Significant Increase In Bone Volume Research Articles

BMP3b regulates bone mass by inhibiting BMP signaling

Bone morphogenetic protein 3b (BMP3b), also known as growth differentiation factor 10 (GDF10), is a non-osteogenic BMP highly expressed in the skeleton. Although in vitro studies have shown that BMP3b suppresses osteoblast differentiation, the physiological role of BMP3b in regulating bone mass in vivo remains unknown. Here, we show that BMP3b deletion in mice leads to a high bone mass phenotype via an unexpected novel mechanism involving de-repression of canonical BMP/Smad signaling. BMP3b null mice were viable, and exhibited no significant difference in body size compared to wildtype control. Trabecular bone parameters assessed by histomorphometry and μCT, revealed a significant increase in bone volume and bone mineral density. Expression of osteoblast-differentiation genes were elevated in bone tissue of BMP3b null mice, whereas expression of osteoclast-related genes remained unchanged. Consistent with this, Bmp3b was highly expressed in osteoblasts relative to osteoclast cells. Ex-vivo culture of primary bone marrow mesenchymal stem cells (BMSCs) and primary bone marrow-derived osteoclasts revealed that inactivation of BMP3b enhances osteogenesis without affecting osteoclastogenesis. Mechanistically, we found that BMP3b suppressed BMP4-induced Smad1/5 phosphorylation and inhibited the activity of a BMP4-driven Id-1 luciferase reporter. Protein-protein interaction assays revealed that BMP3b competitively interfered with the association of BMP4 and BMP type I receptors. These findings suggest that BMP3b regulates bone mass by acting as a BMP receptor antagonist. Thus, maintenance of bone mass involves antagonism of canonical BMP/Smad signaling by a member of the BMP family.

Bone regeneration driven by a nano-hydroxyapatite/chitosan composite bioaerogel for periodontal regeneration.

The most recent progress in reconstructive therapy for the management of periodontitis and peri-implantitis bone defects has relied on the development of highly porous biodegradable bioaerogels for guided bone regeneration. The objective of this work was to evaluate in vitro the osteoinduction of periodontal-originating cells (human dental follicle mesenchymal cells, DFMSCs) promoted by a nano-hydroxyapatite/chitosan (nHAp/CS) bioaerogel, which was purified and sterilized by a sustainable technique (supercritical CO2). Moreover, the in vivo bone regeneration capacity of the nHAp/CS bioaerogel was preliminarily assessed as a proof-of-concept on a rat calvaria bone defect model. The quantification of DNA content of DFMSCs seeded upon nHAp/CS and CS scaffolds (control material) showed a significant increase from the 14th to the 21st day of culture. These results were corroborated through confocal laser scanning microscopy analysis (CLSM). Furthermore, the alkaline phosphatase (ALP) activity increased significantly on the 21st day, similarly for both materials. Moreover, the presence of nHAp promoted a significantly higher expression of osteogenic genes after 21days when compared to CS scaffolds and control. CLSM images of 21days of culture also showed an increased deposition of OPN over the nHAp/CS surface. The in vivo bone formation was assessed by microCT and histological analysis. The in vivo evaluation showed a significant increase in bone volume in the nHAp/CS test group when compared to CS and the empty control, as well as higher new bone formation and calcium deposition within the nHAp/CS structure. Overall, the present study showed that the nHAp/CS bioaerogel could offer a potential solution for periodontal and peri-implant bone regeneration treatments since the in vitro results demonstrated that it provided favorable conditions for DFMSC proliferation and osteogenic differentiation, while the in vivo outcomes confirmed that it promoted higher bone ingrowth.

Alendronate treatment rescues the effects of compressive loading of TMJ in osteogenesis imperfecta mice

BackgroundOsteogenesis imperfecta (OI) is a genetic disorder of connective tissue caused by mutations associated with type I collagen, which results in defective extracellular matrix in temporomandibular joint (TMJ) cartilage and subchondral bone. TMJ is a fibrocartilaginous joint expressing type I collagen both in the cartilage and the subchondral bone. In the present study the effects of alendronate and altered loading of the TMJ was analyzed both in male and female OI mice.Materials and methodsForty-eight, 10-weeks-old male and female OI mice were divided into 3 groups: (1) Control group: unloaded group, (2) Saline + Loaded: Saline was injected for 2 weeks and then TMJ of mice was loaded for 5 days, (3) alendronate + loaded: alendronate was injected for 2 weeks and then TMJ of mice was loaded for 5 days. Mice in all the groups were euthanized 24-h after the final loading.ResultsAlendronate pretreatment led to significant increase in bone volume and tissue density. Histomorphometrically, alendronate treatment led to increase in mineralization, cartilage thickness and proteoglycan distribution. Increased mineralization paralleled decreased osteoclastic activity. Our immunohistochemistry revealed decreased expression of matrix metallopeptidase 13 and ADAM metallopeptidase with thrombospondin type 1 motif 5.ConclusionThe findings of this research support that alendronate prevented the detrimental effects of loading on the extracellular matrix of the TMJ cartilage and subchondral bone.

Effectiveness of pre-alveolar bone graft orthodontics for patients with non-syndromic complete unilateral cleft lip, alveolus and palate: A systematic review and meta-analysis.

Non-syndromic complete unilateral cleft lip, alveolus and palate patients who have had ABG surgery. Orthodontics prior to ABG. Comparison: No orthodontic treatment prior to ABG. Successful eruption of permanent canines. All articles were screened for the title, abstract and full text independently and in duplicate by 2 reviewers. The quality assessment of RCT was performed using Cochrane's risk of bias tool and the CCT was assessed using ROBINS-I tool. Of the 904 studies retrieved in the search, one RCT and one CCT were included. Both studies were judged as high risk of bias. The results from one study showed a statistically significant increase in bone volume and decreased bone defect post-ABG in the orthodontic treatment group. However, there was no difference with respect to other variables. Both included studies were of low quality. There is not enough evidence to recommend orthodontic treatment pre-ABG for patients with complete unilateral cleft lip, alveolus and palate. Future high-quality studies are required to inform patients and clinicians about the effectiveness of pre-graft orthodontic treatment.

Addition of heparin binding sites strongly increases the bone forming capabilities of BMP9 in vivo.

Bone Morphogenetic proteins (BMPs) like BMP2 and BMP7 have shown great potential in the treatment of severe bone defects. In recent in vitro studies, BMP9 revealed the highest osteogenic potential compared to other BMPs, possibly due to its unique signaling pathways that differs from other osteogenic BMPs. However, in vivo the bone forming capacity of BMP9-adsorbed scaffolds is not superior to BMP2 or BMP7. In silico analysis of the BMP9 protein sequence revealed that BMP9, in contrast to other osteogenic BMPs such as BMP2, completely lacks so-called heparin binding motifs that enable extracellular matrix (ECM) interactions which in general might be essential for the BMPs' osteogenic function. Therefore, we genetically engineered a new BMP9 variant by adding BMP2-derived heparin binding motifs to the N-terminal segment of BMP9's mature part. The resulting protein (BMP9 HB) showed higher heparin binding affinity than BMP2, similar osteogenic activity in vitro and comparable binding affinities to BMPR-II and ALK1 compared to BMP9. However, remarkable differences were observed when BMP9 HB was adsorbed to collagen scaffolds and implanted subcutaneously in the dorsum of rats, showing a consistent and significant increase in bone volume and density compared to BMP2 and BMP9. Even at 10-fold lower BMP9 HB doses bone tissue formation was observed. This innovative approach of significantly enhancing the osteogenic properties of BMP9 simply by addition of ECM binding motifs, could constitute a valuable replacement to the commonly used BMPs. The possibility to use lower protein doses demonstrates BMP9 HB's high translational potential.

Rker-050, a Novel Activin Receptor Type II Ligand Trap, Rescued Anemia and Reduced Bone Loss in a Mouse Model of Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS) are a group of hematopoietic stem cell diseases characterized by ineffective hematopoiesis, peripheral blood cytopenias and dysplasias resulting from molecular defects and dysregulated signaling that govern the regulation of hematopoietic stem and progenitor cells (HSPCs). Crosstalk between cells in the osteo-hematopoietic (O-H) niche are necessary for the maintenance and self-renewal of HSPCs. Additionally, MDS patients commonly present with osteopenia and osteoporosis further demonstrating the critical interplay between the O-H cells in the bone marrow (BM). Alterations in the interaction between bone precursors and HSPCs within the BM microenvironment have been linked to the pathophysiology of MDS. Transforming growth factor-beta (TGF-β) superfamily signaling is instrumental in regulating both normal hematopoiesis and bone homeostasis. KER-050 is a modified investigational activin receptor type II ligand trap designed to bind and inhibit select TGF-β superfamily ligands, including activin A, activin B, GDF8 and GDF11, to promote hematopoiesis and bone growth. Preclinical studies have demonstrated that treatment with a research form of KER-050 (RKER-050) increased bone mass and multi-lineage hematopoiesis in healthy mice. To further investigate the effect of RKER-050 on hematopoietic and bone systems within the MDS disease setting, the NUP98/HOX13 MDS mouse model was utilized. Five-month-old male MDS mice were treated with vehicle (VEH, n=10), or RKER-050 (050, 7.5mg/kg, n=9) once weekly for 6 weeks along with a wildtype vehicle (WT, n=7) control group. To assess changes in hematological parameters, CBCs were measured, bone marrow (BM) and splenocytes were characterized via flow cytometry. To investigate alterations of bone microarchitecture, femurs were assessed using microCT imaging. MDS-VEH mice showed a reduction in red blood cells (RBC; 30%), hemoglobin (HGB; 19%), hematocrit (HCT; 13%) and reticulocytes (Retic; 16%) compared to WT; all hallmarks of human MDS. Relative to MDS-VEH, MDS-050 showed increased RBC, HGB, HCT and Retic by 25%, 13%, 10% and 39%, respectively. The erythroid Ter119 positive (Ter119+) cell population was slightly reduced in BM and spleen of MDS-VEH mice compared to WT. The BM Ter119+ cells were unchanged in the MDS-050 cohort relative to MDS-VEH; however, the spleen Ter119+ cells were increased similar to the levels in WT mice. Specific erythroid precursor populations were increased in BM and spleen of MDS-VEH mice versus WT, consistent with the ineffective erythropoiesis in this model. Interestingly, erythroid precursor populations were similar between MDS-VEH and MDS-050, yet MDS-050 exhibited recovered RBC production, suggesting an increase in maturation of erythroid precursors with RKER-050 treatment. Given the importance of the O-H niche in regulating hematopoiesis along with osteoporosis and osteopenia being comorbidities of MDS, bone microarchitecture was also analyzed. In the distal femoral trabecular bone, relative to WT mice, MDS-VEH mice had significant increases in trabecular (Tb) separation (33%) and decreases in Tb number (28%), changes that are consistent with reduced bone strength. Bone volume (34%) and bone volume fraction (26%) decreased but were not statistically significant. MDS-050 treated mice had a significant increase in bone volume (71%), higher bone volume fraction (88%), increased Tb number (44%) and decreased Tb spacing (20%) compared to the MDS-VEH, supporting RKER-050's role in improving bone mass. Taken together, these results suggest that RKER-050 can mitigate anemia and bone loss in an MDS mouse model, potentially by rebalancing hematopoiesis and bone turnover. In particular, the ability of RKER-050 to improve bone microarchitecture within the trabecular region, where the O-H niche resides, could be an important step in reestablishing healthy blood cell production. Furthermore, exploratory PD data from an ongoing phase 2 clinical trial demonstrate improved blood parameters with KER-050 treatment, including platelet and RBC in MDS patients (Tan et al. 2022 EHA #P776), and changes in bone biomarkers are currently being evaluated in the study. KER-050, thus, represents a potentially promising approach for patients with MDS and other hematological diseases, including myelofibrosis, where ineffective hematopoiesis and defects in bone homeostasis occur.

Anabolic Response of Intermittent Parathyroid Hormone and Alendronate on the Osteochondral Tissue of TMJ.

To characterize the effects of parathyroid hormone (PTH) and alendronate (Alend) on the osteochondral tissue of temporomandibular joint (TMJ). Ninety-six male and female transgenic reporter mice, 4 to 5 weeks old were divided into 6 groups: (1) Control group: Saline was injected daily for 14 days; (2) PTH: PTH was injected daily for 14 days; (3) Alend: Alend was injected every alternate days for 14 days; (4) Combined PTH and Alend: PTH was injected daily and Alend injected every alternate days for 14 days; (5) PTH then Alend: PTH was injected daily for 14 days followed by Alend injections in alternate days for 14 days; and (6) PTH wait Alend: PTH was injected daily for 14 days. There was a waiting period of 1 week before administration of Alend in alternate days for 14 days. Mice were injected with 5-ethnyl-2'-deoxyuridine (EdU), 48 and 24 hours prior to euthanization. There was significant increase in bone volume and decrease in osteoclastic activity in groups in which Alend was administered after PTH in both gender. There was significant increase in cartilage thickness with PTH or Alend alone in females, whereas in males, PTH alone led to increase in cartilage thickness. Chondrocyte apoptosis was significantly decreased with PTH or Alend alone in both male and female. Matrix metallopeptidase 13, and aggreganase-2 (ADAMTS5) expression were significantly decreased with PTH and Alend alone in both gender. PTH and Alend administration causes anabolic effects in the osteochondral tissue of TMJ.

Localized Ridge Augmentation with Mandibular Block Autograft and Guided Bone Regeneration: A Case Report

Numerous alveolar ridge defects resulted post extraction, long standing periodontal disease often require surgical intervention before prosthetic rehabilitation. On the other hand, alveolar bone defects affect the prognosis of dental implants and as a result, their reconstruction is must. Although a wide variety of options have been invented, autogenous bone is still the gold standard and has been yielding promising results. The authors report a case of localized alveolar ridge augmentation using autogenous chin block graft in conjunction with other bone substitutes for prosthetic rehabilitation of lower anterior region. Initially, the alveolar ridge was knife edge and the bone volume was insufficient for placement of dental implant. The CBCT analysis post 6 months shows significant increase in bone volume that was now suitable for prosthetic rehabilitation of the edentulous space.

Effect of Demineralized Bone Matrix, Bone Marrow Mesenchymal Stromal Cells, and Platelet-Rich Plasma on Bone Tunnel Healing After Anterior Cruciate Ligament Reconstruction: A Comparative Micro-Computed Tomography Study in a Tendon Allograft Sheep Model

Background:The effect of demineralized bone matrix (DBM), bone marrow–derived mesenchymal stromal cells (BMSCs), and platelet-rich plasma (PRP) on bone tunnel healing in anterior cruciate ligament reconstruction (ACLR) has not been comparatively assessed.Hypothesis:These orthobiologics would reduce tunnel widening, and the effects on tunnel diameter would be correlated with tunnel wall sclerosis.Study Design:Controlled laboratory study.Methods:A total of 20 sheep underwent unilateral ACLR using tendon allograft and outside-in interference screw fixation. The animals were randomized into 4 groups (n = 5 per group): Group 1 received 4mL of DBM paste, group 2 received 10 million BMSCs in fibrin sealant, group 3 received 12 mL of activated leukocyte-poor platelet-rich plasma, and group 4 (control) received no treatment. The sheep were euthanized after 12 weeks, and micro-computed tomography scans were performed. The femoral and tibial tunnels were divided into thirds (aperture, midportion, and exit), and the trabecular bone structure, bone mineral density (BMD), and tunnel diameter were measured. Tunnel sclerosis was defined by a higher bone volume in a 250-µm volume of interest compared with a 4-mm volume of interest surrounding the tunnel.Results:Compared with the controls, the DBM group had a significantly higher bone volume fraction (bone volume/total volume [BV/TV]) (52.7% vs 31.8%; P = .020) and BMD (0.55 vs 0.47 g/cm3; P = .008) at the femoral aperture and significantly higher BV/TV at femoral midportion (44.2% vs 32.9%; P = .038). There were no significant differences between the PRP and BMSC groups versus controls in terms of trabecular bone analysis or BMD. In the controls, widening at the femoral tunnel aperture was significantly greater than at the midportion (46.7 vs 41.7 mm2; P = .034). Sclerosis of the tunnel was common and most often seen at the femoral aperture. In the midportion of the femoral tunnel, BV/TV (r = 0.52; P = .019) and trabecular number (r S = 0.50; P = .024) were positively correlated with tunnel widening.Conclusion:Only DBM led to a significant increase in bone volume, which was seen in the femoral tunnel aperture and midportion. No treatment significantly reduced bone tunnel widening. Tunnel sclerosis in the femoral tunnel midportion was correlated significantly with tunnel widening.Clinical Relevance:DBM might have potential clinical use to enhance healing in the femoral tunnel after ACLR.

Comparing the Effects of Alpha-Tocopherol and Tocotrienol Isomers on Osteoblasts hFOB 1.19 Cultured on Bovine Bone Scaffold

Tocotrienol mixtures have been shown to exert anabolic actions on the skeletal system in animal studies, but it is unclear which tocotrienol isomer shows the most prominent effects. This study aims to investigate the most active tocotrienol isomers using hFOB 1.19 human osteoblasts cultured on a bovine bone scaffold. The bovine trabecular bone was sectioned, demineralised and freeze-dried to form the scaffold. hFOB 1.19 osteoblasts were cultured on the bone scaffolds in humidified condition at 37 °C and 5% carbon dioxide with vitamin E isomers (alpha-, beta-, gamma-, delta-tocotrienol and alpha-tocopherol). The cell differentiation capacity of tocotrienol isomers was investigated through morphological observation, alkaline phosphatase (ALP) activity and osteocalcin expression. Changes in the bone scaffolds were determined using histomorphometry methods. Osteoblast culture treated with gamma- and delta-tocotrienols showed a significant increase in ALP activity and osteocalcin expression. Bone structural histomorphometry analysis showed that bone scaffolds treated with gamma- and delta-tocotrienol showed significant increases in bone volume and trabecular thickness. In conclusion, gamma- and delta-tocotrienol show the most prominent bone anabolic effects by increasing osteoblast differentiation and enhancing bone microstructure.

Spongostan™ Leads to Increased Regeneration of a Rat Calvarial Critical Size Defect Compared to NanoBone® and Actifuse.

Bone substitute materials are becoming increasingly important in oral and maxillofacial surgery. Reconstruction of critical size bone defects is still challenging for surgeons. Here, we compared the clinically applied organic bone substitute materials NanoBone® (nanocrystalline hydroxyapatite and nanostructured silica gel; n = 5) and Actifuse (calcium phosphate with silicate substitution; n = 5) with natural collagen-based Spongostan™ (hardened pork gelatin containing formalin and lauryl alcohol; n = 5) in bilateral rat critical-size defects (5 mm diameter). On topological level, NanoBone is known to harbour nanopores of about 20 nm diameter, while Actifuse comprises micropores of 200–500 µm. Spongostan™, which is clinically applied as a haemostatic agent, combines in its wet form both nano- and microporous topological features by comprising 60.66 ± 24.48 μm micropores accompanied by nanopores of 32.97 ± 1.41 nm diameter. Micro-computed tomography (µCT) used for evaluation 30 days after surgery revealed a significant increase in bone volume by all three bone substitute materials in comparison to the untreated controls. Clearly visual was the closure of trepanation in all treated groups, but granular appearance of NanoBone® and Actifuse with less closure at the margins of the burr holes. In contrast, transplantion of Spongostan™ lead to complete filling of the burr hole with the highest bone volume of 7.98 ccm and the highest bone mineral density compared to all other groups. In summary, transplantation of Spongostan™ resulted in increased regeneration of a rat calvarial critical size defect compared to NanoBone and Actifuse, suggesting the distinct nano- and microtopography of wet Spongostan™ to account for this superior regenerative capacity. Since Spongostan™ is a clinically approved product used primarily for haemostasis, it may represent an interesting alternative in the reconstruction of defects in the maxillary region.

The Effect of Megakaryocytes and Platelets Derived from Human-Induced Pluripotent Stem Cells on Bone Formation.

Introduction Platelet-rich plasma (PRP) is drawing attention as a substance that can promote bone formation. The growth factors present in PRP are stable for a long time after freeze-drying. However, the effects of PRP are inconsistent, and its effects on bone union in spinal surgery remain controversial. The immortalized megakaryocyte cell lines (imMKCLs) derived from human-induced pluripotent stem cells (hiPSCs) have been developed to produce numerous stable and clinically functional platelets. In this study, growth factors present in freeze-dried hiPSC-derived imMKCLs and platelets (iPS-MK/Plts) were evaluated, and their ability to promote bone formation was examined using a rat lumbar artificial bone grafting model. Methods We prepared freeze-dried iPS-MK/Plts and quantified their growth factors by enzyme-linked immunosorbent assays. Surgical grafting of artificial bone to the lumbar transverse processes was performed in 8-week-old female rats, which were divided into two groups: artificial bone graft (control) and artificial bone graft plus freeze-dried iPS-MK/Plts (iPS group). Transplantation was performed only on the left side. Eight weeks after the surgery, we captured computed tomography images and compared bilateral differences in the bone volume of the graft site in each rat. We also compared the left side/right side bone volume ratio between the two groups. Results The freeze-dried iPS-MK/Plts contained numerous growth factors. While there was no significant increase in bone volume on the transplanted side than that on the non-grafted side in the control group, bone volume significantly increased on the transplanted side in the iPS group, as evidenced by augmented mean left/right bone volume ratio of the iPS group compared with that of the control group. But the new bone observed in the iPS group was histologically normal. Conclusions Freeze-dried hiPSC-derived MKCLs and platelets contain several stable growth factors and have the potential for promoting new bone formation.

Bone Mineral Is More Heterogeneously Distributed in the Femoral Heads of Osteoporotic and Diabetic Patients: A Pilot Study.

ABSTRACTOsteoporosis is associated with systemic bone loss, leading to a significant deterioration of bone microarchitecture and an increased fracture risk. Although recent studies have shown that the distribution of bone mineral becomes more heterogeneous because of estrogen deficiency in animal models of osteoporosis, it is not known whether osteoporosis alters mineral distribution in human bone. Type 2 diabetes mellitus (T2DM) can also increase bone fracture risk and is associated with impaired bone cell function, compromised collagen structure, and reduced mechanical properties. However, it is not known whether alterations in mineral distribution arise in diabetic (DB) patients’ bone. In this study, we quantify mineral content distribution and tissue microarchitecture (by μCT) and mechanical properties (by compression testing) of cancellous bone from femoral heads of osteoporotic (OP; n = 10), DB (n = 7), and osteoarthritic (OA; n = 7) patients. We report that though OP cancellous bone has significantly deteriorated compressive mechanical properties and significantly compromised microarchitecture compared with OA controls, there is also a significant increase in the mean mineral content. Moreover, the heterogeneity of the mineral content in OP bone is significantly higher than controls (+25%) and is explained by a significant increase in bone volume at high mineral levels. We propose that these mineral alterations act to exacerbate the already reduced bone quality caused by reduced cancellous bone volume during osteoporosis. We show for the first time that cancellous bone mineralization is significantly more heterogeneous (+26%) in patients presenting with T2DM compared with OA (non‐DB) controls, and that this heterogeneity is characterized by a significant increase in bone volume at low mineral levels. Despite these mineralization changes, bone microarchitecture and mechanical properties are not significantly different between OA groups with and without T2DM. Nonetheless, the observed alterations in mineral heterogeneity may play an important tissue‐level role in bone fragility associated with OP and DB bone. © 2019 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Pretreatment with Pamidronate Decreases Bone Formation but Increases Callus Bone Volume in a Rat Closed Fracture Model.

Clinical concerns have been raised over prior exposure to bisphosphonates impairing fracture healing. To model this, groups of male Wistar rats were assigned to saline control or treatment groups receiving 0.15mg/kg (low dose), 0.5mg/kg (medium dose), and 5 mg/kg (high dose) Pamidronate (PAM) twice weekly for 4weeks. At this point, closed fractures were made using an Einhorn apparatus, and bisphosphonate dosing was continued until the experimental endpoint. Specimens were analyzed at 2 and 6weeks (N = 8 per group per time point). Twice weekly PAM dosing was found to have no effect on early soft callus remodeling at 2weeks post fracture. At this time point, the highest dose PAM group gave significant increases in bone volume (+ 10%, p < 0.05), bone mineral content (+ 30%, p < 0.01), and bone mineral density (+ 10%, p < 0.01). This PAM dosing regimen showed more substantive effects on hard callus at 6weeks post fracture, with PAM treatment groups showing + 46-79% increased bone volume. Dynamic bone labeling showed reduced calcein signal in the PAM-treated calluses (38-63%, p < 0.01) and reduced MAR (32-49%, p < 0.01), suggesting a compensatory reduction in bone anabolism. These data support the concept that bisphosphonates lead to profound decreases in bone turnover in fracture repair, however, this does not affect soft callus remodeling.

The role of magnesium ions in bone regeneration involves the canonical Wnt signaling pathway

Magnesium (Mg)-based implants have become of interest to both academia and the medical industry. The attraction largely is due to Mg’s biodegradability and ability to enhance bone healing and formation. However, the underlying mechanism of how Mg regulates osteogenesis is still unclear. Based on our previous in vivo and molecular signaling work demonstrating the osteogenic effect of Mg, the current study aims to extend this work at the molecular level especially that we also observed and quantified mineral deposits in the bone marrow space in a rabbit ulna fracture model with Mg plates and screws. Histological analysis and quantitative results of micro-CT showed mineralized deposition and a significant increase in bone volume at 8 weeks and 16 weeks post-operative. These in vivo results led us to focus on studying the effect of Mg2+ on human bone marrow stromal cells (hBMSCs). The data presented in this manuscript demonstrate the activation of the canonical Wnt signaling pathway in hBMSCs when treated with 10 mM Mg2+. With additional Mg2+ present, the protein expression of active β-catenin was significantly increased to a level similar to that of the positive control. Immunocytochemistry and the increased expression of LEF1 and Dkk1, downstream target genes that are controlled directly by active β-catenin, demonstrated the protein translocation and the activation of transcription. Taken together, these data suggest that Mg2+ induces an osteogenic effect in the bone marrow space by activating the canonical Wnt signaling pathway, which in turn causes BMSCs to differentiate toward the osteoblast lineage. Statement of SignificanceMagnesium (Mg)-based alloys are being studied to be used in the field of implantable medical devices due to its natural biodegradability and the potential ability to promote bone regeneration. Despite many in vivo studies that demonstrated an increased new bone growth by implanting Mg-based devices, the underlying mechanism of this effect is still unclear. In order to safely use Mg-based implants on human and better control the osteogenic effect, it is necessary to understand the corresponding cellular response in the targeted area. The present study provides the rationale to study Mg ions on bone marrow stromal cells and shows the activation of canonical Wnt signaling pathway that promotes osteogenesis by in vivo and in vitro approaches.

Development and characterization of a humanized mouse model of osteoarthritis

Purpose: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Osteoarthritis (OA) is mainly characterized by cartilage breakdown and synovial inflammation. While OA is one of the most common rheumatic disorders, no efficient disease-modifying drugs still exist today. Mesenchymal stem cells (MSC) have generated significant medical considerations since they exhibit tissue-regenerative properties through the secretion of bioactive factors. Phase I-II clinical trials have shown that intra-articular (IA) injection of MSC is well-tolerated and exhibited promising clinical results. However, MSC display a poor survival and engraftment rate in vivo. Using encapsulating systems could protect MSC and maintain their bioactivity after intra-articular injection. Considering the pivotal role of inflammation and immune system in OA and with regards to the differences between MSC-mediated immunomodulation in human and mice (12), the development of sophisticated animal models that could more closely mimic the human immune system, could be instrumental. Amongst these models, immunologically humanized mice may represent a preclinical bridge between in vitro studies and the in vivo evaluation of human stem cells in OA. Our objective is therefore to develop an experimental model of osteoarthritis by destabilizing the medial meniscus (DMM) in humanized mice.> Methods: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <4 weeks-old NOD/LtSz-scid IL2Rγnull (NSG) mice were humanized as follow. After irradiation (1,5Gy), 5.106 CD34+ human hematopoietic stem cells were injected intravenously. 120 days after the humanization procedure, the level of human immune cells CD45+ (leukocytes), CD3+ (T lymphocytes), CD19+ (B lymphocytes), CD56+ (NK cells) and CD14+ (macrophages) were evaluated by flow cytometry. Humanized NSG mice then had undergone a DMM or a SHAM surgery. 6 and 12 weeks after OA induction the development of osteoarthritis was assessed histologically by safranin-o staining, OA score and histomorphometric analysis of the knees. In addition, to evidence the presence of human immune cells in the joint, Immunohistochemicals stainings for human CD45+, CD3+, CD19+ et CD56+ and CD14+ cells have also been performed.> Results: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Our results showed that 120 days after the humanization procedure, NSG mice exhibit engraftment level (hCD45+) from 10% to 75 % and the presence of human T and B lymphocytes as well as macrophages and NK cells in blood and spleen. Our histological results show that the DMM procedure induce a significant increase in OA score at 12 weeks compared to sham mice. Our histomorphometric results also evidence a significant increase in bone volume of the anterior medial meniscus. However, no significant differences have been observed between DMM and Sham mice on the OA score at 6 weeks and on the subchondral bone morphometry at 6 and 12 weeks’ post-surgery. Immunohistochemical analyses have however revealed the presence of human B lymphocytes and macrophages in the subchondral bone marrow but not in the synovial membrane at 6 and 12 weeks post-surgery.> Conclusions: Normal 0 21 false false false FR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tableau Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman",serif; border:none;} <Finally, using 2 methods of analysis (histology and histomorphometry), we have demonstrated that humanized NSG mice could develop post-traumatic OA after a DMM surgery. The presence of human immune cell in the spleen and subchondral bone marrow confirms the success of the hematopoietic transplant. However, we failed to detect any immune cells in the synovial membrane or articular cartilage although human immune cells were observed near the joint in the subchondral bone marrow. The search for these immune cells in the synovial membrane would be more appropriate during the inflammatory burst, meaning within the first 2 weeks following the DMM surgery. Our results therefore evidence the feasibility to induce OA by DMM in humanized mice. This model could constitute a useful tool to study the therapeutic efficacy of encapsulated human mesenchymal stem cells.>

Intermittent Parathyroid Hormone [1-34] Augments Chondrogenesis of the Mandibular Condylar Cartilage of the Temporomandibular Joint.

To characterize the long-term effects of intermittent parathyroid hormone (I-PTH) on the mandibular condylar cartilage (MCC) and subchondral bone of the temporomandibular joint, in vivo and in vitro. For the in vivo experiments, sixteen 10-week-old mice were divided into 2 groups: (1) I-PTH (n = 8)-subcutaneous daily injection of PTH; (2) control group (n = 8)-subcutaneous daily injection of saline solution. Experiments were carried out for 4 weeks. Mice were injected with calcein, alizarin complexone, and cell proliferation marker before euthanasia. For the in vitro experiments, primary chondrocyte cultures from the MCC of eight 10-week-old mice were treated with I-PTH for 14 days. There was a significant increase in bone volume, tissue density, mineral deposition, osteoclastic activity, cell proliferation in the cartilage, and cartilage thickness in the I-PTH-treated mice when compared with the control group. In addition, immunohistochemistry in cartilage revealed that I-PTH administration led to an increase in expression of vascular endothelial growth factor and to a decreased expression of sclerostin, matrix metallopeptidase 13, and aggreganase-1 (ADAM-TS4). Quantitative polymerase chain reaction analysis of the I-PTH-treated chondrocytes revealed significantly decreased relative expression of collagen type X (Col10a1), alkaline phosphatase (Alp), and Indian Hedgehog (Ihh) and remarkable increased expression of Sox9, fibroblast growth factor 2 (Fgf2), and proteoglycan 4 (Prg4). I-PTH administration causes anabolic effects at the subchondral region of the mandibular condyle while triggers anabolic and protective effects at the MCC.

Harnessing Human Decellularized Blood Vessel Matrices and Cellular Construct Implants to Promote Bone Healing in an Ex Vivo Organotypic Bone Defect Model.

Decellularized matrices offer a beneficial substitute for biomimetic scaffolds in tissue engineering. The current study examines the potential of decellularized placental vessel sleeves (PVS) as a periosteal protective sleeve to enhance bone regeneration in embryonic day 18 chick femurs contained within the PVS and cultured organotypically over a 10 day period. The femurs are inserted into decellularized biocompatibility-tested PVS and maintained in an organotypic culture for a period of 10 days. In femurs containing decellularized PVS, a significant increase in bone volume (p < 0.001) is evident, demonstrated by microcomputed tomography (µCT) compared to femurs without PVS. Histological and immunohistological analyses reveal extensive integration of decellularized PVS with the bone periosteum, and enhanced conservation of bone architecture within the PVS. In addition, the expressions of hypoxia inducible factor-1 alpha (HIF-1α), type II collagen (COL-II), and proteoglycans are observed, indicating a possible repair mechanism via a cartilaginous stage of the bone tissue within the sleeve. The use of decellularized matrices like PVS offers a promising therapeutic strategy in surgical tissue replacement, promoting biocompatibility and architecture of the tissue as well as a factor-rich niche environment with negligible immunogenicity.

Loss of mechanosensitive sclerostin may accelerate cranial bone growth and regeneration.

Cranial defects can result from trauma, infection, congenital malformations, and iatrogenic causes and represent a surgical challenge. The current standard of care is cranioplasty, with either autologous or allogeneic material. In either case, the intrinsic vascularity of the surrounding tissues allows for bone healing. The objective of this study was to determine if mechanotransductive gene manipulation would yield non-weight-bearing bone regeneration in a critical size calvarial defect in mice. A mouse model of Sost deletion in Sost knockout (KO) mice was created in which the osteocytes do not express sclerostin. A critical size calvarial defect (4 mm in diameter) was surgically created in the parietal bone in 8-week-old wild-type (n = 8) and Sost KO (n = 8) male mice. The defects were left undisturbed (no implant or scaffold) to simulate a traumatic calvariectomy model. Eight weeks later, the animals were examined at necropsy by planimetry, histological analysis of new bone growth, and micro-CT scanning of bone thickness. Defects created in wild-type mice did not fill with bone over the study period of 2 months. Genetic downregulation of sclerostin yielded animals that were able to regenerate 40% of the initial critical size defect area 8 weeks after surgery. A thin layer of bone covered a significant portion of the original defect in all Sost KO animals. A statistically significant increase in bone volume (p < 0.05) was measured in Sost KO mice using radiodensitometric analysis. Immunohistochemical analysis also confirmed that this bone regeneration occurred through the Wnt pathway and originated from the edge of the defect; BMP signaling did not appear to be affected by sclerostin. Mechanical loading is an important mechanism of bone formation in the cranial skeleton and is poorly understood. This is partially due to the fact that it is difficult to load bone in the craniomaxillofacial skeleton. This study suggests that modulation of the Wnt pathway, as is able to be done with monoclonal antibodies, is a potentially efficacious method for bone regeneration that requires further study.

Effects of Bisphosphonate Administration on Cleft Bone Graft in a Rat Model.

Bone grafts in patients with cleft lip and palate can undergo a significant amount of resorption. The aim of this study was to investigate the effects of bisphosphonates (BPs) on the success of bone grafts in rats. Thirty-five female 15-week-old Fischer F344 Inbred rats were divided into the following experimental groups, each receiving bone grafts to repair an intraoral CSD: (1) Graft/saline: systemic administration of saline and (2) systemic administration of zoledronic acid immediately following surgery (graft/BP/T0), (3) 1 week postoperatively (graft/BP/T1), and (4) 3 weeks postoperatively (graft/BP/T2). As an additional control, the defect was left empty without bone graft. Microcomputed tomography and histologic analyses were performed in addition to evaluation of osteoclasts through tartrate-resistant acid phosphatase staining. Bone volume fraction (bone volume/tissue volume) for the delayed BP treatment groups (graft/BP/T1 = 45.4% ± 8.8%; graft/BP/T2 = 46.1% ± 12.4%) were significantly greater than that for the graft/saline group (31.0% ± 7.9%) and the graft/BP/T0 (27.6% ± 5.9%) 6 weeks postoperatively (P < .05). Hematoxylin and eosin staining confirmed an evident increase in bone volume and fusion of defect margins with existing palatal bone in the graft/BP/T1 and graft/BP/T2 groups. The graft/BP/T0 group showed the lowest bone volume with signs of acute inflammation. Delayed BP administration following cleft bone graft surgery led to significant increase in bone volume and integration compared with saline controls. However, BP injection immediately after the surgery did not enhance bone volume, and rather, may negatively affect bone graft incorporation.