A rare femoral heterotopic bone formation in a 14th-19th century female skeleton from Constância (Portugal).

OBJECTIVE: This paper aims to: (1) document a rare femoral heterotopic ossification (HO), and (2) discuss its aetiology and impact on the individual's locomotion and daily living activities. MATERIALS: Adult female skeleton (SG.14-SK.7) from the village of Constância (Portugal), and dated from the 14th-19th centuries CE. METHODS: The biological profile and the macroscopic analysis of the bone changes were assessed using standardized methods. RESULTS: The macroscopic analysis revealed a large bony mass (8 cm length) located immediately inferior to the small trochanter of the right femur. The lesion exhibited a compact, tubular appearance located at the site of attachment of the pectineus muscle. No signs of bone fracture were observed. CONCLUSIONS: The morphology of the SG.14-SK.7 femoral lesion is compatible with a probable case of myositis ossificans traumatica (MOT), secondary to acute trauma of the pectineus muscle. The underlying trauma episode, such as random accidental and/or occupation-related injury, is unknown. However, it is highly possible that this self-limiting condition significantly impaired the individual's daily life and mobility. SIGNIFICANCE: Evidence of severe acute muscle trauma is a rare finding compared with HO secondary to bone trauma and other minor muscle injuries. Moreover, no cases of MOT affecting the pectineus muscle have been reported in the paleopathological literature to date. LIMITATIONS: Although unlikely, a case of neurogenic or burn-related HO cannot be completely disregarded. It was not possible to undertake radiography as part of this study. SUGGESTIONS FOR FURTHER RESEARCH: The use of imaging techniques to complement the paleopathological description is advised.

Osseous Metaplasia and Ectopic Bone Formation in a Patient With Hashimoto's Thyroiditis.

Thyroid nodules are frequently accompanied by degenerative changes, such as hemorrhage, cholesterol crystallization, fibrous tissue deposition, or filling with fat. Although calcification is also a common phenomenon, osteogenesis, characterized by mature bone formation, is very rare. Here, we describe a case of Hashimoto's thyroiditis with osseous metaplasia and ectopic bone formation case and discuss its possible causes.

Bone Formation on Murine Cranial Bone by Injectable Cross-Linked Hyaluronic Acid Containing Nano-Hydroxyapatite and Bone Morphogenetic Protein.

New injection-type bone-forming materials are desired in dental implantology. In this study, we added nano-hydroxyapatite (nHAp) and bone morphogenetic protein (BMP) to cross-linkable thiol-modified hyaluronic acid (tHyA) and evaluated its usefulness as an osteoinductive injectable material using an animal model. The sol (ux-tHyA) was changed to a gel (x-tHyA) by mixing with a cross-linker. We prepared two sol−gel (SG) material series, that is, x-tHyA + BMP with and without nHAp (SG I) and x-tHyA + nHAp with and without BMP (SG II). SG I materials in the sol stage were injected into the cranial subcutaneous connective tissues of mice, followed by in vivo gelation, while SG II materials gelled in Teflon rings were surgically placed directly on the cranial bones of rats. The animals were sacrificed 8 weeks after implantation, followed by X-ray analysis and histological examination. The results revealed that bone formation occurred at a high rate (>70%), mainly as ectopic bone in the SG I tests in mouse cranial connective tissues, and largely as bone augmentation in rat cranial bones in the SG II experiments when x-tHyA contained both nHAp and BMP. The prepared x-tHyA + nHAp + BMP SG material can be used as an injection-type osteoinductive bone-forming material. Sub-periosteum injection was expected.

Heterotopic Mesenteric Ossification With Trilineage Hematopoiesis.

Heterotopic ossification (HO) histologically refers to extraskeletal bone formation in non-ossifying tissues, most commonly noted in the extremities, buttocks, abdominal wall, and hip joints. HO developing in the mesentery (heterotopic mesenteric ossification, HMO) is very rare, with fewer than 100 cases reported in the literature. It usually occurs in adult male patients with a history of repeated abdominal trauma. So far, only two cases of HMO have been reported with the development of hematopoietic bone marrow. Here, we report the third case of HMO with true trilineage hematopoiesis in a 66-year-old female with suspicious mesenteric-retained foreign material from prior surgical procedures, including hysterectomy for endometrial adenocarcinoma and multiple repairs for incisional hernia.

BMSCs and Osteoblast-Engineered ECM Synergetically Promotes Osteogenesis and Angiogenesis in an Ectopic Bone Formation Model.

Bone mesenchymal stem cells (BMSCs) have been extensively used in bone tissue engineering because of their potential to differentiate into multiple cells, secrete paracrine factors, and attenuate immune responses. Biomaterials are essential for the residence and activities of BMSCs after implantation in vivo. Recently, extracellular matrix (ECM) modification with a favorable regenerative microenvironment has been demonstrated to be a promising approach for cellular activities and bone regeneration. The aim of the present study was to evaluate the effects of BMSCs combined with cell-engineered ECM scaffolds on osteogenesis and angiogenesis in vivo. The ECM scaffolds were generated by osteoblasts on the small intestinal submucosa (SIS) under treatment with calcium (Ca)-enriched medium and icariin (Ic) after decellularization. In a mouse ectopic bone formation model, the SIS scaffolds were demonstrated to reduce the immune response, and lower the levels of immune cells compared with those in the sham group. Ca/Ic-ECM modification inhibited the degradation of the SIS scaffolds in vivo. The generated Ca/Ic-SIS scaffolds ectopically promoted osteogenesis according to the results of micro-CT and histological staining. Moreover, BMSCs on Ca/Ic-SIS further increased the bone volume percentage (BV/TV) and bone density. Moreover, angiogenesis was also enhanced by the Ca/Ic-SIS scaffolds, resulting in the highest levels of neovascularization according to the data ofCD31 staining. In conclusion, osteoblast-engineered ECM under directional induction is a promising strategy to modify biomaterials for osteogenesis and angiogenesis. BMSCs synergetically improve the properties of ECM constructs, which may contribute to the repair of large bone defects.

Graphene-Oxide Porous Biopolymer Hybrids Enhance In Vitro Osteogenic Differentiation and Promote Ectopic Osteogenesis In Vivo.

Over the years, natural-based scaffolds have presented impressive results for bone tissue engineering (BTE) application. Further, outstanding interactions have been observed during the interaction of graphene oxide (GO)-reinforced biomaterials with both specific cell cultures and injured bone during in vivo experimental conditions. This research hereby addresses the potential of fish gelatin/chitosan (GCs) hybrids reinforced with GO to support in vitro osteogenic differentiation and, further, to investigate its behavior when implanted ectopically. Standard GCs formulation was referenced against genipin (Gp) crosslinked blend and 0.5 wt.% additivated GO composite (GCsGp/GO 0.5 wt.%). Pre-osteoblasts were put in contact with these composites and induced to differentiate in vitro towards mature osteoblasts for 28 days. Specific bone makers were investigated by qPCR and immunolabeling. Next, CD1 mice models were used to assess de novo osteogenic potential by ectopic implantation in the subcutaneous dorsum pocket of the animals. After 4 weeks, alkaline phosphate (ALP) and calcium deposits together with collagen synthesis were investigated by biochemical analysis and histology, respectively. Further, ex vivo materials were studied after surgery regarding biomineralization and morphological changes by means of qualitative and quantitative methods. Furthermore, X-ray diffraction and Fourier-transform infrared spectroscopy underlined the newly fashioned material structuration by virtue of mineralized extracellular matrix. Specific bone markers determination stressed the osteogenic phenotype of the cells populating the material in vitro and successfully differentiated towards mature bone cells. In vivo results of specific histological staining assays highlighted collagen formation and calcium deposits, which were further validated by micro-CT. It was observed that the addition of 0.5 wt.% GO had an overall significant positive effect on both in vitro differentiation and in vivo bone cell recruitment in the subcutaneous region. These data support the GO bioactivity in osteogenesis mechanisms as being self-sufficient to elevate osteoblast differentiation and bone formation in ectopic sites while lacking the most common osteoinductive agents.

Heterotopic Ossification After Arthroscopic Procedures: A Scoping Review of the Literature.

BACKGROUND: Heterotopic ossification (HO) is the formation of bone in soft tissue resultant from inflammatory processes. Lesion formation after arthroscopic procedures is an uncommon but challenging complication. Optimal prophylaxis and management strategies have not been clearly defined. PURPOSE: To present a scoping review of the pathophysiology, risk factors, diagnostic modalities, prophylaxis recommendations, and current treatment practices concerning HO after arthroscopic management of orthopaedic injuries. STUDY DESIGN: Scoping review; Level of evidence, 4. METHODS: A scoping review via a PubMed search was performed according to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-analyses) guidelines. The search strategy was based on the terms "heterotopic ossification" AND "arthroscopy." The clinical outcomes review included studies on the arthroscopic management of orthopaedic injuries in which the primary subject matter or a secondary outcome was the development of HO. An analysis of the pathophysiology, diagnostic modalities, and management options was reported. RESULTS: A total of 43 studies (33,065 patients) reported on HO after hip arthroscopy, while 21 (83 patients) collectively reported on HO after arthroscopic procedures to the shoulder, elbow, knee, or ankle; however, management techniques were not standardized. Identified risk factors for HO included male sex and mixed impingement pathology, while intraoperative capsular management was not suggested as a contributing factor. Diagnosis of ossification foci was performed using radiography and computed tomography. The rate of HO after hip arthroscopy procedures approached 46% without prophylaxis, and administration of nonsteroidal anti-inflammatory drugs (NSAIDs) decreased occurrence rates to 4% but carries associated risks. External beam radiation has not been exclusively studied for use after arthroscopic procedures. CONCLUSION: HO is a known complication after arthroscopic management of orthopaedic injuries. NSAID prophylaxis has been demonstrated to be effective after hip arthroscopy procedures. Patients with persistent symptoms and mature lesions may be indicated for surgical excision, although variability is present in patient-reported outcome scores postoperatively.

Heterotopic Ossification Following Total Ankle Arthroplasty With Fourth-Generation Prostheses.

BACKGROUND: The purpose of the present study was to assess the radiographic incidence, location, and classification of heterotopic ossification (HO) in patients who underwent total ankle arthroplasty (TAA) with a 4th generation prostheses at a minimum of 1-year follow up. Baseline demographic, radiographic, and operative factors between patents with and without HO were compared. METHODS: Ninety ankles that underwent TAA with a 4th generation protheses, INFINITY (n = 62) or CADENCE (n = 28) were followed for an average of 23.7 (range, 12-49) months. Incidence and location of HO was assessed on weightbearing radiographs, and severity graded according to the modified Brooker classification. Data was compared between patents with and without HO to identity any predisposing factors. RESULTS: In 90 ankles that underwent 4th generation TAA, HO incidence was 55.6% (n = 50); 56.5% (n = 35) for INFINITY, and 53.6% (n = 15) for CADENCE. Twenty-five cases of HO were observed posteriorly, 16 anteriorly, and 9 combined. Severity was as follows; class I in 19 cases (38%), class II in 20 (40%), class III in 9 (18%) and class IV in 2 (4%). A single ankle required a non-revisional reoperation for HO debridement; reoperation rate of 2%. CONCLUSION: The present study suggests a similarly high incidence of HO after TAA with two different 4th generation protheses (INFINITY 56.5%, CADENCE 53.6%). A trend for differences in location and severity between the protheses may also be present. Given the paucity of literature, additional studies with longer follow-up are warranted to discern the significance of HO following TAA with 4th generation protheses. LEVEL OF EVIDENCE: Level III: Retrospective cohort study.

Molecular Indicators of Biomaterials Osteoinductivity - Cell Migration, BMP Production and Signalling Turns a Key.

In this work we dissected the osteoinductive properties of selected, PLGA-based scaffolds enriched with gel-derived bioactive glasses (SBGs) of either binary SiO2-CaO or ternary SiO2-CaO-P2O5 system, differing in CaO/SiO2 ratio (i.e. high -or low-calcium SBGs). To assess the inherent ability of the scaffolds to induce osteogenesis of human bone marrow stromal cells (BMSC), the study was designed to avoid any osteogenic stimuli beyond the putative osteogenic SBG component of the studied scaffolds. The bioactivity and porosity of scaffolds were confirmed by SBF test and porosimetry. Condition media (CM) from BMSC-loaded scaffolds exhibited increased Ca and decreased P content corresponding to SBGs CaO/SiO2 ratio, whereas Si content was relatively stable and overall lower in CM from scaffolds containing binary SBGs. CM from cell-loaded scaffolds containing high-calcium, binary SBGs promoted migration of BMSC and BMP-response in reporter osteoblast cell line. BMSC culture on these scaffolds or the ones containing ternary, low-calcium SBGs resulted in the activation of BMP-related signaling and expression of several osteogenic markers. Ectopic bone formation was induced by scaffolds containing binary SBGs, but high-calcium ones produced significantly more osteoid. Scaffolds containing ternary SBGs negatively influenced the expression of osteogenic transcription factors and Cx43, involved in cell-cell interactions. High-calcium scaffolds stimulated overall higher Cx43 expression. We believe the initial cell-cell communication may be crucial to induce and maintain osteogenesis and high BMP signaling on the studied scaffolds. The presented scaffolds' biological properties may also constitute new helpful markers to predict osteoinductive potential of other bioactive implant materials.

Cell-free, quantitative mineralization measurements as a proxy to identify osteoinductive bone graft substitutes.

Some synthetic bone graft substitutes (BGS) can trigger ectopic bone formation, which is the hallmark of osteoinduction and the most important prerequisite for the repair of large bone defects. Unfortunately, measuring or predicting BGS osteoinductive potential based on in vitro experiments is currently impossible. A recent study claimed that synthetic BGS can induce bone formation ectopically if they create a local homeostatic imbalance during their in vivo mineralization. This raised the hope that a simple cell free in vitro mineralization experiment would correlate with osteoinduction. The aim of the present study was therefore to assess the ability of a quantitative in vitro mineralization test to predict and rank the osteoinductive potential of BGS. Eight calcium phosphate BGS already tested ectopically in 9 different in vivo studies were used for that purpose. The experiment was able to identify materials that are reliably osteoinductive from those that are not, but was inaccurate in ranking the osteoinductive materials between each other. Chemical contaminants (Ca2+, Mg2+, H+, OH-, PO43-) present in some of the BGS affected the in vitro mineralization experiment results, but not in a direction that could explain the different rankings. In conclusion, this study suggests that an in vitro experiment can be used as a fast and reliable screening tool to identify osteoinductive BGS and underline the need to study ionic contaminants on calcium phosphate BGS.

Diffuse Pulmonary Ossification with Connective Tissue Weakness Potentially Due to Vascular Ehlers-Danlos Syndrome.

A 30-year-old non-smoking man was referred to our hospital for the further examination of abnormal shadows revealed by chest X-ray. He had mild shortness of breath. Chest computed tomography revealed a fine-grained dendritic shadow with diffuse calcification in both lungs and as well as emphysematous changes in the upper lung lobes. A surgical lung biopsy histology revealed diffuse pulmonary ossification complicated with lung laceration, vascular disruption, hemosiderosis, and emphysema, suggesting vascular Ehlers-Danlos syndrome (vEDS). However, the patient had no external physical signs or family history of vEDS and no COL3A1 gene mutations. We are closely monitoring this patient in the clinic.

Novel three-dimensional bioglass functionalized gelatin nanofibrous scaffolds for bone regeneration.

The clinical use of FDA-approved bone morphogenetic proteins (BMPs) are impeded by high costs, super-high dosage requirement, short half-life, and other undesirable side effects. Therefore, designing a biomaterial that can promote new bone formation without using exogenous BMPs is highly desirable in clinical applications. In the present work, a new kind of nanofibrous scaffold composed of gelatin and 45S5 bioglass (GF/45S5 BG) was prepared through thermally induced phase separation method together with the particle leach technique (TIPS&P). In addition to the significantly higher mechanical strength, the composite scaffolds (GF/45S5 BG) significantly increased osteogenic differentiation of human mesenchymal stem cells (hMSCs) in vitro compared with the neat scaffold (GF) without adding other biological agents, for example, BMPs or hormones. Most importantly, our in vivo studies also indicated that GF/45S5 BG scaffolds could directly promote ectopic bone regeneration in SD rats without exogenous BMP2. In summary, both in vitro and in vivo results indicated that the novel 45S5 bioglass functionalized GF nanofibrous scaffold is a promising alternative for bone tissue engineering.

Zn- or Cu-containing CaP-Based Coatings Formed by Micro-Arc Oxidation on Titanium and Ti-40Nb Alloy: Part II-Wettability and Biological Performance.

This work describes the wettability and biological performance of Zn- and Cu-containing CaP-based coatings prepared by micro-arc oxidation on pure titanium (Ti) and novel Ti-40Nb alloy. Good hydrophilic properties of all the coatings were demonstrated by the low contact angles with liquids, not exceeding 45°. An increase in the applied voltage led to an increase of the coating roughness and porosity, thereby reducing the contact angles to 6° with water and to 17° with glycerol. The free surface energy of 75 ± 3 mJ/m2 for all the coatings were determined. Polar component was calculated as the main component of surface energy, caused by the presence of strong polar PO43- and OH- bonds. In vitro studies showed that low Cu and Zn amounts (~0.4 at.%) in the coatings promoted high motility of human adipose-derived multipotent mesenchymal stromal cells (hAMMSC) on the implant/cell interface and subsequent cell ability to differentiate into osteoblasts. In vivo study demonstrated 100% ectopic bone formation only on the surface of the CaP coating on Ti. The Zn- and Cu-containing CaP coatings on both substrates and the CaP coating on the Ti-40Nb alloy slightly decreased the incidence of ectopic osteogenesis down to 67%. The MAO coatings showed antibacterial efficacy against Staphylococcus aureus and can be arranged as follows: Zn-CaP/Ti > Cu-CaP/TiNb, Zn-CaP/TiNb > Cu-CaP/Ti.

Ectopic bone formation and systemic bone loss in a transmembrane TNF-driven model of human spondyloarthritis.

BACKGROUND: The transmembrane-TNF transgenic mouse, TgA86, has been shown to develop spontaneously peripheral arthritis with signs of axial involvement. To assess similarity to human spondyloarthritis, we performed detailed characterization of the axial, peripheral, and comorbid pathologies of this model. METHODS: TgA86 bone pathologies were assessed at different ages using CT imaging of the spine, tail vertebrae, and hind limbs and characterized in detail by histopathological and immunohistochemical analysis. Cardiac function was examined by echocardiography and electrocardiography and bone structural parameters by µCT analysis. The response of TgA86 mice to either early or late anti-TNF treatment was evaluated clinically, histopathologically, and by µCT analysis. RESULTS: TgA86 mice developed with 100% penetrance spontaneous axial and peripheral pathology which progressed with time and manifested as reduced body weight and body length, kyphosis, tail bendings, as well as swollen and distorted hind joints. Whole-body CT analysis at advanced ages revealed bone erosions of sacral and caudal vertebrae as well as of sacroiliac joints and hind limbs and, also, new ectopic bone formation and eventually vertebral fusion. The pathology of these mice highly resembled that of SpA patients, as it evolved through an early inflammatory phase, evident as enthesitis and synovitis in the affected joints, characterized by mesenchymal cell accumulation, and neutrophilic infiltration. Subsequently, regression of inflammation was accompanied by ectopic bone formation, leading to ankylosis. In addition, both systemic bone loss and comorbid heart valve pathology were evident. Importantly, early anti-TNF treatment, similar to clinical treatment protocols, significantly reduced the inflammatory phase of both the axial and peripheral pathology of TgA86 mice. CONCLUSIONS: The TgA86 mice develop a spontaneous peripheral and axial biphasic pathology accompanied by comorbid heart valvular dysfunction and osteoporosis, overall reproducing the progression of pathognomonic features of human spondyloarthritis. Therefore, the TgA86 mouse represents a valuable model for deciphering the role of transmembrane TNF in the pathogenic mechanisms of spondyloarthritis and for assessing the efficacy of human therapeutics targeting different phases of the disease.

Superior CKIP-1 sensitivity of orofacial bone-derived mesenchymal stem cells in proliferation and osteogenic differentiation compared to long bone-derived mesenchymal stem cells.

Maxillofacial bone defects caused by multiple factors, including congenital deformations and tumors, have become a research focus in the field of oral medicine. Bone tissue engineering is increasingly regarded as a potential approach for maxillofacial bone repair. Mesenchymal stem cells (MSCs) with different origins display various biological characteristics. The aim of the present study was to investigate the effects of casein kinase­2 interaction protein­1 (CKIP­1) on MSCs, including femoral bone marrow­derived MSCs (BMMSCs) and orofacial bone­derived MSCs (OMSCs), isolated from the femoral and orofacial bones of wild­type (WT) and CKIP­1 knockout (KO) mice. MSCs were isolated using collagenase II and the main biological characteristics, including proliferation, apoptosis and osteogenic differentiation, were investigated. Subcutaneous transplantation of MSCs in mice was also performed to assess ectopic bone formation. MTT and clone formation assay results indicated that cell proliferation in the KO group was increased compared with the WT group, and OMSCs exhibited significantly increased levels of proliferation compared with BMMSCs. However, the proportion of apoptotic cells was not significantly different between CKIP­1 KO OMSCs and BMMSCs. Furthermore, it was revealed that osteogenic differentiation was increased in CKIP­1 KO MSCs compared with WT MSCs, particularly in OMSCs. Consistent with the in vitro results, enhanced ectopic bone formation was observed in CKIP­1 KO mice compared with WT mice, particularly in OMSCs compared with BMMSCs. In conclusion, the present results indicated that OMSCs may have a superior sensitivity to CKIP­1 in promoting osteogenesis compared with BMMSCs; therefore, CKIP­1 KO in OMSCs may serve as an efficient strategy for maxillofacial bone repair.

Dendriform pulmonary ossification complicated by recurrent spontaneous pneumothorax: Two case reports and a review of the literature.

BACKGROUND: Dendriform pulmonary ossification (DPO) is a rare disease characterized by the presence of mature bone in the lung parenchyma with typical radiologic findings of diffuse and numerous calcified nodules. We present two cases of primary DPO complicated by recurrent spontaneous pneumothorax. CASE PRESENTATION: Case 1 is a 53-year-old male with recurrent pneumothorax unresponsive to chest tube drainage or subtotal pleurectomy via video assisted thoracoscopy (VATS) who was finally treated with talc pleurodesis. Chest computed tomography (CT) revealed bilateral partially calcified reticulonodular opacities with a basal predominance. VATS biopsy revealed patchy foci of fibrous organizing pneumonia with multifocal ossifications confirming DPO histopathology. Pneumothorax recurred on the same side eight months later managed with talc pleurodesis. Case 2 is a 45-year-old Caucasian male who presented for evaluation of three prior spontaneous left-sided pneumothoraces occurring over eight years, treated with chest tube drainage and eventual talc pleurodesis. Chest CT demonstrated multiple high attenuation peripheral branching opacities greatest in the left lower lobe with several nonspecific, non-calcified pulmonary nodules. VATS biopsy revealed cicatrical organizing pneumonia with associated extensive intraalveolar ossification consistent with DPO. CONCLUSIONS: We describe two cases of DPO complicated by recurrent pneumothorax and reviewed the world literature. Summarized findings included a propensity for middle-aged males with a generally indolent course though pneumothorax was often refractory to initial chest tube drainage requiring more definitive mechanical management. There was also a predominance of primary disease without associated causes other than several reports of obstructive lung disease (asthma).

Minimally Invasive Lumbar Decompression and Removal of Symptomatic Heterotopic Bone Formation After Spinal Fusion with Recombinant Human Bone Morphogenetic Protein-2.

We present a case of symptomatic heterotopic bone formation following revision of posterolateral lumbar fusion/instrumentation and "off-label" use of recombinant human bone morphogenetic protein-2, treated successfully with the use of a minimally invasive tubular approach.

The implication of the notch signaling pathway in biphasic calcium phosphate ceramic-induced ectopic bone formation: A preliminary experiment.

Calcium phosphate (BCP) ceramic is a promising material in bone regeneration because it was proved biocompatible, osteoconductive, osteoinductive, and effective. Although it manifests favorable characteristics in critical-sized bone defects repair, the mechanism of its osteoinduction is still unclear. In the present study, we studied the mechanism of ectopic bone formation, with interest in the Notch signaling pathway. BCP ceramics with or without Notch signaling inhibitor RO4929097 were cocultured with bone marrow-derived stem cells in vitro. The expression of osteogenesis (OPN/Col/Runx2) and Notch signaling pathway-related genes (Hes1/Jagged/Notch1) were increased in the BCP group compared with the control group without BCP but significantly decreased after adding RO4929097. Furthermore, a higher level of alkaline phosphatase activity was observed in the BCP group compared with RO4929097 and control group separately. For further confirmation, the intramuscularly ectopic implantation models of Beagle dogs were used. Quantitative real-time polymerase chain reaction showed a similar trend with the in vitro experiment. Histological and histomorphometric analysis indicated that bone formation was delayed by RO4929097. These findings illustrated that the Notch signaling pathway plays a pivotal role in bone formation induced by BCP; Notch signaling pathway may positively influence ectopic bone formation by promoting BMSCs to differentiate toward osteoblasts.

In vitro measurement of the chemical changes occurring within ß-tricalcium phosphate bone graft substitutes.

Several mechanisms proposed to explain the osteoinductive potential of calcium phosphates involve surface mineralization ("bioactivity") and mention the occurrence of concentration gradients between the inner and the outer part of the implanted material. Determining the evolution of the local chemical environment occurring inside the pores of an implanted bone graft substitute (BGS) is therefore highly relevant. A quantitative and fast method was developed to measure the chemical changes occurring within the pores of ß-Tricalcium Phosphate (ß-TCP) granules incubated in a simulated body fluid. A factorial design of experiment was used to test the effect of particle size, specific surface area, microporosity, and purity of the ß-TCP granules. Large pH, calcium and phosphate concentration changes were observed inside the BGS and lasted for several days. The kinetics and magnitude of these changes (up to 2 pH units) largely depended on the processing and properties of the granules. Interestingly, processing parameters that increased the kinetics and magnitude of the local chemical changes are parameters considered to favor calcium phosphate osteoinduction, suggesting that the model might be useful to predict the osteoinductive potential of BGSs. STATEMENT OF SIGNIFICANCE: Recent results suggest that in situ mineralization of biomaterials (polymers, ceramics, metals) might be key in their ability to trigger ectopic bone formation. This is the reason why the effect on in situ mineralization of various synthesis parameters of ß-tricalcium phosphate granules was studied (size, microporosity, specific surface area, and Ca/P molar ratio). To the best of our knowledge, this is the first article devoted to the chemical changes occurring within the pores of a bone graft substitute. We believe that the manuscript will prove to be highly important in the design and mechanistic understanding of drug-free osteoinductive biomaterials.

Enhanced biocompatibility and improved osteogenesis of coralline hydroxyapatite modified by bone morphogenetic protein 2 incorporated into a biomimetic coating.

OBJECTIVES: (1) To determine whether the biocompatibility of coralline hydroxyapatite (CHA) granules could be improved by using an octacalcium phosphate (OCP) coating layer, and/or functionalized with bone morphogenetic protein 2 (BMP-2), and (2) to investigate if BMP-2 incorporated into this coating is able to enhance its osteoinductive efficiency, in comparison to its surface-adsorbed delivery mode. METHODS: CHA granules (0.25 g per sample) bearing a coating-incorporated depot of BMP-2 (20 µg/sample) together with the controls (CHA bearing an adsorbed depot of BMP-2; CHA granules with an OCP coating without BMP-2; pure CHA granules) were implanted subcutaneously in rats (n = 6 animals per group). Five weeks later, the implants were retrieved for histomorphometric analysis to quantify the volume of newly generated bone, bone marrow, fibrous tissue and foreign body giant cells (FBGCs). The osteoinductive efficiency of BMP-2 and the rates of CHA degradation were also determined. RESULTS: The group with an OCP coating-incorporated depot of BMP-2 showed the highest volume and quality or bone, and the highest osteoinductive efficacy. OCP coating was able to reduce inflammatory responses (improve biocompatibility), and also simple adsorption of BMP-2 to CHA achieved this. CONCLUSIONS: The biocompatibility of CHA granules (reduction of inflammation) was significantly improved by coating with a layer of OCP. Pure surface adsorption of BMP-2 to CHA also reduced inflammation. Incorporation of BMP-2 into the OCP coatings was associated with the highest volume and quality of bone, and the highest biocompatibility degree of the CHA granules. CLINICAL SIGNIFICANCE: Higher osteoinductivity and improved biocompatibility of CHA can be obtained when a layer of BMP-2 functionalized OCP is deposited on the surfaces of CHA granules.