finally inhibited, including NFATc1, CTR, TRAP, cathepsin K, and MMP-9. The inflammatory factors, TNF-a and IL-6, were also reduced by muscone. Taken together, muscone inhibited the binding of TRAF6 to RANK induced by RANKL, thus blocking NF-kB and MAPK pathways, and down-regulating related gene expression. Finally, muscone inhibited osteoclastogenesis and osteoclast function by blocking RANK-TRAF6 binding, as well as downstream signaling pathways in vitro. Muscone also reduced ovariectomy-induced bone loss in vivo.

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Ankylosing spondylitis (AS) is a chronic, progressive, and inflammatory disease that mainly affects the central axis joint. Although this disease has already been well documented and studied, its pathogenesis is still not well understood. This study aimed to screen and identify key candidate genes involved in the progression of AS. For this purpose, expression profiles of GSE39340 and GSE41038 were downloaded from the Gene Expression Omnibus and displayed in the form of volcano plots and heatmaps. Differentially expressed genes (DEGs) were identified by the Limma package in R and functional enrichment analyses were performed. Moreover, STRING and Cytoscape were utilized to construct protein-protein interaction (PPI) networks and screen significant modules. Immunohistochemistry (IHC) in tissue chips of AS and normal human synovial tissues was performed to confirm the major proteins associated with its development. Western blotting (WB) and alizarin red staining were applied to validate the expression level of platelet-derived growth factor receptor beta (PDGFRB) and function during osteogenesis differentiation of fibroblasts in AS. A total of 256 DEGs were screened, including 191 up-regulated genes and 65 down-regulated genes. The enriched functions of these identified genes mainly included adherens junction, focal adhesion, and cell-substrate adherens junction. The pathways most highly associated with the progression of AS were TGF-β signaling pathway, the Hippo signaling pathway, and the AGE-RAGE signaling pathway. In addition, IHC showed that mitogen-activated protein kinase 1 (MAPK1), C-X-C motif chemokine receptor 4 (CXCR4), and PDGFRB were highly expressed in AS. PDGFRB was found upregulated during osteogenesis of fibroblasts and stimulates osteogenesis in AS. These findings may improve our understanding of the molecular mechanisms controlling AS. Pharmacological targeting of PDGFRB may initiate a possible suppression of bone formation in AS.

When the radial waveform was calibrated with the oscillometric brachial pressures, the SphygmoCor system could not provide accurate estimation of central BPs. The inaccurate measurement of cuff pressure was the major limiting factor for the use of the transfer function in the clinical settings.

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