scholarly journals The inhibitory effect of melatonin on osteoclastogenesis of RAW 264.7 cells in low concentrations of RANKL and MCSF

2020 ◽  
Vol 44 (6) ◽  
pp. 427-436
Author(s):  
Hala JARRAR ◽  
Damla ÇETİN ALTINDAL ◽  
Menemşe GÜMÜŞDERELİOĞLU
Keyword(s):  
Cathepsin K ◽  
Suppressive Effect ◽  
Inhibitory Effect ◽  
Raw 264.7 Cells ◽  
Tartrate Resistant Acid Phosphatase ◽  
Raw 264.7 ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Repressive Effect ◽  
Activity Trap

RAW 264.7 cells are one of the most recommended cell lines for investigating the activity and differentiation of osteoclasts. These cells differentiate into osteoclasts in the presence of two critical components: receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony stimulating factor (MCSF). Melatonin (MEL) hormone has recently become one of the small molecules used in the field of bone regeneration and bone disease treatment, as it has the ability to inhibit the differentiation of osteoclasts directly by suppression of the NF-κB signaling pathway. The main aim of the current study is to determine sufficient RANKL/MCSF concentrations for differentiation of the cells to osteoclasts and to describe the repressive effect of MEL on the osteoclastogenesis of these cells. In this regard, it was found that 10 ng/mL of RANKL- and MCSF-containing medium is suitable for inducing osteoclastogenesis of the cells. In addition, melatonin at doses in the range of 100–1000 μM does not have a cytotoxic effect. Subsequently, results of tartrate resistant acid phosphatase (TRAP) activity, TRAP staining, and relative expressions of cathepsin K, nuclear factor of activated T cells one (NFATC1), and TRAP genes showed a suppressive effect of MEL —especially 800 μM— on RANKL-induced osteoclastogenesis of these cells.

scholarly journals Optimized Extract from Corylopsis coreana Uyeki (Hamamelidaceae) Flos Inhibits Osteoclast Differentiation

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Yongjin Lee ◽  
Jung-Eun Kim ◽  
Kwang-Jin Kim ◽  
Seung-Sik Cho ◽  
Young-Jin Son
Keyword(s):  
Osteoclast Differentiation ◽  
Cathepsin K ◽  
Inhibitory Effect ◽  
Osteoclast Formation ◽  
Tartrate Resistant Acid Phosphatase ◽  
Alcoholic Extract ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Fractures Of The Spine ◽  
The Stability

Osteoporosis is a metabolic disorder that decreases the stability against fractures of the spine, femur, and radius by weakening the strength and integrity of bones. Receptor activator of nuclear factor-kappa B ligand signaling ultimately activated nuclear factor-activated T cells c1, a major transcription factor for osteoclast formation. This study researched the effects of Corylopsis coreana (C. coreana) Uyeki flos extracts on the antiosteoclastic potential of macrophages and the phytochemicals contained therein. The alcoholic extract of C. coreana Uyeki flos inhibited the differentiation of osteoclast. We carried out the experiments of the pattern of differentiation of osteoclasts based on the alcoholic percentage of extracts. Among them, 80% alcoholic extract showed the highest inhibitory effect. The alcoholic extract was composed of phytochemicals such as bergenin, quercetin, and quercitrin. This extract inhibited not only mRNA expression levels of NFATc1, osteoclast-associated receptor (OSCAR), cathepsin K, and tartrate-resistant acid phosphatase (TRAP), but also the translational expression of NFATc1. The inhibitory effect for osteoclast differentiation of the alcoholic extract was confirmed using the resorption pit assay. This is the first scientific report of the antiosteoclastic effects of C. coreana Uyeki flos extract, which can be applied therapeutically for the treatment of osteoporosis.

scholarly journals Inhibitory Effects of Gold and Silver Nanoparticles on the Differentiation into Osteoclasts In Vitro

Pharmaceutics ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 462
Author(s):  
Daye Lee ◽  
Wan-Kyu Ko ◽  
Seong Jun Kim ◽  
In-Bo Han ◽  
Je Beom Hong ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Cathepsin K ◽  
Inhibitory Effect ◽  
Tartrate Resistant Acid Phosphatase ◽  
Nuclear Factor ◽  
Inhibitory Effects ◽  
Actin Ring ◽  
Activated T Cells ◽  
Gold And Silver Nanoparticles

Gold nanoparticles (GNPs) have been widely studied to inhibit differentiation into osteoclasts. However, reports of the inhibitory effects of silver nanoparticles (SNPs) during the process of differentiation into osteoclasts are rare. We compared the inhibitory effect of GNPs and SNPs during the process of differentiation into osteoclasts. Bone marrow-derived cells were differentiated into osteoclasts by the receptor activator of the nuclear factor-kappa-Β ligand (RANKL). The inhibitory effect of GNPs or SNPs during the process of differentiation into osteoclasts was investigated using tartrate-resistant acid phosphatase (TRAP) and actin ring staining. The formation of TRAP positive (+) multinuclear cells (MNCs) with the actin ring structure was most inhibited in the SNP group. In addition, the expression of specific genes related to the differentiation into osteoclasts, such as c-Fos, the nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), TRAP, and Cathepsin K (CTSK) were also inhibited in the SNP groups. As a result, the levels related to differentiation into osteoclasts were consistently lower in the SNP groups than in the GNP groups. Our study suggests that SNPs can be a useful material for inhibiting differentiation into osteoclasts and they can be applied to treatments for osteoporosis patients.

scholarly journals Inhibitory Effects of Astaxanthin on CML-HSA-Induced Inflammatory and RANKL-Induced Osteoclastogenic Gene Expression in RAW 264.7 Cells

Biomedicines ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 54
Author(s):  
A. N. M. Mamun-Or-Rashid ◽  
Tanzima Tarannum Lucy ◽  
Masayuki Yagi ◽  
Yoshikazu Yonei
Keyword(s):  
Gene Expression ◽  
Raw 264.7 Cells ◽  
Enzyme Linked Immunosorbent Assay ◽  
Marker Genes ◽  
Tartrate Resistant Acid Phosphatase ◽  
Raw 264.7 ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Inflammatory Gene Expression ◽  
Inflammatory Gene

Objective: Elevated levels of serum Nε-carboxymethyllysine (CML), a well-known advanced glycation end-product (AGE), were observed in patients with inflammation or osteoporosis. Astaxanthin was reported to possess anti-inflammatory and antioxidant effects. In the present study, we investigated the effects of commercially available dietary supplement AstaReal ACTR (ASR) capsule content as astaxanthin on CML-HSA-induced inflammatory and receptor activator of nuclear factor-kappa-Β ligand (RANKL)-induced osteoclastogenic gene expression. Methods: RAW 264.7 murine macrophage cells were stimulated with CML-HSA to trigger inflammatory gene expression and treated with either a vehicle control or varied concentrations of astaxanthin. Inflammatory gene expression was measured using an enzyme-linked immunosorbent assay (ELISA) or qPCR. We triggered osteoclastogenesis using RANKL, and osteoclastogenic gene expression was measured through tartrate-resistant acid phosphatase (TRAP) activity, staining, immunofluorescence, and qPCR analyses. Results: CML-HSA showed a stimulatory effect on inflammatory gene expression, and astaxanthin reduced the expression by at least two-fold. The levels of autoinflammatory gene expression were reduced by astaxanthin. The RANKL-induced osteoclastogenesis was significantly inhibited by astaxanthin, with reductions in the activation of nuclear factor-κB (NF-κB), the expression of NFATc1 (nuclear factor of activated T cells 1), multinucleated cell formation, and the expression of mature osteoclast marker genes. Conclusion: Astaxanthin has potential as a remedy for CML-HSA-induced inflammation and RANKL-induced excessive bone loss.

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

2019 ◽  
Vol 8 (4) ◽  
pp. 431 ◽  
Author(s):  
Mi Kim ◽  
Won Kim ◽  
Jae-Eun Byun ◽  
Jung Choi ◽  
Suk Yoon ◽  
...  
Keyword(s):  
Bone Loss ◽  
Osteoclast Differentiation ◽  
Bone Diseases ◽  
Raw 264.7 Cells ◽  
Tartrate Resistant Acid Phosphatase ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Interacting Protein ◽  
Hematopoietic Stem ◽  
Thioredoxin Interacting Protein

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.

scholarly journals Inhibition of osteoclastogenesis and inflammation by phosvitin and phosvitin hydrolysate via NF-κB and MAPK pathways in RAW 264.7 cells

2021 ◽  
Vol 13 ◽  
Author(s):  
Jiandong Ren ◽  
Subhadeep Chakrabarti ◽  
Jianping Wu
Keyword(s):  
Raw264.7 Cells ◽  
Raw 264.7 Cells ◽  
Tartrate Resistant Acid Phosphatase ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Necrosis Factor Alpha ◽  
Monocyte Chemoattractant Protein 1 ◽  
Tnf Α ◽  
Factor Alpha ◽  
Oxide Synthase

Phosvitin (PV) is an egg protein. Our recent study showed both phosvitin and phosvitin hydrolysate (PVH) could promote osteoblast differentiation in osteoblast cells. The objective of the study was to investigate the effects of PV and PVH on osteoclastogenesis and possible signalling pathways in RAW264.7 cells. Both PV and PVH inhibited osteoclastogenesis (fewer tartrate-resistant acid phosphatase (TRAP) positive cells and lower TRAP activity), reduced levels of transcription factors, c-Fos and NFATc1 (nuclear factor of activated T-cells, cytoplasmic 1), and suppressed inflammatory biomarkers TNF-α (tumor necrosis factor alpha), MCP-1 (monocyte chemoattractant protein 1), RANTES (regulated on activation, normal T cell expressed and secreted), and inducible nitric oxide synthase. The inhibitory effects of PV and PVH on RAW264.7 cells differentiation were likely mediated through p38, c-Jun N-terminal kinases (JNK) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways. These results indicated that PV and PVH might inhibit bone resorption activities.  

scholarly journals DIHYDROTESTOSTERONE DOWNREGULATES BONE RESORPTION ACTIVITY OF OSTEOCLASTS IN DOSE DEPENDENT MANNER: AN IN VITRO MODEL USING RAW 264.7 CELLS

2017 ◽  
Vol 9 (10) ◽  
pp. 86
Author(s):  
Hnin Ei Thu ◽  
Zahid Hussain ◽  
Isa Naina Mohamed ◽  
Ahmad Nazrun Shuid
Keyword(s):  
Bone Resorption ◽  
Cathepsin K ◽  
Raw 264.7 Cells ◽  
Tartrate Resistant Acid Phosphatase ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Sod Activity ◽  
Osteoclastic Differentiation ◽  
Dose Dependent

Objective: Numerous studies have evidenced the bone regulatory potential of dihydrotestosterone in androgen-deficient osteoporosis. The present study was thus aimed to explore the translational mechanism of dihydrotestosterone to down-regulate the bone resorption activity of osteoclasts using RAW 264.7 cells as in vitro model.Methods: Prior to analyze the efficacy of dihydrotestosterone (5α-DHT) to alleviate osteoclastic differentiation, their cell viability and cell proliferative ability was assessed using lactate dehydrogenase (LDH) and MTS assays. The osteoclastic differentiation capacity of dihydrotestosterone was evaluated by measuring TRAP activity and the expression of bone resorption-related proteins such as matrix metallopeptidase-9 (MMP-9), cathepsin-K, tartrate-resistant acid phosphatase (TRAP) and NFATc1. Moreover, the effects of dihydrotestosterone were also evaluated on superoxide (free radicals) generation and superoxide dismutase (SOD) activity in RANKL-induced osteoclasts.Results: Dihydrotestosterone showed no toxicity towards RAW 264.7 cells and significantly enhanced their proliferation and growth rates in a dose-dependent fashion. It was also observed that dihydrotestosterone exhibits a remarkable inhibitory effect on differentiation, maturation and activation of osteoclasts. The marked inhibition of differentiation and activation of osteoclasts caused by 5α-DHT was due to down-regulation of the expression of MMP-9, cathepsin-K, TRAP, NFATc1, generation of superoxide and up-regulation of SOD activity in the RAW 264.7 cells.Conclusion: Resulting data provided substantially in vitroevidence for the pronounced anti-osteoclastogenetic activity of dihydrotestosterone and its therapeutic value in treating osteoporosis and other bone-erosive disorders. 

scholarly journals Stimulation of Resorption in Cultured Mouse Calvarial Bones by Thiazolidinediones

Endocrinology ◽  
2005 ◽  
Vol 146 (10) ◽  
pp. 4349-4361 ◽  
Author(s):  
A. M. Schwab ◽  
S. Granholm ◽  
E. Persson ◽  
B. Wilkes ◽  
U. H. Lerner ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Osteoclast Differentiation ◽  
Cathepsin K ◽  
Nuclear Factor Κb ◽  
Tartrate Resistant Acid Phosphatase ◽  
Peroxisome Proliferator ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Peroxisome Proliferator Activated Receptor ◽  
Stimulation Of

Dosage-dependent release of 45Ca was observed from prelabeled mouse calvarial bones after treatment with two thiazolidinediones, troglitazone and ciglitazone. Release of 45Ca by ciglitazone was decreased by the osteoclast inhibitors acetazolamide, calcitonin, 3-amino-1-hydroxypropylidene-1,1-bisphosphonate, and IL-4, but not affected by the peroxisome proliferator-activated receptor γ antagonist, GW 9662, the mitotic inhibitor, hydroxyurea, or indomethacin. Enhanced expression of receptor activator of nuclear factor-κB ligand (RANKL) mRNA and protein and decreased osteoprotegerin (OPG) mRNA and protein were noted after ciglitazone treatment of calvariae. Ciglitazone and RANKL each caused increased mRNA expression of osteoclast markers: calcitonin receptor, tartrate-resistant acid phosphatase, cathepsin K, matrix metalloproteinase-9, integrin β3, and nuclear factor of activated T cells 2. OPG inhibited mRNA expression of RANKL stimulated by ciglitazone, mRNA expression of osteoclast markers stimulated by ciglitazone and RANKL, and 45Ca release stimulated by troglitazone and ciglitazone. Increased expression of IL-1α mRNA by ciglitazone was not linked to resorption stimulated by the thiazolidinedione. Ciglitazone did not increase adipogenic gene expression but enhanced osteocalcin mRNA in calvariae. In addition to exhibiting sensitivity to OPG, data indicate that stimulation of osteoclast differentiation and activity by thiazolidinediones may occur by a nonperoxisome proliferator-activated receptor γ-dependent pathway that does not require cell proliferation, prostaglandins, or IL-1α but is characterized by an increased RANKL to OPG ratio.

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

2019 ◽  
Vol 20 (6) ◽  
pp. 1439 ◽  
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.

scholarly journals Amaranthus mangostanus Inhibits the Differentiation of Osteoclasts and Prevents Ovariectomy-Induced Bone Loss

2020 ◽  
Vol 2020 ◽  
pp. 1-11 ◽  
Author(s):  
Yeon-Hui Jeong ◽  
Haeng Jeon Hur ◽  
Ae Sin Lee ◽  
Sang Hee Lee ◽  
Mi Jeong Sung
Keyword(s):  
Bone Loss ◽  
Biological Activities ◽  
Osteoclast Differentiation ◽  
Bone Destruction ◽  
Cathepsin K ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Potential Candidate ◽  
Bone Homeostasis ◽  
Activated T Cells

Bone homeostasis is dynamically balanced between bone forming osteoblasts and bone resorbing osteoclasts. Osteoclasts play an important role in bone destruction and osteoporosis, and they are derived from monocyte/macrophages in response to macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor κB (NF-κB) ligand (RANKL). Amaranthus mangostanus L. (AM) is a plant with powerful antioxidant and other biological activities including anti-inflammatory, antidiabetic, and antihyperlipidemic effects. However, its effects on bone health are unknown. In this study, we explored whether AM could affect RANK-mediated osteoclastogenesis. AM significantly suppressed RANKL-induced osteoclast differentiation and expression of osteoclast-specific genes, TRAP, cathepsin K, NF-activated T-cells (NFATc1), and Dc-stamp in RAW 264.7 cells. Moreover, AM significantly inhibited extracellular signal-regulated kinase (ERK), Akt, and NF-κB signaling pathways in RAW 264.7 cells. In addition, AM preserved ovariectomy-induced bone loss in mice. Taken together, our results suggest that AM might be a potential candidate for the treatment of postmenopausal osteoporosis.

scholarly journals RANKL and TNF-α-induced JNK/SAPK Osteoclastogenic Signaling Pathway was Inhibited by Caffeic Acid in RAW-D Cells

2018 ◽  
Vol 9 (2) ◽  
pp. 63
Author(s):  
Ferry Sandra ◽  
Junita Briskila ◽  
Ketherin Ketherin
Keyword(s):  
Caffeic Acid ◽  
Cathepsin K ◽  
Mitogen Activated Protein Kinase ◽  
Control Treatment ◽  
Tartrate Resistant Acid Phosphatase ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Therapeutic Benefits ◽  
Mitogen Activated Protein ◽  
D Cells

Caffeic acid, a natural substance found majorly in fruits, grains, and herbs, is known to have therapeutic benefits. One of which is to inhibit bone resorption by targeting osteoclastogenesis through inhibition of the Cathepsin K, p38 Mitogen-activated Protein Kinase (MAPK), Nuclear Factor of Activated T-cells c1 (NFATc1) and Nuclear Factor kB (NFkB). Besides p38 MAPK, the c-Jun N-terminal kinase (JNK) / stress-activated protein kinases (SAPK), another member of MAPK family, has been reported to play important roles in osteoclastogenesis. Hence, current study was undertaken in order to investigate mechanism of Caffeic Acid towards JNK/SAPK pathway. Tartrate Resistant Acid Phosphatase (TRAP) staining was performed on caffeic acid-treated and RANKL-TNFα-induced RAW-D cells. Western blot analysis was performed to detect JNK/SAPK and phosphorylated-JNK/SAPK. Protein bands were quantified and statistically analyzed. Treatment of 10 μg/mL Caffeic Acid inhibited 20 ng/mL RANKL and 1 ng/mL TNFα-induced RAW-D differentiation into TRAP+ osteoclast-like polynuclear cells. Induction of 20 ng/mL of RANKL and 1 ng/mL of TNFα for 0.2 or 1 hour, significantly increase phosphorylation of JNK/SAPK as compared with control. Treatment of 10 µg/mL Caffeic Acid significantly inhibited the 20 ng/mL of RANKL and 1 ng/mL of TNFα-induced phosphorylation of JNK/SAPK. Taken together, Caffeic Acid could inhibit the RANKL and TNFα-induced osteoclastogenesis through JNK/SAPK.

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