Inhibition of Osteoclastogenesis by Thioredoxin-Interacting Protein-Derived Peptide (TN13)

Overactivated osteoclasts lead to many bone diseases, including osteoporosis and rheumatoid arthritis. The p38 MAPK (p38) is an essential regulator of the receptor activator of nuclear factor-κB ligand (RANKL)-mediated osteoclastogenesis and bone loss. We previously reported TAT conjugated thioredoxin-interacting protein-derived peptide (TAT-TN13) as an inhibitor of p38 in hematopoietic stem cells (HSCs). Here, we examined the role of TAT-TN13 in the differentiation and function of osteoclasts. TAT-TN13 significantly suppressed RANKL-mediated differentiation of RAW 264.7 cells and bone marrow macrophages (BMMs) into osteoclasts. TAT-TN13 also inhibited the RANKL-induced activation of NF-κB and nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), leading to the decreased expression of osteoclast-specific genes, including tartrate-resistant acid phosphatase (TRAP) and Cathepsin K. Additionally, TAT-TN13 treatment protected bone loss in ovariectomized (OVX) mice. Taken together, these results suggest that TAT-TN13 inhibits osteoclast differentiation by regulating the p38 and NF-κB signaling pathway; thus, it may be a useful agent for preventing or treating osteoporosis.

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Orostachys japonicus Suppresses Osteoclast Differentiation by Inhibiting NFATc1 Expression

The American Journal of Chinese Medicine ◽

10.1142/s0192415x15500585 ◽

2015 ◽

Vol 43 (05) ◽

pp. 1013-1030 ◽

Cited By ~ 2

Author(s):

Ki-Shuk Shim ◽

Hyunil Ha ◽

Taesoo Kim ◽

Chung-Jo Lee ◽

Jin Yeul Ma

Keyword(s):

Bone Resorption ◽

Bone Loss ◽

Therapeutic Potential ◽

Osteoclast Differentiation ◽

Water Extract ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Dependent Manner ◽

Nuclear Factor ◽

Activated T Cells

The herb Orostachys japonicus has been traditionally used to treat chronic diseases, such as hepatitis, hemorrhoids, and cancer, in Asia. In this study, we investigated the effect of Orostachys japonicus water extract (OJWE) on the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and bone loss. We found that OJWE inhibited RANKL-induced osteoclast differentiation in a dose-dependent manner without affecting bone resorption in bone marrow-derived macrophage cells. Interestingly, OJWE significantly reduced serum levels of C-terminal telopeptide of type 1 collagen and tartrate-resistant acid phosphatase (TRAP) 5b, markers of bone resorption and osteoclast number, respectively, in an animal model of bone loss. Furthermore, OJWE suppressed the RANKL-induced up-regulation of nuclear factor of activated T cells cytoplasmic 1 (NFATc1) expression, and activation of the p38 signaling pathway, but prevented the RANKL-mediated down-regulation of interferon regulatory factor-8 (IRF-8), which is known to be an anti-osteoclastogenic factor that represses NFATc1 expression. We also identified gallic acid and quercetin-3-O-β-D-glucoside as the OJWE components that inhibit RANKL-induced osteoclast differentiation. These results suggest that OJWE inhibits osteoclast differentiation by inhibiting RANKL-induced NFATc1 expression, which prevents osteoclast differentiation and bone loss. The present study elucidated a mechanism of action underlying the inhibitory effect of OJWE on osteoclast differentiation. Our findings suggest that O. japonicus has therapeutic potential for use in the treatment of bone diseases.

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Kalkitoxin Reduces Osteoclast Formation and Resorption and Protects against Inflammatory Bone Loss

International Journal of Molecular Sciences ◽

10.3390/ijms22052303 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2303

Author(s):

Liang Li ◽

Ming Yang ◽

Saroj Kumar Shrestha ◽

Hyoungsu Kim ◽

William H. Gerwick ◽

...

Keyword(s):

Bone Loss ◽

Transmembrane Protein ◽

Osteoclast Differentiation ◽

Mapk Signaling ◽

Osteoclast Formation ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Nuclear Factor ◽

Mineral Density ◽

Multinucleated Cells

Osteoclasts, bone-specified multinucleated cells produced by monocyte/macrophage, are involved in numerous bone destructive diseases such as arthritis, osteoporosis, and inflammation-induced bone loss. The osteoclast differentiation mechanism suggests a possible strategy to treat bone diseases. In this regard, we recently examined the in vivo impact of kalkitoxin (KT), a marine product obtained from the marine cyanobacterium Moorena producens (previously Lyngbya majuscula), on the macrophage colony-stimulating factor (M-CSF) and on the receptor activator of nuclear factor κB ligand (RANKL)-stimulated in vitro osteoclastogenesis and inflammation-mediated bone loss. We have now examined the molecular mechanism of KT in greater detail. KT decreased RANKL-induced bone marrow-derived macrophages (BMMs) tartrate-resistant acid phosphatase (TRAP)-multinucleated cells at a late stage. Likewise, KT suppressed RANKL-induced pit area and actin ring formation in BMM cells. Additionally, KT inhibited several RANKL-induced genes such as cathepsin K, matrix metalloproteinase (MMP-9), TRAP, and dendritic cell-specific transmembrane protein (DC-STAMP). In line with these results, RANKL stimulated both genes and protein expression of c-Fos and nuclear factor of activated T cells (NFATc1), and this was also suppressed by KT. Moreover, KT markedly decreased RANKL-induced p-ERK1/2 and p-JNK pathways at different time points. As a result, KT prevented inflammatory bone loss in mice, such as bone mineral density (BMD) and osteoclast differentiation markers. These experiments demonstrated that KT markedly inhibited osteoclast formation and inflammatory bone loss through NFATc1 and mitogen-activated protein kinase (MAPK) signaling pathways. Therefore, KT may have potential as a treatment for destructive bone diseases.

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Fucoidan Prevents RANKL-Stimulated Osteoclastogenesis and LPS-Induced Inflammatory Bone Loss via Regulation of Akt/GSK3β/PTEN/NFATc1 Signaling Pathway and Calcineurin Activity

Marine Drugs ◽

10.3390/md17060345 ◽

2019 ◽

Vol 17 (6) ◽

pp. 345 ◽

Cited By ~ 5

Author(s):

Sheng-Hua Lu ◽

Yi-Jan Hsia ◽

Kuang-Chung Shih ◽

Tz-Chong Chou

Keyword(s):

Bone Loss ◽

Molecular Mechanisms ◽

Bone Erosion ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Nuclear Factor ◽

Activated T Cells ◽

Calcineurin Activity

Excessive osteoclast differentiation and/or function plays a pivotal role in the pathogenesis of bone diseases such as osteoporosis and rheumatoid arthritis. Here, we examined whether fucoidan, a sulfated polysaccharide present in brown algae, attenuates receptor activator of nuclear factor-κB ligand (RANKL)-stimulated osteoclastogenesis in vitro and lipopolysaccharide (LPS)-induced bone resorption in vivo, and investigated the molecular mechanisms involved. Our results indicated that fucoidan significantly inhibited osteoclast differentiation in RANKL-stimulated macrophages and the bone resorbing activity of osteoclasts. The effects of fucoidan may be mediated by regulation of Akt/GSK3β/PTEN signaling and suppression of the increase in intracellular Ca2+ level and calcineurin activity, thereby inhibiting the translocation of nuclear factor-activated T cells c1 (NFATc1) into the nucleus. However, fucoidan-mediated NFATc1 inactivation was greatly reversed by kenpaullone, a GSK3β inhibitor. In addition, using microcomputer tomography (micro-CT) scanning and bone histomorphometry, we found that fucoidan treatment markedly prevented LPS-induced bone erosion in mice. Collectively, we demonstrated that fucoidan was capable of inhibiting osteoclast differentiation and inflammatory bone loss, which may be modulated by regulation of Akt/GSK3β/PTEN/NFATc1 and Ca2+/calcineurin signaling cascades. These findings suggest that fucoidan may be a potential agent for the treatment of osteoclast-related bone diseases.

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Transcriptional Modulator Ifrd1 Regulates Osteoclast Differentiation through Enhancing the NF-κB/NFATc1 Pathway

Molecular and Cellular Biology ◽

10.1128/mcb.01075-15 ◽

2016 ◽

Vol 36 (19) ◽

pp. 2451-2463 ◽

Cited By ~ 13

Author(s):

Takashi Iezaki ◽

Kazuya Fukasawa ◽

Gyujin Park ◽

Tetsuhiro Horie ◽

Takashi Kanayama ◽

...

Keyword(s):

Bone Loss ◽

Osteoclast Differentiation ◽

Nuclear Factor Κb ◽

Bone Diseases ◽

Nuclear Factor ◽

Activator Protein 1 ◽

Bone Homeostasis ◽

Activated T Cells

Bone homeostasis is maintained by the synergistic actions of bone-resorbing osteoclasts and bone-forming osteoblasts. Here, we show that the transcriptional coactivator/repressor interferon-related developmental regulator 1 (Ifrd1) is expressed in osteoclast lineages and represents a component of the machinery that regulates bone homeostasis. Ifrd1 expression was transcriptionally regulated in preosteoclasts by receptor activator of nuclear factor κB (NF-κB) ligand (RANKL) through activator protein 1. Global deletion of murineIfrd1increased bone formation and decreased bone resorption, leading to a higher bone mass. Deletion ofIfrd1in osteoclast precursors prevented RANKL-induced bone loss, although no bone loss was observed under normal physiological conditions. RANKL-dependent osteoclastogenesis was impairedin vitroinIfrd1-deleted bone marrow macrophages (BMMs).Ifrd1deficiency increased the acetylation of p65 at residues K122 and K123 via the inhibition of histone deacetylase-dependent deacetylation in BMMs. This repressed the NF-κB-dependent transcription of nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), an essential regulator of osteoclastogenesis. These findings suggest that an Ifrd1/NF-κB/NFATc1 axis plays a pivotal role in bone remodelingin vivoand represents a therapeutic target for bone diseases.

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Berberine Suppresses RANKL-Induced Osteoclast Differentiation by Inhibiting c-Fos and NFATc1 Expression

The American Journal of Chinese Medicine ◽

10.1142/s0192415x19500228 ◽

2019 ◽

Vol 47 (02) ◽

pp. 439-455 ◽

Cited By ~ 3

Author(s):

Sang-Yong Han ◽

Yun-Kyung Kim

Keyword(s):

Bone Loss ◽

Osteoclast Differentiation ◽

Osteoclast Formation ◽

Bone Diseases ◽

Mrna Levels ◽

Nuclear Factor ◽

Activated T Cells ◽

Murine Bone Marrow

Osteoporosis is a common disorder of bone remodeling, marked by excessive osteoclast formation. Recent studies indicated that berberine (BBR) is a potential natural drug for the treatment of various bone diseases. However, it still needs to be further studied for the treatment of osteoporosis. The current study investigated the inhibitory effects of BBR on receptor activator of nuclear factor-[Formula: see text]B ligand (RANKL)-induced osteoclast differentiation in vitro and in vivo. Cell-based assays were performed using osteoclasts generated in cultures of murine bone marrow-derived macrophages (BMMs) treated with RANKL and M-CSF. The effects of BBR on in vivo lipopolysaccharide (LPS)-mediated bone loss were evaluated using ICR mice. BBR significantly inhibited TRAP-positive osteoclast formation induced by RANKL. BBR also inhibited RANKL-induced Akt, p38 and ERK phosphorylation and I[Formula: see text]B degradation, and suppressed RANKL-induced expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which is a key transcription factors for osteoclast formation. BBR reduced the mRNA levels of osteoclast markers, including TRAP, osteoclast-associated receptor (OSCAR), cathepsin K, and ATPase H[Formula: see text] transporting V0 subunit d2 (ATP6v0d2). Moreover, BBR prevented LPS-mediated bone loss in vivo. We suggest BBR as a natural compound that can be a potential therapeutic agent for osteoclast-related bone diseases.

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Austalide K from the Fungus Penicillium rudallense Prevents LPS-Induced Bone Loss in Mice by Inhibiting Osteoclast Differentiation and Promoting Osteoblast Differentiation

International Journal of Molecular Sciences ◽

10.3390/ijms22115493 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5493

Author(s):

Kwang-Jin Kim ◽

Jusung Lee ◽

Weihong Wang ◽

Yongjin Lee ◽

Eunseok Oh ◽

...

Keyword(s):

Bone Loss ◽

Osteoblast Differentiation ◽

Financial Burden ◽

Osteoclast Differentiation ◽

Public Health Problem ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Nuclear Factor ◽

Mineral Density ◽

Trabecular Thickness

Osteoporosis is a chronic disease that has become a serious public health problem due to the associated reduction in quality of life and its increasing financial burden. It is known that inhibiting osteoclast differentiation and promoting osteoblast formation prevents osteoporosis. As there is no drug with this dual activity without clinical side effects, new alternatives are needed. Here, we demonstrate that austalide K, isolated from the marine fungus Penicillium rudallenes, has dual activities in bone remodeling. Austalide K inhibits the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and improves bone morphogenetic protein (BMP)-2-mediated osteoblast differentiation in vitro without cytotoxicity. The nuclear factor of activated T cells c1 (NFATc1), tartrate-resistant acid phosphatase (TRAP), dendritic cell-specific transmembrane protein (DC-STAMP), and cathepsin K (CTSK) osteoclast-formation-related genes were reduced and alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2), osteocalcin (OCN), and osteopontin (OPN) (osteoblast activation-related genes) were simultaneously upregulated by treatment with austalide K. Furthermore, austalide K showed good efficacy in an LPS-induced bone loss in vivo model. Bone volume, trabecular separation, trabecular thickness, and bone mineral density were recovered by austalide K. On the basis of these results, austalide K may lead to new drug treatments for bone diseases such as osteoporosis.

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Nasturtium officinale Extract Suppresses Osteoclastogenesis in RAW 264 Cells by Inhibiting IκB-Kinase β

Natural Product Communications ◽

10.1177/1934578x211020643 ◽

2021 ◽

Vol 16 (6) ◽

pp. 1934578X2110206

Author(s):

Yukino Tsunekage ◽

Masatoshi Takeiri ◽

Yuri Yoshioka ◽

Shinichi Matsumura ◽

Yoshihide Kimura ◽

...

Keyword(s):

Osteoclast Differentiation ◽

Specific Inhibitor ◽

Nuclear Factor Κb ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Nuclear Factor ◽

Activated T Cells ◽

Nasturtium Officinale ◽

Iκb Kinase

Osteoclasts are large, multinucleated, bone-absorbing cells and play a crucial role in osteolytic bone diseases such as osteopetrosis and rheumatoid arthritis. Therefore, controlling osteoclast differentiation and activation has been considered a promising strategy to prevent and treat osteolytic diseases. In this study, we demonstrate, using the mouse monocyte-derived macrophage-like cell line RAW 264, that extract from Nasturtium officinale or watercress, an herb of European origin, suppresses receptor activator of nuclear factor-κB ligand-induced osteoclast differentiation in vitro . N. officinale extract decreased the emergence of tartrate-resistant acid phosphatase-positive differentiated multinuclear cells and inhibited their bone-absorbing activity. The extract decreased expression of genes associated with osteoclast differentiation and function. Induction of nuclear factor of activated T cells c1 (NFATc1), the master transcriptional regulator of osteoclastogenesis, was blunted by N. officinale extract. Activation of nuclear factor-κB and mitogen-activated protein kinases pathways, both of which are necessary for NFATc1 induction and osteoclast differentiation, was also suppressed by the extract. Among upstream kinases, activity of IκB-kinase β (IKKβ), but not that of TGFβ-activated kinase 1, was inhibited by N. officinale extract in vitro. Pharmacological inhibition of IKKβ by a specific inhibitor PS1145 in RAW 264 cells mostly recaptured the inhibitory action of N. officinale extract. These findings provide a novel pharmacological action of N. officinale and its potential usefulness for the prevention of osteoporosis.

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Protective Effects of Fermented Oyster Extract against RANKL-Induced Osteoclastogenesis through Scavenging ROS Generation in RAW 264.7 Cells

International Journal of Molecular Sciences ◽

10.3390/ijms20061439 ◽

2019 ◽

Vol 20 (6) ◽

pp. 1439 ◽

Cited By ~ 10

Author(s):

Jin-Woo Jeong ◽

Sung Choi ◽

Min Han ◽

Gi-Young Kim ◽

Cheol Park ◽

...

Keyword(s):

Signaling Pathways ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Raw 264.7 Cells ◽

Ros Generation ◽

Tartrate Resistant Acid Phosphatase ◽

Raw 264.7 ◽

Nuclear Factor ◽

Protective Effects ◽

Induced Expression

Excessive bone resorption by osteoclasts causes bone loss-related diseases and reactive oxygen species (ROS) act as second messengers in intercellular signaling pathways during osteoclast differentiation. In this study, we explored the protective effects of fermented oyster extract (FO) against receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclast differentiation in murine monocyte/macrophage RAW 264.7 cells. Our results showed that FO markedly inhibited RANKL-induced activation of tartrate-resistant acid phosphatase and formation of F-actin ring structure. Mechanistically, FO has been shown to down-regulate RANKL-induced expression of osteoclast-specific markers by blocking the nuclear translocation of NF-κB and the transcriptional activation of nuclear factor of activated T cells c1 (NFATc1) and c-Fos. Furthermore, FO markedly diminished ROS production by RANKL stimulation, which was associated with blocking the expression of nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1) and its regulatory subunit Rac-1. However, a small interfering RNA (siRNA) targeting NOX1 suppressed RANKL-induced expression of osteoclast-specific markers and production of ROS and attenuated osteoclast differentiation as in the FO treatment group. Collectively, our findings suggest that FO has anti-osteoclastogenic potential by inactivating the NF-κB-mediated NFATc1 and c-Fos signaling pathways and inhibiting ROS generation, followed by suppression of osteoclast-specific genes. Although further studies are needed to demonstrate efficacy in in vivo animal models, FO may be used as an effective alternative agent for the prevention and treatment of osteoclastogenic bone diseases.

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Yukmijihwang-Tang Suppresses Receptor Activator of Nuclear Factor Kappa-B Ligand (RANKL)-Induced Osteoclast Differentiation and Prevents Ovariectomy (OVX)-Mediated Bone Loss

Molecules ◽

10.3390/molecules26247579 ◽

2021 ◽

Vol 26 (24) ◽

pp. 7579

Author(s):

Sang-Yong Han ◽

Yun-Kyung Kim

Keyword(s):

Bone Loss ◽

Nuclear Factor Kappa B ◽

Nuclear Translocation ◽

Transmembrane Protein ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Tartrate Resistant Acid Phosphatase ◽

Nuclear Factor ◽

Nuclear Factor Kappa ◽

Receptor Activator

Yukmijihwang-tang (YJ) has been used to treat diabetes mellitus, renal disorders, and cognitive impairment in traditional medicine. This study aimed to evaluate the anti-osteoporotic effect of YJ on ovariectomy (OVX)-induced bone loss in a rat and receptor activator of nuclear factor kappa-B ligand (RANKL)-mediated osteoclast differentiation in bone marrow macrophages (BMMs). YJ reduced the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells (MNCs) in an osteoclast/osteoblast co-culture system by regulating the ratio of RANKL/osteoprotegerin (OPG) by osteoblasts. Overall, YJ reduced TRAP-positive cell formation and TRAP activity and F-actin ring formation. Analysis of the underlying mechanisms indicated that YJ inhibited the activation of the nuclear factor of activated T cell cytoplasmic 1 (NFATc1) and c-Fos, resulting in the suppression of osteoclast differentiation-related genes such as TRAP, ATPase, H+ transporting, lysosomal 38 kDa, V0 subunit d2, osteoclast-associated receptor, osteoclast-stimulatory transmembrane protein, dendritic cell-specific transmembrane protein, matrix metalloproteinase-9, cathepsin K, and calcitonin receptor. YJ also inhibited the nuclear translocation of NFATc1. Additionally, YJ markedly inhibited RANKL-induced phosphorylation of signaling pathways activated in the early stages of osteoclast differentiation including the p38, JNK, ERK, and NF-κB. Consistent with these in vitro results, the YJ-administered group showed considerably attenuated bone loss in the OVX-mediated rat model. These results provide promising evidence for the potential novel therapeutic application of YJ for bone diseases such as osteoporosis.

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Forsythia suspensa Protects against Bone Loss in Ovariectomized Mice

Nutrients ◽

10.3390/nu11081831 ◽

2019 ◽

Vol 11 (8) ◽

pp. 1831 ◽

Cited By ~ 4

Author(s):

Youn-Hwan Hwang ◽

Seon-A Jang ◽

Taesoo Kim ◽

Hyunil Ha

Keyword(s):

Bone Loss ◽

Trabecular Bone ◽

Osteoclast Differentiation ◽

Water Extract ◽

Nuclear Factor ◽

Micro Computed Tomography ◽

Activated T Cells ◽

Beneficial Effects ◽

Ovariectomized Mice ◽

Forsythia Suspensa

In traditional oriental medicine, the fruit of Forsythia suspensa has been used as a nutritional supplement to alleviate inflammation and treat gastrointestinal diseases. However, there is no information available on its beneficial effects on bone. We investigated the beneficial effects of F. suspensa water extract (WFS) on osteoclast differentiation and bone loss. The microarchitecture of trabecular bone was analyzed by micro-computed tomography. Osteoclast differentiation was evaluated based on tartrate-resistant alkaline phosphatase activity, and bone resorption activity was examined on a bone-like mineral surface. The mechanism of action of WFS was assessed by evaluating the expression and activation of signaling molecules. Phytochemical constituents were identified and quantitated by ultrahigh-performance liquid chromatography–tandem mass spectrometry. WFS reduced ovariectomy-induced trabecular bone loss and inhibited receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation and resorption activity. WFS suppressed RANKL-induced expression of nuclear factor of activated T cells cytoplasmic 1, a crucial transcription factor for osteoclast differentiation by decreasing c-Fos protein levels and suppressing the activation of p38 and c-Jun-N-terminal kinase. We also identified 12 phytochemicals in WFS including lignans, phenylethanoids, and flavonoids. Collectively, these results suggest that WFS inhibits osteoclast differentiation and can potentially be used to treat postmenopausal osteoporosis.

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