Search

The relationship between cartilage growth - mineralization patterns were studied in adult Rajidae with X-ray morphology/morphometry, undecalcified resin-embedded, heat-deproteinated histology and scanning electron microscopy. Morphometry of the wing-fins, nine central rays of the youngest and oldest specimens documented a significant decrement of radials mean length between inner, middle and outer zones, but without a regular progression along the ray. This suggests that single radial length growth is regulated in such a way to align inter-radial joints parallel to the wing metapterygia curvature. Trans-illumination and heat-deproteination techniques showed polygonal and cylindrical morphotypes of tesserae, whose aligned pattern ranged from mono-columnar, bi-columnar, and multi-columnar up to the crustal-like layout. Histology of tessellated cartilage allowed to identify of zones of the incoming mineral deposition characterized by enhanced duplication rate of chondrocytes with the formation of isogenic groups, whose morphology and topography suggested a relationship with the impending formation of the radials calcified column. The morphotype and layout of radial tesserae were related to mechanical demands (stiffening) and the size/mass of the radial cartilage body. The cartilage calcification pattern of the batoids model shares several morphological features with tetrapods' endochondral ossification, that is, (chondrocytes' high duplication rate, alignment in rows, increased volume of chondrocyte lacunae), but without the typical geometry of the metaphyseal growth plates. RESEARCH HIGHLIGHTS: 1. The wing-fins system consists of stiff radials, mobile inter-radial joints and a flat inter-radial membrane adapted to the mechanical demand of wing wave movement. 2. Growth occurs by forming a mixed calcified-uncalcified cartilage texture, developing intrinsic tensional stresses documented by morphoanatomical data.

The authors studied the morphology of the upper and lower jaws, vertebrae and dorsal-fin rays of the teleost fish Xiphias gladius to analyse the skeletal architecture and ossification pattern. The analogies and differences among these segments were investigated to identify a common morphogenetic denominator of the bone tissue osteogenesis and modeling. The large fat glands in the proximal upper jaw and their relationship to the underlying cartilage (absent in the lower jaw) suggested that there is a mechanism that explains rostral overgrowth in the Xiphiidae and Istiophoriidae families. Thus far, the compact structure of the distal rostrum has been interpreted as being the result of remodeling. Nonetheless, no evidence of cutting cones, scalloped outer border of osteons and sequence of bright-dark bands in polarized light was observed in this study, suggesting a primary osteon texture formed by compacting of collagen matrix and mineral deposition in the fat stroma lacunae of the bone, but without being oriented in layers of the collagen fibrils. A similar histology also characterizes the circular structures present in the other examined segments of the skeleton. The early phases of fibrillogenesis carried out by fibroblast-like cells occurred farther from the already-calcified bone surface inside the fat stroma lacunae. The fibrillar matrix was compacted and underwent mineral deposition near the previously calcified bone surface. This pattern of collagen matrix synthesis and calcification was different from that of mammalian osteoblasts, especially concerning the ability to build a lacuno-canalicular system among cells. Necrosis or apoptosis of the latter and refilling of the empty lacunae by mineral deposits might explain the anosteocytic bone formation.

The skeleton of the batoid fish consists of a mixture of calcified and uncalcified cartilage with a typical layout of mineral deposition toward the outer border, leaving an uncalcified central core in most of the skeleton segments. An exception is observed in the radials, where mineral deposition is central. Joints and endoskeleton segments were studied in two adult samples of Raja cf. polystigma. Histomorphology, mineral deposition pattern, and zonal chondrocyte duplication activity were compared among several endoskeleton segments, but with particular attention to the fin rays; in the first, the uncalcified cartilage is central with an outer layer ranging from mineralized tesserae to a continuous calcified coating, whereas in the second, the uncalcified cartilage surrounds one or more central calcified columns. The diarthroses have a joint cavity closed by a fibrous capsule and the sliding surfaces rest on the base of mineralized tesserae, whereas the interradial amphiarthroses show a layer of densely packed chondrocytes between the flat, calcified discs forming the base of neighboring radials. In the endoskeleton segments, three types of tesserae are distinguished, characterizing the phases of skeletal growth and mineralization which present differences in each endoskeleton segment. The chondrocyte density between central core, subtesseral layer, and radial external cartilage did not show significant differences, while there was a significant difference in chondrocyte density between the latter zones and the type c tesserae of the pelvic girdle. The histomorphology and morphometry observed in Raja cf. polystigma suggest a model of cartilage growth associated with structural stiffening without remodeling. A key point of this model is suggested to be the incomplete mineralization of the tesseral layer and the continuous growth of cartilage, both enabling fluid diffusion through the matrix fibril network of scattered, uncalcified cartilage zones inside and between the tesserae.

Primary lymphoma of bone (PLB) is a rare entity, defined as a lymphoma confined to the bone without evidence of systemic involvement. The disease commonly affects middle-aged to elderly population and it accounts for less than 1% of all malignant lymphomas. We present a case of a 10-year-old child affected by PLB of the forearm and the frontal bone. Characteristic imaging features of PLB and the main differential diagnosis were discussed. Keywords: Bone, Lymphoma, Paediatric

Background: Five-fingered hand (5-FH) with completely developed phalanges is a rare phenotype observed so far only in humans and characterised by three phalanges of the 1st ray. A long-lasting, debated question is if the missing element of the normal hand 1st ray is the metacarpal or the phalanx. In this study, comparative X-rays morphometry of long bones in normal and 5-FH is carried out with the aim to face this question through homology analysis of long bone segments in the transverse and longitudinal line of normal hand and 5-FH. Materials and methods: In the normal hand X-rays (n =20) and in a 5-FH X-rays series (n = 9) the relative length of each segment on the ray total length and the index of growth rate (IGR) were assessed. The calculation of the first parameter in normal hand bi-phalangeal thumb was carried out on the 3rd ray total length in the same hand. Results: The parameters of relative length and the proximal/distal growth rate asymmetry in the post-natal period (assessed through the IGR) confirmed in 5-FH the homology of all the five segment on the transverse line. In the normal control hand, the relative length assessment methodology was biased by the missing segment of the thumb, therefore, the reference to the 3rd ray total length in the same hand (instead of the 1st), allowed the homology analysis of the thumb metacarpal and 1st phalanx with the lateral segments (2nd–5th ray) of the same hand. The 5-FH analysis was used to choose the more appropriate reference ray for the normal hand group. Conclusions: The comparative analysis of relative lengths and IGRs in the two groups suggested homology of the (anatomical) 1st metacarpal with the 2nd–5th proximal phalanges in the same hand and that of the (anatomical) 1st proximal phalanx with the 2nd–5th mid phalanges. These data suggest that the missing segment of the normal hand thumb is the metacarpal.

Calcination and decalcification are basic procedures useful to a morphological approach of a biological, composite material like cortical bone. The study was carried out on a whole human femur conserved in liquid (from an educational collection). Cortical fracturing and SEM observation of vascular canals surface collagen texture was used to study bone deproteination at scalar temperatures (400-1,200°C) and acid bone decalcification at crescent time intervals. Heating burned and vaporized the organic matrix with shrinkage of the bone specimens as documented by the weight loss and transverse surface morphometry. SEM showed a pattern of aligned spherulites at 400°C which maintained the collagen fibrils layout (like a mineral cast), followed by a spherulites fusion progression with the temperature increments. At 1200°C a crystalline-like structure of tightly-packed trapezohendron units. XRD analysis supported the SEM morphology displaying the complete Debey rings of hydroxyapatite and spotted Debey rings of withlockite. Surface Ca and P elution was documented after 12 hr of exposition to the acid solution by dissolution of spherulites and the whole canal surface decalcified in depth after 15 days by SEM-EDAX analysis. The periodic pattern of collagen fibrils was still evident up to 15 days of decalcification together with fine granular deposits of a not-collagenic proteic material, while after 30 days no period was observed in the decalcified fibrils. Collagen mineral cast at 400°C calcination. Complete crystalline transformation at 1200°C. Up to 15 days of decalcification fibrils period maintained.

The 'fate' of growth plate hypertrophic chondrocytes has been long debated with two opposing theories: cell apoptosis or survival with transformation into osteogenic cells. This study was carried out on the proximal tibial growth plate of rabbits using light microscopy, scanning and transmission electron microscopy. We focused particularly on the orientation of the specimens included in order to define the mineral deposition and the vascular invasion lines and obtain histological and ultrastructural images at the corresponding height of the plate. Chondrocyte morphology transformation through the maturation process (characterized by vesicles and then large cytoplasmic lacunae before condensation, fragmentation and disappearance of the nuclear chromatin) did not correspond to that observed in the 'in vitro' apoptosis models. These findings rather suggested the passage of free water from the cartilage matrix into a still live cell (swelling). The level of these changes suggested a close relationship with the mineral deposition line. Furthermore, the study provided evidence that the metaphyseal capillaries could advance inside the columns of stacked hypertrophic chondrocytes (delimited by the intercolumnar septa) without the need for calcified matrix resorption because the thin transverse septa between the stacked chondrocyte (below the mineral deposition line) were not calcified. The zonal distribution of cell types (hypertrophic chondrocytes, osteoblasts, osteoclasts and macrophages) did not reveal osteoclasts or chondroclasts at this level. Morphological and morphometric analysis recorded globular masses of an amorphous, necrotic material in a zone 0-70 μm below the vascular invasion line occasionally surrounded by a membrane (indicated as 'hypertrophic chondrocyte ghosts'). These masses and the same material not bound by a membrane were surrounded by a large number of macrophages and other blood cell precursors, suggesting this could be the cause of macrophage recall and activation. The most recent hypotheses based on genetic and lineage tracing studies stating that hypertrophic chondrocytes can survive and transform into osteoblasts and osteocytes (trans-differentiation) were not confirmed by the ultrastructural morphology or by the zonal comparative counting and distribution of cell types below the vascular invasion line.

The progression of mineral phase deposition in hypertrophic cartilage and periosteal bone matrix was studied in human metacarpals primary ossification centers before vascular invasion began. This study aimed to provide a morphologic/morphometric comparative analysis of the calcification process in cartilage and periosteal osteoid used as models of endochondral ossification. Thin, sequential sections from the same paraffin inclusions of metacarpal anlagen (gestational age between the 20th and 22nd weeks) were examined with light microscopy and scanning electron microscopy, either stained or heat-deproteinated. This process enabled the analysis of corresponding fields using the different methods. From the initial CaPO4 nucleation in cartilage matrix, calcification progressed increasing the size of focal, globular, randomly distributed deposits (size range 0.5-5 µm), followed by aggregation into polycyclic clusters and finally forming a dense, compact mass of calcified cartilage. At the same time, the early osteoid calcification was characterized by a fine granular pattern (size range 0.1-0.5 µm), which was soon compacted in the layer of the first periosteal lamella. Scanning electron microscopy of heat-deproteinated sections revealed a rod-like hydroxyapatite crystallite pattern, with only size differences between the early globular deposits of the two calcifying matrices. The morphology of the early calcium deposits was similar in both cartilage and osteoid, with variations in size and density only. However, integration of the reported data with the actual hypotheses of the mechanisms of Ca concentration suggested that ion transport was linked to the progression of the chondrocyte maturation cycle (with recall of H2 O from the matrix) in cartilage, while ions transport was an active process through the cell membrane in osteoid. Other considered factors were the collagen type specificity and the matrix fibrillar texture. Anat Rec, 301:571-580, 2018. © 2017 Wiley Periodicals, Inc.

The purpose of the paper was to assess the morphometric parameters to improve the specificity of the ultrasound (US) signs for the early differential diagnosis between two lethal dysplasias, as thanatophoric dysplasia (TD) and osteogenesis imperfecta type 2 (OI-2). The diaphyseal length and the bowed shape of long bones associated with vertebral body dimension assessment were investigated in a group of 14 pregnancy terminations carried out in the time period 2007–2013. The definitive diagnosis was established after pregnancy termination by means of skeletal standardized X-rays, histopathology and gene analysis. TD and OI-2 long bones were significantly shorter than controls. No significant differences were observed between the two dysplasias. The bowing angle was higher in OI-2; a true angulation or eventually axial displacement was present only in the latter. Furthermore, they did not show any evidence of vertebral collapse. The thanatophoric dysplasia presented less bowed long bones, and never true angulation. The spine was steadily characterized by flattened anterior vertebral bodies. Long bone shortening is not a sufficient and accurate sign for early sonographic differential diagnosis between TD and OI-2. Angled diaphysis, axial diaphyseal displacement and a conserved vertebral body height in the prenatal period support the diagnosis of osteogenesis imperfecta type 2, while moderately regular bowed diaphysis associated with platyspondyly that of thanatophoric dysplasia.

The patterns of longitudinal and peripheral growth were analyzed in human autopod cartilage anlagen (fetal developmental stage 20th-22nd week) through morphometric assessment of chondrocyte parameter size, shape, alignment and orientation between peripheral and central sectors of the anlage transition zone defined by primary ossification center and the epiphyseal basis. The aim was to correlate the chondrocyte dynamics with the longitudinal and peripheral growth. A further comparison was carried out between the corresponding sectors of the postnatal (3-5 months old) growth plate cartilage documenting: (1) the different chondrocyte framework and the new peripheral mechanism; (2) the opposite direction of fetal periosteal ossification versus the Lacroix bone bark. Measurement of multiple parameters (% lac area, % total matrix area, total lac density and mean single lac area), which characterize the cartilage Anlage growth, suggested the following correlations with chondrocyte duplication rate: (a) slow duplication rate ≈ coupled, intralacunar chondrocytes (in central epiphysis); (b) repeated/frequent cell duplications ≈ clusters (in the basal epiphyseal layer); (c) clusters of chondrocytes before becoming hypertrophic were stacked up on the top of each other (both in the Anlage transition zone or in the columns of metaphyseal growth plate); (d) enhanced osteoclastic resorption of the Lacroix bone bark lower end, extended to the more external metaphyseal trabeculae counterbalancing the discrepancy between the epiphyseal and the diaphyseal circumferential growth.

The chondrocyte maturation cycle and endochondral ossification were studied in human, fetal cartilage Anlagen and in postnatal meta-epiphyses. The relationship between the lacunar area, the inter-territorial fibril network variations, and calcium phosphorus nucleation in primary and secondary ossification centers were assessed using light microscopy and scanning electron microscopy (SEM) morphometry. The Anlage topographic, zonal classification was derived from the anatomical nomenclature of the completely developed long bone (diaphysis, metaphyses and epiphyses). A significant increase in the chondrocyte lacunar area was documented in the Anlage of epiphyseal zones 4 and 3 to zone 2 (metaphysis) and zone 1 (diaphysis), with the highest variation from zone 2 to zone 1. An inverse reduction in the intercellular matrix area and matrix interfibrillar empty space was also documented. These findings are consistent with the osmotic passage of free cartilage water from the interfibrillar space into the swelling chondrocytes, which increased the ion concentrations to a critical threshold for mineral precipitation in the matrix. The mineralized cartilage served as a scaffold for osteoblast apposition both in primary and secondary ossification centers and in the metaphyseal growth plate cartilage, though at different periods of bone Anlage development and with distinct patterns for each zone. All developmental processes shared a common initial pathway but progressed at different rates, modes and organization in diaphysis, metaphysis and epiphysis. In the ossification phase the developing vascular supply appeared to play a key role in determining the cortical or trabecular structure of the long bones. J. Morphol. 277:1187-1198, 2016. © 2016 Wiley Periodicals, Inc.

Observation of heat-deproteinized cortical bone specimens in incident light enabled the high definition documentation of the osteonal pattern of diaphyseal Haversian bone. This prompted a study to compare these images with those revealed by polarized light microscopy, carried out either on decalcified or thin, undecalcified, resin-embedded sections. Different bone processing methods can reveal structural aspects of the intercellular matrix, depending on the light diffraction mode: birefringency in decalcified sections can be ascribed to the collagen fibrils orientation alone; in undecalcified sections, to both the ordered layout of collagen and the inorganic phase; in the heat-deproteinized samples, exclusively to the hydroxyapatite crystals aggregation mode. The elemental chemical analysis documented low content of carbon and hydrogen, no detectable levels of nitrogen and significantly higher content of calcium and phosphorus in heat-deproteinized samples, as compared with dehydrated controls. In both samples, the X-ray diffraction (XRD) pattern did not show any significant difference in pattern of hydroxyapatite, with no peaks of any possible decomposition phases. Scanning electron microscopic (SEM) morphology of heat-deproteinized samples could be documented with the fracturing technique facilitated by the bone brittleness. The structure of crystal aggregates, oriented in parallel and with marks of time periods, was documented. Comparative study of deproteinized and undecalcified samples showed that the matrix inorganic phase did not undergo a coarse grain thermal conversion until it reached 500°C, maintaining the original crystals structure and orientation. Incident light stereomicroscopy, combined with SEM analysis of deproteinized bone fractured surfaces, is a new enforceable technique which can be used in morphometric studies to improve the understanding of the osteonal dynamics. Microsc. Res. Tech. 79:691-699, 2016. © 2016 Wiley Periodicals, Inc.

ORTHOUNION is a multicentre, open, comparative, three-arm, randomized clinical trial (EudraCT number 2015-000431-32) to compare the efficacy, at one and two years, of autologous human bone marrow-derived expanded mesenchymal stromal cell (hBM-MSC) treatments versus iliac crest autograft (ICA) to enhance bone healing in patients with diaphyseal and/or metaphysodiaphyseal fracture (femur, tibia, and humerus) status of atrophic or oligotrophic nonunion (more than 9 months after the acute fracture, including recalcitrant cases after failed treatments). The primary objective is to determine if the treatment with hBM-MSCs combined with biomaterial is superior to ICA in obtaining bone healing. If confirmed, a secondary objective is set to determine if the dose of 100 × 106 hBM-MSCs is noninferior to that of 200 × 106 hBM-MSCs. The participants (n=108) will be randomly assigned to either the experimental low dose (n=36), the experimental high dose (n=36), or the comparator arm (n=36) using a central randomization service. The trial will be conducted in 20 clinical centres in Spain, France, Germany, and Italy under the same clinical protocol. The confirmation of superiority for the proposed ATMP in nonunions may foster the future of bone regenerative medicine in this indication. On the contrary, absence of superiority may underline its limitations in clinical use.

Transverse and longitudinal sectioning of undecalcified cortical bone is a commonly employed technique for investigating the lamellar structure of the osteons. Since a flat surface is required, the specimen has to be grinded and then polished. Whereas the smear of debris and inorganic/organic deposits left by these treatments cannot be removed by ultrasonication alone, a chemical treatment of the specimen surface with either a basic or an acid etching solution is currently employed. A further effect of the latter can be the enhancement of the lamellar bone pattern. The kind of etching solution, its pH, the concentration of etchants, and the contact time significantly affect the sectioned surface when it is observed with scanning electron microscopy (SEM). The etching procedures can severely influence the obtained images. Homogeneous cortical bone specimens were sampled from the first metatarsal of two fresh human subjects. One or two cut surfaces were exposed to different acid and basic solutions in bonded conditions. Considering the type of chemical agents, the solution pH, and the exposure time of the specimens, the effects of several etching media have been investigated and compared. Strong etching, either acid or basic produced surface decalcification and severe damage of the collagen matrix, compromising any morphological or morphometric analysis. Weak acid etching (for example citric and acetic acid), even though causing distinctive alteration of the sample, enhanced the visibility of the lamellar pattern, while the polyphosphate treatment of the surface decalcified a thin layer matrix, ensuring a good visibility of fibrils and avoiding rough distortions. Microsc. Res. Tech. 77:653–660, 2014. © 2014 Wiley Periodicals, Inc.

A histological and morphometric analysis of human metacarpal and carpal anlagen between the 16th and 22nd embryonic weeks was carried out with the aim of studying the establishment of the respective anlage architecture. No differences in the pattern of growth were documented between the peripheral and central zones of the metacarpal epiphyses and those of the carpals. The regulation of longitudinal growth in long bone anlagen occurred in the transition zone between the epiphysis and the diaphysis (homologous to the metaphyseal growth plate cartilage in more advanced developmental stage of the bone). Comparative zonal analysis was conducted to assess the chondrocyte density, the mean chondrocyte lacunar area, the paired chondrocyte polarity in the orthogonal longitudinal and transverse planes, and the lacunar shape transformation in the metacarpal. In transition from epiphysis to diaphysis chondrocyte density decreased and mean lacunar area increased. No significant differences in the chondrocyte maturation cycle were observed between proximal/distal metacarpal epiphyses and the carpal anlagen. The number of paired chondrocyte oriented along the growth vector was significantly higher in both proximal/distal transition zones between epiphysis and diaphysis. Human metacarpals shared with experimental models (like mice and nonmammal tetrapods) an early common chondrocyte maturation cycle but with a different timing due to the slower embryonic and fetal developmental rate of human anlagen.

Transformation of osteoblasts into osteocytes is marked by changes in volume and cell shape. The reduction of volume and the entrapment process are correlated with the synthesis activity of the cell which decreases consequently. This transformation process has been extensively investigated by transmission electron microscopy (TEM) but no data have yet been published regarding osteoblast-osteocyte dynamic histomorphometry. Scanning electron microscope (SEM) densitometric analysis was carried out to determine the osteoblast and open osteocyte lacunae density in corresponding areas of a rabbit femur endosteal surface. The lining cell density was 4900.1 ± 30.03 n mm(-2), the one of open osteocyte lacunae 72.89 ± 22.55 n mm(-2). This corresponds to an index of entrapment of one cell every 67.23 osteoblasts (approximated by defect). The entrapment sequence begins with flattening of the osteoblast and spreading of equatorial processes. At first these are covered by the new apposed matrix and then also the whole cellular body of the osteocyte undergoing entrapment. The dorsal aspect of the cell membrane suggests that closure of the osteocyte lacuna may be partially carried out by the same osteoblast-osteocyte which developed a dorsal secretory territory. A significant proportion of the endosteal surface was analysed by SEM, without observing any evidence of osteoblast mitotic figures. This indicates that recruitment of the pool of osteogenic cells in cortical bone lamellar systems occurs prior to the entrapment process. No further additions occurred once osteoblasts were positioned on the bone surface and began lamellar apposition. The number of active osteoblasts on the endosteal surface exceeded that of the cells which become incorporated as osteocytes (whose number was indicated by the number of osteocyte lacunae). Therefore such a balance must be equilibrated by the osteoblasts' transformation in resting lining cells or by apoptosis. The current work characterised osteoblast shape changes throughout the entrapment process, allowing approximate calculation of an osteoblast entrapment index in the rabbit endosteal cortex.

The wedges of the mid-diaphyseal osteotomies carried out to correct the femoral and/or tibial native deformity in type III osteogenesis imperfecta (OI III) were used to study the remodeling patterns and lamellar organization at the level of the major deformity. Histology and scanning electron microscopy (SEM) morphology showed abnormal cortical remodeling characterized by the failure to form a cylinder of compact bone with a regular marrow canal. Atypical, flattened, and large resorption lacunae with a wide resorption front on one side and systems of parallel lamellae on the opposite side were observed, resembling those formerly reported as drifting osteons. SEM morphometry documented a higher percentage of nonossified vascular/resorption area (44.3 %) in OI than in controls (13.6 %), a lower density of secondary osteons, and lower values for the parameters expressing the individual osteon size. The mean osteon total area, the mean central canal area, and the mean osteon bone area of two selected, randomized populations of secondary osteons were significantly higher (p

The lamellar architecture of secondary osteons (Haversian systems) has been studied with scanning electron microscopy (SEM) in transverse sections of human cortical bone. Na3PO4 etching was used to improve the resolution of the interface between neighboring lamellae and the precision of measurements. These technical improvements permitted testing of earlier morphometry assumptions concerning lamellar thickness while revealing the existence of different lamellar patterns. The mean lamellar thickness was 9.0 ± 2.13 μm, thicker and with a wider range of variation with respect to earlier measurements. The number of lamellae showed a direct correlation with the lamellar bone area, and their thickness had a random distribution for osteonal size classes. The circular, concentrical pattern was the more frequently observed, but spiral and crescent-moon-shaped lamellae were also documented. Selected osteons were examined by either SEM or SEM combined with polarized light microscopy allowing comparisons of corresponding sectors of the osteon. The bright bands observed with polarized light corresponded to the grooves observed in etched sections by SEM. The dark bands corresponded to the lamellar surface with the cut fibrils oriented approximately longitudinally along the central canal axis. However, lamellae with large and blurred bright bands could be observed, which did not correspond to a groove observed by SEM. These findings are in contrast with the assumption that all the fibril layers within a lamella are oriented along a constant and unchangeable angle. The different lamellar patterns may be explained by the synchronous or staggered recruitment and activation of osteoblasts committed to the osteon's completion. Anat Rec, 2012. © 2012 Wiley Periodicals, Inc.

The kinetics of osteogenic cells within secondary osteons have been examined within a 2-D model. The linear osteoblast density of the osteons and the osteocyte lacunae density were compared with other endosteal lamellar systems of different geometries. The cell density was significantly greater in the endosteal appositional zone and was always flatter than the central osteonal canals. Fully structured osteons compared with early structuring (cutting cones) did not show any significant differences in density. The osteoblast density may remain constant because some of them leave the row and become embedded within matrix. The overall shape of the Haversian system represented a geometrical restraint and it was thought to be related to osteoblast–osteocyte transformation. To test this hypothesis of an early differentiation and recruitment of the osteoblast pool which completes the lamellar structure of the osteon, the number and density of osteoblasts and osteocyte lacunae were evaluated. In the central canal area, the mean osteoblast linear density and the osteocyte lacunae planar density were not significantly different among sub-classes (with the exclusion of the osteocyte lacunae of the 300–1000 μm2 sub-class). The mean number of osteoblasts compared with osteocyte lacunae resulted in significantly higher numbers in the two sub-classes, no significant difference was seen in the two middle sub-classes with the larger canals, and there were significantly lower levels in the smallest central canal sub-class. The TUNEL technique was used to identify the morphological features of apoptosis within osteoblasts. It was found that apoptosis occurred during the late phase of osteon formation but not in osteocytes. This suggests a regulatory role of apoptosis in balancing the osteoblast–osteocyte equilibrium within secondary osteon development. The position of the osteocytic lacunae did not correlate with the lamellar pattern and the lacunae density in osteonal radial sectors was not significantly different. These findings support the hypothesis of an early differentiation of the osteoblast pool and the independence of the fibrillar lamellation from osteoblast–osteocyte transformation.

The frequency, structure, mode of formation and significance of sealed osteons remain unsettled. Sealed osteons have been reported as an unusual finding in the cortical bone of experimental animals: we extended the observation to human cortical bone studied with SEM. Tibial bone specimens from three patients who sustained a traumatic below-the-knee amputation were used in the study. The observed total mean density of osteons was 19.25/mm2 and the percentage of sealed and partially sealed osteons was 4.2% and 1.7% respectively. The material sealing the central canal showed an X-ray microanalysis spectrum with the same Ca/P ratio as the peripheral lamellae and a lower carbon signal. The morphology suggested a reactivation of bone apposition triggered by exclusion of the occluded canal from blood flow rather than a physiological evolution of the closing process of secondary osteons. This presupposes collapse and degeneration of the central vessel before the osteoblasts resting on the inner surface of the canal could start to lay down new bone matrix. This explanation is consistent with a dynamic model of intracortical blood flow. Anat Rec, 2011. © 2010 Wiley-Liss, Inc.

The intracortical canal/vessel systems of long bones are not yet completely understood in terms of their morphology and physiology, mainly because of the difficulty of injecting the small calibre vessels and cutting the calcified matrix. Here, we apply a novel method combining perfusion of the vessels and fracture of the cortical bone to enlighten the architecture of this system. The femurs of ten rabbits were perfused with a water-soluble dye (China ink) or alcoholic glycerol solution, and the fractured cortex specimens were then examined by scanning electron microscopy (SEM). The results document: (1) the fibrillar structure of the canal surfaces; (2) the perivascular environment with cellular components in different phases of incorporation within the bone matrix; (3) previously unreported filamentous structures on the outer surface of vessels, which could be interpreted as non-myelinic nerve fibres; (4) the inner organisation of the cutting cones. Although based exclusively on morphology, these observation have some relevance to increasing knowledge of bone circulation physiology in the cortical bone.

The vascular anatomy of the cortical bone and the canal system are highly correlated, and the former has an important bearing on shape and microscopic lamellar structure, as it is established in the progression of the remodelling process. The classical description of a longitudinal system of canals (Havers') connected by the transversal Volkmann's canals is the generally acknowledged model of the structural organization of the cortex. However, it is remarkably difficult to study the circulation inside the compact bone in detail owing to its hard, calcified matrix, and the methods thus far applied have represented either the bone morphology and the architecture of the canal system or the injected vessel network. In the present study, the intracortical vessel network was injected with black China ink and evidenced by transillumination of full-thickness, decalcified hemicortices. By making use of the depth of field of the microscope objective, the three-dimensional architecture of the network was highlighted and the morphometry of vessel size measurements and a classification of the network nodes according to the number of arms was made possible. These observations were integrated with data obtained by routine histology on decalcified sections relevant to the connections of the intracortical canal system with the outer environment, with regard to the direction of advancement of new canals and with regard to the mode of formation of the system nodes. The formation of the intracortical vessels network involved two processes: the incorporation of the periosteal network and osteonal remodelling, the latter occurring through the advancement of cutting cones followed by their own vascular loop and by concentric lamellar apposition. The two systems could be distinguished by the diameter of the vessels (the former were significantly larger) and by the network architecture (the former convoluted, and the latter longitudinally orientated and straight). Longitudinal vessels could form branches or create connections with the periosteal derived vessels that occasionally meet on the line of their advancement. They were observed entering from either inside the cortex from the metaphyses or from the endosteal surface of the marrow cavity. The combined observations from different methods of study documented a model of intracortical canal and vessel networks formed by two initially independent systems: one derived from the external, periosteal vessels, and one from metaphyseal and marrow vessels. Connections between the two were established with the advancing of cutting cones from the extremities of the diaphysis. Analysis of the system architecture and the modalities of its progressive organization suggested that the direction of advancement of a forming canal does not necessarily correspond to the final blood flow direction of its central vessel.

We show that Atomic Layer Deposition is a suitable coating technique to prevent metal diffusion from medical implants. The metal distribution in animal bone tissue with inserted bare and coated Co–Cr alloys was evaluated by means of micro X-ray fluorescence mapping. In the uncoated implant, the migration of Co and Cr particles from the bare alloy in the biological tissues is observed just after one month and the number of particles significantly increases after two months. In contrast, no metal diffusion was detected in the implant coated with TiO 2 . Instead, a gradient distribution of the metals was found, from the alloy surface going into the tissue. No significant change was detected after two months of aging. As expected, the thicker is the TiO 2 layer, the lower is the metal migration.

Bone aging was studied in an experimental model (rabbit femur) in three populations aged 0.5, 1.5, and 7.5 years. Cortical bone histology was compared with a data set from a 1.5-month-old population of an earlier published paper. From 0.5-year-old onward, the mean femur length did not increase further. Thereafter, the mean marrow area increased and the cortical area decreased significantly with aging. This was associated with a structural pattern transformation from plexiform to laminar and then Haversian-like type. The distal meta-epiphysis bone trabecular density of the oldest populations also was significantly lower in specific regions of interest (ROI). Percentage sealed primary vascular canals in laminar bone significantly increased with aging without variation of percentage sealed secondary osteons. Remodeling rate reflected by the density of cutting cones did not significantly change among the age populations. These data suggest that laminar bone vascular pattern is more functional in the fast diaphyseal expansion but not much streamlined with the renewal of blood flow during secondary remodeling. Bone aging was characterized by: 1) secondary remodeling subendosteally; 2) increment of sealed primary vascular canals number; 3) increased calcium content of the cortex; 4) cortical and trabecular bone mass loss in specific ROIs. Taken together, the present data may give a morphological and morphometric basis to perform comparative studies on experimental models of osteoporosis in the rabbit. J. Morphol. 276:733–747, 2015. © 2015 Wiley Periodicals, Inc.

Polyethylene (PE) wear is a focal issue in joint replacement, so it is essential to understand how it takes place in vivo. PE wear is a multifactorial process with a complex interaction of variables related to the materials used, the mechanical conditions, operative procedures for implantation, activity of the patient and use of prosthesis. We retrieved 65 PE inserts with the respective femoral and tibial components (50 inserts of total knee prostheses and 15 monocompartmental prostheses) from first revision surgeries. The average age of the patients was 68.3 years; the average time to revision was 41.5 months. Macroscopic observation considered the grade, topography and type of wear. Degradation was also studied with scanning electron microscopy. No direct relationship was found between the level of wear and the survival of total or monocompartmental knee prostheses. The duration of 11 (22%) total prosthetic inserts with grade 2 wear was 42.6 months vs. 51.5 months of 17 (34%) total inserts with grade 1 wear. However, study of the relationship between wear grade and wear topography in total inserts suggested that there was a significant connection. In fact, the 22 (44%) prostheses with central and symmetrical wear never showed wear greater than grade 1. Instead, of the remaining 28 prostheses (56%) without central and symmetric wear, only 3 had grade 0 wear. Finally, considering the relationship between wear grade and type of wear, all 18 inserts (27.7%) with grade 2 or 3 wear had 100% delamination. In conclusion, this study suggests that the correct positioning of the prosthetic components, besides its quality, is an important cause of polyethylene wear.

Documentation through X-ray morphometry and histology of the steady phenotype expressed by FGFR3 gene mutation and interpolation of mechanical factors on spine and long bones dysmorphism. Long bones and spine of eight thanatophoric dysplasia and three age-matched controls without skeletal dysplasia were studied after pregnancy termination between the 18th and the 22nd week with X-ray morphometry, histology, and molecular analysis. Statistical analysis with comparison between TD cases and controls and intraobserver/interobserver variation were applied to X-ray morphometric data. Generalized shortening of long bones was observed in TD. A variable distribution of axial deformities was correlated with chondrocyte proliferation inhibition, defective seriate cell columns organization, and final formation of the primary metaphyseal trabeculae. The periosteal longitudinal growth was not equally inhibited, so that decoupling with the cartilage growth pattern produced the typical lateral spurs around the metaphyseal growth plates. In spine, platyspondyly was due to a reduced height of the vertebral body anterior ossification center, while its enlargement in the transversal plane was not restricted. The peculiar radiographic and histopathological features of TD bones support the hypothesis of interpolation of mechanical factors with FGFR3 gene mutations. The correlated observations of X-ray morphometry, histopathology, and gene analysis prompted the following diagnostic workup for TD: (1) prenatal sonography suspicion of skeletal dysplasia; (2) post-mortem X-ray morphometry for provisional diagnosis; (3) confirmation by genetic tests (hot-spot exons 7, 10, 15, and 19 analysis with 80–90 % sensibility); (4) in negative cases if histopathology confirms TD diagnosis, research of rare mutations through sequential analysis of FGFR3 gene.

Numerous systems of osteosynthesis of olecranon fractures are avaible. The numero-ulnar articulation badly bears the smallest incongruence and easily faces rigidity. Therefore the aim of therapy is the anatomical and permanent reconstruction of the articular surface of the olecranon, in order to mobilize the elbow as soon as possible. We analyzed 85 olecranon fractures, classified according to Merle d'Aubigne and treated with two different surgical techniques (intramedullart screw and AO tension band wiring technique). The results were compared at an average follow-up of 88 months. Excellent clinical results were obtained in 74,12%, good in 29.00%, sufficient 3,53%, insufficient in 2,35% of patients, with a mean of recovery in 2 months and only 2 cases of pseudoarthrosis (with dynamic procedure). Objective results were similar for two techniques adopted, whereas the subjective results in patients with screw. The screw is more suitable in medio-olecranic and basis-olecranic fractures, while the dynamic procedure gives better results in fractures of the apex of olecranon and comminute fracture. Tolerance of the intramedullary screw has generally proved to be better (65% of cases), while the dynamic assembling has been removed in most cases after fracture consolidation. Periarticular calcifications correlated with the type of fracture (e. g. comminuted, associated) rather than with the means of synthesis employed.

The shape and structure of bones is a topic that has been studied for a long time by morphologists and biologists with the goal of explaining the laws governing their development, aging and pathology. The osteonal architecture of tibial and femoral mid-diaphyses was examined morphometrically with scanning electron microscopy in four healthy young male subjects. In transverse sections of the mid-diaphysis, the total area of the anterior, posterior, lateral and medial cortex sectors was measured and analysed for osteonal parameters including osteon number and density, osteon total and bone area and vascular space area. Osteons were grouped into four classes including cutting heads (A), transversely cut osteons (B), longitudinally cut osteons (C) and sealed osteons (D). The morphometric parameters were compared between the inner (endosteal) and outer (periosteal) half of the cortex. Of 5927 examined osteons, 24.4% cutting heads, 71.1% transversely cut osteons, 2.3% longitudinally cut osteons and 2.2% sealed osteons were found. The interosteonic bone (measured as the area in a lamellar system that has lost contact with its own central canal) corresponded to 51.2% of the endosteal and 52.4% of the periosteal half-cortex. The mean number of class A cutting heads and class B osteons was significantly higher in the periosteal than in the endosteal half-cortex (P

A casting technique with methyl-methacrylate (MMA) was applied to the study of the osteon lacunar-canalicular network of human and rabbit cortical bone. The MMA monomer infiltration inside the vascular canals and from these into the lacunar-canalicular system was driven by capillarity, helped by evaporation and the resulting negative pressure in a system of small pipes. There was uniform, centrifugal penetration of the resin inside some osteons, but this was limited to a depth of four to five layers of lacunae. Moreover, not all of the osteon population was infiltrated. This failure can be the result of one of two factors: the incomplete removal of organic debris from the canal and canalicular systems, and lack of drainage at the osteon external border. These data suggest that each secondary osteon is a closed system with a peripheral barrier (represented by the reversal line). As the resin advances into the osteon, the air contained inside the canalicula is compressed and its pressure increases until infiltration is stopped. The casts gave a reliable visualization of the lacunar shape, position and connections between the lacunae without the need for manipulations such as cutting or sawing. Two systems of canalicula could be distinguished, the equatorial, which connected the lacunae (therefore the osteocytes) lying on the same concentric level, and the radial, which established connections between different levels. The equatorial canalicula radiated from the lacunar border forming ramifications on a planar surface around the lacuna, whereas the radial canalicula had a predominantly straight direction perpendicular to the equatorial plane. The mean length of the radial canalicula was 40.12 ± 10.26 μm in rabbits and 38.4 ± 7.35 μm in human osteons; their mean diameter was 174.4 ± 71.12 nm and 195.7 ± 79.58 nm, respectively. The mean equatorial canalicula diameter was 237 ± 66.04 nm in rabbit and 249.7 ± 73.78 nm in human bones, both significantly larger (P < 0.001) than the radial. There were no significant differences between the two species. The lacunar surface measured on the equatorial plane was higher in rabbit than in man, but the difference was not statistically significant. The cast of the lacunar-canalicular network obtained with the reported technique allows a direct, 3-D representation of the system architecture and illustrates how the connections between osteocytes are organized. The comparison with models derived by the assumption of the role of hydraulic conductance and other mechanistic functions provides descriptive, morphological data to the ongoing discussion on the Haversian system biology.

Two cases of pyomyositis in babies, one in the ilio-psoas muscle, the other in the quadriceps, are reported. The presenting sign was a painful hip in both, suggesting osteomyelitis of the proximal femur or arthritis. Pyomyositis is unusual in western countries and in healthy, well-nourished children, but these cases suggest considering this pathology in the differential diagnosis of a painful hip in babies. In the first case an upper respiratory tract infection was followed by haematogenous spread to the ilio-psoas muscle; in the second an injection of vaccine in the quadriceps played a determinant role. The latter case was complicated by secondary osteomyelitis; a metaphyseal radiolucent band just below the growth plate appeared in the early stage together with metaphyseal osteopenia and was interpreted as a consequence of altered blood flow in the metaphysis, not specific osteomyelitis, but indicative of some disturbance of the growth plate-metaphyseal complex.

I report a case of bilateral spontaneous atraumatic anterior subluxation of the sternoclavicular joint in a 19-year-old woman without any known underlying pathology. There was no history of injury. The patient was treated conservatively. One year later the patient was asymptomatic and had returned to her usual activities.

The kinetics of osteogenic cells within secondary osteons have been examined within a 2-D model. The linear osteoblast density of the osteons and the osteocyte lacunae density were compared with other endosteal lamellar systems of different geometries. The cell density was significantly greater in the endosteal appositional zone and was always flatter than the central osteonal canals. Fully structured osteons compared with early structuring (cutting cones) did not show any significant differences in density. The osteoblast density may remain constant because some of them leave the row and become embedded within matrix. The overall shape of the Haversian system represented a geometrical restraint and it was thought to be related to osteoblast-osteocyte transformation. To test this hypothesis of an early differentiation and recruitment of the osteoblast pool which completes the lamellar structure of the osteon, the number and density of osteoblasts and osteocyte lacunae were evaluated. In the central canal area, the mean osteoblast linear density and the osteocyte lacunae planar density were not significantly different among sub-classes (with the exclusion of the osteocyte lacunae of the 300-1000 μm(2) sub-class). The mean number of osteoblasts compared with osteocyte lacunae resulted in significantly higher numbers in the two sub-classes, no significant difference was seen in the two middle sub-classes with the larger canals, and there were significantly lower levels in the smallest central canal sub-class. The TUNEL technique was used to identify the morphological features of apoptosis within osteoblasts. It was found that apoptosis occurred during the late phase of osteon formation but not in osteocytes. This suggests a regulatory role of apoptosis in balancing the osteoblast-osteocyte equilibrium within secondary osteon development. The position of the osteocytic lacunae did not correlate with the lamellar pattern and the lacunae density in osteonal radial sectors was not significantly different. These findings support the hypothesis of an early differentiation of the osteoblast pool and the independence of the fibrillar lamellation from osteoblast-osteocyte transformation.

The lacunar-canalicular system in human secondary osteons was examined by two complementary techniques: light microscopy analysis of undecalcified thick sections and the SEM cortex-fractured surface technique. Unlike the earlier definitions of 'osteoblastic domain' presented as the matrix volume produced by osteoblasts in the process of osteon infilling, this study measured the domain by the length of osteoblast dendritic processes. The domain extension was defined along radial vectors advancing from the reversal line towards the central canal. According to their lengths, domains were divided into three classes: peripheral, intermediate and internal. The mean length of peripheral domains was significantly shorter than those of the intermediate and internal domains. This suggests that the infilling process is modulated by an initial preparatory phase characterised by osteoblast adhesion to the wall of the cutting cone, and a limited matrix synthesis, followed by a regular matrix volume apposition organised in concentric layers. In addition to the radial canaliculae arranged along converging vectors in planes perpendicular to the central canal, we distinguished a further class of canaliculae, the equatorial canaliculae originating from the major perimeter of the lacuna and spreading out radially in the plane of the same lacuna (therefore, perpendicularly to the radial canaliculae). The whole lacunar-canalicular network was structured as a closed system around the vascular axis of the central canal with very few canaliculae crossing the reversal line and connecting the neighbouring osteons. These anatomical observations contribute to our knowledge of lacunar-canalicular system development.

Summary The proliferative impulse of the growth plate cartilage and related structures and its effect on the dimensions of long bones are well documented. The modulation of shape, however, is less known, and in general, it is referred to the coupled resorption/apposition process of bone modelling. A morphometric study was carried out on rabbit tibiae comparing size increments and shape changes in relation to age. Utilizing measurements made using dried bones, radiography and computerized tomography, it was possible to perform a three-dimensional analysis of shape modulation occurring during a period of growth extending from 3 months to 1 year of age. The dynamics of the shape changes related to growth were studied with a fluorescent tetracycline labelling. This enabled correlation of shape modulation with the 3-D distribution of apposition and resorption. The current thinking behind the influences and mechanical forces affecting bone architecture was discussed in the light of these findings. Several factors play a role in the structural organization of the human and upper vertebrates’ skeleton, whose shape is genetically determined in the complex process usually referred as ‘modelling’. This does not conflict with the existing evidence of remodelling being influenced by mechanical stimuli, but the unsolved question remains how physical forces (strains) act on the biological substrate of cartilage and bone cells.

To compare the outcomes of two different surgical treatments for the management of isolated closed Mason Type 2 radial head fractures.Retrospective study. The Student t test and McPearson chi-square test were used to evaluate whether there was a significance difference between the groups.Fifty-nine patients with isolated Mason Type 2 radial head fractures.Twenty-four patients treated with radial head excision (Group I) and 35 treated with open reduction and internal fixation (Group II).Clinical outcomes were assessed using the Broberg and Morrey functional rating scores and the Disabilities of the Arm, Shoulder and Hand (DASH) questionnaire. Orthogonal radiographs were performed on both the elbow and the wrist; these were assessed for the presence of arthritis, heterotopic ossification, and the degree of proximal radial migration.The length of postoperative follow-up was 157 ± 61.84 months (Group I) and 125 ± 39.09 months (Group II). The Broberg and Morrey functional rating score was 86.21 ± 6.10 points and 95.09 ± 4.78 points, respectively. The DASH score was 21.82 ± 6.01 points and 2.81 ± 2.73 points, respectively. Radiologically moderate or severe osteoarthritis was present in the elbows of nine patients in Group I and only two patients in Group II.Patients with isolated Mason Type 2 radial head fractures treated by open reduction and internal fixation (Group II) had less residual pain, greater range of motion, and better strength than patients treated by radial head excision (Group I). Additionally, Group II had a lower incidence of severe posttraumatic arthritis, which contributed to improved DASH and Broberg and Morrey functional scores. These results support open reduction and internal fixation as the treatment of choice for these fractures.Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.

The anchorage of cementless total hip arthroplasty relies on a direct bone to implant bonding. Several factors including material properties and surface treatment determine the interfacial response of the host bone to a foreign material. Another factor that must be taken into account is the bone remodeling after the prosthesis introduction. Considering this bone remodeling, in the last few years the possibility of using anti-osteoporotic drugs has been introduced as a supplementary and integrated treatment in total hip replacement.

Autoptic samples of human bones (from 8 weeks of gestation to 12 years of age) and a second group of serial, skeletal x-rays (required for pathologies not related to bone dysplasia in children from 4 months to 17 years of age) provided the material for the analysis of the physes normal growth mechanism presented in this review. Before the appearance of the ossification centers epiphyseal growth rests exclusively on chondrocytes proliferation (interstitial growth), without any detectable differentiated cellular organization. When endochondral ossification starts a defined spatial disposition of chondrocytes and a corresponding organization of the intercellular matrix is set up, so that it is possible to identify a growth vector corresponding to the columns of piled chondrocytes with direction from hypertrophic toward the proliferative cell layers. The complexity of the tubular bones growth process is well represented by the spatial arrangement of the growth vectors. In the late epiphyseal growth another mechanism is active in addition to endochondral ossification, namely, articular cartilage interstitial growth and subchondral remodelling. The knowledge of the normal mode of organization of the physis and its temporal sequence can help to better understand of the deviaton from the normal development of metaphyseal and epiphyseal dysplasias.

Four commercial hydroxyapatites (both natural and synthetic) were tested to assess transformations of the chemical and crystalline structure following variation of temperature from 20 to 1600 degrees C. The thermal behaviour of hydroxyapatite is relevant for biomedical applications such as plasma spraying of metallic implants. Thermogravimetric analysis showed a weight loss from each hydroxyapatite specimen, due to a release of structural H2O molecules; all the specimens up to 1300 degrees C were made of crystalline hydroxyapatite, determined by X-ray diffraction; at 1470 degrees C they were made of both hydroxyapatite and calcium phosphate, but at 1570 degrees C of calcium phosphate exclusively. The diffractograms of the hydroxyapatite coatings showed the same peaks as the original powders, so at the chosen plasma-spray procedure level no new phases were formed. The peak height was nevertheless lower in the plasma-sprayed hydroxyapatites for all interplanar spacing values, which indicated a lower degree of crystallinity, associated with a random structure derived from an alteration to the original crystalline network.

The intracortical vessel system of the rabbit femur has been studied after perfusion of the vascular tree with a water solution of dye (China ink) with multiplanar analysis. This method utilizes the full depth of field of the microscope objectives focusing different planes of the thick cortex. The microscopic observation even if restricted to a limited volume of cortex allowed to differentiate true 3-D nodes (54.5%) from the superimposition of vessels lying on different planes. The network model with elongated meshes preferentially oriented along the longitudinal axis of the diaphysis in his static configuration is not very different from the vascular anatomy depicted in the 2-D traditional models; however, the semi-quantitative morphometric analysis applied to the former supported the notion of a multidirectional microvascular network allowing change of flow according to the functional requirements. Other peculiar aspects not previously reported were cutting cone loops, blind-end and short-radius-bent vessels, and button-holes figures. The network design and node distribution were consistent with the straight trajectory of the secondary remodeling, with the proximal-to-distal and distal-to-proximal advancement directions of the cutting cones and with two main modes of node formation, namely bifurcation of the cutting cone and interception with pre-existing canals. The general organization of the network and its uninterrupted transformation during bone modeling and remodeling suggested a substantial plasticity of the intracortical vascular system capable to adapt itself to the changeable haemodynamic conditions.

The collagen architecture of secondary osteons was studied with scanning electron microscopy (SEM) employing the fractured cortex technique and osmic maceration. Fibrillar orientation and the change in their direction in sequential lamellae was documented where lamellar formation was ongoing, as well as in resorption pits where osteoclasts had exposed the collagen organisation of the underlying layers. Applying an adaptive stereo matching technique, the mean thickness of matrix layers removed by osteoclasts was 1.36 ± 0.45 μm. It was also documented that osteoclasts do not attack the cellular membrane of the exposed osteocytes. The mean linear osteoblast density in fractured hemicanals was assessed with SEM and no significant differences were observed comparing larger with smaller central canal osteons. These findings suggested a balance between the differentiated osteoblasts that have aligned on the surface of the cutting cone and those that are transformed into osteocytes, because the canal surface is progressively reduced as the lamellar apposition advances.

A patient with a total hip replacement developed optic, acoustic and peripheral neuropathy from metal ions intoxication, due to the wear products released from the prosthesis. Subsequently the kinetics of the metal ions was studied.Massive wear and acute intoxication allowed a study of the metal ions kinetics and of EDTA treatment.Plasma and other organic fluids were saturated by each of the metal ions released from the exposed surface according to the solubility of each ion; a larger fraction of Co ions was bound within red cells, while the plasmatic fraction appeared more movable. In a patient with a prosthesis subjected to wear, the ions released are from the prosthetic and from the debris surface (spread in the body). The latter is a function of the number and size of particles.Revision of the prosthesis from the point of view of the metal ions kinetics corresponded to a reduction of the releasing surface because of debris washed out by irrigation and tissue excision; however, the metal particles spread by lymphatic circulation continued to release ions even though the source of wear had been removed. Early diagnosis of high metal wear can be ascertained with mass spectrometry and after revision high levels of metal ions can only be reduced with repeated chelating treatment. It is preferable not to revise fractured ceramic components with a polyethylene-metal articulation.

The osteonal pattern of cortical bone is gradually built around the intracortical vessels by the progression of the cutting cones (secondary remodelling); therefore, the central canal size can be used as index of the remodelling activity. An experimental model in the rabbit femur was used to investigate, through central canal morphometry and frequency distribution analysis, the remodelling activity, comparing the middle of the diaphysis (mid-shaft) with the extremity (distal-shaft) and at the same level sectors and layers of the cortex in transversal sections. The study documented a higher density of canals in the mid-shaft than in the distal-shaft and a higher remodelling in the distal-shaft. There were no significant differences between dorsal, ventral, medial and lateral sectors at both mid-shaft and distal-shaft levels, while the number of canals was higher in the sub-periosteal layers than in the sub-endosteal. A lower threshold of 40 microm(2) was observed in the central canal area. Sealed osteons in the midshaft were 22.43% of the total number of osteons of the central canal area between 40 and 200 microm(2) and 0.44% of those of the distal-shaft. Micro-CT allowed a 3D reconstruction of the vascular canal system, which confirmed the branched network pattern rather than the trim architecture of the traditional representation. Some aspects like the lower threshold of the central canal size and the sealed osteons documented the plasticity of the system and its capacity for adaptation to changes in the haemodynamic conditions.

Cortex fractured surface and graded osmic maceration techniques were used to study the secretory activity of osteoblasts, the transformation of osteoblast to osteocytes, and the structural organization of the matrix around the cells with scanning electron microscopy (SEM). A specialized membrane differentiation at the base of the cell was observed with finger-like, flattened processes which formed a diffuse meshwork. These findings suggested that this membrane differentiation below the cells had not only functioned in transporting collagen through the membrane but also in orienting the fibrils once assembled. Thin ramifications arose from the large and flat membrane foldings oriented perpendicular to the plane of the osteoblasts. This meshwork of fine filaments could not be visualized with SEM because they were obscured within the matrix substance. Their 3-D structure, however, should be similar to the canalicular system. The meshwork of large, flattened processes was no more evident in the cells which had completed their transformation into osteocytes.

In September, 2007, a 58-year-old woman was admitted to our neurology department because of progressive visual and hearing loss which had started 9 months earlier. She had type 2 diabetes and hypertension, both of which were adequately controlled by medication. In 2001, she had a left hip arthroplasty which was revised in October, 2006 because of rupture of the ceramic head. 3 weeks before admission, investigations showed a mild hypothyroidism of unknown origin, which was treated with levothyroxine sodium. The neurological examination at admission showed impairment of cranial nerves II and VIII bilaterally and mild distal sensory-motor dis turbances. Laboratory investigations ruled out haema tological, infectious, neoplastic, metabolic, and immuno logical diseases. Concomitantly our patient underwent various investigations including brain MRI (showing hyperintensity of optic nerves and tracts), electromyography (showing mild lower limb nerve amplitude reduction), and acoustic and visual evoked potentials (positive for bilateral absence of brainstem acoustic responses and irregular cortical visual responses). A working diagnosis of axonal multi-neuropathy caused by a presumed immune-mediated vasculitis was made and the patient was given prednisone (50 mg/day) for 2 months with little improvement. By December, 2007, our patient became completely blind, severely deaf, and wheel-chair bound because of lower limb hyposthenia. Tests for immune-mediated process remained negative and the case was referred to toxicology for further investigation. Unexpectedly, raised concentrations of cobalt and chromium were found in diff erent biological samples (cobalt: 24 h urine collection 1187 μg/L [0·1–1·5], blood 549 μg/L [0·05–2·7], plasma 90 μg/L [0·1–0·6], CSF 11·4 μg [0·05–0·15]; chromium: 24 h urine collection 510 μg/L [0·05–2·2], blood 54 μg/L [0·1–0·5], plasma 210 μg/L [0·1–0·5], CSF 4·4 μg/L [0·01–0·2]). Cobalt-chromium poisoning due to metal wear debris from her hip prosthesis was proposed, although radiology, including CT, showed no sign of prosthesis loosening. In February, 2008, several metal ion chelating treatments were given with edetic acid. Although metal ion concentrations decreased, neurological improve ment was negligible. Therefore, in April, 2008, resection arthroplasty was done. During surgery infi ltration of the peri-prosthesic tissue by metallic debris was evident (fi gure A); analysis of peri-prosthetic fl uids showed high concentrations of cobalt and chromium, and the removed prosthesis showed wear of the head and neck (fi gure B), supporting the hypothesis of endogenous cobalt-chromium poisoning. During the following 8 months the patient showed progressive improvement, although vision only partially improved. Metal ion concentrations decreased but remained higher than reference values at the last follow-up in November, 2008. The role of cobalt or cobalt-chromium on human tissues has not been defi nitively established. Cobalt can produce various toxicological eff ects including local respiratory symptoms due to inhalation of cobalt containing dusts, and systemic eff ects (thyroiditis, cardiomyopathy, erythropoiesis). Neuro logical toxicity, mainly optic atrophy, nerve deafness, and limb paraesthesia, has been occasionally reported in association with exogenous exposure. Neuro logical toxicity as a reult of endogenous exposure (mainly associated with metal prostheses) has been described. Cobalt can induce a hypoxia-like eff ect, possibly targeting mitochondria; of note, our patient’s symptoms were similar to those observed in some mitochondrial cytopathies. Total hip replacement and hip resurfacing arthroplasty are widely used therapeutic procedures; longer-term follow-up would be necessary to evaluate adverse chronic systemic eff ects due to prolonged exposure to high serum cobalt concentrations. In addition to orthopaedic evaluation, careful neurological and toxicological examinations are recommended whenever a patient with a metallic prosthesis complains of visual loss, hearing disturbance, limb weakness, numbness, or paraesthesia, even in the absence of local osteoarticular symptoms.

The current model of compact bone is that of a system of longitudinal (Haversian) canals connected by transverse (Volkmann's) canals. Models based on histology or microcomputed tomography lack the morphologic detail and sense of temporal development provided by direct observation. Using direct scanning electron microscopy observation, we studied the bone surface and structure of the intracortical canal system in paired fractured surfaces in rabbit femurs, examining density of canal openings on periosteal and endosteal surfaces, internal network nodes and canal sizes, and collagen lining of the inner canal system. The blood supply of the diaphyseal compact bone entered the cortex through the canal openings on the endosteal and periosteal surfaces, with different morphologic features in the midshaft and distal shaft; their density was higher on endosteal than on periosteal surfaces in the midshaft but with no major differences among subregions. The circumference measurements along Haversian canals documented a steady reduction behind the head of the cutting cone but rather random variations as the distance from the head increased. These observations suggested discontinuous development and variable lamellar apposition rate of osteons in different segments of their trajectory. The frequent branching and types of network nodes suggested substantial osteonal plasticity and supported the model of a network organization. The collagen fibers of the canal wall were organized in intertwined, longitudinally oriented bundles with 0.1- to 0.5-mum holes connecting the canal lumen with the osteocyte canalicular system.

A bone-implant interface is influenced by several factors, such as chemical composition of the implanted material, stress transfer at the interface and the biology of the living model. Histological and vascular tree injection methods were used to assess the response to intramedullary pins, cast in different metallic alloys and implanted in the medullary canal of rat femora. A remarkable endosteal reaction, with an absence of major damage of the medullary vascular supply, was observed. Remodelling of this woven bone led to the formation of a lamellar bony shell around the pins, while in sham operations (pin inserted and soon removed) the former was completely resorbed in about 20 days. Mechanical trauma seems to be sufficient to evoke the response of the endosteum, which does not differ from that observed in the normal repair process of diaphyseal fractures. Bone grew very close to the metal surface, but a thin connective membrane or a layer of amorphous material was present at the interface with stainless steel, Co-Cr and titanium pins. Histology of the interface suggests that the implants were not mechanically neutral; these observations may be correlated to the thin layer of bone observed around hip cementless prosthetic stems in patients.