Antioxidant enzyme Prdx1 inhibits osteoclastogenesis via suppressing ROS and NFATc1 signaling pathways.

Bone is a dynamic organ which continuously undergoes remodeling throughout one's lifetime. Cellular production of reactive oxygen species (ROS) is essential for regulating bone homeostasis. Osteoclasts, multinucleated giant cells differentiated from macrophage lineage, are responsible for osteolytic bone conditions which are closely linked to ROS signaling pathways. In this study, an anti-ROS enzyme, peroxiredoxin 1 (Prdx1) was found to be expressed both in bone marrow macrophages and osteoclasts. Recombinant Prdx1 protein was found to dose-dependently inhibit ROS production and osteoclast differentiation. Mechanistically, Prdx1 protein also attenuated NFATc1 activation as well as the expression of C-Fos, V-ATPase-d2, Cathepsin K, and Integrin αV. Collectively, Prdx1 is a negative regulator on osteoclast formation via inhibiting RANKL-mediated ROS activity, thus suggesting its potential application for treating osteoclast related disorders.

Cichoric acid targets RANKL to inhibit osteoclastogenesis and prevent ovariectomy-induced bone loss.

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.

UPLC/Q-TOF-MS-based Metabolomics Study of the Antiosteoporosis Effects of Vaccarin in Ovariectomized Mice.

Osteoporosis is a systemic and metabolic bone disease that usually occurs in postmenopausal women, which mainly manifests as bone loss and increased bone fragility that both facilitate fracture. However, few drugs for osteoporosis have shown good efficacy and limited side effects. Vaccarin has demonstrated its antiosteoporosis effects by inhibiting the formation and osteolytic activities of osteoclasts in our previous investigation. In this study, multivariate statistical analysis and ultrahigh-performance liquid chromatography and quadrupole time-of-flight tandem mass spectrometry were used to analyze the serum metabolites of ovariectomized mice treated with or without vaccarin. As a result, 9 serum metabolites were identified as biomarkers. The metabolic levels of 3 crucial biomarkers, namely, lysophosphatidylcholine [22 : 6, (4Z, 7Z, 10Z, 13Z, 16Z, 19Z)], 1-linoleoylglycerophosphocholine and 1-palmitoyl-Sn-glycero-3-phosphocholine, that were correlated with glycerophospholipid metabolism increased and then decreased significantly after vaccarin treatment. Molecular docking analysis and osteoclasts differentiation experiment further revealed that vaccarin may bind with phospholipase A2 and downregulated its activity to reduce the osteoclastogenesis. Therefore, the occurrence of osteoporosis is closely related with glycerophospholipid metabolism disorders, and vaccarin exerts antiosteoporosis effects by reducing the levels of glycerophospholipid metabolites.

UPLC/Q-TOF-MS-based metabolomics evaluate the efficacy of oroxylin A against postmenopausal osteoporosis.

Post-menopausal osteoporosis (PMOP) is a common metabolic bone malady characterized by bone mass loss and bone microarchitectural deterioration; however, there is currently no effective drug for its management. According to our previous study, oroxylin A (OA) could effectively protect ovariectomized (OVX)-osteoporotic mice from bone loss; however, its therapeutic targets are still unclear. From a metabolomic perspective, we studied serum metabolic profiles to discover potential biomarkers and OVX-related metabolic networks, which could assist us to comprehend the impact of OA on OVX. Five metabolites were identified as biomarkers associated with 10 related metabolic pathways, including phenylalanine, tyrosine and tryptophan biosynthesis, and phenylalanine, tryptophan and glycerophospholipid metabolism. After OA treatment, the expression of multiple biomarkers changed, with lysophosphatidylcholine (18:2) being a major significantly regulated biomarker. Our study demonstrated that OA's effects on OVX are probably related to the regulation of phenylalanine, tyrosine and tryptophan biosynthesis. Our findings explain the role of OA against PMOP in terms of metabolism and pharmacology and provide a pharmacological foundation for OA treatment of PMOP.

Inhibitory effects of biochanin A on titanium particle-induced osteoclast activation and inflammatory bone resorption via NF-κB and MAPK pathways.

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-1ß 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.

Ellagic acid protects ovariectomy-induced bone loss in mice by inhibiting osteoclast differentiation and bone resorption.

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.

Cumambrin A prevents OVX-induced osteoporosis via the inhibition of osteoclastogenesis, bone resorption, and RANKL signaling pathways.

Being the principal cells responsible for bone resorption and pathologic bone loss, osteoclasts have become the main target for antiresorptive treatment. Cumambrin A is a natural compound isolated from Chrysanthemum indicum L. and belongs to a member of the sesquiterpene lactone family. To date, the therapeutic effect of cumambrin A on osteoporosis and its mechanisms of action are not known. In this study, we found that cumambrin A can significantly inhibit osteoclast formation and bone resorption through the suppression of receptor activator of NF-κB ligand (RANKL)-induced NF-κB and nuclear factor of activated T-cell activity and ERK phosphorylation. Furthermore, cumambrin A inhibits the expression of osteoclast marker genes including cathepsin K, calcitonin receptor, and V-ATPase d2. Using an in vivo ovariectomized mouse model, we showed that cumambrin A protects against estrogen withdrawal-induced bone loss. Collectively, our results reveal that cumambrin A can suppress osteoclast formation, bone resorption, and RANKL-induced signaling pathways, suggesting that cumambrin A is a potential therapeutic agent for the treatment of osteoporosis.-Zhou, L., Liu, Q., Hong, G., Song, F., Zhao, J., Yuan, J., Xu, J., Tan, R. X., Tickner, J., Gu, Q., Xu, J. Cumambrin A prevents OVX-induced osteoporosis via the inhibition of osteoclastogenesis, bone resorption, and RANKL signaling pathways.

Retraction of: A005, a novel inhibitor of phosphatidylinositol 3-kinase/mammalian target of rapamycin, prevents osteosarcoma-induced osteolysis.

Osteosarcoma is the most frequent primary bone tumor in children and adolescents. The phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling pathway is an attractive anticancer target because it plays key roles in the regulation of cell growth, division and differentiation. In this study, we demonstrated high expression of PI3K/mTOR signaling pathway-related genes in patients with osteosarcoma. We thus investigated the effects of A005, a newly synthesized dual PI3K/mTOR inhibitor, on osteosarcoma cells and in a mouse xenograft tumor model. The results confirmed that A005 inhibited the proliferation, migration and invasion of human osteosarcoma cells. In addition, A005 also inhibited receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclast differentiation and bone resorption in vitro. Therefore, A005 was further applied to a SaOS-2 osteosarcoma-induced mouse osteolysis model. A005 inhibited tumor growth and prevented osteosarcoma-associated osteolysis via modulation of the PI3K/AKT/mTOR pathway. Overall, our results showed that A005 inhibited osteoclastogenesis and prevented osteosarcoma-induced bone osteolysis by suppressing PI3K/AKT/mTOR signaling. These findings indicated that A005 may be a promising candidate drug for the treatment of human osteosarcoma.

Tiliroside is a new potential therapeutic drug for osteoporosis in mice.

Osteoporosis is a class of metabolic bone disease caused by complexed ramifications. Overactivation of osteoclasts due to a sudden decreased estrogen level plays a pivotal role for postmenopausal women suffering from osteoporosis. Therefore, inhibiting osteoclast formation and function has become a major direction for the treatment of osteoporosis. Tiliroside (Tle) is a salutary dietary glycosidic flavonoid extracted from Oriental Paperbush flower, which has been reported to have an anti-inflammation effect. However, whether Tle affects the osteoclastogenesis and bone resorption remains unknown. Herein, we demonstrate that Tle prevents bone loss in ovariectomy in mice and inhibits osteoclast differentiation and bone resorption stimulated by receptor activator of nuclear factor-κB ligand (RANKL) in vitro. Molecular mechanism studies reveal that Tle reduces RANKL-induced activation of mitogen-activated protein kinase and T-cell nuclear factor 1 pathways, and osteoclastogenesis-related marker gene expression, including cathepsin K (Ctsk), matrix metalloproteinase 9, tartrate-resistant acid phosphatase (Acp5), and Atp6v0d2. Our research indicates that Tle suppresses osteoclastogenesis and bone loss by downregulating the RANKL-mediated signaling protein activation and expression. In addition, Tle inhibits intracellular reactive oxygen species generation which is related to the formation of osteoclasts. Therefore, Tle might serve as a potential drug for osteolytic disease such as osteoporosis.

Pectolinarigenin prevents bone loss in ovariectomized mice and inhibits RANKL-induced osteoclastogenesis via blocking activation of MAPK and NFATc1 signaling.

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.

Salidroside promotes rat spinal cord injury recovery by inhibiting inflammatory cytokine expression and NF-κB and MAPK signaling pathways.

Spinal cord injury (SCI) is a public health problem in the world. The SCI usually triggers an excessive inflammatory response that brings about a secondary tissue wreck leading to further cellular and organ dysfunction. Hence, there is great potential of reducing inflammation for therapeutic strategies of SCI. In this study, we aim to investigate if Salidroside (SAD) exerts an anti-inflammatory effect and promotes recovery of motor function on SCI through suppressing nuclear factor-κB (NF-κB) and the mitogen-activated protein kinase (MAPK) pathways. In vitro, real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) were used to examine the inhibitory effect of SAD on the expression and release of interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) activated by lipopolysaccharide (LPS) in astrocytes. In addition, SAD was found to inhibit NF-κB, p38 and extracellular-regulated protein kinases (ERK) signaling pathways by western blot analysis. Further, in vivo study showed that SAD was able to improve hind limb motor function and reduce tissue damage accompanied by the suppressed expression of inflammatory cytokines IL-1ß, IL-6, and TNF-α. Overall, SAD could reduce the inflammatory response and promote motor function recovery in rats after SCI by inhibiting NF-κB, p38, and ERK signaling pathways.

Scutellarein inhibits RANKL-induced osteoclast formation in vitro and prevents LPS-induced bone loss in vivo.

Osteoporosis, arthritis, Peget's disease, bone tumor, periprosthetic joint infection, and periprosthetic loosening have a common characteristic of osteolysis, which is characterized by the enhanced osteoclastic bone resorptive function. At present, the treatment target of these diseases is to interfere with osteoclastic formation and function. Scutellarein (Scu), a flavonoids compound, can inhibit the progress of tumor and inflammation. However, the role of Scu in inflammatory osteolysis isn't elucidated clearly. Our study showed that Scu inhibited bone destruction induced by LPS in vivo and OC morphology and function induced by RANKL in vitro. Mechanistic studies revealed that Scu suppressed osteoclastic marker gene expression by RANKL-induced, such as Ctsk9, Mmp9, Acp5, and Atp6v0d2. In addition, we found that the inhibition effects of osteoclastogenesis and bone resorption function of Scu were mediated via attenuating NF-κB and NFAT signaling pathways. In conclusion, the results showed that Scu may become a potential new drug for the treatment of inflammatory osteolysis.

Diosmetin inhibits osteoclast formation and differentiation and prevents LPS-induced osteolysis in mice.

Osteolytic bone diseases are closely linked to the over-activation of osteoclasts and enhancement of bone resorption. It has become a major health issue in orthopedic practice worldwide. Inhibition of osteoclasts is proposed to be the main treatment for osteolytic disorders. Diosmetin (DIO) is a natural flavonoid with properties of antioxidant, anti-infection, and antishock. The effect of DIO on osteoclast differentiation is poorly understood. In this study project, we found that DIO could inhibit osteoclastic formation induced by receptor activator of nuclear factor kappa-B ligand (RANKL) in a dose-dependent manner. The expression of the osteoclast differentiation marker genes, cathepsin K, nuclear factor of activated T-cells 1 (NFATc1), Acp5, Ctr, Atp6v0d2, and Mmp9 were also decreased by the treatment of DIO. In addition, DIO attenuated the formation of actin ring and the ability of bone resorption. Further, the western blotting showed that DIO inhibits the phosphorylation of the mitogen-activated protein kinases signaling pathway induced by RANKL, accompanied by the downregulation of NFATc1 and c-Fos expression. We also found that DIO could reduce the accumulation of reactive oxygen species (ROS) induced by RANKL. In vivo, the study revealed that DIO can significantly reduce LPS-induced osteolysis in mice. Collectively, our study shows that DIO can inhibit osteoclast formation and activation, and could serve as a potential therapeutic drug for osteolytic bone diseases.

Vaccarin prevents titanium particle-induced osteolysis and inhibits RANKL-induced osteoclastogenesis by blocking NF-κB and MAPK signaling pathways.

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.

Daphnetin attenuates LPS-induced osteolysis and RANKL mediated osteoclastogenesis through suppression of ERK and NFATc1 pathways.

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.

Cepharanthine suppresses osteoclast formation by modulating the nuclear factor-κB and nuclear factor of activated T-cell signaling pathways.

The increased activation of osteoclasts is the major manifestation of several lytic bone diseases, including osteoporosis, rheumatoid arthritis, aseptic loosening of orthopedic implants, Paget disease and malignant bone diseases. One important bone-protective therapy in these diseases focuses on the inhibition of osteoclast differentiation and resorptive function. Given that the deleterious side-effects of currently available drugs, it is beneficial to search for effective and safe medications from natural compounds. Cepharanthine (CEP) is a compound extracted from Stephania japonica and has been found to have antioxidant and anti-inflammatory effects. In this study, we found that CEP inhibited receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclast formation and bone-resorbing activities using osteoclastogenesis and bone resorption assay. By polymerase chain reaction, we also found that CEP inhibited the expression of osteoclast-differentiation marker genes including Ctsk, Calcr, Atp6v0d2, Mmp9 and Nfatc1. Mechanistic analyses including Western blot and luciferase reporter assay revealed that CEP inhibited RANKL-induced activation of NF-κB and nuclear factor of activated T-cell, which are essential for the formation of osteoclast. Collectively, these data suggested that CEP can potentially be used as an alternative therapy for preventing or treating osteolytic diseases.

Luteoloside prevents lipopolysaccharide-induced osteolysis and suppresses RANKL-induced osteoclastogenesis through attenuating RANKL signaling cascades.

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.

Artesunate inhibits RANKL-induced osteoclastogenesis and bone resorption in vitro and prevents LPS-induced bone loss in vivo.

Osteoclasts are multinuclear giant cells responsible for bone resorption in lytic bone diseases such as osteoporosis, arthritis, periodontitis, and bone tumors. Due to the severe side-effects caused by the currently available drugs, a continuous search for novel bone-protective therapies is essential. Artesunate (Art), the water-soluble derivative of artemisinin has been investigated owing to its anti-malarial properties. However, its effects in osteoclastogenesis have not yet been reported. In this study, Art was shown to inhibit the nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis, the mRNA expression of osteoclastic-specific genes, and resorption pit formation in a dose-dependent manner in primary bone marrow-derived macrophages cells (BMMs). Furthermore, Art markedly blocked the RANKL-induced osteoclastogenesis by attenuating the degradation of IκB and phosphorylation of NF-κB p65. Consistent with the in vitro results, Art inhibited lipopolysaccharide (LPS)-induced bone resorption by suppressing the osteoclastogenesis. Together our data demonstrated that Art inhibits RANKL-induced osteoclastogenesis by suppressing the NF-κB signaling pathway and that it is a promising agent for the treatment of osteolytic diseases.

Monocrotaline Suppresses RANKL-Induced Osteoclastogenesis In Vitro and Prevents LPS-Induced Bone Loss In Vivo.

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.