Active Bone Formation Research Articles

Clinical Utility of Romosozumab in the Management of Osteoporosis: Focus on Patient Selection and Perspectives.

As one of the most potent osteoanabolic agents with a unique mechanism of action, romosozumab has high efficacy for osteoporosis treatment. It is a monoclonal antibody against sclerostin, a natural inhibitor of the Wnt signaling pathway, and by inhibiting sclerostin, activation of Wnt signaling occurs with a cascade of changes ultimately leading to bone mineral density (BMD) gains. Romosozumab stimulates bone modeling and has a dual effect of activating bone formation while inhibiting bone resorption. With this unique mechanism of action, treatment with romosozumab leads to a rapid and significant gain in BMD; these gains are higher than seen with bisphosphonates, denosumab, or parathyroid hormone (PTH) analogs. The FRAME and ARCH studies represent two pivotal trials demonstrating the efficacy of romosozumab in treating osteoporosis. Treatment with romosozumab should be followed by an antiresorptive agent, as this approach has demonstrated maintenance of or greater increases in BMD and reduced fracture risk even after finishing romosozumab treatment. As an osteoanabolic agent, romosozumab has shown superiority to alendronate in reducing fracture risk, increasing bone density, and potentially more rapid fracture risk reduction. Recent data have suggested that romosozumab prior to antiresorptive therapy may be the ideal treatment sequence, especially in high-risk patients and patients at imminent risk of fracture. Carrying a black box warning, romosozumab should be avoided in patients who have had myocardial infarction or stroke in the past year. Further studies are needed to clarify the increased cardiovascular risk attributed to this drug. Romosozumab has expanded our osteoporosis armamentarium and has enabled novel approaches, including "treat to target." Future studies are needed to evaluate the optimal use sequence and to assess its safety, especially in patients with cardiovascular risk factors.

Synergistic effect of cerium and structure directing agent on drug release behavior and kinetics

This work was mainly aimed at the study of the dual effect of cerium and structure directing agent template concentration, on vancomycin release profiles and kinetics from Ce-substituted mesoporous bioactive glasses (MBGs). MBG based on (20-x) B2O3 - 55 SiO2 – 20 CaO – 5 P2O5 – x Ce2O (x = 0, 1 and 3 mol %) was synthesized by the evaporation-induced self-assembly process using two molar ratios (0.01 and 0.02 molar ratio) of nonionic block copolymer Pluronic® 123 (P123) surfactant. The TGA-DTA, FTIR, and textural features analyses were carried out for the glasses. Moreover, the in vitro bone-forming activity and degradation analysis were tested using simulated body fluid (SBF). The drug loading capacity, drug release profile, and kinetics (using different kinetic models such as first order, Higuchi, Hixson-Crowell, and Baker-Lonsdale models) were determined using vancomycin as a drug model. The results showed that the isotherms of all MBGs fit with type IV isotherms, and the surface area of MBGs synthesized by 0.02 M template was higher than that prepared by 0.01 M, where it ranged from 174.05 m2.g−1 to 256.73 m2.g−1. The pore size diameter was decreased as cerium content increased in all MBGs (decreased from 5.44 to 3.54 nm). Moreover, the MBGs induced the formation of a bone-like apatite layer, and their biodegradation properties can be tailored by controlling glass composition. Furthermore, Ce-free MBGs showed the lowest drug adsorption and the highest drug release percentage. The drug release kinetic was best fitted with Higuchi and Baker-Lonsdale models which denoted that the mechanism of drug release from MBGs was a diffusion release from spherical particles. In conclusion, vancomycin release was controlled by the glass composition. Meanwhile, the MBGs synthesized in this study are proposed to be applied for bone regeneration, bone cancer treatment, and reducing the bacterial activity around the tumor site.Graphical abstract

Suffruticosol B Is an Osteogenic Inducer through Osteoblast Differentiation, Autophagy, Adhesion, and Migration.

Suffruticosol B (Suf-B) is a stilbene found in Paeonia suffruticosa ANDR., which has been traditionally used in medicine. Stilbenes and their derivatives possess various pharmacological effects, such as anticancer, anti-inflammatory, and anti-osteoporotic activities. This study aimed to explore the bone-forming activities and mechanisms of Suf-B in pre-osteoblasts. Herein, >99.9% pure Suf-B was isolated from P. suffruticosa methanolic extracts. High concentrations of Suf-B were cytotoxic, whereas low concentrations did not affect cytotoxicity in pre-osteoblasts. Under zero levels of cytotoxicity, Suf-B exhibited bone-forming abilities by enhancing alkaline phosphatase enzyme activities, bone matrix calcification, and expression levels with non-collagenous proteins. Suf-B induces intracellular signal transduction, leading to nuclear RUNX2 expression. Suf-B-stimulated differentiation showed increases in autophagy proteins and autophagosomes, as well as enhancement of osteoblast adhesion and transmigration on the ECM. These results indicate that Suf-B has osteogenic qualities related to differentiation, autophagy, adhesion, and migration. This also suggests that Suf-B could have a therapeutic effect as a phytomedicine in skeletal disorders.

ED-71 Prevents Glucocorticoid-Induced Osteoporosis by Regulating Osteoblast Differentiation via Notch and Wnt/β-Catenin Pathways.

Long-term glucocorticoid- usage can lead to glucocorticoid-induced osteoporosis (GIOP). The study focused on the preventative effects of a novel active vitamin D3 analog, eldecalcitol (ED-71), against GIOP and explored the underlying molecular mechanisms. Intraperitoneal injection of methylprednisolone (MPED) or dexamethasone (DEX) induced the GIOP model within C57BL/6 mice in vivo. Simultaneously, ED-71 was orally supplemented. Bone histological alterations, microstructure parameters, novel bone formation rates, and osteogenic factor changes were evaluated by hematoxylin-eosin (HE) staining, micro-computed tomography, calcein/tetracycline labeling, and immunohistochemical (IHC) staining. The osteogenic differentiation level and mineralization in pre-osteoblast MC3T3-E1 cells were evaluated in vitro using alkaline phosphatase (ALP) staining, alizarin red (AR) staining, quantitative polymerase chain reaction (qPCR), Western blotting, and immunofluorescence staining. ED-71 partially prevented bone mass reduction and microstructure parameter alterations among GIOP-induced mice. Moreover, ED-71 also promoted new bone formation and osteoblast activity while inhibiting osteoclasts. In vitro, ED-71 promoted osteogenic differentiation and mineralization in DEX-treated MC3T3-E1 cells and boosted the levels of osteogenic-related factors. Additionally, GSK3-β and β-catenin expression levels were elevated after ED-71 was added to cells and were accompanied by reduced Notch expression. The Wnt signaling inhibitor XAV939 and Notch overexpression reversed the ED-71 promotional effects toward osteogenic differentiation and mineralization. ED-71 prevented GIOP by enhancing osteogenic differentiation through Notch and Wnt/GSK-3β/β-catenin signaling. The results provide a novel translational direction for the clinical application of ED-71 against GIOP.

SLC38A2 provides proline and alanine to regulate postnatal bone mass accrual in mice.

Amino acids have recently emerged as important regulators of osteoblast differentiation and bone formation. Osteoblasts require a continuous supply of amino acids to sustain biomass production to fuel cell proliferation, osteoblast differentiation and bone matrix production. We recently identified proline as an essential amino acid for bone development by fulfilling unique synthetic demands that are associated with osteoblast differentiation. Osteoblasts rely on the amino acid transporter SLC38A2 to provide proline to fuel endochondral ossification. Despite this, very little is known about the function or substrates of SLC38A2 during bone homeostasis. Here we demonstrate that the neutral amino acid transporter SLC38A2 is expressed in osteoblast lineage cells and provides proline and alanine to osteoblast lineage cells. Genetic ablation of SLC38A2 using Prrx1Cre results in decreased bone mass in both male and female mice due to a reduction in osteoblast numbers and bone forming activity. Decreased osteoblast numbers are attributed to impaired proliferation and osteogenic differentiation of skeletal stem and progenitor cells. Collectively, these data highlight the necessity of SLC38A2-mediated proline and alanine uptake during postnatal bone formation and bone homeostasis.

The Correlation between Total Alkaline Phosphatase and Osteocalcin Levels in Systemic Lupus Erythematosus Patients

Systemic Lupus Erythematosus (SLE) is a chronic autoimmune inflammatory disease with various complications, including osteoporosis. However, Bone Mineral Density (BMD) examination, a gold standard for diagnosing and monitoring osteoporosis, is static. Alkaline phosphatase (ALP) is a membrane-bound glycoprotein that catalysis the hydrolysis of monoester phosphate. Osteocalcin (OC) is a non-collagenic bone protein that binds calcium and phosphate, which are both dynamic bone formation activity markers. This study analyzes the correlation between total ALP and OC serum levels in SLE patients. A cross-sectional observational analytic study was conducted in the Clinical Pathology Installation of Dr. Moewardi Hospital Surakarta in June 2020. The subjects were SLE patients receiving Methylprednisolone (MEP) therapy ≥1 year. Data distribution normality test by Saphiro-Wilk, comparative analysis with unpaired T-test, degree of correlation strength between research variables by Pearson correlation test. There were 41 female subjects, and comparative analysis of total ALP and serum OC levels were not significantly different in inactive and active SLE (ALP p=0.373, serum OC p=0.700). Total ALP and serum OC was found to have a weak positive correlation in all SLE patients (r=0.337; p=0.031), a moderate positive correlation in active SLE (r=0.426; p=0.043), while in inactive SLE there was no significant correlation (r=0.247; p=0.324). There is a significant moderate positive correlation between total ALP and serum OC in SLE patients. Total ALP and serum OC examinations are necessary for osteoporosis screening in SLE patients with > 1-year glucocorticoid (GC) therapy.

Dietary organosulfur compounds: Emerging players in the regulation of bone homeostasis by plant-derived molecules.

The progressive decline of bone mass and the deterioration of bone microarchitecture are hallmarks of the bone aging. The resulting increase in bone fragility is the leading cause of bone fractures, a major cause of disability. As the frontline pharmacological treatments for osteoporosis suffer from low patients’ adherence and occasional side effects, the importance of diet regimens for the prevention of excessive bone fragility has been increasingly recognized. Indeed, certain diet components have been already associated to a reduced fracture risk. Organosulfur compounds are a broad class of molecules containing sulfur. Among them, several molecules of potential therapeutic interest are found in edible plants belonging to the Allium and Brassica botanical genera. Polysulfides derived from Alliaceae and isothiocyanates derived from Brassicaceae hold remarkable nutraceutical potential as anti-inflammatory, antioxidants, vasorelaxant and hypolipemic. Some of these effects are linked to the ability to release the gasotrasmitter hydrogen sulfide (H2S). Recent preclinical studies have investigated the effect of organosulfur compounds in bone wasting and metabolic bone diseases, revealing a strong potential to preserve skeletal health by exerting cytoprotection and stimulating the bone forming activity by osteoblasts and attenuating bone resorption by osteoclasts. This review is intended for revising evidence from preclinical and epidemiological studies on the skeletal effects of organosulfur molecules of dietary origin, with emphasis on the direct regulation of bone cells by plant-derived polysulfides, glucosinolates and isothiocyanates. Moreover, we highlight the potential molecular mechanisms underlying the biological role of these compounds and revise the importance of the so-called ‘H2S-system’ on the regulation of bone homeostasis.

Pathogenic variants of sphingomyelin synthase SMS2 disrupt lipid landscapes in the secretory pathway.

Sphingomyelin is a dominant sphingolipid in mammalian cells. Its production in the trans-Golgi traps cholesterol synthesized in the ER to promote formation of a sphingomyelin/sterol gradient along the secretory pathway. This gradient marks a fundamental transition in physical membrane properties that help specify organelle identify and function. We previously identified mutations in sphingomyelin synthase SMS2 that cause osteoporosis and skeletal dysplasia. Here, we show that SMS2 variants linked to the most severe bone phenotypes retain full enzymatic activity but fail to leave the ER owing to a defective autonomous ER export signal. Cells harboring pathogenic SMS2 variants accumulate sphingomyelin in the ER and display a disrupted transbilayer sphingomyelin asymmetry. These aberrant sphingomyelin distributions also occur in patient-derived fibroblasts and are accompanied by imbalances in cholesterol organization, glycerophospholipid profiles, and lipid order in the secretory pathway. We postulate that pathogenic SMS2 variants undermine the capacity of osteogenic cells to uphold nonrandom lipid distributions that are critical for their bone forming activity.

A 3D-printed molybdenum-containing scaffold exerts dual pro-osteogenic and anti-osteoclastogenic effects to facilitate alveolar bone repair

The positive regulation of bone-forming osteoblast activity and the negative feedback regulation of osteoclastic activity are equally important in strategies to achieve successful alveolar bone regeneration. Here, a molybdenum (Mo)-containing bioactive glass ceramic scaffold with solid-strut-packed structures (Mo-scaffold) was printed, and its ability to regulate pro-osteogenic and anti-osteoclastogenic cellular responses was evaluated in vitro and in vivo. We found that extracts derived from Mo-scaffold (Mo-extracts) strongly stimulated osteogenic differentiation of bone marrow mesenchymal stem cells and inhibited differentiation of osteoclast progenitors. The identified comodulatory effect was further demonstrated to arise from Mo ions in the Mo-extract, wherein Mo ions suppressed osteoclastic differentiation by scavenging reactive oxygen species (ROS) and inhibiting mitochondrial biogenesis in osteoclasts. Consistent with the in vitro findings, the Mo-scaffold was found to significantly promote osteoblast-mediated bone formation and inhibit osteoclast-mediated bone resorption throughout the bone healing process, leading to enhanced bone regeneration. In combination with our previous finding that Mo ions participate in material-mediated immunomodulation, this study offers the new insight that Mo ions facilitate bone repair by comodulating the balance between bone formation and resorption. Our findings suggest that Mo ions are multifunctional cellular modulators that can potentially be used in biomaterial design and bone tissue engineering.

Controlled mechanical loading improves bone regeneration by regulating type H vessels in a S1Pr1-dependent manner.

Despite the best treatment, approximately 10% of fractures still face undesirable repair and result in delayed unions or non-unions. Dynamic mechanical stimulation promotes bone formation, when applied at the correct time frame, with optimal loading magnitude, frequency, and repetition. Controlled mechanical loading significantly increases osteogenic cells during the matrix deposition phase of bone repair. In the bone defect, the blood vessel network guides the initial bone formation activities. A unique blood vessel subtype (Type H) exists in bone, which expresses high levels of CD31 and endomucin, and functions to couple angiogenesis and osteogenesis. However, how this form of controlled mechanical loading regulates the Type H vessels and promotes bone formation is still not clear. Sphingosine 1-phosphate (S1P) participates in the bone anabolic process and is a key regulator of the blood vessel. Its receptor, sphingosine 1-phosphate receptor 1 (S1Pr1), is a mechanosensitive protein that regulates vascular integrity. Therefore, we hypothesis that controlled anabolic mechanical loading promotes bone repair by acting on Type H vessels. To study the effect of S1Pr1 on loading induced-bone repair, we utilized a stabilized tibial defect model, which allows for the application of anabolic mechanical loading. Mechanical loading upregulated S1Pr1 within the entire defect, with up to 80% expressed in blood vessels, as observed by deep tissue imaging. Additionally, S1Pr1 antagonism by W146 inhibited the anabolic effects of mechanical loading. We showed that mechanical loading or activating S1Pr1 could induce YAP nuclear translocation, a key regulator in the cell's mechanical response, in endothelial cells (ECs) in vitro. Inhibition of S1Pr1 in endothelial cells by siRNA reduced loading-induced YAP nuclear translocation and expressions of angiogenic genes. In vivo, YAP nuclear translocation in Type H vessels was up-regulated after mechanical loading but was inhibited by antagonizing S1Pr1. S1Pr1 agonist, FTY720, increased bone volume and Type H vessel volume, similar to that of mechanical stimulation. In conclusion, controlled anabolic mechanical loading enhanced bone formation mainly through Type H vessels in a S1Pr1-dependent manner.

Eosinophil-derived interferon-γ drives transmembrane protein 119-induced new bone formation in chronic rhinosinusitis with nasal polyps.

Chronic rhinosinusitis with nasal polyps (CRSwNP) is a chronic inflammatory sinonasal disease characterized by eosinophilic infiltration and new bone formation. These changes indicate the severity and prognosis of CRSwNP and may be closely linked to each other. We performed RNA sequencing to screen specific osteogenic molecules and validated transmembrane protein 119 (TMEM119) expression by quantitative polymerase chain reaction (qPCR) and immunohistochemistry analyses. TMEM119 knockdown was performed to observe the downregulation of bone mineralization. We validated the bone-forming activity of interferon-γ (IFN-γ) and its signaling pathways in cultured primary sinus bone cells. Cellular sources of IFN-γ were identified using immunohistochemistry and immunofluorescence analyses. Interleukin-4-eosinophil-IFN-γ axis and the effect of dupilumab were investigated in Eol-1 cells. We observed elevated IFN-γ levels and eosinophils in the nasal fluid and predominantly eosinophil-derived IFN-γ in the sinus mucosa of patients with CRSwNP. TMEM119 expression and bone-forming activities were increased in the osteitic and primary sinus bone cells of CRSwNP. IFN-γ treatment enhanced bone mineralization and TMEM119 expression via signal transducer and activator of transcription 1 (STAT1) signaling. Moreover, TMEM119 knockdown inhibited sinus bone cell mineralization and dupilumab attenuated IFN-γ secretion by IL4-stimulated Eol-1 cells. Eosinophil-derived IFN-γ promotes the bone-forming activities of sinus bone cells via the STAT1-TMEM119 signaling pathway. Interleukin-4-eosinophil-IFN-γ axis may be crucial for TMEM119-mediated new bone formation in CRSwNP.

SIRS tanılı hastalarda enflamasyon ve kemik döngüsü arasındaki ilişkinin sklerostin ve Dickkopf-1 (DKK-1) düzeyleri ile değerlendirilmesi

Introduction: In intensive care units (ICU), patients remain bedridden for a long time. In addition, severe infections are frequently seen in ICUs. Both prolonged immobilization and serious infections are associated with bone tissue loss. The Wnt pathway has recently been focused on evaluating bone tissue loss. The Wnt pathway participates in both infections and the formation of bone tissue. Wnt pathway inhibitors sclerostin and Dickkopf-1 (DKK-1) inhibit bone formation and increase osteoclastic activity. In this study, we aimed to examine bone turnover by the Wnt inhibitors sclerostin and DKK-1 and their possible associations with inflammation in SIRS patients.Methods: We included 30 patients diagnosed with systemic inflammatory response syndrome (SIRS) in the study group and 16 in the control group. Serum sclerostin, DKK-1, white blood cell (WBC), and C-Reactive Protein (CRP) levels on the day of SIRS diagnosis (basal), the 7th, 14th, and 21stdays were evaluated in the study group, and the results were compared with the control group.Results: When the control group was compared with the basal SIRS, there was a significant elevation in both sclerostin (p=0.003) and DKK-1 (p=0.001). Statistical analysis showed significant decreases in sclerostin levels between basal and the 7th, 14th, and 21st days (p=0.033, p=0.003, p=0.002, respectively). Similarly, significant decreases in DKK-1 levels between basal and the 7th and 21st days (p=0.015, p=0.001, respectively) and an insignificant decrease on the 14th day (p=0.191) was observed. Sclerostin was positively and significantly correlated with WBC and CRP in basal and 7th-day measurements and WBC in 7th and 14th days. DKK-1 is positively and significantly correlated with WBC in basal and 7th-daymeasurements, while DKK-1 negatively correlates with CRP in basal-7th-day measurements.Conclusion: In this study, it was shown for the first time that the Wnt antagonists sclerostin and DKK-1 values are high in SIRS patients in ICU. Both biomarker levels decreased in parallel with the treatment. However, it could not be associated with disease severity and inflammatory marker levels. We believe that monitoring the change of Wnt antagonists will be useful in demonstrating bone turnover in patients with SIRS.Keywords: Dickkopf-1, Intensive care unit, Sclerostin, Systemic inflammatory response syndrome, Wnt signaling pathway, Bone turnover

Sec-O-glucosylhamaudol promotes the osteogenesis of pre-osteoblasts via BMP2 and Wnt3a signaling

• SOGH was isolatedfrom the dried roots of Saposhnikoviadivaricata . • SOGH promotes osteoblast differentiation and mineralization. • SOGH promotes cell migration and adhesion. • The osteogenic properties of SOGH are mediated via the BMP2/Wnt3a signaling pathways. • SOGH has potential medicinal applications as a bone-protective compound. The perennial herb Saposhnikovia divaricata SCHISCHKIN, used as a source of traditional medicines, is known for its anti-bone erosion, anti-inflammatory, and analgesic effects. We examined the osteogenic properties of Sec- O -glucosylhamaudol (SOGH) extracted from S. divaricata roots as a bone-protective compound. We found that SOGH promoted osteoblast differentiation and mineralization and induced the expression of Bsp , Ocn , and Opn without any cytotoxic effects. SOGH also increased nuclear RUNX2 levels in pre-osteoblasts via the BMP2 and Wnt3a signaling pathways, but it had no evident effects on the autophagic and necroptotic signaling pathways. Furthermore, SOGH promoted bone-forming activities by inducing the adhesion and migration of osteoblasts and the expression of RUNX2 target proteins, including VEGF and MMP13. In sum, SOGH has osteogenic properties mediated via the BMP2 and Wnt3a signaling pathways and RUNX2 expression, indicating the bone-protective potential of this compound in diseases such as periodontitis and osteoporosis.

The effect of ethyl acetate fraction of Marsilea crenata presl. leaves in increasing osterix expression in hFOB 1.19 cells

Marsilea crenata Presl. leaves contain phytoestrogens that is suspected to play a role in increasing bone formation. Bone formation activity is defined by the expression of Osterix, a transcription factor that plays a key role in bone development. The aim of this study is to show that the ethyl acetate fraction of Marsilea crenata Presl. leaves can increase bone formation process in hFOB 1.19 cells in a TNF-α dependent manner, by measurement of Osterix. The hFOB 1.19 cells were grown in 24-well microplates and treated with 10 ng/ml TNF-α for 24 hours. The ethyl acetate fraction of Marsilea crenata Presl. leaves were also added in concentrations of 62.5, 125, and 250 ppm. A positive control was employed, at a concentration of 2.5 g/L of genistein. The expression of osterix was examined using an immunocytochemistry technique with CLSM to assess bone forming activity. The results show that ethyl acetate fraction Marsilea crenata Presl. leaves can boost osterix expression in hFOB 1.19 cells, with optimal concentration was 250 ppm at p<0.005. The ethyl acetate fraction of Marsilea crenata Presl. leaves can increase Osterix in osteoblast hFOB 1.19 cell, indicating improved bone forming activity.

Nano-encapsulation of melatonin into polydiacetylene-phospholipid assembly for sustained-release and enhanced bone formation in zebrafish

The low bioavailability of melatonin limits its effective antiosteoporosis activity. In the present study, the polymer-supported liposome is used to encapsulate the melatonin in order to enhance its bone formation activity. More specifically, the melatonin drug has enwrapped into a polymer/lipid hybrid nanovesicle (Lip-mel) by the self-assembly of 10,12-pentacosadiynoic acid (PCDA) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) through thin-film hydration method. The liposome-encapsulated melatonin was characterised well by UV–Vis absorption spectroscopy, fluorescence microscopy, energy-dispersive X-ray analysis (EDX), and dynamic light scattering (DLS) measurements. The conjugated yne-ene chain of polymeric backbone of polydiacetylene (PDA) facilitates the formation of stable nanoaggregate from which only 50% of the encapsulated melatonin was leached out even after 72 h under physiological conditions. The Lip-mel shows alkaline phosphatase activity (ALP) and calcium deposition in mouse mesenchymal stem cells more efficient than non-encapsulated melatonin. Furthermore, the Lip-mel elevated the expression of key transcription factors (Runx2, type1 col mRNAs) and secretion of extracellular matrix proteins that are related to osteoblast differentiation. Interestingly, the bone formation in zebrafish model was also enhanced after exposure of Lip-mel compared to melatonin. Thus, the nano formulation of melatonin could be developed as efficient antiosteoporosis drug with better pharmacokinetic properties than that of free form of melatonin.

Bone Regeneration of Critical-Size Calvarial Defects in Rats Using Highly Pressed Nano-Apatite/Collagen Composites.

Osteo-conductive bone substitute materials are required in dentistry. In this study, highly pressed nano-hydroxyapatite/collagen (P-nHAP/COL) composites were formed by a hydraulic press. Critical-size bone defects (Φ = 6 mm) were made in the cranial bones of 10-week-old Wistar rats, in which P-nHAP/COL and pressed collagen (P-COL) specimens were implanted. Defect-only samples (DEF) were also prepared. After the rats had been nourished for 3 days, 4 weeks, or 8 weeks, ossification of the cranial defects of the rats was evaluated by micro-computed tomography (micro-CT) (n = 6 each). Animals were sacrificed at 8 weeks, followed by histological examination. On micro-CT, the opacity of the defect significantly increased with time after P-nHAP/COL implantation (between 3 days and 8 weeks, p < 0.05) due to active bone regeneration. In contrast, with P-COL and DEF, the opacity increased only slightly with time after implantation, indicating sluggish bone regeneration. Histological inspections of the defect zone implanted with P-nHAP/COL indicated the adherence of multinucleated giant cells (osteoclasts) to the implant with phagocytosis and fragmentation of P-nHAP/COL, whereas active bone formation occurred nearby. Fluorescent double staining indicated dynamic bone-formation activities. P-nHAP/COL is strongly osteo-conductive and could serve as a useful novel bone substitute material for future dental implant treatments.

Alendronate delays activation of bone formation in cortical bone in postmenopausal osteoporosis

Alendronate delays activation of bone formation in cortical bone in postmenopausal osteoporosis

Glucocorticoids inhibit the maturation of committed osteoblasts via SOX2.

During bone formation, mesenchymal progenitor cells mature into bone-forming osteoblasts after undergoing several stages of differentiation. Impaired bone formation is a predominant finding in glucocorticoid (GC)-induced osteoporosis (GIO). Osteoblasts at different stages of maturation can be affected by excessive endogenous or therapeutic GCs. Sex-determining region Y-box 2 (SOX2) is normally expressed in immature osteoblasts, but its overexpression can suppress osteoblast differentiation. This study aimed to evaluate whether GC affects SOX2 expression in osteoblasts, and whether SOX2 contributes to GC-induced inhibition of osteoblast differentiation. Treatment with GCs such as dexamethasone (Dex) or hydrocortisone enhanced SOX2 expression. Silencing SOX2 improved inhibition of GC-induced osteoblast differentiation, whereas SOX2 overexpression decreased mineralized nodule formation and RUNX2 and Osterix expression in MC3T3-E1 cells. On the contrary, when C3H10T1/2 uncommitted mesenchymal stem cells were subjected to SOX2 overexpression, RUNX2 expression increased. As a mechanism of Dex-induced SOX2 upregulation in preosteoblasts, we found that the STAT3 pathway or GC receptor (GR) is involved, using a GR antagonist, STAT3 regulators, and chromatin immunoprecipitation assays. Moreover, mice treated with Dex for 4 weeks showed a notable increase in SOX2 expression in the bones and an increased ratio of procollagen type 1 N-terminal propeptide to osteocalcin in the plasma than in control mice. This study demonstrated that GC enhances SOX2 expression in vitro in osteoblast and in vivo in the mice bone, which affects bone-forming activity differently depending on the differentiation stage of osteoblast-lineage cells. Our results provide new insights into prevention and treatment against impaired bone formation in GIO.

The Role Of BMPs in the Regulation of Osteoclasts Resorption and Bone Remodeling: From Experimental Models to Clinical Applications.

In response to mechanical forces and the aging process, bone in the adult skeleton is continuously remodeled by a process in which old and damaged bone is removed by bone-resorbing osteoclasts and subsequently is replaced by new bone by bone-forming cells, osteoblasts. During this essential process of bone remodeling, osteoclastic resorption is tightly coupled to osteoblastic bone formation. Bone-resorbing cells, multinuclear giant osteoclasts, derive from the monocyte/macrophage hematopoietic lineage and their differentiation is driven by distinct signaling molecules and transcription factors. Critical factors for this process are Macrophage Colony Stimulating Factor (M-CSF) and Receptor Activator Nuclear Factor-κB Ligand (RANKL). Besides their resorption activity, osteoclasts secrete coupling factors which promote recruitment of osteoblast precursors to the bone surface, regulating thus the whole process of bone remodeling. Bone morphogenetic proteins (BMPs), a family of multi-functional growth factors involved in numerous molecular and signaling pathways, have significant role in osteoblast-osteoclast communication and significantly impact bone remodeling. It is well known that BMPs help to maintain healthy bone by stimulating osteoblast mineralization, differentiation and survival. Recently, increasing evidence indicates that BMPs not only help in the anabolic part of bone remodeling process but also significantly influence bone catabolism. The deletion of the BMP receptor type 1A (BMPRIA) in osteoclasts increased osteoblastic bone formation, suggesting that BMPR1A signaling in osteoclasts regulates coupling to osteoblasts by reducing bone-formation activity during bone remodeling. The dual effect of BMPs on bone mineralization and resorption highlights the essential role of BMP signaling in bone homeostasis and they also appear to be involved in pathological processes in inflammatory disorders affecting bones and joints. Certain BMPs (BMP2 and -7) were approved for clinical use; however, increased bone resorption rather than formation were observed in clinical applications, suggesting the role BMPs have in osteoclast activation and subsequent osteolysis. Here, we summarize the current knowledge of BMP signaling in osteoclasts, its role in osteoclast resorption, bone remodeling, and osteoblast–osteoclast coupling. Furthermore, discussion of clinical application of recombinant BMP therapy is based on recent preclinical and clinical studies.

Thelephoric acid, p-terphenyl, induces bone-forming activities in pre-osteoblasts

• Thelephoric acid is a p-terphenyl compound isolated from the mushrooms of Polyozellus multiplex. • Thelephoric acid enhances ALP enzymatic activities in pre-osteoblasts. • Thelephoric acid enhances bone matrix mineralization in pre-osteoblasts. • Thelephoric acid enhances nuclear RUNX2 expression and accumulation via Smad1/5/8 phosphorylation. • Thelephoric acid enhances cell adhesion and migration in osteoblast differentiation. • Thelephoric acid is a potentially beneficial compound for prevention and treatment in bone diseases. Polyozellus multiplex , a wild mushroom belonging to Thelephoracea, is distributed in Korea, Japan, and China. Thelephoric acid (Thel), a terphenylquinone found in P. multiplex , reportedly exhibits beneficial effects, including antioxidant, anticancer, and neuroprotective activities. Herein, we extracted Thel from P. multiplex (>95.8% purity) and examined its osteogenic effects on pre-osteoblasts. Thel exerted no cytotoxicity in pre-osteoblasts. Based on alkaline phosphatase staining and activity, Thel promoted osteoblast differentiation and bone matrix mineralization. Biochemical investigations revealed that Thel enhanced nuclear runt-related transcription factor 2 (RUNX2) expression and accumulation via Smad1/5/8 phosphorylation, while exhibiting synergistic activities following cotreatment with bone morphogenetic protein 2 or fibroblast growth factor 2 in pre-osteoblast cultures. Thel increased cell adhesion and migration in differentiating osteoblasts by upregulating matrix metalloproteinase 13 (MMP13). Collectively, these findings indicate that Thel induced osteogenic properties by regulating RUNX2 and MMP13 expression, suggesting that Thel could be further developed to treat bone disorders.