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Purpose: To estimate the effect of simvastatin gel in the osseointegration of dental implants using bone scintigraphy. Materials and Methods: A total of 20 participants with missing mandibular first molars and D2 type bone were assigned equally to two groups. Group A received 1.2% simvastatin and group B received placebo gels during the placement of implants. The participants were subjected to bone scintigraphy to determine the osteoblastic activity at baseline, 30 days, and 90 days after implant placement. Results: Group A had a significant increase in osteoblastic activity between baseline, day 30, and day 90 (P < .05), with a higher mean of 100.06% ± 21.644% on day 30. Group B had a significant increase in osteoblastic activity only between baseline and day 30 and between baseline and day 90 (P < .05). There was no difference between days 30 and 90 (P > .05), with a higher mean of 79.20% ± 18.255% on day 30. A bivariate analysis performed at different time periods revealed a significant difference between groups A and B on day 30. Conclusions: Implants placed with 1.2% simvastatin gel showed enhanced osteoblastic activity in the 4th week after implant placement, indicating that there was a faster rate of osseointegration at an early stage. [ABSTRACT FROM AUTHOR]

Considering the biological activity of osteoblasts is crucial when devising new approaches to enhance the osseointegration of implant surfaces, as their behavior profoundly influences clinical outcomes. An established inverse correlation exists between osteoblast proliferation and their functional differentiation, which constrains the rapid generation of a significant amount of bone. Examining the surface morphology of implants reveals that roughened titanium surfaces facilitate rapid but thin bone formation, whereas smooth, machined surfaces promote greater volumes of bone formation albeit at a slower pace. Consequently, osteoblasts differentiate faster on roughened surfaces but at the expense of proliferation speed. Moreover, the attachment and initial spreading behavior of osteoblasts are notably compromised on microrough surfaces. This review delves into our current understanding and recent advances in nanonodular texturing, meso-scale texturing, and UV photofunctionalization as potential strategies to address the biological dilemma of osteoblast kinetics, aiming to improve the quality and quantity of osseointegration. We discuss how these topographical and physicochemical strategies effectively mitigate and even overcome the dichotomy of osteoblast behavior and the biological challenges posed by microrough surfaces. Indeed, surfaces modified with these strategies exhibit enhanced recruitment, attachment, spread, and proliferation of osteoblasts compared to smooth surfaces, while maintaining or amplifying the inherent advantage of cell differentiation. These technology platforms suggest promising avenues for the development of future implants.

Dental implant therapy, established as standard-of-care nearly three decades ago with the advent of microrough titanium surfaces, revolutionized clinical outcomes through enhanced osseointegration. However, despite this pivotal advancement, challenges persist, including prolonged healing times, restricted clinical indications, plateauing success rates, and a notable incidence of peri-implantitis. This review explores the biological merits and constraints of microrough surfaces and evaluates the current landscape of nanofeatured dental implant surfaces, aiming to illuminate strategies for addressing existing impediments in implant therapy. Currently available nanofeatured dental implants incorporated nano-structures onto their predecessor microrough surfaces. While nanofeature integration into microrough surfaces demonstrates potential for enhancing early-stage osseointegration, it falls short of surpassing its predecessors in terms of osseointegration capacity. This discrepancy may be attributed, in part, to the inherent dichotomy kinetics of osteoblasts, wherein increased surface roughness by nanofeatures enhances osteoblast differentiation but concomitantly impedes cell attachment and proliferation. We also showcase a controllable, hybrid micro-nano titanium model surface and contrast it with commercially-available nanofeatured surfaces. Unlike the commercial nanofeatured surfaces, the controllable micro-nano hybrid surface exhibits superior potential for enhancing both cell differentiation and proliferation. Hence, present nanofeatured dental implants represent an evolutionary step from conventional microrough implants, yet they presently lack transformative capacity to surmount existing limitations. Further research and development endeavors are imperative to devise optimized surfaces rooted in fundamental science, thereby propelling technological progress in the field.

Purpose: To evaluate the usefulness of ridge augmentation using a customized titanium mesh (CTM) that was preformed by trimming and bending the commercial titanium mesh on a virtually reconstructed 3D acrylic resin model using clinical, radiologic, histologic, and histomorphometric analyses. Materials and Methods: This study was designed prospectively for patients who required vertical ridge augmentation using a staged approach before implant surgery. After installation of the CTM, grafting was performed using deproteinized porcine bone mineral covered with an absorbable membrane. Computed tomography was performed preoperatively and 6 months after simultaneous/staged guided bone regeneration to measure planned, reconstructed, and lacking bone volume, and the reconstruction rate was calculated based on these values. Clinical complications were also recorded, particularly the mesh exposure rate. At re-entry, the bone core was obtained using a trephine bur, and histologic and histomorphometric analyses were performed. Results: A total of 10 sites in eight patients were used for the study analysis. The mean planned bone volume was 1.15 cm3 (range: 0.78 to 1.56 cm3), mean lacking bone volume was 0.13 cm3 (range: 0 to 0.59 cm3), and mean reconstructed bone volume was 1.02 cm3 (range: 0.56 to 1.43 cm3). The exposure rate was 30% (3 out of 10 sites). The reconstruction rate was over 80%, except for one case that showed suppuration. From histomorphometric analysis, 27.52% ± 16.87% of new bone, 7.62% ± 5.19% of residual graft, and 64.86% ± 23.76% of connective tissue were observed. The core biopsy samples demonstrated different pseudoperiosteum layer appearances based on the healing stage of the augmented sites. In the premature bone, the inner osteogenic layer consisted of multiple layers of osteoblast cells with adjacent large blood vessels. However, in the mature augmented site, there was no specific inner osteogenic layer, and the outer fibrous layer was dominant. Conclusions: The fabrication of CTM based on the application of the 3D-printing technique makes vertical ridge augmentation easier and can reduce complications and achieve target bone acquisition. In addition, it is expected that quantitative analysis of the pseudoperiosteum layer will be facilitated using the CTM. [ABSTRACT FROM AUTHOR]

Androgen binding to the androgen receptor (AR) in the cytoplasm induces the AR to translocate to the nucleus, where it regulates the expression of target genes. Here, we found that androgens rapidly activated a plasma membrane-associated signaling node that enhanced nuclear AR functions. In murine primary osteoblasts, dihydrotestosterone (DHT) binding to a membrane-associated form of AR stimulated plasma membrane-associated protein kinase G type 2 (PKG2), leading to the activation of multiple kinases, including ERK. Phosphorylation of AR at Ser515 by ERK increased the nuclear accumulation and binding of AR to the promoter of Ctnnb1, which encodes the transcription factor β-catenin. In male mouse osteoblasts and human prostate cancer cells, DHT induced the expression of Ctnnb1 and CTNN1B, respectively, as well as β-catenin target genes, stimulating the proliferation, survival, and differentiation of osteoblasts and the proliferation of prostate cancer cells in a PKG2-dependent fashion. Because β-catenin is a master regulator of skeletal homeostasis, these results explain the reported male-specific osteoporotic phenotype of mice lacking PKG2 in osteoblasts and imply that PKG2-dependent AR signaling is essential for maintaining bone mass in vivo. Our results suggest that widely used pharmacological PKG activators, such as sildenafil, could be beneficial for male and estrogen-deficient female patients with osteoporosis but detrimental in patients with prostate cancer.

Three-dimensional cultures are widely used to study bone and cartilage. These models often focus on the interaction between osteoblasts and osteoclasts or osteoblasts and chondrocytes. A culture of osteoblasts, osteoclasts and chondrocytes would represent the cells that interact in the joint and a model with these cells could be used to study many diseases that affect the joints. The goal of this study was to develop 3D bone-cartilage interface (3D-BCI) that included osteoblasts, osteocytes, osteoclasts, and cartilage. Fluorescently tagged cell lines were developed to assess the interactions as cells differentiate to form bone and cartilage. Mouse cell line, MC3T3, was labeled with a nuclear GFP tag and differentiated into osteoblasts and osteocytes in Matrigel. Raw264.7 cells transfected with a red cytoplasmic tag were added to the system and differentiated with the MC3T3 cells to form osteoclasts. A new method was developed to differentiate chondrocyte cell line ATDC5 in a cartilage spheroid, and the ATDC5 spheroid was added to the MC3T3 and Raw264.7 cell model. We used an Incucyte and functional analysis to assess the cells throughout the differentiation process. The 3D-BCI model was found to be positive for TRAP, ALP, Alizarin red and Alcian blue staining to confirm osteoblastogenesis, osteoclastogenesis, and cartilage formation. Gene expression confirmed differentiation of cells based on increased expression of osteoblast markers: Alpl , Bglap , Col1A2 , and Runx 2 , cartilage markers: Acan , Col2A1 , Plod2 , and osteoclast markers: Acp5 , Rank and Ctsk. Based on staining, protein expression and gene expression results, we conclude that we successfully developed a mouse model with a 3D bone-cartilage interface. [ABSTRACT FROM AUTHOR]

We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast differentiation was analyzed. Using catalytically inactive Cas12a, the CRISPRa system induced targeted overexpression of endogenous miRNA and repressed the luciferase activities of reporters that contained functional miRNA target sites. The delivery of miR-2861 into MSCs enhanced osteoblast differentiation by decreased expressions of the HDAC1, 4 and 5 genes. The mechanism of HDAC5 repression by miR-2861 in humans has not been fully elucidated. To this end, the HDAC5 mRNA sequence was analyzed and a putative primate-specific miR-2861 binding site was identified in the 3' untranslated region (3'-UTR). CRISPRa was applied to validate the putative binding site and an increase in endogenous miR-2861 was found to repress the expression of a reporter that contained the novel miR-2861 binding site. The delivery of miR-2861 to human MSCs enhanced osteoblast differentiation. In the 3'-UTR, the HDAC5 repression was mediated by the miR-2861 binding site, and miR-2861 promoted osteoblast differentiation via the inhibition of HDAC5 through a primate-specific miRNA binding site. Therefore, miRNAmiR-2861 with the CRISPRa methods might be a good biomaterial for osteogenesis augmentation. [ABSTRACT FROM AUTHOR]

AbstractBackgroundMethodsResultsConclusionThe aberrant H-type vessel formation was found to be intimately linked to subchondral bone remodeling during osteoarthritis (OA) development. Herein, we investigated the role and mechanism of osteoblast-secreted slit guidance ligand 3 (Slit3) in H-type vessel formation during OA progression.Slit3 protein levels in subchondral bone samples of OA patients were detected. The isolated osteoblasts were transfected with Slit3 overexpression or knockdown plasmids, and their conditioned medium was cultured with endothelial progenitor cells (EPCs). The migration, tube formation, VEGF, and H-type vessel marker protein CD31 and endomucin (EMCN) levels in EPCs were accessed. The interactions between Slit3 and roundabout (Robo) family members were validated by Co-IP assay. Besides, whether the Slit3/Robo signaling affects the transforming growth factor β1 (TGF-β1)/SMADs pathway was determined. Additionally, sh-Slit3 was injected into OA mice, followed by the detection of articular cartilage degradation, subchondral bone remodeling, and H-type vessel formation.Slit3 was upregulated in subchondral bone tissues of OA patients. Slit3 overexpression in osteoblasts intensified the migration and H-type vessel formation of EPCs, while Slit3 knockdown showed the opposite results. Slit3 overexpression enhanced Robo1 protein level. Robo1 knockdown abrogated Slit3-mediated migration and H-type vessel formation in EPCs. Slit3 activated the TGF-β1/SMADs pathway in EPCs, which might be associated with H-type vessel formation in EPCs. Additionally, Slit3 silencing restrained articular cartilage degradation, aberrant subchondral bone formation, and H-type vessel formation in OA mice.Inhibition of Slit3/Robo1 signaling alleviates osteoarthritis in mice by reducing abnormal H-type vessel formation in the subchondral bone. [ABSTRACT FROM AUTHOR]

Dental implants fabricated from titanium have several limitations and therefore, alternative materials that fulfil the criteria of successful dental implant (bioactivity and anti-bacterial activity) need to be considered. Polyether ether ketone (PEEK) has been suggested to replace titanium implants. However, this material needs surface modification to meet the appropriate criteria. A nano-sized zirconium phosphate/GO (nZrP/GO) composite coating was prepared to improve PEEK's biological qualities. Polished and cleaned PEEK discs were coated with the composite of nZrP doped with 1.25 wt% GO by the soft-template method. To analyze the composite coating, X-ray, atomic force microscopy, and field emission scanning electron microscopy-energy dispersive spectroscopy were used. The adhesion of the coating to PEEK was measured by adhesive tape test. By measuring the optical contact angle, the coated and non-coated samples' differences in wettability were evaluated. Antimicrobial activity was evaluated against S. aureus and E. coli and cytotoxicity tested employing gingival fibroblasts and osteoblast-like cells. The nZrP/GO composite coating was 23.45 µm thick, was irregular and attached strongly to the PEEK surface. Following coating, the water contact angle dropped to 34° and surface roughness to 13 nm. The coating reduced the count of bacteria two-fold and was non-cytotoxic to mammalian osteoblast-like cells and fibroblasts. A precipitation of nano-calcium-deficient apatite was observed on the surface of the nZrP/GO coating following a 28-day immersion in SBF. PEEK-coated with nZr/GO coating is a good candidate as dental implant. • nZrP/GO converts PEEK surface from bionert to a bioactive surface. • Addition of GO into nZrP potentiates its anti-bacterial activity. • PEEK implant coated with nZrP/GO is not cytotoxic to mammalian cells. • nZrP/GO increases surface wettability of PEEK implant. • Addition of GO has no effect on adhesion of nZrP coating to PEEK surface. [ABSTRACT FROM AUTHOR]

Bone diseases such as osteoporosis, osteoarthritis, and rheumatoid arthritis are a class of diseases that have long plagued people. Its pathogenesis may be multi-factorial, such as genetics, living environment, and drug effects. It is generally believed that bone metabolic disorders are an important part of the pathogenesis of bone diseases. With the deepening of research, it has been found that ferroptosis and mitochondrial homeostasis play a crucial role in regulating osteogenesis, osteoclasts, chondrocyte proliferation, differentiation and function, and bone-related diseases. This article will summarize the current research on mitochondrial homeostasis and ferroptosis and their mechanisms of action in bone metabolism disorders, and elaborate on the relationship between the three. The aim is to explore the potential and new research directions of mitochondrial homeostasis and ferroptosis in the treatment of diseases caused by bone metabolic disorders. [ABSTRACT FROM AUTHOR]

Background: The incidence of osteoarthritis (OA) is increasing, yet its pathogenesis remains largely unknown. Recent studies suggest that abnormal subchondral bone remodeling plays a crucial role in OA development, highlighting a gap in clinical treatments targeting this aspect. Soybean Isoflavone (SI) has shown potential in treating OA, although its mechanisms are not fully understood. Methods: This research investigated the effects of SI on subchondral bone remodeling in an OA rat model, assessing joint damage, OARSI scores, and type H vessel formation (CD31hiEmcnhi expression). Additionally, the expression of ALP, OCN, BMP, and TSC1 was evaluated to determine involvement of the mTORC1 pathway. In vitro studies on IL-1β-induced osteoblasts further examined the impact of SI on TSC1/mTORC1 signaling and related markers. Results: SI treatment reduced joint damage and OARSI scores in the rat OA model, significantly decreasing CD31hiEmcnhi expression, indicating a reduction in type H vessel formation. SI also downregulated ALP, OCN, and BMP expression while upregulating TSC1, suggesting inhibition of the mTORC1 signaling pathway and VEGF release. In vitro, SI increased TSC1 expression and decreased mTORC1 signaling, VEGF, ALP, OCN, and BMP levels in IL-1β-induced osteoblasts. Conclusion: SI targets the TSC1/mTORC1 signaling pathway to suppress osteoblast activation and VEGF release, inhibiting type H vessel formation and slowing abnormal subchondral bone remodeling. These findings provide a novel therapeutic approach for OA by focusing on subchondral bone remodeling mechanisms. [ABSTRACT FROM AUTHOR]

The development of effective biomaterials for tissue regeneration has led to the exploration of blood derivatives such as leucocyte- and platelet-rich fibrin (L-PRF). A novel variant, Albumin-Enriched Platelet-Rich Fibrin (Alb-PRF), has been introduced to improve structural stability and bioactivity, making it a promising candidate for bone regeneration. This study aimed to evaluate Alb-PRF's capacity for cytokine and growth factor release, along with its effects on the proliferation, differentiation, and mineralization of human osteoblasts in vitro. Alb-PRF membranes were analyzed using histological, scanning electron microscopy, and fluorescence microscopy techniques. Cytokine and growth factor release was quantified over seven days, and osteoinductive potential was evaluated with MG-63 osteoblast-like cells. Structural analysis showed Alb-PRF as a biphasic, highly cellularized material that releases lower levels of inflammatory cytokines and higher concentrations of platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) compared to L-PRF. Alb-PRF exhibited higher early alkaline phosphatase activity and in vitro mineralization (p < 0.05) and significantly increased the OPG/RANKL mRNA ratio (p < 0.05). These results indicate that Alb-PRF has promising potential as a scaffold for bone repair, warranting further in vivo and clinical assessments to confirm its suitability for clinical applications. [ABSTRACT FROM AUTHOR]

Objective: Bone incision leads to interrupted and sluggish blood flow in the process of distraction osteogenesis (DO), creating a hypoxia (0–2% oxygen tension) at the center of the bone callus. This hypoxia is critical in the coupling of osteogenesis and angiogenesis during DO. This study aimed to investigate the effect of Uniaxial Static Strain (USS) on osteogenesis in osteoblasts under hypoxic conditions, with a focus on the expression of osteogenic markers and angiogenic factors. Methods: The USS was made by a multi-unit tension compression device.Osteoblasts were subjected to 10% USS made under hypoxic conditions to mimic the process of DO in vitro. The cell proliferation, alkaline phosphatase (ALP) activity, mineralized nodule formation, and expression of osteogenic and angiogenic markers were evaluated by using a CCK-8 assay, alkaline phosphatase (ALP) staining, ALP activity assay, alizarin red S staining, qRT-PCR, Western blotting and ELISA. Results: Hypoxia inhibited osteoblast cell proliferation, ALP activity, mineralized nodule formation, and the expression of runt-related transcription factor 2 (Runx- 2), osteopontin(OPN), osteocalcin (OCN), collagen type I (Col1a1). Conversely, hypoxia upregulated the expression of hypoxia-inducible factor 1-alpha (HIF-1α) and vascular endothelial growth factor (VEGF), which are associated with angiogenesis. However, the application of USS enhanced osteoblasts' osteogenic capacity and upregulated angiogenic factors under hypoxic conditions. Conclusion: USS can enhance osteogenesis in osteoblasts under hypoxic conditions. Moreover, it may stimulate angiogenesis by promoting the expression of VEGF, which further contributes to bone formation. This finding provides important implications for understanding the mechanisms involved in bone regeneration and may have clinical applications in optimizing the effectiveness of DO techniques. [ABSTRACT FROM AUTHOR]

Osteoblasts originate from mesenchymal stem cells (MSCs) and their maturation is regulated by numerous genes. However, specific genes that play decisive roles in osteoblast maturation are not completely understood. Herein, we propose a new algorithm called SGCAL to identify sensitive gene combinations during osteoblast development and decode the regulatory roles of sensitive gene combinations in cell development. In contrast to conventional gene identification algorithms, the SGCAL algorithm, which is based on a combination of ARNN-LNE iterative cycles, quantitatively evaluates the sensitivity of genes by analyzing the average network entropy before and after gene perturbation. We verified that osteoblast development is fundamentally regulated by an overall sensitive network formed by specific genes. Moreover, we explored the roles of these sensitive genes and their combinations in the transition time point of the initial state of network change. Our algorithm was validated on two independent mouse osteoblast development datasets, successfully identifying sensitive genes and their combinations in both datasets and revealing that gene combinations containing Col1a1 can exhibit significant sensitivity. This study demonstrated the universality of cell fate and network regulation, offering new insights and a theoretical basis for the treatment and repair of bone diseases An algorithm called SGCAL is proposed to check effectively the sensitive genes and their combination during osteoblast development. The gene combinations containing Col1a1 can exhibit significant sensitivity for the process of Osteoblasts. We perturb expression data to discern the sensitivity of the maker gene by measuring the network entropy. [ABSTRACT FROM AUTHOR]

This study involved the proliferation and differentiation of osteoblasts treated with low-intensity pulsed ultrasound (LIPUS) and iron (Fe3+) ions, respectively. The biological effects of LIPUS and Fe3+ ions on the proliferation and differentiation of osteoblasts were also evaluated. MC3T3-E1 cells were seeded in six-well plates with the medium, which contained different concentrations of Fe3+ (0, 100, 200, 300, 400, 500, 600 and 700 μg L-1, respectively). LIPUS treatment was directed at the bottom of the plate for 20 min at an intensity of 80 mW cm-2 every day. Viability results showed that a dose of 400 μg L-1 Fe3+ ions had the best effect at promoting osteogenic proliferation in cell culture. The results of alkaline phosphatase staining and mineralization indicated that the differentiation of osteoblasts was promoted by LIPUS and Fe3+ ions. Fluorescence staining results showed that the number of cell nuclei in the LIPUS, Fe3+ and LIPUS-Fe groups increased by 37.20%, 55.81% and 89.76%, respectively. Migration data indicated that migration and proliferation rates were increased by LIPUS and Fe3+, and the results of protein expression indicated that LIPUS and Fe3+ may increase the expression of Wnt, β-catenin, and Runx2, hence promoting normal bone regeneration and development. The combination of LIPUS (1.5 MHz, 80 mW cm-2) and Fe3+ accelerates the proliferation and differentiation of osteoblasts significantly compared with single-factor treatment (stimulated by LIPUS and Fe3+ ions, respectively). This study could establish a foundation for LIPUS-responsive biomaterials in the repair and regeneration of bone tissues. [ABSTRACT FROM AUTHOR]

Skeletal tissue development and regeneration in mammals are intricate, multistep, and highly regulated processes. Various signaling pathways have been implicated in the regulation of these processes, including redox. Redox signaling is the signal transduction by electron transfer reactions involving free radicals or related species. Redox homeostasis is essential to cell metabolic states, as the ROS not only regulates cell biological processes but also mediates physiological processes. Following a bone fracture, redox signaling is also triggered to regulate bone healing and regeneration by targeting resident stromal cells, osteoblasts, osteoclasts and endothelial cells. This review will focus on how the redox signaling impact the bone development and bone regeneration. [ABSTRACT FROM AUTHOR]

Spermine synthase, encoded by the SMS gene, is involved in polyamine metabolism, as it is required for the synthesis of spermine from its precursor molecule spermidine. Pathogenic variants of SMS are known to cause Snyder–Robinson syndrome (SRS), an X-linked recessive disorder causing various symptoms, including intellectual disability, muscular hypotonia, infertility, but also skeletal abnormalities, such as facial dysmorphisms and osteoporosis. Since the impact of a murine SMS deficiency has so far only been analyzed in Gy mice, where a large genomic deletion also includes the neighboring Phex gene, there is only limited knowledge about the potential role of SMS in bone cell regulation. In the present manuscript, we describe 2 patients carrying distinct SMS variants, both diagnosed with osteoporosis. Whereas the first patient displayed all characteristic hallmarks of SRS, the second patient was initially diagnosed, based on laboratory findings, as a case of adult-onset hypophosphatasia. To study the impact of SMS inactivation on bone remodeling, we took advantage of a newly developed mouse model carrying a pathogenic SMS variant (p.G56S). Compared to their wildtype littermates, 12-wk-old male SMSG56S/0 mice displayed reduced trabecular bone mass and cortical thickness, as assessed by μCT analysis of the femur. This phenotype was histologically confirmed by the analysis of spine and tibia sections, where we also observed a moderate enrichment of non-mineralized osteoid in SMSG56S/0 mice. Cellular and dynamic histomorphometry further identified a reduced bone formation rate as a main cause of the low bone mass phenotype. Likewise, primary bone marrow cells from SMSG56S/0 mice displayed reduced capacity to form a mineralized matrix ex vivo, thereby suggesting a cell-autonomous mechanism. Taken together, our data identify SMS as an enzyme with physiological relevance for osteoblast activity, thereby demonstrating an important role of polyamine metabolism in the control of bone remodeling. [ABSTRACT FROM AUTHOR]

A correction to the article "Mir-381-3p aggravates ovariectomy-induced osteoporosis by inhibiting osteogenic differentiation through targeting KLF5/Wnt/βcatenin signaling pathway," published online is presented.

Osteoblasts are mainly derived from mesenchymal stem cells in the bone marrow. These stem cells can differentiate into osteoblasts, which have the functions of secreting bone matrix, promoting bone formation, and participating in bone remodeling. The abnormality of osteoblasts can cause a variety of bone-related diseases, including osteoporosis, delayed fracture healing, and skeletal deformities. In recent years, with the side effects caused by the application of PTH drugs, biphosphonate drugs, and calmodulin drugs, people have carried out more in-depth research on the mechanism of osteoblast differentiation, and are actively looking for natural compounds for the treatment of osteoporosis. The Wnt/β-catenin signaling pathway is considered to be one of the important pathways of osteoblast differentiation, and has become an important target for the treatment of osteoporosis. The Wnt/β-catenin signaling pathway, whether its activation is enhanced or its expression is weakened, will cause a variety of diseases including tumors. This review will summarize the effect of Wnt/β-catenin signaling pathway on osteoblast differentiation and the correlation between the related proteins in the pathway and human diseases. At the same time, the latest research progress of natural compounds targeting Wnt/β-catenin signaling pathway against osteoporosis is summarized. [ABSTRACT FROM AUTHOR]

Mesenchymal stem cells (MSCs) have gained tremendous interest due to their overall potent pro‐regenerative and immunomodulatory properties. In recent years, various in vitro and preclinical studies have investigated different priming ("licensing") approaches to enhance MSC functions for specific therapeutic purposes. In this study, we primed bone marrow‐derived human MSCs (hMSCs) with an inflammation cocktail designed to mimic the elevated levels of inflammatory mediators found in serum of patients with severe injuries, such as bone fractures. We observed a significantly enhanced osteogenic differentiation potential of primed hMSCs compared to untreated controls. By RNA‐sequencing analysis, we identified the immediate early response 3 (IER3) gene as one of the top‐regulated genes upon inflammatory priming. Small interfering RNA knockdown experiments established IER3 as a novel positive regulator of osteogenic differentiation. Mechanistic analysis further revealed that IER3 deletion significantly downregulated bone marrow stromal cell antigen 2 (BST2) expression and extracellular signal‐related kinase 1/2 (ERK1/2) phosphorylation in hMSCs, suggesting that IER3 regulates osteogenic differentiation through BST2 and ERK1/2 signaling pathway activation. On the basis of these findings, we propose IER3 as a novel therapeutic target to promote hMSC osteoblastogenesis, which might be of high clinical relevance, for example, in patients with osteoporosis or compromised fracture healing. [ABSTRACT FROM AUTHOR]

Diabetes osteoporosis (DOP) is a chronic metabolic bone disease. This study aimed to identify potential biomarkers of DOP and explore their underlying mechanisms through bioinformatics methods and experimental verification. Bioinformatics methods were used to identify differentially expressed genes (DEGs) for DOP based on GEO data and the GeneCards database. GO and KEGG enrichment analyses were used to search the key pathways. The STRING website was used to construct a protein–protein interaction (PPI) network and identify key genes. Then, 50 mg/mL glucose was used to interveneosteoblasts (OBs).CCK‐8 and Alizarin Red staining were used to investigate the proliferation and differentiation changes in OBs. Flowcytometry was used to investigate apoptosis. The membrane protein chip, WB, and RT‐PCR were used to verify the expression of key targets or pathways about DOP. Forty‐two common genes were screened between DOP‐related targets and DEGs. GO and KEGG enrichment analysis showed that DOP was mainly associated with cytokine‐cytokine receptor interactions, and apoptosis. PPI network analysis showed that TNF, IL1A, IL6, IL1B, IL2RA, Fas ligand (FASLG), and Fas cell surface death receptor (FAS) were key up‐regulated genes in the occurrence of DOP. The experiment results show that 50 mg/mL glucose significantly inhibited OBs proliferation but presented an increase in apoptosis. Membrane protein chip, WB, and RT‐PCR‐verified a significantly active in the expression of TNF/FASLG/FAS pathway. High glucose activated the TNF‐α/FAS/FASLG pathway and induced the inflammatory microenvironment and apoptosis, then impaired osteogenic differentiation of OBs. These may be an important mechanism for the occurrence and development of DOP. [ABSTRACT FROM AUTHOR]

To explore the potential mechanisms which O‐linked‐N‐acetylglucosaminylation (O‐GlcNAcylation) regulates osteogenesis, a publicly RNA‐seq dataset was re‐analyzed with literature‐mining and showed the primary targets of O‐GlcNAcylation in osteoblasts are mitochondria/cytoskeleton. Although the O‐GlcNAcylation‐regulated mitochondria/cytoskeleton has been extensively studied, its specific role during osteogenesis remains unclear. To address this, we knocked out Ogt (Ogt‐KO) in MC3T3‐E1 osteoblastic cells. Then, significantly reduced osteoblast differentiation, motility, proliferation, mitochondria–endoplasmic reticulum (Mito–ER) coupling, volume of ER, nuclear tubulins, and oxygen metabolism were observed in Ogt‐KO cells. Through artificial intelligence (AI)‐predicted cellular structures, the time‐lapse live cells imaging with reactive‐oxygen‐species/hypoxia staining showed that lower cell proliferation and altered oxygen metabolism in the Ogt‐KO cells were correlated with the Mito–ER coupling. Bioinformatics analysis, combined with correlated mRNA and protein expression, suggested that Ezh2 and its downstream targets (Opa1, Gsk3a, Wnt3a, Hif1a, and Hspa9) may be involved in O‐GlcNAcylation‐regulated Mito–ER coupling, ultimately impacting osteoblast differentiation. In conclusion, our findings indicate that O‐GlcNAcylation‐regulated osteoblast differentiation is linked to morphological changes in mitochondria, cytoskeleton, and ER, with Ezh2 potentially playing a crucial role. [ABSTRACT FROM AUTHOR]

Background: Craniofacial bone regeneration represents a dynamic area within tissue engineering and regenerative medicine. Central to this field, is the continual exploration of new methodologies for template fabrication, leveraging established bio ceramic materials, with the objective of restoring bone integrity and facilitating successful implant placements. Methods: Photopolymerized templates were prepared using three distinct bio ceramic materials, specifically a wet chemically synthesized bioactive glass and two commercially sourced hydroxyapatite variants. These templates underwent comprehensive characterization to assess their physicochemical and mechanical attributes, employing techniques including Fourier transform infrared spectroscopy, scanning electron microscopy, and nano-computed tomography. Evaluation of their biocompatibility was conducted through interaction with primary human osteoblasts (hOB) and subsequent examination using scanning electron microscopy. Results: The results demonstrated that composite showed intramolecular hydrogen bonding interactions with the photopolymer, while computerized tomography unveiled the porous morphology and distribution within the templates. A relatively higher porosity percentage (31.55 ± 8.70%) and compressive strength (1.53 ± 0.11 MPa) was noted for bioactive glass templates. Human osteoblast cultured on bioactive glass showed higher viability compared to other specimens. Scanning micrographs of human osteoblast on templated showed cellular adhesion and the presence of filopodia and lamellipodia. Conclusion: In summary these templates have the potential to be used for alveolar bone regeneration in critical size defect. Photopolymerization of bioceramics may be an interesting technique for scaffolds fabrication for bone tissue engineering application but needs more optimization to overcome existing issues like the ideal ratio of the photopolymer to bioceramics. [ABSTRACT FROM AUTHOR]

Bone is a dynamic tissue that is constantly remodeled throughout adult life. Recently, it has been shown that bone turnover decreases shortly after food consumption. This process has been linked to the fermentation of non-digestible food ingredients such as inulin by gut microbes, which results in the production of the short-chain fatty acids (SCFAs) acetate, propionate and butyrate. SCFAs exert various metabolic functions, which in part can be explained by activation of G protein-coupled receptors (Gpr) 41 and 43. However, the potential relevance of a SCFA-Gpr41/43 signaling axis for bone metabolism has not been established. The aim of our study is to investigate the role of Gpr41/43 in bone metabolism and osteogenic differentiation of mesenchymal stem cells. For this purpose, we analyzed the skeletal phenotype of wild type controls (WT) and Gpr41/43 double knockout (Gpr41/43 dKO) mice fed either a chow or an inulin-enriched diet. In addition, we isolated bone marrow derived mesenchymal stem cells from WT and Gpr41/43 dKO mice and differentiated them into osteoblasts in the absence or presence of acetate. MicroCT scanning of femoral bones of Gpr41/43 dKO mice revealed a significant increase of trabecular bone volume and trabecular compared to WT controls. Treatment of WT bone marrow-derived osteoblasts with acetate resulted in decreased mineralization and substantial downregulation of bone formation markers such as Phex, Ptgs2 and Col1a1. Notably, this effect was strongly attenuated in differentiated osteoblasts lacking Gpr41/43. Inversely, acetate supplementation resulted in higher levels of adipocyte marker genes including Pparg, Lpl and Adipoq in bone marrow-derived cells from WT mice, an effect blunted in differentiated cells isolated from Gpr41/43 dKO mice. Overall, these data indicate that acetate regulates bone architecture via SCFA-Gpr41/43 signaling by modulating the osteogenic versus adipogenic differentiation of mesenchymal stem cells. [ABSTRACT FROM AUTHOR]

Introduction: Hyaluronic acid as a socket preservation material can absorb large amounts of hydration and regulate appropriate pressure in the surrounding tissue, resulting in expansion of the extracellular space. The socket preservation technique used is a solution to bone loss due to tooth extraction. However, research on the hyaluronic acid (Gengigel) material in sockets with periodontitis has not yet been clearly studied. In this study, we will see an increase in the number of osteoblasts in the alveolar bone after tooth extraction in Wistar rats with periodontitis. Objective: To determine the increase in the number of osteoblasts after administering Hyaluronic acid to the tooth sockets of Wistar rats with periodontitis. Methods: An increase in the number of osteoblast cells was observed using a multi-head light microscope in 5 areas of the preparation at 400x magnification. The measurement results from 5 areas were averaged and a calculation was obtained for one preparation. There are 2 groups that will be used in this research, namely the Control group (without treatment), HA (given Gengigel). Each group was tested at 14 and 21 days after administering the material. Results: The research results showed that the Hyaluronic Acid (Gengigel) group had a higher number of osteoblast cells than the control group. The oneway ANOVA test showed an average significance result of 0.000 (p < 0.05) for all groups. This indicates that there was a significant change in each treatment group. Conclusion: Giving hyaluronic acid to the tooth sockets of Wistar rats with periodontitis can increase the number of osteoblast cells. [ABSTRACT FROM AUTHOR]

Background/Objectives: Although there have been some clinical observations made, the mechanistic effects on bone metabolism of whole-body cryostimulation and high-intensity interval training (HIIT), either alone or in combination, are still debated. Here, we have investigated their effects on circulating osteo-immune and bone metabolic markers (osteopontin, osteocalcin, sclerostin, dikkopf-related protein 1, and fibroblast-growth factor 23) and their potential effects on osteoblast differentiation and function, in vitro, by treating SaOS-2 osteoblast-like cells with the sera obtained from the subjects who had undergone the different interventions or untreated control subjects. Methods: Sixty-seven inactive, overweight-to-obese participants (body mass index = 31.9 ± 5.0 kg·m−2, 42 ± 13 years old) were recruited and randomly assigned to one group: control (CTRL, n = 14), training (HIIT, 6 sessions, n = 13), WBC (CRYO, 10 sessions, n = 17) or training combined with WBC (CRYO-HIIT, n = 23). The interventions lasted 14 days. Results: While circulating markers analysis revealed more protective potential against resorption in HIIT than in WBC alone or combined, gene expression from in vitro analysis showed an induction of late bone metabolic markers in the HIIT group. Conclusions: These data suggest a potentially protective effect of HIIT in bone against resorption, while WBC maintains homeostasis by preventing any resorptive phenomena and limiting any anabolic activity even when stimulated by intensive exercise. [ABSTRACT FROM AUTHOR]

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic bone disorder characterized by episodic flare-ups in connective tissue, which are frequently followed by the formation of heterotopic ossification. The absence of available plasma-soluble biomarkers for flare-ups or heterotopic bone formation poses severe challenges to the monitoring of disease activity to measure or predict disease progression. Recently, 18-fluor-sodium fluoride positron emission tomography/computed tomography ([18F]NaF PET/CT) was introduced as a potential marker for ossifying FOP activity. This review discusses the pharmacokinetics of [18F]NaF in relation to the pathophysiology of FOP, and its use as a marker of local bone metabolism in a variety of bone-related disorders. In addition, the review specifically addresses the applicability of [18F]NaF PET/CT imaging in FOP as a monitoring modality. [ABSTRACT FROM AUTHOR]

Osteoporosis (OP) was long viewed as an inevitable process of aging, due to an imbalance between osteoclast bone resorbing and osteoblast bone formation function, leading to a negative balance in bone remodeling. This leads to low bone mass and increased bone fragility putting the patient at risk for fracture. While this view still holds, a better understanding disclosed that OP can occur at any age, as a comorbidity or a complication of many diseases and treatments. Differentiation, maturation, and function of osteoclasts and osteoblasts are affected by many factors from different morbidities: endocrine, metabolic, mechanical and inflammatory. Inflammatory diseases are often complicated by a generalized bone loss that subsequently leads to OP. Factors such as glucocorticoid treatment, immobilization, malnutrition, and insufficient intake of vitamin D play a role. However, the inflammatory process itself is involved and the resulting bone loss is termed immune‐mediated bone loss. Experiments on animals and on humans, in addition to clinical studies, shed light on the role of inflammation in OP. [ABSTRACT FROM AUTHOR]

Osteoporosis, a prevalent ailment worldwide, compromises bone strength and resilience, particularly afflicting the elderly population. This condition significantly heightens susceptibility to fractures even from trivial incidents, such as minor falls or impacts. A major challenge in diagnosing osteoporosis is the absence of discernible symptoms, allowing osteoporosis to remain undetected until the occurrence of a fracture event. Early symptom detection and swift diagnosis are critical for preventing severe issues related to bone diseases. Assessing bone turnover markers aids in identifying, diagnosing, and monitoring these conditions, guiding treatment decisions. However, conventional techniques for measuring bone mineral density are costly, time-consuming, and require specialized expertise. The integration of sensor technologies into medical practices has transformed how we monitor, diagnose, and treat various health conditions, including bone health and orthopedics. This review aims to provide a comprehensive overview of the current state of sensor technologies used in bone, covering their integration with bone tissue, various applications, recent advancements, challenges, and future directions. [ABSTRACT FROM AUTHOR]

Bone health is ensured by the coordinated action of two types of cells—the osteoblasts that build up bone structure and the osteoclasts that resorb the bone. The loss of balance in their action results in pathological conditions such as osteoporosis. Central to this study is a class of RNA-binding proteins (RBPs) that regulates the biogenesis of miRNAs. In turn, miRNAs represent a critical level of regulation of gene expression and thus control multiple cellular and biological processes. The impact of miRNAs on the pathobiology of various multifactorial diseases, including osteoporosis, has been demonstrated. However, the role of RBPs in bone remodeling is yet to be elucidated. The aim of this study is to dissect the transcriptional landscape of genes encoding the compendium of 180 RBPs in bone cells. We developed and applied a multi-modular integrative analysis algorithm. The core methodology is gene expression analysis using the GENEVESTIGATOR platform, which is a database and analysis tool for manually curated and publicly available transcriptomic data sets, and gene network reconstruction using the Ingenuity Pathway Analysis platform. In this work, comparative insights into gene expression patterns of RBPs in osteoblasts and osteoclasts were obtained, resulting in the identification of 24 differentially expressed genes. Furthermore, the regulation patterns upon different treatment conditions revealed 20 genes as being significantly up- or down-regulated. Next, novel gene–gene associations were dissected and gene networks were reconstructed. Additively, a set of osteoblast- and osteoclast-specific gene signatures were identified. The consolidation of data and information gained from each individual analytical module allowed nominating novel promising candidate genes encoding RBPs in osteoblasts and osteoclasts and will significantly enhance the understanding of potential regulatory mechanisms directing intracellular processes in the course of (patho)physiological bone turnover. [ABSTRACT FROM AUTHOR]

The neuropeptide kisspeptin and its cognate receptor have been extensively studied in reproductive physiology, with diverse and well‐established functions, including as an upstream regulator of pubertal onset, reproductive hormone secretion, and sexual behavior. Besides classical reproduction, both kisspeptin and its receptor are extensively expressed in bone‐resorbing osteoclasts and bone‐forming osteoblasts, which putatively permits direct bone effects. Accordingly, this sets the scene for recent compelling findings derived from in vitro experiments through to in vivo and clinical studies revealing prominent regulatory interactions for kisspeptin signaling in bone metabolism, as well as certain oncological aspects of bone metabolism. Herein, we comprehensively examine the experimental evidence obtained to date supporting the interaction between kisspeptin and bone. A comprehensive understanding of this emerging facet of kisspeptin biology is fundamental to exploiting the future therapeutic potential of kisspeptin‐based medicines as a novel strategy for treating bone‐related disorders. [ABSTRACT FROM AUTHOR]

Recent evidence suggests an association between age‐related osteoporosis and cellular senescence in the bone; however, the specific bone cells that play a critical role in age‐related osteoporosis and the mechanism remain unknown. Results revealed that age‐related osteoporosis is characterized by the loss of osteoblast Men1. Osteoblast‐specific inducible knockout of Men1 caused structural changes in the mice bones, matching the phenotypes in patients with age‐related osteoporosis. Histomorphometrically, Men1‐knockout mice femurs decreased osteoblastic activity and increased osteoclastic activity, hallmarks of age‐related osteoporosis. Loss of Men1 induces cellular senescence via mTORC1 activation and AMPK suppression, rescued by metformin treatment. In bone morphogenetic protein‐indued bone model, loss of Men1 leads to accumulation of senescent cells and osteoporotic bone formation, which are ameliorated by metformin. Our results indicate that cellular senescence in osteoblasts plays a critical role in age‐related osteoporosis and that osteoblast‐specific inducible Men1‐knockout mice offer a promising model for developing therapeutics for age‐related osteoporosis. [ABSTRACT FROM AUTHOR]

Objectives: To explore the role of fibrocytes in the recurrence and calcification of fibrous epulides. Methods: Different subtypes of fibrous epulides and normal gingival tissue specimens were first collected for histological and immunofluorescence analyses to see if fibrocytes were present and whether they differentiated into myofibroblasts and osteoblasts upon stimulated by transforming growth factor‐β1 (TGF‐β1). Electron microscopy and elemental analysis were used to characterize the extracellular microenvironment in different subtypes of fibrous epulides. Human peripheral blood mononuclear cells (PBMCs) were subsequently isolated from in vitro models to mimic the microenvironment in fibrous epulides to identify whether TGF‐β1 as well as the calcium and phosphorus ion concentration in the extracellular matrix (ECM) of a fibrous epulis trigger fibrocyte differentiation. Results: Fibrous epulides contain fibrocytes that accumulate in the local inflammatory environment and have the ability to differentiate into myofibroblasts or osteoblasts. TGF‐β1 promotes fibrocytes differentiation into myofibroblasts in a concentration‐dependent manner, while TGF‐β1 stimulates the fibrocytes to differentiate into osteoblasts when combined with a high calcium and phosphorus environment. Conclusions: Our study revealed fibrocytes play an important role in the fibrogenesis and osteogenesis in fibrous epulis, and might serve as a therapeutic target for the inhibition of recurrence of fibrous epulides. [ABSTRACT FROM AUTHOR]

Modulating macrophage phenotype based on implant surface characteristics, including topography and chemistry, has been employed to enhance osseointegration and long‐term functional outcomes for titanium (Ti)‐based implants. An excessive and/or prolonged M1 macrophage response can lead to damaging immune‐inflammatory reactions, negatively influencing the fate of the implant, and hence, modulating these responses via nanoscale implant surface modification is an emerging paradigm. Herein, an anodized titanium implant surface based on single‐step electrochemical anodization, with preserved underlying microfeatures and superimposed nanopores (50 and 70 nm), compared with irregular rough and microrough (machined‐like) surfaces, is investigated for its effect on the functions of primary macrophages in vitro. Significantly reduced macrophage proliferation and increased tissue‐reparative M2 phenotype polarization are confirmed for the nanopores, which are more pronounced for 70 nm diameter. Moreover, osteoclastogenesis is reduced while osteogenic differentiation of osteoblasts is enhanced for the nanopores (higher for 70 nm pores). Advanced nanoengineered Ti implants can enhance titanium implant tissue integration by modulating the inflammatory response at the implant–cell interface. [ABSTRACT FROM AUTHOR]

Bone formation is tightly modulated by genetically encoded molecular proteins that interact to regulate cellular differentiation and secretion of bony matrix. Many transcription factors are known to coordinate these events by controlling gene transcription within networks. However, not all factors involved are known. Here, we identified a novel function for Zinc Finger Homeobox 3 (Zfhx3), a gene encoding a transcription factor, as a regulator of bone metabolism. We knocked out Zfhx3 conditionally in mice in either chondrocytes or osteoblasts and characterized their bones by micro-CT in 12-week-old mice. We observed a negative effect in linear bone growth in both knockout mice but reduced bone mass only in mice with Zfhx3 deleted in osteoblasts. Loss of Zfhx3 expression in osteoblasts affected trabecular bone mass in femurs and vertebrae in both sexes but influenced cortical bone volume fraction only in females. Moreover, transcriptional analysis of femoral bones in osteoblast Zfhx3 conditional knockout mice revealed a reduced expression of osteoblast genes, and histological evaluation of trabecular bones suggests that Zfhx3 causes changes in bone formation and not resorption. The loss of Zfhx3 causes reductions in trabecular bone area and osteoid volume, but no changes in the expression of osteoclast differentiation markers or number of TRAP stained osteoclasts. These studies introduce Zfhx3 as a relevant factor toward understanding gene regulatory networks that control bone formation and development of peak bone mass. [ABSTRACT FROM AUTHOR]

Skeletal fluorosis is a chronic metabolic bone disease caused by long-term excessive fluoride intake. Abnormal differentiation of osteoblasts plays an important role in disease progression. Research on the mechanism of fluoride-mediated bone differentiation is necessary for the prevention and treatment of skeletal fluorosis. In the present study, a rat model of fluorosis was established by exposing it to drinking water containing 50 mg/L F−. We found that fluoride promoted Runt-related transcription factor 2 (RUNX2) as well as superoxide dismutase 2 (SOD2) and sirtuin 3 (SIRT3) expression in osteoblasts of rat bone tissue. In vitro, we also found that 4 mg/L sodium fluoride promoted osteogenesis-related indicators as well as SOD2 and SIRT3 expression in MG-63 and Saos-2 cells. In addition, we unexpectedly discovered that fluoride suppressed the levels of reactive oxygen species (ROS) and mitochondrial reactive oxygen species (mtROS) in osteoblasts. When SOD2 or SIRT3 was inhibited in MG-63 cells, fluoride-decreased ROS and mtROS were alleviated, which in turn inhibited fluoride-promoted osteogenic differentiation. In conclusion, our results suggest that SIRT3/SOD2 mediates fluoride-promoted osteoblastic differentiation by down-regulating reactive oxygen species. [ABSTRACT FROM AUTHOR]

Cementum shares many properties with bone; however, in contrast to bone, it is not innervated or vascularized and has a limited capacity for remodeling. Osteocytes located in the lacunae-canalicular system of bone tissue play a central role in bone remodeling by communicating with osteoblasts and osteoclasts. Although cementocytes are present in cellular cementum and are morphologically similar to osteocytes, it remains unclear whether they are involved in the dynamic functional regulation of metabolism in cementum. The present study focused on the extracellular vesicles (EVs) secreted by cementocytes and examined their effects on osteoclasts and osteoblasts. EVs were extracted from the mouse cementocyte cell line, IDG-CM6. The effects of EVs on recombinant RANKL-induced osteoclastogenesis and recombinant Bone morphogenetic protein (BMP)-2-mediated osteoblastogenesis were investigated using the mouse osteoclast progenitor cell line, RAW264.7 and mouse pre-osteoblast cell line, MC3T3-E1, respectively. EVs enhanced the formation of tartrate-resistant acid phosphatase activity-positive cells. Real-time PCR revealed that EVs up-regulated the expression of osteoclast-related genes. On the other hand, the cell culture supernatant of cementocytes significantly inhibited the differentiation of osteoclasts. Regarding osteoblastogenesis, EVs suppressed the expression of alkaline phosphatase, bone sialoprotein, and osteocalcin induced by recombinant BMP-2 at the gene and protein levels. A network of cementocytes, osteoblasts, and osteoclasts may exist in cellular cementum, which suggests the involvement of cementocytes in dynamic metabolism of cementum through EVs. [ABSTRACT FROM AUTHOR]

The machining of implants and parts for dental prostheses to eliminate biofilm in the implantoplasty process causes a loss of mechanical properties and also characteristics of the surfaces, making tissue regeneration difficult. In the present work, treatments consisting of elements that can reduce infection, such as citric acid and magnesium, together with elements that can improve cell adhesion and proliferation, such as collagen, are proposed for implant–crown assembly. Titanium, zirconia, composite (PMMA + feldspar) and cobalt–chromium discs were immersed in four different solutions: 25% citric acid, 25% citric acid with the addition of collagen 0.25 g/L, 25% citric acid with the addition of 0.50 g/L and the latter with the addition of 1% Mg (NO3)2. After immersion was applied for 2 and 10 min, the roughness was determined by interferometric microscopy and the contact angle (CA) was evaluated. Human fibroblastic and osteoblastic line cells (HFFs and SaOS-2) were used to determine cell viability and proliferation capacity. Cell binding and cytotoxicity were determined by resazurin sodium salt assay (Alamar Blue) and cell morphology by confocal assay (immunofluorescence F-actin (phalloidin)) after 3 days of incubation. For the evaluation of bacterial activity, the bacterial strains Sptreptococcus gordonii (Gram+) and Pseudomonas aeruginosa (Gram−) were used. The antibacterial properties of the proposed treatments were determined by means of the resazurin sodium salt (Alamar Blue) assay after 1 day of incubation. The treatments considerably decreased the contact angle of the treated samples with respect to the control samples. The treatments endowed the surfaces of the samples with a hydrophilic/super-hydrophilic character. The combination of elements proposed for this study provided cell viability greater than 70%; considering the absence of cytotoxicity, it therefore promotes the adhesion and proliferation of fibroblasts and osteoblasts. In addition, it also endows the surface with antibacterial characteristics against from Gram+ and Gram− bacteria without damaging the cells. These results show that this mouthwash can be useful in oral applications to produce a new passivation layer that favors the hydrophilicity of the surface and promotes cellular activity for the formation of fibroblasts and osteoblasts, as well as showing bactericidal activity. [ABSTRACT FROM AUTHOR]

Osteoporosis and other metabolic bone diseases are prevalent in the aging population. While bone has the capacity to regenerate throughout life, bone formation rates decline with age and contribute to reduced bone density and strength. Identifying mechanisms and pathways that increase bone accrual in adults could prevent fractures and accelerate healing. G protein-gated inwardly rectifying K+ (GIRK) channels are key effectors of G protein-coupled receptor signaling. Girk3 was recently shown to regulate endochondral ossification. Here, we demonstrate that deletion of Girk3 increases bone mass after 18 weeks of age. Male 24-week-old Girk3-/- mice have greater trabecular bone mineral density and bone volume fraction than wildtype (WT) mice. Osteoblast activity is moderately increased in 24-week-old Girk3-/- mice compared to WT mice. In vitro, Girk3-/- bone marrow stromal cells (BMSCs) are more proliferative than WT BMSCs. Calvarial osteoblasts and BMSCs from Girk3-/- mice are also more osteogenic than WT cells, with altered expression of genes that regulate the wingless-related integration site (Wnt) family. Wnt inhibition via Dickkopf-1 (Dkk1) or β-catenin inhibition via XAV939 prevents enhanced mineralization, but not proliferation, in Girk3-/- BMSCs and slows these processes in WT cells. Finally, selective ablation of Girk3 from cells expressing Cre recombinase from the 2.3 kb-Col1a1 promoter, including osteoblasts and osteocytes, is sufficient to increase bone mass and bone strength in male mice at 24 weeks of age. Taken together, these data demonstrate that Girk3 regulates progenitor cell proliferation, osteoblast differentiation, and bone mass accrual in adult male mice. Lay Summary: Osteoporosis is a prevalent, debilitating disease characterized by low bone mass. Identifying pathways that enhance bone formation could lead to therapies that prevent fractures and increase bone mass. In this study, we show that whole-body or osteoblast-specific deletion of G protein-gated inwardly rectifying K+ channel 3 (Girk3) increases bone mass in male adult mice. Osteoblast progenitor cells from Girk3-/- animals are more proliferative and differentiate more quickly into mature osteoblasts. Taken together, we show that Girk3 is a novel regulator of bone mass in adult mice. Graphical Abstract [ABSTRACT FROM AUTHOR]

Background: Tibial bone tunnel widening (TW) is a common postoperative phenomenon after anterior cruciate ligament reconstruction (ACLR). Purpose: To compare the physical, biomechanical, osteoinductive, and histological characteristics of 2 fabricated bioabsorbable interference screws: (1) a modified poly(l -lactide- co - d, l -lactide) and hydroxyapatite (mPLA/HA) screw and (2) a poly(l -lactide- co -glycolide) and β-tricalcium phosphate (PLGA/β-TCP) screw; and to evaluate the effect of the PLA/HA screw on ameliorating postoperative TW in a canine ACLR model. Study Design: Controlled laboratory study. Methods: In vitro, the physical and biomechanical properties of the mPLA/HA and PLGA/β-TCP screws were tested. The osteoinductive activity of the screws was studied by cell experiments. In vivo, ACLR was performed on 48 beagle dogs, divided into the mPLA/HA group and the PLGA/β-TCP group. The femoral and tibial ends of the graft were both fixed with screws. Six animals in each group were sacrificed after live computed tomography (CT) scanning at 1, 3, 6, and 12 months postoperatively. For six knee samples of each group, three knee samples underwent biomechanical testing, and 1 of them, along with the other 3 samples, underwent micro-CT and histological examination to evaluate tibial TW. Results: The mPLA/HA screw exhibited better particle dispersion, bending strength, desirable self-locking effect, and optimized degradation behavior both in vivo and in vitro. Histologically, the mPLA/HA screw had comparative osteoinductive activity. There was good screw-bone integration using the mPLA/HA screw, while most fibrous scar healing was in the PLGA/β-TCP group. There were significant differences between the mPLA/HA and PLGA/β-TCP groups in tibial bone tunnel diameter at the screw body (6 months postoperatively: 5.09 ± 0.44 vs 7.12 ± 0.67; 12 months postoperatively: 4.83 ± 0.27 vs 6.23 ± 0.56; P <.01 for both) and the screw tail (6 months postoperatively: 4.84 ± 0.28 vs 5.97 ± 0.73; 12 months postoperatively: 4.77 ± 0.29 vs 5.92 ± 0.56; P <.01 for both). Conclusion: Compared with the PLGA/β-TCP screw commonly used in clinics at present, the mPLA/HA screw had comparative biosafety and mechanical properties, satisfactory biomechanical properties, and osteoinductive activity in vivo and in vitro. It effectively ameliorated the postoperative tibial TW in a canine ACLR model and increased the quality of screw-bone integration. Clinical Relevance: The good mechanical and biological properties of the mPLA/HA screws may provide an option to reduce the incidence of complications after ACLR. [ABSTRACT FROM AUTHOR]

Only in the past decade have skeletal stem cells (SSCs), a cell type displaying formal evidence of stemness and serving as the ultimate origin of mature skeletal cell types such as osteoblasts, been defined. Here, we discuss a pair of recent reports that identify that SSCs do not represent a single cell type, but rather a family of related cells that each have characteristic anatomic locations and distinct functions tailored to the physiology of those sites. The distinct functional properties of these SSCs in turn provide a basis for the diseases of their respective locations. This concept emerges from one report identifying a distinct vertebral skeletal stem cell driving the high rate of breast cancer metastasis to the spine over other skeletal sites and a report identifying 2 SSCs in the calvaria that interact to mediate both physiologic calvarial mineralization and pathologic calvarial suture fusion in craniosynostosis. Despite displaying functional differences, these SSCs are each united by shared features including a shared series of surface markers and parallel differentiation hierarchies. We propose that this diversity at the level of SSCs in turn translates into a similar diversity at the level of mature skeletal cell types, including osteoblasts, with osteoblasts derived from different SSCs each displaying different functional and transcriptional characteristics reflecting their cell of origin. In this model, osteoblasts would represent not a single cell type, but rather a family of related cells each with distinct functions, paralleling the functional diversity in SSCs. Lay Summary: Only in the past decade have the stem cells in the skeleton been identified. Here, we discuss a pair of recent reports that identify that skeletal stem cells are actually a family of related cells that each have distinct locations and functions. These site-specific skeletal stem cells account for the signature diseases occurring in different regions of the skeleton. Specifically, one of these stem cells forms the spine and establishes that this stem cell drives the high rate of breast cancer metastasis to the spine over other skeletal sites. There are also at least 2 skeletal stem cells in the flat bones of the skull, with mutations alerting how these 2 stem cells "talk" to each other serving as a cause for disorders of premature skull fusion. Despite displaying differences in their function, these stem cells are each united by shared features including a partially shared series marker genes. We also here propose that this diversity at the level of skeletal stem cells translates into a similar diversity in mature skeletal cell types, including osteoblasts. In this model, osteoblasts are not a single cell type, but rather a family of related cells each with distinct functions. [ABSTRACT FROM AUTHOR]

Critically‐sized segmental bone defects represent significant challenges requiring grafts for reconstruction. 3D‐printed synthetic bone grafts are viable alternatives to structural allografts if engineered to provide appropriate mechanical performance and osteoblast/osteoclast cell responses. Novel 3D‐printable nanocomposites containing acrylated epoxidized soybean oil (AESO) or methacrylated AESO (mAESO), polyethylene glycol diacrylate, and nanohydroxyapatite (nHA) were produced using masked stereolithography. The effects of volume fraction of nHA and methacrylation of AESO on interactions of differentiated MC3T3‐E1 osteoblast (dMC3T3‐OB) and differentiated RAW264.7 osteoclast cells with 3D‐printed nanocomposites were evaluated in vitro and compared with a control biomaterial, hydroxyapatite (HA). Higher nHA content and methacrylation significantly improved the mechanical properties. All nanocomposites supported dMC3T3‐OB cells' adhesion and proliferation. Higher amounts of nHA enhanced cell adhesion and proliferation. mAESO in the nanocomposites resulted in greater adhesion, proliferation, and activity at day 7 compared with AESO nanocomposites. Excellent osteoclast‐like cells survival, defined actin rings, and large multinucleated cells were only observed on the high nHA fraction (30%) mAESO nanocomposite and the HA control. Thus, mAESO‐based nanocomposites containing higher amounts of nHA have better interactions with osteoblast‐like and osteoclast‐like cells, comparable with HA controls, making them a potential future alternative graft material for bone defect repair. [ABSTRACT FROM AUTHOR]

Heterotopic ossification (HO) is a pathological process that generates ectopic bone in soft tissues. Hedgehog signaling (Hh signaling) is a signaling pathway that plays an important role in embryonic development and involves three ligands: sonic hedgehog (Shh), Indian hedgehog (Ihh) and desert hedgehog (Dhh). Hh signaling also has an important role in skeletal development. This paper discusses the effects of Hh signaling on the process of HO formation and describes several signaling molecules that are involved in Hh-mediated processes: parathyroid Hormone-Related Protein (PTHrP) and Fkbp10 mediate the expression of Hh during chondrogenesic differentiation. Extracellular signal-regulated kinase (ERK), GNAs and Yes-Associated Protein (YAP) interact with Hh signaling to play a role in osteogenic differentiation. Runt-Related Transcription Factor 2 (Runx2), Mohawk gene (Mkx) and bone morphogenetic protein (BMP) mediate Hh signaling during both chondrogenic and osteogenic differentiation. This paper also discusses possible therapeutic options for HO, lists several Hh inhibitors and explores whether they could serve as emerging targets for the treatment of HO. [ABSTRACT FROM AUTHOR]

This study assessed the effects of titanium (Ti) surface modification with sodium hydroxide (NaOH) associated or not with Naringenin (NA) citrus flavonoid-coating on osteoblastic-like cells (Ob) metabolism. Ti discs were submitted to alkalinization by NaOH solution (5 M, 60 °C) for 24 h; then, the discs were impregnated or not with 100 µg/mL of NA and dried for 1 h at room temperature. The chemical composition, surface topography, and NA release were evaluated. For the biological assays, the discs were placed on 24-well cell culture plates and Ob (Saos-2; ATCC HTB-85) was seeded onto the discs. After different periods, cell adhesion and viability, alkaline phosphatase activity (ALP), and mineralized nodules deposition (MND) were assessed. In addition, cells stimulated with tumor necrosis factor-alpha (TNF-α) were submitted to matrix metalloproteinase (MMP)-2 synthesis and ALP gene expression assessment. Since data presented normal distribution and homogeneity (Shapiro-Wilk; Levene), Student's t-test or one-way ANOVA/post-hoc tests were selected for data analysis (α = 0.05). Higher roughness was observed on Ti discs submitted to NaOH treatment, while the chemical and NA release evaluations indicated the successful adsorption of NA to alkali-treated Ti surface. Higher cell adhesion, cell viability (after 7 days of culture), ALP activity, and MND were observed on Ti NaOH coated with NA compared to the control group (Ti NaOH) (p < 0.05). Moreover, NA coating also promoted decreased MMP-2 synthesis and increased ALP gene expression in the presence of the inflammatory stimulus TNF-α (p < 0.05). The modification of Ti disks with NaOH associated with NA-coating enhanced bone cell metabolism, suggesting that this type of surface modification has a promising potential to accelerate bone repair and formation around dental implants. [ABSTRACT FROM AUTHOR]

Background/Objectives: Degenerative musculoskeletal diseases represent a global health problem due to the progressive deterioration of affected individuals. As a bioactive compound, catechins have shown osteoprotective properties by stimulating osteoblastic cells and inhibiting bone resorption. Thus, this review aimed to address the mechanism of action of catechins on bone tissue. Methods: The search was applied to PubMed without limitations in date, language, or article type. Fifteen articles matched the topic and objective of this review. Results: EGCG (epigallocatechin gallate) and epicatechin demonstrated action on the osteogenic markers RANKL, TRAP, and NF-κβ and expression of BMPs and ALP, thus improving the bone microarchitecture. Studies on animals showed the action of EGCG in increasing calcium and osteoprotegerin levels, in addition to regulating the transcription factor NF-ATc1 associated with osteoclastogenesis. However, it did not show any effect on osteocalcin and RANK. Regarding human studies, EGCG reduced the risk of fracture in a dose-dependent manner. In periodontal tissue, EGCG reduced IL-6, TNF, and RANKL in vitro and in vivo. Human studies showed a reduction in periodontal pockets, gingival index, and clinical attachment level. The action of EGCG on membranes and hydrogels showed biocompatible and osteoinductive properties on the microenvironment of bone tissue by stimulating the expression of osteogenic growth factors and increasing osteocalcin and alkaline phosphate levels, thus promoting new bone formation. Conclusions: EGCG stimulates cytokines related to osteogenes, increasing bone mineral density, reducing osteoclastogenesis factors, and showing great potential as a therapeutic strategy for reducing the risk of bone fractures. [ABSTRACT FROM AUTHOR]

Background: Osteoporosis (OP) is a systemic bone disease characterized by reduced bone mass and deterioration of bone microstructure, leading to increased bone fragility. Platelets can take up and release cytokines, and a high platelet count has been associated with low bone density. Obesity is strongly associated with OP, and adipose tissue can influence platelet function by secreting adipokines. However, the biological relationship between these factors remains unclear. Methods: We conducted differential analysis to identify OP platelet-related plasma proteins. And, making comprehensive analysis, including functional enrichment, protein-protein interaction network analysis, and Friends analysis. The key protein, Tetranectin (TNA/CLEC3B), was identified through screening. Then, we analyzed TNA's potential roles in osteogenic and adipogenic differentiation using multiple RNA-seq data sets and validated its effect on osteoclast differentiation and bone resorption function through in vitro experiments. Results: Six OP-platelet-related proteins were identified via differential analysis. Then, we screened the key protein TNA, which was found to be highly expressed in adipose tissue. RNA-seq data suggested that TNA may promote early osteoblast differentiation. In vitro experiments showed that knockdown of TNA expression significantly increased the expression of osteoclast markers, thereby promoting osteoclast differentiation and bone resorption. Conclusions: We identified TNA as a secreted protein that inhibits osteoclast differentiation and bone resorption. While, it potentially promoted early osteoblast differentiation from bioinformatic results. TNA may play a role in bone metabolism through the adipose-bone axis. [ABSTRACT FROM AUTHOR]

Background: Shortening retention time and minimizing relapse rates are ongoing challenges in orthodontics. This study investigated the effects of natural fulvic acids (FAs) and low-level laser therapy (LLLT) on orthodontic retention in rats. Methods: Seventy-two male Sprague-Dawley rats underwent mesial movement of the left maxillary first molar using a 50 g force via a nickel-titanium tension spring. After three weeks of movement, the rats entered the retention phase with retainer wires and were divided into four groups: Control (no intervention), FAs (80 mg/kg orally daily), LLLT (808 nm laser twice weekly), and FAs + LLLT (both treatments). Retainers were removed on days 7, 14, and 21 for a 3-day relapse assessment. Maxillary impressions were analyzed for relapse rates using 3Shape software, alongside histological and immunohistochemical evaluations of bone morphogenetic protein-2 (BMP-2) expression in periodontal tissues, with differences among groups analyzed using an ordinary two-way analysis of variance (ANOVA). Results: The relapse rate decreased over time, particularly at 10, 17, and 24 days (p < 0.001). The FAs group did not significantly affect relapse rates compared to the control group (p = 0.084). In contrast, both the LLLT and FAs + LLLT groups significantly reduced relapse rate (p < 0.001), with no significant difference between these groups (p = 0.555). Histological examination revealed active osteoclasts on day 10, decreasing by days 17 and 24. The LLLT and FAs + LLLT groups showed less local cementum resorption and better periodontal fiber arrangement. All treatment groups significantly increased BMP-2 expression (P < 0.05) compared to controls. with LLLT and FAs + LLLT differing significantly from FAs (P < 0.001), though no difference was observed between LLLT and FAs + LLLT (P = 0.578). Conclusions: FAs did not significantly reduce relapse rate with retainers, while LLLT effectively reduced relapse rates, showing no additional benefit from combining FAs with LLLT. Both FAs and LLLT increased BMP-2 expression in PDL fibroblasts but with no synergistic effect. [ABSTRACT FROM AUTHOR]

Background: Polyetheretherketone (PEEK) is well known for its excellent physical–chemical properties and biosafety. The study aimed to open up a new method for clinical application of PEEK to reconstruct large-scale bone defects. Methods: A bilayer scaffold for bone regeneration was prepared by combining a sulfonated PEEK barrier framework (SPEEK) with a hydrogel layer loaded with aspirin (ASA) and nano-hydroxyapatite (nHAP) by the wet-bonding of Polydopamine (PDA). Results: The hydrogel was successfully adhered to the surface of SPEEK, resulting in significant changes including the introduction of bioactive groups, improved hydrophilicity, and altered surface morphology. Subsequent tests confirmed that the bilayer scaffold exhibited enhanced compression resistance and mechanical compatibility with bone compared to a single hydrogel scaffold. Additionally, the bilayer scaffold showed stable and reliable bonding properties, as well as excellent biosafety verified by cell proliferation and viability experiments using mouse embryo osteoblast precursor (MC3T3-E1) cells. Conclusion: The bilayer bone regeneration scaffold prepared in this study showed promising potential in clinical application for bone regeneration. [ABSTRACT FROM AUTHOR]

Percutaneous implants osseointegrated into the residuum of a person with limb amputation need to provide mechanical stability and protection against infections. Although significant progress has been made in the biointegration of percutaneous implants, the problem of forming a reliable natural barrier at the level of the surface of the implant and the skin and bone tissues remains unresolved. The use of a microporous implant structure incorporated into the Skin and Bone Integrated Pylon (SBIP) should address the issue by allowing soft and bone tissues to grow directly into the implant structure itself, which, in turn, should form a reliable barrier to infections and support strong osseointegration. To evaluate biological interactions between dermal fibroblasts and MC3T3-E1 osteoblasts in vitro, small titanium discs (with varying pore sizes and volume fractions to achieve deep porosity) were fabricated via 3D printing and sintering. The cell viability MTT assay demonstrated low cytotoxicity for cells co-cultured in the pores of the 3D-printed and sintered Ti samples during the 14-day follow-up period. A subsequent Quantitative Real-Time Polymerase Chain Reaction (RT-PCR) analysis of the relative gene expression of biomarkers that are associated with cell adhesion (α2, α5, αV, and β1 integrins) and extracellular matrix components (fibronectin, vitronectin, type I collagen) demonstrated that micropore sizes ranging from 200 to 500 µm of the 3D printed and sintered Ti discs were favorable for dermal fibroblast adhesion. For example, for representative 3D-printed Ti sample S6 at 72 h the values were 4.71 ± 0.08 (α2 integrin), 4.96 ± 0.08 (α5 integrin), 4.71 ± 0.08 (αV integrin), and 1.87 ± 0.12 (β1 integrin). In contrast, Ti discs with pore sizes ranging from 400 to 800 µm demonstrated the best results (in terms of marker expression related to osteogenic differentiation, including osteopontin, osteonectin, osteocalcin, TGF-β1, and SMAD4) for MC3T3-E1 cells. For example, for the representative 3D sample S4 on day 14, the marker levels were 11.19 ± 0.77 (osteopontin), 7.15 ± 0.29 (osteonectin), and 6.08 ± 0.12 (osteocalcin), while for sintered samples the levels of markers constituted 5.85 ± 0.4 (osteopontin), 4.45 ± 0.36 (osteonectin), and 4.46 ± 0.3 (osteocalcin). In conclusion, the data obtained show the high biointegrative properties of porous titanium structures, while the ability to implement several pore options in one structure using 3D printing makes it possible to create personalized implants for the best one-time integration with both skin and bone tissues. [ABSTRACT FROM AUTHOR]

Simple Summary: This study aimed to determine the gene expression profiles associated with chemotherapeutic responses in conventional osteosarcomas (COS) within South Africa. We observed a significant downregulation in the ATP binding cassette subfamily C members (ABCC3 and ABCB1-p-glycoprotein), excision repair cross-complimenting group 1 (ERCC 1), replication factor C subunit 1 (RFC1), and tumour protein 53 (p53) genes in the COS tumours compared to the healthy donors. Furthermore, an upregulated ERCC1 gene expression level predicted a poor chemotherapeutic response. Additionally, the predictors of COS chemotherapeutic response comprised age, chondroblastic and osteoblastic histological subtypes, and ABCC3, ERCC1, and RFC1 gene expression. Background: We determined the predictive gene expression profiles associated with chemo-response in conventional osteosarcomas (COS) within South Africa. Materials and methods: In 28 patients, we performed an RNA extraction, cDNA synthesis, and quantitative analysis using the RT-PCR 2−∆∆CT method to determine the fold change in gene expression alongside GAPDH (housekeeping gene). Results: We observed a significant downregulation in the mRNA expression profiles of ABCB1-p-glycoprotein (p = 0.0007), ABCC3 (p = 0.002), ERCC1 (p = 0.007), p-53 (p = 0.007), and RFC1 (p = 0.003) in the COS patients compared to the healthy donors. Furthermore, ABCB1-p-glycoprotein (p = 0.008) and ABCC3 (p = 0.020) exhibited a significant downregulation in the COS tumour tissues when compared to the healthy donors. In our univariate logistic regression, the predictors of chemotherapeutic response comprised ERCC1 [restricted cubic spline (RCS) knot: OR −0.27; CI −0.504 to −0.032; p = 0.036]; osteoblastic subtype [OR −0.36; CI −0.652 to −0.092; p = 0.026); fibroblastic subtype [OR 0.91; CI 0.569 to 1.248; p < 0.001]; and mixed subtype [OR 0.53; CI 0.232 to 0.032; p = 0.032]. In our multivariable logistic regression, the significant predictors of chemotherapeutic response comprised age [RCS knot: OR −2.5; CI −3.616 to −1.378; p = 0.022]; ABCC3 [RCS knot: OR 0.67; CI 0.407 to 0.936, p = 0.016]; ERCC1 [RCS knot: OR 0.57; CI 0.235 to 0.901; p = 0.044]; RFC1 [RCS knot: OR −1.04; CI −1.592 to −0.487; p = 0.035]; chondroblastic subtype [OR −0.83; CI −1.106 to −0.520; p = 0.012]; and osteoblastic subtype [OR −1.28; CI −1.664 to −0.901; p = 0.007]. Conclusions: In this South African cohort, we observed the unique gene expression profiles of osteosarcoma tumourigenesis and chemotherapeutic responses. These may serve as prognostication and therapeutic targets. Larger-scale research is needed on the African continent. [ABSTRACT FROM AUTHOR]