β-Boswellic Acid Inhibits RANKL-Induced Osteoclast Differentiation and Function by Attenuating NF-κB and Btk-PLCγ2 Signaling Pathways

Osteoporosis is a systemic metabolic bone disorder that is caused by an imbalance in the functions of osteoclasts and osteoblasts and is characterized by excessive bone resorption by osteoclasts. Targeting osteoclast differentiation and bone resorption is considered a good fundamental solution for overcoming bone diseases. β-boswellic acid (βBA) is a natural compound found in Boswellia serrata, which is an active ingredient with anti-inflammatory, anti-rheumatic, and anti-cancer effects. Here, we explored the anti-resorptive effect of βBA on osteoclastogenesis. βBA significantly inhibited the formation of tartrate-resistant acid phosphatase-positive osteoclasts induced by receptor activator of nuclear factor-B ligand (RANKL) and suppressed bone resorption without any cytotoxicity. Interestingly, βBA significantly inhibited the phosphorylation of IκB, Btk, and PLCγ2 and the degradation of IκB. Additionally, βBA strongly inhibited the mRNA and protein expression of c-Fos and NFATc1 induced by RANKL and subsequently attenuated the expression of osteoclast marker genes, such as OC-STAMP, DC-STAMP, β3-integrin, MMP9, ATP6v0d2, and CtsK. These results suggest that βBA is a potential therapeutic candidate for the treatment of excessive osteoclast-induced bone diseases such as osteoporosis.

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Inhibitory Effects of N-[2-(4-acetyl-1-piperazinyl) phenyl]-2-(2-chlorophenoxy) acetamide on Osteoclast Differentiation In Vitro via the Downregulation of TRAF6

International Journal of Molecular Sciences ◽

10.3390/ijms20205196 ◽

2019 ◽

Vol 20 (20) ◽

pp. 5196 ◽

Cited By ~ 5

Author(s):

Zhihao Chen ◽

Eunjin Cho ◽

Jinkyung Lee ◽

Sunwoo Lee ◽

Tae-Hoon Lee

Keyword(s):

Bone Resorption ◽

Mononuclear Cells ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Marker Genes ◽

Tartrate Resistant Acid Phosphatase ◽

Specific Marker ◽

Potential Candidate ◽

Dependent Manner

Osteoclasts are poly-nuclear cells that resorb mineral components from old or damaged bone tissue. Primary mononuclear cells are activated by receptor activator of nuclear factor kappa-Β ligand (RANKL) and differentiate into large multinucleated cells. Dysregulation of osteoclast differentiation can lead to pathological bone loss and destruction. Many studies have focused on the development of new molecules to regulate RANKL-mediated signaling. In this study, N-[2-(4-acetyl-1-piperazinyl)phenyl]-2-(2-chlorophenoxy) acetamide (PPOA-N-Ac-2-Cl) led to a significant decrease in the formation of multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cells in a dose-dependent manner, without inducing significant cytotoxicity. PPOA-N-Ac-2-Cl affected the expression of osteoclast-specific marker genes, such as TRAF6, c-fos, DC-STAMP, NFATc1, MMP9, CtsK, and TRAP (Acp5), during RANKL-mediated osteoclastogenesis. Moreover, PPOA-N-Ac-2-Cl significantly attenuated the protein levels of CtsK, a critical protease involved in bone resorption. Accordingly, bone resorption activity and F-actin ring formation decreased in the presence of PPOA-N-Ac-2-Cl. In conclusion, this study shows that PPOA-N-Ac-2-Cl acts as an inhibitor of osteoclast differentiation and may serve as a potential candidate agent for the treatment of osteoclast-related bone diseases by virtue of attenuating bone resorption.

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

Cellular Physiology and Biochemistry ◽

10.1159/000491892 ◽

2018 ◽

Vol 48 (2) ◽

pp. 644-656 ◽

Cited By ~ 4

Author(s):

Cheng-Ming Wei ◽

Yi-Ji Su ◽

Xiong Qin ◽

Jia-Xin Ding ◽

Qian Liu ◽

...

Keyword(s):

Critical Role ◽

Osteoclast Differentiation ◽

Osteoclast Formation ◽

Bone Diseases ◽

Marker Genes ◽

Tartrate Resistant Acid Phosphatase ◽

Dependent Manner ◽

And Function

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.

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Piceatannol attenuates RANKL-induced osteoclast differentiation and bone resorption by suppressing MAPK, NF-κB and AKT signalling pathways and promotes Caspase3-mediated apoptosis of mature osteoclasts

Royal Society Open Science ◽

10.1098/rsos.190360 ◽

2019 ◽

Vol 6 (6) ◽

pp. 190360 ◽

Cited By ~ 5

Author(s):

Liuliu Yan ◽

Lulu Lu ◽

Fangbin Hu ◽

Dattatrya Shetti ◽

Kun Wei

Keyword(s):

Bone Resorption ◽

Osteoclast Differentiation ◽

Giant Cells ◽

Inhibitory Effect ◽

Osteoclast Formation ◽

Bone Diseases ◽

Signalling Pathways ◽

Dependent Manner ◽

Underlying Mechanisms ◽

And Function

Osteoclasts are multinuclear giant cells that have unique ability to degrade bone. The search for new medicines that modulate the formation and function of osteoclasts is a potential approach for treating osteoclast-related bone diseases. Piceatannol (PIC) is a natural organic polyphenolic stilbene compound found in diverse plants with a strong antioxidant and anti-inflammatory effect. However, the effect of PIC on bone health has not been scrutinized systematically. In this study, we used RAW264.7, an osteoclast lineage of cells of murine macrophages, to investigate the effects and the underlying mechanisms of PIC on osteoclasts. Here, we demonstrated that PIC treatment ranging from 0 to 40 µM strongly inhibited osteoclast formation and bone resorption in a dose-dependent manner. Furthermore, the inhibitory effect of PIC was accompanied by the decrease of osteoclast-specific genes. At the molecular level, PIC suppressed the phosphorylation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK1/2), NF-κB p65, IκBα and AKT. Besides, PIC promoted the apoptosis of mature osteoclasts by inducing caspase-3 expression. In conclusion, our results suggested that PIC inhibited RANKL-induced osteoclastogenesis and bone resorption by suppressing MAPK, NF-κB and AKT signalling pathways and promoted caspase3-mediated apoptosis of mature osteoclasts, which might contribute to the treatment of bone diseases characterized by excessive bone resorption.

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Role of the Endocannabinoid/Endovanilloid System in the Modulation of Osteoclast Activity in Paget’s Disease of Bone

International Journal of Molecular Sciences ◽

10.3390/ijms221810158 ◽

2021 ◽

Vol 22 (18) ◽

pp. 10158

Author(s):

Marco Paoletta ◽

Antimo Moretti ◽

Sara Liguori ◽

Alessandra Di Paola ◽

Chiara Tortora ◽

...

Keyword(s):

Cannabinoid Receptors ◽

Cannabinoid Receptor ◽

Disease Model ◽

Paget’S Disease ◽

Paget's Disease ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Metabolic Bone Disorder ◽

Metabolic Bone

The role of the endocannabinoid/endovanilloid (EC/EV) system in bone metabolism has recently received attention. Current literature evidences the modulation of osteoclasts and osteoblasts through the activation or inhibition of cannabinoid receptors in various pathological conditions with secondary involvement of bone tissue. However, this role is still unclear in primary bone diseases. Paget’s disease of the bone (PDB) could be considered a disease model for analyzing the role of the EC/EV system on osteoclasts (OCs), speculating the potential use of specific agents targeting this system for managing metabolic bone disorders. The aim of the study is to analyze OCs expression of EC/EV system in patients with PDB and to compare OCs activity between this population and healthy people. Finally, we investigate whether specific agents targeting EC/EV systems are able to modulate OCs activity in this metabolic bone disorder. We found a significant increase in cannabinoid receptor type 2 (CB2) protein expression in patients with PDB, compared to healthy controls. Moreover, we found a significant reduction in multi-nucleated tartrate-resistant acid phosphatase (TRAP)–positive OCs and resorption areas after treatment with JWH-133. CB2 could be a molecular target for reducing the activity of OCs in PDB, opening new therapeutic scenarios for the management of this condition.

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Inhibitory Effects of 2N1HIA (2-(3-(2-Fluoro-4-Methoxyphenyl)-6-Oxo-1(6H)-Pyridazinyl)-N-1H-Indol-5-Ylacetamide) on Osteoclast Differentiation via Suppressing Cathepsin K Expression

Molecules ◽

10.3390/molecules23123139 ◽

2018 ◽

Vol 23 (12) ◽

pp. 3139 ◽

Cited By ~ 6

Author(s):

Sun-Hee Ahn ◽

Zhihao Chen ◽

Jinkyung Lee ◽

Seok-Woo Lee ◽

Sang Min ◽

...

Keyword(s):

Bone Resorption ◽

Osteoclast Differentiation ◽

Cell Formation ◽

Cathepsin K ◽

Bone Diseases ◽

Specific Gene ◽

Tartrate Resistant Acid Phosphatase ◽

Potential Candidate ◽

Dependent Manner ◽

Gene Markers

Osteoclasts are large multinucleated cells which are induced by the regulation of the receptor activator of nuclear factor kappa-Β ligand (RANKL), which is important in bone resorption. Excessive osteoclast differentiation can cause pathologic bone loss and destruction. Numerous studies have targeted molecules inhibiting RANKL signaling or bone resorption activity. In this study, 11 compounds from commercial libraries were examined for their effect on RANKL-induced osteoclast differentiation. Of these compounds, only 2-(3-(2-fluoro-4-methoxyphenyl)-6-oxo-1(6H)-pyridazinyl)-N-1H-indol-5-ylacetamide (2N1HIA) caused a significant decrease in multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cell formation in a dose-dependent manner, without inducing cytotoxicity. The 2N1HIA compound neither affected the expression of osteoclast-specific gene markers such as TRAF6, NFATc1, RANK, OC-STAMP, and DC-STAMP, nor the RANKL signaling pathways, including p38, ERK, JNK, and NF-κB. However, 2N1HIA exhibited a significant impact on the expression levels of CD47 and cathepsin K, the early fusion marker and critical protease for bone resorption, respectively. The activity of matrix metalloprotease-9 (MMP-9) decreased due to 2N1HIA treatment. Accordingly, bone resorption activity and actin ring formation decreased in the presence of 2N1HIA. Taken together, 2N1HIA acts as an inhibitor of osteoclast differentiation by attenuating bone resorption activity and may serve as a potential candidate in preventing and/or treating osteoporosis, or other bone diseases associated with excessive bone resorption.

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Alpinetin Inhibits RANKL-Induced Osteoclastogenesis and Ovariectomy-Induced Bone Loss by Modulating NFATc1 Transcription and Lysosomal Function

10.21203/rs.3.rs-132706/v1 ◽

2020 ◽

Author(s):

Rongxin He ◽

Jinwei Lu ◽

Yazhou Chen ◽

Yong Li ◽

Chenyi Ye ◽

...

Keyword(s):

Bone Resorption ◽

Bone Loss ◽

Bone Matrix ◽

Osteoclast Differentiation ◽

Estrogen Deficiency ◽

Bone Diseases ◽

Dependent Manner ◽

Metabolic Bone Diseases ◽

Lysosomal Function ◽

Metabolic Bone

Abstract BackgroundPostmenopausal osteoporosis is a chronic metabolic bone disease caused by excessive osteoclast activation, and osteoclasts are considered to be the sole participants in the degeneration and resorption of bone matrix for controlling bone integrity and continuity. The biological functions of osteoclasts depend critically on the number and activity of fused polykaryon. Hence, targeting osteoclast differentiation and activity can modulate bone resorption and alleviate osteoporosis. Alpinetin is widely used for excellent anti-inflammatory activities and little side-effect, but its role in osteoporosis remains unknown.ResultsIn this study, we investigated for the first time the ability of alpinetin to inhibit estrogen deficiency-induced bone loss. Alpinetin significantly reduced the expression levels of NFATc1 and its downstream genes, thereby inhibiting osteoclast differentiation in a concentration- and time-dependent manner. Additionally, alpinetin inhibited F-actin ring formation and bone resorption, as well as reduced the activation levels of NF-κB, ERK, and AKT signaling cascades. In mature osteoclasts, alpinetin remarkably inhibited integrin-mediated migration and lysosomal biogenesis and trafficking by modulating the PKCβ/TFEB and ATG5/LC3 axes. Importantly, alpinetin treatment in mice alleviated ovariectomy-induced bone volume loss. ConclusionOur findings strongly suggest that alpinetin plays a significant role in the regulation of NFATc1 production for the differentiation of osteoclasts and inhibits integrin-mediated cell migration and lysosomal function in mature osteoclasts, thus weaken the increased osteolytic ability due to estrogen deficiency. Alpinetin may represent a promising agent for the treatment of osteoporosis and other metabolic bone diseases.

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Picrasidine I from Picrasma Quassioides Suppresses Osteoclastogenesis via Inhibition of RANKL Induced Signaling Pathways and Attenuation of ROS Production

Cellular Physiology and Biochemistry ◽

10.1159/000481874 ◽

2017 ◽

Vol 43 (4) ◽

pp. 1425-1435 ◽

Cited By ~ 9

Author(s):

Lingbo Kong ◽

Biao Wang ◽

Xiaobin Yang ◽

Hua Guo ◽

Ke Zhang ◽

...

Keyword(s):

Bone Resorption ◽

Signaling Pathways ◽

Osteoclast Differentiation ◽

Inhibitory Effect ◽

Ring Structure ◽

Osteoclast Formation ◽

Ros Generation ◽

Tartrate Resistant Acid Phosphatase ◽

Metabolic Bone Disorder ◽

Picrasma Quassioides

Background/Aims: Osteoporosis is a metabolic bone disorder that tortures about millions of people worldwide. Recent study demonstrated agents derived from picrasma quassioides is a promising drug for targets multiple signaling pathways. However its potential in treatment of bone loss has not been fully understood. Methods: The bone marrow macrophages (BMMs) were cultured and induced with M-CSF and RANKL followed by picrasidine I (PI) treatment. Then the effects of PI on osteoclast formation were evaluated by counting tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells. Moreover, effects of PI on bone resorption activity of mature osteoclast were studied through bone resorption pit counting and actin ring structure analysis. Further, the involved potential signaling pathways cross-talking were investigated by performed Western blotting and quantitative real-time PCR examination. Results: Results demonstrated PI strongly inhibited RANKL induced osteoclast formation from its precursors. Mechanistically, the inhibitory effect of PI on osteoclast differentiation was due to the suppression of osteoclastogenic transcription factors, c-Fos and NFATc1. Moreover, PI markedly blocked the RANKL-induced osteoclastogenesis by attenuating MAPKs and NF-κB signaling pathways. In addition, PI decreased the ROS generation in osteoclast and osteoblast. Conclusion: Taken together our data demonstrate that PI has antiosteoclastogenic effect by inhibiting inflammation induced activation of MAPKs, NF-κB and ROS generation followed by suppressing the gene expression of c-Fos and NFATc1 in osteoclast precursors.

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PSTP-3,5-Me Inhibits Osteoclast Differentiation and Bone Resorption

Molecules ◽

10.3390/molecules24183346 ◽

2019 ◽

Vol 24 (18) ◽

pp. 3346 ◽

Cited By ~ 1

Author(s):

Eunjin Cho ◽

Zhihao Chen ◽

Jinkyung Lee ◽

Sunwoo Lee ◽

Tae-Hoon Lee

Keyword(s):

Osteoclast Differentiation ◽

Cathepsin K ◽

Osteoporosis Treatment ◽

Bone Diseases ◽

Marker Genes ◽

Tartrate Resistant Acid Phosphatase ◽

Mouse Bone Marrow ◽

Nuclear Factor ◽

Activated T Cell ◽

Macrophage Colony

Osteogenesis is an orchestrated process regulated by osteoclastogenesis and osteoblastogenesis. Excessive osteoclastogenesis causes bone diseases, such as osteoporosis. Although a few drugs are effective in osteoporosis treatment, these drugs lead to side effects, including cellulitis, flatulence, and hypocalcemia. In this study, we reported a 2-(N-Phenylmethylsulfonamido)-N-(2-(phenylthio)phenyl)propanamide (PSTP) compound, PSTP-3,5-Me, as a potential therapeutic agent for osteoporosis. Mouse bone marrow-derived macrophages (BMMs) were differentiated into osteoclasts by receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) in the presence of PSTP-3,5-Me. PSTP-3,5-Me inhibited osteoclast differentiation by reduced tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts, and suppressed the expression of osteoclast marker genes, such as cathepsin K (Ctsk) and TRAP (Acp5). We investigated signaling pathways mediated by RANKL and its receptor, RANK, and found that PSTP-3,5-Me inhibits nucleus translocation of nuclear factor of activated T cell cytoplasmic-1 (NFATc1). Moreover, PSTP-3,5-Me inhibited F-actin ring formation and mineral resorption. Overall, our data suggests that PSTP-3,5-Me attenuates osteoclast differentiation by blocking the activation of NFATc1.

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N-[2-(4-Acetyl-1-Piperazinyl)Phenyl]-2-(3-Methylphenoxy)Acetamide (NAPMA) Inhibits Osteoclast Differentiation and Protects against Ovariectomy-Induced Osteoporosis

Molecules ◽

10.3390/molecules25204855 ◽

2020 ◽

Vol 25 (20) ◽

pp. 4855

Author(s):

Jinkyung Lee ◽

Sun-Hee Ahn ◽

Zhihao Chen ◽

Sohi Kang ◽

Dong Kyu Choi ◽

...

Keyword(s):

Bone Resorption ◽

Bone Loss ◽

Osteoclast Differentiation ◽

Cathepsin K ◽

Bone Diseases ◽

Drug Candidate ◽

Tartrate Resistant Acid Phosphatase ◽

Dependent Manner ◽

Osteoclast Activity ◽

Protein Levels

Osteoclasts are large, multinucleated cells responsible for bone resorption and are induced in response to the regulatory activity of receptor activator of nuclear factor-kappa B ligand (RANKL). Excessive osteoclast activity causes pathological bone loss and destruction. Many studies have investigated molecules that specifically inhibit osteoclast activity by blocking RANKL signaling or bone resorption. In recent years, we screened compounds from commercial libraries to identify molecules capable of inhibiting RANKL-induced osteoclast differentiation. Consequently, we reported some compounds that are effective at attenuating osteoclast activity. In this study, we found that N-[2-(4-acetyl-1-piperazinyl)phenyl]-2-(3-methylphenoxy)acetamide (NAPMA) significantly inhibited the formation of multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cells from bone marrow-derived macrophages in a dose-dependent manner, without cytotoxic effects. NAPMA downregulated the expression of osteoclast-specific markers, such as c-Fos, NFATc1, DC-STAMP, cathepsin K, and MMP-9, at the transcript and protein levels. Accordingly, bone resorption and actin ring formation were decreased in response to NAPMA treatment. Furthermore, we demonstrated the protective effect of NAPMA against ovariectomy-induced bone loss using micro-CT and histological analysis. Collectively, the results showed that NAPMA inhibited osteoclast differentiation and attenuated bone resorption. It is thus a potential drug candidate for the treatment of osteoporosis and other bone diseases associated with excessive bone resorption.

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Inhibition of Osteoclast Differentiation and Bone Resorption by N-Methylpyrrolidone

Journal of Biological Chemistry ◽

10.1074/jbc.m111.223297 ◽

2011 ◽

Vol 286 (27) ◽

pp. 24458-24466 ◽

Cited By ~ 37

Author(s):

Chafik Ghayor ◽

Rita M. Correro ◽

Katrin Lange ◽

Lindsay S. Karfeld-Sulzer ◽

Klaus W. Grätz ◽

...

Keyword(s):

Acid Phosphatase ◽

Bone Resorption ◽

Osteoclast Differentiation ◽

Cathepsin K ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Treatment Of Osteoporosis ◽

Multinucleated Cells ◽

Antiresorptive Agents ◽

Actin Rings

Regulation of RANKL (receptor activator of nuclear factor κB ligand)-induced osteoclast differentiation is of current interest in the development of antiresorptive agents. Osteoclasts are multinucleated cells that play a crucial role in bone resorption. In this study, we investigated the effects of N-methylpyrrolidone (NMP) on the regulation of RANKL-induced osteoclastogenesis. NMP inhibited RANKL-induced tartrate-resistant acid phosphatase activity and the formation of tartrate-resistant acid phosphatase-positive multinucleated cells. The RANKL-induced expression of NFATc1 (nuclear factor of activated T cells, cytoplasmic 1) and c-Fos, which are key transcription factors for osteoclastogenesis, was also reduced by treatment with NMP. Furthermore, NMP induced disruption of the actin rings and decreased the mRNAs of cathepsin K and MMP-9 (matrix metalloproteinase-9), both involved in bone resorption. Taken together, these results suggest that NMP inhibits osteoclast differentiation and attenuates bone resorption. Therefore, NMP could prove useful for the treatment of osteoporosis or other bone diseases associated with excessive bone resorption.

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