Saikosaponin D Inhibited IL-1ß Induced ATDC 5 Chondrocytes Apoptosis In Vitro and Delayed Articular Cartilage Degeneration in OA Model Mice In Vivo.

Osteoarthritis (OA) is the most common joint disease in the elderly, characterized by cartilage degradation and proliferation of subchondral bone. The pathogenesis of OA involves a variety of inflammatory mediators, including nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor (TNF)-α, and interleukin (IL)-1ß. From the molecular mechanism, the nuclear factor-erythroid 2-related factor (Nrf2)/heme oxygenase-1 (HO-1) pathway and the expression of ROS regulated the production of the above inflammatory mediators. Saikosaponin D (SSD), which is an active ingredient isolated from Bupleurum, has various biological functions. In this study, IL-1ß was used as a pro-inflammatory factor to create an in vitro OA model. According to the results of high-density culture, qPCR, ROS measurement, Western blot, and immunofluorescence, SSD activated the Nrf2/HO-1/ROS axis, inhibited the production of inflammatory mediators, and protected against ECM destruction. The DMM mouse model was used as a model of OA in mice. From the results of safranin O/fast green staining, hematoxylin-eosin staining, tartrate-resistant acid phosphatase (TRAP) staining, and OARSI scores, SSD protected against the mice knee articular cartilage degeneration and reduced the number of osteoclasts in the subchondral bone. Experimental results found that SSD suppressed IL-1ß-induced differentiated ATDC 5 chondrocytes apoptosis via the Nrf2/HO-1/ROS axis in vitro. SSD delayed the progression of OA in DMMs model mice in vivo. Therefore, SSD has the potential to become a drug for clinical treatment of OA.

Inhibition of Lipopolysaccharide-Induced Inflammatory Bone Loss by Saikosaponin D is Associated with Regulation of the RANKL/RANK Pathway.

BACKGROUND: Osteolytic diseases such as osteoporosis are featured with accelerated osteoclast differentiation and strong bone resorption. Considering the complications and other limitations of current drug treatments, it is necessary to develop a safer and more reliable drug to deal with osteoclast-related diseases. Saikosaponin D (SSD) is the active extract of Bupleurum, which has anti-inflammation, anti-tumor and liver protection functions. However, the role of SSD in regulating the differentiation and function of osteoclasts is not clear. PURPOSE: To explore whether SSD could prevent osteoclast differentiation and bone resorption induced by M-CSF and RANKL, and further evaluate the potential therapeutic properties of SSD in LPS-induced inflammatory bone loss mouse models. METHODS: BMMs were cultured in complete medium stimulated by RANKL with different concentrations of SSD. TRAP staining, bone resorption determination, qRT-PCR, immunofluorescence and Western blotting were performed. A mouse model of LPS-induced calvarial bone loss was established and treated with different doses of SSD. The excised calvaria bones were used for TRAP staining, micro-CT scan and histological analysis. RESULTS: SSD inhibited the formation and bone resorption of osteoclasts induced by RANKL in vitro. SSD suppressed LPS-induced inflammatory bone loss in vivo. CONCLUSION: SSD inhibited osteoclastogenesis and LPS-induced osteolysis in mice both which served as a new potential agent for the treatment of osteoclast-related conditions.

TAZ inhibits osteoclastogenesis by attenuating TAK1/NF-κB signaling.

Osteoporosis is an osteolytic disorder commonly associated with excessive osteoclast formation. Transcriptional coactivator with PDZ-binding motif (TAZ) is a key downstream effector of the Hippo signaling pathway; it was suggested to be involved in the regulation of bone homeostasis. However, the exact role of TAZ in osteoclasts has not yet been established. In this study, we demonstrated that global knockout and osteoclast-specific knockout of TAZ led to a low-bone mass phenotype due to elevated osteoclast formation, which was further evidenced by in vitro osteoclast formation assays. Moreover, the overexpression of TAZ inhibited RANKL-induced osteoclast formation, whereas silencing of TAZ reduced it. Mechanistically, TAZ bound to TGF-activated kinase 1 (TAK1) and reciprocally inhibited NF-κB signaling, suppressing osteoclast differentiation. Collectively, our findings highlight an essential role of TAZ in the regulation of osteoclastogenesis in osteoporosis and its underlying mechanism.

Pristimerin Inhibits Osteoclast Differentiation and Bone Resorption in vitro and Prevents Ovariectomy-Induced Bone Loss in vivo.

INTRODUCTION: Osteoporosis is a metabolic bone disease characterized by reduced bone quantity and microstructure, typically owing to increased osteoclastogenesis and/or enhanced osteoclastic bone resorption, resulting in uncontrolled bone loss, which primarily affects postmenopausal women. In consideration of the severe side effects of current drugs for osteoporosis, new safe and effective medications are necessary. Pristimerin (Pri), a quinone methide triterpene extracted from Celastraceae and Hippocrateaceae members, exhibits potent antineoplastic and anti-inflammatory effects. However, its effect on osteoclasts remains unknown. MATERIALS AND METHODS: We evaluated the anti-osteoclastogenic and anti-resorptive effect of Pri on bone marrow-derived osteoclasts and its underlying mechanism in vitro. In addition, the protective effect of Pri on ovariectomy model was also explored in vivo. RESULTS: In vitro, Pri inhibited osteoclast differentiation and mature osteoclastic bone resorption in a time- and dose-dependent manner. Further, Pri suppressed the expression of osteoclast-related genes and the activation of key proteins. Pri also inhibited the early activation of ERK, JNK MAPK, and AKT signaling pathways in bone marrow-derived macrophages (BMMs), ultimately inhibiting the induction and activation of the crucial osteoclast transcriptional factor nuclear factor of activated T-cell cytoplasmic 1 (NFATc1). In vivo, consistent with our in vitro data, Pri clearly prevented ovariectomy-induced bone loss. CONCLUSION: Our data showed that Pri inhibits the differentiation and activation of osteoclasts in vitro and in vivo, and could be a promising candidate for treating osteoporosis.

Sarsasapogenin Suppresses RANKL-Induced Osteoclastogenesis in vitro and Prevents Lipopolysaccharide-Induced Bone Loss in vivo.

INTRODUCTION: Osteoclasts are giant polynuclear cells; their main function is bone resorption. An increased number of osteoclasts and enhanced bone resorption exert significant effects on osteoclast-related bone-lytic diseases, including osteoporosis. Given the limitations of current therapies for osteolytic diseases, it is urgently required to develop safer and more effective alternatives. Sarsasapogenin, a major sapogenin from Anemarrhena asphodeloides Bunge, possesses potent antitumor effects and inhibits NF-κB and MAPK signaling. However, the manner in which it affects osteoclasts is unclear. METHODS: We investigated the effects of anti-osteoclastogenic and anti-resorptive of sarsasapogenin on bone marrow-derived osteoclasts. RESULTS: Sarsasapogenin inhibited multiple RANKL-induced signaling cascades, thereby inhibiting the induction of key osteoclast transcription factor NFATc1. The in vivo and in vitro results were consistent: sarsasapogenin treatment protected against bone loss in a mouse osteolysis model induced by lipopolysaccharide. CONCLUSION: Our research confirms that sarsasapogenin can be used as a new treatment for osteoclast-related osteolytic diseases.

Correction to: Garcinol Suppresses IL-1ß-Induced Chondrocyte Inflammation and Osteoarthritis via Inhibition of the NF-κB Signaling Pathway.

The original version of this article contained mistakes, and the authors would like to correct them.

Disulfiram suppressed ethanol promoted RANKL-induced osteoclastogenesis in vitro and ethanol-induced osteoporosis in vivo via ALDH1A1-NFATc1 axis.

Excessive alcohol consumption is positively related to osteoporosis, and its treatment strategies are poorly developed. Disulfiram inhibits receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis; however, whether it can be used for ethanol-induced osteoclastogenesis and its underlying mechanism are still unclear. In this study, we demonstrated that ethanol promoted RANKL-induced osteoclast formation and bone resorption, whereas, disulfiram suppressed ethanol-induced osteoclastogenesis by abrogating the expression of nuclear factor of activated T cell c1 (NFATc1) in vitro. Further analysis revealed that aldehyde dehydrogenase 1A1 (ALDH1A1) is important for the expression of NFATc1, the master regulator of osteoclast differentiation. Furthermore, we showed that disulfiram protected ethanol-induced osteoporosis in vivo. Overall, our study provides promising evidence that disulfiram can be used as a treatment strategy for alcohol-related osteoporosis via the ALDH1A1T-NFATc1 axis.

Garcinol Suppresses IL-1ß-Induced Chondrocyte Inflammation and Osteoarthritis via Inhibition of the NF-κB Signaling Pathway.

Osteoarthritis (OA), which is characterized as a common degenerative joint disease, is presently the most prevalent chronic degenerative joint disease. Accumulating evidence has shown a biological function for Garcinol in a variety of diseases; however, whether it could be used to treat OA remains unclear. In this study, we explored the protective effects of garcinol on the progression of OA and explored the underlying mechanism. In vitro, garcinol reduced the expression of pro-inflammatory cytokines, such as IL-6 and tumor necrosis factor alpha (TNF-α). It also decreased the expression of inducible nitric oxide synthase (iNOS), as well as cyclooxygenase-2 (COX-2). Furthermore, garcinol inhibited the expression of thrombospondin motifs 5(ADAMTS5) and metalloproteinase (MMPs), both of which regulate extracellular matrix degradation. These changes could be attributed to garcinol-related suppression of the IL-1ß-induced NF-κB signaling pathway. Moreover, we investigated the protective effects of garcinol on the surgical destabilization of the medial meniscus (DMM) of the mouse, an in vivo model of OA. Taken together, our data suggest garcinol as a potential future agent for the treatment of OA.

Anacardic acid inhibits RANKL-induced osteoclastogenesis in vitro and prevents ovariectomy-induced bone loss in vivo.

Postmenopausal osteoporosis is the most common form of primary osteoporosis, and the incidence of the condition is rapidly increasing. In consideration of the limitations of current therapeutic options for the treatment of postmenopausal osteoporosis, there is an urgent need to develop safer alternatives. Anacardic acid, a natural phenolic acid compound extracted from cashew nut shell, possesses potent antitumor and anti-inflammatory effects and inhibits NF-κB signaling. However, its effect on osteoclasts remains unknown. This study reports the first evidence for the antiosteoclastogenic and antiresorptive effects of anacardic acid on bone marrow-derived macrophage-derived osteoclasts. Mechanistically, anacardic acid disrupts the phosphorylation of TGF-ß activated kinase 1 and subsequently suppresses multiple receptor activator of NF-κB ligand-induced signaling cascades, ultimately inhibiting the induction and activation of the crucial osteoclast transcriptional factor nuclear factor of activated T-cell cytoplasmic 1. Consistent with cellular results in vitro, anacardic acid treatment improves bone density in the murine model of ovariectomy-induced bone loss. Taken together, our study provides promising evidence for the therapeutic application of anacardic acid as a new potential pharmacological treatment for osteoporosis.-Zhao, K., Jia, Y., Peng, J., Pang, C., Zhang, T., Han, W., Jiang, J., Lu, X., Zhu, J., Qian, Y. Anacardic acid inhibits RANKL-induced osteoclastogenesis in vitro and prevents ovariectomy-induced bone loss in vivo.

Vitexin suppresses RANKL-induced osteoclastogenesis and prevents lipopolysaccharide (LPS)-induced osteolysis.

Osteolytic diseases are characterized by an increase in the number and/or activity of bone-resorbing osteoclasts. Identification of natural compounds that can suppress osteoclast formation and function is crucial for the prevention and treatment of osteolytic diseases. Vitexin, a naturally-derived flavonoid extracted from various medicinal plant species, demonstrates a broad range of pharmacological properties including anticancer and anti-inflammatory effects. Here in this study, we showed that vitexin exerts antiosteoclastogenic effects by directly inhibiting receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation and bone resorption in vitro and protected against lipopolysaccharide (LPS)-induced inflammatory osteolysis in vivo. Vitexin suppressed the early activation of ERK and p38 MAPK pathways in response to RANKL thereby attenuating the downstream induction of c-Fos and NFATc1, and abrogating the expression of osteoclast marker genes. Collectively, these results provide evidence for the therapeutic application of vitexin in the treatment of osteoclast-mediated bone lytic diseases.

Psoralen accelerates bone fracture healing by activating both osteoclasts and osteoblasts.

Bone fracture healing is a complex, dynamic process that involves various cell types, with osteoclasts and osteoblasts playing indispensable roles. In this study, we found that psoralen, the main active ingredient in Psoralea corylifolia L. fruit extract, enhanced bone fracture healing through activation of osteoclast and osteoblast activity via the ERK signaling pathway. In detail, psoralen promoted receptor activator of nuclear factor-κB ligand-induced osteoclastogenesis, mRNA expression of osteoclast-specific genes, and osteoclastic bone resorption in primary bone marrow-derived macrophages. Meanwhile, psoralen induced osteogenic differentiation by promoting the mRNA expression of the osteoblast differentiation markers alkaline phosphatase, runt-related transcription factor 2, osterix, and osteocalcin. At the molecular level, psoralen preferentially activated ERK1/2 but not JNK or p38 MAPKs. Further experiments revealed that psoralen-induced osteoclast and osteoblast differentiation was abrogated by a specific inhibitor of phosphorylated ERK. In addition, psoralen accelerated bone fracture healing in a rat tibial fracture model, and the numbers of osteoclasts and osteoblasts were increased in psoralen-treated fracture callus. Taken together, our findings indicate that psoralen accelerates bone fracture healing through activation of osteoclasts and osteoblasts via ERK signaling and has potential as a novel drug in the orthopedic clinic for the treatment of bone fractures.-Zhang, T., Han, W., Zhao, K., Yang, W., Lu, X., Jia, Y., Qin, A., Qian, Y. Psoralen accelerates bone fracture healing by activating both osteoclasts and osteoblasts.

Deguelin inhibits RANKL-induced osteoclastogenesis in vitro and prevents inflammation-mediated bone loss in vivo.

Excessive bone resorption by osteoclasts (OCs) plays an important role in lytic bone diseases, such as osteoporosis. Although the pharmacological treatment of osteoporosis has been extensively developed, alternative treatments are still needed. Deguelin, a rotenoid isolated from several plant species, is a strong antitumor agent; however, its effect on OCs remains unclear. To the best of our knowledge, this is the first study to report that deguelin inhibits the receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclastogenesis, messenger RNA expression of osteoclastic-specific genes, and osteoclastic bone resorption, in primary bone marrow-derived macrophages. At the molecular level, deguelin markedly blocked RANKL-induced osteoclastogenesis by attenuating the phosphorylation of NF-κB p65 and inhibiting p65 nuclear translocation. In addition, deguelin suppressed the downstream expression of nuclear factor of activated T-cell cytoplasmic 1, which is a crucial transcription factor in OC differentiation. Consistent with the in vitro results, deguelin inhibited lipopolysaccharide-induced bone resorption by suppressing osteoclastogenesis. Taken together, our findings reveal that deguelin has antiosteoclastic effects in vitro and in vivo and possesses potential as a new therapeutic option for osteolytic bone diseases.

Garcinol suppresses RANKL-induced osteoclastogenesis and its underlying mechanism.

Osteoclasts (OCs) are multinuclear giant cells responsible for bone resorption, and an excessive bone resorption by OCs plays an important role in osteoporosis. Commonly used drugs for the treatment of osteoporosis have severe side effects. As such, identification of alternative treatments is essential. Garcinol, a polyisoprenylated benzophenone extracted from the fruit of Garcinia indica, has shown a strong antitumor effect through the nuclear factor-κB (NF-κB) and mitogen-associated protein kinases (MAPK) signaling pathways. However, the role of garcinol in the osteoclastogenesis is still unclear. Here, we demonstrated that garcinol can inhibit the receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis, osteoclastogenesis-related gene expression, the f-actin ring, and resorption pit formation. In addition, garcinol abrogated RANKL-induced osteoclastogenesis by attenuating the degradation of the MAPK, NF-κB, and PI3K-AKT signaling pathway as well as downstream factors c-jun, c-fos, and NFATC1. In vivo, suppression of osteoclastogenesis by garcinol was evidenced by marked inhibition of lipopolysaccharide-induced bone resorption. In conclusion, our data demonstrated that garcinol inhibited the RANKL-induced osteoclastogenesis by suppressing the MAPK, NF-κB, and PI3K-AKT signaling pathways and thus has potential as a novel therapeutic option for osteolytic bone diseases.