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Bone turnover markers (BTMs) are commonly used in osteoporosis treatment as indicators of cell activities of bone-resorbing osteoclasts and bone-forming osteoblasts. The wide variability in their values due to multiple factors, such as aging and diseases, makes it difficult for physicians to utilize them for clinical decision-making. The progenitors of osteoclasts and osteoblasts are indispensable for a comprehensive interpretation of the variability in BTM values because these upstream progenitors strongly regulate the downstream cell activities of bone turnover. However, understanding the complex interactions among the multiple populations of bone cells is challenging. In this study, we aimed to gain a fundamental understanding of the mechanism by which the progenitor dynamics affect the variability in bone turnover through in silico experiments by exploring the cell dynamics with aging effects on osteoporosis. Negative feedback control driven by the consumptive loss of progenitors prevents rapid bone loss due to excessive bone turnover, and through feedback regulation, aging effects on osteoclast differentiation and osteoclast progenitor proliferation cause variability in the osteoclast and osteoblast activity balance and its temporal transition. By expressing the variability in the bone turnover status, our model describes the individualities of patients based on their clinical backgrounds. Therefore, our model could play a powerful role in assisting tailored treatment and has the potential to resolve the various health problems associated with osteoporosis worldwide. [ABSTRACT FROM AUTHOR]

Extracellular matrix metalloproteinase inducer (EMMPRIN) plays critical roles in the regulation of inflammation and bone metabolism. The roles of EMMPRIN signaling in osteoclasts are worthy of deep study. The present study aimed to investigate bone resorption in periodontitis through the intervention of EMMPRIN signaling. The distribution of EMMPRIN in human periodontitis was observed. RANKL-induced osteoclast differentiation of mouse bone marrow-derived macrophages (BMMs) were treated with EMMPRIN inhibitor in vitro. Rats with ligation-induced periodontitis were treated with EMMPRIN inhibitor and harvested for microcomputed tomography scanning, histologic observation, immunohistochemistry, and double immunofluorescence analysis. Positive expressions of EMMPRIN could be found in the CD68+-infiltrating cells. Downregulated EMMPRIN restrained osteoclast differentiation of BMMs in vitro, which also inhibited MMP-9 expression (*P < 0.05). In vivo, EMMPRIN inhibitor restrained ligation-induced bone resorption by decreasing tartrate-resistant acid phosphatase-positive osteoclasts. Both EMMPRIN-positive and MMP-9-positive osteoclasts were less common in the EMMPRIN inhibitor groups than in the control groups. Intervention of EMMPRIN signaling in osteoclasts could probably provide a potential therapeutic target for attenuating ligation-induced bone resorption. [ABSTRACT FROM AUTHOR]

Numerous pieces of evidence have supported the therapeutic potential of photobiomodulation (PBM) to modulate bone remodeling on mechanically stimulated teeth, proving PBM's ability to be used as a coadjuvant treatment to accelerate orthodontic tooth movement (OTM). However, there are still uncertainty and discourse around the optimal PBM protocols, which hampers its optimal and consolidated clinical applicability. Given the differential expression and metabolic patterns exhibited in the tension and compression sides of orthodontically stressed teeth, it is plausible that different types of irradiation may be applied to each side of the teeth. In this sense, this study aimed to design and implement an optimization protocol to find the most appropriate PBM parameters to stimulate specific bone turnover processes. To this end, three levels of wavelength (655, 810 and 940 nm), two power densities (5 and 10 mW/cm2) and two regimens of single and multiple sessions within three consecutive days were tested. The biological response of osteoblasts and periodontal ligament (PDL) fibroblasts was addressed by monitoring the PBM's impact on the cellular metabolic activity, as well as on key bone remodeling mediators, including alkaline phosphatase (ALP), osteoprotegerin (OPG) and receptor activator of nuclear factor κ-B ligand (RANK-L), each day. The results suggest that daily irradiation of 655 nm delivered at 10 mW/cm2, as well as 810 and 940 nm light at 5 mW/cm2, lead to an increase in ALP and OPG, potentiating bone formation. In addition, irradiation of 810 nm at 5 mW/cm2 delivered for two consecutive days and suspended by the third day promotes a downregulation of OPG expression and a slight non-significant increase in RANK-L expression, being suitable to stimulate bone resorption. Future studies in animal models may clarify the impact of PBM on bone formation and resorption mediators for longer periods and address the possibility of testing different stimulation periodicities. The present in vitro study offers valuable insights into the effectiveness of specific PBM protocols to promote osteogenic and osteoclastogenesis responses and therefore its potential to stimulate bone formation on the tension side and bone resorption on the compression side of orthodontically stressed teeth. [ABSTRACT FROM AUTHOR]

Considering the conflicting evidence regarding the potential long-term detrimental effect of swimming during growth on femur quality and fracture risk, our aim was to investigate the effect of eight months of swimming on femur quality. Twenty male eight-week-old Wistar rats were assigned into a swimming (SW; n = 10; 2 h/day, 5 days/week) or active control group (CG; n = 10, housed with running wheel) for eight months. Plasma osteocalcin and C-terminal telopeptide of type I collagen concentrations (ELISA) were assessed at baseline, four, and eight months of protocol. Femur structure (micro-computed tomography), biomechanical properties (three-point bending), and cellular density (histology) were determined after the protocol. SW displayed a lower uncoupling index, suggesting higher bone resorption, lower empty lacunae density, cortical and trabecular femur mass, femur length and cortical thickness, and higher cortical porosity than CG (p < 0.05). Although both biomarkers' concentrations decreased in both groups throughout the experiment (p < 0.001), there were no significant differences between groups (p > 0.05). No differences were also found regarding biomechanical properties, bone marrow adiposity, and osteocyte and osteoclast densities (p > 0.05). Long-term swimming was associated with unbalanced bone turnover and compromised femur growth, lower femur mass, and deteriorated cortical bone microarchitecture. However, femur trabecular microarchitecture and biomechanical properties were not affected by swimming. [ABSTRACT FROM AUTHOR]

Bcl2l1 (Bcl-XL) belongs to the Bcl-2 family, Bcl2 and Bcl2-XL are major anti-apoptotic proteins, and the apoptosis of osteoblasts is a key event for bone homeostasis. As the functions of Bcl2l1 in osteoblasts and bone homeostasis remain unclear, we generated osteoblast-specific Bcl2l1-deficient (Bcl2l1fl/flCre) mice using 2.3-kb Col1a1 Cre. Trabecular bone volume and the trabecular number were lower in Bcl2l1fl/flCre mice of both sexes than in Bcl2l1fl/fl mice. In bone histomorphometric analysis, osteoclast parameters were increased in Bcl2l1fl/flCre mice, whereas osteoblast parameters and the bone formation rate were similar to those in Bcl2l1fl/fl mice. TUNEL-positive osteoblastic cells and serum TRAP5b levels were increased in Bcl2l1fl/flCre mice. The deletion of Bcl2l1 in osteoblasts induced Tnfsf11 expression, whereas the overexpression of Bcl-XL had no effect. In a co-culture of Bcl2l1-deficient primary osteoblasts and wild-type bone-marrow-derived monocyte/macrophage lineage cells, the numbers of multinucleated TRAP-positive cells and resorption pits increased. Furthermore, serum deprivation or the deletion of Bcl2l1 in primary osteoblasts increased apoptosis and ATP levels in the medium. Therefore, the reduction in trabecular bone in Bcl2l1fl/flCre mice may be due to enhanced bone resorption through osteoblast apoptosis and the release of ATP from apoptotic osteoblasts, and Bcl2l1 may inhibit bone resorption by preventing osteoblast apoptosis. [ABSTRACT FROM AUTHOR]

Classically, particle-induced periprosthetic osteolysis at the implant-bone interface has explained the aseptic loosening of joint replacement. This response is preceded by triggering both the innate and acquired immune response with subsequent activation of osteoclasts, the bone-resorbing cells. Although particle-induced periprosthetic osteolysis has been considered a foreign body chronic inflammation mediated by myelomonocytic-derived cells, current reports describe wide heterogeneous inflammatory cells infiltrating the periprosthetic tissues. This review aims to discuss the role of those non-myelomonocytic cells in periprosthetic tissues exposed to wear particles by showing original data. Specifically, we discuss the role of T cells (CD3+, CD4+, and CD8+) and B cells (CD20+) coexisting with CD68+/TRAP- multinucleated giant cells associated with both polyethylene and metallic particles infiltrating retrieved periprosthetic membranes. This review contributes valuable insight to support the complex cell and molecular mechanisms behind the aseptic loosening theories of orthopedic implants. [ABSTRACT FROM AUTHOR]

Osteoclasts are the primary target for osteoporosis drug development. Recent animal studies revealed the crucial roles of osteoblasts in regulating osteoclastogenesis and the longer lifespans of osteoclasts than previously thought with fission and recycling. However, existing culture platforms are limited to replicating these newly identified cellular processes. We report a demineralized bone paper (DBP)-based osteoblast culture and osteoclast assay platform that replicates osteoclast fusion, fission, resorption, and apoptosis with high fidelity and analytical power. An osteoid-inspired DBP supports rapid and structural mineral deposition by osteoblasts. Coculture osteoblasts and bone marrow monocytes under biochemical stimulation recapitulate osteoclast differentiation and function. The DBP-based bone model allows longitudinal quantitative fluorescent monitoring of osteoclast responses to bisphosphonate drug, substantiating significantly reducing their number and lifespan. Finally, we demonstrate the feasibility of humanizing the bone model. The DBP-based osteo assay platforms are expected to advance bone remodeling-targeting drug development with improved prediction of clinical outcomes. Here, authors report demineralized bone paper-based in vitro osteogenic culture and assay platforms that replicate essential bone tissue complexity, osteoclast processes, and drug responses with high fidelity and predictive power. [ABSTRACT FROM AUTHOR]

Objectives: Colony‐stimulating factor‐1 receptor (CSF1R) is vital for the recruitment of monocytes, and their proliferation and differentiation into functional osteoclasts. Mouse studies, where CSF1R and its cognate ligand are absent, have significant craniofacial phenotypes, but these have not been studied in detail. Materials and Methods: Pregnant CD1 mice were fed diets laced with CSF1R inhibitor—PLX5622 starting at embryonic day 3.5 (E3.5) up to birth. Pups were collected at E18.5 to study CSF1R expression using immunofluorescence. Additional pups were studied at postnatal day 21 (P21) and P28 using microcomputed tomography (μCT) and Geometric Morphometrics, to evaluate craniofacial form. Results: CSF1R‐positive cells were present throughout the developing craniofacial region, including the jaw bones, surrounding teeth, tongue, nasal cavities, brain, cranial vault and base regions. Animals exposed to the CSF1R inhibitor in utero had severe depletion of CSF1R‐positive cells at E18.5 and had significant differences in craniofacial form (size and shape) at postnatal timepoints. Centroid sizes for the mandibular and cranio‐maxillary regions were significantly smaller in CSF1R‐inhibited animals. Proportionally, these animals had a domed skull, with taller and wider cranial vaults and shortening of their midfacial regions. Mandibles were smaller vertically and anterio‐posteriorly, with proportionally wider inter‐condylar distances. Conclusions: Embryonic inhibition of CSF1R impacts postnatal craniofacial morphogenesis, with significant influences on the mandibular and cranioskeletal size and shape. These data indicate that CSF1R plays a role in early cranio‐skeletal patterning, likely through osteoclast depletion. [ABSTRACT FROM AUTHOR]

Critical-size defects (CSDs) are challenging oral clinical issues that need to be solved. Adipose-derived mesenchymal stem cells (ADSCs) and gene therapy offer a new target to solve these issues. Consequently, ADSCs attract more and more attention because of advantages such as easy obtainability and no ethical concerns. TNF receptor-associated factor 6 (TRAF6) is a significant binding protein both of tumour necrosis factor superfamily and of the toll/interleukin-1 receptor superfamily. Evidence is accumulating that TRAF6 inhibited osteoclast formation and promoted the proliferation of multiple myeloma cell lines and bone resorption. Here, we reported that overexpression of TRAF6 enhanced the proliferation, migration and osteogenesis of ADSCs through Raf-Erk-Merk-Hif1a pathway. Cell sheet of ADSCs combined with TRAF6 accelerated the healing of CSDs. In a word, TRAF6 enhanced osteogenesis, migration and proliferation through Raf-Erk-Merk-Hif1a pathway. [ABSTRACT FROM AUTHOR]

Previous researches have shown that patients with schizophrenia who were using antipsychotics in their treatment developed changes in Bone Mineral Density and body composition, leading to osteoporosis and modifications in weight, skeletal muscle mass index and percent of fat tissue. Results of many studies suggested that the use of antipsychotic causes hyperprolactinemia and consequently lower bone mineral density values were observed. Further, antipsychotics increase food intake and have possible effects on metabolism, causing changes in weight, glucose level and lipid status, all of which can be a risk for developing metabolic syndrome and result in changing of bone mineral density. Antipsychotics change cytokine profiles in patients with schizophrenia and on the other hand the influence of T cells, B cells and inflammatory cytokines on osteoclasts and on osteoblasts was also established. In assessing the effects of antipsychotic on bone metabolism very important is to consider the duration of the treatment and clinical course of the disease, but undeniable effect is careless life style and inadequate physical activity that patients with schizophrenia have. Our attempt is to give an overview of the newest findings in this field, regarding the direct effects of antipsychotics on the bone metabolism, but also through prolactine elevation, metabolic and immune changes. Better understanding of the underlying mechanisms of schizophrenia and changes in bone mineral density could improve our clinical practice: affect to choice of the individually most appropriate antipsychotic, point to the need to monitor possible immunmetabolic changes during the treatment and improvement of the life quality of this vulnerable population. [ABSTRACT FROM AUTHOR]

Objective: IĸB protein B cell lymphoma 3‐encoded protein (BCL3) is a regulator of the NF‐κB family of transcription factors. NF‐κB signaling fundamentally influences the fate of bone‐forming osteoblasts and bone‐resorbing osteoclasts, but the role of BCL3 in bone biology has not been investigated. The objective of this study was to evaluate BCL3 in skeletal development, maintenance, and osteoarthritic pathology. Methods: To assess the contribution of BCL3 to skeletal homeostasis, neonatal mice (n = 6–14) lacking BCL3 (Bcl3−/−) and wild‐type (WT) controls were characterized for bone phenotype and density. To reveal the contribution to bone phenotype by the osteoblast compartment in Bcl3−/− mice, transcriptomic analysis of early osteogenic differentiation and cellular function (n = 3–7) were assessed. Osteoclast differentiation and function in Bcl3−/− mice (n = 3–5) was assessed. Adult 20‐week Bcl3−/− and WT mice bone phenotype, strength, and turnover were assessed. A destabilization of the medial meniscus model of osteoarthritic osteophytogenesis was used to understand adult bone formation in Bcl3−/− mice (n = 11–13). Results: Evaluation of Bcl3−/− mice revealed congenitally increased bone density, long bone dwarfism, increased bone biomechanical strength, and altered bone turnover. Molecular and cellular characterization of mesenchymal precursors showed that Bcl3−/− cells displayed an accelerated osteogenic transcriptional profile that led to enhanced differentiation into osteoblasts with increased functional activity, which could be reversed with a mimetic peptide. In a model of osteoarthritis‐induced osteophytogenesis, Bcl3−/− mice exhibited decreased pathological osteophyte formation (P < 0.05). Conclusion: Cumulatively, these findings demonstrate that BCL3 controls developmental mineralization to enable appropriate bone formation, whereas in a pathological setting, it contributes to skeletal pathology. [ABSTRACT FROM AUTHOR]

Structural variations, including copy number variations (CNVs), affect around 20 million bases in the human genome and are common causes of rare conditions. CNVs are rarely investigated in complex disease research because most CNVs are not targeted on the genotyping arrays or the reference panels for genetic imputation. In this study, we characterize CNVs in a Swedish cohort (N = 1,021) using short-read whole-genome sequencing (WGS) and use long-read WGS for validation in a subcohort (N = 15), and explore their effect on 438 plasma proteins. We detected 184,182 polymorphic CNVs and identified 15 CNVs to be associated with 16 proteins (P < 8.22×10−10). Of these, 5 CNVs could be perfectly validated using long-read sequencing, including a CNV which was associated with measurements of the osteoclast-associated immunoglobulin-like receptor (OSCAR) and located upstream of OSCAR, a gene important for bone health. Two other CNVs were identified to be clusters of many short repetitive elements and another represented a complex rearrangement including an inversion. Our findings provide insights into the structure of common CNVs and their effects on the plasma proteome, and highlights the importance of investigating common CNVs, also in relation to complex diseases. [ABSTRACT FROM AUTHOR]

Objectives The variable region of most ACPA IgG molecules in the serum of RA patients carries N -glycan (N -glycanV). To analyse the pathogenicity of N -glycanV of ACPAs, we analysed the pathogenicity of a monoclonal ACPA, CCP-Ab1, with or without N -glycanV, which had been isolated from a patient with RA. Methods CCP-Ab1 with no N -glycosylation site in the variable region (CCP-Ab1 N-rev) was generated, and antigen binding, the effect on in vitro differentiation of osteoclasts from bone marrow mononuclear cells of autoimmune arthritis–prone SKG mice (the cell size of TRAP+ cells and bone resorption capacity) and the in vivo effect on the onset or exacerbation of autoimmune arthritis in SKG mice were evaluated in comparison with glycosylated CCP-Ab1. Results Amino acid residues in citrullinated peptide (cfc1), which are essential for binding to CCP-Ab1 N-rev and original CCP-Ab1, were almost identical. The size of TRAP+ cells was significantly larger and osteoclast bone resorption capacity was enhanced in the presence of CCP-Ab1, but not with CCP-Ab1 N-rev. This enhancing activity required the sialic acid of the N -glycan and Fc region of CCP-Ab1. CCP-Ab1, but not CCP-Ab1 N-rev, induced the exacerbation of experimental arthritis in the SKG mouse model. Conclusions These data showed that N -glycanV was required for promoting osteoclast differentiation and bone resorption activity in both in vitro and in vivo assays. The present study demonstrated the important role of N -glycanV in the exacerbation of experimental arthritis by ACPAs. [ABSTRACT FROM AUTHOR]

Simple Summary: Multiple myeloma (MM) accounts for about 10% of all hematologic malignancies and is usually accompanied by severe bone disease. We analyzed the circulating levels of sRANKL, periostin, and osteopontin as osteoclast activators in newly diagnosed MM (NDMM) patients at diagnosis and in the course of treatment. Given the significantly higher levels of tested proteins at diagnosis, their correlations with the stage and grade of the disease, as well as their gradual decrease in the course of treatment, we suggest the applicability of these proteins as potential biomarkers for myeloma related bone disease (MBD) monitoring. The hallmark of multiple myeloma is myeloma related bone disease. Interactions between myeloma plasma cells (MPCs), stromal cells, and the bone marrow (BM) microenvironment play a critical role in the pathogenesis of MBD. Bone remodeling is severely dysregulated with the prevalence of osteoclast activity. We aimed to assess circulating levels of sRANKL, periostin, and osteopontin as osteoclast activators in NDMM patients at diagnosis and in the course of treatment, correlations with clinical and laboratory data, and to evaluate their potential as additional biomarkers for the assessment of MBD. The current study involved 74 subjects (41 NDMM patients, 33 controls). MBD was assessed by whole-body low-dose computed tomography. sRANKL, periostin, and osteopontin were assayed by commercial ELISA kits. At diagnosis, all tested parameters were significantly higher in NDMM patients compared to the controls (p < 0.0001), correlating with disease stage, MBD grade, and BM infiltration by MPCs. During therapy, the serum levels of all tested proteins decrease, most prominently after autologous stem cell transplantation (p < 0.0001). A significant reduction was established in patients achieving complete and very-good partial response compared to all others (p < 0.05). In conclusion, sRANKL, periostin, and osteopontin reflect MBD severity and could be promising markers for MBD monitoring and the effect of myeloma treatment. [ABSTRACT FROM AUTHOR]

Background: Excess reactive oxygen species generated by NADPH oxidase 2 (Nox2) in response to ethanol exposure mediate aspects of skeletal toxicity including increased osteoclast differentiation and activity. Because perturbation of chondrocyte differentiation in the growth plate by ethanol could be prevented by dietary antioxidants, we hypothesized that Nox2 in the growth plate was involved in ethanol‐associated reductions in longitudinal bone growth. Methods: Nox2 conditional knockout mice were generated, where the essential catalytic subunit of Nox2, cytochrome B‐245 beta chain (Cybb), is deleted in chondrocytes using a Cre‐Lox model with Cre expressed from the collagen 2a1 promoter (Col2a1‐Cre). Wild‐type and Cre‐Lox mice were fed an ethanol Lieber‐DeCarli‐based diet or pair‐fed a control diet for 8 weeks. Results: Ethanol treatment significantly reduced the number of proliferating chondrocytes in the growth plate, enhanced bone marrow adiposity, shortened femurs, reduced body length, reduced cortical bone volume, and decreased mRNA levels of a number of osteoblast and chondrocyte genes. Conditional knockout of Nox2 enzymatic activity in chondrocytes did not consistently prevent any ethanol effects. Rather, knockout mice had fewer proliferating chondrocytes than wild‐type mice in both the ethanol‐ and control‐fed animals. Additional analysis of tibia samples from Nox4 knockout mice showed that loss of Nox4 activity also reduced the number of proliferating chondrocytes and altered chondrocyte size in the growth plate. Conclusions: Although Nox enzymatic activity regulates growth plate development, ethanol‐associated disruption of the growth plate morphology is independent of ethanol‐mediated increases in Nox2 activity. [ABSTRACT FROM AUTHOR]

Aim: Our previous study revealed that the C‐C motif chemokine receptor 2 (CCR2) is a promising target for periodontitis prevention and treatment. However, CCR2 is a receptor with multiple C‐C motif chemokine ligands (CCLs), including CCL2, CCL7, CCL8, CCL13 and CCL16, and which of these ligands plays a key role in periodontitis remains unclear. The aim of the present study was to explore the key functional ligand of CCR2 in periodontitis and to evaluate the potential of the functional ligand as a therapeutic target for periodontitis. Materials and Methods: The expression levels and clinical relevance of CCR2, CCL2, CCL7, CCL8, CCL13 and CCL16 were studied using human samples. The role of CCL2 in periodontitis was evaluated by using CCL2 knockout mice and overexpressing CCL2 in the periodontium. The effect of local administration of bindarit in periodontitis was evaluated by preventive and therapeutic medication in a mouse periodontitis model. Microcomputed tomography, haematoxylin and eosin staining, tartrate‐resistant acid phosphatase staining, real‐time quantitative polymerase chain reaction, enzyme‐linked immunosorbent assay, bead‐based immunoassays and flow cytometry were used for histomorphology, molecular biology and cytology analysis. Results: Among different ligands of CCR2, only CCL2 was significantly up‐regulated in periodontitis gingival tissues and was positively correlated with the severity of periodontitis. Mice lacking CCL2 showed milder inflammation and less bone resorption than wild‐type mice, which was accompanied by a reduction in monocyte/macrophage recruitment. Adeno‐associated virus‐2 vectors overexpressing CCL2 in Ccl2−/− mice gingiva reversed the attenuation of periodontitis in a CCR2‐dependent manner. In ligation‐induced experimental periodontitis, preventive or therapeutic administration of bindarit, a CCL2 synthesis inhibitor, significantly inhibited the production of CCL2, decreased the osteoclast number and bone loss and reduced the expression levels of proinflammatory cytokines TNF‐α, IL‐6 and IL‐1β. Conclusions: CCL2 is a pivotal chemokine that binds to CCR2 during the progression of periodontitis, and targeting CCL2 may be a feasible option for controlling periodontitis. [ABSTRACT FROM AUTHOR]

Aim: To determine the effects of RVX‐208, a selective bromodomain and extra‐terminal domain (BET) inhibitor targeting bromodomain 2 (BD2), on periodontal inflammation and bone loss. Materials and Methods: Macrophage‐like cells (RAW264.7) and human gingival epithelial cells were challenged by Porphyromonas gingivalis (Pg) with or without RVX‐208. Inflammatory gene expression and cytokine production were measured by reverse transcription polymerase chain reaction and enzyme‐linked immunosorbent assay, respectively. RAW264.7 cells were induced to osteoclast differentiation. After RVX‐208 treatment, osteoclast differentiation was evaluated by histology, tartrate‐resistant‐acid‐phosphatase (TRAP) activity and the expression of osteoclast‐specific genes. The effect of RVX‐208 on osteoclast transcriptome was studied by RNA sequencing. Periodontitis was induced in rats by ligature and local RVX‐208 treatment was administered every other day. Alveolar bone loss was measured by micro‐computed tomography. Results: RVX‐208 inhibited inflammatory gene expression and cytokine production in Pg‐infected cells. Osteoclast differentiation was inhibited by RVX‐208, as evidenced by reduced osteoclast number, TRAP activity and osteoclast‐specific gene expression. RVX‐208 displayed a more selective and less profound suppressive impact on transcriptome compared with pan‐BET inhibitor, JQ1. RVX‐208 administration prevented the alveolar bone loss in vivo. Conclusions: RVX‐208 regulated both upstream (inflammatory cytokine production) and downstream (osteoclast differentiation) events that lead to periodontal tissue destruction, suggesting that it may be a promising 'epi‐drug' for the prevention of periodontitis. [ABSTRACT FROM AUTHOR]

Osteoclasts are specialized cells responsible for bone resorption, a highly energy-demanding process. Focus on osteoclast metabolism could be a key for the treatment of osteolytic diseases including osteoporosis. In this context, AMP-activated protein kinase α1 (AMPKα1), an energy sensor highly expressed in osteoclasts, participates in the metabolic reconfiguration during osteoclast differentiation and activation. This study aimed to elucidate the role of AMPKα1 during osteoclastogenesis in vitro and its impact in bone loss in vivo. Using LysMcre/0AMPKα1f/f animals and LysMcre/0 as control, we evaluated how AMPKα1 interferes with osteoclastogenesis and bone resorption activity in vitro. We found that AMPKα1 is highly expressed in the early stages of osteoclastogenesis. Genetic deletion of AMPKα1 leads to increased gene expression of osteoclast differentiation and fusion markers. In addition, LysMcre/0AMPKα1f/f mice had an increased number and size of differentiated osteoclast. Accordingly, AMPKα1 negatively regulates bone resorption in vitro, as evidenced by the area of bone resorption in LysMcre/0AMPKα1f/f osteoclasts. Our data further demonstrated that AMPKα1 regulates mitochondrial fusion and fission markers upregulating Mfn2 and downre-gulating DRP1 (dynamics-related protein 1) and that Ctskcre/0AMPKα1f/f osteoclasts lead to an increase in the number of mitochondria in AMPKα1-deficient osteoclast. In our in vivo study, femurs from Ctskcre/0AMPKα1f/f animals exhibited bone loss associated with the increased number of osteoclasts, and there was no difference between Sham and ovariectomized group. Our data suggest that AMPKα1 acts as a negative regulator of osteoclastogenesis, and the depletion of AMPKα1 in osteoclast leads to a bone loss state similar to that observed after ovariectomy. [ABSTRACT FROM AUTHOR]

Taking advantage of the well-known Komarova's type model, it is proposed here to analyze again the bone dynamics process within the context of a new parameter introduced in order to act only on the production/removal activities of osteoblasts and osteoclasts (BMU) while saving the net effectiveness of osteoclast-or-osteoblast-derived autocrine or paracrine factors. The effects of this new parameter upon simulation of the bone remodeling cycle as well as stable, intrinsically regulated oscillatory changes in bone cell numbers and bone mass are analyzed and from which unstable oscillations, similar to pathologically accelerated bone remodeling of Paget's disease appear. One can say that the introduced d parameter, with 0 ≤ d < 1 , can be viewed as a new parameter driven by Pathological conditions. On the other hand, the parameter d can probably be linked to the complex mechanisms that regulate communication between osteoblasts and osteoclasts, which are known as critical to bone cell biology. [ABSTRACT FROM AUTHOR]

Prior work demonstrated that Phlpp1 deficiency alters limb length and bone mass, but the cell types involved and requirement of Phlpp1 for this effect were unclear. To understand the function of Phlpp1 within bone‐forming osteoblasts, we crossed Phlpp1 floxed mice with mice harboring type 1 collagen (Col1a12.3kb)‐Cre. Mineralization of bone marrow stromal cell cultures derived from Phlpp1 cKOCol1a1 was unchanged, but levels of inflammatory genes (eg, Ifng, Il6, Ccl8) and receptor activator of NF‐κB ligand/osteoprotegerin (RANKL/OPG) ratios were enhanced by either Phlpp1 ablation or chemical inhibition. Micro‐computed tomography of the distal femur and L5 vertebral body of 12‐week‐old mice revealed no alteration in bone volume per total volume, but compromised femoral bone microarchitecture within Phlpp1 cKOCol1a1 conditional knockout females. Bone histomorphometry of the proximal tibia documented no changes in osteoblast or osteoclast number per bone surface but slight reductions in osteoclast surface per bone surface. Overall, our data show that deletion of Phlpp1 in type 1 collagen–expressing cells does not significantly alter attainment of peak bone mass of either males or females, but may enhance inflammatory gene expression and the ratio of RANKL/OPG. Future studies examining the role of Phlpp1 within models of advanced age, inflammation, or osteocytes, as well as functional redundancy with the related Phlpp2 isoform are warranted. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research. [ABSTRACT FROM AUTHOR]

The maintenance of bone homeostasis is dynamically regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. Abnormal differentiation of osteoclast and insufficient osteoblast production can cause bone diseases such as osteoporosis. As one of the highly conserved catabolic pathways in eukaryotic cells, autophagy plays an important role in maintaining cell homeostasis, stress injury repair, proliferation and differentiation. Numerous studies have found that autophagy activity is essential for the survival, differentiation and function of bone cells, and that regulation of autophagy can affect the metabolism of osteoblasts and osteoclasts, thus affecting bone homeostasis. Therefore, using autophagy as a theme, this review outlines the basic process of autophagy, the relationship between autophagy and osteoblasts and osteoclasts, and summarizes the latest research progress of common autophagic signaling pathways in osteoblasts and osteoclasts. The regulatory effects of protein molecules and natural compounds on the autophagy pathway of osteoblasts and osteoclasts discovered in current research are summarized and discussed. This will help to further clarify the mechanism of osteoporosis, understand the relationship between autophagy and osteoporosis, and propose new therapeutic strategies and new ideas for anti-osteoporosis. [ABSTRACT FROM AUTHOR]

Purpose of Review: Metformin is an anti-glycemic agent, which is widely prescribed to diabetes patients. Although its alleged role on bone strength has been reported for some time, this review focuses primarily on the recent mechanistical insights of metformin on osteocytes, osteoblasts, and osteoclasts. Recent Findings: Overall, metformin contributed to steering anabolic activity in osteocytes. It caused lower expression in osteocytes of the negative regulators of bone formation sclerostin and DKK1. Likewise, the osteoclastogenesis function of osteoblasts was also skewed towards lower RANKL and higher OPG expressions. Osteoblast lineage cells generally responded to metformin by activating bone formation parameters, such as alkaline phosphatase activity, higher expression of anabolic members of the Wnt pathway, transcription factor Runx2, bone matrix protein proteins, and subsequent mineralization. Metformin affected osteoclast formation and activity in a negative way, reducing the number of multinucleated cells in association with lower expression of typical osteoclast markers and with inhibited resorption. A common denominator studied in all three cell types is its beneficial effect on activating phosphorylated AMP kinase (AMPK) which is associated with the coordination of energy metabolism. Summary: Metformin differentially affects bone cells, shifting the balance to more bone formation. Although metformin is a drug prescribed for diabetic patients, the overall bone anabolic effects on osteocytes and osteoblasts and the anti-catabolic effect on osteoclast suggest that metformin could be seen as a promising drug in the bone field. [ABSTRACT FROM AUTHOR]

Purpose of Review: Osteoclasts are crucial for the dynamic remodeling of bone as they resorb old and damaged bone, making space for new bone. Metabolic reprogramming in these cells not only supports phenotypic changes, but also provides the necessary energy for their highly energy-consuming activity, bone resorption. In this review, we highlight recent developments in our understanding of the metabolic adaptations that influence osteoclast behavior and the overall remodeling of bone tissue. Recent Findings: Osteoclasts undergo metabolic reprogramming to meet the energy demands during their transition from precursor cells to fully mature bone-resorbing osteoclasts. Recent research has made considerable progress in pinpointing crucial metabolic adaptations and checkpoint proteins in this process. Notably, glucose metabolism, mitochondrial biogenesis, and oxidative respiration were identified as essential pathways involved in osteoclast differentiation, cytoskeletal organization, and resorptive activity. Furthermore, the interaction between these pathways and amino acid and lipid metabolism adds to the complexity of the process. These interconnected processes can function as diverse fuel sources or have independent regulatory effects, significantly influencing osteoclast function. Summary: Energy metabolism in osteoclasts involves various substrates and pathways to meet the energetic requirements of osteoclasts throughout their maturation stages. This understanding of osteoclast biology may provide valuable insights for modulating osteoclast activity during the pathogenesis of bone-related disorders and may pave the way for the development of innovative therapeutic strategies. [ABSTRACT FROM AUTHOR]

Background: MicroRNA-155 (miR-155) is a multifunctional miRNA whose expression is known to be involved in a range of physiological and pathological processes. Its association with several oral diseases has been established. However, the specific role of miR-155 in orthodontic tooth movement remains unclear. In this study, we investigated the impact of miR-155 on osteoclast differentiation and orthodontic tooth movement models, aiming to explore the underlying mechanisms. Methods: In this experiment, we utilized various agents including miR-155 mimic, miR-155 inhibitor, as well as non-specific sequences (NC mimic & NC inhibitor) to treat murine BMMNCs. Subsequently, osteoclast induction (OC) was carried out to examine the changes in the differentiation ability of monocytes under different conditions. To assess these changes, we employed RT-PCR, Western blotting, and TRAP staining techniques. For the orthodontic tooth movement model in mice, the subjects were divided into two groups: the NaCl group (injected with saline solution) and the miR-155 inhibitor group (injected with AntagomiR-155). We observed the impact of orthodontic tooth movement using stereoscopic microscopy, micro-CT, and HE staining. Furthermore, we performed RT-PCR and Western blotting analyses on the tissues surrounding the moving teeth. Additionally, we employed TargetScan to predict potential target genes of miR-155. Results: During osteoclast induction of BMMNCs, the expression of miR-155 exhibited an inverse correlation with osteoclast-related markers. Overexpression of miR-155 led to a decrease in osteoclast-related indexes, whereas underexpression of miR-155 increased those indexes. In the mouse orthodontic tooth movement model, the rate of tooth movement was enhanced following injection of the miR-155 inhibitor, leading to heightened osteoclast activity. TargetScan analysis identified SOCS1 as a target gene of miR-155. Conclusions: Our results suggest that miR-155 functions as an inhibitor of osteoclast differentiation, and it appears to regulate osteoclasts during orthodontic tooth movement. The regulatory mechanism of miR-155 in this process involves the targeting of SOCS1. [ABSTRACT FROM AUTHOR]

Posttraumatic osteomyelitis and the ensuing bone defects are a debilitating complication after open fractures with little therapeutic options. We have recently identified potent osteoanabolic effects of sphingosine‐1‐phosphate (S1P) signalling and have now tested whether it may beneficially affect bone regeneration after infection. We employed pharmacological S1P lyase inhibition by 4‐deoxypyrodoxin (DOP) to raise S1P levels in vivo in an unicortical long bone defect model of posttraumatic osteomyelitis in mice. In a translational approach, human bone specimens of clinical osteomyelitis patients were treated in organ culture in vitro with DOP. Bone regeneration was assessed by μCT, histomorphometry, immunohistology and gene expression analysis. The role of S1P receptors was addressed using S1PR3 deficient mice. Here, we present data that DOP treatment markedly enhanced osteogenesis in posttraumatic osteomyelitis. This was accompanied by greatly improved osteoblastogenesis and enhanced angiogenesis in the callus accompanied by osteoclast‐mediated bone remodelling. We also identified the target of increased S1P to be the S1PR3 as S1PR3−/− mice showed no improvement of bone regeneration by DOP. In the human bone explants, bone mass significantly increased along with enhanced osteoblastogenesis and angiogenesis. Our data suggest that enhancement of S1P/S1PR3 signalling may be a promising therapeutic target for bone regeneration in posttraumatic osteomyelitis. [ABSTRACT FROM AUTHOR]

Aim: Multiple myeloma is a malignancy of clonal plasmacytes. Osteolytic lesions represent a criterion for symptomatic myeloma and are associated with bone loss, pathological fractures, and osteoporosis. Skeletal surveys with other sophisticated techniques and dual-energy x-ray absorptiometry (DEXA) are used to screen lytic lesions, and bone mineral loss, respectively. Here, we aimed to investigate the rates of detection regarding osteolytic lesions and bone mineral loss by several imaging techniques. Materials and Methods: The study was carried out in Aydın Adnan Menderes University Hospital/Turkey, between the years 2004-2020. Three-hundred and ten symptomatic myeloma patients were screened retrospectively. The results of radiological techniques were recorded. The detection rate of osteolytic lesions, fractures, and plasmacytomas by imaging techniques, as well as the detection rate of bone mineral loss with DEXA were recorded. Also, the relationship between the detected lytic lesions and gender, myeloma type, and the sensitivity and specificity of DEXA to detect osteoporosis in cases with lytic lesions were investigated. Results: Skeletal survey and PET-CT detected lytic lesions in 71.3% and 81.2% of patients, respectively. PET-CT had a sensitivity of 96.1% and specificity of 90.6% to detect lytic lesions. MRI was only used for patients with suspicious fractures and detected them for all patients who underwent MRI. The osteoporosis rate was 83.1% for 113 patients who underwent DEXA. No association was detected between lytic lesions and gender/myeloma type. The sensitivity and specificity of BMD-DEXA to detect osteoporosis among patients who have lytic lesions were 85.3% and 23.5%, respectively. Conclusion: Our study demonstrated that osteolytic lesions are not correlated with gender or myeloma type. PET-CT is a sensitive and specific method for detecting osteolytic lesions. Although DEXA is sensitive, its specificity is limited in patients with lytic lesions. Giriş: Multipl myelom, klonal plazma hücrelerinin bir malignitesidir. Osteolitik lezyon varlığı, semptomatik myelom için bir tanı kriteri olarak kabul edilmektedir. Ayrıca, kemik kaybı, patolojik kırıklar ve osteoporoz ile ilişkilidir. Litik lezyon varlığı ve kemik mineral kaybını saptamak için çeşitli görüntüleme yöntemleri ve çift enerjili x-ışını absorbsiyometrisi (DEXA) kullanılır. Bu çalışmada görüntüleme yöntemleri ve DEXA ile osteolitik lezyonların ve kemik mineral kaybının saptanma oranlarının araştırılması amaçlanmıştır. Gereç ve Yöntem: Bu çalışma 2004-2020 yılları arasında Aydın Adnan Menderes Üniversitesi Hastanesi/Türkiye'de gerçekleştirildi. Üç yüz on semptomatik myelom hastası retrospektif olarak incelendi. Osteolitik lezyon, fraktür ve plazmasitomların görüntüleme yöntemleriyle tespit oranı ve ayrıca DEXA ile kemik mineral kaybı saptama oranı kaydedildi. Ayrıca tespit edilen litik lezyonlar ile cinsiyet, myelom tipi arasındaki ilişki ve litik lezyon saptanan olgularda DEXA'nın osteoporozu saptama duyarlılığı ve özgüllüğü araştırıldı. Bulgular: Litik lezyonlar, direkt grafi ve PET-BT ile hastaların sırasıyla %71,3'ünde ve %81,2'sinde saptadı. PET-BT'nin litik lezyonları saptamada duyarlılığı %96,1 ve özgüllüğü %90,6 idi. MR görüntülemesine, sadece fraktür şüphesi olan hastalarda başvuruldu ve MR çekilen tüm hastalarda fraktür tespit edildi. DEXA çekilen 113 hastada osteoporoz oranı %83,1 idi. Litik lezyon varlığı ile cinsiyet/myelom tipi arasında herhangi bir ilişki saptanmadı. Osteolitik lezyon saptanan olgularda DEXA' nın osteoporozu saptamadaki duyarlılığı %83,3, özgüllüğü %23,5 olarak bulundu. Sonuç: Çalışmamız, osteolitik lezyonların cinsiyet veya myelom tipi ile korele olmadığını göstermiştir. PET-BT, osteolitik lezyonları saptamak için hassas ve spesifik bir yöntemdir. DEXA, osteoporozu saptamada duyarlı bir yöntem olmasına ragmen; özgüllüğü, litik lezyonları olan hastalarda sınırlı bir değere sahiptir. [ABSTRACT FROM AUTHOR]

Introduction: The gastrointestinal neuroectodermal tumor (GNET) is a rare mesenchymal tumor mainly arising in the gastrointestinal tract. Case Presentation: Herein, we describe a case of GNET that had been previously diagnosed as metastatic primitive neuroectodermal tumor (PNET) to the lung and liver and received 30 cycles of adjuvant chemotherapy. The physical exam showed a palpable mass in the lower right quadrant of the abdomen. The tumor mass was composed of diffuse sheets of epithelioid tumor cells, in which osteoclast-like giant cells scattered unevenly. Immunohistochemically, the tumor cells were diffusely positive for SOX10, CD99, and CD56, patchy positive forpan-CK, and negative fordesmin, HMB-45, Melan-A, and chromogranin. Conclusions: Consistent with previous studies that reported a secondary GNET following chemotherapy, we assume that the chemotherapy might trigger the tumorigenesis of GNET in this case. [ABSTRACT FROM AUTHOR]

Bone formation is a constant procedure in which osteoblasts lay new bone and osteoclasts resorb it. Mineralization takes place at active bone formation sites where the extracellular matrix vesicles, the major sites of apatite mineral deposition, are present. Turnover of bone, which maintains its structure and integrity, has several events such as activation, resorption and osteogenesis by osteoblasts. The bone crystals proliferation depends on the presence of collagen, hormones, minerals and vitamins such as phosphorus, magnesium, calcium, zinc, fluoride, potassium, manganese, boron, copper, iron, calcium and a number of vitamins such as B,K,C,A,D, etc. Information regarding the elements involved in bone formation was retrieved through studies published up to 2017 in PubMed, Medline and other authentic search engines available in the University. This review highlights the individual roles of specific vitamins and minerals at the respective steps of bone formation, insufficiency at all particular stages result in various bone pathologies, leading to deficiency disorders, fractures and poor friable bones. Since the role of vitamin D and calcium is well established therefore these were not included in this review.

BackgroundPostmenopausal osteoporosis is a prevalent disease that affects the bone health of middle-aged and elderly women. The link between gut microbiota and bone health, known as the gut-bone axis, has garnered widespread attention.MethodsWe employed a two-sample Mendelian randomization approach to assess the associations between gut microbiota with osteoclasts and postmenopausal osteoporosis, respectively. Single nucleotide polymorphisms associated with the composition of gut microbiota were used as instrumental variables. By analyzing large-scale multi-ethnic GWAS data from the international MiBioGen consortium, and combining data from the eQTLGen consortium and the GEFOS consortium, we identified microbiota related to osteoclasts and postmenopausal osteoporosis. Key genes were further identified through MAGMA analysis, and validation was performed using single-cell data GSE147287.ResultsThe outcomes of this study have uncovered significant associations within the gut microbiome community, particularly with the Burkholderiales order, which correlates with both an increase in osteoclasts and a reduced risk of postmenopausal osteoporosis. with an odds ratio (OR) of 0.400, and a P-value of 0.011. Further analysis using single-cell data allowed us to identify two key genes, FMNL2 and SRBD1, that are closely linked to both osteoclasts and osteoporosis.ConclusionThis study utilizing Mendelian randomization and single-cell data analysis, provides new evidence of a causal relationship between gut microbiota and osteoclasts, as well as postmenopausal osteoporosis. It was discovered that the specific microbial group, the Burkholderiales order, significantly impacts both osteoporosis and osteoclasts. Additionally, key genes FMNL2 and SRBD1 were identified, offering new therapeutic strategies for the treatment of postmenopausal osteoporosis.

The transcription factor, signal transducer, and stimulator of transcription 3 (STAT3) is a potential target in osteoarthritis (OA) treatment. Although xanthatin (XA), a biologically active substance derived from Xanthium strumarium L, specifically inhibits STAT3 phosphorylation at Tyr705, the mechanism underlying its inhibitory effect on OA progression remains unclear. In this study, our objective was to explore the therapeutic effects exerted by XA on OA and the underlying molecular mechanisms. The effects of XA treatment on mouse OA models subjected to destabilization of the medial meniscus using medial collateral ligament transection, as well as on interleukin-1β (IL-1β)-induced mouse chondrocytes, were examined. Histological changes in cartilage and subchondral bone (SCB), as well as changes in the expression levels of osteophytes, cartilage degeneration- and osteoclast differentiation-related factors, and the role of XA-related signaling pathways in human cartilage tissue, were studied using different techniques. XA inhibited STAT3 phosphorylation at Tyr705 and further attenuated the activity of nuclear factor-κB (NF-κB) in chondrocytes and osteoclasts. In vitro, XA administration alleviated pro-inflammatory cytokine release, extracellular matrix catabolism, and RANKL-mediated osteoclast differentiation. In vivo, intraperitoneal injection of XA exerted a protective effect on cartilage degeneration and SCB loss. Similarly, XA exerted a protective effect on human cartilage tissue by inhibiting the STAT3/NF-κB signaling pathway. Overall, our study elucidated the therapeutic potential of XA as a small-molecule inhibitor of STAT3-driven OA progression. This discovery may help enhance innovative clinical interventions against OA.

Bone remodeling is a complex process involving the coordinated actions of osteoblasts and osteoclasts to maintain bone homeostasis. While the influence of osteoblasts on osteoclast differentiation is well established, the reciprocal regulation of osteoblasts by osteoclasts has long remained enigmatic. In the past few years, a fascinating new role for osteoclasts has been unveiled in promoting bone formation and facilitating osteoblast migration to the remodeling sites through a number of different mechanisms, including the release of factors from the bone matrix following bone resorption and direct cell–cell interactions. Additionally, considerable evidence has shown that osteoclasts can secrete coupling factors known as clastokines, emphasizing the crucial role of these cells in maintaining bone homeostasis. Due to their osteoprotective function, clastokines hold great promise as potential therapeutic targets for bone diseases. However, despite long-standing work to uncover new clastokines and their effect in vivo, more substantial efforts are still required to decipher the mechanisms and pathways behind their activity in order to translate them into therapies. This comprehensive review provides insights into our evolving understanding of the osteoclast function, highlights the significance of clastokines in bone remodeling, and explores their potential as treatments for bone diseases suggesting future directions for the field.

It is well established that inflammatory processes in the vicinity of bone often induce osteoclast formation and bone resorption. Effects of inflammatory processes on bone formation are less studied. Therefore, we investigated the effect of locally induced inflammation on bone formation. Toll-like receptor (TLR) 2 agonists LPS from Porphyromonas gingivalis and PAM2 were injected once subcutaneously above mouse calvarial bones. After five days, both agonists induced bone formation mainly at endocranial surfaces. The injection resulted in progressively increased calvarial thickness during 21 days. Excessive new bone formation was mainly observed separated from bone resorption cavities. Anti-RANKL did not affect the increase of bone formation. Inflammation caused increased bone formation rate due to increased mineralizing surfaces as assessed by dynamic histomorphometry. In areas close to new bone formation, an abundance of proliferating cells was observed as well as cells robustly stained for Runx2 and alkaline phosphatase. PAM2 increased the mRNA expression of Lrp5, Lrp6 and Wnt7b, and decreased the expression of Sost and Dkk1. In situ hybridization demonstrated decreased Sost mRNA expression in osteocytes present in old bone. An abundance of cells expressed Wnt7b in Runx2-positive osteoblasts and ß-catenin in areas with new bone formation. These data demonstrate that inflammation, not only induces osteoclastogenesis, but also locally activates canonical WNT signaling and stimulates new bone formation independent on bone resorption.

Abstract Background Ever since their discovery, induced pluripotent stem cells (iPSCs) have been extensively differentiated into a large variety of cell types. However, a limited amount of work has been dedicated to differentiating iPSCs into osteoclasts. While several differentiation protocols have been published, it remains unclear which protocols or differentiation methods are preferable regarding the differentiation of osteoclasts. Methods In this study, we compared the osteoclastogenesis capacity of a peripheral blood mononuclear cell (PBMC)-derived iPSC line to a fibroblast-derived iPSC line in conjunction with either embryoid body-based or monolayer-based differentiation strategies. Both cell lines and differentiation protocols were investigated regarding their ability to generate osteoclasts and their inherent robustness and ease of use. The ability of both cell lines to remain undifferentiated while propagating using a feeder-free system was assessed using alkaline phosphatase staining. This was followed by evaluating mesodermal differentiation and the characterization of hematopoietic progenitor cells using flow cytometry. Finally, osteoclast yield and functionality based on resorptive activity, Cathepsin K and tartrate-resistant acid phosphatase (TRAP) expression were assessed. The results were validated using qRT-PCR throughout the differentiation stages. Results Embryoid body-based differentiation yielded CD45+, CD14+, CD11b+ subpopulations which in turn differentiated into osteoclasts which demonstrated TRAP positivity, Cathepsin K expression and mineral resorptive capabilities. This was regardless of which iPSC line was used. Monolayer-based differentiation yielded lower quantities of hematopoietic cells that were mostly CD34+ and did not subsequently differentiate into osteoclasts. Conclusions The outcome of this study demonstrates the successful differentiation of osteoclasts from iPSCs in conjunction with the embryoid-based differentiation method, while the monolayer-based method did not yield osteoclasts. No differences were observed regarding osteoclast differentiation between the PBMC and fibroblast-derived iPSC lines.

Aims: Currently, the effect of drug treatment for osteoporosis is relatively poor, and the side effects are numerous and serious. Melatonin is a potential drug to improve bone mass in postmenopausal women. Unfortunately, the mechanism by which melatonin improves bone metabolism remains unclear. The aim of this study was to further investigate the potential mechanism of melatonin in the treatment of osteoporosis. Methods: The effects of melatonin on mitochondrial apoptosis protein, bmal1 gene, and related pathway proteins of RAW264.7 (mouse mononuclear macrophage leukaemia cells) were analyzed by western blot. Cell Counting Kit-8 was used to evaluate the effect of melatonin on cell viability. Flow cytometry was used to evaluate the effect of melatonin on the apoptosis of RAW264.7 cells and mitochondrial membrane potential. A reactive oxygen species (ROS) detection kit was used to evaluate the level of ROS in osteoclast precursors. We used bmal1-small interfering RNAs (siRNAs) to downregulate the Bmal1 gene. We established a postmenopausal mouse model and verified the effect of melatonin on the bone mass of postmenopausal osteoporosis in mice via micro-CT. Bmal1 lentiviral activation particles were used to establish an in vitro model of overexpression of the bmal1 gene. Results: Melatonin promoted apoptosis of RAW264.7 cells and increased the expression of BMAL1 to inhibit the activation of ROS and phosphorylation of mitogen-activated protein kinase (MAPK)-p38. Silencing the bmal1 gene weakened the above effects of melatonin. After that, we used dehydrocorydaline (DHC) to enhance the activation of MAPK-p38, and the effects of melatonin on reducing ROS levels and promoting apoptosis of RAW264.7 cells were also blocked. Then, we constructed a mouse model of postmenopausal osteoporosis and administered melatonin. The results showed that melatonin improves bone loss in ovariectomized mice. Finally, we established a model of overexpression of the bmal1 gene, and these results suggest that the bmal1 gene can regulate ROS activity and change the level of the MAPK-p38 signalling pathway. Conclusion: Our study confirmed that melatonin promotes the apoptosis of RAW264.7 cells through BMAL1/ROS/MAPK-p38, and revealed the therapeutic effect and mechanism of melatonin in postmenopausal osteoporosis. This finding enriches BMAL1 as a potential target for the treatment of osteoporosis and the pathogenesis of postmenopausal osteoporosis. Cite this article: Bone Joint Res 2023;12(11):677–690.

Aims: Osteoporosis is characterized by decreased trabecular bone volume, and microarchitectural deterioration in the medullary cavity. Interleukin-19 (IL-19), a member of the IL-10 family, is an anti-inflammatory cytokine produced primarily by macrophages. The aim of our study was to investigate the effect of IL-19 on osteoporosis. Methods: Blood and femoral bone marrow suspension IL-19 levels were first measured in the lipopolysaccharide (LPS)-induced bone loss model. Small interfering RNA (siRNA) was applied to knock down IL-19 for further validation. Thereafter, osteoclast production was stimulated with IL-19 in combination with mouse macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). The effect of IL-19 was subsequently evaluated using tartrate-resistant acid phosphatase (TRAP) staining and quantitative real-time polymerase chain reaction (RT-qPCR). The effect of IL-19 on osteoprotegerin (OPG) was then assessed using in vitro recombinant IL-19 treatment of primary osteoblasts and MLO-Y4 osteoblast cell line. Finally, transient transfection experiments and chromatin immunoprecipitation (ChIP) experiments were used to examine the exact mechanism of action. Results: In the LPS-induced bone loss mouse model, the levels of IL-19 in peripheral blood serum and femoral bone marrow suspension were significantly increased. The in vivo results indicated that global IL-19 deletion had no significant effect on RANKL content in the serum and bone marrow, but could increase the content of OPG in serum and femoral bone marrow, suggesting that IL-19 inhibits OPG expression in bone marrow mesenchymal stem cells (BMSCs) and thus increases bone resorption. Conclusion: IL-19 promotes bone resorption by suppressing OPG expression in BMSCs in a LPS-induced bone loss mouse model, which highlights the potential benefits and side effects of IL-19 for future clinical applications. Cite this article: Bone Joint Res 2023;12(11):691–701.

Yi-Ming Gao,1 Yan Pei,1 Fei-Fei Zhao,2 Ling Wang1 1Department of Orthopedic Oncology, The Third Hospital of Hebei Medical University, Shijiazhuang, Hebei, People’s Republic of China; 2Department of Orthopedics, The Fourth Hospital of Hebei Medical University, Shijiazhuang, Hebei, People’s Republic of ChinaCorrespondence: Ling Wang, Tel +86031188603682, Email wangling2016uw@126.comAbstract: Osteosarcoma is an extremely malignant tumor, and its pathogenesis is complex and remains incompletely understood. Most cases of osteosarcoma are accompanied by symptoms of bone loss or result in pathological fractures due to weakened bones. Enhancing the survival rate of osteosarcoma patients has proven to be a long-standing challenge. Numerous studies mentioned in this paper, including in-vitro, in-vivo, and in-situ studies have consistently indicated a close association between the symptoms of bone loss associated with osteosarcoma and the presence of osteoclasts. As the sole cells capable of bone resorption, osteoclasts participate in a malignant cycle within the osteosarcoma microenvironment. These cells interact with osteoblasts and osteosarcoma cells, secreting various factors that further influence these cells, disrupting bone homeostasis, and shifting the balance toward bone resorption, thereby promoting the onset and progression of osteosarcoma. Moreover, the interaction between osteoclasts and various other cells types, such as tumor-associated macrophages, myeloid-derived suppressor cells, DCs cells, T cells, and tumor-associated fibroblasts in the osteosarcoma microenvironment plays a crucial role in disease progression. Consequently, understanding the role of osteoclasts in osteosarcoma has sparked significant interest. This review primarily examines the physiological characteristics and functional mechanisms of osteoclasts in osteosarcoma, and briefly discusses potential therapies targeting osteoclasts for osteosarcoma treatment. These studies provide fresh ideas and directions for future research on the treatment of osteosarcoma.Keywords: osteosarcoma, osteoclasts, bone remodeling, osteoimmunology, bone microenvironment

Aims: The management of periprosthetic joint infection (PJI) remains a major challenge in orthopaedic surgery. In this study, we aimed to characterize the local bone microstructure and metabolism in a clinical cohort of patients with chronic PJI. Methods: Periprosthetic femoral trabecular bone specimens were obtained from patients suffering from chronic PJI of the hip and knee (n = 20). Microbiological analysis was performed on preoperative joint aspirates and tissue specimens obtained during revision surgery. Microstructural and cellular bone parameters were analyzed in bone specimens by histomorphometry on undecalcified sections complemented by tartrate-resistant acid phosphatase immunohistochemistry. Data were compared with control specimens obtained during primary arthroplasty (n = 20) and aseptic revision (n = 20). Results: PJI specimens exhibited a higher bone volume, thickened trabeculae, and increased osteoid parameters compared to both control groups, suggesting an accelerated bone turnover with sclerotic microstructure. On the cellular level, osteoblast and osteoclast parameters were markedly increased in the PJI cohort. Furthermore, a positive association between serum (CRP) but not synovial (white blood cell (WBC) count) inflammatory markers and osteoclast indices could be detected. Comparison between different pathogens revealed increased osteoclastic bone resorption parameters without a concomitant increase in osteoblasts in bone specimens from patients with Staphylococcus aureus infection, compared to those with detection of Staphylococcus epidermidis and Cutibacterium spp. Conclusion: This study provides insights into the local bone metabolism in chronic PJI, demonstrating osteosclerosis with high bone turnover. The fact that Staphylococcus aureus was associated with distinctly increased osteoclast indices strongly suggests early surgical treatment to prevent periprosthetic bone alterations. Cite this article: Bone Joint Res 2023;12(10):644–653.

Maintenance of skeletal integrity requires the coordinated activity of multinucleated bone-resorbing osteoclasts and bone-forming osteoblasts. Osteoclasts form resorption lacunae on bone surfaces in response to cytokines by fusion of precursor cells. Osteoblasts are derived from mesenchymal precursors and lay down new bone in resorption lacunae during bone remodeling. Nuclear factor-kappa B (NF-κB) signaling regulates osteoclast and osteoblast formation and is activated in osteoclast precursors in response to the essential osteoclastogenic cytokine, receptor activator of NF-κB ligand (RANKL), which can also control osteoblast formation through RANK-RANKL reverse signaling in osteoblast precursors. RANKL and some pro-inflammatory cytokines, including tumor necrosis factor (TNF), activate NF-κB signaling to positively regulate osteoclast formation and functions. However, these cytokines also limit osteoclast and osteoblast formation through NF-κB signaling molecules, including TNF receptor-associated factors (TRAFs). TRAF6 mediates RANKL-induced osteoclast formation through canonical NF-κB signaling. In contrast, TRAF3 limits RANKL- and TNF-induced osteoclast formation, and it restricts transforming growth factor β (TGFβ)-induced inhibition of osteoblast formation in young and adult mice. During aging, neutrophils expressing TGFβ and C-C chemokine receptor type 5 (CCR5) increase in bone marrow of mice in response to increased NF-κB-induced CC motif chemokine ligand 5 (CCL5) expression by mesenchymal progenitor cells and injection of these neutrophils into young mice decreased bone mass. TGFβ causes degradation of TRAF3, resulting in decreased glycogen synthase kinase-3β/β-catenin-mediated osteoblast formation and age-related osteoporosis in mice. The CCR5 inhibitor, maraviroc, prevented accumulation of TGFβ+/CCR5+ neutrophils in bone marrow and increased bone mass by inhibiting bone resorption and increasing bone formation in aged mice. This paper updates current understanding of how NF-κB signaling is involved in the positive and negative regulation of cytokine-mediated osteoclast and osteoblast formation and activation with a focus on the role of TRAF3 signaling, which can be targeted therapeutically to enhance bone mass.

Abstract Background Though articular cartilage stem cell (ACSC)-based therapies have been demonstrated to be a promising option in the treatment of diseased joints, the wide variety of cell isolation, the unknown therapeutic targets, and the incomplete understanding of the interactions of ACSCs with diseased microenvironments have limited the applications of ACSCs. Methods In this study, the human ACSCs have been isolated from osteoarthritic articular cartilage by advantage of selection of anatomical location, the migratory property of the cells, and the combination of traumatic injury, mechanical stimuli and enzymatic digestion. The protective effects of ACSC infusion into osteoarthritis (OA) rat knees on osteochondral tissues were evaluated using micro-CT and pathological analyses. Moreover, the regulation of ACSCs on osteoarthritic osteoclasts and the underlying mechanisms in vivo and in vitro were explored by RNA-sequencing, pathological analyses and functional gain and loss experiments. The one-way ANOVA was used in multiple group data analysis. Results The ACSCs showed typical stem cell-like characteristics including colony formation and committed osteo-chondrogenic capacity. In addition, intra-articular injection into knee joints yielded significant improvement on the abnormal subchondral bone remodeling of osteoarthritic rats. Bioinformatic and functional analysis showed that ACSCs suppressed osteoarthritic osteoclasts formation, and inflammatory joint microenvironment augmented the inhibitory effects. Further explorations demonstrated that ACSC-derived tumor necrosis factor alpha-induced protein 3 (TNFAIP3) remarkably contributed to the inhibition on osteoarhtritic osteoclasts and the improvement of abnormal subchondral bone remodeling. Conclusion In summary, we have reported an easy and reproducible human ACSC isolation strategy and revealed their effects on subchondral bone remodeling in OA rats by releasing TNFAIP3 and suppressing osteoclasts in a diseased microenvironment responsive manner. Graphical abstract

Abstract The occurrence of osteoporosis gradually increases within the aging population. As the side effects of therapeutic agents currently used for osteoporosis are increasing, the development of preventive and therapeutic agents derived from natural products without any long-term side effects is important. Here, we investigated the effect of wheat seedling extract (WSE) on the RANKL-mediated differentiation, fusion, and function of osteoclasts. WSE inhibited the differentiation of RANKL-induced bone marrow macrophages and phosphorylation of AKT and ERK. Moreover, the protein and mRNA expression levels of c-Fos and NFATc1 as well as RANKL-induced transcription of TRAP and OSCAR were suppressed by WSE treatment. DC-STAMP and cathepsin K, which are essential for cell fusion and bone degradation, were also inhibited by WSE. Furthermore, eight components constituting WSE were confirmed to decrease the osteoclast TRAP activity. Taken together, WSE may have potential implications as a useful therapeutic or preventive agent for inhibition of bone loss.

Osteoarthritis (OA) is mainly caused by ageing, strain, trauma, and congenital joint abnormalities, resulting in articular cartilage degeneration. During the pathogenesis of OA, the changes in subchondral bone (SB) are not only secondary manifestations of OA, but also an active part of the disease, and are closely associated with the severity of OA. In different stages of OA, there were microstructural changes in SB. Osteocytes, osteoblasts, and osteoclasts in SB are important in the pathogenesis of OA. The signal transduction mechanism in SB is necessary to maintain the balance of a stable phenotype, extracellular matrix (ECM) synthesis, and bone remodelling between articular cartilage and SB. An imbalance in signal transduction can lead to reduced cartilage quality and SB thickening, which leads to the progression of OA. By understanding changes in SB in OA, researchers are exploring drugs that can regulate these changes, which will help to provide new ideas for the treatment of OA. Cite this article: Bone Joint Res 2023;12(9):536–545.

Rare skeletal diseases are still in need of proper clinically available transfection agents as the major challenge for first-in-human translation relates to intrinsic difficulty in targeting bone without exacerbating any inherent toxicity due to used vector. SiSaf’s silicon stabilized hybrid lipid nanoparticles (sshLNPs) constitute next-generation non-viral vectors able to retain the integrity and stability of constructs and to accommodate considerable payloads of biologicals, without requiring cold-chain storage. sshLNP was complexed with a small interfering RNA (siRNA) specifically designed against the human CLCN7G215R mRNA. When tested via single intraperitoneal injection in pre-puberal autosomal dominant osteopetrosis type 2 (ADO2) mice, carrying a heterozygous mutation of the Clcn7 gene (Clcn7G213R), sshLNP, this significantly downregulated the Clcn7G213R related mRNA levels in femurs at 48 h. Confirmatory results were observed at 2 weeks and 4 weeks after treatments (3 intraperitoneal injections/week), with rescue of the bone phenotype and demonstrating safety. The pre-clinical results will enable advanced preclinical development of RNA-based therapy for orphan and genetic skeletal disorders by safely and effectively delivering biologicals of interest to cure human systemic conditions.

Sexually dimorphic tissues are formed by cells that are regulated by sex hormones. While a number of systemic hormones and transcription factors are known to regulate proliferation and differentiation of osteoblasts and osteoclasts, the mechanisms that determine sexually dimorphic differences in bone regeneration are unclear. To explore how sex hormones regulate bone regeneration, we compared bone fracture repair between adult male and female mice. We found that skeletal stem cell (SSC) mediated regeneration in female mice is dependent on estrogen signaling but SSCs from male mice do not exhibit similar estrogen responsiveness. Mechanistically, we found that estrogen acts directly on the SSC lineage in mice and humans by up-regulating multiple skeletogenic pathways and is necessary for the stem cell's ability to self- renew and differentiate. Our results also suggest a clinically applicable strategy to accelerate bone healing using localized estrogen hormone therapy.

Bone cement containing benzoyl peroxide (BPO) as a polymerization initiator are commonly used to fix orthopedic metal implants. However, toxic complications caused by bone cement are a clinically significant problem. Poly (methyl methacrylate) tri-n-butylborane (PMMA-TBB), a newly developed material containing TBB as a polymerization initiator, was found to be more biocompatible than conventional PMMA-BPO bone cements due to reduced free radical generation during polymerization. However, free radicals might not be the only determinant of cytotoxicity. Here, we evaluated the response and functional phenotypes of cells exposed to extracts derived from different bone cements. Bone cement extracts were prepared from two commercial PMMA-BPO cements and an experimental PMMA-TBB. Rat bone marrow-derived osteoblasts and osteoclasts were cultured in a medium supplemented with bone cement extracts. More osteoblasts survived and attached to the culture dish with PMMA-TBB extract than in the culture with PMMA-BPO extracts. Osteoblast proliferation and differentiation were higher in the culture with PMMA-TBB extract. The number of TRAP-positive multinucleated cells was significantly lower in the culture with PMMA-TBB extract. There was no difference in osteoclast-related gene expression in response to different bone cement extracts. In conclusion, PMMA-TBB extract was less toxic to osteoblasts than PMMA-BPO extracts. Although extracts from the different cement types did not affect osteoclast function, PMMA-TBB extract seemed to reduce osteoclastogenesis, a possible further advantage of PMMA-TBB cement. These implied that the reduced radical generation during polymerization is not the only determinant for the improved biocompatibility of PMMA-TBB and that the post-polymerization chemical elution may also be important.

The article discusses the link between periodontitis and rheumatoid arthritis (RA) and asks if regular periodontal treatment is needed for RA patients. It explains how periodontal pathogens can worsen RA and suggests that periodontal therapy, especially non-surgical treatment, can improve symptoms in patients with both conditions. It recommend regular periodontal care for RA patients to manage inflammation and counteract the effects of periodontal pathogens.

Glycosphingolipids (GSLs) are key constituents of membrane bilayers playing a role in structural integrity, cell signalling in microdomains, endosomes and lysosomes, and cell death pathways. Conversion of ceramide into GSLs is controlled by GCS (glucosylceramide synthase) and inhibitors of this enzyme for the treatment of lipid storage disorders and specific cancers. With a diverse range of functions attributed to GSLs, the ability of the GSC inhibitor, eliglustat, to reduce myeloma bone disease was investigated. In pre-clinical models of multiple myeloma, osteoclast-driven bone loss was reduced by eliglustat in a mechanistically separate manner to zoledronic acid, a bisphosphonate that prevents osteoclast-mediated bone destruction. Autophagic degradation of TNF receptor-associated factor 3 (TRAF3), a key step for osteoclast differentiation, was inhibited by eliglustat as evidenced by TRAF3 lysosomal and cytoplasmic accumulation. By altering GSL composition, eliglustat prevented lysosomal degradation whilst exogenous addition of missing GSLs rescued TRAF3 degradation to restore osteoclast formation in bone marrow cells from myeloma patients. This work highlights the clinical potential of eliglustat as a therapy for myeloma bone disease. Furthermore, using eliglustat as a lysosomal inhibitor in osteoclasts may widen its therapeutic uses to other bone disorders such as bone metastasis, osteoporosis and inflammatory bone loss. [ABSTRACT FROM AUTHOR]