Capacitively coupled electrical stimulation of rat chondroepiphysis explants: A histomorphometric analysis.

The growth plate is a cartilaginous layer present from the gestation period until the end of puberty where it ossifies joining diaphysis and epiphysis. During this period several endocrine, autocrine, and paracrine processes within the growth plate are carried out by chondrocytes; therefore, a disruption in cellular functions may lead to pathologies affecting bone development. It is known that electric fields impact the growth plate; however, parameters such as stimulation time and electric field intensity are not well documented. Accordingly, this study presents a histomorphometrical framework to assess the effect of electric fields on chondroepiphysis explants. Bones were stimulated with 3.5 and 7 mV/cm, and for each electric field two exposure times were tested for 30 days (30 min and 1 h). Results evidenced that electric fields increased the hypertrophic zones compared with controls. In addition, a stimulation of 3.5 mV/cm applied for 1 h preserved the columnar cell density and its orientation. Moreover, a pre-hypertrophy differentiation in the center of the chondroepiphysis was observed when explants were stimulated during 1 h with both electric fields. These findings allow the understanding of the effect of electrical stimulation over growth plate organization and how the stimulation modifies chondrocytes morphophysiology.

On the influence of surface coating on tissue biomechanics - effects on rat bones under routine conditions with implications for image-based deformation detection.

BACKGROUND: Biomechanical testing using image-based deformation detection techniques such as digital image correlation (DIC) offer optical contactless methods for strain and displacement measurements of biological tissues. However, given the need of most samples to be speckled for image correlation using sprays, chemical alterations with impact on tissue mechanicals may result. The aim of this study was to assess the impact of such surface coating on the mechanical properties of rat bones, under routine laboratory conditions including multiple freeze-thaw cycles. METHODS: Two groups of rat bones, highly-uniform and mixed-effects, were assigned to six subgroups consisting of three types of surface coating (uncoated, commercially-available water- and solvent-based sprays) and two types of bone conditions (periosteum attached and removed). The mixed-effects group had undergone an additional freeze-thaw cycle at - 20 degrees. All bones underwent a three-point bending test ranging until material failure. RESULTS: Coating resulted in similar and non-significantly different mechanical properties of rat bones, indicated by elastic moduli, maximum force and bending stress. Scanning electron microscopy showed more pronounced mechanical alterations related to the additional freeze-thaw cycle, with fewer cracks being present in a bone from the highly-uniform group. CONCLUSIONS: This study has concluded that surface coating with water- or solvent-based sprays for enhancing image correlation for DIC and having an additional freeze-thaw cycle do not significantly alter mechanical properties of rat bones. Therefore, this method may be recommended as an effective way of obtaining a speckled pattern.

A method for measuring the three-dimensional orientation of cortical canals with implications for comparative analysis of bone microstructure in vertebrates.

The orientation of vascular canals in cortical bone can reveal information about the growth rate and loading environment of a bone. For example, in birds it has been proposed that a high proportion of circumferential canals (a laminar cortex) is related to fast growth or torsional loading related to active flight. In this paper we present a method to measure the three dimensional (3D) orientation of vascular canals. Image data are obtained from micro-CT and two angles are measured: phi, determining how longitudinal a canal is; and theta, determining whether a canal is radial or circumferential. This method can measure the orientation of each canal contained in the scanned images. Here we demonstrate the approach on two samples - a rat tibia and a hawk humerus. This method offers a direct (3D) method for quantifying features of canal orientation, such as the degree of laminarity, and can be applied easily and non-destructively to multiple species and bones. The growth and development of the cortical canal network and its impact on factors such as bone strength and bone quality remains relatively unexplored. Our method provides a new tool to examine the impact of the orientation of cortical bone canals on bone and explore the origins of cortical canals formed during modelling and remodeling. This method has applications in comparative bone biology, small animal models, and human bone studies.

Bone structural similarity score: a multiparametric tool to match properties of biomimetic bone substitutes with their target tissues.

BACKGROUND: One of the hardest tasks in developing or selecting grafts for bone substitution surgery or tissue engineering is to match the structural and mechanical properties of tissue at the recipient site, because of the large variability of tissue properties with anatomical site, sex, age and health conditions of the patient undergoing implantation. We investigated the feasibility of defining a quantitative bone structural similarity score based on differences in the structural properties of synthetic grafts and bone tissue. METHODS: Two biocompatible hydroxyapatite porous scaffolds with different nominal pore sizes were compared with trabecular bone tissues from equine humerus and femur. Images of samples' structures were acquired by high-resolution micro-computed tomography and analyzed to estimate porosity, pore size distribution and interconnectivity, specific surface area, connectivity density and degree of anisotropy. Young's modulus and stress at break were measured by compression tests. Structural similarity distances between sample pairs were defined based on scaled and weighted differences of the measured properties. Their feasibility was investigated for scoring structural similarity between considered scaffolds or bone tissues. RESULTS: Manhattan distances and Quadrance generally showed sound and consistent similarities between sample pairs, more clearly than simple statistical comparison and with discriminating capacity similar to image-based scores to assess progression of pathologies affecting bone structure. CONCLUSIONS: The results suggest that a quantitative and objective bone structural similarity score may be defined to help biomaterials scientists fabricate, and surgeons select, the graft or scaffold best mimicking the structure of a given bone tissue.

Stochastic interdigitation as a toughening mechanism at the interface between tendon and bone.

Reattachment and healing of tendon to bone poses a persistent clinical challenge and often results in poor outcomes, in part because the mechanisms that imbue the uninjured tendon-to-bone attachment with toughness are not known. One feature of typical tendon-to-bone surgical repairs is direct attachment of tendon to smooth bone. The native tendon-to-bone attachment, however, presents a rough mineralized interface that might serve an important role in stress transfer between tendon and bone. In this study, we examined the effects of interfacial roughness and interdigital stochasticity on the strength and toughness of a bimaterial interface. Closed form linear approximations of the amplification of stresses at the rough interface were derived and applied in a two-dimensional unit-cell model. Results demonstrated that roughness may serve to increase the toughness of the tendon-to-bone insertion site at the expense of its strength. Results further suggested that the natural tendon-to-bone attachment presents roughness for which the gain in toughness outweighs the loss in strength. More generally, our results suggest a pathway for stochasticity to improve surgical reattachment strategies and structural engineering attachments.

The cn/cn dwarf mouse. Histomorphometric, ultrastructural, and radiographic study in mutants corresponding to human acromesomelic dysplasia Maroteaux type (AMDM).

BACKGROUND: The cn/cn dwarf mouse is caused by a loss-of-function mutation in the natriuretic peptide receptor 2 (NPR-2) gene which helps positively regulate endochondral longitudinal bone growth. The gene mutation corresponds to that in the human skeletal dysplasia Acromesomelic Dysplasia Maroteaux type (AMDM). This study assesses histomorphometric, ultrastructural and radiographic correlates of the growth abnormality. METHODS: Ten litters of cn/cn and cn/+littermates at ages ranging from 2.5 to 6.5 weeks were studied by skeletal radiographs, histomorphometry and physeal ultrastructure. Skeletal radiographs were done on 2 cn/cn and 2 cn/+littermates at 5 weeks of age. Humeral, femoral, and tibial lengths were measured from 34 intact bones (17 cn/cn, 17 cn/+) at 2.5 to 6.5 weeks. Growth plate histomorphometry in 50 bones (26 cn/cn and 24 cn/+) determined the hypertrophic zone/entire physeal cartilage ratios in 204 sections (87 cn/+, 117 cn/cn) at 3 time periods (2.5-3, 4-4.5, and 6-6.5 weeks). Electron microscopy assessed 6 cn/cn and 6 cn/+age and site-matched physeal cartilage. RESULTS: Cn/cn mice were two thirds the size of the cn/+. Cn/cn bones were normal in shape or only minimally deformed except for the radius with mid-diaphyseal bowing. Length ratios of cn/cn humeri, femurs, and tibias were a mean of 0.65 (± 0.03, n = 34, 17 ratios) compared to cn/+bones. The main physeal abnormality was a markedly shortened hypertrophic zone with the ratio of hypertrophic zone to entire physis 0.17 (± 0.063) in the cn/cn and 0.30 (± 0.052) in the cn/+mice. Ratio assessments were similar comparing humeral, femoral, and tibial growth plates as were ratios from each of the 3 time periods. Ultrastructural assessments from the resting zone to the lower hypertrophic zone-metaphyseal junction showed no specific individual cell abnormalities in cn/cn compared to cn/+physes. CONCLUSIONS: The disorder causes a shortened physeal hypertrophic zone but normal ultrastructure of cn/cn chondrocytes points to abnormality primarily affecting the hypertrophic zone rather than a structural cell or matrix synthesis problem.

Bone ablation without thermal or acoustic mechanical injury via a novel picosecond infrared laser (PIRL).

BACKGROUND AND OBJECTIVE: A precise means to cut bone without significant thermal or mechanical injury has thus far remained elusive. A novel non-ionizing ultrafast pulsed picosecond infrared laser (PIRL) may provide the solution. Tissue ablation with the PIRL occurs via a photothermal process with thermal and stress confinement, resulting in efficient material ejection greatly enhanced through front surface spallation photomechanical effects. By comparison, the Er:YAG laser (EYL) ablates via photothermal and cavitation-induced photomechanical effects without thermal or acoustic confinement, leading to significant collateral tissue injury. This study compared PIRL and EYL bone ablation by infrared thermography (IRT), environmental scanning electron microscopy (ESEM), and histology. STUDY DESIGN: Prospective, comparative, ex vivo animal model. SETTING: Optics laboratory. SUBJECTS AND METHODS: Ten circular area defects were ablated in ex vivo chicken humeral cortex using PIRL and EYL at similar average power (~70 mW) under IRT. Following fixation, ESEM and undecalcified light microscopy images were obtained and examined for signs of cellular injury. RESULTS: Peak rise in surface temperature was negligible and lower for PIRL (1.56 °C; 95% CI, 0.762-2.366) compared to EYL ablation (12.99 °C; 95% CI, 12.189-13.792) (P < .001). ESEM and light microscopy demonstrated preserved cortical microstructure following PIRL ablation in contrast to diffuse thermal injury seen with EYL ablation. Microfractures were not observed. CONCLUSION: Ablation of cortical bone using the PIRL generates negligible and significantly less heat than EYL ablation while preserving cortical microstructure. This novel laser has great potential in advancing surgical techniques where precision osseous manipulation is required.

The Otto Aufranc Award: Demineralized bone matrix around porous implants promotes rapid gap healing and bone ingrowth.

BACKGROUND: Noncemented revision arthroplasty is often complicated by the presence of bone implant gaps that reduce initial stability and biologic fixation. Demineralized bone matrix has osteoinductive properties and therefore the potential to enhance gap healing and porous implant fixation. QUESTIONS/PURPOSES: We determined at what times and to what extent demineralized bone matrix promotes gap healing and bone ingrowth around a porous implant. METHODS: We inserted porous titanium implants into the proximal metaphyses of canine femora and humeri, with an initial 3-mm gap between host cancellous bone and implants. We left the gaps empty (control; n = 12) or filled them with either demineralized bone matrix (n = 6) or devitalized demineralized bone matrix (negative control; n = 6) and left them in situ for 4 or 12 weeks. We quantified volume healing of the gap with new bone using three-dimensional micro-CT scanning and quantified apposition and ingrowth using backscattered scanning electron microscopy. RESULTS: The density of bone inside gaps filled with demineralized bone matrix reached 64% and 93% of surrounding bone density by 4 and 12 weeks, respectively. Compared with empty controls and negative controls at 4 and 12 weeks, gap healing using demineralized bone matrix was two to three times greater and bone ingrowth and apposition were up to 15 times greater. CONCLUSIONS: Demineralized bone matrix promotes rapid bone ingrowth and gap healing around porous implants. CLINICAL RELEVANCE: Demineralized bone matrix has potential for enhancing implant fixation in revision arthroplasty.

Implant stability is affected by local bone microstructural quality.

It is known that low bone quality, caused for instance by osteoporosis, not only increases the risk of fractures, but also decreases the performance of fracture implants; yet the specific mechanisms behind this phenomenon are still largely unknown. We hypothesized that especially peri-implant bone microstructure affects implant stability in trabecular bone, to a greater degree than more distant bone. To test this hypothesis we performed a computational study on implant stability in trabecular bone. Twelve humeral heads were measured using micro-computed tomography. Screws were inserted digitally into these heads at 25 positions. In addition, at each screw location, a virtual biopsy was taken. Bone structural quality was quantified by morphometric parameters. The stiffness of the 300 screw-bone constructs was quantified as a measure of implant stability. Global bone density correlated moderately with screw-bone stiffness (r2=0.52), whereas local bone density was a very good predictor (r2=0.91). The best correlation with screw-bone stiffness was found for local bone apparent Young's modulus (r2=0.97), revealing that not only bone mass but also its arrangement in the trabecular microarchitecture are important for implant stability. In conclusion, we confirmed our hypothesis that implant stability is affected by the microstructural bone quality of the trabecular bone in the direct vicinity of the implant. Local bone density was the best single morphometric predictor of implant stability. The best predictability was provided by the mechanical competence of the peri-implant bone. A clinical implication of this work is that apparently good bone stock, such as assessed by DXA, does not guarantee good local bone quality, and hence does not guarantee good implant stability. New tools that could quantify the structural or mechanical quality of the peri-implant bone may help improve the surgical intervention in reaching better clinical outcomes for screw fixation.

Primary kaposiform hemangioendothelioma of a long bone: two cases in unusual locations with long-term follow up.

Kaposiform hemangioendothelioma (KHE) is a rare vascular neoplasm of low malignant potential that mainly affects infants and adolescents. The tumor almost exclusively occurs in somatic soft tissue or the retroperitoneum. We report herein two cases of primary KHE occurring in a long bone without cutaneous changes with long-term follow up in young patients. The patients were a 9-year-old girl and 5-year-old boy presenting with lytic lesions of the femur and humerus, respectively, without cutaneous lesions. Histologically, the neoplasms were comprised of nodules of spindle- to oval-shaped cells growing in an infiltrative fashion. The neoplastic cells formed poorly canalized or slit-like blood vessels alternating with solid spindle areas. Immunohistochemical studies showed that the tumor cells expressed CD31, CD34 and Fli1, but not HHV8, LNA-1 or GLUT1. D2-40 stained the neoplastic spindle cells and lymphatic channels adjacent to vascular lobules. The girl remains well with 15 years and 6 months follow up after a second complete excision. The boy has no signs of recurrence or metastasis nearly 5 years after local complete excision. To our best knowledge, this is the first report in the English literature of primary long bone occurrences of KHE without cutaneous changes with long-term follow up.

Incidence of humeral head of biceps brachii muscle: anatomical insight / Incidencia de la cabeza humeral del músculo bíceps braquial - visión anatómica

Biceps brachii is stated as one of the muscles that shows most frequent anatomical variations. Its most commonly reported anomaly is the presence of an accessory fascicle arising from the humerus which is termed as the humeral head of biceps brachii. Evidence shows a clear racial trend in the incidence of the humeral head of biceps brachii. Therefore, detailed knowledge of this variation in different populations is important for surgical interventions of the arm, nerve compression syndromes and in unexplained pain syndromes in the arm or shoulder region. The goal of this study was to elucidate the incidence and morphological features of this muscle in an adult Sri Lankan population. Upper extremities of the total of one hundred thirty five cadavers were dissected and studied for the presence of accessory heads of the biceps brachii muscle. The proximal and distal attachments of the humeral heads as well as their cranio-caudal, antero-posterior and medio-lateral dimensions were recorded. The incidence of humeral head of biceps brachii was found to be 3.7 percent. In all cases, it was found unilaterally and only in male subjects. The humeral head originated from the antero-medial aspect of the humeral shaft and descended and merged with the other two heads to form a common tendon. The results of the present study further highlight the racial variations in the incidence of humeral head of biceps brachii among Sri Lankans. Knowledge of the occurrence of humeral head of biceps brachii may facilitate preoperative diagnosis as well as the surgical procedures of the upper limb thus avoiding iatrogenic injuries.

El músculo bíceps braquial se conoce como uno de los músculos que muestra las variaciones anatómicas más frecuentes. Su anomalía más común es la presencia de un fascículo accesorio proveniente del húmero, que se denomina cabeza humeral del músculo bíceps braquial. La evidencia muestra una clara tendencia racial en la incidencia de la cabeza humeral del músculo bíceps braquial. El conocimiento acabado de esta variación, en las diferentes poblaciones, es importante para las intervenciones quirúrgicas del brazo, en los síndromes de compresión nerviosa y en los síndromes de dolor inexplicable en la región del brazo o del hombro. El objetivo de este estudio fue determinar la incidencia y las características morfológicas de este músculo en una población adulta de Sri Lanka. Fueron estudiados los miembros superiores en 135 cadáveres, disecados para evaluar la presencia de las cabezas del músculo bíceps braquial accesorio. Fueron registrados el origen e inserción de la cabeza humeral del músculo bíceps braquial, así como su dimensión cráneo-caudal, anteroposterior y mediolateral. La incidencia de la cabeza humeral del músculo bíceps braquial se encontró en el 3,7 por ciento de los miembros estudiados. En todos los casos, su presencia era unilateral y sólo presente en hombres. La cabeza humeral se originó en la región antero-medial de la diáfisis del húmero, descendió y se fusionó con las otras dos cabezas para formar un tendón común. Los resultados de este estudio resaltan aún más las variaciones raciales en la incidencia de la cabeza humeral del músculo bíceps braquial, entre los habitantes de Sri Lanka. El conocimiento de la presencia de la cabeza humeral del músculo bíceps braquial puede facilitar el diagnóstico preoperatorio, así como los procedimientos quirúrgicos del miembro superior, evitando las lesiones iatrogénicas.

Trabecular bone structure in the humeral and femoral heads of anthropoid primates.

The functional significance of three-dimensional trabecular bone architecture in the primate postcranial skeleton has received significant interest over the last decade. Some previous work has produced promising results, finding significant relationships between femoral head trabecular bone structure and hypothesized locomotor loading in leaping and nonleaping strepsirrhines. Conversely, most studies of anthropoid femoral head bone structure have found broad similarity across taxonomic and locomotor groups. The goal of this study is to expand on past analyses of anthropoid trabecular bone structure by assessing the effects of differential limb usage on the trabecular bone architecture of the forelimb and hindlimb across taxa characterized by diverse locomotor behaviors, including brachiation, quadrupedalism, and climbing. High-resolution x-ray computed tomography scans were collected from the proximal humerus and proximal femur of 55 individuals from five anthropoid primate species, including Symphalangus syndactylus, Papio sp., Presbytis rubicunda, Alouatta caraya, and Pan troglodytes. Trabecular bone structural features including bone volume fraction, anisotropy, trabecular thickness, and trabecular number were quantified in large volumes positioned in the center of the humeral or femoral head. Femoral head trabecular bone volume is consistently and significantly higher than trabecular bone volume in the humerus in all taxa independent of locomotor behavior. Humeral trabecular bone is more isotropic than femoral trabecular bone in all species sampled, possibly reflecting the emphasis on a mobile shoulder joint and manipulative forelimb. The results indicate broad similarity in trabecular bone structure in these bones across anthropoids.

What humeri are suitable for comparative testing of suture anchors? An ultrastructural bone analysis and biomechanical study of ovine, bovine and human humeri and four different anchor types.

For testing of fixation devices such as suture anchors used in rotator cuff repair often animal bones are used. They are easily obtained, inexpensive and some have been found to be similar to human bone. But can we rely on the results drawn from these studies in our daily surgical practice? The purpose of this study was to compare the trabecular bone mineral density, the trabecular bone volume fraction and the cortical layer thickness in the greater tubercle in different species to evaluate their influence on primary stability of suture anchors under a cyclic loading protocol representing the physiologic forces placed on rotator cuff repairs in vivo. Bovine and ovine humeri are not suitable for suture anchor testing. The statistical significances for pullout forces between the anchors varied from species to species. Therefore, no very applicable information can be obtained from testing suture anchors in ovine or bovine humeri with regard to ultimate failure loads in human humeri. The ultimate failure load seems to depend mainly on the cortical thickness and on the subcortical trabecular bone quality.

The influence of severe prolonged exercise restriction on the mechanical and structural properties of bone in an avian model.

Many studies have described the effects of exercise restriction on the mammalian skeleton. In particular, human and animal models have shown that reduction in weight bearing leads to generalised bone loss and deterioration of its mechanical properties. The aim of this study was to assess the effect of prolonged exercise restriction coupled with heavy calcium demands on the micro-structural, compositional and mechanical properties of the avian skeleton. The tibiae and humeri of 2-year-old laying hens housed in conventional caging (CC) and free-range (FR) housing systems were compared by mechanical testing and micro-computed tomography (microCT) scanning. Analyses of cortical, cancellous and medullary bone were performed. Mechanical testing revealed that the tibiae and humeri of birds from the FR group had superior mechanical properties relative to those of the CC group, and microCT scanning indicated larger cortical and lower medullary regions in FR group bones. Cancellous bone analysis revealed higher trabecular thickness and a higher bone volume fraction in the FR group, but no difference in mineral density. The biomechanical superiority of bones from the FR group was primarily due to structural rather than compositional differences, and this was reflected in both the cortical and cancellous components of the bones. The study demonstrated that prolonged exercise restriction in laying hens resulted in major structural and mechanical effects on the bird skeleton.

Bone microstructure in juvenile chimpanzees.

The growth, development, and maintenance of bone are influenced by genetic and environmental variables. Understanding variability in bone microstructure among primates may help illuminate the factors influencing the number and size of secondary osteons. The purpose of this study is to assess the bone microstructure in 8 humeral and 12 femoral sections of 12 juvenile chimpanzees, aged 2-15.3 years, and one adult chimp. Secondary osteons were counted and measured for 16 fields per section. Results indicate that the femur exhibits a mean osteon population density (OPD) of 4.46 +/- 2.34/mm(2), mean Haversian canal area of 0.0016 +/- 0.0007 mm(2), and mean osteon area of 0.033 +/- 0.006 mm(2). The humerus has a mean OPD of 4.72 +/- 1.57/mm(2), mean Haversian canal area of 0.0013 +/- 0.0003 mm(2), and mean osteon area of 0.033 +/- 0.005 mm(2). Differences are not significant between the humerus and femur, possibly indicating similar mechanical demands during locomotion. Osteon population density exhibits a moderate correlation with age (r = 0.498) in the femur of the juvenile chimps, but the adult chimp has an OPD of 10.28/mm(2), suggesting that osteons likely accumulate with age. Females exhibit higher osteon densities in the periosteal envelope compared to males in the humerus, indicating more remodeling during periosteal expansion. Overall similarities between chimpanzees and humans as well as previously published data on Late Pleistocene hominids (Abbott et al.: Am J Phys Anthropol 99 1996 585-601) suggest that bone microstructure has been stable throughout human evolution.

Comparative bone histology of adult horses (Equus caballus) and cows (Bos taurus).

Bone microstructure of domestic herbivores is still not completely understood. Indeed, works focused on the bone histology of numerous Mammalian species frequently led to misunderstandings because of the high number of variations such as the kind of bone, section orientation, species, breed and age. Moreover, attempts to identify the species in archaeozoological studies by a mere qualitative approach have not been encouraging and in recent years quantitative methods, based on image processing and statistical analysis, have appeared. The present study was undertaken to determine whether morphometrical and morphological differences exist in the compact bone structure of the femur and humerus between horses and cows. Measurements such as area, perimeter, minimum and maximum diameter of osteons and Haversian canals as well as the osteonal density were carried out on cross sections of eight humeri and eight femurs of the two herbivores investigated. In agreement with other authors, the qualitative investigation confirmed that the compact bone of horses and cows can be classified as dense Haversian tissue. Osteons of the horse were more numerous and composed of a higher number of well-defined lamellae when compared with the cow. Diameter, perimeter and area of osteons and Haversian canals were always higher in horses than in cows and this pattern could be related to the different locomotor behaviour of these animals.

Fracture length scales in human cortical bone: the necessity of nonlinear fracture models.

Recently published data for fracture in human humeral cortical bone are analyzed using cohesive-zone models to deal with the nonlinear processes of material failure. Such models represent the nonlinear deformation processes involved in fracture by cohesive tractions exerted by the failing material along a fracture process zone, rather than attributing all damage to a process occurring at a single point, as in conventional linear-elastic fracture mechanics (LEFM). The relationship between the tractions and the net displacement discontinuity across the process zone is hypothesized to be a material property for bone. To test this hypothesis, the cohesive law was evaluated by analyzing published load vs. load-point displacement data from one laboratory; the calibrated law was then used to predict similar data taken for a different source of bone using a different specimen geometry in a different laboratory. Further model calculations are presented to illustrate more general characteristics of the nonlinear fracture of bone and to demonstrate in particular that LEFM is not internally consistent for all cases of interest. For example, the fracture toughness of bone deduced via LEFM from test data is not necessarily a material constant, but will take different values for different crack lengths and test configurations. LEFM is valid when the crack is much longer than a certain length scale, representative of the length of the process zone in the cohesive model, which for human cortical bone ranges from 3 to 10mm. Since naturally occurring bones and the specimens used to test them are not much larger than this dimension for most relevant orientations, it is apparent that only nonlinear fracture models can give an internally consistent account of their fracture. The cohesive law is thus a more complete representation of the mechanics of material failure than the single-parameter fracture toughness and may therefore provide a superior measure of bone quality. The analysis of fracture data also requires proper representation of the approximately orthotropic elasticity of the bone specimen; if the specimen is incorrectly assumed to be isotropic, the initial measured compliance cannot be reproduced to within a factor of four and the fracture toughness deduced from the measured work of fracture will be overestimated by approximately 30%.

A light and electron microscopic study of the limb long bones perichondral ossification in the quail embryo (Coturnix coturnix japonica).

The perichondral ossification of the limb long bones in the quail embryo is investigated, in this study, by means of light and electron microscopy. Longitudinal sections of the humerus, radius, ulna, femur, tibia and fibula stained with haematoxylin-eosin were examined by the light microscope. Ultrathin cross sections were selected for the electron microscope as well. Light microscopic analysis showed that the ossification began at the same time in the long bones of the wing and leg. At the embryonic day 6, all the cartilaginous rudiments consisted of three zones. The central zone composed of hypertrophic chondrocytes, a second zone on either side of the central zone, which consisted of flattened cells and a third zone, which represented the epiphyseal region. A thin sheath of osteoid and a bi-layered perichondrium-periosteum surrounded the central zone of the cartilaginous rudiments of the long bones. The perichondrium consisted of a layer of osteoblasts, in contact with the cartilage, and a layer of fibroblasts. At the embryonic day 7, the thickness of the calcified osteoid ring increased and a vasculature appeared between the layer of osteoblasts and the layer of fibroblasts. At the embryonic day 8, a second sheath of periosteal bone began to be formed. Concurrently, vascular and perivascular elements began to invade the cartilage. The ossification spread towards the distal ends of both the diaphysis. At the electron microscopic level, the osteoblasts of the perichondium showed cytoplasmatic characteristics of cells involved in protein synthesis. The perichondral ossification is the first hallmark of the osteogenesis in the long bones. The observations reported above, are in accordance with previous studies in the chick embryo.

Depressed bone mineralization following high dose TGF-beta1 application in an orthopedic implant model.

Several previous studies of bone repair have shown 2- to 4-fold increases in bone formation following local delivery of exogenous transforming growth factor-beta (TGF-beta). Here, we use quantitative backscatter electron microscopy to test the effect of TGF-beta1 on mineralization of regenerated bone by examining tissue samples from a previously published canine study in which we found increased bone formation. In the experiment, the proximal humeri of 10 male canines were implanted bilaterally for 28 days with porous-coated implants in the presence of a 3 mm gap between the surface of the implant and the host bone. Implants placed in the left humeri were treated with TGF-beta1 at a dose of either 120 microg (n = 5) or 335 microg (n = 5), and the implants placed in the contralateral humeri served as untreated controls. Quantitative backscatter scanning electron microscopy was used to assess the volume fraction of bone and its degree of mineralization in the 3 mm gaps. The calibrated grayscale mean and median values were depressed compared to the controls in the high dose group (p = 0.048 and p = 0.041, respectively), suggesting that high dose TGF-beta delayed or inhibited mineralization of newly formed osteoid.

[Optical, scanning electron and atomic force microscopic observation of the microstructure of human humeral bone].

OBJECTIVE: To observe the microstructure of human long bone with scanning electron and atomic force microscopes to understand the ultrastructural organization and the composition and the morphology of the bone collagen and minerals. METHOD: Fresh human cadaveric humeral bone were fixed and dehydrated to prepare the ground sections and thin slices, respectively, which were observed for bone microstructure using optical, scanning electron and atomic force microscopy, respectively. RESULTS: Under optical microscope, the bone lacunae were in circular permutation around the Haversian canal, and the canaliculi communicated the lacunae and Haversian canal or between the lacunae. Atomic force microscopy represented distinct morphology of the large clusters of collagen, and the collagen fibers thickened in the mineralized zone and appeared in the shape of overlapping discs, whereas the calcium-phosphorus crystal displayed ellipse cylinders. The size, shape and relation of the canaliculi and lacunae were observed clearly. The bone matrix was observed in circular arrangement along the Haversian canal and the calcium-phosphorus crystal was in regular alignment under scanning electron microscope. CONCLUSION: Atomic force microscopy can help analyze the three-dimensional configuration of calcium-phosphorus crystal and bone collagen as well as the canaliculi and lacuna. The canaliculi serve as the channels mediating alimentation and molecular signals into the lacuna.