Bone Formation In Humans Research Articles

Delivery of a Jagged1-PEG-MAL hydrogel with pediatric human bone cells regenerates critically sized craniofacial bone defects

Current treatments for congenital and acquired craniofacial (CF) bone abnormalities are limited and costly. Conventional methods involve surgical correction, short-term stabilization, and long-term bone grafting, which may include problematic allografts and limited autografts. While bone morphogenetic protein 2 (BMP2) has been used for bone regeneration, it can cause bone overgrowth and life-threatening inflammation. Bone marrow-derived mesenchymal stem cell therapies, though promising, are not Food and Drug Administration approved and are resource intensive. Thus, there is a need for effective, affordable, and less side-effect-prone bone regenerative therapies. Previous research demonstrated that JAGGED1 induces osteoblast commitment in murine cranial neural crest cells through a NOTCH-dependent non-canonical pathway involving JAK2–STAT5. We hypothesize that delivery of JAGGED1 and induction of its downstream NOTCH non-canonical signaling in pediatric human osteoblasts constitutes an effective bone regenerative treatment. Delivering pediatric human bone-derived osteoblast-like cells to an in vivo murine bone loss model of a critically sized cranial defect, we identified that JAGGED1 promotes human pediatric osteoblast commitment and bone formation through p70 S6K phosphorylation. This approach highlights the potential of JAGGED1 and its downstream activators as innovative treatments for pediatric CF bone loss.

Delivery of a Jagged1-PEG-MAL hydrogel with pediatric human bone cells regenerates critically sized craniofacial bone defects.

Current treatments for congenital and acquired craniofacial (CF) bone abnormalities are limited and costly. Conventional methods involve surgical correction, short-term stabilization, and long-term bone grafting, which may include problematic allografts and limited autografts. While bone morphogenetic protein 2 (BMP2) has been used for bone regeneration, it can cause bone overgrowth and life-threatening inflammation. Bone marrow-derived mesenchymal stem cell therapies, though promising, are not Food and Drug Administration approved and are resource intensive. Thus, there is a need for effective, affordable, and less side-effect-prone bone regenerative therapies. Previous research demonstrated that JAGGED1 induces osteoblast commitment in murine cranial neural crest cells through a NOTCH-dependent non-canonical pathway involving JAK2-STAT5. We hypothesize that delivery of JAGGED1 and induction of its downstream NOTCH non-canonical signaling in pediatric human osteoblasts constitutes an effective bone regenerative treatment. Delivering pediatric human bone-derived osteoblast-like cells to an in vivo murine bone loss model of a critically sized cranial defect, we identified that JAGGED1 promotes human pediatric osteoblast commitment and bone formation through p70 S6K phosphorylation. This approach highlights the potential of JAGGED1 and its downstream activators as innovative treatments for pediatric CF bone loss.

LINC00963 Represses Osteogenic Differentiation of hBMSCs via the miR-10b-5p/RAP2A/AKT Axis.

Osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs) is important for human bone formation. Long non-coding RNAs (lncRNAs) are critical regulators in osteogenic differentiation. This study aimed to explore the function and mechanisms of long intergenic non-protein coding RNA 963 (LINC00963) in affecting osteogenesis. Cell differentiation was assessed by alkaline phosphatase (ALP) activity detection and ALP staining assay. Meanwhile, levels of osteogenic marker genes, including RUNX family transcription factor 2 (RUNX2), osteocalcin (OCN), and osteopontin (OPN), were detected by RT-qPCR and western blot. Cell proliferation and apoptosis were measured using CCK-8 assay and flow cytometry analysis. RNA immunoprecipitation (RIP), RNA pull-down and luciferase reporter assays were used to investigate the interaction between genes. LINC00963 expression was down-regulated in hBMSCs treated with osteogenic induction. LINC00963 overexpression inhibited hBMSCs differentiation, proliferation, and elevated apoptosis. LINC00963 acted as a competing endogenous RNA (ceRNA) to interact with miR-10b-5p and thereby regulated the expression level of Ras-related protein Rap-2a (RAP2A). LINC00963 regulated RAP2A to inhibit the level of phosphorylated AKT (p-AKT). LINC00963 inhibited hBMSCs differentiation, proliferation, and elevated apoptosis via the miR-10b-5p/RAP2A/AKT signaling, which might help improve the treatment of osteoporosis.

Disease aggravation following surgery in a rare patient suspected to Fibrodysplasia (Myositis) ossificans progressiva: a case report

BackgroundFibrodysplasia ossificans progressiva (FOP) as a rare and heritable disorder with the infrequent genetic transmission of the condition is a catastrophic disorder of heterotopic ossification (HO) and a cause of extraskeletal bone formation in humans. Given the lack of effective treatment for this disease, the important point is to avoid aggravating factors such as bone biopsy, surgery, and intramuscular injection.Case presentationIn this report, we present a 52-year-old female patient, Kurdish ethnic, suspected to FOP who had a surgical intervention on the second toe of the right foot, which subsequently, it caused further deterioration of the disease in the person including necrosis and amputation of the distal phalanx of the second toe.ConclusionsAlthough, based on our investigation and the available scientific evidence, surgery may a cause for faster progression and worsening of the FOP disorder, but its proof requires further studies.

Human Wharton’s jelly-derived mesenchymal stromal cells promote bone formation in immunodeficient mice when administered into a bone microenvironment

BackgroundWharton’s Jelly (WJ) Mesenchymal Stromal Cells (MSC) have emerged as an attractive allogeneic therapy for a number of indications, except for bone-related conditions requiring new tissue formation. This may be explained by the apparent recalcitrance of MSC,WJ to differentiate into the osteogenic lineage in vitro, as opposed to permissive bone marrow (BM)-derived MSCs (MSC,BM) that readily commit to bone cells. Consequently, the actual osteogenic in vivo capacity of MSC,WJ is under discussion.MethodsWe investigated how physiological bone environments affect the osteogenic commitment of recalcitrant MSCs in vitro and in vivo. To this end, MSC of BM and WJ origin were co-cultured and induced for synchronous osteogenic differentiation in vitro using transwells. For in vivo experiments, immunodeficient mice were injected intratibially with a single dose of human MSC and bone formation was evaluated after six weeks.ResultsCo-culture of MSC,BM and MSC,WJ resulted in efficient osteogenesis in both cell types after three weeks. However, MSC,WJ failed to commit to bone cells in the absence of MSC,BM’s osteogenic stimuli. In vivo studies showed successful bone formation within the medullar cavity of tibias in 62.5% of mice treated with MSC, WJ. By contrast, new formed trabeculae were only observed in 25% of MSC,BM-treated mice. Immunohistochemical staining of human COXIV revealed the persistence of the infused cells at the site of injection. Additionally, cells of human origin were also identified in the brain, heart, spleen, kidney and gonads in some animals treated with engineered MSC,WJ (eMSC,WJ). Importantly, no macroscopic histopathological alterations, ectopic bone formation or any other adverse events were detected in MSC-treated mice.ConclusionsOur findings demonstrate that in physiological bone microenvironment, osteogenic commitment of MSC,WJ is comparable to that of MSC,BM, and support the use of off-the-shelf allogeneic MSC,WJ products in bone repair and bone regeneration applications.

3D relationship between hierarchical canal network and gradient mineralization of shark tooth osteodentin.

Osteodentin is a dominant mineralized collagenous tissue in the teeth of many fishes, with structural and histological characteristics resembling those of bone. Osteodentin, like bone, comprises osteons as basic structural building blocks, however, it lacks the osteocytes and the lacuno-canalicular network (LCN), which are known to play critical roles in controlling the mineralization of the collagenous matrix in bone. Although numerous vascular canals exist in osteodentin, their role in tooth maturation and the matrix mineralization process remain poorly understood. Here, high resolution micro-computed tomography (micro-CT) and focused ion beam-scanning electron microscopy (FIB-SEM) were used to obtain 3D structural information of osteodentin in shark teeth at multiple scales. We observed a complex 3D network of primary canals with a diameter ranging from ∼10 µm to ∼120 µm, where the canals are surrounded by osteon-like concentric layers of lamella, with 'interosteonal' tissue intervening between neighboring osteons. In addition, numerous hierarchically branched secondary canals extended out radially from the primary canals into the interosteonal tissue, with a decreasing diameter from ∼10 µm to hundreds of nanometers. Interestingly, the mineralization degree increases from the periphery of primary canals to the interosteonal tissue, suggesting that mineralization begins in the interosteonal tissue. Correspondingly, the hardness and elastic modulus of the interosteonal tissue is higher than those of the osteonal tissue. These results demonstrate that the 3D hierarchical canal network is positioned to play a critical role in controlling the gradient mineralization of osteodentin, also providing valuable insight into the formation of mineralized collagenous tissue without osteocytes and LCN. STATEMENT OF SIGNIFICANCE: : Bone is a composite material with versatile mechanical properties. Osteocytes and lacuno-canalicular network (LCN) are kown to play critical roles during formation of human bone. However, the skeleton of many fishes, including bone and osteodentin, although lacking osteocytes and LCN, could exhibit similar osteon-like structure and mechanical functions. Here, using various high resolution 3D characterization techniques, we reveal that the 3D network of primary canals and numerous hierarchically branched secondary canals correlate with the mineralization gradient and micromechanical properties of osteonal and interosteonal tissues of shark tooth osteodentin. This work significantly improves our understanding of the construction of bone-like mineralized tissue without osteocytes and LCN, and provides inspirations for the fabrication of functional materials with hierarchical structure.

Engineering 3D Printed Bioceramic Scaffolds to Reconstruct Critical-Sized Calvaria Defects in a Skeletally Immature Pig Model.

Three-dimensional printed bioceramic scaffolds composed of 100% β-tricalcium phosphate augmented with dipyridamole (3DPBC-DIPY) can regenerate bone across critically sized defects in skeletally mature and immature animal models. Before human application, safe and effective bone formation should be demonstrated in a large translational animal model. This study evaluated the ability of 3DPBC-DIPY scaffolds to restore critically sized calvarial defects in a skeletally immature, growing minipig. Unilateral calvarial defects (~1.4 cm) were created in 6-week-old Göttingen minipigs ( n = 12). Four defects were filled with a 1000 μm 3DPBC-DIPY scaffold with a cap (a solid barrier on the ectocortical side of the scaffold to prevent soft-tissue infiltration), four defects were filled with a 1000 μm 3DPBC-DIPY scaffold without a cap, and four defects served as negative controls (no scaffold). Animals were euthanized 12 weeks postoperatively. Calvariae were subjected to micro-computed tomography, 3D reconstruction with volumetric analysis, qualitative histologic analysis, and nanoindentation. Scaffold-induced bone growth was statistically greater than in negative controls ( P ≤ 0.001), and the scaffolds with caps produced significantly more bone generation compared with the scaffolds without caps ( P ≤ 0.001). Histologic analysis revealed woven and lamellar bone with haversian canals throughout the regenerated bone. Cranial sutures were observed to be patent, and there was no evidence of ectopic bone formation or excess inflammatory response. Reduced elastic modulus and hardness of scaffold-regenerated bone were found to be statistically equivalent to native bone ( P = 0.148 for reduced elastic modulus of scaffolds with and without caps and P = 0.228 and P = 0.902 for hardness of scaffolds with and without caps, respectively). 3DPBC-DIPY scaffolds have the capacity to regenerate bone across critically sized calvarial defects in a skeletally immature translational pig model. This study assessed the bone generative capacity of 3D-printed bioceramic scaffolds composed of 100% β-tricalcium phosphate and augmented with dipyridamole placed within critical-sized calvarial defects in a growing porcine model.

Positive and Negative Regulators of Sclerostin Expression.

Sclerostin is secreted from osteocytes, binds to the Wnt co-receptor Lrp5/6, and affects the interaction between Wnt ligands and Lrp5/6, which inhibits Wnt/β-catenin signals and suppresses bone formation. Sclerostin plays an important role in the preservation of bone mass by functioning as a negative regulator of bone formation. A sclerostin deficiency causes sclerosteosis, which is characterized by an excess bone mass with enhanced bone formation in humans and mice. The expression of sclerostin is positively and negatively regulated by many factors, which also govern bone metabolism. Positive and negative regulators of sclerostin expression and their effects are introduced and discussed herein based on recent and previous findings, including our research.

Three-dimensional bioprinting of bioactive scaffolds with thermally embedded abalone shell particles for bone tissue engineering

Abalone shells, which contain both organic and inorganic matter, can facilitate bone remodeling and have been used to fabricate three-dimensional (3D)-printed scaffolds for bone regeneration. Herein, polycaprolactone (PCL) scaffolds were fabricated using 3D printing with abalone shell particles (ASPs) used in high-temperature processing. ASPs were heated to approximately the melting point of PCL and thermally embedded in 3D-printed PCL using a relatively simple process. The morphology and roughness of the composite scaffold changed according to the weight of ASPs used. The scaffolds were grouped as follows: ASP25 (25 mg), ASP50 (50 mg), and ASP100 (100 mg). The ASP25 group exhibited optimum cell viability and proliferation because of the direct influence of roughness and rapid pH changes. The ASP25 and ASP100 groups showed the highest alkaline phosphatase activity. This could be attributed to the effect of the alkaline environment, dissolution of calcium ions, and presence of bioactive molecules in the ASPs that could support bone regeneration. Therefore, the ASP25 group was the most suitable for fabricating bone scaffolds. This study revealed the potential applicability of ASP-embedded scaffolds in bone tissue engineering involving natural bio-organisms that self-mineralize in a process similar to human bone formation.

Gli1+ progenitors mediate bone anabolic function of teriparatide via Hh and Igf signaling.

SUMMARYTeriparatide is the most widely prescribed bone anabolic drug in the world, but its cellular targets remain incompletely defined. The Gli1+ metaphyseal mesenchymal progenitors (MMPs) are a main source for osteoblasts in postnatal growing mice, but their potential response to teriparatide is unknown. Here, by lineage tracing, we show that teriparatide stimulates both proliferation and osteoblast differentiation of MMPs. Single-cell RNA sequencing reveals heterogeneity among MMPs, including an unexpected chondrocyte-like osteoprogenitor (COP). COP expresses the highest level of Hedgehog (Hh) target genes and the insulin-like growth factor 1 receptor (Igf1r) among all cell clusters. COP also expresses Pth1r and further upregulates Igf1r upon teriparatide treatment. Inhibition of Hh signaling or deletion of Igf1r from MMPs diminishes the proliferative and osteogenic effects of teriparatide. The study therefore identifies COP as a teriparatide target wherein Hh and insulin-like growth factor (Igf) signaling are critical for the osteoanabolic response in growing mice.

Tissue-level mechanical stimuli drive bone formation and resorption in humans and mice

Tissue-level mechanical stimuli drive bone formation and resorption in humans and mice

Maternal vitamin B12 in mice positively regulates bone, but not muscle mass and strength in post-weaning and mature offspring.

Vitamin B12 deficiency has been shown to affect bone mass in rodents and negatively impact bone formation in humans. In this study using mouse models, we define the effect of B12 supplementation in the wild-type mother and B12 deficiency in a mouse genetic model (Gif−/− mice) during gestation on bone and muscle architecture and mechanical properties in the offspring. Analysis of bones from 4-wk-old offspring of the wild-type mother following vehicle or B12 supplementation during gestation (from embryonic day 0.5 to 20.5) showed an increase in bone mass caused by an isolated increase in bone formation in the B12-supplemented group compared with vehicle controls. Analysis of the effect of B12 deficiency in the mother in a mouse genetic model (Gif−/− mice) on the long bone architecture of the offspring showed a compromised cortical and trabecular bone mass, which was completely prevented by a single injection of B12 in the B12-deficient Gif−/− mothers. Biomechanical analysis of long bones of the offspring born from B12-supplemented wild-type mothers showed an increase in bone strength, and conversely, offspring born from B12-deficient Gif−/− mothers revealed a compromised bone strength, which could be rescued by a single injection of B12 in the B12-deficient Gif−/− mother. Muscle structure and function analysis however revealed no significant effect on muscle mass, structure, and grip strength of B12 deficiency or supplementation in Gif−/− mice compared with littermate controls. Together, these results demonstrate the beneficial effect of maternally derived B12 in the regulation of bone structure and function in the offspring.

Type 1 diabetic Akita mice have low bone mass and impaired fracture healing.

Type 1 diabetes (T1DM) impairs bone formation and fracture healing in humans. Akita mice carry a mutation in one allele of the insulin-2 (Ins2) gene, which leads to pancreatic beta cell dysfunction and hyperglycemia by 5-6weeks age. We hypothesized that T1DM in Akita mice is associated with decreased bone mass, weaker bones, and impaired fracture healing. Ins2±(Akita) and wildtype (WT) males were subjected to femur fracture at 18-weeks age and healing assessed 3-21days post-fracture. Non-fractured left femurs were assessed for morphology (microCT) and strength (bending or torsion) at 19-21weeks age. Fractured right femurs were assessed for callus mechanics (torsion), morphology and composition (microCT and histology) and gene expression (qPCR). Both Akita and WT mice gained weight from 3 to 18weeks age, but Akita mice weighed less starting at 5weeks (-5.2%, p<0.05). At 18-20weeks age Akita mice had reduced serum osteocalcin (-30%), cortical bone area (-16%), and thickness (-17%) compared to WT, as well as reduced cancellous BV/TV (-39%), trabecular thickness (-23%) and vBMD (-31%). Mechanical testing of non-fractured femurs showed decreased structural (stiffness, ultimate load) and material (ultimate stress) properties of Akita bones. At 14 and 21days post fracture Akita mice had a significantly smaller callus than WT mice (~30%), with less cartilage and bone area. Assessment of torsional strength showed a weaker callus in Akita mice with lower stiffness (-42%), maximum torque (-44%) and work to fracture (-44%). In summary, cortical and cancellous bone mass were reduced in Akita mice, with lower bone mechanical properties. Fracture healing in Akita mice was impaired by T1DM, with a smaller, weaker fracture callus due to decreased cartilage and bone formation. In conclusion, the Akita mouse mimics some of the skeletal features of T1DM in humans, including osteopenia and impaired fracture healing, and may be useful to test interventions.

MicroRNAs in Combat-Related Heterotopic Ossification: Commentary on an article by J.F. de Vasconcellos, PhD, et al.: "A microRNA Signature for Impaired Wound-Healing and Ectopic Bone Formation in Humans".

MicroRNAs in Combat-Related Heterotopic Ossification: Commentary on an article by J.F. de Vasconcellos, PhD, et al.: "A microRNA Signature for Impaired Wound-Healing and Ectopic Bone Formation in Humans".

LncRNA LINC01535 upregulates BMP2 expression levels to promote osteogenic differentiation via sponging miR‑3619‑5p.

Osteoporosis is a debilitating skeletal disease that causes bones to collapse and is accompanied by a high risk of bone fracture. It was previously demonstrated that the osteogenic differentiation of human bone marrow‑derived mesenchymal stem cells (hBMSCs) serves an important role in the process of human bone formation. Accumulating research has indicated that long non‑coding RNAs (lncRNAs) participate in hBMSC osteogenic differentiation. For example, LINC01535 was reported to serve as a carcinogenic factor in cervical cancer; however, its latent function and molecular mechanism in the osteogenesis of hBMSCs remain to be investigated. The present study showed that the expression levels of LINC01535 were upregulated upon increasing osteogenic differentiation time. In addition, the inhibition of LINC01535 inhibited hBMSC proliferation and osteogenic differentiation and promoted cell apoptosis. Using bioinformatics analysis, LINC01535 was discovered to have complementary binding sites for microRNA (miR)‑3619‑5p, and further experiments demonstrated that LINC01535 functioned as a sponge of miR‑3619‑5p. Additionally, bone morphogenetic protein2 (BMP2) was confirmed to be a target of miR‑3619‑5p. The results revealed that LINC01535 regulated the expression levels of BMP2 via sponging miR‑3619‑5p. In conclusion, the findings of the present study suggested that LINC01535 may accelerate the osteogenic process of hBMSCs via targeting the miR‑3619‑5p/BMP2 axis, which may offer an innovative therapeutic method for osteoporosis.

γ-Aminobutyric Acid Promotes Osteogenic Differentiation of Mesenchymal Stem Cells by Inducing TNFAIP3.

Osteoporosis is the most common metabolic bone disease. There is still an unmet need for novel therapeutic agents that could be beneficial as osteoporosis treatments. It has been reported that the neurotransmitter γ-aminobutyric acid (GABA) might be associated with human bone formation. However, the precise mechanism remains unclear. To investigate the effect of GABA on bone metabolism and explore the possible role of TNFAIP3 in this process. GABA had little effect on the proliferation of human mesenchymal stem cells (hMSCs) and RAW 264.7 cells, as indicated by the cell counting kit-8 (CCK-8) assay. The results showed that GABA enhanced the intensity of ALP staining, ALP activity, and accumulation of Ca2+ mineralized nodules in hMSCs during osteogenic induction. The qRT-PCR results indicated that GABA treatment significantly increased the mRNA expression of osteogenic genes in hMSCs. In RAW 264.7 cells, TRAP staining showed that GABA did not alter the number or size of osteoclasts or the expression of osteoclastic genes, which suggests that GABA does not affect osteoclastic differentiation. Mechanistically, GABA treatment significantly induced the sustained expression of TNFAIP3. Furthermore, by knocking down TNFAIP3, the osteogenic effect of GABA was antagonized, which suggests that TNFAIP3 mediates the effects of GABA in hMSCs. Our results suggested that GABA treatment positively regulated osteogenic differentiation by upregulating TNFAIP3, while no obvious effect on osteoclastic differentiation was detected. Therefore, our results provide a potential gene therapy for the treatment of osteoporosis and low bone mineral density.

Human Umbilical Vein Endothelial Cell Support Bone Formation of Adipose-Derived Stem Cell-Loaded and 3D-Printed Osteogenic Matrices in the Arteriovenous Loop Model.

Introduction: For the regeneration of large volume tissue defects, the interaction between angiogenesis and osteogenesis is a crucial prerequisite. The surgically induced angiogenesis by means of an arteriovenous loop (AVL), is a powerful methodology to enhance vascularization of osteogenic matrices. Moreover, the AVL increases oxygen and nutrition supply, thereby supporting cell survival as well as tissue formation. Adipose-derived stem cells (ADSCs) are interesting cell sources because of their simple isolation, expansion, and their osteogenic potential. This study targets to investigate the coimplantation of human ADSCs after osteogenic differentiation and human umbilical vein endothelial cells (HUVECs), embedded in a vascularized osteogenic matrix of hydroxyapatite (HAp) ceramic for bone tissue engineering. Materials and Methods: An osteogenic matrix consisting of HAp granules and fibrin has been vascularized by means of an AVL. Trials in experimental groups of four settings were performed. Control experiments without any cells (A) and three cell-loaded groups using HUVECs (B), ADSCs (C), as well as the combination of ADSCs and HUVECs (D) were performed. The scaffolds were implanted in a porous titanium chamber, fixed subcutaneously in the hind leg of immunodeficient Rowett Nude rats and explanted after 6 weeks. Results: In all groups, the osteogenic matrix was strongly vascularized. Moreover, remodeling processes and bone formation in the cell-containing groups with more bone in the coimplantation group were proved successful. Conclusion: Vascularization and bone formation of osteogenic matrices consisting of ADSCs and HUVECs in the rat AVL model could be demonstrated successfully for the first time. Hence, the coimplantation of differentiated ADSCs with HUVECs may therefore be considered as a promising approach for bone tissue engineering.

A microRNA Signature for Impaired Wound-Healing and Ectopic Bone Formation in Humans.

Heterotopic ossification (HO) is characterized by the abnormal growth of ectopic bone in soft tissues, frequently occurring within the military population because of extensive orthopaedic combat trauma. MicroRNAs (miRNAs) are small noncoding RNAs that act as post-transcriptional regulators of gene expression. We hypothesized that a clinically relevant miRNA signature could be detected in patients following injury that progressed to form HO (HO+) or did not form HO (HO-). Tissue samples were obtained from injured servicemembers during their initial surgical debridements, and miRNA profiling was performed using a real-time miRNA polymerase chain reaction (PCR) array. Primary mesenchymal progenitor cells (MPCs) were harvested from debrided traumatized human muscle tissue, and cells were isolated and cultured in vitro. Mimic miRNAs were transfected into MPCs, followed by downstream in vitro analyses. The investigation of the miRNA expression profile in the tissue of HO+ compared with HO- patients demonstrated a molecular signature that included the upregulation of miR-1, miR-133a, miR-133b, miR-206, miR-26a, and miR-125b. Transfection of each of these mature miRNAs into MPCs followed by osteogenic induction demonstrated that miR-1, miR-133a, miR-133b, and miR-206 enhanced osteogenic differentiation compared with control treatments. In silico and in vitro analyses identified the transcription factor SOX9 as a candidate downstream target of miR-1 and miR-206 miRNAs. Our data demonstrated a molecular signature of miRNAs in the soft tissue of wounded servicemembers that was associated with the development of HO, providing novel insights into the underlying molecular mechanisms associated with posttraumatic HO. Prognostic Level II. See Instructions for Authors for a complete description of levels of evidence.

Downregulation of LINC00707 promotes osteogenic differentiation of human bone marrow‑derived mesenchymal stem cells by regulating DKK1 via targeting miR‑103a‑3p.

Human bone marrow-derived mesenchymal stem cells (HBMSCs) have the potential of multidirectional differentiation and self-renewal, which is important for the formation of human bone. It has been reported that long non-coding RNAs (lncRNAs) serve important roles in HBMSC osteogenic differentiation. The current study aimed to investigate the roles of long intergenic non-protein coding RNA 00707 (LINC00707) and microRNA (miR)-103a-3p in the osteogenic differentiation of HBMSCs. Reverse transcription-quantitative PCR (RT-qPCR) was performed to detect the expression levels of LINC00707, miR-103a-3p and osteogenesis-related genes (Alkaline phosphatase, osteocalcin, osteopontin and RUNX family transcription factor 2) in HBMSCs cultured in proliferation medium (PM) and osteogenic medium (OM). Mineralized matrix deposition was measured using Alizarin Red S staining. The protein expression levels of osteogenesis-related genes were detected by western blotting. The relationships between LINC00707, miR-103a-3p and dickkopf WNT signaling pathway inhibitor 1 (DKK1) were predicted using Starbase and TargetScan7.2, and were further assessed with a dual-luciferase reporter assay. After 21 days of cell culture, the results indicated that expression of LINC00707 was downregulated, and those of miR-103a-3p and osteogenesis-related genes were upregulated in OM-cultured HBMSCs. However, there was no significant difference in the aforementioned gene expression levels in PM-cultured HBMSCs. Small interfering (si)LINC00707 increased the deposition of mineralized matrix and promoted the expression levels of osteogenesis-related proteins. Furthermore, miR-103a-3p was predicted to be a target gene of LINC00707, its expression was significantly upregulated by siLINC00707, while overexpression of miR-103a-3p increased the expression levels of osteogenesis-related proteins. DKK1 was also predicted to be a target gene of miR-103a-3p and could inhibit the expression levels of osteogenesis-related proteins, but such effect of DKK1 could be reversed by the miR-103a-3p mimic. In conclusion, the present results suggested that LINC00707 regulated DKK1 expression by targeting miR-103a-3p to regulate osteogenic differentiation.

Synthesis, Characterization and Bioactivity of Fluoride Containing Borosilicate Glass Matrix Composite

Fluoride containing borosilicates find interesting applications in biomedical transplants and dentistry, wherein calcium and magnesium play pivotal role in human bone formation. In the present work, calcium and magnesium fluoride containing borosilicate glass matrix composites were synthesized via sol-gel route. The as synthesized and heat-treated samples were characterized using various techniques such as X-ray diffractometer (XRD), Fourier transform infrared spectroscopy (FT-IR), and thermogravimetric analysis (TGA). and Scanning Electron Microscopy-Energy dispersive spectroscopy (SEM-EDS). The samples were stabilized at suitable temperature based on the thermogravimetric analysis (TGA). The XRD patterns of as prepared samples showed various crystalline phases with varied degree of crystallinity. Heat-treated samples were tested for their in–vitro bioactivity. Interaction at sample-SBF interface was investigated by dipping the samples in simulated body fluid (SBF) at 37 °C. The analysis of the samples dipped in SBF and that of the residual SBF was carried out using various characterization techniques. Increasing the ageing time, the samples exhibited better in-vitro bioactivity.