Regulation Of Bone Resorption Research Articles

Energy Metabolism of Osteocytes.

In this review, we provide a recent update on bioenergetic pathways in osteocytes and identify potential future areas of research interest. Studies have identified a role for regulation of bone formation and bone resorption through osteocyte mechanosensing and osteocyte secreted factors. Nevertheless, there is a paucity of studies on the bioenergetics and energy metabolism of osteocytes, which are required for the regulation of bone remodeling. Osteocytes are cells of the osteoblast lineage embedded in bone. The osteocyte lacunocanalicular network within the skeletal matrix is exposed to a unique hypoxic environment. Therefore, the bioenergetic requirements of these cells could differ from other bone cells due to its location in the ossified matrix and its role in bone regulation transduced by mechanical signals. Recent findings highlighted in this review provide some evidence that metabolism of these cells is dependent on their location due to the substrates present in the microenvironment and metabolic cues from stress pathways. Both glycolysis (glucose metabolism) and oxidative phosphorylation (mitochondrial dynamics, ROS generation) affect osteocyte function and viability. In this review, we provide evidence that is currently available about information regarding bioenergetics pathways in osteocytes. We discuss published studies showing a role for hypoxia-driven glucose metabolism in regulating osteocyte bioenergetics. We also provide information on various substrates that osteocytes could utilize to fuel energetic needs, namely pyruvate, amino acids, and fatty acids. This is based on some preliminary experimental evidence that is available in literature. The role of parathyroid hormone PTH and parathryoid hormone-related peptide PTHrP in bone anabolism and resorption, along with regulation of metabolic pathways in the cells of the skeletal niche, needs to be explored further. Mitochondrial metabolism has a role in osteocyte bioenergetics through substrate utilization, location of the osteocyte in the bone cortex, and mitochondrial biogenesis. While there are limitations in studying metabolic flux in traditional cell lines, there are now novel cell lines and sophisticated tools available to study osteocyte bioenergetics to help harness its potential in vivo in the future.

The Role of GH/IGF Axis in Dento-Alveolar Complex from Development to Aging and Therapeutics: A Narrative Review.

The GH/IGF axis is a major regulator of bone formation and resorption and is essential to the achievement of normal skeleton growth and homeostasis. Beyond its key role in bone physiology, the GH/IGF axis has also major pleiotropic endocrine and autocrine/paracrine effects on mineralized tissues throughout life. This article aims to review the literature on GH, IGFs, IGF binding proteins, and their respective receptors in dental tissues, both epithelium (enamel) and mesenchyme (dentin, pulp, and tooth-supporting periodontium). The present review re-examines and refines the expression of the elements of the GH/IGF axis in oral tissues and their in vivo and in vitro mechanisms of action in different mineralizing cell types of the dento-alveolar complex including ameloblasts, odontoblasts, pulp cells, cementoblasts, periodontal ligament cells, and jaw osteoblasts focusing on cell-specific activities. Together, these data emphasize the determinant role of the GH/IGF axis in physiological and pathological development, morphometry, and aging of the teeth, the periodontium, and oral bones in humans, rodents, and other vertebrates. These advancements in oral biology have elicited an enormous interest among investigators to translate the fundamental discoveries on the GH/IGF axis into innovative strategies for targeted oral tissue therapies with local treatments, associated or not with materials, for orthodontics and the repair and regeneration of the dento-alveolar complex and oral bones.

Beraprost ameliorates postmenopausal osteoporosis by regulating Nedd4-induced Runx2 ubiquitination

Bone health requires adequate bone mass, which is maintained by a critical balance between bone resorption and formation. In our study, we identified beraprost as a pivotal regulator of bone formation and resorption. The administration of beraprost promoted differentiation of mouse bone mesenchymal stem cells (M-BMSCs) through the PI3K–AKT pathway. In co-culture, osteoblasts stimulated with beraprost inhibited osteoclastogenesis in a rankl-dependent manner. Bone mass of p53 knockout mice remained stable, regardless of the administration of beraprost, indicating that p53 plays a vital role in the bone mass regulation by beraprost. Mechanistic in vitro studies showed that p53 binds to the promoter region of neuronal precursor cell-expressed developmentally downregulated 4 (Nedd4) to promote its transcription. As a ubiquitinating enzyme, Nedd4 binds to runt-related transcription factor 2 (Runx2), which results in its ubiquitination and subsequent degradation. These data indicate that the p53–Nedd4–Runx2 axis is an effective regulator of bone formation and highlight the potential of beraprost as a therapeutic drug for postmenopausal osteoporosis.

Transcriptomics\u2010based analysis of the mechanism by which Wang-Bi capsule alleviates joint destruction in rats with collagen\u2010induced arthritis

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disease accompanied with joint destruction that often leads to disability. Wang-Bi capsule (WB), a traditional Chinese medicine-based herbs formula, has exhibited inhibition effect on joint destruction of collagen-induced arthritis (CIA) animal model in our previous study. But its molecular mechanisms are still obscure.MethodsCIA rats were treated intragastrical with WB for eight weeks, and the effect of joints protection were evaluated by hematoxylin and eosin (H&E) staining, safranin O fast green staining, tartrate-resistant acid phosphatase (TRAP) staining and micro‑CT scanning analysis. The transcriptomic of tarsal joints were used to investigate how WB alleviated joint destruction.ResultsThe histological examination of ankle joints showed WB alleviated both cartilage damage and bone destruction of CIA rats. This protective effect on joints were further evidenced by micro-CT analysis. The transcriptomic analysis showed that WB prominently changed 12 KEGG signaling pathways (“calcium signaling pathway”, “cAMP signaling pathway”, “cell adhesion molecules”, “chemokine signaling pathway”, “complement and coagulation cascades”, “MAPK signaling pathway”, “NF-kappa B signaling pathway”, “osteoclast differentiation”, “PI3K-Akt signaling pathway”, “focal adhesion”, “Gap junction” and “Rap1 signaling pathway”) associated with bone or cartilage. Several genes (including Il6, Tnfsf11, Ffar2, Plg, Tnfrsf11b, Fgf4, Fpr1, Siglec1, Vegfd, Cldn1, Cxcl13, Chad, Arrb2, Fgf9, Egfr) regulating bone resorption, bone formation and cartilage development were identified by further analysis. Meanwhile, these differentially expressed genes were validated by real-time quantitative PCR.ConclusionsOverall, the protective effect of WB treatment on joint were confirmed in CIA rats, and its basic molecular mechanisms may be associated with regulating some genes (including Il6, Tnfsf11, Ffar2 and Plg etc.) involved in bone resorption, bone formation and cartilage development.

Osteoclasts recycle via osteomorphs during RANKL-stimulated bone resorption

SummaryOsteoclasts are large multinucleated bone-resorbing cells formed by the fusion of monocyte/macrophage-derived precursors that are thought to undergo apoptosis once resorption is complete. Here, by intravital imaging, we reveal that RANKL-stimulated osteoclasts have an alternative cell fate in which they fission into daughter cells called osteomorphs. Inhibiting RANKL blocked this cellular recycling and resulted in osteomorph accumulation. Single-cell RNA sequencing showed that osteomorphs are transcriptionally distinct from osteoclasts and macrophages and express a number of non-canonical osteoclast genes that are associated with structural and functional bone phenotypes when deleted in mice. Furthermore, genetic variation in human orthologs of osteomorph genes causes monogenic skeletal disorders and associates with bone mineral density, a polygenetic skeletal trait. Thus, osteoclasts recycle via osteomorphs, a cell type involved in the regulation of bone resorption that may be targeted for the treatment of skeletal diseases.

Dental pulp-derived stem cells inhibit osteoclast differentiation by secreting osteoprotegerin and deactivating AKT signalling in myeloid cells.

Osteoclasts (OCs) differentiate from the monocyte/macrophage lineage, critically regulate bone resorption and remodelling in both homeostasis and pathology. Various immune and non‐immune cells help initiating activation of myeloid cells for differentiation, whereas hyper‐activation leads to pathogenesis, and mechanisms are yet to be completely understood. Herein, we show the efficacy of dental pulp–derived stem cells (DPSCs) in limiting RAW 264.7 cell differentiation and underlying molecular mechanism, which has the potential for future therapeutic application in bone‐related disorders. We found that DPSCs inhibit induced OC differentiation of RAW 264.7 cells when co‐cultured in a contact‐free system. DPSCs reduced expression of key OC markers, such as NFATc1, cathepsin K, TRAP, RANK and MMP‐9 assessed by quantitative RT‐PCR, Western blotting and immunofluorescence detection methods. Furthermore, quantitative RT‐PCR analysis revealed that DPSCs mediated M2 polarization of RAW 264.7 cells. To define molecular mechanisms, we found that osteoprotegerin (OPG), an OC inhibitory factor, was up‐regulated in RAW 264.7 cells in the presence of DPSCs. Moreover, DPSCs also constitutively secrete OPG that contributed in limiting OC differentiation. Finally, the addition of recombinant OPG inhibited OC differentiation in a dose‐dependent manner by reducing the expression of OC differentiation markers, NFATc1, cathepsin K, TRAP, RANK and MMP9 in RAW 264.7 cells. RNAKL and M‐CSF phosphorylate AKT and activate PI3K‐AKT signalling pathway during osteoclast differentiation. We further confirmed that OPG‐mediated inhibition of the downstream activation of PI3K‐AKT signalling pathway was similar to the DPSC co‐culture–mediated inhibition of OC differentiation. This study provides novel evidence of DPSC‐mediated inhibition of osteoclastogenesis mechanisms.

PMEPA1 and NEDD4 control the proton production of osteoclasts by regulating vesicular trafficking.

Osteoclast bone resorption activity is critically regulated to maintain bone homeostasis. Osteoclasts resorb bone by producing protons and acid hydrolase via lysosomal secretion, however, a detailed mechanism remains elusive. PMEPA1 is a vesicular membrane protein, which binds to the NEDD4 family member of ubiquitin ligases. We have previously reported that Pmepa1 is highly expressed in bone resorbing osteoclasts, and regulates bone resorption. Here, we investigated the mechanism of bone resorption regulated by PMEPA1. Mutant mice lacking NEDD4-binding domains of PMEPA1 displayed enhanced bone volume, and reduced bone resorption activity in comparison with those of WT mice. Analysis with pH-sensitive fluorescence probe revealed that proton secretion from osteoclasts significantly decreased in Pmepa1 mutant osteoclasts. Immunofluorescence analysis revealed that PMEPA1 was colocalized with NEDD4, V0A3, and V0D2 subunits of vacuolar ATPase, which regulate the proton production of osteoclasts. In addition, Nedd4 knockdown reduced bone resorption and proton secretion of osteoclasts. Furthermore, Pmepa1 mutation and Nedd4 knockdown altered the cytoplasmic distribution of components of V-ATPase and expression of autophagy-related proteins, suggesting that lysosomal secretion is affected. Collectively, these findings indicate that PMEPA1 controls proton secretion from osteoclasts via NEDD4 by regulating vesicular trafficking, and NEDD4 is an important regulator of bone resorption.

Anti-perimenopausal osteoporosis effects of Erzhi formula via regulation of bone resorption through osteoclast differentiation: A network pharmacology-integrated experimental study

Ethnopharmacological relevanceErzhi formula (EZF) consists of Ecliptae herba (EH) and Fructus Ligustri Lucidi (FLL) at a ratio 1:1, and constitutes a well-known formula in China that is commonly used for treating menopausal diseases. Aim of the studyIn this study, we explored the pharmacologic actions and potential molecular mechanisms underlying EZF's action in preventing and treating osteoporosis. Materials and methodsThe active components and related targets of EZF's anti-osteoporotic effects were predicted by network pharmacology, and functional enrichment analysis was also performed. We then used an osteoporosis model of ovariectomized (OVX) mice to detect the effects of EZF on osteoporosis. ResultsThe results from network pharmacology identified a total of 10 active ingredients from EH and 13 active ingredients from FLL that might affect 65 potential therapeutic targets. GO enrichment analysis revealed that EZF affected bone tissue primarily via hormone (particularly estradiol)-related pathways and bone resorption by osteoclast differentiation. KEGG analysis demonstrated that bone-related factors such as Runt-related transcription factor 2 (Runx2), Ca2, estrogen receptor1 (ESR1), androgen receptors (AR), and TNFα served as the primary targets during osteoclastic differentiation. In vivo experiments showed that the formula significantly improved the diminution in estrogen and the subsequent uterine atrophy induced by ovariectomy (P < 0.01 or 0.05), implying that the EZF exerted its actions via regulation of estradiol and the nourishing effects of the uterus in OVX mice. Dual-energy X-ray absorptiometry and micro-CT showed that EZF significantly inhibited bone loss and improved bone micro-architecture by statistically increasing the number of bone trabeculae and decreasing the separation of bone trabeculae in OVX mice (P < 0.01 or 0.05); EZF also inhibited bone loss and enhanced bone-fracture load. Furthermore, we confirmed that EZF reduced the calcium concentrations, augmented protein and mRNA levels for Runx2 in the bone marrow, and reduced PPARγ levels. RANKL—a key downstream regulatory protein of many targets that was referred to in our results of network pharmacology as being involved in the regulation of osteoclastogenesis—was significantly diminished by EZF; it also elevated OPG content. In addition, we used monocytes of bone-marrow origin to detect the effects of the potential components of EZF on osteoclast differentiation and found that wedelolactone, oleanolic acid, echinocystic acid, luteolin, and luteolin-7-o-glucoside significantly inhibited osteoclast differentiation from monocytes induced by 25 ng/mL MCSF and 50 ng/mL RANKL (P < 0.01 or 0.05). ConclusionsOur present study indicated that EZF significantly inhibited the bone loss induced by OVX in mice by its regulation of estradiol combined with the nourishing effect of the uterus, and that it also attenuated bone resorption by decreasing the RANKL/OPG ratio so as to inhibit osteoclast maturation.

USF3 modulates osteoporosis risk by targeting WNT16, RANKL, RUNX2, and two GWAS lead SNPs rs2908007 and rs4531631.

Osteoporotic fractures cause major morbidity and mortality in the aging population. Genome-wide association studies (GWAS) have identified USF3 as the novel susceptibility gene of osteoporosis. However, the functional role in bone metabolism and the target gene of the basic helix-loop-helix transcription factor USF3 are unclear. Here, we show that USF3 enhances osteoblast differentiation and suppresses osteoclastogenesis in cultured human osteoblast-like U-2OS cells. Mechanistic studies revealed that transcription factor USF3 antagonistically interacts with anti-osteogenic TWIST1/TCF12 heterodimer in the WNT16 and RUNX2 promoter, and counteracts CREB1 and JUN/FOS in the RANKL promoter. Importantly, the osteoporosis GWAS variant g.1744A>G (rs2908007A>G) located in the WNT16 promoter confers G-allele-specific transcriptional modulation by USF3, TWIST1/TCF12 and TBX5/TBX15, and USF3 transactivates the osteoclastogenesis suppressor WNT16 promoter activity and antagonizes the repression of WNT16 by TWIST1 and TCF12. The risk G allele of osteoporosis GWAS variant g.3260A>G (rs4531631A>G) in the RANKL promoter facilitates the binding of CREB1 and JUN/FOS and enhances transactivation of the osteoclastogenesis contributor RANKL that is inhibited by USF3. Our findings uncovered the functional mechanisms of osteoporosis novel GWAS-associated gene USF3 and lead single nucleotide polymorphisms rs2908007 and rs4531631 in the regulation of bone formation and resorption.

The molecular structure and function of sorting nexin 10 in skeletal disorders, cancers, and other pathological conditions.

SNX10 is a member of the phox homology domain-containing family of phosphoinositide-binding proteins. Intracellularly, SNX10 localizes to endosomes where it mediates intracellular trafficking, endosome organization, and protein localization to the centrosome and cilium. It is highly expressed in bone and the gut where it participates in bone mineral and calcium homeostasis through the regulation of osteoclastic bone resorption and gastric acid secretion, respectively. Not surprisingly, patients harboring mutations in SNX10 mutation manifest a phenotype of autosomal recessive osteopetrosis or malignant infantile osteopetrosis, which is clinically characterized by dense bones with increased cortical bone into the medullary space with bone marrow occlusion or depletion, bone marrow failure, and anemia. Accordingly, SNX10 mutant osteoclasts exhibit impaired bone resorptive capacity. Beyond the skeleton, there is emerging evidence implicating SNX10 in cancer development, metabolic disorders, inflammation, and chaperone-mediated autophagy. Understanding the structural basis through which SNX10 exerts its diverse biological functions in both cell and tissue-specific manners may therefore inform new therapeutic opportunities towardthe treatment and management of SNX10-related diseases.

Siglec-15 on Osteoclasts Is Crucial for Bone Erosion in Serum-Transfer Arthritis.

Siglec-15 is a conserved sialic acid-binding Ig-like lectin, which is expressed on osteoclasts. Deficiency of Siglec-15 leads to an impaired osteoclast development, resulting in a mild osteopetrotic phenotype. The role of Siglec-15 in arthritis is still largely unclear. To address this, we generated Siglec-15 knockout mice and analyzed them in a mouse arthritis model. We could show that Siglec-15 is directly involved in pathologic bone erosion in the K/BxN serum-transfer arthritis model. Histological analyses of joint destruction provided evidence for a significant reduction in bone erosion area and osteoclast numbers in Siglec-15-/- mice, whereas the inflammation area and cartilage destruction was comparable to wild-type mice. Thus, Siglec-15 on osteoclasts has a crucial function for bone erosion during arthritis. In addition, we generated a new monoclonal anti-Siglec-15 Ab to clarify its expression pattern on immune cells. Whereas this Ab demonstrated an almost exclusive Siglec-15 expression on murine osteoclasts and hardly any other expression on various other immune cell types, human Siglec-15 was more broadly expressed on human myeloid cells, including human osteoclasts. Taken together, our findings show a role of Siglec-15 as a regulator of pathologic bone resorption in arthritis and highlight its potential as a target for future therapies, as Siglec-15 blocking Abs are available.

The calcium channel TRPV6 is a novel regulator of RANKL-induced osteoclastic differentiation and bone absorption activity through the IGF-PI3K-AKT pathway.

ObjectivesCalcium ion signals are important for osteoclast differentiation. Transient receptor potential vanilloid 6 (TRPV6) is a regulator of bone homeostasis. However, it was unclear whether TRPV6 was involved in osteoclast formation. Therefore, the aim of this study was to evaluate the role of TPRV6 in bone metabolism and to clarify its regulatory role in osteoclasts at the cellular level.Materials and methodsBone structure and histological changes in Trpv6 knockout mice were examined using micro‐computed tomography and histological analyses. To investigate the effects of Trpv6 on osteoclast function, we silenced or overexpressed Trpv6 in osteoclasts via lentivirus transfection, respectively. Osteoclast differentiation and bone resorption viability were measured by tartrate‐resistant acid phosphatase (TRAP) staining and pit formation assays. The expression of osteoclast marker genes, including cathepsin k, DC‐STAMP, Atp6v0d2 and TRAP, was measured by qRT‐PCR. Cell immunofluorescence and Western blotting were applied to explore the mechanisms by which the IGF‐PI3K‐AKT pathway was involved in the regulation of osteoclast formation and bone resorption by Trpv6.ResultsWe found that knockout of Trpv6 induced osteoporosis and enhanced bone resorption in mice, but did not affect bone formation. Further studies showed that Trpv6, which was distributed on the cell membrane of osteoclasts, acted as a negative regulator for osteoclast differentiation and function. Mechanistically, Trpv6 suppressed osteoclastogenesis by decreasing the ratios of phosphoprotein/total protein in the IGF–PI3K–AKT signalling pathway. Blocking of the IGF–PI3K–AKT pathway significantly alleviated the inhibitory effect of Trpv6 on osteoclasts formation.ConclusionsOur study confirmed the important role of Trpv6 in bone metabolism and clarified its regulatory role in osteoclasts at the cellular level. Taken together, this study may inspire a new strategy for the treatment of osteoporosis.

RANKL as a target for the treatment of osteoporosis.

Osteoporosis is characterized by compromised bone strength, predisposing to an increased risk of fracture. Because bone is constantly remodeled, and bone mass and structure are determined by the balance between bone resorption and bone formation, it is important to maintain normal bone turnover. Therefore, therapies that reduce bone resorption have been the mainstream of osteoporosis treatment. Receptor activator of nuclear factor-kappa B ligand (RANKL)-RANK signaling was found to play a pivotal role in the regulation of osteoclastic bone resorption, and inhibition of RANKL-RANK system has become an important therapeutic target for the treatment of osteoporosis. Denosumab, a fully human monoclonal anti-RANKL neutralizing antibody, is developed as a drug for the treatment of osteoporosis. This review summarized pharmacokinetic and pharmacodynamic properties of denosumab, clinical studies including phase 2 dose-ranging and its extension study, phase 3 fracture prevention study (FREEDOM) with extension up to 10years, studies on male osteoporosis (ADAMO study), and on glucocorticoid-induced osteoporosis, along with relevant clinical studies in Japan. In addition, mechanism of denosumab action that can explain its long-term sustained effects, combination and sequential treatment as well as the problems in discontinuation of denosumab, and finally safety of denosumab therapy is discussed.

Osteocyte-Related Cytokines Regulate Osteoclast Formation and Bone Resorption.

The process of bone remodeling is the result of the regulated balance between bone cell populations, namely bone-forming osteoblasts, bone-resorbing osteoclasts, and the osteocyte, the mechanosensory cell type. Osteoclasts derived from the hematopoietic stem cell lineage are the principal cells involved in bone resorption. In osteolytic diseases such as rheumatoid arthritis, periodontitis, and osteoporosis, the balance is lost and changes in favor of bone resorption. Therefore, it is vital to elucidate the mechanisms of osteoclast formation and bone resorption. It has been reported that osteocytes express Receptor activator of nuclear factor κΒ ligand (RANKL), an essential factor for osteoclast formation. RANKL secreted by osteocytes is the most important factor for physiologically supported osteoclast formation in the developing skeleton and in pathological bone resorption such as experimental periodontal bone loss. TNF-α directly enhances RANKL expression in osteocytes and promotes osteoclast formation. Moreover, TNF-α enhances sclerostin expression in osteocytes, which also increases osteoclast formation. These findings suggest that osteocyte-related cytokines act directly to enhance osteoclast formation and bone resorption. In this review, we outline the most recent knowledge concerning bone resorption-related cytokines and discuss the osteocyte as the master regulator of bone resorption and effector in osteoclast formation.

Antiresorptive activity of osteoprotegerin requires an intact heparan sulfate-binding site

Osteoprotegerin (OPG), a secreted decoy receptor for receptor activator of nuclear factor B ligand (RANKL), plays an essential role in regulating bone resorption. While much is known about the function of the N-terminal domains of OPG, which is responsible for binding to RANKL, the exact biological functions of the three C-terminal domains of OPG remain uncertain. We have previously shown that one likely function of the C-terminal domains of OPG is to bind cell surface heparan sulfate (HS), but the in vivo evidence was lacking. To investigate the biological significance of OPG-HS interaction in bone remodeling, we created OPG knock-in mice (opgAAA ). The mutated OPG is incapable of binding to HS but binds RANKL normally. Surprisingly, opgAAA/AAA mice displayed a severe osteoporotic phenotype that is very similar to opg-null mice, suggesting that the antiresorption activity of OPG requires HS. Mechanistically, we propose that the HS immobilizes secreted OPG at the surface of osteoblasts lineage cells, which facilitates binding of OPG to membrane-anchored RANKL. To further support this model, we altered the structure of osteoblast HS genetically to make it incapable of binding to OPG. Interestingly, osteocalcin-Cre;Hs2stf/f mice also displayed osteoporotic phenotype with similar severity to opgAAA/AAA mice. Combined, our data provide strong genetic evidence that OPG-HS interaction is indispensable for normal bone homeostasis.

MiR-1-3p regulates the differentiation of mesenchymal stem cells to prevent osteoporosis by targeting secreted frizzled-related protein 1.

Osteoporosis (OP) is a systemic skeletal disorder with the characteristics of bone mass reduction and microarchitecture deterioration, resulting in bone fragility and increased fracture risk. A reduction in the osteoblast-differentiation of bone marrow mesenchymal stem cells (BMSCs) is considered as a basic pathogenesis of osteoporosis. miRNAs play a substantial role in the development and differentiation of BMSCs. In the present study, we found that miR-1-3p was significantly downregulated in the bones of Chinese osteoporotic patients (n=29). Secreted frizzled-related protein 1 (SFRP1) was predicted as a target gene of miR-1-3p via the TargetScan and PicTar softwares and validated by dual-luciferase reporter assays. The findings revealed that the expression of SFRP1 was inversely correlated with miR-1-3p in osteoporotic patients. We induced mouse MSCs (mMSCs) to osteogenesis or adipogenesis and found that miR-1-3p was upregulated during osteogenesis but downregulated during adipogenesis. The overexpression of miR-1-3p stimulated osteogenesis and inhibited adipogenesis of mMSCs. In addition, ovariectomized (OVX) mice were tested and the function of miR-1-3p in vivo was explored. Immunohistochemistry and histomorphometric assays showed that in vivo inhibition of miR-1-3p increased the expression level of SFRP1 and reduced bone formation and bone mass. Furthermore, tartrate-resistant acid phosphatase (TRAP) staining indicated that the in vivo suppression of miR-1-3p promoted osteoclast activity, suggesting that miR-1-3p may influence bone mass by regulating bone resorption. It can be concluded that miR-1-3p plays a pivotal role in the pathogenesis of osteoporosis via targeting SFRP1 and may be a potential therapeutic target for osteoporosis.

The Zinc-Finger Domain Containing Protein ZC4H2 Interacts with TRPV4, Enhancing Channel Activity and Turnover at the Plasma Membrane.

The Ca2+-permeable Transient Receptor Potential channel vanilloid subfamily member 4 (TRPV4) is involved in a broad range of physiological processes, including the regulation of systemic osmotic pressure, bone resorption, vascular tone, and bladder function. Mutations in the TRPV4 gene are the cause of a spectrum of inherited diseases (or TRPV4-pathies), which include skeletal dysplasias, arthropathies, and neuropathies. There is little understanding of the pathophysiological mechanisms underlying these variable disease phenotypes, but it has been hypothesized that disease-causing mutations affect interaction with regulatory proteins. Here, we performed a mammalian protein–protein interaction trap (MAPPIT) screen to identify proteins that interact with the cytosolic N terminus of human TRPV4, a region containing the majority of disease-causing mutations. We discovered the zinc-finger domain-containing protein ZC4H2 as a TRPV4-interacting protein. In heterologous expression experiments, we found that ZC4H2 increases both the basal activity of human TRPV4 as well as Ca2+ responses evoked by ligands or hypotonic cell swelling. Using total internal reflection fluorescence (TIRF) microscopy, we further showed that ZC4H2 accelerates TRPV4 turnover at the plasma membrane. Overall, these data demonstrate that ZC4H2 is a positive modulator of TRPV4, and suggest a link between TRPV4 and ZC4H2-associated rare disorders, which have several neuromuscular symptoms in common with TRPV4-pathies.

Regulation of Bone Resorption by the Actin-Bundling Protein L-Plastin in Osteoclasts

Osteoporosis is related to estrogen deficiency and aging. Bone loss can also occur as a result of inflammation-associated diseases such as rheumatoid arthritis and periodontitis, which share several pathologic features with osteoporosis. Estrogen deficiency is associated with increased osteoclast activation, decreased osteoblast function, and increased inflammatory bone-resorbing cytokines (e.g., interleukin-1, -6, and tumor necrosis factor –α). The differentiation of osteoclasts is regulated by the cytokines macrophage colony-stimulating factor, RANK ligand, and osteoprotegerin secreted by osteoblasts. Bone resorption is the unique function of osteoclasts. Podosomes are essential features of osteoclast migration. Podosomes are F-actin rich structures joined radially by actin fibers called F-actin cloud. Upon attachment to the bone surface, osteoclasts reorganize their cytoskeleton to form sealing zones. Sealing zone formation is required for efficient bone resorption to occur by osteoclasts. Integrin αvβ3 and TNF-alpha mediated signaling mechanisms regulate the assembly/disassembly of podosomes during migration and the organization of sealing zones during bone resorption. A brief description is provided on these aspects in this review.

The opposite effects of Antarctic krill oil and arachidonic acid-rich oil on bone resorption in ovariectomized mice.

Osteoporosis, a chronic disease that affects over 200 million people worldwide, presents a substantial medical and socioeconomic burden on the modern society. However, long-term intake of diets supplemented with different polyunsaturated fatty acids (PUFAs) can affect bone metabolism; thus, this study investigated the comparative effects of Antarctic krill oil (AKO, containing n-3 PUFAs) and arachidonic acid-rich oil (AAO, containing n-6 PUFAs) on bone resorption in a mice model of postmenopausal osteoporosis. Mice were orally administered with AKO (200 mg kg-1) or AAO (220 mg kg-1) once daily for 30 days, ovariectomized, followed by the continued administration of the respective samples for 90 days. Biomechanical and histomorphometric analyses revealed that AKO increased the bone mineral density (BMD) to enhance the biomechanical properties by increasing the mineral apposition rate and repairing the microstructure of the trabecular bone, whereas AAO had the opposite effect. The fatty acid analysis of the vertebra showed that AKO increased the n-3 PUFA (especially for DHA) content, thereby decreasing the ratio of n-6/n-3 PUFAs, which was negatively correlated with the BMD. However, AAO had the opposite effect due to high amounts of arachidonic acid. To explore the underlying mechanism responsible for these observations, we compared the classical bone resorption OPG/RANKL/NF-κB pathway mediated by PGE2/EP4. The ratio of n-6/n-3 PUFAs in the bone affected the production of PGE2, a factor regulating the OPG/RANKL pathway, thereby regulating osteoclastogenesis by stimulating the NF-κB pathway. The results of ELISA, qRT-PCR, and western blot demonstrated that AKO reduced the secretion of PGE2 and the expression of EP4, upregulating the ratio of OPG/RANKL in the bone, thereby decreasing TRAF6 expression to inhibit the activation of the NF-κB signaling pathway, and finally inhibiting the expression of nuclear transcription factors (c-fos and NFATc1) to prevent excessive osteoclastogenesis (TRACP, MMP-9, and Cath-K). Arachidonic acid is a precursor of PGE2 synthesis. AAO showed the opposite trend through the same pathway. Thus, AKO could significantly improve osteoporosis via the OPG/RANKL/NF-κB pathway mediated by PGE2/EP4 to inhibit osteoclastogenesis, whereas AAO aggravated osteoporosis via the same pathway. This is the first study to systematically compare the effects and mechanism of AKO and AAO in regulating bone resorption in osteoporotic mice to support recommendations on fatty acid types in dietary oils for an osteoporotic population.

Evidence for Gender-Specific Bone Loss Mechanisms in Periprosthetic Osteolysis.

Osteolysis adjacent to total hip replacement (THR) prostheses is a major cause of their eventual failure. Periprosthetic osteolysis is associated with the production of bioactive particles, produced by the wear of articulating prosthesis surfaces. Wear particles invade the periprosthetic tissue, inducing inflammation and bone resorption. Previous studies have shown that osteocytes, the most numerous cell type in mineralised bone, can respond to wear particles of multiple orthopaedic material types. Osteocytes play important roles in bone resorption, regulating bone resorption by osteoclasts and directly through osteocytic osteolysis, also known as perilacunar remodelling. In this study, we perform a histological analysis of bone biopsies obtained from cohorts of male and female patients undergoing either primary THR surgery or revision THR surgery for aseptic loosening. The osteocyte lacunae area (Ot.Lac.Ar) and percentage lacunar area/bone area (%Ot.Lac.Ar/B.Ar) were significantly larger overall in revision THR bone than bone from similar sites in primary THR. Analysis by patient gender showed that increased Ot.Lac.Ar, indicative of increased perilacunar remodelling, was restricted to female revision samples. No significant differences in osteoclast parameters were detectable between the cohorts. These findings suggest previously unrecognised gender-specific mechanisms of bone loss in orthopaedic wear particle-induced osteolysis in humans.