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Summary Porous biomaterials promote osseointegration. We have prepared porous titanium cylinders by additive manufacturing from titanium beads. Two types of morphology were tested: cylinders with geometric pores or mimicking trabecular microarchitecture. Cylinders were decontaminated and cleaned by HF/HNO3 to remove unmelted balls. Surgical implantation in ewes was performed under general anesthesia and the animals were housed for 90 and 270 days. The femoral condyles were collected and analyzed by nanoCT, embedded in pMMA and analyzed by histomorphometry. No significant difference was found in terms of bone volume or bone/titanium interface between the two types of cylinders. There was no evolution over time except for the mineralization rates which decreased, reflecting the effect of the aging of the animals. The influence of the pores (geometrical or “natural”) did not influence osseointegration. HF/HNO3 etching treatments are effective on the outermost surfaces but do not seem to reach the central cavities of the samples. Finally, osseointegration seems to occur only in the few millimeters around the periphery of the implants and does not extend in the center. This is explained by the absence of stress transmission within the very rigid metal cylinders, preventing bone modeling and remodeling.

Supplementary Data from Pharmacologic inhibitors of IκB kinase suppress growth and migration of mammary carcinosarcoma cells in vitro and prevent osteolytic bone metastasis in vivo

The NF-κB signaling pathway is known to play an important role in the regulation of osteoclastic bone resorption and cancer cell growth. Previous studies have shown that genetic inactivation of IκB kinase (IKK), a key component of NF-κB signaling, inhibits osteoclastogenesis, but the effects of pharmacologic IKK inhibitors on osteolytic bone metastasis are unknown. Here, we studied the effects of the IKK inhibitors celastrol, BMS-345541, parthenolide, and wedelolactone on the proliferation and migration of W256 cells in vitro and osteolytic bone destruction in vivo. All compounds tested inhibited the growth and induced apoptosis of W256 cells as evidenced by caspase-3 activation and nuclear morphology. Celastrol, BMS-345541, and parthenolide abolished IL1β and tumor necrosis factor α–induced IκB phosphorylation and prevented nuclear translocation of NF-κB and DNA binding. Celastrol and parthenolide but not BMS-345541 prevented the activation of both IKKα and IKKβ, and celastrol inhibited IKKα/β activation by preventing the phosphorylation of TAK1, a key receptor–associated factor upstream of IKK. Celastrol and parthenolide markedly reduced the mRNA expression of matrix metalloproteinase 9 and urinary plasminogen activator, and inhibited W256 migration. Administration of celastrol or parthenolide at a dose of 1 mg/kg/day suppressed trabecular bone loss and reduced the number and size of osteolytic bone lesions following W256 injection in rats. Histomorphometric analysis showed that both compounds decreased osteoclast number and inhibited bone resorption. In conclusion, pharmacologic inhibitors of IKK are effective in preventing osteolytic bone metastasis in this model and might represent a promising class of agents to the prevention and treatment of metastatic bone disease associated with breast cancer. [Mol Cancer Ther 2009;8(8):2339–47]

ObjectivesBotulinum toxin (BTX) induces muscle paralysis. It is used in human in masticatory muscles injections performed often repeatedly. A single BTX injection in masticatory muscles in animal induces mandibular bone loss (alveolar and condylar) with a muscle enthesis hypertrophic metaplasia. Our aim was to evaluate mandibular bone changes after unilateral repeated injections of BTX in temporal and masseter muscles in adult rats.Materials and MethodsMature male rats were randomized into 3 groups: one, two or three injections. Each injection was performed 4 weeks after the prior injection. Each rat received injections in right masseter and temporalis muscles. The left side was the control side. Microcomputed tomography was used to perform 2D and 3D analyses.ResultsBone loss was evidenced on the right sides of alveolar and condylar bone. Alveolar bone volume increased in both control left side and injected right side whereas condylar bone volume remained constant in all groups, for both left and right sides. Enthesis bone hypertrophic metaplasias were evidenced on the BTX injected sides without any modification with the number of injections.ConclusionsBTX repeated injections in masticatory muscles lead to major mandibular condylar and alveolar bone loss that does not worsen. They lead to the occurrence of an enthesis bone proliferation that is not dependent on the number of injections.Clinical relevanceThese results are an argument for the safety of BTX injections in masticatory muscles in human.

Aseptic osteonecrosis of the hip (AON) is a rare, but well-known pathology in rheumatology and orthopedic surgery that is a necrosis of the articular cartilage secondary to a necrosis of the subchondral bone. The microscopic aspect is well known, but the microCT aspect has never been reported or correlated with histopathological findings. The objective of this study was to improve the knowledge of the pathophysiology of AON using histochemistry and microCT. One hundred and sixty femoral heads with stage 3 or 4 AON were analyzed: one half of the head was sent for microCT analysis after impregnation with phosphotungstic acid (PTA) and the other half was used for histological analysis without decalcification. The microCT analysis provides relevant information on the cracked articular cartilage and the relationship with the necrotic subchondral trabecular bone well illustrated on three videos. In histology, Goldner's trichrome showed that the articular cartilage remains well preserved for a long time. In addition, toluidine blue staining reveals a modeling process, i.e. the apposition of new bone without prior resorption by osteoclasts. Rhodamine B staining (fluorescence analysis) reveals that the osteonecrotic trabeculae and subchondral bone were devoid of osteocytes. Areas of peri-necrotic osteosclerosis are due to direct bone formation on the surface of pre-existing necrotic trabeculae.

Resume Contexte Les etudes tribologiques ont montre que les couples les plus utilises pour les protheses de hanche sont les protheses metal-polyethylene et alumine-alumine. Avec le temps, les produits d’usure s’accumulent dans la cavite articulaire et dans les tissus peri-prothetiques. Bien que le polyethylene et les metaux soient facilement identifiables en microscopie, les particules d’alumine restent tres difficiles a identifier. Hypothese Nous avons evalue le colorant azoique fluorescent, lumogallion, comme methode histochimique specifique pour les particules d’alumine dans les tissus peri-prothetiques. Materiel et methodes Chez 28 patients ayant eu une revision d’une prothese alumine-alumine, les tissus peri-prothetiques ont ete preleves et inclus en paraffine; les coupes ont ete colorees en HPS ou au lumogallion. Elles ont ete examinees en microscopie optique et a fluorescence pour detecter les particules d’usure. Certaines coupes ont ete contre-colorees par le DAPI, detectant les noyaux cellulaires. Resultats Les particules d’alumine, tres difficiles a identifier sur les coupes colorees en HPS, etaient clairement mises en evidence par le lumogallion avec une fluorescence orange-vif. La coloration a revele la presence de plusieurs milliers de particules par coupe. Seuls deux patients n’avaient pas de particule decelable. Le lumogallion identifie de nombreuses particules, invisibles en HPS, dans les macrophages, les synoviocytes et les fibroblastes. Conclusion Cette coloration, validee en pathologie neuromusculaire pour l’identification des particules d’alumine utilisees comme adjuvant vaccinal, a donne des resultats probants ici. Les particules d’alumine sont alterees lorsqu’elles sont phagocytees par les macrophages. Le lumogallion montre la presence de milliers de particules d’usure liberees par les protheses alumine-alumine dans les tissus peri-prothetiques. Niveau de preuve V: etude d’opinion d’experts.

International audience; The use of resorbable devices for osteosynthesis has become a subject of interest. Nacre has been proposed as a resorbable and osteoconductive material favoring bone apposition without triggering an inflammatory reaction. We compared the in vivo osseointegration and erosion of nacre screws in an animal model with titanium screws. Implantation of similar nacre and titanium screws was performed in the femoral condyles of adult rats. Animals (n = 41) were randomized in four groups sacrificed at day one, 1, 6, and 12 months. Microcomputed tomography (microCT) allowed 3D morphometry of erosion of nacre. Osseointegration was measured as the volume of trabecular bone bone volume/tissue volume (BV/TV) in a standardized volume of interest around each screw. Undecalcified bone histol-ogy was also done. Gross examination revealed a similar clinical osseointegration for titanium and nacre screws. A progressive erosion of nacre screws, but no erosion of titanium screws, was observed in microCT. The volume of nacre screws progressively decreased over time whereas no modification occurred for titanium. For titanium screws, BV/TV remained stable throughout the study. For nacre screws, the BV/TV decrease was not statistically different. A significant difference was found between nacre and titanium screws at 6 months but not at 12 months. The screw heads, outside the bone shaft, were not eroded even after 12 months. Erosion of nacre occurred during the entire study period, only within the bone shaft in direct contact with bone marrow. Bone apposition was observed on nacre surfaces without signs of erosion. Nacre is a promising biomaterial in maxillofacial surgery. K E Y W O R D S erosion, maxillofacial surgery, mother of pearl, nacre, osseointegration, resorbable device, titanium screw

International audience; Aim: To compare head-to-head the effects of dapagliflozin and liraglutide on bone strength and bone material properties in a pre-clinical model of diabetes-obesity. Materials and methods: Combined low-dose streptozotocin and high fat feeding were employed in mice to promote obesity, insulin resistance and hyperglycaemia. Mice were administered daily for 28 days with saline vehicle, 1mg/kg dapagliflozin or 25 nmol/kg liraglutide. Bone strength was assessed by three-point bending and nanoindentation. Bone material properties were investigated by Fourier transform infrared microspectroscopy/imaging. Results: Although diabetic controls presented with dramatic reductions in mechanical strength, no deterioration of bone microarchitecture was apparent. At the tissue level, significant alterations in phosphate/amide ratio, carbonate/phosphate ratio, tissue water content, crystal size index, collagen maturity and collagen glycation were observed and linked to alteration of matrix biomechanics. Dapagliflozin and liraglutide failed to improve bone strength by 3-point bending or bone microarchitecture during the 28-day-treatment period. At bone formation site, dapagliflozin enhanced phosphate/amide ratio, mineral maturity, and reduced tissue water content, crystal size index and collagen glycation. Liraglutide had significant effects on phosphate/amide ratio, tissue water content, crystal size index, mature collagen crosslinks, collagen maturity and collagen glycation. At bone formation site, both drugs modulated matrix biomechanics. Conclusions: This study highlighted that these two molecules are effective in improving bone material properties and modulating matrix biomechanics at bone formation site. This study also highlighted that the resulting effects on bone material properties are not identical between dapagliflozin and liraglutide and not only mediated by lower blood glucose.

Objective Botulinum toxin (BTX) is injected into masticatory muscles to treat various conditions. Animal studies have demonstrated bone loss at the condylar and alveolar regions of the mandible after BTX injection into masticatory muscles. The aim of the present study was to investigate mandibular bone changes in patients who received BTX injections in masticatory muscles. Study Design Twelve adult patients who received BTX injections into masticatory muscles were included in this study. Cone beam computed tomography (CBCT) was performed before and 12 months after the injection. The condylar and alveolar regions of the mandible were analyzed by using texture analysis of the CBCT images with the run length method. Condylar cortical thickness was measured, and 3-dimensional analysis of the mandible was also performed. Six patients who did not receive BTX injections were used as controls. Results A run length parameter (gray level nonuniformity) was found to be increased in condylar and alveolar bones. A significant cortical thinning was found at the anterior portion of the right condyle. Three-dimensional analysis showed significant changes in the condylar bone and at the digastric fossa. No changes in mandibular angles were found. Conclusions This study identified mandibular bone changes in adult patients who received BTX injection into masticatory muscles.

Diabetes mellitus has become a serious problem associated with health complications, such as metabolism disorders and liver-kidney dysfunction. The inadequacies associated with conventional medicines have led to a determined search for alternative natural therapeutic agents. The present study was conducted to evaluate the hypoglycemic, antilipidemic, and antioxidant effects of EGCG in surviving diabetic mice. Alloxan diabetic mice were treated with EGCG. Their bloods were collected and submitted to various biochemical measurements, including blood glucose, cholesterol, triglycerides, urea, creatinine, and transaminases. Their livers and kidneys were isolated to assess oxidative damage and to perform histological analysis. Both EGCG and insulin treatment of diabetic mice resulted in a significant reduction in fasting blood glucose levels. EGCG supplementation also ameliorated hepatic as well as renal toxicity indices. Moreover, diabetic mice injected with EGCG exhibited significant changes in antioxidant enzyme activities in the liver and kidney. Histological analyses also showed that it exerted an ameliorative action on these organs and efficiently protected the liver-kidney functions of diabetic mice. EGCG was found to bind α-amylase, PTP1B, and α-glucosidase with good affinities ranging from -6.1 to -8.4 kcal/mol. The findings revealed that EGCG administration induced attractive curative effects on diabetic mice, particularly in terms of liver-kidney function. EGCG can, therefore, be considered as a potential strong candidate for future applications to treat and alleviate diabetic burden. Its pharmacokinetics, high affinities, and molecular interactions with the targeted receptors satisfactory explain the in vivo findings.

Large bone defects are a challenge for orthopedic surgery. Natural (bone grafts) and synthetic biomaterials have been proposed but several problems arise such as biomechanical resistance or viral/bacterial safety. The use of metallic foams could be a solution to improve mechanical resistance and promote osseointegration of large porous metal devices. Titanium cylinders have been prepared by additive manufacturing (3D printing/rapid prototyping) with a geometric or trabecular microarchitecture. They were implanted in the femoral condyles of aged ewes; the animals were left in stabling for 90 and 270 days. A double calcein labeling was done before sacrifice; bones were analyzed by histomorphometry. Neither bone volume, bone/titanium interface nor mineralization rate were influenced by the cylinder’s microarchitecture; the morphometric parameters did not significantly increase over time. Bone anchoring occurred on the margins of the cylinders and some trabeculae extended in the core of the cylinders but the amount of bone inside the cylinders remained low. The rigid titanium cylinders preserved bone cells from strains in the core of the cylinders. Additive manufacturing is an interesting tool to prepare 3D metallic scaffolds, but microarchitecture does not seem as crucial as expected and anchoring seems limited to the first millimeters of the graft.

Death of osteocytes is synonymous of bone death. Aseptic osteonecrosis of the femoral head is a lesion characterized by the death of osteocytes occurring after major vascular changes. The evolution may lead to hip osteoarthritis, which requires total hip arthroplasty in most cases. Evolution of aseptic osteonecrosis in four radiological stages is well known. We analyzed 24 femoral heads from patients with osteonecrosis or osteoarthritis, retrieved at the time of surgery for a hip arthroplasty. The aim of the study was to clearly identify the necrotic bone from the living bone in the histological samples. The femoral heads were sawed, and a large sample was harvested in the superior zone; it was stained en-bloc with rhodamine dissolved in formalin to make the osteocytes fluorescent under UV light microscopy. Undecalcified sections, 7 μm thick, were obtained on a heavy-duty microtome. A micrographic analysis using two UV excitation wavelengths visualized the living osteocytes (in green) and the bone matrix (in blue). A simple method to prepare combined images is described. In addition, the blocks can be analyzed by confocal microscopy to visualize more details. It is possible to identify at low magnification the osteocytes within the bone matrix and the osteonecrotic areas where osteocytes have disappeared. Identification of osteocytes showed that newly formed bone packets are laid on dead trabeculae in patients with aseptic osteonecrosis or with osteoarthritis. In the osteosclerotic areas, the enlarged trabeculae have a dead central core surrounded by recently apposed bone structure units.

The involvement of a gut-bone axis in controlling bone physiology has been long suspected, although the exact mechanisms are unclear. We explored whether glucose-dependent insulinotropic polypeptide (GIP)-producing enteroendocrine K cells were involved in this process. The bone phenotype of transgenic mouse models lacking GIP secretion (GIP-GFP-KI) or enteroendocrine K cells (GIP-DT) was investigated. Mice deficient in GIP secretion exhibited lower bone strength, trabecular bone mass, trabecular number, and cortical thickness, notably due to higher bone resorption. Alterations of microstructure, modifications of bone compositional parameters, represented by lower collagen cross-linking, were also apparent. None of these alterations were observed in GIP-DT mice lacking enteroendocrine K cells, suggesting that another K-cell secretory product acts to counteract GIP action. To assess this, stable analogues of the known K-cell peptide hormones, xenin and GIP, were administered to mature NIH Swiss male mice. Both were capable of modulating bone strength mostly by altering bone microstructure, bone gene expression, and bone compositional parameters. However, the two molecules exhibited opposite actions on bone physiology, with evidence that xenin effects are mediated indirectly, possibly via neural networks. Our data highlight a previously unknown interaction between GIP and xenin, which both moderate gut-bone connectivity. © 2020 American Society for Bone and Mineral Research.

Total hip arthroplasty uses commercial devices that combine different types of biomaterials. Among them, metals, ceramics and metal oxides can be used either in the prosthesis itself or in the cement used to anchor them in the bone. Over time, all of these materials can wear out and release particles that accumulate in the periprosthetic tissues or can migrate away. We used histology blocks from 15 patients (5 titanium metallosis, 5 alumina prostheses, 5 with altered methacrylic cement) to perform a microCT study and compare it with conventional histology data. An EDS-SEM analysis was done to characterise the atomic nature of the materials involved. A morphometric analysis was also performed in 3D to count the particles and assess their density and size. The metallic particles appeared to be the largest and the ceramic particles the finest. However, microCT could not reveal the wear particles of radiolucent biomaterials such as polyethylene and the very fine zirconia particles from cement fragmentation. MicroCT analysis can reveal the extent of the accumulation of these debris in the periprosthetic tissues. LAYOUT DESCRIPTION: Hip prostheses progressively degrade in the body by releasing wear debris. They accumulate in the periprosthetic tissues. Microcomputed tomography was used to image three types of radio-opaque wear debris: metal, ceramic and zirconia used in the bone cements.

Summary Microcomputed X-ray tomography (microCT), developed since the late 1990s, is a miniaturized version of the tomographs used daily in medical imaging. It produces vascular images that are different from those obtained by microradiography, in particular by facilitating the vision in space, thus understanding microvascularisation. The anatomical specimens, once treated with formalin, are injected with a mixture made of gelatin containing a contrast product (barium) and then analyzed by microCT. The acquisition times that can exceed 24 hours and metal sheets used for X-ray filtering vary according to the sample. The projection images are reconstructed to produce 2D sections. These are combined for the reconstruction of 3D models using a volume rendering software. Four examples will allow the imaging of microvascularization: the inferior alveolar nerve, the cerebral cortex and pia-mother, brain stem, central gray nuclei (ganglia at the base of the brain). Small capillaries are highlighted using high-end software for reconstruction. Conventional software or freeware cause a considerable loss of information on small vessels that are not visualized. The VGStudio max high-end software allows the production of videos that are particularly useful for 3D exploration and teaching (four videos are provided with this article).

Background The donkey pericardium is considered a good candidate to manufacture percutaneous heart valves based upon its thinness, low cellularity and undulating collagen bundles and laminates. Decellularization represents an avenue worth exploring, should its superiority to glutaraldehyde-treated pericardium be demonstrated. Materials and methods Donkey pericardium was divided into two groups: regular glutaraldehyde fixation and mild decellularization. The treated pericardia were observed using scanning electron microscopy, histology and transmission electron microscopy. Tensile tests were performed along the axial and perpendicular directions, with the data fitted into both the Gasser–Ogden–Holzapfel (GOH) material model and the Fung’s anisotropic one. Results The microstructures of the pericardia processed by the two protocols were similar, showing collagen bundles and laminates free of flaws. The decellularization eliminated most of the cells, however leaving the structure somehow compressed. The collagen filaments in bundles were slightly blurry. The anisotropy rates of the non-decellularized specimens were almost identical to the decellularized ones. The decellularized pericardium appeared stiffer. Conclusion The decellularization proved to be effective. However, it makes the tissue stiffer, which may lead to higher shear concentration during cardiac cycles and reduce its wavy microstructure. Therefore, it appears premature to select decellularized donkey pericardium to manufacture heart valves.

Background Cementless total hip arthroplasty (THA) is a common procedure producing excellent clinical results. Their long-term survival is nevertheless burdened by loosening of the acetabular part caused by changes in the distribution of strains around the cup. In this context the RM-Pressfit® cup has been developed, resulting in a more harmonious distribution of the strains. Hypothesis Texture analysis of X-ray films can evaluate the evolution of trabecular bone micro-architecture during the five years following THA with a RM-Pressfit® cup. Material and method A monocentric series of 46 hips was reviewed regularly within five years post-surgery. Radiographic evaluation of the operated hip was done on frontal digitized radiographs of the pelvis to follow evolution of bone micro-architecture in the #2 zone of De Lee and Charnley. Texture analysis using fractal algorithms was done at D0, 6 months, 1, 2 and 5 years post-THA. The fractal methods used included the skyscrapers and the dynamic blanket methods with 3 different structuring elements (a cross, a horizontal and a vertical vector). Results The RM-Pressfit® caused significant changes in the distribution of strains around the acetabulum that preserved the bone volume over a 5-year period post-surgery. This corresponds to an improvement of the trabecular micro-architecture around the acetabular cups. Conclusion A statistically significant increase in the four fractal dimensions considered corresponded to an improved trabecular bone micro-architecture revealed by texture analysis, a non-invasive method that can be used on digitized X-ray images. Level of evidence IIIb, Case control study, retrospective design.

Background Cementless total hip arthroplasty (THA) is a common procedure producing excellent clinical results. Their long-term survival is nevertheless burdened by loosening of the acetabular part caused by changes in the distribution of strains around the cup. In this context the RM-Pressfit® cup has been developed, resulting in a more harmonious distribution of the strains. Hypothesis Texture analysis of X-ray films can evaluate the evolution of trabecular bone micro-architecture during the five years following THA with a RM-Pressfit® cup. Material and method A monocentric series of 46 hips was reviewed regularly within five years post- surgery. Radiographic evaluation of the operated hip was done on frontal digitized radiographs of the pelvis to follow evolution of bone micro-architecture in the #2 zone of De Lee and Charnley. Texture analysis using fractal algorithms was done at D0, 6 months, 1, 2 and 5 years post-THA. The fractal methods used included the skyscrapers and the dynamic blanket methods with 3 different structuring elements (a cross, a horizontal and a vertical vector). Results The RM-Pressfit® caused significant changes in the distribution of strains around the acetabulum that preserved the bone volume over a 5-year period post-surgery. This corresponds to an improvement of the trabecular micro-architecture around the acetabular cups. Conclusion A statistically significant increase in the four fractal dimensions considered corresponded to an improved trabecular bone micro-architecture revealed by texture analysis, a non-invasive method that can be used on digitized X-ray images. Level of evidence IIIb, Case control study, retrospective design.

International audience; Most osteolytic tumors are in fact mixed and contain an osteoblastic component associated with the predominant osteolytic areas. This metaplastic woven bone is always evidenced by histological analysis even in the absence of radiological expression. Metaplastic bone formation reflects the activation of new osteoblasts coming from the stimulation of the dormant lining cells. Twelve patients with secondary metastases of the iliac crest evidenced by hot spots on a Tc-MBP san were diagnosed by histomorphometry on bone biopsies. Fourier Transformed InfraRed analysis and Imaging (FTIRI) was used on 4μm thick sections of undecalcified bone. The mineralization degree, carbonate substitution, crystallinity and the cross-links ratio of collagen (1660/1690cm bands) were determined. The matrix characteristics were analyzed and imaged in the pre-existing residual bone and in the metaplastic woven bone in the vicinity of the tumor cells. FTIRI provided images of the phosphate, amide and combination of peak ratio after having selected the peaks of interest. In addition, the matrix properties can be measured and compared between the old and newly-formed bones. Woven bone appeared poorly calcified with a low phosphate/amide ratio (P=0.03) crystallinity (P; La plupart des tumeurs ostéolytiques sont en fait mixtes et contiennent un composant ostéoblastique associé aux zones ostéolytiques qui prédominent. Cet os métaplasique fibreux, non lamellaire, est toujours mis en évidence histologiquement même en l’absence d’expression radiologique. La formation d’os métaplasique reflète l’activation de nouveaux ostéoblastes provenant de la stimulation des cellules bordantes. Douze patients présentant des métastases osseuses secondaires mises en évidence par des foyers chauds en scintigraphie au 99Tc-MBP ont été diagnostiqués en histomorphométrie sur des biopsies osseuses de la crête iliaque. L’analyse et l’imagerie infrarouges à transformée de Fourier (FTIRI) ont été utilisées sur des coupes de 4μm d’épaisseur d’os non décalcifié. La minéralisation, le taux de substitution par des carbonates, la cristallinité et le rapport des cross-links du collagène (bandes à 1660/1690cm−1) ont été déterminés. Les caractéristiques de la matrice osseuse ont été analysées et imagées dans l’os résiduel préexistant et dans l’os métaplasique au voisinage des cellules tumorales. La FTIRI a fourni des images de la distribution des phosphates, des amides et des rapports de combinaisons de pics après avoir sélectionné les pics d’intérêt. De plus, les propriétés de la matrice ont pu être mesurées et comparées entre l’os résiduel et l’os métaplasique. Ce dernier est apparu faiblement calcifié avec un rapport phosphate/amide (p=0,03), une cristallinité (p&nbsp

Biomaterials are used in the granular form to fill small bone defects. Granules can be prepared with a grinder from trabecular bone samples or provided as synthetic biomaterials by industry. Granules occupy the 3D-space and create a macroporosity allowing invasion of vascular and bone cells when the inter-granular pores are larger than 300 µm. We compared the 3D-porosity of granule stacks obtained or prepared with nine biomaterials Osteopure® , Lubboc® , Bio-Oss® , CopiOs® , TCP Dental® , TCP Dental HP® , KeraOs® , and TCH® in comparison with that of human trabecular bone. For each biomaterial, two sizes of granules were analyzed: 250-1000 and 1000-2000 µm. Microcomputed tomography determined porosity and microarchitectural characteristics of granular stacks and Raman microspectroscopy was used to analyze their composition. Stacks of 250-1000 µm granules had a much lower porosity than 1000-2000 µm granules and the maximum frequency of pores was always centered at 200-250 µm. One biomaterial contained substantial amount of cortical bone (Bio-Oss® ). The highest porosity and pore size was obtained with TCP Dental HP. Raman spectroscopy found differences in biomaterials of the same composition. Stacks of granules represent 3D scaffolds resembling trabecular bone with an interconnected porous microarchitecture. Small granules have created pores

Introduction Les surcharges en fer, genetiques ou lors des thalassemies, ont pour consequence une osteoporose. Le mecanisme en cause n’est pas completement elucide. Notre objectif est d’identifier le role des differents acteurs du metabolisme du fer et particulierement de l’hepcidine mais aussi d’autres metaux que le fer dans le developpement de la perte osseuse. Materiels et methodes Nous avons etudie un modele de surcharge en fer severe, consecutif a une hepcidino-deficience, chez des souris mâles CD1 âgees de 6 mois invalidees pour le gene Bmp6 (Bmp6-/-) compare a des souris sauvages controles CD1. Ont ete evalues : le fer serique et la saturation de la transferrine, l’expression hepatique de l’ARNm de l’hepcidine par RT PCR, les concentrations en metaux (manganese, molybdene, cuivre, magnesium, zinc, aluminium) par inductively coupled plasma-Mass Spectrometry (ICP-MS). La localisation de fer dans les tissus a ete realisee par coloration de Perls. Une etude de la micro-architecture osseuse a ete realisee par microtomographie aux rayons X (micro-CT). Resultats Chez les souris Bmp6-/-, la concentration hepatique en fer (CHF) et la concentration osseuse en fer (COF) etaient significativement augmentees par rapport aux controles (p Conclusion L’invalidation du gene Bmp6 entraine : – une hepcidino deficience ; – une surcharge en fer severe au niveau osseux et tissulaire ; – une perte osseuse tres significative. La diminution de la quantite osseuse ainsi que les alterations de la micro-architecture etaient correlees au niveau d’expression hepatique d’hepcidine, mais pas aux autres parametres du fer. La perte osseuse observee pourrait etre liee a l’hepcidino-deficience elle-meme et/ou a un role de BMP6 sur le remodelage osseux.

International audience; Microcomputed tomography (microCT) is well adapted to quantitative analysis of calcified tissues but soft tissues (such as cartilage) are radiolucent and need a contrast enhancement procedure for microCT. We developed a "staining" method allowing microCT imaging of articular cartilage using uranyl acetate (UA). The method was used to see whether adult rats with a botulinum toxin (BTX) injection in masticatory muscles present a change at the condylar cartilage of the mandible in association with a localized trabecular bone loss. Human femoral head samples were used to develop the enhanced contrast method using UA or lanthanides (recently proposed as a substitute for UA). The method was then applied to the condylar cartilage of rat mandibles. Mature male rats (n=11) were randomized into 2 groups: control (CTRL; n=4) and BTX group (n=7). Rats of the BTX group received a single injection of BTX into the right M. Masseter and M. Temporalis. Rats of the CTRL group were similarly injected with saline. Rats were sacrificed 4 weeks after injection. Condyles were harvested, fixed in formalin and immersed in UA. MicroCT was performed for bone and cartilage measurements. After UA impregnation, articular cartilage of human femoral head samples was clearly seen on its full thickness whereas lanthanides produced a much less pronounced contrast, with a faint labeling at the upper layer. In BTX rats, microCT analysis showed a significant bone loss at the right condyles. After UA, the whole thickness of articular cartilage was clearly evidenced. Cartilage thickness measurement showed no difference when comparing the right with the left sides of the BTX group nor between the two sides of the CTRL group. Contrast enhancement with UA is a simple technique allowing quantitative analysis of cartilage by microCT.290 words.

International audience; Non-biodegradable porous polystyrene (PS) scaffolds, composed of microfibers, have been prepared by electrospinning for the reconstruction of large bone defects. PS microfibers were prepared by incorporating β-TCP grains inside the polymer or grafting gold nanoparticles surface functionalized with mercaptosuccinic acid. Cytocompatibility of the three types of scaffolds (PS, β-TCP-PS and Au-PS) was studied by seeding human mesenchymal stem cells. Biocompatibility was evaluated by implanting β-TCP-PS and Au-PS scaffolds into a critical size (4mm) calvarial defect in mice. Calvaria were taken 6, 9, and 12 weeks after implantation; newly formed bone and cellular response was analyzed by microcomputed tomography (microCT) and histology. β-TCP-PS scaffolds showed a significantly higher cell proliferation in vitro than on PS or Au-PS alone; clearly, the presence of β-TCP grains improved cytocompatibility. Biocompatibility study in the mouse calvaria model showed that β-TCP-PS scaffolds were significantly associated with more newly-formed bone than Au-PS. Bone developed by osteoconduction from the defect margins to the center. A dense fibrous connective tissue containing blood vessels was identified histologically in both types of scaffolds. There was no inflammatory foci nor giant cell in these areas. AuNPs aggregates were identified histologically in the fibrosis and also incorporated in the newly-formed bone matrix. Although the different types of PS microfibers appeared cytocompatible during the in vitro experiment, they appeared biotolerated in vivo since they induced a fibrotic reaction associated with newly formed bone.

Objectives: Multiple myeloma (MM) is characterized by the occurrence of osteolytic lesions. MM treatment usually involves antiresorptive drugs (mainly bisphosphonates). Case Report: A patient with an MM presented osteolytic lesions of the mandible. Extraction of teeth 45 and 46 was performed 5 years after the diagnosis of periodontitis. Four months later, osteonecrosis of the jaw (ONJ) was diagnosed at the extraction site. X-ray showed an extension of osteolytic lesions on the right side, close to the extraction site, without modification of the lesions on the left side. Two months later, a curettage was performed because of a painful bone sequestration. X-ray showed an extension of the osteolytic lesions on the right side. Results: Histological analysis found a vascularized plasmacytoma of the soft tissues around the ONJ. Analysis of the bone showed mixed lesions with osteonecrotic areas and living bone resorbed by active osteoclasts surrounding a plasmacytoma. The surface area of the osteolytic foci has considerably increased only close to the extraction site. Conclusions: Tooth extraction triggered an ONJ associated with bisphosphonate treatment. However, it also seemed to induce a considerable proliferation of plasma cells at the extraction site; we hypothesize that it is due to the increase in bone remodeling related to the surgical trauma.

Resume Introduction Des biomateriaux biodegradables ont ete proposes pour preparer des dispositifs orthopediques. La nacre est un materiau naturel bioerodable a base d’aragonite (carbonate de calcium). Hypothese Nous postulons que la nacre est biodegradable sans provoquer d’osteolyse et favorise l’apposition osseuse. Materiel et methodes Nous avons prepare des vis orthopediques avec la nacre de l’huitre geante Pinctada maxima. Des vis filetees (diametre 3,5 mm) ont ete implantees chez 6 brebis dans la metaphyse tibiale superieure (3 a 4 vis par animal). Leur trajectoire etait transcorticale et intramedullaire jusqu’a la corticale opposee. Les animaux ont ete stabules pendant 3 mois (n = 2) et 6 mois (n = 4). Ils n’ont pas developpe d’inflammation locale. Avant l’euthanasie, ils ont recu un double marquage par la calceine. Les echantillons osseux ont ete analyses par nanotomographie aux rayons X et histologie apres inclusion en poly(methyl methacrylate). La dimension fractale des contours des vis (mesuree par la methode des boites) a ete utilisee pour quantifier l’erosion de surface. Resultats La nanotomographie 3D a montre une erosion progressive des filets des vis qui a ete confirmee par une reduction de la dimension fractale. Histologiquement, des cellules multinucleees (d’aspect non osteoclastique) etaient visibles a la surface des vis. Aucune membrane ondulee n’a ete vue au pole basal de ces cellules, mais elles avaient des extensions dans la matiere organique de la nacre, entre les plaquettes d’aragonite. L’apposition osseuse a ete notee dans la zone transcorticale des vis avec osteoconduction a partir de l’endoste. La vitesse de mineralisation etait augmentee dans ces zones composees d’os non lamellaire en contact avec la nacre. Discussion et conclusion Des vis preparees a partir de nacre ont l’avantage d’une resorption in vivo par des cellules de nature macrophagique et une osteoconduction en contact avec le materiau sans declenchement de reaction inflammatoire locale.

OBJECTIVES: To evaluate whether the effects of botulinum neurotoxin (Botox) injection into the masseter in the mandibular condylar cartilage (MCC) and subchondral bone are transient. MATERIALS AND METHODS: Botox (0.3 U) was injected into the right masseter of 6-week-old female mice (C57BL/6; n = 16). Additionally, 16 mice were used as control and received no injections. Experimental and matching control mice were sacrificed 4 or 8 weeks after the single botox injection. Mandibles and mandibular condyles were analyzed by micro-CT and histology. Sagittal sections of condyles were stained for TRAP, toluidine blue, EdU and TUNEL. RESULTS: Bone volume fraction was significantly decreased on the subchondral bone of botox injected side when compared to control side and control mice, 4 and 8 weeks after injection. Furthermore, histological analysis revealed decrease in mineralization, cartilage thickness, TRAP activity and EdU positive cells in the MCC of the botox injected side, 4 and 8 weeks after injection. CONCLUSION: The effects of Botox injection into the masseter muscle in the MCC and subchondral bone persisted for 8 weeks after injection and were not considered transient.

Summary Osteoporosis is considered the most frequent skeletal manifestation of systemic mastocytosis (SM). We performed a retrospective analysis of sixty patients (37 males and 23 females) who underwent a bone biopsy in the assessment of SM or in the assessment of unexplained bone fragility. Thirty-three had simultaneously a bone marrow biopsy with a Jamshidi's needle; this sample was used for immunohistochemical analysis (tryptase, c-KIT. CD20, VCAM-1). Bone biopsy was realized in 42 cases in the assessment of SM to provide histologic proof of the disease and in 18 cases in the assessment of unexplained bone fragility and surprisingly revealed a SM. An increased bone turnover was observed in patients with SM with elevated eroded surfaces, osteoclast number and bone formation rate. In addition to nodules of mast cells (MC), a high number of MC was directly apposed on the trabeculae, affixed on the osteoblasts or the lining cells. The VCAM-1 adhesion protein recognizing α4β7 and α4β1 integrins may be a candidate to explain this particular adherence. One third of the bone marrow biopsies did not exhibit MC nodules or MC infiltration and led to a false negative diagnosis for SM. SM can be discovered in the assessment of fracture or osteoporosis. Transiliac bone biopsy allows for the diagnosis of the disease more accurately than bone marrow biopsy; it also provides a histomorphometric analysis of bone remodeling.

In osteogenesis imperfecta (OI), vertebrae brittleness causes thorax deformations and leads to cardiopulmonary failure. As sclerostin-neutralizing antibodies increase bone mass and strength in animal models of osteoporosis, their administration in two murine models of severe OI enhanced the strength of vertebrae in growing female Crtap−/− mice but not in growing male Col1a1Jrt/+ mice. However, these two studies ignored the impact of antibodies on spine growth, fracture rates, and compressive mechanical properties. Here, we conducted a randomized controlled trial in oim/oim mice, an established model of human severe OI type III due to a mutation in Col1a2. Five-week-old female WT and oim/oim mice received either PBS or sclerostin antibody (Scl-Ab) for 9 weeks. Analyses included radiography, histomorphometry, pQCT, microcomputed tomography, and biomechanical testing. Though it did not modify vertebral axial growth, Scl-Ab treatment markedly reduced the fracture prevalence in the pelvis and caudal vertebrae, enhanced osteoblast activity (L4), increased cervico-sacral spine BMD, and improved the lumbosacral spine bone cross-sectional area. Scl-Ab did not impact vertebral height and body size but enhanced the cortical thickness and trabecular bone volume significantly in the two Scl-Ab groups. At lumbar vertebrae and tibial metaphysis, the absolute increase in cortical and trabecular bone mass was higher in Scl-Ab WT than in Scl-Ab oim/oim. The effects on trabecular bone mass were mainly due to changes in trabecular number at vertebrae and in trabecular thickness at metaphyses. Additionally, Scl-Ab did not restore a standard trabecular network, but improved bone compressive ultimate load with more robust effects at vertebrae than at metaphysis. Overall, Scl-Ab treatment may be beneficial for reducing vertebral fractures and spine deformities in patients with severe OI.

Cross-linked hyaluronic acid (HyAR) increases the local concentration of growth factors. We compared β-TCP osseointegration in old and young ewes with/without HyAR addition. A blind tunnel was drilled on the medial femoral condyle of each knee in nine young and nine old ewes and was filled with β-TCP, β-TCP + HyAR or left unfilled. Double labeling with calcein allowed histodynamic analysis. Ewes were sacrificed at 84 days and the knees were harvested. MicroCT provided histomorphometric parameters: trabecular bone volume, residual volume of biomaterial. Histodynamic parameters were: mineralization rate, mineralized surfaces, bone formation rate. A non-parametric ANOVA and post hoc test analyzed differences between subgroups. Osseointegration of β-TCP was similar in the aged/young grafted groups. Trabecular bone volume was significantly increased versus ungrafted animals (p

Nacre (mother of pearl) is a natural biomaterial used to prepare orthopedic devices. We have implanted screws and plates made with nacre in five sheeps. Bone were harvested after two months and embedded in poly(methyl methacrylate). Blocks were saws and the thick slabs were grinded, polished and surface stained. Sections were photographed at an ×1000 magnification. Giant cells were found in contact with nacre in eroded areas and true osteoclasts were found at distance in the neighboring bone in Howship lacunae. A texture analysis of the nuclei of giant cells and osteoclasts was done using the run-length method of the MaZda freeware. The size of the nuclei was reduced in osteoclast and their mean gray level appeared reduced. Texture analysis revealed that chromatin had a completely different pattern in giant cells when compared to osteoclasts. Giant cells had a fine repartition of the chromatin with large clear areas around prominent nucleoli. On the contrary, osteoclast nuclei had chromatin blocks evenly dispersed in the nuclei. This reflects the different origin of these cells expressing different functions.

International audience; Osteogenesis imperfecta type III (OI) is a serious genetic condition with poor bone quality and a high fracture rate in children. In a previous study, it was shown that a monoclonal antibody neutralizing sclerostin (Scl-Ab) increases strength and vertebral bone mass while reducing the number of axial fractures in oim/oim, a mouse model of OI type III. Here, we analyze the impact of Scl-Ab on long bones in OI mice. After 9 weeks of treatment, Scl-Ab significantly reduced long bone fractures (3.6 ± 0.3 versus 2.1 ± 0.8 per mouse, p

International audience; β-TCP is a resorbable bony biomaterial but its biodegradation mechanisms in vivo remains unclear. Osteoclast can resorb β-TCP but a role for macrophages has also been suggested by in vivo studies. However no in vitro study has clearly evidenced the action of macrophages in the resorption process. We prepared flat β-TCP tablets with a smooth surface to investigate the in vitro capability of murine (RAW 264.7) and human macrophage cells (PBMCs) to resorb the biomaterial. In parallel, these cells were differentiated into multinucleated osteoclasts with M-CSF and RANK-L. The action of these cells was evaluated by scanning electron microscopy and Raman microspectroscopy after a 21 day culture on the tablets. Human macrophages and osteoclasts derived from PBMCs appeared able to resorb β-TCP by forming resorption pits at the surface of the flat tablets. RAW macrophages were unable to resorb β-TCP but they exhibited this possibility when they have been differentiated into osteoclasts. These cells can engulf β-TCP grains in their cytoplasm as evidenced by light and TEM microscopy with production of carbonic anhydrase (revealed by the immunogold technique in TEM). The resorbed areas were characterized by severe degradation of the grains showing speckled and stick-like aspects indicating a chemical corrosion. The effect was maximal at the grain boundaries which have a slightly different chemical composition. Changes in the Raman spectrum were observed between the resorbed and un-resorbed β-TCP suggesting crystal modifications. In contrast, un-differentiated murine macrophages were not able to chemically attack β-TCP and no resorption pit was observed. RAW cell is not a representative model of the macrophage-biomaterial interactions that occur in human. This in vitro study evidences that both human osteoclasts and macrophages represent active cell populations capable to resorb β-TCP.

International audience; Sinus elevation is a common procedure to increase bone volume in the atrophic maxilla to allow placement of dental implants. Autogenous bone is the gold standard but is limited in quantity and causes morbidity at the donor site. β-TCP is a synthetic biomaterial commonly used in that purpose. It appears to induce a poor inflammatory response. This study aimed to evaluate the degree of edema of the sinus mucosa after sinus lift surgery according to the type of biomaterial. Forty sinuses (20 patients) were included retrospectively and divided into 2 groups according to the biomaterial that was used: synthetic biomaterial (BTCP group), natural bone (BONE group). A control group (CTRL group) was constituted by the non-grafted maxillary sinuses. Twelve measurements per sinus were realized on pre- and post-operative computed tomography and averaged to provide the sinus membrane thickness value (SM.Th). SM.Th was thicker post-operatively in the BTCP and BONE groups in comparison with the CTRL group and in comparison with pre-operative measurements. No difference was found post operatively between the BTCP and BONE groups. We found that a synthetic biomaterial (β-TCP) induced the same degree of edema, and thus of inflammation, as natural bone. It constitutes therefore an interesting alternative to autogenous bone for maxillary sinus lifts.