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Inflammatory osteolysis is often caused by the excessive activation of osteoclasts stimulated by bacterial products such as lipopolysaccharide. The natural flavonoid trifolirhizin (TRI) has anti‐inflammatory properties; however, its function in inflammatory bone lysis remains unclear. This study aimed to elucidate the potential regulatory mechanisms of TRI in osteoclasts.Tartrate‐resistant acid phosphatase (TRAP) staining, acid secretion assays, podosomal actin belt fluorescence staining, and bone resorption assays were used to investigate the effects of TRI on osteoclast differentiation and bone resorption. A reactive oxygen species (ROS) measurement kit was used to detect the effect of TRI on ROS levels in osteoclasts. The effects of TRI on genes and signaling pathways related to osteoclast differentiation were determined by quantitative polymerase chain reaction (qPCR) and western blotting. A mouse model of lipopolysaccharide‐mediated inflammatory osteolysis was established, and the effects of TRI treatment on bone mass were observed using micro‐CT and histological examination. Mechanistically, TRI reduced ROS production by inhibiting receptor activator of nuclear factor‐κB ligand (RANKL)‐induced activation of the nuclear factor‐κB (NF‐κB) and mitogen‐activated protein kinase (MAPK) signaling pathways, and by upregulating the expression levels of the anti‐ROS enzymes heme oxygenase‐1 (HO‐1) and catalase (CAT), which contributed to the degradation of ROS, ultimately leading to a decrease in osteoclastogenesis. TRI inhibited osteoclast formation and ameliorated lipopolysaccharide (LPS)‐mediated inflammatory osteolysis. Thus, TRI may be a candidate agent for anti‐inflammatory osteolysis. [ABSTRACT FROM AUTHOR]

Background and Aim: Osteoporosis, a systemic metabolic bone disease, is characterized by the decline of bone mass and quality due to excessive osteoclast activity. Currently, drug‐targeting osteoclasts show promising therapy for osteoporosis. In this study, we investigated the effect of cichoric acid (CA) on receptor activator of nuclear kappa‐B ligand (RANKL)‐induced osteoclastogenesis and the bone loss induced by ovariectomy in mice. Experimental Procedure: Molecular docking technologies were employed to examine the interaction between CA and RANKL. CCK8 assay was used to evaluate the cell viability under CA treatment. TRAcP staining, podosome belt staining, and bone resorption assays were used to test the effect of CA on osteoclastogenesis and osteoclast function. Further, an OVX‐induced osteoporosis mice model was employed to identify the effect of CA on bone loss using micro‐CT scanning and histological examination. To investigate underlying mechanisms, network pharmacology was applied to predict the downstream signaling pathways, which were verified by Western blot and immunofluorescence staining. Key Results: The molecular docking analysis revealed that CA exhibited a specific binding affinity to RANKL, engaging multiple binding sites. CA inhibited RANKL‐induced osteoclastogenesis and bone resorption without cytotoxic effects. Mechanistically, CA suppressed RANKL‐induced intracellular reactive oxygen species, nuclear factor‐kappa B, and mitogen‐activated protein kinase pathways, followed by abrogated nuclear factor activated T‐cells 1 activity. Consistent with this finding, CA attenuated post‐ovariectomy‐induced osteoporosis by ameliorating osteoclastogenesis. Conclusions and Implications: CA inhibited osteoclast activity and bone loss by targeting RANKL. CA might represent a promising candidate for treating osteoclast‐related diseases, such as osteoporosis. [ABSTRACT FROM AUTHOR]

Obacunone (OB) targets macrophage migration inhibitory factor (MIF) to impede osteoclastogenesis and alleviate osteoporosis. [Display omitted] • Obacunone (OB) effectively inhibits osteoclast formation and function in vitro. • OB attenuates RANKL-induced osteoclast differentiation signaling pathways. • Macrophage migration inhibitory factor (MIF) is identified as the molecular target of OB. • OB alleviates estrogen deficiency-induced bone loss by impeding excessive osteoclast activity. • OB may serve as an alternative therapeutic candidate for osteoporosis. Osteoporosis is the most common bone disorder where the hyperactive osteoclasts represent the leading role during the pathogenesis. Targeting hyperactive osteoclasts is currently the primary therapeutic strategy. However, concerns about the long-term efficacy and side effects of current frontline treatments persist. Alternative therapeutic agents are still needed. Obacunone (OB) is a small molecule with a broad spectrum of biological activities, particularly antioxidant and anti-inflammatory effects. This study aims to examine OB's therapeutic potential on osteoporosis and explore the rudimentary mechanisms. Osteoclast formation and osteoclastic resorption assays were carried out to examine OB's inhibitory effects in vitro , followed by the in-vivo studies of OB's therapeutic effects on ovariectomy-induced osteoporotic preclinical model. To further study the underlying mechanisms, mRNA sequencing and analysis were used to investigate the changes of downstream pathways. The molecular targets of OB were predicted, and in-silico docking analysis was performed. Ligand-target binding was verified by surface plasmon resonance (SPR) assay and Western Blotting assay. The results indicated that OB suppressed the formation of osteoclast and its resorptive function in vitro. Mechanistically, OB interacts with macrophage migration inhibitory factor (MIF) which attenuates receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL)-induced signaling pathways, including reactive oxygen species (ROS), NF-κB pathway, and mitogen-activated protein kinases (MAPKs). These effects eventually caused the diminished expression level of the master transcriptional factor of osteoclastogenesis, nuclear factor of activated T cells 1 (NFATc1), and its downstream osteoclast-specific proteins. Furthermore, our data revealed that OB alleviated estrogen deficiency-induced osteoporosis by targeting MIF and thus inhibiting hyperactive osteoclasts in vivo. These results together implicated that OB may represent as a therapeutic candidate for bone disorders caused by osteoclasts, such as osteoporosis. [ABSTRACT FROM AUTHOR]

This study was designed to determine whether the use of acetylcholinesterase inhibitors (AChEIs), a group of drugs that stimulate acetylcholine receptors and are used to treat Alzheimer's disease (AD), is associated with osteoporosis protection and inhibition of osteoclast differentiation and function. Firstly, we examined the effects of AChEIs on RANKL‐induced osteoclast differentiation and function with osteoclastogenesis and bone resorption assays. Next, we investigated the impacts of AChEIs on RANKL‐induced nuclear factor κB and NFATc1 activation and expression of osteoclast marker proteins CA‐2, CTSK and NFATc1, and dissected the MAPK signaling in osteoclasts in vitro by using luciferase assay and Western blot. Finally, we assessed the in vivo efficacy of AChEIs using an ovariectomy‐induced osteoporosis mouse model, which was analyzed using microcomputed tomography, in vivo osteoclast and osteoblast parameters were assessed using histomorphometry. We found that Donepezil and Rivastigmine inhibited RANKL‐induced osteoclastogenesis and impaired osteoclastic bone resorption. Moreover, AChEIs reduced the RANKL‐induced transcription of Nfatc1, and expression of osteoclast marker genes to varying degrees (mainly Donepezil and Rivastigmine but not Galantamine). Furthermore, AChEIs variably inhibited RANKL‐induced MAPK signaling accompanied by downregulation of AChE transcription. Finally, AChEIs protected against OVX‐induced bone loss mainly by inhibiting osteoclast activity. Taken together, AChEIs (mainly Donepezil and Rivastigmine) exerted a positive effect on bone protection by inhibiting osteoclast function through MAPK and NFATc1 signaling pathways through downregulating AChE. Our findings have important clinical implications that elderly patients with dementia who are at risk of developing osteoporosis may potentially benefit from therapy with the AChEI drugs. Our study may influence drug choice in those patients with both AD and osteoporosis. [ABSTRACT FROM AUTHOR]

Bone homeostasis only exists when the physical function of osteoblast and osteoclast stays in the balance between bone formation and resorption. Bone resorption occurs when the two processes are uncoupled, shifting the balance in favour of bone resorption. Excessive activation of osteoclasts leads to a range of osteolytic bone diseases including osteoporosis, aseptic prosthesis loosening, rheumatoid arthritis, and osteoarthritis. Receptor activator of nuclear factor kappa-B ligand (RANKL) and its downstream signaling pathways are recognized as key mediators that drive the formation and activation of osteoclastic function. Hence, osteoclast formation and/or its function remain as dominant targets for research and development of agents reaching the treatment towards osteolytic diseases. Chrysin (CHR) is a flavonoid with a wide range of anti-inflammatory and anti-tumor effects. However, its effect on osteoclasts remains unknown. In this study, we found the effects of CHR on inhibiting osteoclast differentiation which were assessed in terms of the number and size of TRAcP positive multinucleated osteoclasts (OCs). Further, the inhibitory effects of CHR on bone resorption and osteoclast fusion of pre-OC were assessed by hydroxyapatite resorption pit assay and F-actin belts staining; respectively. Western blotting analysis of RANKL-induced signaling pathways and immunofluorescence analysis for p65 nuclear translocation in response to RANKL-induced osteoclasts were used to analyze the mechanism of action of CHR affecting osteoclasts. Lastly, the murine calvarial osteolysis model revealed that CHR could protect against particle-induced bone destruction in vivo. Collectively, our data strongly suggested that CHR with its promising anti-tumor effects would also be a potential therapeutic agent for osteolytic diseases. [ABSTRACT FROM AUTHOR]

Osteoclasts are of hematopoietic lineage and have the ability to degrade mineralized bone tissues. Abnormalities in osteoclastic activity under certain pathological conditions are common in bone diseases such as osteoporosis, osteosclerosis, and arthritis. Although many kinds of drugs are currently used to treat osteoporosis, they have obvious adverse reactions and limitations. CYT387 is a new small-molecule Janus kinase (JAK) inhibitor involved in hematopoiesis, immune modulation, fertility, lactation, and embryonic development. However, it has remained unclear whether CYT387 functionally impacts osteoclast formation. Our study demonstrated through osteoclast formation assay in vitro , that the use of CYT387 is a potential drug candidate for treating osteoclast-associated bone disease. The effects of CYT387 on osteoclast formation, bone resorption, NFATc1 activation, and especially intracellular ROS levels were investigated in vitro. Further, we examined the preclinical prospects of CYT387 using an oophorectomy (OVX) mouse model of osteoporosis with its anti-osteoclast activity in vivo. On the whole, this study shows that CYT387 holds promise for treating osteoclast-related bone illnesses including osteoporosis. [ABSTRACT FROM AUTHOR]

Wear debris after total joint arthroplasty can attract the recruitment of macrophages, which release pro-inflammatory substances, triggering the activation of osteoclasts, thereby leading to periprosthetic osteolysis (PPOL) and aseptic loosening. However, the development of pharmacological strategies targeting osteoclasts to prevent periprosthetic osteolysis has not been fruitful. In this study, we worked toward researching the effects and mechanisms of a farnesyltransferase (FTase) inhibitor Lonafarnib (Lon) on receptor activator of nuclear factor κB (NF-κB) ligand (RANKL)-induced osteoclastogenesis and bone resorption, as well as the impacts of Lon on titanium particle-induced osteolysis. To investigate the impacts of Lon on bone resorption and osteoclastogenesis in vitro , bone marrow macrophages were incubated and stimulated with RANKL and macrophage colony-stimulating factor (M-CSF). The influence of Lon on osteolysis prevention in vivo was examined utilizing a titanium particle-induced mouse calvarial osteolysis model. The osteoclast-relevant genes expression was explored by real-time quantitative PCR. Immunofluorescence was used to detect intracellular localization of nuclear factor of activated T cells 1 (NFATc1). SiRNA silence assay was applied to examine the influence of FTase on osteoclasts activation. Related signaling pathways, including NFATc1 signaling, NF-κB, mitogen-activated protein kinases pathways were identified by western blot assay. Lon was illustrated to suppress bone resorptive function and osteoclastogenesis in vitro , and it also reduced the production of pro-inflammatory substances and prevented titanium particle-induced osteolysis in vivo. Lon decreased the expression of osteoclast-relevant genes and suppressed NFATc1 nuclear translocation and auto-amplification. Mechanistically, Lon dampened FTase, and inhibition of FTase reduced osteoclast formation by suppressing ERK signaling. Lon is a promising treatment option for osteoclast-related osteolysis diseases including periprosthetic osteolysis by targeted inhibition of FTase through suppressing ERK signaling. [ABSTRACT FROM AUTHOR]

Excessive activity of osteoclasts contributes to skeletal diseases such as osteoporosis and osteolysis. However, current drugs targeting osteoclast have various deficiencies, placing natural compounds as substitutions of great potential. Roburic acid (RA) is a triterpenoid exacted from Radix Gentianae Macrophyllae, which exhibits inhibitory effects on inflammation and oxidation. By employing an in vitro osteoclastogenesis model, this study investigates the effects and mechanisms of RA on intracellular signaling induced by receptor activator of nuclear factor‐κB ligand (RANKL). As expected, RA at a concentration scope from 1 to 10 μM dampened the osteoclast differentiation of bone marrow macrophages (BMMs) but without cell toxicity. Interestingly, RA showed no effect on osteoblastogenesis in vitro. Furthermore, RA mitigated F‐actin ring formation, hydroxyapatite resorption, and gene expression in osteoclasts. Mechanistically, RA suppressed TNF receptor‐associated factor 6 (TRAF6), the crucial adaptor protein following RANKL‐RANK binding. On the one hand, RA downregulated the nuclear factor‐κB (NF‐κB) activity, extracellular regulated protein kinases (ERK) phosphorylation, and calcium oscillations. On the other hand, RA upregulated the antioxidative response element (ARE) response and the protein expression of heme oxygenase (HO)‐1. These upstream alterations eventually led to the suppression of the nuclear factor of activated T cells 1 (NFATc1) activity and the expression of proteins involved in osteoclastogenesis and bone resorption. Furthermore, by using an ovariectomized (OVX) mice model, RA was found to have therapeutic effects against bone loss. On account of these findings, RA could be used to restrain osteoclasts for treating osteoporosis and other osteolytic diseases. [ABSTRACT FROM AUTHOR]

Osteoporosis, which typically affects postmenopausal women, is an osteolytic disease due to over-activation of osteoclasts. However, current drugs targeting osteoclast inhibition face various side effects, making natural compounds with great interest as alternative treatment options. Cycloastragenol (CAG) is a triterpenoid with multiple biological activities. Previously, CAG's activity against aging-related osteoporosis was reported, but the mechanisms of actions for the activities were not understood. This study demonstrated that CAG dose-dependently inhibited osteoclast formation in receptor activator of nuclear factor-κB ligand (RANKL)-stimulated bone marrow macrophage (BMMs). Mechanism studies showed that CAG inhibited NF-κB, calcium, and nuclear factor of activated T cells 1 (NFATc1) pathways. Additionally, CAG also promoted the nuclear factor-erythroid 2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1)/anti-oxidative response element (ARE) pathway that scavenges reactive oxygen species (ROS). Furthermore, CAG was also found to prevent bone loss of postmenopausal osteoporosis (PMO) in a preclinical model of ovariectomized (OVX) mice. Collectively, our research confirms that CAG inhibits the formation and function of osteoclasts by regulating RANKL-induced intracellular signaling pathways, which may represent a promising alternative for the therapy of osteoclast-related disease. [ABSTRACT FROM AUTHOR]

Morin is a natural compound isolated from moraceae family members and has been reported to possess a range of pharmacological activities. However, the effects of morin on bone-associated disorders and the potential mechanism remain unknown. In this study, we investigated the anti-osteoclastogenic effect of morin in vitro and the potential therapeutic effects on ovariectomy (OVX)-induced osteoporosis in vivo. In vitro, by using a bone marrow macrophage-derived osteoclast culture system, we determined that morin attenuated receptor activator of nuclear factor (NF)-κB ligand (RANKL)-induced osteoclast formation via the inhibition of the mitogen-activated protein kinase (MAPK), NF-κB and calcium pathways. In addition, the subsequent expression of nuclear factor of activated T cells c1 (NFATc1) and c-fos was significantly suppressed by morin. In addition, NFATc1 downregulation led to the reduced expression of osteoclastogenesis-related marker genes, such as V-ATPase-d2 and Integrin β3. In vivo, results provided that morin could effectively attenuate OVX-induced bone loss in C57BL/6 mice. In conclusion, our results demonstrated that morin suppressed RANKL-induced osteoclastogenesis via the NF-κB, MAPK and calcium pathways, in addition, its function of preventing OVX-induced bone loss in vivo, which suggested that morin may be a potential therapeutic agent for postmenopausal osteoporosis treatment. [ABSTRACT FROM AUTHOR]

The extravagant osteoclast formation and resorption is the main cause of osteoporosis. Inhibiting the hyperactive osteoclastic resorption is considered as an efficient treatment for osteoporosis. Rhaponticin (RH) is a small molecule that has been reported to possess anti-inflammatory, anti-allergic, anti-fibrotic, and anti-diabetic activities. However, the influence of RH on osteoclasts differentiation and function is still unclear. To this end, an array of assays including receptor activator of nuclear factor kappa-Β (NF-κB) ligand (RANKL) induced osteoclastogenesis, tartrate-resistant acidic phosphatase (TRAcP) staining, immunofluorescence, and hydroxyapatite resorption were performed in this study. It was found that RH had significant anti-catabolic effects by inhibiting osteoclastogenesis and bone resorption without cytotoxicity. Mechanistically, the expression of NADPH oxidase 1 (Nox1) was found to be suppressed and antioxidant enzymes including catalase, superoxide dismutase 2 (SOD-2), and heme oxygenase-1(HO-1) were enhanced following RH treatment, suggesting RH exhibited antioxidant activity by reducing the generation of reactive oxygen species (ROS) as well as enhancing the depletion of ROS. In addition, MAPKs, NF-κB, and intracellular Ca2+ oscillation pathways were significantly inhibited by RH. These changes led to the deactivation of osteoclast master transcriptional factor-nuclear factor of activated T cells 1 (NFATc1), as examined by qPCR and Western blot assay, which led to the decreased expression of downstream integrin β3, c-Fos, cathepsin K, and Atp6v0d2. These results suggested that RH could effectively suppress RANKL-regulated osteoclast formation and bone resorption. Therefore, we propose that RH can represent a novel natural small molecule for the treatment of osteoporosis by inhibiting excessive osteoclast activity. [ABSTRACT FROM AUTHOR]

Giant cell tumor of bone (GCTB) is benign tumor that can cause significant osteolysis and bone destruction at the epiphysis of long bones. Osteoclasts are thought to be highly associated with osteolysis in GCTB. However, the migration of osteoclasts in GCTB remains unclear. A deeper understanding of the complex tumor microenvironment is required in order to delineate the migration of osteoclasts in GCTB. In this study, samples were isolated from one patient diagnosed with GCTB. Single-cell RNA sequencing (scRNA-seq) was used to detect the heterogeneity of GCTB. Multiplex immunofluorescence staining was used to evaluate the cell subtypes identified by scRNA-seq. A total of 8,033 cells were obtained from one patient diagnosed with GCTB, which were divided into eight major cell types as depicted by a single-cell transcriptional map. The osteoclasts were divided into three subsets, and their differentiation trajectory and migration status were further analyzed. Osteoclast migration may be regulated via a series of genes associated with cell migration. Furthermore, four signaling pathways (RANKL, PARs, CD137 and SMEA3 signaling pathway) were found to be highly associated with osteoclast migration. This comprehensive single-cell transcriptome analysis of GCTB identified a series of genes associated with cell migration as well as four major signaling pathways that were highly related to the migration of osteoclasts in GCTB. Our findings broaden the understanding of GCTB bionetworks and provides a theoretical basis for anti-osteolysis therapy against GCTB in the future. [ABSTRACT FROM AUTHOR]

This article was submitted to Experimental Pharmacology and Drug Discovery, a section of the journal Frontiers in Pharmacology. Postmenopausal osteoporosis (PMOP), which increases the risk of fracture, is the most common bone disease in women. PMOP not only increases the risk of death but also imposes a financial burden on countless families. At present, most of the drugs used to treat osteoporosis have significant side effects, so it is important to find effective anti-osteoporosis medications without major side effects. Sesamolin (Ses) is a kind of natural lignan extracted from sesame oil. Many researches have shown that Ses has anti-inflammatory, antioxidative, and anticancer effects, however it is still unknown whether it has any effect on osteoporosis. In this research, we explored the therapeutic effect of Ses in the process of osteoclast formation and bone resorption and found that Ses effectively inhibited osteoclast formation in vitro through TRAcP staining and hydroxyapatite resorption assays. Through Western blot analysis of the NF-κB pathway, MAPK pathway, c-Fos and NFATc1, it was found that Ses not only effectively inhibited the activation of NF-κB and MAPK signaling pathways induced by RANKL but also significantly reduced the protein expression of c-Fos and NFATc1. Several genes specifically expressed in osteoclasts were determined by qPCR, and Ses was also found to play a significant inhibitory role on the expression of these genes. Besides, an osteoporosis model induced in ovariectomized (OVX) mice was employed to verify that Ses could effectively reduce bone loss caused by estrogen deficiency in vivo. In conclusion, Ses showed promise as a new treatment for postmenopausal osteoporosis. [ABSTRACT FROM AUTHOR]

The incidence of osteoporosis, which is primarily characterized by plethoric osteoclast (OC) formation and severe bone loss, has increased in recent years. Millions of people worldwide, especially postmenopausal women, suffer from osteoporosis. The drugs commonly used to treat osteoporosis still exist many disadvantages, but natural extracts provide options for the treatment of osteoporosis. Therefore, the identification of cost-effective natural compounds is important. Patchouli alcohol (PA), a natural compound extracted from Pogostemon cablin that exerts anti-inflammatory effects, is used as a treatment for gastroenteritis. However, no research on the use of Patchouli alcohol in osteoporosis has been reported. We found that PA dose-dependently inhibited the receptor activator of nuclear factor kappa-B ligand (RANKL)-induced formation and function of OCs without cytotoxicity. Furthermore, these inhibitory effects were reflected in the significant effect of PA on the NF-κB signaling pathway, as PA suppressed the transcription factors NFATc1 and c-Fos. We also determined that PA activated expression of the nuclear receptor pregnane X receptor (PXR) and promoted the PXR/Toll-like receptor 4 (TLR4) axis to inhibit the nuclear import of NF-κB (p50 and p65). Additionally, PA exerted therapeutic effects against osteoporosis in ovariectomized (OVX) mice, supporting the use of PA as a treatment for osteoporosis in the future. [ABSTRACT FROM AUTHOR]

Revision operations have become a new issue after successful artificial joint replacements, and periprosthetic osteolysis leading to prosthetic loosening is the main cause of why the overactivation of osteoclasts (OCs) plays an important role. The effect of biochanin A (BCA) has been examined in osteoporosis, but no study on the role of BCA in prosthetic loosening osteolysis has been conducted yet. In this study, we utilised enzyme‐linked immunosorbent assay, computed tomography imaging, and histological analysis. Results showed that BCA downregulated the secretion levels of tumor necrosis factor‐α, interleukin‐1α (IL‐1α), and IL‐1β to suppress inflammatory responses. The secretion levels of receptor‐activated nuclear factor‐κB ligand, CTX‐1, and osteoclast‐associated receptor as well as Ti‐induced osteolysis were also reduced. BCA effectively inhibited osteoclastogenesis and suppressed hydroxyapatite resorption by downregulating OC‐related genes in vitro. Analysis of mechanisms indicated that BCA inhibited the signalling pathways of mitogen‐activated protein kinase (P38, extracellular signal‐regulated kinase, and c‐JUN N‐terminal kinase) and nuclear factor‐κB (inhibitor κB‐α and P65), thereby downregulating the expression of nuclear factor of activated T cell 1 and c‐Fos. In conclusion, BCA may be an alternative choice for the prevention of prosthetic loosening caused by OCs. [ABSTRACT FROM AUTHOR]

Microfibrillar‐associated proteins (MFAPs) are extracellular matrix glycoproteins, which play a role in microfibril assembly, elastinogenesis, and tissue homeostasis. MFAPs consist of five subfamily members, including MFAP1, MFAP2, MFAP3, MFAP4, and MFAP5. Among these, MFAP2 and MFAP5 are most closely related, and exhibit very limited amino acid sequence homology with MFAP1, MFAP3, and MFAP4. Gene expression profiling analysis reveals that MFAP2, MFAP5, and MFAP4 are specifically expressed in osteoblastic like cells, whereas MFAP1 and MFAP3 are more ubiquitously expressed, indicative of their diverse role in the tropism of tissues. Molecular structural analysis shows that each MFAP family member has distinct features, and functional evidence reveals discrete purposes of individual MFAPs. Animal studies indicate that MFAP2‐deficient mice exhibit progressive osteopenia with elevated receptor activator of NF‐κB ligand (RANKL) expression, whereas MFAP5‐deficient mice are neutropenic, and MFAP4‐deficient mice displayed emphysema‐like pathology and the impaired formation of neointimal hyperplasia. Emerging data also suggest that MFAPs are involved in cancer progression and fat metabolism. Further understanding of tissue‐specific pathophysiology of MFAPs might offer potential novel therapeutic targets for related diseases, such as skeletal and metabolic disorders, and cancers. [ABSTRACT FROM AUTHOR]

Osteolytic diseases are typified by over‐enhanced formation and resorbing function of osteoclasts and have a major impact on human health. Inhibition of osteoclastic differentiation and function is a key strategy for clinical therapy of osteolytic conditions. Maackiain is a natural compound extracted from Sophora flavescens, which has been applied to anti‐allergic and anti‐tumour treatments. The present results showed that Maackiain could restrain receptor activator of nuclear factor‐κB ligand (RANKL)‐stimulated osteoclast formation and hydroxyapatite resorption dose‐dependently, and interrupt the structures of F‐actin belts in the mature osteoclasts. It also repressed the expressions of osteoclast‐specific genes and proteins. Furthermore, Maackiain could inhibit RANKL‐stimulated NF‐κB and calcium signalling pathways, and dampen Nuclear factor of activated T cell cytoplasmic 1 activity, protein expression and translocation into the nucleus. These results revealed that Maackiain may have a potential therapeutic effect on osteoclast‐related disorders. [ABSTRACT FROM AUTHOR]

Osteoporosis is a devastating disease that features reduced bone quantity and microstructure, which causes fragility fracture and increases mortality, especially in the aged population. Due to the long‐term side‐effects of current drugs for osteoporosis, it is of importance to find other safe and effective medications. Ellagic acid (EA) is a phenolic compound found in nut galls, plant extracts, and fruits, and exhibits antioxidant and antineoplastic effects. Here, we showed that EA attenuated the formation and function of osteoclast dose‐dependently. The underlying mechanism was further discovered by western blot, immunofluorescence assay, and luciferase assay, which elucidated that EA suppressed osteoclastogenesis and bone resorption mainly through attenuating receptor activator of nuclear factor‐κB (NF‐κB) ligand‐induced NF‐κB activation and extracellular signal‐regulated kinase signaling pathways, accompanied by decreased protein expression of nuclear factor of activated T‐cells calcineurin‐dependent 1 and c‐Fos. Moreover, EA inhibits osteoclast marker genes expression including Dc‐stamp, Ctsk, Atp6v0d2, and Acp5. Intriguingly, we also found that EA treatment could significantly protect ovariectomy‐induced bone loss in vivo. Conclusively, this study suggested that EA might have the therapeutic potentiality for preventing or treating osteoporosis. [ABSTRACT FROM AUTHOR]

Osteoclasts with elevated bone resorption are commonly present in postmenopausal osteoporosis, and other osteolytic pathologies. Therefore, suppressing osteoclast generation and function has been the main focus of osteoporosis treatment. Betulinic acid (BA) represents a triterpenoid mainly purified from the bark of Betulaceae. BA shows multiple biological activities, including antitumor and anti-HIV properties, but its effect on osteolytic conditions is unknown. Here, BA suppressed receptor activator of nuclear factor‐κB ligand (RANKL)‐associated osteoclastogenesis and bone resorptive function, as assessed by tartrate‐resistant acid phosphatase (TRAP) staining, fibrous actin ring generation, and hydroxyapatite resorption assays. Mechanistically, BA downregulated the expression of osteoclastic-specific genes. Western blot analysis revealed that BA significantly interrupted ERK, JNK and p38 MAPK activation as well as intracellular reactive oxygen species (ROS) production, thus altering c-Fos and NFATc1 activation. Corroborating the above findings in cell-based assays, BA prevented ovariectomy-associated bone loss in an animal model. In conclusion, these findings suggest that BA can inhibit osteoclast generation and function as well as the RANKL signaling pathway, and might be used for treating osteoclast-related osteoporosis. [ABSTRACT FROM AUTHOR]

Asiatic acid is a triterpenoid compound extracted from a medicinal plant Centella asiatica. It has been used as a highly efficient compound for the treatment of cancer and hyperlipidemia, as well as possessing potential antiinflammatory properties. However, its effects on bone metabolism and osteoporosis haven't been reported. The purpose of our research were to reveal the biomolecular effects of asiatic acid on osteoclasts, and its underlying molecular mechanisms regulating its effects on receptor activator of NF-κB ligand (RANKL)-induced signaling pathways. We found that asiatic acid inhibited multinucleated tartrate-resistant acid phosphatase (TRAcP)-positive osteoclast differentiation and osteoclast induced bone loss. Real time PCR showed that asiatic acid reduced the expression of down-cascade target genes including Ctsk , Nfatc1 , Calcr , and Atp6v0d2. Western blot and luciferase reporter gene assays revealed that asiatic acid inhibits RANKL mediated NF-κB and NFATc1 signalings. Further, in vivo study demonstrated asiatic acid attenuates estrogen deficiency-induced bone loss in ovariectomized mice. MicroCT and histology analyses revealed that osteoclast numbers were significantly suppressed in asiatic acid treated groups. Furthermore, serum levels of TRAcP and CTX-1 were downregulated in treated groups. Taken together, our data show that asiatic acid can inhibit osteoclastic formation and reduce OVX-induced bone resorption through RANKL-activated NF-κB or NFATc1 signaling, suggesting that asiatic acid may be a potential and effective natural compound for the therapy of excessive RANKL-related osteolytic diseases. [ABSTRACT FROM AUTHOR]

Osteolytic skeletal disorders are caused by an imbalance in the osteoclast and osteoblast function. Suppressing the differentiation and resorptive function of osteoclast is a key strategy for treating osteolytic diseases. Dracorhodin perchlorate (D.P), an active component from dragon blood resin, has been used for facilitating wound healing and anti‐cancer treatments. In this study, we determined the effect of D.P on osteoclast differentiation and function. We have found that D.P inhibited RANKL‐induced osteoclast formation and resorbed pits of hydroxyapatite‐coated plate in a dose‐dependent manner. D.P also disrupted the formation of intact actin‐rich podosome structures in mature osteoclasts and inhibited osteoclast‐specific gene and protein expressions. Further, D.P was able to suppress RANKL‐activated JNK, NF‐κB and Ca2+ signalling pathways and reduces the expression level of NFATc1 as well as the nucleus translocation of NFATc1. Overall, these results indicated a potential therapeutic effect of D.P on osteoclast‐related conditions. [ABSTRACT FROM AUTHOR]

Excessive bone resorption conducted by osteoclasts is considered as the main cause of osteoclast-related bone diseases such as osteoporosis. Therefore, the suppression of excessive osteoclast formation and function is one of the strategies to treat osteoclast-related bone diseases. Fumitremorgin C (Fum) is a mycotoxin extracted from Aspergillus fumigatus. It has been shown to have extensive pharmacological properties, but its role in the treatment of osteoclast-related bone diseases remains unclear. In this study, we aim to find out whether Fum can inhibit the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation and function. The results showed that Fum could significantly attenuate osteoclast formation and function at concentrations from 2.5 to 10 µM. The protein expression of bone resorption factors such as NFATc1, cathepsin K, V-ATPase-d2, and c-Fos was suppressed with the treatment of Fum at a concentration of 10 µM. In addition, Fum was also shown to suppress the activity of NF-κB, intracellular reactive oxygen species level, and MAPK pathway. Taken together, the present study showed that Fum could attenuate the formation and function of osteoclast via suppressing RANKL-induced signaling pathways, suggesting that Fum might be a potential novel drug in the treatment of osteoclast-related bone diseases. [ABSTRACT FROM AUTHOR]

Osteolytic bone diseases, for example postmenopausal osteoporosis, arise from the imbalances between osteoclasts and osteoblasts in the bone remodeling process, whereby osteoclastic bone resorption greatly exceeds osteoblastic bone formation resulting in severe bone loss and deterioration in bone structure and microarchitecture. Therefore, the identification of agents that can inhibit osteoclast formation and/or function for the treatment of osteolytic bone disease has been the focus of bone and orthopedic research. Vindoline (Vin), an indole alkaloid extracted from the medicinal plant Catharanthus roseus , has been shown to possess extensive biological and pharmacological benefits, but its effects on bone metabolism remains to be documented. Our study demonstrated for the first time, that Vin could inhibit osteoclast differentiation from bone marrow macrophages (BMMs) precursor cells as well as mature osteoclastic bone resorption. We further determined that the underlying molecular mechanism of action of Vin is in part due to its inhibitory effect against the activation of MAPK including p38, JNK, and ERK and intracellular reactive oxygen species (ROS) production. This effect ultimately suppressed the induction of c-Fos and NFATc1, which consequently downregulated the expression of the genes required for osteoclast formation and bone resorption. Consistent with our in vitro findings, in vivo administration of Vin protected mice against ovariectomy (OVX)-induced bone loss and trabecular bone deterioration. These results provided promising evidence for the potential therapeutic application of Vin as a novel treatment option against osteolytic diseases. [ABSTRACT FROM AUTHOR]

Osteoporosis is the most common osteolytic disease characterized by excessive osteoclast formation and resultant bone loss, which afflicts millions of patients around the world. Astilbin, a traditional herb, is known to have anti‐inflammatory, antioxidant and antihepatic properties, but its role in osteoporosis treatment has not yet been confirmed. In our study, astilbin was found to have an inhibitory effect on the RANKL‐induced formation and function of OCs in a dose‐dependent manner without cytotoxicity. These effects were attributed to its ability to suppress the activity of two transcription factors (NFATc1 and c‐Fos) indispensable for osteoclast formation, followed by inhibition of the expression of bone resorption‐related genes and proteins (Acp5/TRAcP, CTSK, V‐ATPase‐d2 and integrin β3). Furthermore, we examined the underlying mechanisms and found that astilbin repressed osteoclastogenesis by blocking Ca2+ oscillations and the NF‐κB and MAPK pathways. In addition, the therapeutic effect of MA on preventing bone loss in vivo was further confirmed in an ovariectomized mouse model. Therefore, considering its ability to inhibit RANKL‐mediated osteoclastogenesis and the underlying mechanisms, astilbin might be a potential candidate for treating osteolytic bone diseases. [ABSTRACT FROM AUTHOR]

Imbalance of osteoblast and osteoclast in adult leads to a variety of bone‐related diseases, including osteoporosis. Thus, suppressing the activity of osteoclastic bone resorption becomes the main therapeutic strategy for osteoporosis. Asperpyrone A is a natural compound isolated from Aspergillus niger with various biological activities of antitumour, antimicrobial and antioxidant. The present study was designed to investigate the effects of Asperpyrone A on osteoclastogenesis and to explore its underlining mechanism. We found that Asperpyrone A inhibited RANKL‐induced osteoclastogenesis in a dose‐dependent manner when the concentration reached 1 µm, and with no cytotoxicity until the concentration reached to 10 µm. In addition, Asperpyrone A down‐regulated the mRNA and protein expression of NFATc1, c‐fos and V‐ATPase‐d2, as well as the mRNA expression of TRAcP and Ctsk. Furthermore, Asperpyrone A strongly attenuated the RNAKL‐induced intracellular Ca2+ oscillations and ROS (reactive oxygen species) production in the process of osteoclastogenesis and suppressed the activation of MAPK and NF‐κB signalling pathways. Collectively, Asperpyrone A attenuates RANKL‐induced osteoclast formation via suppressing NFATc1, Ca2+ signalling and oxidative stress, as well as MAPK and NF‐κB signalling pathways, indicating that this compound may become a potential candidate drug for the prevention or treatment of osteoporosis. [ABSTRACT FROM AUTHOR]

Aseptic prosthetic loosening and periprosthetic infection resulting in inflammatory osteolysis is a leading complication of total joint arthroplasty (TJA). Excessive bone destruction around the bone and prosthesis interface plays a key role in the loosening prostheses leading to revision surgery. The bacterial endotoxins or implant‐derived wear particles‐induced inflammatory response is the major cause of the elevated osteoclast formation and activity. Thus, agents or compounds that can attenuate the inflammatory response and/or inhibit the elevated osteoclastogenesis and excessive bone resorption would provide a promising therapeutic avenue to prevent aseptic prosthetic loosening in TJA. Daphnetin (DAP), a natural coumarin derivative, is clinically used in Traditional Chinese Medicine for the treatment of rheumatoid arthritis due to its anti‐inflammatory properties. In this study, we report for the first time that DAP could protect against lipopolysaccharide‐induced inflammatory bone destruction in a murine calvarial osteolysis model in vivo. This protective effect of DAP can in part be attributed to its direct inhibitory effect on RANKL‐induced osteoclast differentiation, fusion, and bone resorption in vitro. Biochemical analysis found that DAP inhibited the activation of the ERK and NFATc1 signaling cascades. Collectively, our findings suggest that DAP as a natural compound has potential for the treatment of inflammatory osteolysis. [ABSTRACT FROM AUTHOR]

Prosthesis loosening is a highly troublesome clinical problem following total joint arthroplasty. Wear‐particle‐induced osteoclastogenesis has been shown to be the primary cause of periprosthetic osteolysis that eventually leads to aseptic prosthesis loosening. Therefore, inhibiting osteoclastogenesis is a promising strategy to control periprosthetic osteolysis. The possible mechanism of action of rhoifolin on osteoclastogenesis and titanium particle‐induced calvarial osteolysis was examined in this study. The in vitro study showed that rhoifolin could strongly suppress the receptor activators of nuclear factor‐κB (NF‐κB) ligand‐stimulated osteoclastogenesis, hydroxyapatite resorption, F‐actin formation, and the gene expression of osteoclast‐related genes. Western blot analysis illustrated that rhoifolin could attenuate the NF‐κB and mitogen‐activated protein kinase pathways, and the expression of transcriptional factors nuclear factor of activated T cells 1 (NFATc1) and c‐Fos. Further studies indicated that rhoifolin inhibited p65 translocation to the nucleus and the activity of NFATc1 and NF‐κB rhoifolin could decrease the number of tartrate‐resistant acid phosphate‐positive osteoclasts and titanium particle‐induced C57 mouse calvarial bone loss in vivo. In conclusion, our results suggest that rhoifolin can ameliorate the osteoclasts‐stimulated osteolysis, and may be a potential agent for the treatment of prosthesis loosening. [ABSTRACT FROM AUTHOR]

Osteoporosis (OP) is a metabolic disease caused by multiple factors, which is characterized by a reduction of bone mass per unit volume and destruction of bone microstructure. Aberrant osteoclast function is the main cause of OP, therefore, regulating the differentiation and function of osteoclast is one of the treatment strategies for OP. Pectolinarigenin (PEC) is a medicinal implant isolated from Fragrant Eupatorium. Our experimental data showed that PEC was able to inhibit receptor activator of nuclear factor‐κB ligand (RANKL)‐induced osteoclastogenesis in vitro, by tartrate‐resistant acid phosphatase (TRAcP) staining, Fibrous actin ring formation, and hydroxyapatite resorption assays. In terms of mechanism, PEC inhibited the expression of the osteoclastogenesis‐related gene, including cathepsin K (Ctsk), matrix metalloproteinase 9 (Mmp9), and TRAcP (Acp5). Western blot analysis demonstrated that PEC could significantly block the activation of RANKL‐induced mitogen‐activated protein kinase signaling cascades and was able to suppress the protein expression of nuclear factor of activated T‐cells and c‐Fos. Meanwhile, the intracellular reactive oxygen species levels were also reduced by PEC in a concentration‐dependent manner. Further, PEC could prevent the ovariectomy‐induced bone loss in vivo. Summarizing all, our data suggested that PEC inhibits osteoclast formation and function and RANKL signaling pathways, and thus could potentially be used in the treatment the osteoclast‐related bone loss diseases. [ABSTRACT FROM AUTHOR]

Wearing titanium particle‐induced osteoclastogenesis, accompanied by peri‐implant osteolysis, is the main cause of long‐term failure of hip prosthesis. Currently, medications used for the prevention and treatment of peri‐implant osteolysis show serious side effects. Therefore, development for more effective new drugs with less side effects is extremely urgent. Vaccarin is a natural flavonoid extracted from Vaccaria segetalis, with various biological functions, including antioxidantory, anti‐inflammatory, and promotion of angiogenesis. However, the putative role of vaccarin in the inhibition of titanium particle‐induced osteolysis has not been reported. In this study, it was indicated that vaccarin could effectively inhibit RANKL‐induced osteoclastogenesis, fusion of F‐actin rings, bone resorption, and expression of osteoclast marker genes in a dose‐dependent manner in vitro. Moreover, vaccarin could also inhibit RANKL‐induced osteoclastogenesis via the inhibition of NF‐κB and MAPK (p38, ERK, and JNK) signaling pathways, and inhibit the transcription of downstream transcription factors, such as c‐Fos and NFATc1. Consistent with in vitro results, this in vivo study showed that vaccarin exhibited an inhibitory effect on titanium particle‐induced osteolysis by antiosteoclastogenesis. In conclusion, vaccarin could be a promising agent for preventing and treating peri‐implant osteolysis. [ABSTRACT FROM AUTHOR]

Osteoporosis is a form of osteolytic disease caused by an imbalance in bone homeostasis, with reductions in osteoblast bone formation, and augmented osteoclast formation and resorption resulting in reduced bone mass. Cajaninstilbene acid (CSA) is a natural compound derived from pigeon pea leaves. CSA possesses beneficial properties as an anti‐inflammatory, antibacterial, antihepatitis, and anticancer agent; however, its potential to modulate bone homeostasis and osteoporosis has not been studied. We observed that CSA has the ability to suppress RANKL‐mediated osteoclastogenesis, osteoclast marker gene expression, and bone resorption in a dose‐dependent manner. Mechanistically, it was revealed that CSA attenuates RANKL‐activated NF‐κB and nuclear factor of activated T‐cell pathways and inhibited phosphorylation of key signaling mediators c‐Fos, V‐ATPase‐d2, and ERK. Moreover, in osteoclasts, CSA blocked RANKL‐induced ROS activity as well as calcium oscillations. We further evaluated the therapeutic effect of CSA in a preclinical mouse model and showed that in vivo treatment of ovariectomized C57BL/6 mice with CSA protects the mice from osteoporotic bone loss. Thus, this study demonstrates that osteolytic bone diseases can potentially be treated by CSA. Cajaninstilbene acid (CSA), a natural compound derived from pigeon pea leaves, has the ability to suppress RANKL‐mediated osteoclastogenesis, osteoclast marker gene expression, and bone resorption in a dose‐dependent manner. It attenuates RANKL‐activated NF‐κB and nuclear factor of activated T‐cell (NFAT) pathways and inhibited phosphorylation of key signaling mediators c‐Fos, V‐ATPase‐d2, and ERK. Moreover, in osteoclasts, CSA blocked the RANKL‐induced ROS activity as well as calcium oscillations.The treatment of ovariectomized C57BL/6 mice with CSA protects the mice from osteoporotic bone loss. Thus, this study demonstrates that osteolytic bone diseases can potentially be treated by CSA. [ABSTRACT FROM AUTHOR]

Excessive osteoclast formation and function are considered as the main causes of bone lytic disorders such as osteoporosis and osteolysis. Therefore, the osteoclast is a potential therapeutic target for the treatment of osteoporosis or other osteoclast‐related diseases. Helvolic acid (HA), a mycotoxin originally isolated from Aspergillus fumigatus , has been discovered as an effective broad‐spectrum antibacterial agent and has a wide range of pharmacological properties. Herein, for the first time, HA was demonstrated to be capable of significantly inhibiting receptor activator of nuclear factor‐κB ligand (RANKL)‐induced osteoclastogenesis and bone resorption in vitro by suppressing nuclear factor of activated T cells 1 (NFATc1) activation. This inhibition was followed by the dramatically decreased expression of NFATc1‐targeted genes including Ctr (encoding calcitonin receptor), Acp5 (encoding tartrate‐resistant acid phosphatase [TRAcP]), Ctsk (encoding cathepsin K), Atp6v0d2 (encoding the vacuolar H+ ATPase V0 subunit d2 [V‐ATPase‐d2]) and Mmp9 (encoding matrix metallopeptidase 9) which are osteoclastic‐specific genes required for osteoclast formation and function. Mechanistically, HA was shown to greatly attenuate multiple upstream pathways including extracellular signal‐regulated kinase (ERK) phosphorylation, c‐Fos signaling, and intracellular Ca 2+ oscillation, but had little effect on nuclear factor‐κB (NF‐κB) activation. In addition, HA also diminished the RANKL‐induced generation of intracellular reactive oxygen species. Taken together, our study indicated HA effectively suppressed RANKL‐induced osteoclast formation and function. Thus, we propose that HA can be potentially used in the development of a novel drug for osteoclast‐related bone diseases. Helvolic acid (HA), a mycotoxin originally isolated from Aspergillus fumigatus, is demonstrated to be capable of significantly inhibiting receptor activator of nuclear factor‐κB ligand (RANKL)‐induced osteoclastogenesis and bone resorption in vitro by suppressing nuclear factor of activated T cells 1 (NFATc1) activation without affecting nuclear factor‐κB (NF‐κB) activation. Mechanistically, HA greatly attenuated multiple upstream pathways of NFATc1, including extracellular‐signal‐regulated kinase (ERK) phosphorylation, c‐Fos signaling, and intracellular Ca2+oscillation. Taken together, HA can be potentially used in the development of a novel drug for osteoclast‐related bone diseases. [ABSTRACT FROM AUTHOR]

Identification of natural compounds that inhibit osteoclastogenesis will facilitate the development of antiresorptive treatment of osteolytic bone diseases. Asiaticoside is a triterpenoid derivative isolated from Centella asiatica, which exhibits varying biological effects like angiogenesis, anti‐inflammation, wound healing, and osteogenic differentiation. However, its role in osteoclastogenesis remains unknown. Here, we show that Asiaticoside can suppress RANKL‐induced osteoclast formation and bone resorption in a dose‐dependent manner. Asiaticoside attenuated the expression of osteoclast marker genes including Ctsk, Atp6v0d2, Nfatc1, Acp5, and Dc‐stamp. Furthermore, Asiaticoside inhibited RANKL‐mediated NF‐κB and NFATc1 activities, and RANKL‐induced calcium oscillation. Collectively, this study demonstrates that Asiaticoside inhibited osteoclast formation and function through attenuating RANKL‐induced key signaling pathways, which may indicate that Asiaticoside is a potential antiresorptive agent against osteoclast‐related osteolytic bone diseases. Asiaticoside can suppress RANKL‐induced osteoclast formation and bone resorption in a dose‐dependent manner. Asiaticoside attenuated the expression of osteoclast marker genes including Ctsk, V‐ATPase d2, NFATc1, TRAcP, and DC‐STAMP. Furthermore, Asiaticoside inhibited RANKL‐mediated NF‐κB and NFATc1 activities, and RANKL‐induced calcium oscillation. [ABSTRACT FROM AUTHOR]

Postmenopausal osteoporosis (PMO) is a progressive bone disease characterized by the over‐production and activation of osteoclasts in elderly women. In our study, we investigated the anti‐osteoclastogenic effect of evodiamine (EVO) in vivo and in vitro, as well as the underlying mechanism. By using an in vitro bone marrow macrophage (BMM)‐derived osteoclast culture system, we found that EVO inhibited osteoclast formation, hydroxyapatite resorption and receptor activator of NF‐κB ligand (RANKL)‐induced osteoclast marker gene and protein expression. Mechanistically, we found that EVO inhibited the degradation and RANKL‐induced transcriptional activity of IκBα. RANKL‐induced Ca2+ oscillations were also abrogated by EVO. In vivo, an ovariectomized (OVX) mouse model was established to mimic PMO, and OVX mice received oral administration of either EVO (10 mg/kg) or saline every other day. We found that EVO can attenuate bone loss in OVX mice by inhibiting osteoclastogenesis. Taken together, our findings suggest that EVO suppresses RANKL‐induced osteoclastogenesis through NF‐κB and calcium signalling pathways and has potential value as a therapeutic agent for PMO. [ABSTRACT FROM AUTHOR]

Osteoporosis is the most common osteolytic disease characterized by excessive osteoclast formation and resultant bone loss, which afflicts millions of patients around the world. Madecassoside (MA), isolated from Centella asiatica, was reported to have anti‐inflammatory and antioxidant activities, but its role in osteoporosis treatment has not yet been confirmed. In our study, MA was found to have an inhibitory effect on the RANKL‐induced formation and function of OCs in a dose‐dependent manner without cytotoxicity. These effects were attributed to its ability to suppress the activity of two transcription factors (NFATc1 and c‐Fos) indispensable for osteoclast formation, followed by inhibition of the expression of bone resorption‐related genes and proteins (Acp5/TRAcP, CTSK, ATP6V0D2/V‐ATPase‐d2, and integrin β3). Furthermore, we examined the underlying mechanisms and found that MA represses osteoclastogenesis by blocking Ca2+ oscillations and the NF‐κB and MAPK pathways. In addition, the therapeutic effect of MA on preventing bone loss in vivo was further confirmed in an ovariectomized mouse model. Therefore, considering its ability to inhibit RANKL‐mediated osteoclastogenesis and the underlying mechanisms, MA might be a potential candidate for treating osteolytic bone diseases. [ABSTRACT FROM AUTHOR]

Osteoclasts are responsible for bone resorption during the process of bone remodeling. Increased osteoclast numbers and bone resorption activity are the main factors contributing to bone loss–related diseases such as osteoporosis. Therefore, modulating the formation and function of osteoclasts is critical for the effective treatment of osteolysis and osteoporosis. Kavain is the active ingredient extracted from the root of the kava plant, which possesses known anti‐inflammatory properties. However, the effects of kavain on osteoclastogenesis and bone resorption remain unclear. In this study, we found that kavain inhibits receptor activator of nuclear factor‐κB ligand (RANKL)–induced osteoclast differentiation and fusion using tartrate‐resistant acid phosphatase staining and immunofluorescence. Furthermore, kavain inhibited bone resorption performed by osteoclasts. Using reverse transcription‐polymerase chain reaction and western blot analysis, we found that kavain downregulates the expression of osteoclast marker genes, such as nuclear factor of activated T cells, cytoplasmic 1 (Nfatc1), v‐atpase d2 (Atp6v0d2), dendrocyte expressed seven transmembrane protein (Dcstamp), matrix metallopeptidase 9 (Mmp9), cathepsin K (Ctsk), and Acp5. Additionally, kavain repressed RANKL‐induced calcium oscillations, nuclear factor of activated T cells activation, and mitogen‐activated protein kinase phosphorylation, while leaving NF‐κB unaffected. We found no effects of kavain on either osteoblast proliferation or differentiation. Besides, kavain inhibited bone loss in ovariectomized mice by suppressing osteoclastogenesis. Collectively, these data suggest a potential use for kavain as a candidate drug for the treatment of osteolytic diseases. Kavain suppresses NFATc1 expression and phosphorylation of P38 and JNK, while leaving ERK and NF‐κB unaffected. Besides, kavain attenuates NFATc1 activation via modulating intracellular calcium oscillations, hence inhibits osteoclastogenesis. [ABSTRACT FROM AUTHOR]

MiR‐214 belongs to a family of microRNA (small, highly conserved noncoding RNA molecules) precursors that play a pivotal role in biological functions, such as cellular function, tissue development, tissue homeostasis, and pathogenesis of diseases. Recently, miR‐214 emerged as a critical regulator of musculoskeletal metabolism. Specifically, miR‐214 can mediate skeletal muscle myogenesis and vascular smooth muscle cell proliferation, migration, and differentiation. MiR‐214 also modulates osteoblast function by targeting specific molecular pathways and the expression of various osteoblast‐related genes; promotes osteoclast activity by targeting phosphatase and tensin homolog (Pten); and mediates osteoclast‐osteoblast intercellular crosstalk via an exosomal miRNA paracrine mechanism. Importantly, dysregulation in miR‐214 expression is associated with pathological bone conditions such as osteoporosis, osteosarcoma, multiple myeloma, and osteolytic bone metastasis of breast cancer. This review discusses the cellular targets of miR‐214 in bone, the molecular mechanisms governing the activities of miR‐214 in the musculoskeletal system, and the putative role of miR‐214 in skeletal diseases. Understanding the biology of miR‐214 could potentially lead to the development of miR‐214 as a possible biomarker and a therapeutic target for musculoskeletal diseases. This review discusses the cellular targets of miR‐214 in bone, the molecular mechanisms governing the activities of miR‐214 in the musculoskeletal system and the putative role of miR‐214 in skeletal diseases. Understanding the biology of miR‐214 could potentially lead to the development of miR‐214 as a possible biomarker and a therapeutic target for musculoskeletal diseases. Highlights: MiR‐214 is an important regulator of the musculoskeletal system and is involved in muscle development, bone homeostasis and bone‐related disorders.MiR‐214 is able to mediate skeletal muscle myogenesis and vascular smooth muscle cell proliferation, migration and differentiation.MiR‐214 modulates osteoblast function by targeting specific molecular pathways and the expression of various osteoblast‐related genes.MiR‐214 promotes osteoclast activity by targeting specific genes and mediates osteoblast‐osteoclast intercellular crosstalk via an exosomal miRNA paracrine mechanism.Dysregulation in miR‐214 expression has been associated with bone diseases such as osteoporosis, osteosarcoma and multiple myeloma.Osteoclastic miR‐214 is involved in regulating osteoclast‐mediated osteolytic bone metastasis of breast cancer. [ABSTRACT FROM AUTHOR]

Osteoporosis is a metabolic disease characterized by osteopenia and bone microstructural deterioration. Osteoclasts are the primary effector cells that degrade bone matrix and their abnormal function leads to the development of osteoporosis. Reactive oxygen species (ROS) accumulation during cellular metabolism promotes osteoclast proliferation and differentiation, therefore, playing an important role in osteoporosis. Cistanche deserticola polysaccharide (CDP) possesses antitumor, anti‐inflammatory, and antioxidant activity. However, the impact of CDP on osteoclasts is unclear. In this study, tartrate‐resistant acid phosphatase staining, immunofluorescence, reverse transcription‐polymerase chain reaction, and western blot analysis were utilized to demonstrate that CDP inhibited osteoclastogenesis and hydroxyapatite resorption. In addition, CDP also inhibited the expression of osteoclast maker genes including Ctsk, Mmp9, and Acp5 and had no effect on receptor activator of nuclear factor κB (RANK) expression. Mechanistic analyses revealed that CDP increases the expression of antioxidant enzymes to attenuate RANKL‐mediated ROS production in osteoclasts and inhibits nuclear factor of activated T cells and mitogen‐activated protein kinase activation. These results suggest that CDP may represent a candidate drug for the treatment of osteoporosis caused by excessive osteoclast activity. CDP suppresses RANKL‐induced ROS production, as well as NFATc1 and MAPK activation, hence inhibits osteoclastogenesis. [ABSTRACT FROM AUTHOR]

The dynamic balance between bone resorption and bone formation is crucial to maintain bone mass. Osteoclasts are key cells that perform bone resorption while osteoblasts and osteocytes function in bone formation. Osteoporosis, a bone metabolism disease characterized by bone loss and degradation of bone microstructure, occurs when osteoclastic bone resorption outstrips osteoblastic bone synthesis. The interaction between receptor activator of nuclear factor κB ligand (RANKL) and RANK on the surface of bone marrow macrophages promotes osteoclast differentiation and activation. In this study, we found that lumichrome, a photodegradation product of riboflavin, inhibits RANKL‐induced osteoclastogenesis and bone resorption as determined by tartrate‐resistant acid phosphatase staining, immunofluorescence, reverse transcription‐polymerase chain reaction, and western blot. Our results showed that lumichrome represses the expression of osteoclast marker genes, including cathepsin K (Ctsk) and Nfatc1. In addition, lumichrome suppressed RANKL‐induced calcium oscillations, NFATc1, NF‐κB, and MAPK signaling activation. Moreover, lumichrome promoted osteoblast differentiation at an early stage, as demonstrated by upregulated expression of osteoblast marker genes Alp, Runx2, and Col1a1. We also found that lumichrome reduces bone loss in ovariectomized mice by inhibiting osteoclastogenesis. In summary, our data suggest the potential of lumichrome as a therapeutic drug for osteolytic diseases. [ABSTRACT FROM AUTHOR]

Background/Aims: Extensive osteoclast formation plays a critical role in bone diseases, including rheumatoid arthritis, periodontitis and the aseptic loosening of orthopedic implants. Thus, identification of agents that can suppress osteoclast formation and bone resorption is important for the treatment of these diseases. Monocrotaline (Mon), the major bioactive component of crotalaria sessiliflora has been investigated for its anti-cancer activities. However, the effect of Mon on osteoclast formation and osteolysis is not known. Methods: The bone marrow macrophages (BMMs) were cultured with M-CSF and RANKL followed by Mon treatment. Then the effects of Mon on osteoclast differentiation were evaluated by counting TRAP (+) multinucleated cells. Moreover, effects of Mon on hydroxyapatite resorption activity of mature osteoclast were studied through resorption areas measurement. The involved potential signaling pathways were analyzed by performed Western blotting and quantitative real-time PCR examination. Further, we established a mouse calvarial osteolysis model to measure the osteolysis suppressing effect of Mon in vivo. Results: In this study, we show that Mon can inhibit RANKL-induced osteoclast formation and function in a dose-dependent manner. Mon inhibits the expression of osteoclast marker genes such as tartrate-resistant acid phosphatase (TRAP) and cathepsin K. Furthermore, Mon inhibits RANKL-induced the activation of p38 and JNK. Consistent with in vitro results, Mon exhibits protective effects in an in vivo mouse model of LPS-induced calvarial osteolysis. Conclusion: Taken together our data demonstrate that Mon may be a potential prophylactic anti-osteoclastic agent for the treatment of osteolytic diseases caused by excessive osteoclast formation and function. [ABSTRACT FROM AUTHOR]

Over-production and activation of osteoclasts is a common feature of osteolytic conditions such as osteoporosis, tumor-associated osteolysis, and inflammatory bone erosion. Cyanidin Chloride, a subclass of anthocyanin, displays antioxidant and anti-carcinogenesis properties, but its role in osteoclastic bone resorption and osteoporosis is not well understood. In this study, we showed that Cyanidin Chloride inhibits osteoclast formation, hydroxyapatite resorption, and receptor activator of NF-κB ligand (RANKL)-induced osteoclast marker gene expression; including ctr, ctsk, and trap. Further investigation revealed that Cyanidin Chloride inhibits RANKL-induced NF-κB activation, suppresses the degradation of IκB-α and attenuates the phosphorylation of extracellular signal-regulated kinases (ERK). In addition, Cyanidin Chloride abrogated RANKL-induced calcium oscillations, the activation of nuclear factor of activated T cells calcineurin-dependent 1 (NFATc1), and the expression of c-Fos. Further, we showed that Cyanidin Chloride protects against ovariectomy-induced bone loss in vivo. Together our findings suggest that Cyanidin Chloride is capable of inhibiting osteoclast formation, hydroxyapatite resorption and RANKL-induced signal pathways in vitro and OVX-induced bone loss in vivo, and thus might have therapeutic potential for osteolytic diseases. [ABSTRACT FROM AUTHOR]

Bone destruction or osteolysis marked by excessive osteoclastic bone resorption is a very common medical condition. Identification of agents that can effectively suppress excessive osteoclast formation and function is crucial for prevention and treatment of osteolytic conditions such as periprosthetic joint infection and periprosthetic loosening. Luteoloside, a flavonoid, is a natural bioactive compound with anti-inflammation and anti-tumor properties. However, the effect of Luteoloside on inflammation-induced osteolysis is unknown. Here, we found that Luteoloside exhibited a strong inhibitory effect on lipopolysaccharide (LPS)-induced osteolysis in vivo. In addition, Luteoloside suppressed RANKL-induced osteoclast differentiation and abrogated bone resorption in a dose-dependent manner. Further, we found that the anti-osteoclastic and anti-resorptive actions of Luteoloside are mediated via blocking NFATc1 activity and the attenuation of RANKL-mediated Ca2+ signaling as well as NF-κB and MAPK pathways. Taken together, this study shows that Luteoloside may be a potential therapeutic agent for osteolytic bone diseases associated with abnormal osteoclast formation and function in inflammatory conditions. [ABSTRACT FROM AUTHOR]

ABSTRACT Osteoporosis is an osteolytic disease that features enhanced osteoclast formation and bone resorption. Identification of agents that can inhibit osteoclast formation and function is important for the treatment of osteoporosis. Dihydroartemisinin is a natural compound used to treat malaria but its role in osteoporosis is not known. Here, we found that dihydroartemisinin can suppress RANKL-induced osteoclastogenesis and bone resorption in a dose-dependent manner. Dihydroartemisinin inhibited the expression of osteoclast marker genes such as cathepsin K, calcitonin receptor, and tartrate-resistant acid phosphatase (TRAcP). Furthermore, dihydroartemisinin inhibited RANKL-induced NF-κB and NFAT activity. In addition, using an in vivo ovariectomized mouse model, we show that dihydroartemisinin is able to reverse the bone loss caused by ovariectomy. Together, this study shows that dihydroartemisinin attenuates bone loss in ovariectomized mice through inhibiting RANKL-induced osteoclast formation and function. This indicates that dihydroartemisinin, the first physiology or medicine nobel prize discovery of China, is a potential treatment option against osteolytic bone disease. © 2015 American Society for Bone and Mineral Research. [ABSTRACT FROM AUTHOR]

Highlights: [•] HtrA1 is secreted by osteoclasts. [•] HtrA1 inhibits osteoblast differentiation. [•] HtrA1 inhibits osteoblast activities through the inhibition of BMP2-induced signaling. [•] HtrA1 is identified as a new osteoclast-derived factor regulating osteoblasts. [Copyright &y& Elsevier]

Osteoclastic bone resorption requires strict interplay between acidified carrier vesicles, motor proteins, and the underlying cytoskeleton in order to sustain the specialized structural and functional polarization of the ruffled border. Cytoplasmic dynein, a large processive mechanochemical motor comprising heavy, intermediate, and light chains coupled to the dynactin cofactor complex, powers unilateral motility of diverse cargos to microtubule minus-ends. We have recently shown that regulators of the dynein motor complex constitute critical components of the osteoclastic bone resorptive machinery. Here, by selectively modulating endogenous dynein activity, we show that the integrity of the dynein-dynactin motor complex is an essential requirement for both osteoclast formation and function. Systematic dissection of the osteoclast dynein-dynactin complex revealed that it is differentially localized throughout RANKL-induced osteoclast formation and activation, undergoing microtubule-coupled reorganization upon the establishment of cellular polarization. In osteoclasts actively resorbing bone, dynein-dynactin intimately co-localizes with the CAP-Gly domain-containing microtubule plus-end protein CLIP-170 at the resorptive front, thus orientating the ruffled border as a microtubule plus-end domain. Unexpectedly, disruption of the dynein-dynactin complex by exogenous p50/dynamitin expression retards osteoclast formation in vitro, owing largely to prolonged mitotic stasis of osteoclast progenitor cells. More importantly, loss of osteoclastic dynein activity results in a drastic redistribution of key intracellular organelles, including the Golgi and lysosomes, an effect that coincides with impaired cathepsin K secretion and diminished bone resorptive function. Collectively, these data unveil a previously unrecognized role for the dynein-dynactin motor complex in osteoclast formation and function, serving not only to regulate their timely maturation but also the delivery of osteolytic cargo that is essential to the bone resorptive process. © 2013 American Society for Bone and Mineral Research [ABSTRACT FROM AUTHOR]

Reactive oxidative species (ROS) generation triggers pyroptosis and induces development of inflammatory osteolysis. Hecogenin (HG) has anti-inflammatory and antioxidative property, but its effects on inflammatory osteolysis remains unclear. In our study, we investigated the mechanism of HG on pyroptosis and its effect on inflammatory osteolysis in vitro and in vivo. The impact of HG on osteoclastogenesis was evaluated using cytotoxicity, TRAcP staining and bone resorption assays. The RNA-sequencing was employed to identify potential signaling pathways, and then RT-qPCR, western blot, immunofluorescence, and ELISA were used to verify. To determine the protective effect of HG in vivo , Lipopolysaccharide (LPS)-induced animal models were utilized, along with micro-CT and histological examination. HG suppressed RANKL-induced osteoclast differentiation, bone resorption, NFATc1 activity and downstream factors. RNA-sequencing results showed that HG inhibited osteoclastogenesis by modulating the inflammatory response and macrophage polarization. Furthermore, HG inhibited the NF-κB pathway, and deactivated the NLRP3 inflammasome. HG activated the expression of nuclear factor E2-related factor 2 (Nrf2) to eliminate ROS generation. Importantly, the inhibitory effect of HG on NLRP3 inflammasome could be reversed by treatment with the Nrf2 inhibitor ML385. In viv o, HG prevented the mice against LPS-induced osteolysis by suppressing osteoclastogenesis and inflammatory factors. In conclusion, HG could activate Nrf2 to eliminate ROS generation, inactivate NLRP3 inflammasome and inhibit pyroptosis, thereby suppressing osteoclastogenesis in vitro and alleviating inflammatory osteolysis in vivo, which indicating that HG might be a promising candidate to treat inflammatory osteolysis. [Display omitted] • HG suppressed osteoclast formation and resorption function. • HG modulated the inflammatory response and macrophage polarization. • HG restrained pyroptosis by inactivating NLRP3 inflammasome. • HG suppressed NLRP3 inflammasome involved Nrf2 activation • HG exhibited protective effects against LPS-induced osteolysis in vivo. [ABSTRACT FROM AUTHOR]

Abstract: Advanced glycation end products (AGEs) have been observed to accumulate in bone with increasing age and may impose effects on bone resorption activities. However, the underlying mechanism of AGEs accumulation in bone is still poorly understood. In this study, human cortical bone specimens from young (31±6years old), middle-aged (51±3years old) and elderly (76±4years old) groups were examined to determine the spatial–temporal distribution of AGEs in bone matrix and its effect on bone resorption activities by directly culturing osteoclastic cells on bone slices. The results of this study indicated that the fluorescence intensity (excitation wave length 360nm and emission wave length 470±40nm) could be used to estimate the relative distribution of AGEs in bone (pentosidine as its marker) under an epifluorescence microscope. Using the fluorescence intensity as the relative measure of AGEs concentration, it was found that the concentration of AGEs varied with biological tissue ages, showing the greatest amount in the interstitial tissue, followed by the old osteons, and the least amount in newly formed osteons. In addition, AGEs accumulation was found to be dependent on donor ages, suggesting that the younger the donor the less AGEs were accumulated in the tissue. Most interestingly, AGEs accumulation appeared to initiate from the region of cement lines, and spread diffusively to the other parts as the tissue aged. Finally, it was observed that the bone resorption activities of osteoclasts were positively correlated with the in situ concentration of AGEs and such an effect was enhanced with increasing donor age. These findings may help elucidate the mechanism of AGEs accumulation in bone and its association with bone remodeling process. [Copyright &y& Elsevier]

[Display omitted] Reactive Oxygen Species (ROS) play an essential role in the pathogenesis of osteoporosis mainly characterized by excessive osteoclasts (OCs) activity. OCs are rich in mitochondria for energy support, which is a major source of total ROS. Tussilagone (TSG), a natural Sesquiterpenes from the flower of Tussilago farfara , has plentiful beneficial pharmacological characteristics with anti-inflammatory and anti-oxidative activity, but its effects and mechanism in osteopathology are still unclear. In our study, we investigated the regulation of ROS generated from the mitochondria in OCs. We found that TSG inhibited OCs differentiation and bone resorption without any cytotoxicity. Mechanistically, TSG reduced RANKL-mediated total ROS level by down-regulating intracellular ROS production and mitochondrial function, leading to the suppression of NFATc1 transcription. We also found that nuclear factor erythroid 2-related factor 2 (Nrf2) could enhance ROS scavenging enzymes in response to RANKL-induced oxidative stress. Furthermore, TSG up-regulated the expression of Nrf2 by inhibiting its proteosomal degradation. Interestingly, Nrf2 deficiency reversed the suppressive effect of TSG on mitochondrial activity and ROS signaling in OCs. Consistent with this finding, TSG attenuated post-ovariectomy (OVX)- and lipopolysaccharide (LPS) induced bone loss by ameliorating osteoclastogenesis. Taken together, TSG has an anti-bone resorptive effect by modulating mitochondrial function and ROS production involved Nrf2 activation. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Osteoporosis is a chronic bone lytic disease, because of inadequate bone ossification and/or excessive bone resorption. Even though drugs are currently available for the treatment of osteoporosis, there remains an unmet need for the development of more specific novel agents with less adverse effects. Dehydrocostus lactone (DHC), a natural sesquiterpene lactone, was previously found to affect the differentiation of inflammatory cells by inhibiting NF-κB pathways, and garnered much interest for its anti-cancer properties via SOCS-mediated cell cycle arrest and apoptosis. As NF-κB pathway plays an essential role in osteoclast differentiation, we sought to discover the biological effects of DHC on osteoclast differentiation and resorptive activity, as well as the underlying mechanisms on these effects. Our research found that DHC inhibited RANKL-induced osteoclast differentiation, bone resorption and osteoclast specific genes expression via suppression of NF-κB and NFAT signaling pathways in vitro. We further demonstrated that DHC protected against ovariectomy (OVX)-induced bone loss in mice and the protective effect was mediated at least in part through the attenuation of NF-κB signaling pathway. Thus, this study provides insight that DHC might be used as a potential pharmacological treatment for osteoporosis. [ABSTRACT FROM AUTHOR]

Osteoporosis, as a severe public health problem worldwide, causes systemic damage to bone mass, strength, and microstructure with an increased propensity for fragility fractures. Given the inherent adverse effects associated with long-term use of current prescription medications for osteoporosis treatment, identifying natural alternatives to existing treatment methods is imperative. Pteryxin (PTX), a natural coumarin, is isolated from the Peucedanum species belonging to the family Apiaceae. PTX has been reported to have antioxidant, anti-inflammatory and anti-obesity properties. However, its effect on osteoporosis has not been clarified. Our study confirmed that PTX could attenuate the formation of osteoclasts and bone resorption on a dose-dependent basis in vitro. Consistently, in vivo ovariectomy (OVX)-induced osteoporosis models simulating the physiological characteristics of postmenopausal women showed that PTX could partially reverse the bone loss caused by OVX. Further study of its mechanism revealed that PTX might block the MAPK and Ca2+-calcineurin-NFATc1 signaling pathways by decreasing the reactive oxygen species (ROS) level in osteoclasts to dampen the expression of critical transcriptional NFATc1 and downstream osteoclast-specific genes. Overall, PTX may present a new or alternative treatment option for osteoporosis. [Display omitted] • Intracellular ROS play an important role in regulating osteoclast differentiation. • ROS could affect the level of intracellular Ca2+ in osteoclasts. • Pteryxin suppresses osteoclast ogenesis by inhibiting MAPK/Ca2+ signaling pathways via decreasing intracellular ROS levels. • Pteryxin plays a therapeutic effect on ovariectomy-induced osteoporosis in mice. [ABSTRACT FROM AUTHOR]