IL-33 is expressed in human osteoblasts, but has no direct effect on bone remodeling.

The aim of the present study was to investigate the potential role of the recently discovered IL-1 family member IL-33 in bone remodeling. Our results indicate that IL-33 mRNA is expressed in osteocytes in non-inflammatory human bone. Moreover, IL-33 levels are increased by TNF-α and IL-1ß in human bone marrow stromal cells, osteoblasts and adipocytes obtained from three healthy donors. Experiments with the inhibitor GW-9662 suggested that expression of IL-33, in contrast to that of IL-1ß, is not repressed by PPARγ likely explaining why IL-33, but not IL-1ß, is expressed in adipocytes. The IL-33 receptor ST2L is not constitutively expressed in human bone marrow stromal cells, osteoblasts or CD14-positive monocytes, and IL-33 has no effect on these cells. In addition, although ST2L mRNA is induced by TNF-α and IL-1ß in bone marrow stromal cells, IL-33 has the same effects as TNF-α and IL-1ß, and, therefore, the biological activity of IL-33 may be redundant in this system. In agreement with this hypothesis, MC3T3-E1 osteoblast-like cells constitutively express ST2L mRNA, and IL-33 and TNF-α/IL-1ß similarly decrease osteocalcin RNA levels in these cells. In conclusion, our results suggest that IL-33 has no direct effects on normal bone remodeling.

Microradiographic aspect on iliac bone tissue in postmenopausal women with and without vertebral crush fractures.

Bone mass, microstructure and microconnectivity of cancellous bone tissue, microporosity, and microdensity of cortical bone have been evaluated with an image analysing computer on microradiographs in 34 iliac crest bone biopsies from two individual, paired age-matched groups of postmenopausal women. Group 1 comprised 17 cases without vertebral crush fractures (mean age 66 years, range 45-80 years) and time since menopause 20.5 +/- 11.5 years. The second group were 17 cases with vertebral crush fractures (mean age 67 years, range 48-86 years) and time since menopause 16.5 +/- 7.6 years. No differences were observed either in bone mass or in Parfitt's indices of the microarchitecture of their cancellous bone tissue. However, both groups 1 and 2 were different by the number of the structure elements per 30 mm2 of tissue volume and the trabecular bone pattern factor (TBPF) being higher in group 1 than in group 2 (p < 0.005)--showing a better connectivity of bone network in group 2 with crush vertebral fractures than in group 1. The number of terminus to terminus struts of the trabecular bone pattern is increased in group 1 (p < 0.001). On 7-microns stained sections the active cancellous eroded perimeter (Cn.Oc.Pm/B.Pm) is greater in group 1 than in group 2 (p < 0.02). Therefore, in these two populations with a similar degree of osteopenia, a decrease of trabecular connectivity does not provide an explanation of bone fragility and the occurrence of vertebral crushed fractures.

Osteomesopyknosis: report of a new case with bone histology.

A new case of osteomesopyknosis, a rare autosomal dominant axial osteosclerosis is reported, with 4 affected members of the same family. Biochemical investigations, bone mineral content (BMC) measurement, 99mTc HMDP bone scan and microscopy of iliac crest bone and femoral head have been performed on 1 subject. A marked increase of BMC was found, without abnormality of biochemical data. Microscopy of bone showed an increase of trabecular thickness, and a low rate of bone turnover. No abnormality of mineralization was found on microradiographs.

Volume effect on biological properties of a calcium phosphate hydraulic cement: experimental study in sheep.

Injectable calcium phosphate hydraulic cements (CPHC) are a new family of bone substitutes within the class of bone reconstruction biomaterials. In this work, CPHC were tested in two consistencies (preset blocks or liquid paste) in an experimental model of cancellous bone defect in sheep. The defects were eight times larger than those investigated previously in rabbits. Three delays (12, 24, and 52 weeks) were used. Before death, a double label of oxytetracycline and alizarine was made intravenously. The distribution of implants was randomized, histomorphometric evaluation was performed and compared with micrographic observation, and optical microscopy of stained sections was performed either under visible, ultraviolet, or polarized light. The results were compared with spontaneous healing of empty defects and with a control group of normal cancellous bone from sheeps of the same age. No significant difference has been observed between premolded and injected implants. In the sheep model, the degradation and new bone formation rates are three times slower, compared with those observed previously in rabbits. New bone formation increased from 5.9% (12 weeks) up to 11.0% (24 weeks) in the empty defect group. In the cement groups, 28.3% new bone was obtained at 12 weeks, which seemed then to level off (27.8% new bone at 24 weeks). Cement residues appear as radio-opaque cylinders on microradiographs. In all cases, a radiolucent layer was observed at the cement/bone interface at 24 weeks. Stained sections showed the formation of a fibroconnective capsule around the residual cement, which presumably slows down new bone formation. Nevertheless, quantitative bone remodeling was accelerated in the cement group; mineral apposition as well as adjusted apposition rates were higher, and the formation period as well as the mineralization of osteoid tissue were faster compared with empty cavities and controls. These results point to higher osteoblast activity and better exchange with surrounding tissues in the defects filled with cement.

Association of porous hydroxyapatite and bone marrow cells for bone regeneration.

The preparation of hybrid material with osteoinductive capacity may be achieved by association of cultured autologous bone cells with a porous ceramic vehicle. We optimized culture conditions for rabbit marrow stromal stem cells (MSCs), notably by selection from batches of fetal calf serum. Rabbit MSCs formed colony-forming unit-ribroblastic (CFU-Fs) in vitro. Their alkaline phosphatase (ALP) activity was doubled in the presence of dexamethasone. Autologous rabbit serum allowed the formation of ALP-positive CFU-Fs, but results were highly variable depending on the rabbit. We tested the osteogenic potential of autologous cultured (with or without dexamethasone addition in the culture medium) and noncultured rabbit MSCs associated with a porous hydroxyapatite ceramic after a dorsal intramuscular implantation. Nucleated cells (10(7) or 10(8)/mL) were used for the preparation of autologous hybrid material. A significantly higher number of implants containing bone was obtained with a suspension of 10(7) cells/mL cultured in the presence of 10(-8) M dexamethasone. Some positive implants were also obtained with a suspension of 10(8) noncultured cells/mL. We demonstrated the feasibility of preparing rabbit autologous hybrid materials following a process for controlling culture conditions, cell characterization and cell/material association.

Raman microspectrometry studies of brushite cement: in vivo evolution in a sheep model.

Calcium phosphate hydraulic cements are promising synthetic bone grafting materials. Brushite-based cements were implanted for 6 and 12 months in the distal condyle of sheep femur, and their in vivo evolution was investigated by Raman microspectrometry. This new technique can probe small volumes in the cubic micrometer range. Its resolution allows a very fine analysis of crystalline changes in calcium phosphate mixtures at the microscopic level. First, Raman spectra of pure brushite, monetite, and beta-tricalcium phosphate (beta-TCP) were recorded, in order to set a data base for the basic components of brushite cements. These spectra show significant differences in the vibration mode v1 for the phosphate ion (988 and 878 cm(-1) for brushite, 988 and 900 cm(-1) for monetite, 968 and 948 cm(-1) for beta-TCP). These differences are strong enough as to allow the qualitative and quantitative analysis of these crystalline phases in the cement. Implanted sheep femur samples were harvested after 24 and 52 weeks post-op, and prepared for Raman analysis in the form of 1-mm-thick sections. Implants at 24 weeks show a core of residual cement isolated from the surrounding bone by fibroconnective tissue. No trace of brushite was detected by micro-Raman analysis in this area, but instead, a mixture of beta-TCP and Type-B carbonated apatite, the latter being very close in composition and structure to the mineral fraction of normal bone in the vicinity of the implant. Implants recovered after 52 weeks show a decrease of the bone/residual cement perimeter, whereas new trabeculations are formed in the implanted zone; the small amounts of residual cement still present are substantially transformed into Type-B carbonated apatite containing small amounts of proteins. In the same area, some beta-TCP particles are also detected showing that, contrary to brushite, the excess beta-TCP originally present in the cement is not completely metabolized. In the implanted zone already converted into trabecular bone, Raman microspectrometry shows the characteristic spectrum of normal bone.

Time-dependent morphology and adhesion of osteoblastic cells on titanium model surfaces featuring scale-resolved topography.

The role of micrometer and submicrometer surface roughness on the interaction of cells with titanium model surfaces of well-defined topography was investigated using human bone-derived cells (MG63 cells). The early phase of interactions was studied using a kinetic morphological analysis of adhesion, spreading and proliferation of the cells. By SEM and double immunofluorescent labeling of vinculin and actin, it was found that the cells responded to nanoscale roughness by a higher cell thickness and a delayed apparition of the focal contacts. A singular behavior was observed on nanoporous oxide surfaces, where the cells were more spread and displayed longer and more numerous filopods. On electrochemically microstructured surfaces with hemispherical cavities, arranged in a hexagonal pattern, the MG63 cells were able to go inside, adhere and proliferate in cavities of 30 or 100 microm in diameter, whereas they did not recognize the 10 microm diameter cavities. Cells adopted a 3D shape when attaching inside the 30 microm diameter cavities. Condensation of actin cytoskeleton correlated with vinculin-positive focal contacts on cavity edges were observed on all microstructured surfaces. Nanotopography on surfaces with 30 microm diameter cavities had little effect on cell morphology compared to flat surfaces with same nanostructure, but cell proliferation exhibited a marked synergistic effect of microscale and nanoscale topography.

Improvement in the morphology of Ti-based surfaces: a new process to increase in vitro human osteoblast response.

Surface roughness has been shown to be an influencing parameter for cell response. In this experience we attempted to compare the effect of roughness organization of Ti6A14V or pure titanium substrates on human osteoblast (hOB) response (proliferation, adhesion). Surface roughness was extensively analyzed at scales above the cell size (macro-roughness) or below the cell size (micro-roughness) by calculation of relevant classic amplitude parameters (Ra, Rt) and original frequency parameters (Order, Delta). We developed a new process to prepare isotropic surfaces (electro-erosion), which were compared to isotropic surfaces obtained by polishing and anisotropic surfaces obtained by machine-tooling. The hOB response on electro-eroded (EE) Ti6A14V surfaces or pure titanium (Ti) surfaces was largely increased when compared to polished or machine-tooled surfaces after 21 days of culture. Moreover, the polygonal morphology of hOB on these EE surfaces was very close to the aspects of hOB in vivo on human bone trabeculae. By a complete description of the surface topography of EE surfaces, we concluded that when the topography was considered below the cell scale, hOB appreciated their isotropic smooth aspect, although when the topography was considered above the cell scale they appreciated their rough isotropic 'landscape' formed by many 'bowl-like nests' favouring cell adhesion and growth. Electro-erosion is a promising method for preparation of bone implant surfaces, as it could easily be applied to preparation of most biomaterials with complex geometries.

Fate of bioresorbable poly(lactic acid) microbeads implanted in artificial bone defects for cortical bone augmentation in dog mandible.

The fate was examined of poly(lactic acid) microbeads implanted in large artificial defects created in cortical bone of dog mandibles. Two poly(lactic acid) polymers--poly(L-lactic acid) (PLA 100) and poly(DL-lactic acid) (PLA 50)--were used to make microbeads by solvent evaporation with poly(vinyl alcohol) as surfactant. Histological observation of non-decalcified mandibular bone showed that no real bone regeneration existed in the experimental bone defects 18 months after PLA 100 microbeads implantation. The same observation was made 6 months after implantation of PLA 50 microbeads. PLA 100 and PLA 50 microbeads appeared unable to induce regeneration of cortical bone defects of dog mandible, in contrast to previous observations in man for PLA 50 large implants. The failure is tentatively assigned to the presence of poly(vinyl alcohol) at the surface of microbeads.

The relative influence of the topography and chemistry of TiAl6V4 surfaces on osteoblastic cell behaviour.

Proliferation and adhesion of mouse (MC3T3-E1) osteoblastic cells and primary human osteoblastic cells were carried out on Ti6Al4V titanium alloy samples with varied surface roughnesses. Mechanically or manually polished surfaces were prepared to produce respectively non-oriented or oriented residual polishing grooves. Sand-blasted surfaces were prepared using 500 microm or 3 mm alumina particles. Surface roughness parameters showed a negative correlation in comparison to proliferation and adhesion parameters. X-ray microprobe chemical surface microanalysis showed complete disturbance of the surface element composition of the Ti6Al4V alloy following sand-blasting treatment. An AlOx-enriched layer was observed on sample surfaces. This may lead to the suspicion that the concomittant effect of surface roughness amplitude and AlOx surface concentration has an effect on osteoblastic cell proliferation and adhesion. These findings show the significance of chemical surface analysis after any surface treatment of titanium-based implants before any biological use.

An unscaled parameter to measure the order of surfaces: a new surface elaboration to increase cells adhesion.

We present a new parameter to quantify the order of a surface. This parameter is scale-independent and can be used to compare the organization of a surface at different scales of range and amplitude. To test the accuracy of this roughness parameter versus a hundred existing ones, we created an original statistical bootstrap method. In order to assess the physical relevance of this new parameter, we elaborated a great number of surfaces with various roughness amplitudes on titanium and titanium-based alloys using different physical processes. Then we studied the influence of the roughness amplitude on in vitro adhesion and proliferation of human osteoblasts. It was then shown that our new parameter best discriminates among the cell adhesion phenomena than others' parameters (Average roughness (Ra em leader )): cells adhere better on isotropic surfaces with a low order, provided this order is quantified on a scale that is more important than that of the cells. Additionally, on these low ordered metallic surfaces, the shape of the cells presents the same morphological aspect as that we can see on the human bone trabeculae. The method used to prepare these isotropic surfaces (electroerosion) could be undoubtedly and easily applied to prepare most biomaterials with complex geometries and to improve bone implant integration. Moreover, the new order parameter we developed may be particularly useful for the fundamental understanding of the mechanism of bone cell installation on a relief and of the formation of bone cell-material interface.

Genetic mapping of 66 new microsatellite (SSR) loci in bread wheat.

In hexaploid bread wheat ( Triticum aestivum L. em. Thell), ten members of the IWMMN ( International Wheat Microsatellites Mapping Network) collaborated in extending the microsatellite (SSR = simple sequence repeat) genetic map. Among a much larger number of microsatellite primer pairs developed as a part of the WMC ( Wheat Microsatellite Consortium), 58 out of 176 primer pairs tested were found to be polymorphic between the parents of the ITMI ( International Triticeae Mapping Initiative) mapping population W7984 x Opata 85 (ITMI pop). This population was used earlier for the construction of RFLP ( Restriction Fragment Length Polymorphism) maps in bread wheat (ITMI map). Using the ITMI pop and a framework map (having 266 anchor markers) prepared for this purpose, a total of 66 microsatellite loci were mapped, which were distributed on 20 of the 21 chromosomes (no marker on chromosome 6D). These 66 mapped microsatellite (SSR) loci add to the existing 384 microsatellite loci earlier mapped in bread wheat.

Epidemiology and optimal management of polymyalgia rheumatica.

Polymyalgia rheumatica (PMR) is a disease of unknown aetiology that occurs in elderly patients, predominantly affecting the Caucasian population. The disease has a slightly higher prevalence in women than in men. There is ongoing discussion regarding the relationship between PMR and giant cell arteritis; an increasing number of studies indicate that they are closely related. PMR has also been linked with rheumatoid arthritis, myopathy and malignant disease. Oral corticosteroids remain the mainstay of drug therapy for PMR. These drugs usually induce prompt relief of symptoms, and some authors consider this dramatic response to be diagnostic for PMR. However, the ideal initial dosage, the duration of treatment and the optimal tapering schedule are much debated. Other drugs, such as methotrexate and azathioprine, have been suggested as corticosteroid sparing agents. Nonsteroidal anti-inflammatory drugs are generally considered to be unsuitable for the long term treatment of PMR.

Influence of polymeric additives on the biological properties of brushite cements: an experimental study in rabbit.

The resorbability and ability of calcium phosphate hydraulic cements to promote new bone formation was investigated in vivo. The effects of two hydrosoluble polymeric additives (hyaluronic acid, and xanthan gum,) on the biological response of two brushite cement formulations (BHC-A vs BHC-B) was investigated. The brushite cements differed in P/Ca (0.71 vs 0.98) and S/Ca (0.10 vs 0.005) atomic ratios and by the presence of calcium sulfate hemihydrate in BHC-A. Polymer-free cements were used as controls. Cement specimens were injected in cylindrical bone defects manually drilled in the distal condyle of rabbit femora. The implants were harvested at 12 and 24 weeks after implantation and subjected to quantitative histomorphometry. The study showed a significantly lower resorption rate for cement BHC-A, which induces the formation of well-mineralized bone in close apposition to the residual material. In contrast, cement BHC-B showed a significant increase of bone formation period and the formation of a thick layer of unmineralized osteoid tissue at the bone/residual cement interface. The presence of xanthan gum made the biological response even worse, particularly in the case of cement BHC-B. The presence of hyaluronic acid has little effect, except for a slight decrease in initial resorption rate, in the case of cement BHC-A.

Experimental model of posterolateral spinal arthrodesis in sheep. Part 1. Experimental procedures and results with autologous bone graft.

OBJECTIVES: The authors evaluated the reliability in obtaining a posterolateral spinal arthrodesis (PSA) with autologous bone graft. SUMMARY OF BACKGROUND DATA: Posterolateral spinal arthrodesis using autogenous cancellous bone graft is the most simple and efficient technique to get a spinal graft. No extensive biomechanical study of PSA is available. Thus, an experimental model of PSA is needed. METHODS: Eleven sheep underwent lumbar autologous bone grafts and Cotrel-Dubousset instrumentations, and four sheep were used as controls. Sacrifice and biomechanical evaluation of the lumbar spines were performed after 1 year. RESULTS: All grafts appeared continuous. A large decrease of flexibility (in rotation and in translation) was found in grafted spines in every direction. Failure in extension occurred at a mean value of 35.26 +/- 3.71 Nm. CONCLUSION: A constant and homogenous PSA appears to be obtained in sheep under conditions close to the human surgery.

Experimental model of posterolateral spinal arthrodesis in sheep. Part 2. Application of the model: evaluation of vertebral fusion obtained with coral (Porites) or with a biphasic ceramic (Triosite).

OBJECTIVES: The authors evaluated two bone substitutes in a posterolateral spinal arthrodesis (PSA) model in sheep: coral porites (99% calcium carbonate, Biocoral, Inoteb, France) and a biphasic ceramic (BCP) (65% hydroxyapatite and 35% B tricalcium phosphate, Triosite, Zimmer International). SUMMARY OF BACKGROUND DATA: Bone substitutes would be of great interest for PSA. Previous trials began with two kinds of biomaterials: natural coralline calcium carbonate, and phosphate calcium ceramic. METHODS: A lumbar PSA was performed in 11 sheep (coral group) and in 9 sheep (BCP group). Sacrifice and biomechanical tests were performed after 1 year. RESULTS: A large decrease of flexibility in all directions was obtained with both coral PSA and BCP PSA similarly to autologous graft. No nonfusion case was observed. CONCLUSION: In conditions close to the human surgery, a PSA can be obtained using either coral porites or BCP as bone substitutes.

Dialysis-related beta 2 microglobulin-amyloid arthropathy. Improvement of clinical symptoms after a switch of dialysis membranes.

Amyloidosis containing beta 2 microglobulin (beta 2m) and joint pains are frequent complications of long-term hemodialysis. In a prospective study the authors planned to act on dialysis arthropathies (DA) by improving beta 2m depuration: for 8 patients suffering from DA we replaced usual cuprophane membranes (CU) by high permeability polyacrylonitrile membranes (PAN). DA was diagnosed on the presence of severe disabling arthralgias in the absence of any other obvious etiology, and on the presence of at least 3 among 6 criteria. The efficacy of the switch of DM was assessed on 6 criteria including the patient's own evaluation at the end of the study and a joint pain severity index. Clinical evaluation and beta 2m measurement were carried out at the start of the trial and after a mean duration of dialysis on PAN of 6 months. An improvement in joint symptomatology and a decrease of joint severity index were noted in the 8 cases. Our results suggest that the change of DM has a favorable influence on DA. This improvement could result either from a better beta 2m depuration or from a better biocompatibility of PAN than the CU previously used.

Migration of polyethylene debris along well-fixed cemented implants.

Implants, consisting of smooth Inox cylinders, were cemented into the lower femur and upper tibia of nine sheep to study the distal migration of polyethylene particles. Some implants had a titanium-bead porous coat at the proximal end. These were of three types: In the first type, the porous coat was covered with hydroxyapatite to obtain a bony seal; the second type was prepared for a polymethylmethacrylate seal; in the third type, the porous zone was surrounded by a 2-mm-thick space to allow the formation of a fibrous seal. Small polyethylene particles were injected into the knees once a week during the third and fourth months after implantation. The animals were euthanized 2 months later. Major longitudinal sections of the implants and the surrounding bone were examined under a polarized light microscope. Birefringent particles were counted at the cement-bone and cement-implant interfaces. Osteolysis was not observed. None of the seals significantly decreased the migration of particles around the cemented part of the implants. Particles were observed in cement fissures and vacuoles. They migrated at both interfaces and in the bone itself. They were visible in marrow spaces between bone trabeculae.

Should percutaneous vertebroplasty be used to treat osteoporotic fractures? An update.

Acrylic cement vertebroplasty is being increasingly used to treat osteoporotic vertebral compression fractures (VCFs), although no controlled studies supporting this trend have been published. Vertebroplasty remains controversial as a treatment for osteoporotic fractures because it is a local response to a systemic disease and because the pain caused by osteoporotic fractures usually subsides within a few days or weeks. Current data suggest that pain severity may decrease by half, on average, in 90-100% of patients. Although vertebroplasty is usually well tolerated, serious neurological complications have been reported in a few patients. The most common adverse event is nerve root pain, usually caused by leakage of the cement into the intervertebral foramen. Whether vertebroplasty is followed by an increased risk of osteoporotic fractures in the adjacent vertebras remains unclear. Resorbable cements are being developed and may provide better results than the acrylic cements used today. At present, acrylic cement vertebroplasty to treat osteoporotic VCFs is appropriate in only a minority of patients selected carefully by a multidisciplinary team including a rheumatologist.

Tissue engineering and skeletal diseases.

Tissue engineering, a cross between the science of the living organism and that of engineering, aims to replace, maintain or improve human tissue functions, by means of tissue substitutes containing living elements. Thus, it is about production of artificial tissue, using (alone or in combination) cells, matrix or bioactive factors. Their association gives rise to a hybrid biomaterial combining biological components (cells, growth factors or adhesion proteins) and materials (polymers, ceramics). The applications are wide-ranging, from the skin, to the liver, or to the cornea as well as to the locomotor system. Bone tissue engineering has advanced the most in this field, partly because of the progress made by research into bone substitutes, although cartilage and tendons are also concerned. This technology requires cell culture (committed cells or more often bone marrow stem cells), biomaterials (porous materials with controlled architecture and cements), growth factors (such as 'Bone Morphogenetic Proteins'), the proteins implicated in cell adhesion (such as fibronectin or the aminoacid sequences specifically recognised by integrin subunits) or gene therapy (notably using transfected stem cells). Tissue engineering and regenerative stimulation of tissue are now booming on experimental and industrial levels and clinical applications are increasingly numerous. Considering the potential of these technologies, they should continue to develop widely.