Hyperthyroidism-driven bone loss depends on BMP receptor Bmpr1a expression in osteoblasts.

Hyperthyroidism is a well-known trigger of high bone turnover that can lead to the development of secondary osteoporosis. Previously, we have shown that blocking bone morphogenetic protein (BMP) signaling systemically with BMPR1A-Fc can prevent bone loss in hyperthyroid mice. To distinguish between bone cell type-specific effects, conditional knockout mice lacking Bmpr1a in either osteoclast precursors (LysM-Cre) or osteoprogenitors (Osx-Cre) were rendered hyperthyroid and their bone microarchitecture, strength and turnover were analyzed. While hyperthyroidism in osteoclast precursor-specific Bmpr1a knockout mice accelerated bone resorption leading to bone loss just as in wildtype mice, osteoprogenitor-specific Bmpr1a deletion prevented an increase of bone resorption and thus osteoporosis with hyperthyroidism. In vitro, wildtype but not Bmpr1a-deficient osteoblasts responded to thyroid hormone (TH) treatment with increased differentiation and activity. Furthermore, we found an elevated Rankl/Opg ratio with TH excess in osteoblasts and bone tissue from wildtype mice, but not in Bmpr1a knockouts. In line, expression of osteoclast marker genes increased when osteoclasts were treated with supernatants from TH-stimulated wildtype osteoblasts, in contrast to Bmpr1a-deficient cells. In conclusion, we identified the osteoblastic BMP receptor BMPR1A as a main driver of osteoporosis in hyperthyroid mice promoting TH-induced osteoblast activity and potentially its coupling to high osteoclastic resorption.

Circadian Regulation of Bone Remodeling.

Adult bones are continuously remodeled by the balance between bone resorption by osteoclasts and subsequent bone formation by osteoblasts. Many studies have provided molecular evidence that bone remodeling is under the control of circadian rhythms. Circadian fluctuations have been reported in the serum and urine levels of bone turnover markers, such as digested collagen fragments and bone alkaline phosphatase. Additionally, the expressions of over a quarter of all transcripts in bones show circadian rhythmicity, including the genes encoding master transcription factors for osteoblastogenesis and osteoclastogenesis, osteogenic cytokines, and signaling pathway proteins. Serum levels of calcium, phosphate, parathyroid hormone, and calcitonin also display circadian rhythmicity. Finally, osteoblast- and osteoclast-specific knockout mice targeting the core circadian regulator gene Bmal1 show disrupted bone remodeling, although the results have not always been consistent. Despite these studies, however, establishing a direct link between circadian rhythms and bone remodeling in vivo remains a major challenge. It is nearly impossible to repeatedly collect bone materials from human subjects while following circadian changes. In addition, the differences in circadian gene regulation between diurnal humans and nocturnal mice, the main model organism, remain unclear. Filling the knowledge gap in the circadian regulation of bone remodeling could reveal novel regulatory mechanisms underlying many bone disorders including osteoporosis, genetic diseases, and fracture healing. This is also an important question for the basic understanding of how cell differentiation progresses under the influence of cyclically fluctuating environments.

Sclerostin blockade inhibits bone resorption through PDGF receptor signaling in osteoblast lineage cells.

While sclerostin-neutralizing antibodies (Scl-Abs) transiently stimulate bone formation by activating Wnt signaling in osteoblast lineage cells, they exert sustained inhibition of bone resorption, suggesting an alternate signaling pathway by which Scl-Abs control osteoclast activity. Since sclerostin can activate platelet-derived growth factor receptors (PDGFRs) in osteoblast lineage cells in vitro and PDGFR signaling in these cells induces bone resorption through M-CSF secretion, we hypothesized that the prolonged anticatabolic effect of Scl-Abs could result from PDGFR inhibition. We show here that inhibition of PDGFR signaling in osteoblast lineage cells is sufficient and necessary to mediate prolonged Scl-Ab effects on M-CSF secretion and osteoclast activity in mice. Indeed, sclerostin coactivates PDGFRs independently of Wnt/ß-catenin signaling inhibition, by forming a ternary complex with LRP6 and PDGFRs in preosteoblasts. In turn, Scl-Ab prevents sclerostin-mediated coactivation of PDGFR signaling and consequent M-CSF upregulation in preosteoblast cultures, thereby inhibiting osteoclast activity in preosteoblast/osteoclast coculture assays. These results provide a potential mechanism explaining the dissociation between anabolic and antiresorptive effects of long-term Scl-Ab.

RUFY4 deletion prevents pathological bone loss by blocking endo-lysosomal trafficking of osteoclasts.

Mature osteoclasts degrade bone matrix by exocytosis of active proteases from secretory lysosomes through a ruffled border. However, the molecular mechanisms underlying lysosomal trafficking and secretion in osteoclasts remain largely unknown. Here, we show with GeneChip analysis that RUN and FYVE domain-containing protein 4 (RUFY4) is strongly upregulated during osteoclastogenesis. Mice lacking Rufy4 exhibited a high trabecular bone mass phenotype with abnormalities in osteoclast function in vivo. Furthermore, deleting Rufy4 did not affect osteoclast differentiation, but inhibited bone-resorbing activity due to disruption in the acidic maturation of secondary lysosomes, their trafficking to the membrane, and their secretion of cathepsin K into the extracellular space. Mechanistically, RUFY4 promotes late endosome-lysosome fusion by acting as an adaptor protein between Rab7 on late endosomes and LAMP2 on primary lysosomes. Consequently, Rufy4-deficient mice were highly protected from lipopolysaccharide- and ovariectomy-induced bone loss. Thus, RUFY4 plays as a new regulator in osteoclast activity by mediating endo-lysosomal trafficking and have a potential to be specific target for therapies against bone-loss diseases such as osteoporosis.

ED-71 Improves Bone Mass in Ovariectomized Rats by Inhibiting Osteoclastogenesis Through EphrinB2-EphB4-RANKL/OPG Axis.

Purpose: Estrogen deficiency is the main reason of postmenopausal osteoporosis. Eldecalcitol (ED-71) is a new active vitamin D analogue clinically used in the treatment of postmenopausal osteoporosis. We aimed to investigate whether EphrinB2-EphB4 and RANKL/RANK/OPG signaling cooperate in mediating the process of osteoporosis by ED-71. Methods: In vivo, the ovariectomized (OVX) rats were administered orally with 30 ng/kg ED-71 once a day for 8 weeks. HE staining, Masson staining and Immunofluorescence staining were used to evaluate bone mass, bone formation, osteoclastogenesis associated factors and the expression of EphrinB2, EphB4, RANKL and OPG. In vitro, H2O2 stimulation was used to simulate the cell environment in osteoporosis. Immunofluorescence, quantitative real time PCR (qRT-PCR), enzyme-linked immunosorbent assay (ELISA) and Western Blot were applied to detect the expression of EphrinB2, EphB4, RANKL and OPG. In osteoblasts, EphB4 was knocked down by EphB4 small-interfering RNA (siRNA) transfection. LY294002 (PI3K inhibitor) or ARQ092 (AKT inhibitor) was used to block PI3K/AKT pathway. An indirect co-culture system of osteoblasts and osteoclasts was established. The mRNA and protein expression of osteoclastogenes is associated factors were tested by qRT-PCR and Western Blot. Results: ED-71 increased bone mass and decreased the number of osteoclasts in OVX rats. Moreover, ED-71 promoted the expression of EphrinB2, EphB4, and decreased the RANKL/OPG ratio in osteoblasts. Osteoclastogenesis was restrained when osteoclasts were indirectly co-cultured with ED-71-treated osteoblasts. After silencing of EphB4 expression in osteoblasts, ED-71 inhibited the expression of P-PI3K and P-AKT and increased the ratio of RANKL/OPG. This reversed the inhibitory effect of ED-71 on osteoclastogenes. Therefore, in ED-71-inhibited osteoclastogenes, EphB4 is a key factor affecting the secretion of RANKL and OPG by osteoblasts. EphB4 suppressed the RANKL/OPG ratio through activating PI3K/AKT signaling in osteoblasts. Conclusion: ED-71 inhibits osteoclastogenesis through EphrinB2-EphB4-RANKL/OPG axis, improving bone mass in ovariectomized rats. PI3K/AKT pathway is involved this process.

mmu-miR-185 regulates osteoclasts differentiation and migration by targeting Btk.

BACKGROUND: Bones undergo a constant remodeling, a process involving osteoclast-mediated bone resorption and osteoblast-mediated bone formation, crucial for maintaining healthy bone mass. We previously observed that miR-185 depletion may promote bone formation by regulating Bgn expression and the BMP/Smad signaling pathway. However, the effects of miR-185-5p on the osteoclasts and bone remodeling have not been elucidated, warranting further exploration. METHODS: Tartrate-resistant acid phosphatase staining was utilized to assess the differentiation ability of bone marrow mononuclear macrophages (BMMs) from mmu-miR-185 gene knockout (KO) mice and wild-type (WT) mice. A reverse transcriptase-quantitative PCR was conducted to compare differences in miR-185-5p and osteoclast marker molecules, including Trap, Dcstamp, Ctsk and Nfatc1, between the KO group and WT group BMMs. Western blot analysis was employed to observe the expression of osteoclast marker molecules. A cell-counting kit-8 was used to analyze cell proliferation ability. Transwell experiments were conducted to detect cell migration. Dual-luciferase reporter assays were employed to confirm whether Btk is a downstream target gene of miR-185-5p. RESULTS: miR-185 depletion promoted osteoclast differentiation in bone marrow-derived monocytes/macrophages. Overexpression of miR-185-5p in RAW264.7 cells inhibited differentiation and migration of osteoclasts. Furthermore, Btk was identified as a downstream target gene of miR-185-5p, suggesting that miR-185-5p may inhibit osteoclast differentiation and migration by targeting Btk. CONCLUSIONS: miR-185 regulates osteoclasts differentiation, with overexpression of miR-185-5p inhibiting osteoclast differentiation and migration in vitro. Additionally, miR-185-5p may modulate osteoclastic differentiation and migration by regulating Btk expression.

Natural Compounds for Bone Remodeling: A Computational and Experimental Approach Targeting Bone Metabolism-Related Proteins.

Osteoporosis, characterized by reduced bone density and increased fracture risk, affects over 200 million people worldwide, predominantly older adults and postmenopausal women. The disruption of the balance between bone-forming osteoblasts and bone-resorbing osteoclasts underlies osteoporosis pathophysiology. Standard treatment includes lifestyle modifications, calcium and vitamin D supplementation and specific drugs that either inhibit osteoclasts or stimulate osteoblasts. However, these treatments have limitations, including side effects and compliance issues. Natural products have emerged as potential osteoporosis therapeutics, but their mechanisms of action remain poorly understood. In this study, we investigate the efficacy of natural compounds in modulating molecular targets relevant to osteoporosis, focusing on the Mitogen-Activated Protein Kinase (MAPK) pathway and the gut microbiome's influence on bone homeostasis. Using an in silico and in vitro methodology, we have identified quercetin as a promising candidate in modulating MAPK activity, offering a potential therapeutic perspective for osteoporosis treatment.

Pleurotus sajor-caju (Fr.) Singer ß-1,3-Glucanoligosaccharide (Ps-GOS) Suppresses RANKL-Induced Osteoclast Differentiation and Function in Pre-Osteoclastic RAW 264.7 Cells by Inhibiting the RANK/NFκB/cFOS/NFATc1 Signalling Pathway.

Edible grey oyster mushroom, Pleurotus sajor-caju, ß (1,3), (1,6) glucan possesses a wide range of biological activities, including anti-inflammation, anti-microorganism and antioxidant. However, its biological activity is limited by low water solubility resulting from its high molecular weight. Our previous study demonstrated that enzymatic hydrolysis of grey oyster mushroom ß-glucan using Hevea ß-1,3-glucanase isozymes obtains a lower molecular weight and higher water solubility, Pleurotus sajor-caju glucanoligosaccharide (Ps-GOS). Additionally, Ps-GOS potentially reduces osteoporosis by enhancing osteoblast-bone formation, whereas its effect on osteoclast-bone resorption remains unknown. Therefore, our study investigated the modulatory activities and underlying mechanism of Ps-GOS on Receptor activator of nuclear factor kappa-Β ligand (RANKL) -induced osteoclastogenesis in pre-osteoclastic RAW 264.7 cells. Cell cytotoxicity of Ps-GOS on RAW 264.7 cells was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and its effect on osteoclast differentiation was determined by tartrate-resistant acid phosphatase (TRAP) staining. Additionally, its effect on osteoclast bone-resorptive ability was detected by pit formation assay. The osteoclastogenic-related factors were assessed by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), Western blot and immunofluorescence. The results revealed that Ps-GOS was non-toxic and significantly suppressed the formation of mature osteoclast multinucleated cells and their resorption activity by reducing the number of TRAP-positive cells and pit formation areas in a dose-dependent manner. Additionally, Ps-GOS attenuated the nuclear factor kappa light chain-enhancer of activated B cells' P65 (NFκB-P65) expression and their subsequent master osteoclast modulators, including nuclear factor of activated T cell c1 (NFATc1) and Fos proto-oncogene (cFOS) via the NF-κB pathway. Furthermore, Ps-GOS markedly inhibited RANK expression, which serves as an initial transmitter of many osteoclastogenesis-related cascades and inhibited proteolytic enzymes, including TRAP, matrix metallopeptidase 9 (MMP-9) and cathepsin K (CTK). These findings indicate that Ps-GOS could potentially be beneficial as an effective natural agent for bone metabolic disease.

Eldecalcitol Induces Minimodeling-Based Bone Formation and Inhibits Sclerostin Synthesis Preferentially in the Epiphyses Rather than the Metaphyses of the Long Bones in Rats.

This study aimed to examine minimodeling-based bone formation between the epiphyses and metaphyses of the long bones of eldecalcitol (ELD)-administered ovariectomized rats. Sixteen-week-old female rats were divided into four groups: sham-operated rats receiving vehicle (Sham group), ovariectomized (OVX) rats receiving vehicle (Vehicle group), or ELDs (30 or 90 ng/kg BW, respectively; ELD30 and ELD90 groups). ELD administration increased bone volume and trabecular thickness, reducing the number of osteoclasts in both the epiphyses and metaphyses of OVX rats. The Sham and Vehicle groups exhibited mainly remodeling-based bone formation in both regions. The epiphyses of the ELD groups showed a significantly higher frequency of minimodeling-based bone formation than remodeling-based bone formation. In contrast, the metaphyses exhibited significantly more minimodeling-based bone formation in the ELD90 group compared with the ELD30 group. However, there was no significant difference between minimodeling-based bone formation and remodeling-based bone formation in the ELD90 group. While the minimodeling-induced new bone contained few sclerostin-immunoreactive osteocytes, the underlying pre-existing bone harbored many. The percentage of sclerostin-positive osteocytes was significantly reduced in the minimodeling-induced bone in the epiphyses but not in the metaphyses of the ELD groups. Thus, it seems likely that ELD could induce minimodeling-based bone formation in the epiphyses rather than in the metaphyses, and that ELD-driven minimodeling may be associated with the inhibition of sclerostin synthesis.

The immune cells in modulating osteoclast formation and bone metabolism.

Osteoclasts are pivotal in regulating bone metabolism, with immune cells significantly influencing both physiological and pathological processes by modulating osteoclast functions. This is particularly evident in conditions of inflammatory bone resorption, such as rheumatoid arthritis and periodontitis. This review summarizes and comprehensively analyzes the research progress on the regulation of osteoclast formation by immune cells, aiming to unveil the underlying mechanisms and pathways through which diseases, such as rheumatoid arthritis and periodontitis, impact bone metabolism.

Epstein-barr virus infections induce aberrant osteoclastogenesis in immune system-humanized NOD/Shi-scid/IL-2RγCnull mice.

Epstein-Barr virus (EBV) and other viral infections are possible triggers of autoimmune diseases, such as rheumatoid arthritis (RA). To analyze the causative relationship between EBV infections and RA development, we performed experiment on humanized NOD/Shi-scid/IL-2RγCnull (hu-NOG) mice reconstituted human immune system components and infected with EBV. In EBV-infected hu-NOG mice, breakdown of knee joint bones was found to be accompanied by the accumulation of receptor activator of nuclear factor-κB (NF-κB) (RANK) ligand (RANKL), a key factor in osteoclastogenesis, human CD19 and EBV-encoded small RNA (EBER)-bearing cells. Accumulation of these cells expanded in the bone marrow adjacent to the bone breakage, showing a histological feature like to that in bone marrow edema. On the other hand, human RANK/human matrix metalloprotease-9 (MMP-9) positive, osteoclast-like cells were found at broken bone portion of EBV-infected mouse knee joint. In addition, human macrophage-colony stimulating factor (M-CSF), an essential factor in development of osteoclasts, evidently expressed in spleen and bone marrow of EBV-infected humanized mice. Furthermore, RANKL and M-CSF were identified at certain period of EBV-transformed B lymphoblastoid cells (BLBCs) derived from umbilical cord blood lymphocytes. Co-culturing bone marrow cells of hu-NOG mice with EBV-transformed BLBCs resulted in the induction of a multinucleated cell population positive for tartrate-resistant acid phosphatase and human MMP-9 which indicating human osteoclast-like cells. These findings suggest that EBV-infected BLBCs induce human aberrant osteoclastogenesis, which cause erosive arthritis in the joints.

Lactobacillus plantarum 45 activates SHP2 through inhibition of oxidative stress to regulate osteoblast and osteoclast differentiation.

BACKGROUND: The purpose of this study is to observe LP45 (Lactobacillus plantarum 45) to investigate the mechanism by which LP45 attenuates oxidative stress-induced damage and regulates the osteoblast-osteoclast balance. MATERIALS AND METHODS: The oxidative stress level and osteoblast- and osteoclast-related proteins were detected by immunofluorescence staining, Western blotting, ROS fluorescent probe and ELISA. Osteoblast cell proliferation capacity was determined by the CCK-8 assay. X-ray observation and HE staining were used to detect the effect of LP45 on osteoporosis. RESULTS: The expression level of SHP2 and Src was significantly increased, and the expression levels of NOX4, P22, P47, IL-1ß, NLRP3, IRF3, RANK, ß-catenin and INF-ß were inhibited in LP45 group and LPS + LP45 group as compared to those in LPS group. Compared with that in LPS group, the concentration of SOD was increased and the concentration of MDA was decreased in LPS + LP45 group. The protein expressions of OPG, RANKL, RUNX3, RANK and ß-catenin in LP45 group and LPS + LP45 group increased. The protein expressions of NF-κB, CREB and AP-1 in LP45 group and LPS + LP45 group decreased significantly. The results were also confirmed by immunofluorescence staining and ROS fluorescent probe. X-ray observation and HE staining showed that LP45 could inhibit the progression of osteoporosis. CONCLUSION: LP45 can exert its antioxidant effect by inhibiting the production of oxidative stress to activate the SHP2 signaling pathway, thus promoting osteoblast differentiation and repressing osteoclast formation to maintain bone homeostasis and improve bone metabolism.

A 70% Ethanol Neorhodomela munita Extract Attenuates RANKL-Induced Osteoclast Activation and H2O2-Induced Osteoblast Apoptosis In Vitro.

The rapid aging of the population worldwide presents a significant social and economic challenge, particularly due to osteoporotic fractures, primarily resulting from an imbalance between osteoclast-mediated bone resorption and osteoblast-mediated bone formation. While conventional therapies offer benefits, they also present limitations and a range of adverse effects. This study explores the protective impact of Neorhodomela munita ethanol extract (EN) on osteoporosis by modulating critical pathways in osteoclastogenesis and apoptosis. Raw264.7 cells and Saos-2 cells were used for in vitro osteoclast and osteoblast models, respectively. By utilizing various in vitro methods to detect osteoclast differentiation/activation and osteoblast death, it was demonstrated that the EN's potential to inhibit RANKL induced osteoclast formation and activation by targeting the MAPKs-NFATc1/c-Fos pathway and reducing H2O2-induced cell death through the downregulation of apoptotic signals. This study highlights the potential benefits of EN for osteoporosis and suggests that EN is a promising natural alternative to traditional treatments.

[Research Progress in the Regulatory Role of circRNA-miRNA Network in Bone Remodeling]. / circRNA-miRNAç½‘ç»œè°ƒæŽ§éª¨é‡å¡‘çš„ç ”ç©¶è¿›å±•.

The dynamic balance between bone formation and bone resorption is a critical process of bone remodeling. The imbalance of bone formation and bone resorption is closely associated with the occurrence and development of various bone-related diseases. Under both physiological and pathological conditions, non-coding RNAs (ncRNAs) play a crucial regulatory role in protein expression through either inhibiting mRNAs translation or promoting mRNAs degradation. Circular RNAs (circRNAs) are a type of non-linear ncRNAs that can resist the degradation of RNA exonucleases. There is accumulating evidence suggesting that circRNAs and microRNAs (miRNAs) serve as critical regulators of bone remodeling through their direct or indirect regulation of the expression of osteogenesis-related genes. Additionally, recent studies have revealed the involvement of the circRNAs-miRNAs regulatory network in the process by which mesenchymal stem cells (MSCs) differentiate towards the osteoblasts (OB) lineage and the process by which bone marrow-derived macrophages (BMDM) differentiate towards osteoclasts (OC). The circRNA-miRNA network plays an important regulatory role in the osteoblastic-osteoclastic balance of bone remodeling. Therefore, a thorough understanding of the circRNA-miRNA regulatory mechanisms will contribute to a better understanding of the regulatory mechanisms of the balance between osteoblastic and osteoclastic activities in the process of bone remodeling and the diagnosis and treatment of related diseases. Herein, we reviewed the functions of circRNA and microRNA. We also reviewed their roles in and the mechanisms of the circRNA-miRNA regulatory network in the process of bone remodeling. This review provides references and ideas for further research on the regulation of bone remodeling and the prevention and treatment of bone-related diseases.

Current perspectives on the multiple roles of osteoclasts: Mechanisms of osteoclast-osteoblast communication and potential clinical implications.

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.

Systematic review of the osteogenic effect of rare earth nanomaterials and the underlying mechanisms.

Rare earth nanomaterials (RE NMs), which are based on rare earth elements, have emerged as remarkable biomaterials for use in bone regeneration. The effects of RE NMs on osteogenesis, such as promoting the osteogenic differentiation of mesenchymal stem cells, have been investigated. However, the contributions of the properties of RE NMs to bone regeneration and their interactions with various cell types during osteogenesis have not been reviewed. Here, we review the crucial roles of the physicochemical and biological properties of RE NMs and focus on their osteogenic mechanisms. RE NMs directly promote the proliferation, adhesion, migration, and osteogenic differentiation of mesenchymal stem cells. They also increase collagen secretion and mineralization to accelerate osteogenesis. Furthermore, RE NMs inhibit osteoclast formation and regulate the immune environment by modulating macrophages and promote angiogenesis by inducing hypoxia in endothelial cells. These effects create a microenvironment that is conducive to bone formation. This review will help researchers overcome current limitations to take full advantage of the osteogenic benefits of RE NMs and will suggest a potential approach for further osteogenesis research.

[Latest Findings on the Role of RUNX1 in Bone Development and Disorders]. / RUNX1åœ¨éª¨å‘è‚²åŠéª¨ç›¸å ³ç–¾ç— ä¸­çš„ä½œç”¨ç ”ç©¶è¿›å±•.

Runt-related transcription factor (RUNX1) is a transcription factor closely involved in hematopoiesis. RUNX1 gene mutation plays an essential pathogenic role in the initiation and development of hematological tumors, especially in acute myeloid leukemia. Recent studies have shown that RUNX1 is also involved in the regulation of bone development and the pathological progression of bone-related diseases. RUNX1 promotes the differentiation of mesenchymal stem cells into chondrocytes and osteoblasts and modulates the maturation and extracellular matrix formation of chondrocytes. The expression of RUNX1 in mesenchymal stem cells, chondrocytes, and osteoblasts is of great significance for maintaining normal bone development and the mass and quality of bones. RUNX1 also inhibits the differentiation and bone resorptive activities of osteoclasts, which may be influenced by sexual dimorphism. In addition, RUNX1 deficiency contributes to the pathogenesis of osteoarthritis, delayed fracture healing, and osteoporosis, which was revealed by the RUNX1 conditional knockout modeling in mice. However, the roles of RUNX1 in regulating the hypertrophic differentiation of chondrocytes, the sexual dimorphism of activities of osteoclasts, as well as bone loss in diabetes mellitus, senescence, infection, chronic inflammation, etc, are still not fully understood. This review provides a systematic summary of the research progress concerning RUNX1 in the field of bone biology, offering new ideas for using RUNX1 as a potential target for bone related diseases, especially osteoarthritis, delayed fracture healing, and osteoporosis.

The effect of Schizophyllan on the differentiation of osteoclasts and osteoblasts.

Schizophyllan (SPG), a ß-glucan from Schizophyllum commune, is recognized for its antioxidant, immunoregulatory, and anticancer activities. In this study, its effects on bone cells, particularly osteoclasts and osteoblasts, were examined. We demonstrated that SPG dose-dependently inhibited osteoclastogenesis and reduced gene expression associated with osteoclast differentiation. SPG also decreased bone resorption and F-actin ring formation. This inhibition could have been due to the downregulation of transcription factors c-Fos and nuclear factor of activated T cells 1 (NFATc1) via the MAPKs (JNK and p38), IκBα, and PGC1ß/PPARγ pathways. In coculture, SPG lowered osteoclastogenic activity in calvaria-derived osteoblasts by reducing macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL) expression. In addition, SPG slightly enhanced osteoblast differentiation, as evidenced by increased differentiation marker gene expression and alizarin red staining. It also exhibited antiresorptive effects in a lipopolysaccharide-induced calvarial bone loss model. These results indicated a dual role of SPG in bone cell regulation by suppressing osteoclastogenesis and promoting osteoblast differentiation. Thus, SPG could be a therapeutic agent for bone resorption-related diseases such as osteoporosis, rheumatoid arthritis, and periodontitis.

Glycogen Synthase Kinase-3 Beta (GSK3ß) as a Potential Drug Target in Regulating Osteoclastogenesis: An Updated Review on Current Evidence.

Glycogen synthase kinase 3-beta (GSK3ß) is a highly conserved protein kinase originally involved in glucose metabolism, insulin activity, and energy homeostasis. Recent scientific evidence demonstrated the significant role of GSK3ß in regulating bone remodelling through involvement in multiple signalling networks. Specifically, the inhibition of GSK3ß enhances the conversion of osteoclast progenitors into mature osteoclasts. GSK3ß is recognised as a pivotal regulator for the receptor activator of nuclear factor-kappa B (RANK)/receptor activator of nuclear factor-kappa B ligand (RANKL)/osteoprotegerin (OPG), phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT), nuclear factor-kappa B (NF-κB), nuclear factor-erythroid 2-related factor 2 (NRF2)/Kelch-like ECH-associated protein 1 (KEAP1), canonical Wnt/beta (ß)-catenin, and protein kinase C (PKC) signalling pathways during osteoclastogenesis. Conversely, the inhibition of GSK3ß has been shown to prevent bone loss in animal models with complex physiology, suggesting that the role of GSK3ß may be more significant in bone formation than bone resorption. Divergent findings have been reported regarding the efficacy of GSK3ß inhibitors as bone-protecting agents. Some studies demonstrated that GSK3ß inhibitors reduced osteoclast formation, while one study indicated an increase in osteoclast formation in RANKL-stimulated bone marrow macrophages (BMMs). Given the discrepancies observed in the accumulated evidence, further research is warranted, particularly regarding the use of GSK3ß silencing or overexpression models. Such efforts will provide valuable insights into the direct impact of GSK3ß on osteoclastogenesis and bone resorption.

Young osteocyte-derived extracellular vesicles facilitate osteogenesis by transferring tropomyosin-1.

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can undergo inadequate osteogenesis or excessive adipogenesis as they age due to changes in the bone microenvironment, ultimately resulting in decreased bone density and elevated risk of fractures in senile osteoporosis. This study aims to investigate the effects of osteocyte senescence on the bone microenvironment and its influence on BMSCs during aging. RESULTS: Primary osteocytes were isolated from 2-month-old and 16-month-old mice to obtain young osteocyte-derived extracellular vesicles (YO-EVs) and senescent osteocyte-derived EVs (SO-EVs), respectively. YO-EVs were found to significantly increase alkaline phosphatase activity, mineralization deposition, and the expression of osteogenesis-related genes in BMSCs, while SO-EVs promoted BMSC adipogenesis. Neither YO-EVs nor SO-EVs exerted an effect on the osteoclastogenesis of primary macrophages/monocytes. Our constructed transgenic mice, designed to trace osteocyte-derived EV distribution, revealed abundant osteocyte-derived EVs embedded in the bone matrix. Moreover, mature osteoclasts were found to release osteocyte-derived EVs from bone slices, playing a pivotal role in regulating the functions of the surrounding culture medium. Following intravenous injection into young and elderly mouse models, YO-EVs demonstrated a significant enhancement of bone mass and biomechanical strength compared to SO-EVs. Immunostaining of bone sections revealed that YO-EV treatment augmented the number of osteoblasts on the bone surface, while SO-EV treatment promoted adipocyte formation in the bone marrow. Proteomics analysis of YO-EVs and SO-EVs showed that tropomyosin-1 (TPM1) was enriched in YO-EVs, which increased the matrix stiffness of BMSCs, consequently promoting osteogenesis. Specifically, the siRNA-mediated depletion of Tpm1 eliminated pro-osteogenic activity of YO-EVs both in vitro and in vivo. CONCLUSIONS: Our findings suggested that YO-EVs played a crucial role in maintaining the balance between bone resorption and formation, and their pro-osteogenic activity declining with aging. Therefore, YO-EVs and the delivered TPM1 hold potential as therapeutic targets for senile osteoporosis.