Daniel Chappard

Publications of Daniel Chappard

• Thiazolidinediones induced osteocyte apoptosis by a GPR40-dependent mechanism.

  Authors: Aleksandra Mieczkowska, Michel F Basle, Daniel Chappard, Guillaume Mabilleau

  The Journal of biological chemistry. 05/2012;

  Thiazolidinediones represent an interesting treatment in type 2 diabetes mellitus. However adverse effects such as heart problems and bone fractures have already been reported. Previously, weThiazolidinediones represent an interesting treatment in type 2 diabetes mellitus. However adverse effects such as heart problems and bone fractures have already been reported. Previously, we reported that pioglitazone and rosiglitazone induce osteocyte apoptosis and sclerostin up-regulation; however, the molecular mechanisms leading to such effects are unknown. In this study, we found that thiazolidinediones rapidly activated Erk 1/2 and p38. These activations were mediated through ras proteins and GPR40, a receptor expressed at the surface of osteocytes. Activation of this pathway conducted only to osteocyte apoptosis but not sclerostin up-regulation. On the other hand, thiazolidinediones were capable of activating PPAR-γ, and activation of this signalling pathway led to sclerostin up-regulation but not osteocyte apoptosis. This study evidenced two distinct signalling pathways activated in osteocytes in response to thiazolidinediones that could participate to the observed increase in fractures in thiazolidinedione-treated patients.

• Glucocorticoids reduce alveolar and trabecular bone in mice.

  Authors: Béatrice Bouvard, Yves Gallois, Erick Legrand, Maurice Audran, Daniel Chappard

  Joint, bone, spine : revue du rhumatisme. 02/2012;

  Glucocorticoid (GC) treatment is the main cause of secondary osteoporosis. There are some controversies about the relationships between alveolar bone loss and bone loss at the appendicular and axialGlucocorticoid (GC) treatment is the main cause of secondary osteoporosis. There are some controversies about the relationships between alveolar bone loss and bone loss at the appendicular and axial skeleton. OBJECTIVE: To assess, in parallel, the effects of GCs on alveolar bone and on the tibia in a mice model. METHODS: Five-month-old male Swiss-Webster mice were randomized into two groups. Pellets releasing 5mg/kg/day of prednisolone or control pellets were subcutaneously implanted for 28days. After euthanasia, the right tibia and the right hemimandible of each mouse were analyzed by histomorphometry and microcomputed tomography. Alveolar bone consists of a thin slab between the incisor and the molar roots connected with the alveolar processes. A 2D-frontal section was done through the pulp chamber of the first molar and was used to measure the thickness of the alveolar bone slab. A 2D-sagittal section was done through the pulp chamber of the three molars and was used to measure bone volume in the alveolar processes. RESULTS: At day 28, thickness and bone volume of alveolar bone were significantly decreased in the GC group (P<0.05). At the tibia, GCs decreased bone formation with a reduced mineral apposition rate and bone formation rate and a significant decrease in BV/TV and Tb.Th (P<0.05). CONCLUSION: Although the amount of alveolar bone is very low in the mouse, this study shows that GCs can induce an alveolar bone loss in long-term treated animals.

• Is transiliac bone biopsy a painful procedure?

  Authors: Maurice Audran, Emilie Maury, Béatrice Bouvard, Erick Legrand, Michel Felix Baslé, Daniel Chappard

  Clinical nephrology. 02/2012; 77(2):97-104.

  Despite an increased availability of non-invasive procedures to assess bone mass, histological examination of undecalcified transiliac bone biopsies remains a very valuable tool in the diagnosis ofDespite an increased availability of non-invasive procedures to assess bone mass, histological examination of undecalcified transiliac bone biopsies remains a very valuable tool in the diagnosis of metabolic or malignant bone disorders. Nonetheless, clinicians are sometimes reluctant to perform this "invasive" examination, arguing that it might be a painful procedure. The aim of our study was to evaluate pain and anxiety described by patients in the months following the biopsy and to characterize potential early or late side effects. A single interviewer conducted a phone survey (19 items questionnaire) in 117 patients in whom a bone biopsy had been performed by two experienced physicians, with the same material and similar anesthetic and technical procedure. The topics covered pain during or after the biopsy, anxiety, comparison of other potentially painful procedures, early or late side effects as well as global evaluation by the patients. Bone biopsy was judged as non-painful by almost 70% of patients; some discomfort was present in 25% in the following days. The procedure was described as similar as or less painful than bone marrow aspiration, venipuncture or tooth extraction. About 90% of the patients estimated that it was a quite bearable diagnostic procedure. Side effects were not serious. About 7% remembered a vasovagal episode, 47% of local bruising in the following days. There was no report of hematoma or infection. In experienced hands and adapted trephine, transiliac bone biopsy is a safe procedure that brings invaluable information in bone disorders.

• Depth and volume of resorption induced by osteoclasts generated in the presence of RANKL, TNF-alpha/IL-1 or LIGHT.

  Authors: Guillaume Mabilleau, Florence Pascaretti-Grizon, Michel Félix Baslé, Daniel Chappard

  Cytokine. 12/2011; 57(2):294-9.

  Rheumatoid arthritis (RA) is associated with pathological bone destruction mediated by osteoclasts. Although RANKL has been reported as a crucial factor for osteoclastogenesis, several other factorsRheumatoid arthritis (RA) is associated with pathological bone destruction mediated by osteoclasts. Although RANKL has been reported as a crucial factor for osteoclastogenesis, several other factors increased in RA support osteoclast formation and resorption in the absence of RANKL such as TNF-alpha and LIGHT. To date, in vitro bone resorption experiments are reported as the mean area of bone resorption per cortical or dentine slices and do not provide any information about depth and volume of resorption. The aims of this study were to assess these parameters by light microscopy and vertical scanning profilometry (VSP). Peripheral blood mononuclear cells were used as a source of osteoclast precursors and were cultured for up to 21 days in the presence of RANKL, TNF-alpha/IL-1 or LIGHT. Mean area, depth and volume of resorption were assessed by light microscopy and vertical scanning profilometry. As expected, RANKL induced large resorption pits (10,876 ± 2190μm(2)) whereas TNF-alpha/IL-1 and LIGHT generated smaller pits (respectively 1328 ± 210 and 1267 ± 173μm(2)) with no noticeable differences between these two cytokines. Depth and volume of resorption measured by VSP showed that RANKL promoted deep resorption pits resulting in large volume of resorption. Interestingly, although mean area of resorption was similar between TNF-alpha/IL-1 and LIGHT, the depth and volume of resorption of these lacunae were significantly increased by 2-fold with TNF-alpha/IL-1. These results provide evidence that although LIGHT appeared elevated in the synovial fluid of RA patients, its role in bone resorption is less than TNF-alpha/IL-1 or RANKL.

• Evaluation of the bone status in high-level cyclists.

  Authors: Gérard Guillaume, Daniel Chappard, Maurice Audran

  Journal of clinical densitometry : the official journal of the International Society for Clinical Densitometry. 11/2011; 15(1):103-7.

  The purpose of this study was to evaluate the bone status in highly trained professional cyclists subjected to regular training and tough competitions. Bone mineral density (BMD) was measured atThe purpose of this study was to evaluate the bone status in highly trained professional cyclists subjected to regular training and tough competitions. Bone mineral density (BMD) was measured at different regions of interest by dual-energy X-ray absorptiometry, and main biological parameters related to bone metabolism were obtained in 29 cyclists. Lumbar BMD was 0.94 ± 0.01g/cm(2) (Z-score=-1.28 ± 0.07), and 1 cyclist out of 4 had an abnormally low value (Z-score <-2). The mean Z-score at the total femoral site was -1.22 ± 0.21, and 45% of athletes had an Z-score of <-2. All femoral neck BMD values were within normal boundaries. The lowest BMD Z-score was measured at the midradius or 1/3 proximal site with a mean Z-score of -1.77 ± 0.78, but only 3 cyclists (15%) had Z-scores <-2. Biochemical parameters of bone formation (serum osteocalcin and alkaline phosphatase) were normal. Three cyclists had low 25-hydroxyvitamin D levels. Blood testosterone and thyroid stimulating hormone were in the normal range. Insulin-like growth factor 1 levels were in the normal range; however, a significant inverse correlation was found with lumbar BMD (r=0.495; p=0.003). We confirm that cycling has no positive effect on BMD, BMD being often lower than in normal controls at the lumbar site; femoral BMD is less concerned. The absence of beneficial changes at the spine can be explained by biomechanical conditions related to the cyclists' position, reducing loading strains. It is necessary to pay greater attention to the bone status of high-level athletes to prevent an increased risk of fractures.

• Relationships between bone mass and micro-architecture at the mandible and iliac bone in edentulous subjects: a dual X-ray absorptiometry, computerised tomography and microcomputed tomography study.

  Authors: François Bodic, Yves Amouriq, Marie Gayet-Delacroix, Yves Maugars, Luc Hamel, Michel F Baslé, Daniel Chappard

  Gerodontology. 06/2011;

  Gerodontology 2011; 00; doi: 10.1111/j.1741-2358.2011.00527.x Relationships between bone mass and micro-architecture at the mandible and iliac bone in edentulous subjects: a dual X-rayGerodontology 2011; 00; doi: 10.1111/j.1741-2358.2011.00527.x Relationships between bone mass and micro-architecture at the mandible and iliac bone in edentulous subjects: a dual X-ray absorptiometry, computerised tomography and microcomputed tomography study Objectives:  To compare bone volume, bone mineral density, cortical thickness and bone micro-architecture in a series of paired mandibular and iliac bone samples analysed by various imagery techniques to see whether relationships exist between the various techniques and between mandibular and iliac bone. Materials and methods:  Bone samples from the mandible and ilium were harvested in 20 cadavers and analysed by dual energy X-ray absorptiometry (DXA), computerised tomography (CT) on a conventional hospital machine and microCT. Results:  Significant correlations were found between Hounsfield density obtained by CT, and bone mass determined by microCT but not with DXA values. Cortical thickness measurements were well correlated between CT and microCT. No relationships were found between mandibular and iliac bone, when considering mineral density, cortical thickness, bone volume or micro-architecture. Conclusion:  In clinical practice, CT remains the most appropriate routine means for bone qualitative and quantitative evaluation at the mandible. In this ex vivo study, these results confirm that mandibular bone status does not reflect the axial skeletal one and assist in the placement of implants with dental prostheses in old or osteoporotic patients.

• Effects of risedronate in Runx2 overexpressing mice, an animal model for evaluation of treatment effects on bone quality and fractures.

  Authors: Valerie Geoffroy, Eleftherios P Paschalis, Helene Libouban, Stephane Blouin, Agnes Ostertag, Daniel Chappard, Magali Cros, Roger Phipps, Marie-Christine de Vernejoul

  Calcified tissue international. 04/2011; 88(6):464-75.

  Young mice overexpressing Runx2 specifically in cells of the osteoblastic lineage failed to gain bone mass and exhibited a dramatic increase in bone resorption, leading to severe osteopenia andYoung mice overexpressing Runx2 specifically in cells of the osteoblastic lineage failed to gain bone mass and exhibited a dramatic increase in bone resorption, leading to severe osteopenia and spontaneous vertebral fractures. The objective of the current study was to determine whether treatment with a bisphosphonate (risedronate, Ris), which reduces fractures in postmenopausal as well as in juvenile osteoporosis, was able to improve bone quality and reduce vertebral fractures in mice overexpressing Runx2. Four-week-old female Runx2 mice received Ris at 2 and 10 μg/kg subcutaneously twice a week for 12 weeks. Runx2 and wild-type mice received vehicle (Veh) as control. We measured the number of new fractures by X-ray and bone mineral density (BMD) by DEXA. We evaluated bone quality by histomorphometry, micro-CT, and Fourier transform infrared imaging (FTIRI). Ris at 20 μg/kg weekly significantly reduced the average number of new vertebral fractures compared to controls. This was accompanied by significantly increased BMD, increased trabecular bone volume, and reduced bone remodeling (seen in indices of bone resorption and formation) in the vertebrae and femoral metaphysis compared to Runx2 Veh. At the femur, Ris also increased cortical thickness. Changes in collagen cross-linking seen on FTIRI confirmed that Runx2 mice have accelerated bone turnover and showed that Ris affects the collagen cross-link ratio at both forming and resorbing sites. In conclusion, young mice overexpressing Runx2 have high bone turnover-induced osteopenia and spontaneous fractures. Ris at 20 μg/kg weekly induced an increase in bone mass, changes in bone microarchitecture, and decreased vertebral fractures.

• Role of the A20-TRAF6 axis in lipopolysaccharide-mediated osteoclastogenesis.

  Authors: Guillaume Mabilleau, Daniel Chappard, Afsie Sabokbar

  The Journal of biological chemistry. 02/2011; 286(5):3242-9.

  Bacterial lipopolysaccharide (LPS) has long been suggested as a potent inducer of bone loss in vivo despite controversial effects on osteoclast precursors. Recently, the role of the deubiquitinatingBacterial lipopolysaccharide (LPS) has long been suggested as a potent inducer of bone loss in vivo despite controversial effects on osteoclast precursors. Recently, the role of the deubiquitinating protease A20 in regulating the LPS response in various organs was reported. In the present study, we investigated whether A20 is expressed in osteoclast cultures in response to RANKL or LPS and whether this protein plays a role in osteoclast formation and activation. Human peripheral blood mononuclear cells were cultured in the presence of M-CSF ± RANKL ± LPS. Although LPS induced the formation of multinucleated TRAP-positive cells expressing OSCAR, cathepsin K, and the calcitonin receptor, these cells were not capable of lacunar resorption. Release of TNF-α was noted in LPS-treated cultures, and the addition of a neutralizing anti-TNF-α antibody abrogated osteoclast formation in these cultures. A20 appeared to be a late-expressed gene in LPS-treated cultures and was associated with TRAF6 degradation and NF-κB inhibition. Silencing of A20 restored TRAF6 expression and NF-κB activation and resulted in increased bone resorption in LPS-treated cultures. A20 appeared important in the control of bone resorption and could represent a therapeutic target to treat patients with bone resorption associated with inflammatory diseases.

• Extraskeletal effects of vitamin D: facts, uncertainties, and controversies.

  Authors: Béatrice Bouvard, Cédric Annweiler, Agnès Sallé, Olivier Beauchet, Daniel Chappard, Maurice Audran, Erick Legrand

  Joint, bone, spine : revue du rhumatisme. 01/2011; 78(1):10-6.

  Vitamin D was long viewed as a hormone acting chiefly to regulate calcium-phosphate metabolism and bone mineralization. Over the last decade, however, basic science and clinical researchers haveVitamin D was long viewed as a hormone acting chiefly to regulate calcium-phosphate metabolism and bone mineralization. Over the last decade, however, basic science and clinical researchers have produced a bewildering amount of information on the extraskeletal effects of vitamin D. This article is a review of the clinical and biological actions of vitamin D including effects on the immune system, auto-immune diseases, infections, cancer, metabolic syndrome, fall risk, cognitive function, and muscle function.

• Cellular and molecular effects of thiazolidinediones on bone cells: a review.

  Authors: Guillaume Mabilleau, Daniel Chappard, Michel F Baslé

  International journal of biochemistry and molecular biology. 01/2011; 2(3):240-6.

  Thiazolidinediones represent a class of molecules used in the treatment of type 2 diabetes mellitus. Despite interesting effects in lowering blood glucose and HbA1c levels durably, an augmentation ofThiazolidinediones represent a class of molecules used in the treatment of type 2 diabetes mellitus. Despite interesting effects in lowering blood glucose and HbA1c levels durably, an augmentation of the fracture risk in women has emerged in the past years. This review is providing the readers with information about the cellular and molecular mechanisms involved in bone and bone cells in response to these drugs.

• In vitro assessment of osteoblast and macrophage mobility in presence of β-TCP particles by videomicroscopy.

  Authors: Johanne Beuvelot, Florence Pascaretti-Grizon, Robert Filmon, Marie-Fraçoise Moreau, Michel F Baslé, Daniel Chappard

  Journal of biomedical materials research. Part A. 01/2011; 96(1):108-15.

  β-TCP is widely used to repair bone defects due to its good biocompatibility, macroporosity (favoring bone ingrowth) and bioresorbability. However, cell interactions with the biomaterial at the firstβ-TCP is widely used to repair bone defects due to its good biocompatibility, macroporosity (favoring bone ingrowth) and bioresorbability. However, cell interactions with the biomaterial at the first times of implantation remain largely unknown. We have observed cell behaviors in direct contact with β-TCP particles using long-term culture under videomicroscopy. Osteoblastlike cells (SaOs-2) and macrophages (J774.2 and mouse peritoneal macrophages) were cultured in the presence of β-TCP particles. For each experiment, images from 20 independent fields were acquired and stored every 15 min during 8 days. At the end of the culture, they were combined to generate time lapse videos; coverslips were fixed and observed by scanning electron microscopy (SEM). SaOs-2 proliferation was determined by counting cells on six different and independent fields at days 1, 3, and 6. Videos showed the capacity of cells to displace the particles. Dynamic follow-up showed active proliferation of SaOs-2 occurring in the direction of the particles. J774.2 and peritoneal macrophages did not proliferate but came in direct contact with the particles and actively eroded them. SEM showed that cells were stretched and fixed onto the surface and seemed to climb from the coverslip to the particles. The long-term culture under videomicroscopy allowed a better understanding of the colonization process of β-TCP particles by osteoblastlike cells and macrophages. Data obtained from long-term videomicroscopy are in agreement with in vivo observations confirming the interest of β-TCP to promote osteogenesis.

• Three-dimensional characterization of the vascular bed in bone metastasis of the rat by microcomputed tomography (MicroCT).

  Authors: Hervé Nyangoga, Philippe Mercier, Hélène Libouban, Michel Félix Baslé, Daniel Chappard

  PloS one. 01/2011; 6(3):e17336.

  Angiogenesis contributes to proliferation and metastatic dissemination of cancer cells. Anatomy of blood vessels in tumors has been characterized with 2D techniques (histology or angiography). TheyAngiogenesis contributes to proliferation and metastatic dissemination of cancer cells. Anatomy of blood vessels in tumors has been characterized with 2D techniques (histology or angiography). They are not fully representative of the trajectories of vessels throughout the tissues and are not adapted to analyze changes occurring inside the bone marrow cavities. We have characterized the vasculature of bone metastases in 3D at different times of evolution of the disease. Metastases were induced in the femur of Wistar rats by a local injection of Walker 256/B cells. Microfil®, (a silicone-based polymer) was injected at euthanasia in the aorta 12, 19 and 26 days after injection of tumor cells. Undecalcified bones (containing the radio opaque vascular casts) were analyzed by microCT, and a first 3D model was reconstructed. Bones were then decalcified and reanalyzed by microCT; a second model (comprising only the vessels) was obtained and overimposed on the former, thus providing a clear visualization of vessel trajectories in the invaded metaphysic allowing quantitative evaluation of the vascular volume and vessel diameter. Histological analysis of the marrow was possible on the decalcified specimens. Walker 256/B cells induced a marked osteolysis with cortical perforations. The metaphysis of invaded bones became progressively hypervascular. New vessels replaced the major central medullar artery coming from the diaphyseal shaft. They sprouted from the periosteum and extended into the metastatic area. The newly formed vessels were irregular in diameter, tortuous with a disorganized architecture. A quantitative analysis of vascular volume indicated that neoangiogenesis increased with the development of the tumor with the appearance of vessels with a larger diameter. This new method evidenced the tumor angiogenesis in 3D at different development times of the metastasis growth. Bone and the vascular bed can be identified by a double reconstruction and allowed a quantitative evaluation of angiogenesis upon time.

• [Bone microarchitecture].

  Authors: Daniel Chappard

  Bulletin de l'Académie nationale de médecine. 11/2010; 194(8):1469-80; discussion 1480-1.

  Bone mass is determined throughout life by modeling and remodeling mechanisms. Trabecular bone consists of a network of trabeculae (plates and rods) whose distribution is highly anisotropic:Bone mass is determined throughout life by modeling and remodeling mechanisms. Trabecular bone consists of a network of trabeculae (plates and rods) whose distribution is highly anisotropic: trabeculae are disposed parallel to the resultant of stress lines (Wolff's law). The trabecular microarchitecture appears to be conditioned by mechanical stresses exerted on skeletal bones. Few clinically validated methods are available to assess and monitor the bone microarchitecture in bone diseases. The best-developed is bone histomorphometry, with the use of new algorithms to appreciate various 2D characteristics of the trabeculae, such as thickness and connectivity. Several studies have shown that several independent techniques should be used to study microarchitecture parameters. X-ray microtomography (micro CT), microMRI and synchrotron devices can give non destructive 3D measurements of the trabecular microarchitecture of bone specimens. This review describes advances in our knowledge of bone microarchitecture, its role in bone diseases such as osteoporosis, and methods of 2D and 3D histological evaluation.

• In vitro calcification of chemically functionalized carbon nanotubes.

  Authors: Johanne Beuvelot, Céline Bergeret, Romain Mallet, Vincent Fernandez, Jack Cousseau, Michel Félix Baslé, Daniel Chappard

  Acta biomaterialia. 10/2010; 6(10):4110-7.

  Bone is composed of two phases. The organic phase is made of collagen fibrils assembled in broad fibers acting as a template for mineralization. The mineral phase comprises hydroxyapatite (HAP)Bone is composed of two phases. The organic phase is made of collagen fibrils assembled in broad fibers acting as a template for mineralization. The mineral phase comprises hydroxyapatite (HAP) crystals grown between and inside the collagen fibers. We have developed a biomimetic material using functionalized carbon nanotubes as scaffold to initiate in vitro mineralization. Biomimetic formation of HAP was performed on single-walled carbon nanotubes (SWCNTs) which have been grafted with carboxylic groups. Two types of nanotubes, HiPco(R) and Carbon Solutions(R), were oxidized via various acidic processes, leading to five different groups of carboxylated nanotubes, fully characterized by physical methods (thermogravimetric analysis, attenuated total reflectance infrared spectroscopy and X-ray photoelectron spectroscopy). All samples were dispersed in ultra-pure water and incubated for 2weeks in a synthetic body fluid, in order to induce the calcification of the SWCNTs. Scanning electron microscopy (SEM) and energy-dispersive X-ray analysis studies showed that Ca(2+) and PO(4)(3-) ions were deposited as round-shaped nodules (calcospherites) on the carboxylated SWCNTs. Fourier transform infrared and Raman spectroscopic studies confirmed the HAP formation, and image analysis made on SEM pictures showed that calcospherites and carboxylated SWCNTs were packed together. The size of calcospherites thus obtained in vitro from the HiPco(R) series was close to that issued from calcospherites observed in vivo. Functionalization of SWCNTs with carboxylic groups confers the capacity to induce calcification similar to woven bone.

• A non-steroidal anti-inflammatory drug (ketoprofen) does not delay beta-TCP bone graft healing.

  Authors: Herve Nyangoga, Eric Aguado, Eric Goyenvalle, Michel Felix Baslé, Daniel Chappard

  Acta biomaterialia. 08/2010; 6(8):3310-7.

  Beta-tricalcium phosphate (beta-TCP) is a suitable biomaterial in oral and maxillofacial surgery since it can induce a rapid proliferation of woven bone. Granules, prepared by the polyurethane foamBeta-tricalcium phosphate (beta-TCP) is a suitable biomaterial in oral and maxillofacial surgery since it can induce a rapid proliferation of woven bone. Granules, prepared by the polyurethane foam method, were implanted in critical size defects performed in the femoral condyles of New Zealand rabbits. Animals were studied after 8 and 28 days. Ketoprofen (a non-steroidal anti-inflammatory drug (NSAID)) was given for 8 and 28 days to evaluate its effects on the healing of the graft. Before euthanasia, the rabbits received an intravenous injection of fluorescent microbeads. Bones were analyzed by microcomputed tomography. beta-TCP granules induced metaplastic bone trabeculae as early as 8 days post-surgery. At 28 days, the amount of bone was increased and the biomaterial volume decreased due to simultaneous macrophagic resorption. The amount of macrophages labeled with microbeads was less in the grafted area than in the vicinal intact marrow spaces. Ketoprofen had no effect on the amount of bone formed and on the number of labeled macrophages. The influence of small doses of NSAID, given in a short duration period, did not present deleterious effects on bone graft healing.

• Computed microtomography of bone specimens for rapid analysis of bone changes associated with malignancy.

  Authors: Daniel Chappard, Hélène Libouban, Erick Legrand, Norbert Ifrah, Charles Masson, Michel Felix Baslé, Maurice Audran

  Anatomical record (Hoboken, N.J. : 2007). 07/2010; 293(7):1125-33.

  Breast and prostate cancers are specially metastasizing to bone. Metastases from breast cancer usually exhibit a mixed osteolytic/osteosclerotic aspect, with osteolysis predominating. OsteosclerosisBreast and prostate cancers are specially metastasizing to bone. Metastases from breast cancer usually exhibit a mixed osteolytic/osteosclerotic aspect, with osteolysis predominating. Osteosclerosis is a common finding in prostatic cancer although osteolysis occurs within the sclerotic lesions. B-cell malignancies (lymphoma, myeloma) are also associated with marked osteolysis. Histopathological examination of bone biopsies was used for the diagnosis of malignancies and, prior to embedding, microcomputed tomography (microCT) was done on the bone specimens. Patients (247) who presented either a bone metastasis, an overt myeloma, a lymphoma or a monoclonal gammopathy of undetermined significance were studied. All patients had a bone biopsy studied by 2D histomorphometry for the histopathology. During the fixation time, the bone cores were analyzed by microCT. On the 3D reconstructed models provided by microCT, signs of osteolysis/osteosclerosis were searched: excess of bone resorption, focal disorganization of microarchitecture, bone metaplasia, osteosclerosis. A strong agreement was obtained between histomorphometry and microCT results using Cohen's kappa test (kappa = 0.713). MicroCT identified excess bone resorption on trabecular surfaces when eroded surfaces were >10.5% by histomorphometry. MicroCT failed to identify some patients with smoldering myeloma or some lymphomas with microresorption. MicroCT data are obtained within 4 hr and suggest the malignant invasion of bone marrow when excess of bone resorption/formation is obtained. MicroCT can be used in the immediate postbiopsy period making possible a fast identification of malignancy. However these signs are not specific and must be confirmed by histopathological analysis.

• Sinus lift augmentation and beta-TCP: a microCT and histologic analysis on human bone biopsies.

  Authors: Daniel Chappard, Bernard Guillaume, Romain Mallet, Florence Pascaretti-Grizon, Michel F Baslé, Hélène Libouban

  Micron (Oxford, England : 1993). 06/2010; 41(4):321-6.

  Sinus lift elevation is an interesting method to restore bone mass at the maxilla in edentulated patients. We have investigated the histological effects of beta tricalcium phosphate (beta-TCP)Sinus lift elevation is an interesting method to restore bone mass at the maxilla in edentulated patients. We have investigated the histological effects of beta tricalcium phosphate (beta-TCP) combined with autograft bone for sinus lift elevation. A series of 14 patients who were candidate for dental implantation were grafted with beta-TCP granules and morcellized autograft bone harvested at the chin. beta-TCP was characterized by scanning electron microscopy and optical profilometry. Before implant placement, a small bone biopsy (2mm in diameter) was done. The amount of residual material and newly formed bone were determined by microcomputed tomography. Histological analysis was done on undecalcified sections stained by Goldner's trichrome and osteoclast identification (TRAcP). beta-TCP served as a template for bone apposition by osteoblasts onto the granules' surface. The material was simultaneously resorbed by TRAcP positive osteoclasts and macrophages. Fragments of the material remained buried in bone trabeculae as long as 12 months post-graft but the formed bone onto the granules surface had a lamellar texture. beta-TCP combined with autograft bone appears a suitable biomaterial for sinus lift augmentation before the placement of bone implants. The material favors the apposition of lamellar bone by osteoblasts and is simultaneous resorbed by two types of cells.

• Monoclonal gammopathy of undetermined significance, multiple myeloma, and osteoporosis.

  Authors: Béatrice Bouvard, Mathieu Royer, Daniel Chappard, Maurice Audran, Emmanuel Hoppé, Erick Legrand

  Joint, bone, spine : revue du rhumatisme. 03/2010; 77(2):120-4.

  The finding of monoclonal gammopathy of undetermined significance (MGUS) is not infrequent during an evaluation for osteoporosis or a fracture. In most cases, the diagnosis is MGUS, whose prevalenceThe finding of monoclonal gammopathy of undetermined significance (MGUS) is not infrequent during an evaluation for osteoporosis or a fracture. In most cases, the diagnosis is MGUS, whose prevalence increases with age. Although the impact of MGUS on bone mineral density, bone remodeling, and the fracture risk remains unclear, this asymptomatic hematological disorder may constitute a risk factor for osteoporosis. Furthermore, each year, 1% of patients with MGUS progress to multiple myeloma, a disease whose pathophysiology and association with bone loss and pathological fractures are increasingly well understood. Osteoporotic fractures, although probably common in myeloma patients, are less likely to be recognized. Here, we discuss the pathophysiology of myeloma and MGUS and their impact in terms of bone mineral density, osteoporotic fractures, and bone turnover markers.

• The cathepsin K inhibitor AAE581 induces morphological changes in osteoclasts of treated patients.

  Authors: Daniel Chappard, Hélène Libouban, Linda Mindeholm, Michel F Baslé, Erick Legrand, Maurice Audran

  Microscopy research and technique. 12/2009;

  Inhibitors of Cathepsin K (Cat-K) are recognized as an interesting way to inhibit osteoclast (OC) activity. OCs from patients treated with the anticathepsin-K inhibitor AAE581 (balicatib) were foundInhibitors of Cathepsin K (Cat-K) are recognized as an interesting way to inhibit osteoclast (OC) activity. OCs from patients treated with the anticathepsin-K inhibitor AAE581 (balicatib) were found enlarged. They contained numerous vacuoles filled with tartrate resistant acid phosphatase (TRAcP), an intracellular enzyme that terminates the degradation of collagen internalized in OC transcytotic vesicles. In a phase 2 clinical study, 675 patients with postmenopausal osteoporosis received the Cat-K inhibitor AAE581 at 0, 5, 10, 25, or 50 mg/D during 1 year. Eleven patients had a transiliac bone biopsy, studied undecalcified. Histoenzymatic detection of TRAcP was used to identify and count OC number. The histomorphometrist was not aware of the randomization of patients at the time of analysis. OC were unstained in one patient because of a failure in the fixation protocol, but easily observable in the 10 remaining patients. Whatever the received dose, treated patients exhibited a characteristic aspect of the OC cytoplasm which appeared filled of deeply-stained brown vacuoles, making cells looking like bunches of grape. These round vacuoles, evidenced on TRAcP-stained sections, were due to the accumulation of intracytoplasmic TRAcP. This led to a moderate enlargement of the OC size when compared to a series of control osteoporotic patients. AAE581 did not induce OC apoptosis at any dosage but it modified OC morphology. Cat-K inhibition (inhibiting the extracellular collagen breakdown) is associated with a compensatory accumulation of intracellular TRAcP that could not be used to complete protein degradation. TRAcP is also known to be degraded by Cat-K. Microsc. Res. Tech., 2010. (c) 2009 Wiley-Liss, Inc.

• A single pretreatment by zoledronic acid converts metastases from osteolytic to osteoblastic in the rat.

  Authors: Hervé Nyangoga, Stéphane Blouin, Hélène Libouban, Michel Félix Baslé, Daniel Chappard

  Microscopy research and technique. 12/2009;

  Bone metastases are severe complications of cancers associated with increased morbidity, pain, risk fracture, and reduced life span for patients. Bisphosphonates emerged as a relief treatment in boneBone metastases are severe complications of cancers associated with increased morbidity, pain, risk fracture, and reduced life span for patients. Bisphosphonates emerged as a relief treatment in bone metastases. A single dose of zoledronic acid (78 mug/kg) was injected into six Copenhagen rats 4 days before receiving an intraosseous inoculation of metastatic anaplastic tumor of lymph node and lung cell (MLL) prostate cancer cells. Rat femurs were analyzed for changes by microCT and histomorphometry; trabecular volume, trabecular characteristics, osteoid parameters, osteoblastic surfaces, and osteoclast number were measured. Values were compared to a group of SHAM animals, a group of SHAM animals having received zoledronic acid and animals inoculated with MLL cells. All rats were euthanized after 1 month. MLL cells induced osteolysis in the metaphysis with extension of the tumor to soft tissues through cortical perforations. Zoledronic acid induced a marked osteosclerosis in the primary spongiosa in both SHAM and rats inoculated with MLL. Osteosclerosis was obtained in the secondary spongiosa of MLL rats. The bisphosphonate preserved cortical integrity in all animals, and no extension to soft tissues was observed in most animals. The number of osteoclasts was elevated, indicating that there was no apoptosis of osteoclasts but they became inactive. Osteosclerosis was associated with increased osteoblastic surfaces. A single zoledronic acid injection turned osteolytic metastases into osteosclerotic and preserved cortical integrity. Microsc. Res. Tech., 2010. (c) 2009 Wiley-Liss, Inc.