Induced Osteoclast Differentiation Research Articles

Ginseng (Panax ginseng) leaf extract modulates the expression of heme oxygenase-1 to attenuate osteoclast differentiation

Osteoporosis is an aging disease characterized by an imbalance between bone formation and resorption. However, drugs that inhibit bone resorption have various adverse effects. Ginseng (Panax ginseng), a prominent herbal medicine in East Asia for >2000 years, is renowned for its manifold beneficial properties, including antioxidant, anti-cancer, anti-diabetic, and anti-adipogenic activities. Despite its long history of use, the pharmacological functions of ginseng leaves are not yet fully comprehended. In this study, we evaluated the potential effects of ginseng leaf extract (GLE) on receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclast differentiation in RAW264.7 macrophage cells. Tartrate-resistant acid phosphatase (TRAP) staining revealed that GLE had significant anti-osteoclastogenic activity. GLE significantly reduced mRNA levels of osteoclast differentiation markers including TRAP, nuclear factor of activated T cell cytoplasmic 1, and cathepsin K. It also suppressed the production of reactive oxygen species (ROS) and secretion of high mobility group box-1 (HMGB1) in RANKL-treated RAW264.7 cells. In addition, GLE upregulated dose- and time-dependently the expression of heme oxygenase-1 (HO-1), eventually suppressing ROS production and HMGB1 secretion. This effects of GLE were significantly reversed by Tin Protoporphyrin IX dichloride, an inhibitor of HO-1, and HO-1 shRNA, indicating that HO-1 potently inhibits RANKL-induced osteoclast differentiation by inhibiting ROS production and HMGB1 secretion. Taken together, these observations suggest that GLE could have therapeutic potential as a natural product-derived medicine for the treatment of bone disorders.

Oxyresveratrol attenuates bone resorption by inhibiting the mitogen-activated protein kinase pathway in ovariectomized rats

BackgroundBone is continuously produced by osteoblasts and resorbed by osteoclasts to maintain homeostasis. Impaired bone resorption by osteoclasts causes bone diseases such as osteoporosis and arthritis. Most pharmacological treatment of osteoporosis focuses on inhibiting osteoclast differentiation, often to restore osteoclast/osteoclast balance. However, recent osteoporosis treatments have various side effects. According to a recent study, resveratrol, known as a stilbenoid family, is known to increase bone density, and the osteoclast inhibitory effect was confirmed using oxyresveratrol, a stilbenoid family. Here, we investigated the effect of oxyresveratrol on osteoclast differentiation and an ovariectomized mouse model.MethodsMouse leukemia monocyte/macrophage cell line RAW 264.7 was treated with oxyresveratrol, and cell cytotoxicity was confirmed by measuring MTT assay. Tartrate-resistant acid phosphatase (TRAP), an enzyme marker for osteoclasts, was confirmed by staining. In addition, osteoclast differentiation markers and MAPK-related markers were confirmed at the mRNA level and protein expression. The effect of oxyresveratrol was confirmed using ovariectomized mice. Deoxypyridinoline (DPD) was measured using mouse urine and TRAP activity was observed using serum. Bone mineral density was also measured using Micro-CT.ResultsThe polyphenol oxyresveratrol inhibited receptor activator of nuclear factor kappa-Β ligand (RANKL)-induced osteoclast differentiation of RAW 264.7 cells. Furthermore, oxyresveratrol inhibited TRAP activity and actin-ring formation. Moreover, oxyresveratrol suppressed the phosphorylation of the RANKL-induced mitogen-activated protein kinases (MAPKs) p38, JNK, and ERK and significantly reduced the expression of bone differentiation markers (NFATc1, cathepsin K, and TRAP).ConclusionOxyresveratrol inhibits osteoclast differentiation via MAPK and increases bone density in ovariectomized rats, suggesting it has therapeutic potential for bone diseases such as osteoporosis. We confirmed the osteoporosis prevention effect of OR in Raw 264.7 cells, and future studies should confirm the effect of OR using rat bone marrow-derived cells.

MCP-5 suppresses osteoclast differentiation through Ccr5 upregulation.

Human monocyte chemoattractant protein-1 (MCP-1) in mice has two orthologs, MCP-1 and MCP-5. MCP-1, which is highly expressed in osteoclasts rather than in osteoclast precursor cells, is an important factor in osteoclast differentiation. However, the roles of MCP-5 in osteoclasts are completely unknown. In this study, contrary to MCP-1, MCP-5 was downregulated during receptor activator of nuclear factor kappa B ligand(RANKL)-induced osteoclast differentiation and was considered an inhibitory factor in osteoclast differentiation. The inhibitory role of MCP-5 in osteoclast differentiation was closely related to the increase in Ccr5 expression and the inhibition of IκB degradation by RANKL. Transgenic mice expressing MCP-5 controlled by Mx-1 promoter exhibited an increased bone mass because of a decrease in osteoclasts. This result strongly supported that MCP-5 negatively regulated osteoclast differentiation. MCP-5 also prevented severe bone loss caused by RANKL.

Expression and functional characterization of bovine receptor activator of NF-κB ligand (RANKL)

Receptor activator of nuclear factor Kappa-B Ligand (RANKL) is a member of the tumor necrosis factor ligand (TNF) family involved in immune responses and immunomodulation. Expressed in various cells types around the body, RANKL plays a crucial role in bone remodeling and development of the thymus, lymph nodes and mammary glands. Research in other species demonstrates that RANKL is required for the development of microfold cells (M cells) in the gut, however limited information specific to cattle is available. Cloning and expression of bovine RANKL (BoRANKL) was carried out and bioactivity of the protein was demonstrated in the induction of osteoclast differentiation from both bovine and ovine bone marrow cells. The effects of BoRANKL on particle uptake in bovine enteroids was also assessed. The production of cross-reactive bovine RANKL protein will enable further investigations into cell differentiation using the available ruminant organoid systems, and their role in investigating host-pathogen interactions in cattle and sheep.

Nrf2 Mitigates RANKL and M-CSF Induced Osteoclast Differentiation via ROS-Dependent Mechanisms.

Nuclear factor-erythroid 2-related factor 2 (Nrf2) has been shown to be a negative regulator of osteoclast differentiation, but the precise mechanisms have not yet been established. We examined the precise roles of Nrf2 in regulating antioxidants and reactive oxygen species (ROS) levels, especially the cytoplasmic and mitochondrial ROS during osteoclastogenesis in vitro. In the current study, we found that the absence of Nrf2 promotes osteoclast differentiation in bone-marrow-derived macrophages (BMMs) and RAW 264.7 cells. The receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) significantly lowered the levels of Nrf2 and its downstream antioxidant enzymes at mRNA and/or protein levels during osteoclast differentiation in the BMMs of mice and RAW 264.7 mouse leukemic monocytes. Compared to the wild-type cells, Nrf2-deficient cells exhibited heightened sensitivity to both transient RANKL-induced cytoplasmic ROS and prolonged RANKL and M-CSF-induced cytoplasmic and mitochondrial ROS accumulation. Furthermore, exogenous antioxidant agents, including N-acetyl-cysteine (NAC), diphenyleneiodonium chloride (DPI), and mitoquinone mesylate (MitoQ), exhibited substantial capability to suppress the elevation of ROS levels during osteoclast differentiation induced by Nrf2 deficiency, and they consequently inhibited osteoclast differentiation augmented by the lack of Nrf2. The activation of phosphorylated c-FOS resulting from elevated ROS promoted osteoclast differentiation. The inhibition of c-FOS blocked osteoclast differentiation, which was elevated by Nrf2-deficiency. Taken together, these data reveal that Nrf2 effectively decreased the accumulation of intracellular ROS and the phosphorylation of c-FOS during osteoclastic differentiation by regulating antioxidant enzymes and subsequently inhibited RANKL-induced osteoclast differentiation.

Vascular Pericyte-Derived Exosomes Inhibit Bone Resorption via Traf3.

Blood vessels distribute cells, oxygen, and nutrients throughout the body to support tissue growth and balance. Pericytes and endothelial cells form the inner wall of blood vessels, crucial for organ development and tissue homeostasis by producing paracrine signaling molecules. In the skeletal system, pericyte-derived vascular factors along with angiogenic factors released by bone cells regulate angiogenesis and bone formation. Although the involvement of angiogenic factors and skeletal blood vessels in bone homeostasis is relatively clear, the role of pericytes and the underlying mechanisms remain unknown. Here, our objective was to elucidate the significance of pericytes in regulating osteoclast differentiation. We used tissue staining to detect the coverage of pericytes and osteoclasts in femoral tissues of osteoporotic mice and mice of different ages, analyzing their correlation. We developed mice with conditionally deleted pericytes, observing changes in bone mass and osteoclast activity using micro-computer tomography and tissue staining to detect the regulatory effect of pericytes on osteoclasts. Pericytes-derived exosomes (PC-EVs) were collected and co-cultured with monocytes that induce osteoclast differentiation to detect the effect of the former on the exosomes. Finally, the specific mechanism of PC-EVs regulating osteoclast differentiation was verified using RNA sequencing and Western blotting. Our study indicates a significant correlation between pericytes and age-related bone resorption. Conditional deletion of pericytes activated bone resorption and led to osteopenia in vivo. We discovered that PC-EVs inhibited the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway, which is mediated by tumor necrosis factor receptor-associated factor 3 (Traf3), negatively regulating osteoclast development and bone resorption. Silencing Traf3 in PC-EVs canceled their inhibitory effect on osteoclast differentiation. Our study provides a novel perspective into the regulatory role of pericytes on bone resorption and may provide potential strategies for developing novel anti-bone resorption therapies.

Staphylococcus aureus enhances osteoclast differentiation and bone resorption by stimulating the NLRP3 inflammasome pathway.

Osteomyelitis is one of the most challenging infectious diseases and is mainly caused by Staphylococcus aureus (S. aureus). In this study, we analyzed the effect of S. aureus on osteoclast differentiation and its possible molecular mechanism. We cultured RAW 264.7 cells with live S. aureus for 5 days. We assessed cell viability and the formation of resorption pits. We tested the NLRP3 inflammasome signaling pathways and measured the mRNA expression levels of osteoclastspecific genes, including TRAP, MMP9, cathepsin K, calcitonin receptor and ATP6V0d2. Furthermore, we analyzed the protein expression levels of the protein in the NF-κB and p38 MAPK signaling pathways to clarify the signaling pathways by which S. aureus promotes osteoclast differentiation. Staphylococcus aureus induced NLRP3 inflammasome activation. S. aureus promoted bone resorption and enhanced the expression of osteoclastspecific genes, such as TRAP, MMP9, cathepsin K, calcitonin receptor and ATP6V0d2. MCC950 was used to inhibit NLRP3 inflammasome activity. Osteoclast differentiation and the expression of osteoclastspecific genes induced by S. aureus were inhibited by MCC950 pretreatment. The degradation of IκBα and phosphorylation of P65 were increased under the induction of S. aureus, but proteins in the p38 MAPK signaling pathway did not change significantly. Staphylococcus aureus induces osteoclast differentiation and promotes bone resorption in vitro, and the NLRP3 inflammasome signaling pathway plays a significant role in this process. S. aureus-induced NLRP3 inflammasome activation was mainly dependent on the NF-κB signaling pathway during osteoclastogenesis.

The Capacity of Magnesium to Induce Osteoclast Differentiation Is Greatly Enhanced by the Presence of Zoledronate.

Bisphosphonates (BPs) are successfully used to cure a number of diseases characterized by a metabolic reduction in bone density, such as Osteoporosis, or a neoplastic destruction of bone tissue, such as multiple myeloma and bone metastases. These drugs exert their therapeutic effect by causing a systemic osteoclast depletion that, in turn, is responsible for reduced bone resorption. Unfortunately, in addition to their beneficial activity, BPs can also determine a frightening side effect known as osteonecrosis of the jaw (ONJ). It is generally believed that the inability of osteoclasts to dispose of inflamed/necrotic bone represents the main physiopathological aspect of ONJ. In principle, a therapeutic strategy able to elicit a local re-activation of osteoclast production could counteract ONJ and promote the healing of its lesions. Using an experimental model of Vitamin D3-dependent osteoclastogenesis, we have previously demonstrated that Magnesium is a powerful inducer of osteoclast differentiation. Here we show that, surprisingly, this effect is greatly enhanced by the presence of Zoledronate, chosen for our study because it is the most effective and dangerous of the BPs. This finding allows us to hypothesize that Magnesium might play an important role in the topical therapy of ONJ.

Surface proteins of Propionibacterium freudenreichii MJ2 inhibit RANKL-induced osteoclast differentiation by lipocalin-2 upregulation and lipocalin-2-mediated NFATc1 inhibition

Osteoclasts degrade bone and osteoclast differentiation has been implicated in bone destruction in rheumatoid arthritis. The dairy bacterium Propionibacterium freudenreichii MJ2 (MJ2) isolated from raw milk inhibits osteoclast differentiation and ameliorates collagen-induced arthritis. This study aimed to investigate the inhibitory effect of the surface proteins of MJ2 on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and explain the underlying mechanism. The murine macrophage cell line RAW 264.7 was used to study the inhibition of osteoclast differentiation. The surface proteins significantly inhibited RANKL-induced osteoclast differentiation in a protein concentration-dependent manner by inhibiting the expression of genes and proteins related to osteoclast differentiation. RNA microarray analysis showed that the surface proteins significantly upregulated lipocalin-2 (lcn2) expression. In addition, they downregulated c-fos and NFATc1 and inhibited the expression of NFATc1-downstream genes Atp6v0d2, Calcr, and Ctsk. siRNA silencing of lcn2 decreased the extent of surface protein inhibition on osteoclast differentiation, suggesting that lcn2 plays an important role in the inhibition of RANKL-induced osteoclast differentiation. In conclusion, surface proteins of MJ2 show inhibitory effects on RANKL-induced osteoclast differentiation by upregulating lcn2 expression, in turn downregulating NFATc1, leading to the inhibition of NFATc1-downstream osteoclastogenesis-related gene expression.

P7C3 Ameliorates Bone Loss by Inhibiting Osteoclast Differentiation and Promoting Osteogenesis.

Bone homeostasis, the equilibrium between bone resorption and formation, is essential for maintaining healthy bone tissue in adult humans. Disruptions of this process can lead to pathological conditions such as osteoporosis. Dual-targeted agents, capable of inhibiting excessive bone resorption and stimulating bone formation, are being explored as a promising strategy for developing new treatments to address osteoporosis. In this study, we investigated the effects of P7C3 on bone remodeling and its potential therapeutic role in osteoporosis treatment in mice. Specifically, P7C3 can remarkably suppress receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclast differentiation in bone marrow macrophages via the Akt-NF-κB-NFATc1 signaling pathway. Additionally, RNA sequencing (RNAseq) analysis revealed that P7C3 promoted osteoblast differentiation and function through the Wnt/β-catenin signaling pathway, thereby enhancing bone formation. Furthermore, μCT analysis and histological examination of bone tissues from P7C3-treated mice showed attenuation of both Ti-induced bone erosion and ovariectomy (OVX)-induced bone loss. These findings suggest that P7C3 may have a novel function in bone remodeling and may be a promising therapeutic agent for the treatment of osteoporosis. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Macrophage-derived exosomes modulate wear particle-induced osteolysis via miR-3470b targeting TAB3/NF-κB signaling

Aseptic prosthesis loosening (APL) is one of the most prevalent complications associated with arthroplasty. The main cause is the periprosthetic osteolysis induced by wear particles. However, the specific mechanisms of crosstalk between immune cells and osteoclasts/osteoblasts during osteolysis are unclear. In this study, we report the role and mechanism of macrophage-derived exosomes in wear particle-induced osteolysis. The results of exosomes up-taken experiments revealed that osteoblast and mature osteoclasts capture macrophage-derived exosomes (M-Exo). Next-generation sequencing and RT-qPCR on M-Exo revealed that exosomal microRNA miR-3470b was downregulated in wear particle-induced osteolysis. The results of analysis on Luciferase reporter assays/fluorescence in situ hybridization (FISH)/immunofluorescence (IF)/immunohistochemistry (IHC) and co-culture experiments demonstrated that wear particles induced osteoclast differentiation by increasing the expression of NFatc1 via M-Exo miR-3470b targeting TAB3/ NF-κB signaling. We also illustrate that engineered exosomes enriching miR-3470b facilitated to suppressed the osteolysis; the microenvironment enriching with miR-3470b could suppress wear particle-induced osteolysis via inhibition of TAB3/ NF-κB in vivo. In summary, our findings indicate that macrophage-derived exosomes transfer to osteoclasts to induce osteolysis in wear particle-induced APL. Engineering exosomes enriching with miR-3470b might be a novel strategy for the targeting treatment of bone resorption-related diseases.

Pharmacological inhibition of protein S-palmitoylation suppresses osteoclastogenesis and ameliorates ovariectomy-induced bone loss

BackgroundExcessive osteoclast formation disrupts bone homeostasis, thereby significantly contributing to pathological bone loss associated with a variety of diseases. Protein S-palmitoylation is a reversible post-translational lipid modification catalyzed by ZDHHC family of palmitoyl acyltransferases, which plays an important role in various physiological and pathological processes. However, the role of palmitoylation in osteoclastogenesis has never been explored. Consequently, it is unclear whether this process can be targeted to treat osteolytic bone diseases that are mainly caused by excessive osteoclast formation. Materials and methodsIn this study, we employed acyl-biotin exchange (ABE) assay to reveal protein S-palmitoylation in differentiating osteoclasts (OCs). We utilized 2-bromopalmitic acid (2-BP), a pharmacological inhibitor of protein S-palmitoylation, to inhibit protein palmitoylation in mouse bone marrow-derived macrophages (BMMs), and tested its effect on receptor activator of nuclear factor κβ ligand (RANKL)-induced osteoclast differentiation and activity by TRAP staining, phalloidin staining, qPCR analyses, and pit formation assays. We also evaluated the protective effect of 2-BP against estrogen deficiency-induced bone loss and bone resorption in ovariectomized (OVX) mice using μCT, H&E staining, TRAP staining, and ELISA assay. Furthermore, we performed western blot analyses to explore the molecular mechanism underlying the inhibitory effect of 2-BP on osteoclastogenesis. ResultsWe found that many proteins were palmitoylated in differentiating OCs and that pharmacological inhibition of palmitoylation impeded RANKL-induced osteoclastogenesis, osteoclast-specific gene expression, F-actin ring formation and osteoclastic bone resorption in vitro, and to a lesser extent, osteoblast formation from MC3T3-E1 cells. Furthermore, we demonstrated that administration of 2-BP protected mice from ovariectomy-induced osteoporosis and bone resorption in vivo. Mechanistically, we showed that 2-BP treatment inhibited osteoclastogenesis partly by downregulating the expression of c-Fos and NFATc1 without overtly affecting RANKL-induced activation of osteoclastogenic AKT, MAPK, and NF-κB pathways. ConclusionPharmacological inhibition of palmitoylation potently suppresses RANKL-mediated osteoclast differentiation in vitro and protects mice against OVX-induced osteoporosis in vivo. Mechanistically, palmitoylation regulates osteoclast differentiation partly by promoting the expression of c-Fos and NFATc1. Thus, palmitoylation plays a key role in promoting osteoclast differentiation and activity, and could serve as a potential therapeutic target for the treatment of osteoporosis and other osteoclast-related diseases. The translational potential of this articleThe translation potential of this article is that we first revealed palmitoylation as a key mechanism regulating osteoclast differentiation, and therefore provided a potential therapeutic target for treating osteolytic bone diseases.

AMPK negatively regulates RANKL-induced osteoclast differentiation by controlling oxidative stress

AMP-activated protein kinase (AMPK) is a crucial energy sensor of cellular metabolism under various metabolic stresses, such as oxidative stress and inflammation. AMPK deficiency increases osteoclast numbers and reduces bone mass; however, the precise mechanisms remain unclear. This study aimed to clarify the mechanistic connection between AMPK and osteoclast differentiation, and the potential role of AMPK in the anti-resorptive effects of several phytochemicals. We found that receptor activator of nuclear factor-kappa B (NF-κB) ligand (RANKL)-induced osteoclast differentiation, osteoclastic gene expression, and activation of mitogen-activated protein kinase (MAPK) and NF-κB were promoted in cells transfected with AMPK siRNA. AMPK knockdown led to defective synthesis of heme oxygenase-1, an antioxidant enzyme, and the upstream mediator, nuclear factor erythroid-2-related factor 2. Furthermore, treatment with N-acetyl-l-cysteine, an antioxidant, abolished osteoclast differentiation and MAPK/NF-κB activation induced by AMPK knockdown. AMPK activators, hesperetin, gallic acid, resveratrol, and curcumin, suppressed osteoclast differentiation via the activation of AMPK. These results suggest that AMPK inhibits RANKL-induced osteoclast differentiation by enhancing antioxidant defense system and regulating oxidative stress. AMPK activation by dietary-derived phytochemicals may be effective for the treatment of bone diseases.

Citropten Ameliorates Osteoclastogenesis Related to MAPK and PLCγ/Ca2+ Signaling Pathways through the Regulation of Amyloid Beta.

Osteoporosis and Alzheimer's disease are typical types of dementia in seniors, which share common risk factors. Previous studies have shown that citizens with osteoporosis are more likely than healthy individuals to be at risk of Alzheimer's disease. Citropten, found in Citrus aurantifolia, has been reported to have several pharmacological activities; however, its antiosteoclastogenic activity remains unknown. Here, receptor activator nuclear factor κB ligand (RANKL)-induced osteoclast differentiation, formation, and function in the presence of amyloid beta (Aβ) were attenuated by citropten in the RAW 264.7 cell line. The expression of osteoclast specific genes and proteins indicated that citropten pretreatment lowers the MAPK and PLCγ/Ca2+ signaling pathways. Molecular docking simulations revealed that citropten interacts with the active sites of proteins in the calcium signaling pathway, which have negative binding affinities. These findings indicate that, through Aβ regulation, the RANKL-induced osteoclast can be suppressed by citropten, suggesting that citropten is a potential candidate for treating osteoclastogenesis-related diseases.

New Meroterpenoids and Anti-Osteoclastogenic Polyketides from the Mangrove-Derived fungus Arthrinium sp. SCSIO 41306.

Two new meroterpenoids, arthrinones A and B (1 and 2), along with six known compounds (3-8), were obtained from the fungus Arthrinium sp. SCSIO 41306. Comprehensive methods such as chiral-phase HPLC analysis and ECD calculations were applied to determine the absolute configurations. Griseofulvin (5), kojic acid (6), and 1H-indole-3-carboxaldehyde (8) showed inhibition of NF-κB in RAW 264.7 macrophages induced by lipopolysaccharide (LPS) with IC50 values of 22.21, 13.87 and 19.31 μM, respectively. In addition, griseofulvin (5) inhibited receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation in a dose-dependent manner without visible evidence of cytotoxicity in bone marrow macrophages (BMMs). This is the first report on the activity of griseofulvin (5) to inhibit osteoclast formation (IC50 10.09±0.21 μM).

CUL4A-mediated ZEB1/microRNA-340-5p/HMGB1 axis promotes the development of osteoporosis.

Understanding the molecular mechanisms underlying osteoclast differentiation provides insights into bone loss and even osteoporosis. The specific mechanistic actions of cullin 4A (CUL4A) in osteoclast differentiation and resultant osteoporosis is poorly explored. We developed a mouse model of osteoporosis using bilateral ovariectomy (OVX) and examined CUL4A expression. It was noted that CUL4A expression was increased in the bone marrow of OVX mice. Overexpression of CUL4A promoted osteoclast differentiation, and knockdown of CUL4A alleviated osteoporosis symptoms of OVX mice. Bioinformatic analyses were applied to identify the downstream target genes of microRNA-340-5p (miR-340-5p), followed by interaction analysis. The bone marrow macrophages (BMMs) were isolated from femur of OVX mice, which were transfected with different plasmids to alter the expression of CUL4A, Zinc finer E-box binding homeobox 1 (ZEB1), miR-340-5p, and Toll-like receptor 4 (TLR4). ChIP assay was performed to detect enrichment of ZEB1 promoter by H3K4me3 antibody in BMMs. ZEB1 was overexpressed in the bone marrow of OVX mice. Overexpression of CUL4A mediated H3K4me3 methylation to increase ZEB1 expression, thus promoting osteoclast differentiation. Meanwhile, ZEB1 could inhibit miR-340-5p expression and upregulate HMGB1 to induce osteoclast differentiation. Overexpressed ZEB1 activated the TLR4 pathway by regulating the miR-340-5p/HMGB1 axis to induce osteoclast differentiation, thus promoting the development of osteoporosis. Overall, E3 ubiquitin ligase CUL4A can upregulate ZEB1 to repress miR-340-5p expression, leading to HMGB1 upregulation and the TLR4 pathway activation, which promotes osteoclast differentiation and the development of osteoporosis.

Chronic arthritides and bone structure: focus on rheumatoid arthritis-an update.

Normal bone remodeling depends of a balance between bone forming cells, osteoblasts and bone resorbing cells, the osteoclasts. In chronic arthritides and some inflammatory and autoimmune diseases such as rheumatoid arthritis, there is a great constellation of cytokines produced by pannus that impair bone formation and stimulate bone resorption by inducing osteoclast differentiation and inhibiting osteoblast maturation. Patients with chronic inflammation have multiple causes that lead to low bone mineral density, osteoporosis and a high risk of fracture including circulating cytokines, impaired mobility, chronic administration of glucocorticoids, low vitamin D levels and post-menopausal status in women, among others. Biologic agents and other therapeutic measures to reach prompt remission might ameliorate these deleterious effects. In many cases, bone acting agents need to be added to conventional treatment to reduce the risk of fractures and to preserve articular integrity and independency for daily living activities. A limited number of studies related to fractures in chronic arthritides were published, and future investigation is needed to determine the risk of fractures and the protective effects of different treatments to reduce this risk.

Monocyte Differentiation on Atomic Layer-Deposited (ALD) Hydroxyapatite Coating on Titanium Substrate.

Hydroxyapatite (HA; Ca10(PO4)6(OH)2) coating of bone implants has many beneficial properties as it improves osseointegration and eventually becomes degraded and replaced with new bone. We prepared HA coating on a titanium substrate with atomic layer deposition (ALD) and compared monocyte differentiation and material resorption between ALD-HA and bone. After stimulation with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa-B ligand (RANKL), human peripheral blood monocytes differentiated into resorbing osteoclasts on bovine bone, but non-resorbing foreign body cells were observed on ALD-HA. The analysis of the topography of ALD-HA and bone showed no differences in wettability (water contact angle on ALD-HA 86.2° vs. 86.7° on the bone), but the surface roughness of ALD-HA (Ra 0.713 µm) was significantly lower compared to bone (Ra 2.30 µm). The cellular reaction observed on ALD-HA might be a consequence of the topographical properties of the coating. The absence of resorptive osteoclasts on ALD-HA might indicate inhibition of their differentiation or the need to modify the coating to induce osteoclast differentiation.

Protective Effects of the Postbiotic Lactobacillus plantarum MD35 on Bone Loss in an Ovariectomized Mice Model

Postmenopausal osteoporosis is caused by estrogen deficiency, which impairs bone homeostasis, resulting in increased osteoclastic resorption without a corresponding increase in osteoblastic activity. Postbiotics have several therapeutic properties, including anti-obesity, anti-diabetic, anti-inflammatory, and anti-osteoporotic effects. However, the beneficial effects of the postbiotic MD35 of Lactobacillus plantarum on bone have not been studied. In this study, we demonstrated that the postbiotic L. plantarum MD35, isolated from young radish water kimchi, influences osteoclast differentiation in mouse bone marrow-derived macrophage (BMM) culture. In addition, it was effective protecting against estrogen deficiency-induced bone loss in ovariectomized (OVX) mice, an animal model of postmenopausal osteoporosis. In BMM cells, postbiotic MD35 inhibited the receptor activator of nuclear factor-kappa B of NF-κB ligand (RANKL)-induced osteoclast differentiation by attenuating the phosphorylation of extracellular signal-related kinase, significantly suppressing the resorption activity and down-regulating the expression of RANKL-mediated osteoclast-related genes. In the animal model, the oral administration of postbiotic MD35 remarkably improved OVX-induced trabecular bone loss and alleviated the destruction of femoral plate growth. Therefore, postbiotic MD35 could be a potential therapeutic candidate for postmenopausal osteoporosis by suppressing osteoclastogenesis through the regulation of osteoclast-related molecular mechanisms.

Periplocin targets low density lipoprotein receptor-related protein 4 to attenuate osteoclastogenesis and protect against osteoporosis

Osteoporosis is a common inflammaging-related condition, where long-term accumulation of pro-inflammatory cytokines causes massive bone loss. Periplocin, a cardiotonic steroid isolated from Periploca forrestii, has been proved to reduce inflammation in several inflammatory diseases, such as rheumatoid arthritis. However, its effect and mechanism of inflammation in osteoporosis, in which pro-inflammatory factors accelerate bone loss, has not been well demonstrated. In this study, periplocin attenuated receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation of bone marrow-derived macrophages (BMMs) and RAW264.7 cells in vitro. It reduced osteoclast numbers and bone resorption in a concentration- and time-dependent manner. Further, periplocin treatment resulted in reduced bone loss on mice with ovariectomy-induced osteoporosis in vivo. By transcriptome sequencing, periplocin was indicated to function through inhibition of the mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) signaling pathways and attenuating interactions between NF-κB and nuclear factor of activated T-cells 1 (NFATc1). It was further detected to bind low density lipoprotein receptor-related protein 4 (LRP4) in osteoclasts to exert anti-inflammatory and anti-osteoclastic effects. Overall, the findings have highlighted a better understanding for the anti-inflammatory and anti-osteoclastic role of periplocin in osteoporosis and its mechanism, bringing new possibilities for osteoporosis treatment.