Osteoclasts Research Articles

Macrophage Polarization and the Regulation of Bone Immunity in Bone Homeostasis.

Bone homeostasis is a dynamic equilibrium state of bone formation and absorption, ensuring skeletal development and repair. Bone immunity encompasses all aspects of the intersection between the skeletal and immune systems, including various signaling pathways, cytokines, and the crosstalk between immune cells and bone cells under both homeostatic and pathological conditions. Therefore, as key cell types in bone immunity, macrophages can polarize into classical pro-inflammatory M1 macrophages and alternative anti-inflammatory M2 macrophages under the influence of the body environment, participating in the regulation of bone metabolism and playing various roles in bone homeostasis. M1 macrophages can not only act as precursors of osteoclasts (OCs), differentiate into mature OCs, but also secrete pro-inflammatory cytokines to promote bone resorption; while M2 macrophages secrete osteogenic factors, stimulating the differentiation and mineralization of osteoblast precursors and mesenchymal stem cells (MSCs), and subsequently increase bone formation. Once the polarization of macrophages is imbalanced, the resulting immune dysregulation will cause inflammatory stimulation, and release a large amount of inflammatory factors affecting bone metabolism, leading to pathological conditions such as osteoporosis (OP), rheumatoid arthritis (RA), and steroid-induced femoral head necrosis (SANFH). In this review, we introduce the signaling pathways and related factors of macrophage polarization, as well as their relationships with immune factors, OB, OC, and MSC. We also discuss the roles of macrophage polarization and bone immunity in various diseases of bone homeostasis imbalance, as well as the factors regulating them, which may help to develop new methods for treating bone metabolic disorders.

Eicosapentaenoic acid supplementation modulates the osteoblast/osteoclast balance in inflammatory environments and protects against estrogen deficiency-induced bone loss in mice

An imbalance of osteoblasts (OBs) and osteoclasts (OCs) in a chronic inflammatory microenvironment is an important pathological factor leading to osteoporosis. Eicosapentaenoic acid (EPA) has been shown to suppress inflammation in macrophages and adipocytes. However, the effect of EPA on OBs and OCs has yet to be fully elucidated. We explored the roles of EPA in the differentiation of OBs and OCs, as well as the coupling between OBs and OCs in an inflammatory microenvironment. The effects of EPA on estrogen deficiency-induced osteoporosis were also evaluated. Mouse bone marrow mesenchymal stem cells (mBMSCs) and mouse bone marrow-derived macrophages (mBMMs) were used for invitro OBs and OCs differentiation. TNF-α was used to create an inflammatory microenvironment. We examined the effects of EPA on osteoblastogenesis in the absence or presence of TNF-α and collect OBs' culture medium as the conditioned medium (CM). Then we examined the effects of EPA and CM on RANKL-induced osteoclastogenesis. The invivo effects of EPA were determined using an ovariectomized (OVX) mouse model treated with EPA or vehicle. High-dose EPA was shown to promote osteoblastogenesis in an inflammatory environment invitro, as well as upregulate expression of OBs-specific proteins and genes. ARS and ALP staining also showed that high-dose EPA-treated groups restored mBMSCs' impaired osteogenic capacity caused by TNFa. Mechanistically, EPA suppressed the NF-κB pathway activated by TNF-α in mBMSCs and rescued TNF-α-mediated inhibition of osteoblastogenesis. EPA was also shown to inhibit expression of RANKL and decrease the RANKL/OPG ratio in OBs in an inflammatory environment. CM from TNF-α-stimulated OBs promoted osteoclastogenesis of mBMMs; EPA-treated CM prevented this. In the OVX mouse model, EPA supplementation prevented bone loss in an estrogen deficiency-induced inflammatory environment. EPA was demonstrated for the first time to restore mBMSCs' impaired osteogenic capacity caused by TNFa-induced inflammation and rescue the OBs/OCs balance via regulation of RANKL and OPG expression in OBs. EPA showed a remarkable ability to prevent bone loss in OVX mice, suggesting a potential application of EPA in postmenopausal osteoporosis.

Abr, a Rho-regulating protein, modulates osteoclastogenesis by enhancing lamellipodia formation by interacting with poly(ADP-ribose) glycohydrolase.

Osteoclasts are multinucleated bone-resorbing cells formed by the fusion of monocyte/macrophage lineage. During osteoclast differentiation, Rho GTPases are involved in various processes, including cell migration, adhesion, and polarity. However, the role of Rho-regulatory molecules in the regulation of osteoclast differentiation remains unclear. In this study, among these genes, we focused on active breakpoint cluster region-related (Abr) protein that is a multifunctional regulator of Rho GTPases. We examined using knockdown and overexpression experiments in RANKL-stimulated RAW-D macrophages whether Abr regulates osteoclast differentiation and cell morphology. We observed an increase in Abr expression during osteoclast differentiation and identified expression of a variant of the Abr gene in osteoclasts. Knockdown of Abr suppressed osteoclast differentiation and resorption. Abr knockdown markedly inhibited the expression of osteoclast markers, such as Nfatc1, c-fos, Src, and Ctsk in osteoclasts. Conversely, overexpression of Abr enhanced the formation of multinucleated osteoclasts, bone resorption activity, and osteoclast marker gene expression. Moreover, Abr overexpression accelerated lamellipodia formation and induced the formation of well-developed actin in osteoclasts. Importantly, the Abr protein interacted with poly(ADP-ribose) glycohydrolase (PARG) and Rho GTPases, including RhoA, Rac1/2/3, and Cdc42 in osteoclasts. Taken together, these results indicate that Abr modulates osteoclastogenesis by enhancing lamellipodia formation via its interaction with PARG.

Osteoclast-derived IGF1 induces RANKL production in osteocytes and contributes to pagetic lesion formation.

We previously reported that measles virus nucleocapsid protein (MVNP) expression in osteoclasts (OCLs) of patients with Paget disease (PD) or targeted to the OCL lineage in MVNP-transgenic mice (MVNP mice) increases IGF1 production in osteoclasts (OCL-IGF1) and leads to development of PD OCLs and pagetic bone lesions (PDLs). Conditional deletion of Igf1 in OCLs of MVNP mice fully blocked development of PDLs. In this study, we examined whether osteocytes (OCys), key regulators of normal bone remodeling, contribute to PD. OCys in PDLs of patients and of MVNP mice expressed less sclerostin, and had increased RANKL expression compared with OCys in bones from WT mice or normal patients. To test whether increased OCL-IGF1 is sufficient to induce PDLs and PD phenotypes, we generated TRAP-Igf1 (T-Igf1) transgenic mice to determine whether increased IGF1 expression in the absence of MVNP in OCLs is sufficient to induce PDLs and pagetic OCLs. We found that T-Igf1 mice at 16 months of age developed PD OCLs, PDLs, and OCys, with decreased sclerostin and increased RANKL, similar to MVNP mice. Thus, pagetic phenotypes could be induced by OCLs expressing increased IGF1. OCL-IGF1 in turn increased RANKL production in OCys to induce PD OCLs and PDLs.

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

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

Clinical masks of marble disease

Marble disease, or osteopetrosis (OPT), is rare in the practice of a rheumatologist, internist or pediatrician. This group of hereditary diseases is based on a defect in the formation, development and functioning of osteoclasts (OCL), which leads to disruption of the processes of resorption and remodeling of bone tissue. Disturbance of resorption leads to increased density and changes in the quality of the bones, as a result of which they become more fragile. As a rule, the disease manifests with pathological fractures. In recent decades, 70% of patients with OPT have been found to have mutations in at least 10 genes that lead to impaired functioning of the OCL. Depending on the variant of inheritance, autosomal dominant, autosomal recessive and intermediate types of OPT are distinguished. Autosomal dominant OPT has a benign course that can be asymptomatic or characterized by multiple bone fractures and other spinal anomalies. The disease usually manifests in adulthood or adolescence. Life expectancy in patients of this group does not differ from that in the general population. Malignant, or infantile, OPT is associated with an autosomal recessive inheritance pattern. Its clinical manifestations are observed from the moment of birth, without treatment, patients die within the first decade of life. In such patients, in addition to the skeletal pathology, there is involvement of the hematopoietic system, compression of the cranial nerves and their function disturbance.The article presents a clinical case of autosomal dominant OPT diagnosed in adulthood (at the age of 38), when the patient referred to the doctor for the first time. Differential diagnosis with ankylosing spondylitis and paraneoplastic spondyloarthritis was performed.

Mechanotransduction in osteoclasts: Novel strategies of bone repairs

Mechanotransduction, the transfer of mechanical stimuli into various biological signals, is a vital biological process in multiple organ systems. The osteoclast (OC) plays a vital role in bone metabolism and repair. The role of mechanotransduction in osteoclasts and other bone cells is emerging. This commentary highlights a recent research report on a novel strategy for the precise regulation of OC formation via modulating matrix stiffness. Modulation of the mechanotransduction pathways in the skeletal system will pave the way for the development of a matrix stiffness-based strategy for bone tissue regeneration.

In Vitro and In Vivo Bone Regeneration Assessment of Titanium-Doped Waste Eggshell-Derived Hydroxyapatite in the Animal Model.

In this work, a titanium-doped hydroxyapatite (HAp) scaffold was produced from two different sources (natural eggshell and laboratory-grade reagents) to compare the efficacy of natural and synthetic resources of HAp materials on new bone regeneration. This comparative study also reports the effect of Ti doping on the physical, mechanical, and in vitro as well as in vivo biological properties of the HAp scaffold. Pellets were prepared in the conventional powder metallurgy route, compacted, and sintered at 900 °C, showing sufficient porosity for bony ingrowth. The physical-mechanical characterizations were performed by density, porosity evaluation, XRD, FTIR, SEM analysis, and hardness measurement. In vitro interactions were evaluated by bactericidal assay, hemolysis, MTT assay, and interaction with simulated body fluid. All categories of pellets showed absolute nonhemolytic and nontoxic character. Furthermore, significant apatite formation was observed on the Ti-doped HAp samples in the simulated body fluid immersion study. The developed porous pellets were implanted to assess the bone defect healing in the femoral condyle of healthy rabbits. A 2 month study after implantation showed no marked inflammatory reaction for any samples. Radiological analysis, histological analysis, SEM analysis, and oxytetracycline labeling studies depicted better invasion of mature osseous tissue in the pores of doped eggshell-derived HAp scaffolds as compared to the undoped HAp, and laboratory-made samples. Quantification using oxytetracycline labeling depicted 59.31 ± 1.89% new bone formation for Ti-doped eggshell HAp as compared to Ti-doped pure HAp (54.41 ± 1.93) and other undoped samples. Histological studies showed the presence of abundant osteoblastic and osteoclastic cells in Ti-doped eggshell HAp in contrast to other samples. Radiological and SEM data also showed similar results. The results indicated that Ti-doped biosourced HAp samples have good biocompatibility, new bone-forming ability, and could be used as a bone grafting material in orthopedic surgery.

Histopathological assessment of the preventive effect of leukocyte-platelet-rich fibrin on bisphosphonate-related osteonecrosis of the jaw following dental extraction: An animal study

BackgroundLeukocyte- and platelet-rich fibrin (L-PRF) could be considered a preventive measure in Bisphosphonate-Related Osteonecrosis of the Jaw (BRONJ). The present experiment aimed to assess the preventive effects of L-PRF on osteonecrosis of the jaw in rats. MethodsIn this interventional animal study with a split-mouth design, 28 rats were randomly allocated to saline (negative control), bisphosphonate (positive control), and Bis + L-PRF (case) groups. Bilateral extraction of maxillary molar teeth was performed followed by random application of L-PRF to one of the extraction sockets treated with Zoledronic acid for four weeks. Clinical occurrence of BRONJ and histopathologic evaluations were done, and data were subjected to the Kruskal-Wallis test, Mann-Whitney U test and exact Fisher test performed using SPSS 25. The significance level was set at 0.05. ResultsThe application of L-PRF resulted in a 41.67% reduction in osteonecrosis centers and the number of osteoclast cells. Also, Kruskal Wallis test results showed a significant difference among the three groups regarding the frequency distribution of inflammation severity. However, no significant difference was detected regarding the frequency distribution of the blood vessels (Kruskal Wallis test, P-value = 0.649). ConclusionIt could be inferred that possible preventive effects on the clinical occurrence of osteonecrosis could be expected from the application of L-PRF.

Mangosteen peel extract (mpe) and dfdbbx applied to cavia cobaya tooth extraction sockets for guided bone regeneration

Objectives: The objective of this study was to examine the clinical and microbiological effects of combining mangosteen peel extract (MPE) and demineralized freeze-dried bovine bone xenograft (DFDBBX) on the expression of fibroblast growth factor-2 (FGF-2), the amount of osteoblast, and the amount of osteoclast in the alveolar bone following Cavia cobaya tooth extraction. Tooth extraction followed by alveolar ridge resorption is an unavoidable physiological process. The combination of MPE and DFDBBX is expected to primarily provide osteoconductive properties as well as strengthen the graft in order to stimulate new bone formation. Materials and Methods: Fifty-six Cavia cobaya were divided into two groups for 7-day (group A) and 30-day (group B) examinations, respectively. Each group consisted of twenty-eight animals that received four treatments. In each treatment, seven Cavia cobaya were used. The bottom right incisor was extracted. In groups AI and BI, tooth sockets were filled with 25 grams of polyethylene glycol (PEG). Tooth sockets in groups AII and BII were filled with 0.5 grams of DFDBBX and 24.5 grams of PEG. Tooth sockets in groups AIII and BIII were filled with 0.5 grams of MPE and 24.5 grams of PEG. In groups AIV and BIV, tooth sockets were filled with a mixture of 0.5 grams of DFDBBX, 0.5 grams of MPE, and 24 grams of PEG. Cavia cobaya's mandible was decalcified with ethylene diamine tetraacetic acid (EDTA) at 7 and 30 days. During this time, histopathology (HPA) and immunohistochemistry (IHC) tests were also performed. Statistical analysis: The calculated amounts of osteoblasts, osteoclasts, and FGF-2 expression data were used in the one-way ANOVA test. Results: FGF2 expression and the number of osteoblasts increased significantly after 7-day and 30-day of examination. On the other hand, the number of osteoclasts decreased. Conclusion: The combination of MPE and DFDBBX effectively increases FGF-2 expression and the number of osteoblast cells while decreasing the number of osteoclast cells on the alveolar ridge of Cavia cobaya.

Aneurysmal bone cyst of dorsal spine presenting with paraparesis in a pediatric patient: a case report

Introduction: Aneurysmal bone cyst (ABC) are rare benign tumours of spine. They most commonly presents with backache. However neurological deficit in spinal ABCs are not uncommon. Magnetic resonance imaging of ABC may sometimes be confused with osteosarcoma or Giant cell tumours. Treatment options are varied but complete excision of tumour with spine stabilisation should be the goal. Clinical case: Here we presented the case of a 13 year old girl presenting with spastic paraparesis and bladder incontinence with MRI dorsal spine suggestive of an expansile lesion of D5 body. She underwent subtotal corpectomy with fixation of spine . Biopsy was suggestive of aneurysmal bone cyst. Post operative neurological outcome was excellent and followup scans over 2 year period suggest complete remission. Discussion: ABC are non neoplastic expansile bone lesion consisting of blood filled spaces separated by connective tissue septa of bony trabeculation and osteoclastic giant cells. Similar to our case it is seen more commonly in young females. Lumbar spine has higher incidence of ABC as compared to dorsal and cervical spine. Most common presentation is backache. Surgical options for ABC include intralesional curettage, en bloc resection followed by fixation. Vertebroplasty is also a treatment option to reinforce spinal stability. Post operative radiotherapy also can be considered in partially resected cases. Although ABC have high recurrence rate of 10-44% which has been reported with 2 years following surgery. However our patients continue to have no radiological evidence of recurrence on 2 year follow up. Conclusion: ABCs are benign tumour which may present with neurological deficit. Although treatment of ABC remains controversial but early surgery with complete removal of tumour in patients with neurological deficit provides excellent recovery. However due to high recurrence rate regular follow up is necessary.

Fanca deficiency is associated with alterations in osteoclastogenesis that are rescued by TNFα

BackgroundHematopoietic stem cells (HSCs) reside in the bone marrow (BM) niche, which includes bone-forming and bone-resorbing cells, i.e., osteoblasts (OBs) and osteoclasts (OCs). OBs originate from mesenchymal progenitors, while OCs are derived from HSCs. Self-renewal, proliferation and differentiation of HSCs are under the control of regulatory signals generated by OBs and OCs within the BM niche. Consequently, OBs and OCs control both bone physiology and hematopoiesis. Since the human developmental and bone marrow failure genetic syndrome fanconi anemia (FA) presents with skeletal abnormalities, osteoporosis and HSC impairment, we wanted to test the hypothesis that the main pathological abnormalities of FA could be related to a defect in OC physiology and/or in bone homeostasis.ResultsWe revealed here that the intrinsic differentiation of OCs from a Fanca−/− mouse is impaired in vitro due to overactivation of the p53–p21 axis and defects in NF-kB signaling. The OC differentiation abnormalities observed in vitro were rescued by treating Fanca−/− cells with the p53 inhibitor pifithrin-α, by treatment with the proinflammatory cytokine TNFα or by coculturing them with Fanca-proficient or Fanca-deficient osteoblastic cells.ConclusionsOverall, our results highlight an unappreciated role of Fanca in OC differentiation that is potentially circumvented in vivo by the presence of OBs and TNFα in the BM niche.

Graded levels of Eimeria challenge altered the microstructural architecture and reduced the cortical bone growth of femur of Hy-Line W-36 pullets at early stage of growth (0–6 wk of age)

An experiment was carried out to evaluate the impact of mixed Eimeria challenge on skeletal health of Hy-Line W-36 pullets. A total of 540, 16-day-old pullets were randomly allocated into 5 treatment groups, including a nonchallenged control. A mixed Eimeria species solution containing 50,000E. maxima, 50,000E. tenella, and 250,000E. acervulinaoocysts per mL was prepared and challenged to 1 group as a high-dose treatment. The 2-fold serial dilution was done to prepare the medium-high (25,000E. maxima; 25,000E. tenella; 125,000E. acervulina), the medium-low (12,500E. maxima; 12,500E. tenella; 62,500E. acervulina), and the low (6,250E. maxima; 6,250E. tenella; 31,250E. acervulina) dose treatments which were challenged to 3 corresponding groups, respectively. The mineral apposition rate (MAR) was measured from 0 to 14 d post inoculation (DPI) and 14 to 28 DPI using calcein injection. The microstructural architecture of the femur was analyzed using the Skyscan X-ray microtomography (microCT) on 6, 14, and 28 DPI. The results showed that the MAR decreased linearly with an increase in the challenged dose (P < 0.05) during 0 to 14 DPI. The results of microCT revealed that cortical and total BMD, BMC, bone volume (BV), and bone volume as a fraction of tissue volume (BV/TV) of femur decreased both linearly (P < 0.05). Conversely, the total number of pores increased linearly with an increase in challenge dosages on 6 and 14 DPI. Trabecular BMD, BV, BV/TV, trabecular number, and trabecular thickness decreased linearly with an increase in the challenge dosages (P < 0.05) on 6 DPI. Furthermore, Eimeria infection significantly increased the number of osteoclasts and osteoclastic activity (P=0.001). The result of this study suggests that the mixed Eimeria challenge negatively impacts the quality of skeletal health in a linear or quadratic manner with an increase in the concentration of Eimeria oocysts. The negative impact on long bone development might be due to malabsorption, nutrient deficiency during the infection, along with oxidative stress/inflammation disrupting the balance of osteoblastic and osteoclastic cells and their functions.

Staphylococci planktonic and biofilm environments differentially affect osteoclast formation

IntroductionThe pathophysiology of chronic implant-related bone infections is characterized by an increase in osteoclast numbers and enhanced bone resorption. Biofilms are a major reason for chronicity of such infections as the biofilm matrix protects bacteria against antibiotics and impairs the function of immune cells. Macrophages are osteoclast precursor cells and therefore linked to inflammation and bone destruction.Objective and methodInvestigations on the impact of biofilms on the ability of macrophages to form osteoclasts are yet missing and we, therefore, analyzed the effect of Staphylococcus aureus (SA) and Staphylococcus epidermidis (SE) planktonic and biofilm environments on osteoclastogenesis using RAW 264.7 cells and conditioned media (CM).ResultsPriming with the osteoclastogenic cytokine RANKL before CM addition enabled the cells to differentiate into osteoclasts. This effect was highest in SE planktonic or SA biofilm CM. Simultaneous stimulation with CM and RANKL, however, suppressed osteoclast formation and resulted in formation of inflammation-associated multinucleated giant cells (MGCs) which was most pronounced in SE planktonic CM.ConclusionOur data indicate that the biofilm environment and its high lactate levels are not actively promoting osteoclastogenesis. Hence, the inflammatory immune response against planktonic bacterial factors through Toll-like receptors seems to be the central cause for the pathological osteoclast formation. Therefore, immune stimulation or approaches that aim at biofilm disruption need to consider that this might result in enhanced inflammation-mediated bone destruction.

Inhibition of Osteoclast Differentiation and Promotion of Osteogenic Formation by Wolfiporia extensa Mycelium.

Osteoporosis, Greek for "porous bone," is a bone disease characterized by a decrease in bone strength, microarchitectural changes in the bone tissues, and an increased risk of fracture. An imbalance of bone resorption and bone formation may lead to chronic metabolic diseases such as osteoporosis. Wolfiporia extensa, known as "Bokryung" in Korea, is a fungus belonging to the family Polyporaceae and has been used as a therapeutic food against various diseases. Medicinal mushrooms, mycelium and fungi, possess approximately 130 medicinal functions, including antitumor, immunomodulating, antibacterial, hepatoprotective, and antidiabetic effects, and are therefore used to improve human health. In this study, we used osteoclast and osteoblast cell cultures treated with Wolfiporia extensa mycelium water extract (WEMWE) and investigated the effect of the fungus on bone homeostasis. Subsequently, we assessed its capacity to modulate both osteoblast and osteoclast differentiation by performing osteogenic and anti-osteoclastogenic activity assays. We observed that WEMWE increased BMP-2-stimulated osteogenesis by inducing Smad-Runx2 signal pathway axis. In addition, we found that WEMWE decreased RANKL-induced osteoclastogenesis by blocking c-Fos/NFATc1 via the inhibition of ERK and JNK phosphorylation. Our results show that WEMWE can prevent and treat bone metabolic diseases, including osteoporosis, by a biphasic activity that sustains bone homeostasis. Therefore, we suggest that WEMWE can be used as a preventive and therapeutic drug.

A single-institution analysis of a rare variant of pancreatic neoplasm-osteoclastic giant cell tumor.

e16315 Background: Pancreatic osteoclastic giant cell tumor (OGCT) is a rare variant (&lt;1% cases) of pancreatic neoplasm. Due to its rarity, prospective studies on management are limited. The prognosis and molecular characteristics may differ from pancreatic adenocarcinoma. Methods: We analyzed demographics, clinical features, treatment modalities, and outcomes of overall survival (OS) and progression-free survival (PFS) in patients with pancreatic OGCT at our institution from 2005-2022. Results: We identified 8 OGCT patients with a median age of 67, of whom 7 had locoregional disease (resectable- 5, borderline resectable- 1, locally advanced-1), and 1 had denovo metastatic disease at diagnosis. The borderline resectable and locally advanced patients eventually developed metastatic disease. The majority of patients were male (n=5), Caucasian (n=7), former smokers (n=5), had no significant alcohol use (n=8) and had no pre-existing pancreatitis (n=8). 4 of the 5 resectable patients had surgery, and 2 received adjuvant chemotherapy. The median OS was 54 months for locoregional disease (n=5) vs.16 months for metastatic disease (denovo metastatic and progression to metastatic disease, n=3). The median OS was 54 months for surgical resection (n=4) vs. 25 months for those without surgical resection (n=4) based on quartile computations from the Kaplan-Meier survival curve. Conclusions: Our patients had a longer OS than the previously reported OS of less than 12 months for all stages of pancreatic OGCT. There was no progression in resectable patients, indicating an indolent disease with an overall good prognosis. Patients with locoregional disease or those who underwent surgical resection had longer OS, but due to the small sample size and censored patient survival times (i.e., patients still alive), the statistical significance of this trend could not be calculated. Studies are required to understand tumor biology and determine the optimal multidisciplinary treatment. [Table: see text]

Unravelling the Relacatib activity against the CTSK proteins causing pycnodysostosis: a molecular docking and dynamics approach

Pycnodysostosis is an atypical autosomal recessive condition of Lysosomal storage disorder that originated due to the deficit of the enzyme Cathepsin K which is vital for normal osteoclast action in bone resorption. Abnormal degradation of type 1 collagen and accumulation of toxic undigested collagen fibers in lysosomes of the osteoclast cells resulting in high bone density, brittle bones, and a short stature is caused in CTSK protein-carrying individuals. The broad aim of this study is to identify the most significant variant through various computational pipelines. This study was initiated by retrieving a total number of thirty-six variants from NCBI, HGMD, and UniProt databases, and the Y283C variant was found to be more significant by various standard computational tools. A structural investigation was performed to understand and gain a better knowledge about the interaction profile for the native (1BY8) and variant (Y283C) with Relacatib (a small-molecule drug that blocks the function of Cathepsin K, an enzyme that has been linked to osteoporosis, osteoarthritis, and other bone-degrading diseases). The interaction profile was analyzed using molecular docking. Relacatib (ligand) had an average binding affinity for both native (-7.16 kcal/mol) and Y283C (-6.76 kcal/mol). Finally, Molecular dynamics simulations were done in duplicates to recognize the variant (Y283C) activity of the protein structure against Relacatib for 100 ns. This study assists in comprehending the most pathogenic amino-acid variant, the ligand interaction with the protein structure, and paves the way for understanding the steadiness of the ligand with the native and selected significant amino-acid variant. Communicated by Ramaswamy H. Sarma

DCIR suppresses osteoclastic proliferation and resorption by downregulating M-CSF and RANKL signaling.

Dendritic cell immunoreceptor (DCIR) is an inhibitory C-type lectin receptor that acts as a negative regulator in the immune system and bone metabolism. We previously revealed that DCIR deficiency enhanced osteoclastogenesis and antigen presentation of dendritic cells, and that asialo-biantennary N-glycan (NA2) functions as a ligand for DCIR. NA2 binding to DCIR suppressed murine and human osteoclastogenesis that occurs in the presence of M-CSF and RANKL. The DCIR-NA2 axis, therefore, plays an important role in regulating osteoclastogenesis in both mice and humans, although the underlying mechanisms remain unclear. Here we found that Dcir -/- bone marrow-derived macrophages (BMMs) exhibited greater proliferative and differentiation responses to M-CSF and RANKL, respectively, than wild-type (WT) BMMs. Moreover, Dcir -/- osteoclasts (OCs) increased resorptive activity and cell fusion more significantly than WT OCs. DCIR deficiency affects gene expression patterns in OCs, and we found that the expression of neuraminidase 4 was increased in Dcir -/- OCs. Furthermore, DCIR-NA2 interaction in WT BMMs, but not Dcir -/- BMMs, decreased Akt phosphorylation in response to M-CSF and RANKL. These data suggest that DCIR regulates osteoclastogenesis by downregulating M-CSF and RANKL signaling, and that DCIR-mediated signaling may contribute to the terminal modification of oligosaccharides by controlling the expression of glycosylation enzymes.

澱粉補強リン酸カルシウム骨ペーストへの異種カチオン種の導入

Bone cement is applied to bone detect caused by injury and disease. Our group has developed a novel bone paste by kneading a gelatinized wheat starch and calcium phosphate (CP) powders in which CaHPO4, Ca4(PO4)2O and β-Ca3(PO4)2 are equimolarly mixed. This bone paste is easily handled and can be set to form hydroxyapatite (HAp) under physiological conditions. On the other hand, previous literature reported that strontium ion can promote and regulate the activity of osteoblastic and osteoclastic cells in the living bone, respectively. In addition, copper ion is reported antibacterial performance. In this work, strontium and copper ions were fed into the bone paste as starting materials to promote cell activity as well as antibacterial properties. Paste samples were prepared by kneading a starch gelatinized with mixed powders of CP, α-SrHPO4 and Cu2+-substituted β-Ca3(PO4)2. Phase change, release behavior of metal cations and antibacterial properties were investigated. A paste sample immersed in Tris-buffered NaCl solution was set through forming HAp, and the amount of strontium ion released from the paste sample was a level expected to enhance the cell activity. However, all the pastes didn’t show antibacterial properties in spite of introducing of copper ion to the samples.

Interaction gene set between osteoclasts and regulatory CD4+ T cells can accurately predict the prognosis of patients with osteosarcoma.

Osteoclasts (OCs) and regulatory CD4+ T cells (CD4+ Tregs) are important components in the tumor microenvironment (TME) of osteosarcoma. In this study, we collected six osteosarcoma samples from our previous study (GSE162454). We also integrated a public database (GSE152048), which included single cell sequencing data of 11 osteosarcoma patients. We obtained 138,192 cells and then successfully identified OCs and CD4+ Tregs. Based on the interaction gene set between OCs and CD4+ Tregs, patients from GSE21257 were distinguished into two clusters by consensus clustering analysis. Both the tumor immune microenvironment and survival prognosis between the two clusters were significantly different. Subsequently, five model genes were identified by protein-protein interaction network based on differentially upregulated genes of cluster 2. Quantitative RT-PCR was used to detect their expression in human osteoblast and osteosarcoma cells. A prognostic model was successfully established using these five genes. Kaplan-Meier survival analysis found that patients in the high-risk group had worse survival (p = 0.029). Therefore, our study first found that cell-cell communication between OCs and CD4+ Tregs significantly alters TME and is connected to poor prognosis of OS. The model we constructed can accurately predict prognosis for osteosarcoma patients.