Microcrack surface density in the human otic capsule: An unbiased stereological quantification

Bone is continuously remodeled to repair and strengthen degenerative bone with accumulating dead osteocytes and microfractures. Inner ear osteoprotegerin (OPG)-mediated inhibition of otic capsular bone remodeling causes excessive perilabyrinthine bone degeneration. Consequently, microcracks accumulate around the inner ear. Microcracks cause osteocyte apoptosis and may disrupt the canalicular network connecting osteocytes. Despite their linear microscopic appearance, microcracks are three-dimensional disruption planes and represent surface areas inside a tissue space. With an elevated microcrack burden the number of disconnected osteocytes is expected to increase. This may prove relevant to ongoing research in otic focal pathologies like otosclerosis. Therefore, an unbiased quantification of the microcrack surface density (mm²/mm³) in the human otic capsule is essential. In this study unbiased stereology was applied to undecalcified bulk stained human temporal bones to demonstrate its feasibility in describing the three-dimensional reality behind two dimensional observations of microcracks. A total of 28 human temporal bones and five ribs were bulk stained in basic fuchsin, serially sectioned and hand-ground to a thickness of 80–120 μm. Both horizontal and vertical sections were produced and compared. This study showed that surface density of microcracks was significantly higher around the inner ear compared to ribs. Furthermore, no significant difference in microcrack surface density between horizontal and vertical sections in the temporal bone was demonstrated.     

Bone mineral 31P and matrix‐bound water densities measured by solid‐state 31P and 1H MRI

Bone is a composite material consisting of mineral and hydrated collagen fractions. MRI of bone is challenging because of extremely short transverse relaxation times, but solid‐state imaging sequences exist that can acquire the short‐lived signal from bone tissue. Previous work to quantify bone density via MRI used powerful experimental scanners. This work seeks to establish the feasibility of MRI‐based measurement on clinical scanners of bone mineral and collagen‐bound water densities, the latter as a surrogate of matrix density, and to examine the associations of these parameters with porosity and donors’ age. Mineral and matrix‐bound water images of reference phantoms and cortical bone from 16 human donors, aged 27–97 years, were acquired by zero‐echo‐time 31‐phosphorus (³¹P) and 1‐hydrogen (¹H) MRI on whole body 7T and 3T scanners, respectively. Images were corrected for relaxation and RF inhomogeneity to obtain density maps. Cortical porosity was measured by micro‐computed tomography (μCT), and apparent mineral density by peripheral quantitative CT (pQCT). MRI‐derived densities were compared to X‐ray‐based measurements by least‐squares regression. Mean bone mineral ³¹P density was 6.74 ± 1.22 mol/l (corresponding to 1129 ± 204 mg/cc mineral), and mean bound water ¹H density was 31.3 ± 4.2 mol/l (corresponding to 28.3 ± 3.7 %v/v). Both ³¹P and bound water (BW) densities were correlated negatively with porosity (³¹P: R² = 0.32, p < 0.005; BW: R² = 0.63, p < 0.0005) and age (³¹P: R² = 0.39, p < 0.05; BW: R² = 0.70, p < 0.0001), and positively with pQCT density (³¹P: R² = 0.46, p < 0.05; BW: R² = 0.50, p < 0.005). In contrast, the bone mineralization ratio (expressed here as the ratio of ³¹P density to bound water density), which is proportional to true bone mineralization, was found to be uncorrelated with porosity, age or pQCT density. This work establishes the feasibility of image‐based quantification of bone mineral and bound water densities using clinical hardware. Copyright © 2014 John Wiley & Sons, Ltd.     

The Sealed Osteons of Cortical Diaphyseal Bone. Early Observations Revisited With Scanning Electron Microscopy

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.     

Analysis of the Effect of Osteon Diameter on the Potential Relationship of Osteocyte Lacuna Density and Osteon Wall Thickness

An important hypothesis is that the degree of infilling of secondary osteons (Haversian systems) is controlled by the inhibitory effect of osteocytes on osteoblasts, which might be mediated by sclerostin (a glycoprotein produced by osteocytes). Consequently, this inhibition could be proportional to cell number: relatively greater repression is exerted by progressively greater osteocyte density (increased osteocytes correlate with thinner osteon walls). This hypothesis has been examined, but only weakly supported, in sheep ulnae. We looked for this inverse relationship between osteon wall thickness (On.W.Th) and osteocyte lacuna density (Ot.Lc.N/B.Ar) in small and large osteons in human ribs, calcanei of sheep, deer, elk, and horses, and radii and third metacarpals of horses. Analyses involved: (1) all osteons, (2) smaller osteons, either ≤150 μm diameter or less than or equal to the mean diameter, and (3) larger osteons (>mean diameter). Significant, but weak, correlations between Ot.Lc.N/B.Ar and On.W.Th/On.Dm (On.Dm = osteon diameter) were found when considering all osteons in limb bones (r values ?0.16 to ?0.40, P < 0.01; resembling previous results in sheep ulnae: r = ?0.39, P < 0.0001). In larger osteons, these relationships were either not significant (five/seven bone types) or very weak (two/seven bone types). In ribs, a negative relationship was only found in smaller osteons (r = ?0.228, P < 0.01); this inverse relationship in smaller osteons did not occur in elk calcanei. These results do not provide clear or consistent support for the hypothesized inverse relationship. However, correlation analyses may fail to detect osteocyte‐based repression of infilling if the signal is spatially nonuniform (e.g., increased near the central canal). Anat Rec,, 2011. © 2011 Wiley‐Liss, Inc.     

Localisation of prostaglandin endoperoxide H synthase (PGHS)-1 and PGHS-2 in bone following mechanical loading in vivo

Recent data suggests that induction of prostaglandin endoperoxide H synthase-2 (PGHS-2) is critical for the anabolic response of lamellar bone elicited by mechanical strain in vivo. The aim of the present study was to localise PGHS-1 and PGHS-2 in rat tibiae following four-point bending in vivo. Right tibiae of 19 adult female rats were subjected to 300 cycles of bending or sham loading at 2.0 Hz with an applied load of 65 N. At 0, 6, and 24 hr postloading, rats were anaesthetised and perfused with Bouin's fixative. Left and right tibiae were dissected, postfixed for 4 hr at 4°C, decalcified in EDTA, and embedded in paraffin. Serial 5 μM sections were stained for PGHS-1 and PGHS-2 using standard immunoperoxidase procedures. For the first time, immunoreactivity for both PGHS-1 and PGHS-2 was localised in bone cells in situ, in the rat tibia. PGHS-1 was distributed widely in all tibiae, while PGHS-2 showed sparse localisation. At the endocortical surfaces (EcS), osteoblasts, lining cells, and osteocytes close to the surface reacted strongly for PGHS-1, as did intracortical osteocytes. At the periosteal surface (PsS), osteoblasts and cells of the osteogenic region were immunopositive. Immediately after loading, the numerical density (n.mm⁻²) of osteocytes labeled with PGHS-1 was significantly greater in loaded tibiae compared to controls. This increase was not seen after sham loading. At 6 and 24 hr postloading, this difference was no longer evident. Staining for PGHS-2 was sparse compared to PGHS-1. Light to moderate reactivity was observed in osteocytes and canaliculae, but the numerical density of labeled cells was significantly less than that for PGHS-1. Moderate staining was seen in lining cells and osteoblasts at the EcS and PsS of some tibiae. Osteoclasts at the PsS reacted strongly for both PGHS-1 and PGHS-2. There was a similar load-related increase in the density of PGHS-2-labeled osteocytes 0 hr postloading. The labeled osteocyte density had decreased at 6 hr, but remained significantly greater in loaded bones. These results show that both forms of PGHS can be localised in bone cells, with PGHS-1 expressed to a greater extent than PGHS-2. The data also suggest that both PGHS-1 and PGHS-2 may play important roles in the early response of bone to mechanical loading in vivo. Anat. Rec. 252:580–586, 1998. © 1998 Wiley-Liss, Inc.     

Overexpression of RANKL in osteoblasts: a possible mechanism of susceptibility to bone disease in cystic fibrosis

Bone fragility and loss are a significant cause of morbidity in patients with cystic fibrosis (CF), and the lack of effective therapeutic options means that treatment is more often palliative rather than curative. A deeper understanding of the pathogenesis of CF‐related bone disease (CFBD) is necessary to develop new therapies. Defective CF transmembrane conductance regulator (CFTR) protein and chronic inflammation in bone are important components of the CFBD development. The receptor activator of nuclear factor kappa‐B ligand (RANKL) and osteoprotegerin (OPG) drive the regulation of bone turnover. To investigate their roles in CFBD, we evaluated the involvement of defective CFTR in their production level in CF primary human osteoblasts with and without inflammatory stimulation, in the presence or not of pharmacological correctors of the CFTR. No major difference in cell ultrastructure was noted between cultured CF and non‐CF osteoblasts, but a delayed bone matrix mineralization was observed in CF osteoblasts. Strikingly, resting CF osteoblasts exhibited strong production of RANKL protein, which was highly localized at the cell membrane and was enhanced in TNF (TNF‐α) or IL‐17‐stimulated conditions. Under TNF stimulation, a defective response in OPG production was observed in CF osteoblasts in contrast to the elevated OPG production of non‐CF osteoblasts, leading to an elevated RANKL‐to‐OPG protein ratio in CF osteoblasts. Pharmacological inhibition of CFTR chloride channel conductance in non‐CF osteoblasts replicated both the decreased OPG production and the enhanced RANKL‐to‐OPG ratio. Interestingly, using CFTR correctors such as C18, we significantly reduced the production of RANKL by CF osteoblasts, in both resting and TNF‐stimulated conditions. In conclusion, the overexpression of RANKL and high membranous RANKL localization in osteoblasts are related to defective CFTR, and may worsen bone resorption, leading to bone loss in patients with CF. Targeting osteoblasts with CFTR correctors may represent an effective strategy to treat CFBD. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Regional maps of rib cortical bone thickness and cross‐sectional geometry

Here we present detailed regional bone thickness and cross‐sectional measurements from full adult ribs using high resolution CT scans processed with a cortical bone mapping technique. Sixth ribs from 33 subjects ranging from 24 to 99 years of age were used to produce average cortical bone thickness maps and to provide average ± 1SD corridors for expected cross‐section properties (cross‐sectional areas and inertial moments) as a function of rib length. Results obtained from CT data were validated at specific rib locations using direct measurements from cut sections. Individual thickness measurements from CT had an accuracy (mean error) and precision (SD error) of ?0.013 ± 0.167 mm (R² coefficient of determination of 0.84). CT‐based measurement errors for rib cross‐sectional geometry were ?0.1 ± 13.1% (cortical bone cross‐sectional area) and 4.7 ± 1.8% (total cross‐sectional area). Rib cortical bone thickness maps show the expected regional variation across a typical rib's surface. The local mid‐rib maxima in cortical thickness along the pleural rib aspect ranged from range 0.9 to 2.6 mm across the study population with an average map maximum of 1.4 mm. Along the cutaneous aspect, rib cortical bone thickness ranged from 0.7 to 1.9 mm with an average map thickness of 0.9 mm. Average cross‐sectional properties show a steady reduction in total cortical bone area from 10% along the rib's length through to the sternal end, whereas overall cross‐sectional area remains relatively constant along the majority of the rib's length before rising steeply towards the sternal end. On average, male ribs contained more cortical bone within a given cross‐section than was seen for female ribs. Importantly, however, this difference was driven by male ribs having larger overall cross‐sectional areas, rather than by sex differences in the bone thickness observed at specific local cortex sites. The cortical bone thickness results here can be used directly to improve the accuracy of current human body and rib models. Furthermore, the measurement corridors obtained from adult subjects across a wide age range can be used to validate future measurements from more widely available image sources such as clinical CT where gold standard reference measures (e.g. such as direct measurements obtained from cut sections) are otherwise unobtainable.     

A comparative study of pneumatization of Temporal bone

Introduction

The present review is based on the study of various classifications of pneumatization of temporal bone and their comparison. The air cells are classified based on their location in the temporal bone in a radiograph or based on their interpretation by a radiologist or otolaryngologist with the help of different reference structures.

A potential mechanism for allometric trabecular bone scaling in terrestrial mammals

Trabecular bone microstructural parameters, including trabecular thickness, spacing, and number, have been reported to scale with animal size with negative allometry, whereas bone volume fraction is animal size‐invariant in terrestrial mammals. As for the majority of scaling patterns described in animals, its underlying mechanism is unknown. However, it has also been found that osteocyte density is inversely related to animal size, possibly adapted to metabolic rate, which shows a negative relationship as well. In addition, the signalling reach of osteocytes is limited by the extent of the lacuno‐canalicular network, depending on trabecular dimensions and thus also on animal size. Here we propose animal size‐dependent variations in osteocyte density and their signalling influence distance as a potential mechanism for negative allometric trabecular bone scaling in terrestrial mammals. Using an established and tested computational model of bone modelling and remodelling, we run simulations with different osteocyte densities and influence distances mimicking six terrestrial mammals covering a large range of body masses. Simulated trabecular structures revealed negative allometric scaling for trabecular thickness, spacing, and number, constant bone volume fraction, and bone turnover rates inversely related to animal size. These results are in agreement with previous observations supporting our proposal of osteocyte density and influence distance variation as a potential mechanism for negative allometric trabecular bone scaling in terrestrial mammals. The inverse relationship between bone turnover rates and animal size further indicates that trabecular bone scaling may be linked to metabolic rather than mechanical adaptations.     

Internal rib structure can be predicted using mathematical models: An anatomic study comparing the chest to a shell dome with application to understanding fractures

The human rib cage resembles a masonry dome in shape. Masonry domes have a particular construction that mimics stress distribution. Rib cortical thickness and bone density were analyzed to determine whether the morphology of the rib cage is sufficiently similar to a shell dome for internal rib structure to be predicted mathematically. A finite element analysis (FEA) simulation was used to measure stresses on the internal and external surfaces of a chest‐shaped dome. Inner and outer rib cortical thickness and bone density were measured in the mid‐axillary lines of seven cadaveric rib cages using computerized tomography scanning. Paired t tests and Pearson correlation were used to relate cortical thickness and bone density to stress. FEA modeling showed that the stress was 82% higher on the internal than the external surface, with a gradual decrease in internal and external wall stresses from the base to the apex. The inner cortex was more radio‐dense, P < 0.001, and thicker, P < 0.001, than the outer cortex. Inner cortical thickness was related to internal stress, r = 0.94, P < 0.001, inner cortical bone density to internal stress, r = 0.87, P = 0.003, and outer cortical thickness to external stress, r = 0.65, P = 0.035. Mathematical models were developed relating internal and external cortical thicknesses and bone densities to rib level. The internal anatomical features of ribs, including the inner and outer cortical thicknesses and bone densities, are similar to the stress distribution in dome‐shaped structures modeled using FEA computer simulations of a thick‐walled dome pressure vessel. Fixation of rib fractures should include the stronger internal cortex. Clin. Anat. 28:1008–1016, 2015. © 2015 Wiley Periodicals, Inc.     

Laterality and grip strength influence hand bone micro‐architecture in modern humans,an HRpQCT study

It is widely hypothesized that mechanical loading, specifically repetitive low‐intensity tasks, influences the inner structure of cancellous bone. As such, there is likely a relationship between handedness and bone morphology. The aim of this study is to determine patterns in trabecular bone between dominant and non‐dominant hands in modern humans. Seventeen healthy patients between 22 and 32 years old were included in the study. Radial carpal bones (lunate, capitate, scaphoid, trapezium, trapezoid, 1st, 2nd and 3rd metacarpals) were analyzed with high‐resolution micro‐computed tomography. Additionally, crush and pinch grip were recorded. Factorial analysis indicated that bone volume ratio, trabeculae number (Tb.N), bone surface to volume ratio (BS.BV), body weight, stature and crush grip were all positively correlated with principal components 1 and 2 explaining 78.7% of the variance. Volumetric and trabecular endostructural parameters (BV/TV, BS/BV or Tb.Th, Tb.N) explain the observed inter‐individual variability better than anthropometric or clinical parameters. Factors analysis regressions showed correlations between these parameters and the dominant side for crush strength for the lunate (r² = 0.640, P < 0.0001), trapezium (r² = 0.836, P < 0.0001) and third metacarpal (r² = 0.763). However, despite a significant lateralization in grip strength for all patients, the endostructural variability between dominant and non‐dominant sides was limited in perspective to inter‐individual differences. In conclusion, handedness is unlikely to generate trabecular patterns of asymmetry. It appears, however, that crush strength can be considered for endostructural analysis in the modern human wrist.     

The macroscopic and histomorphological properties of periosteal rib lesions and its relation with disease duration: evidence from the Luis Lopes Skeletal Collection (Lisbon,Portugal)

Periosteal new bone formation (PNBF) is a common finding in a large spectrum of diseases. In clinical practice, the morphology and location of periosteal lesions are frequently used to assist in the differential diagnosis of distinct bone conditions. Less commonly reported is the presence of PNBF on the ribs. This contrasts with the data retrieved from the study of skeletonized human remains that shows a high frequency of cases and a strong, albeit not specific, association between periosteal rib lesions and pulmonary conditions (e.g. tuberculosis). Despite that, an overall disagreement regarding the specificity and non‐specificity of periosteal reactions exists in the study of dry bone remains. The insufficient number of clinical models exploring the morphology and the pathophysiology of PNBF's and the lack of systematic studies of pathological samples with a known diagnosis are claimed as major reasons for the disagreements. This study aimed to describe and compare the macroscopic and the histomorphologic appearance of periosteal rib lesions and to discuss their usefulness as diagnostic indicators. To pursue this goal, an assemblage of 13 rib samples (males = 11, females = 2, mean age‐at‐death = 36.6 years old) was collected from the Luis Lopes Skeletal Collection (Museu Nacional de História Natural e da Ciência, Universidade de Lisboa, Portugal). The assemblage belongs to individuals who died from pulmonary‐TB (group 1), non‐TB pulmonary infections (group 2) and other conditions (group 3). Prior to sample preparation, the ribs were visually inspected and the PNBF described according to its thickness, the degree of cortical integration and the type of new bone formed (e.g. woven, lamellar or both). After sampling, each bone sample was prepared for histological analysis under plane and polarized light microscopy. Macroscopically, the results showed no differences in the new bone composition between cause‐of‐death groups. Only slight differences in the degree of cortical integration, which was most frequently classified as mild to high in the pulmonary‐TB group, were observed. Histologically, no distinguishing features were identified by pathological group. However, new bone microarchitectures were observed compatible with (1) acute, fast‐growing processes (e.g. spiculated reactions), (2) long‐standing processes with a rapid bone formation (e.g. appositional layering of bone) and/or (3) chronic, slow‐growing processes (e.g. layers of compact lamellae). To some extent, these distinct rates of disease progression resonate with the cause‐of‐death listed for some individuals. Despite the small sample size, the results of this investigation are in agreement with previous studies, according to which the macroscopic and histological appearance of periosteal formations are not specific for a particular pathological conditions. Nevertheless, the results support the conclusion that the morphology of periosteal lesions is a good biological indicator for inferring the rate of progression and duration of pathological processes. This study provides important reference data regarding the histomorphology of periosteal lesions that can be used for comparative purposes, as well as to narrow down the differential diagnosis in unidentified skeletal remains.     

3D‐Microarchitectural patterns of Hyperostosis frontalis interna: a micro‐computed tomography study in aged women

Although seen frequently during dissections and autopsies, Hyperostosis frontalis interna (HFI) – a morphological pattern of the frontal bone thickening – is often ignored and its nature and development are not yet understood sufficiently. Current macroscopic classification defines four grades/stages of HFI based on the morphological appearance and size of the affected area; however, it is unclear if these stages also depict the successive phases in the HFI development. Here we assessed 3D‐microarchitecture of the frontal bone in women with various degrees of HFI expression and in an age‐ and sex‐matched control group, hypothesizing that the bone microarchitecture bears imprints of the pathogenesis of HFI and may clarify the phases of its development. Frontal bone samples were collected during routine autopsies from 20 women with HFI (age: 69.9 ± 11.1 years) and 14 women without HFI (age: 74.1 ± 9.7 years). We classified the HFI samples into four groups, each group demonstrating different macroscopic type or stage of HFI. All samples were scanned by micro‐computed tomography to evaluate 3D bone microarchitecture in the following regions of interest: total sample, outer table, diploe and inner table. Our results revealed that, compared to the control group, the women with HFI showed a significantly increased bone volume fraction in the region of diploe, along with significantly thicker and more plate‐like shaped trabeculae and reduced trabecular separation and connectivity density. Moreover, the inner table of the frontal bone in women with HFI displayed significantly increased total porosity and mean pore diameter compared to controls. Microstructural reorganization of the frontal bone in women with HFI was also reflected in significantly higher porosity and lower bone volume fraction in the inner vs. outer table due to an increased number of pores larger than 100 μm. The individual comparisons between the control group and different macroscopic stages of HFI revealed significant differences only between the control group and the morphologically most pronounced type of HFI. Our microarchitectural findings demonstrated clear differences between the HFI and the control group in the region of diploe and the inner table. Macroscopic grades of HFI could not be distinguished at the level of bone microarchitecture and their consecutive nature cannot be supported. Rather, our study suggests that only two different types of HFI (moderate and severe HFI) have microstructural justification and should be considered further. It is essential to record HFI systematically in human postmortem subjects to provide more data on the mechanisms of its development.     

Morphologische Veränderungen der Beckenkammspongiosa beim primären Hyperparathyreoidismus und ihre Bedeutung für die Diagnostik

Summary Iliac crest bone biopsy specimens of 391 patients with surgically proven primary hyperparathyroidism were investigated. In 60 unselected cases quantitative analysis of trabecular bone changes was performed. The age of the patients ranged between 12 and 85 years. The observed morphological findings were divided into four stages. In a few cases no differences from normal bone tissue could be observed. In 46% occurrence of a nonspecific increase of osteoid seams, osteoblasts, and osteoclasts was observed. Of the cases 50% showed a specific, but very often mild endosteal fibrosis. Only in 4% was there a severe fibroosteoclasia with development of so-called brown tumors. The quantitative analysis showed an increase of trabecular bone mass as well as of remodeling surfaces. But there was an overlap of up to 25% with the normal controls. The results demonstrate the influence of parathyroid hormone peptides on bone morphology. However, the investigation of a bone biopsy specimen is not generally very useful for diagnostic purposes.

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Abkürzungen Dtrab mittlerer Trabekeldurchmesser () - NaOcl Zahl der Osteoclasten pro cm² Spongiosaoberfläche (n/cm²) - Ocl Zahl der Osteoklasten pro cm Spongiosaumfang (n/cm) - PHPT primärer Hyperparathyreoidismus - S mittlere Osteoidsaumbreite () - Sv Oberflächendichte der Spongiosa (mm²/cm³) - S/V spezifische Trabekeloberfläche (mm²/mm³) - Svos Oberflächendichte der osteoiden Säume (mm²/cm³) - Svob Oberflächendichte der mit Osteoblasten besetzten osteoiden Säume (mm²/cm³) - Svho Oberflächendichte der mit Osteoklasten besetzten Resorptionslakunen (mm²/cm³) - Svht Oberflächendichte aller Resorptionslakunen (mm²/cm³) - Vv Volumendichte der Spongiosa (%) - Vvo Volumendichte des Osteoids (%) Mit Unterstützung der Deutschen Forschungsgemeinschaft     

Bone Signaling in Middle Ear Development: A Genome‐Wide Differential Expression Analysis

Common middle ear diseases may affect bone behavior in the middle ear air cell system. To understand this pathologic pneumatization, the normal development of bone in the middle ear should be investigated. The objective of this study was to analyze gene expression of bone‐related signaling factors and gene sets in the developing middle ear. Microarray technology was used to identify bone‐related genes and gene sets, which were differentially expressed between the lining tissue of adult (quiescent) bulla and young (resorbing/forming) bulla. Data were analyzed using tools of bioinformatics and expression levels of selected genes were validated using quantitative polymerase chain reaction. The candidate gene products were compared with previously published data on middle ear bone metabolism. No differentially expressed genes were found on the outer surface of bulla. On the inner lining a total of 260 genes were identified of which 22 genes were involved in bone metabolism. Gene set analysis revealed five enriched bone‐related gene sets. The identified differentially expressed bone‐related mRNAs and gene sets are of potential significance in the normally developing bulla. These factors and gene sets may also play important roles during pathologic pneumatization of the middle ear air cell system in common middle ear diseases. In addition, this study suggests that the control of growth rate and wall thickness from resorptive as well as formative signals all originate from the inner lining cells of the bulla wall. Anat Rec, 297:2349–2355, 2014. © 2014 Wiley Periodicals, Inc.     

Intracortical remodelling increases in highly loaded bone after exercise cessation

Resorption within cortices of long bones removes excess mass and damaged tissue and increases during periods of reduced mechanical loading. Returning to high-intensity exercise may place bones at risk of failure due to increased porosity caused by bone resorption. We used point-projection X-ray microscopy images of bone slices from highly loaded (metacarpal, tibia) and minimally loaded (rib) bones from 12 racehorses, 6 that died during a period of high-intensity exercise and 6 that had a period of intense exercise followed by at least 35 days of rest prior to death, and measured intracortical canal cross-sectional area (Ca.Ar) and number (N.Ca) to infer remodelling activity across sites and exercise groups. Large canals that are the consequence of bone resorption (Ca.Ar >0.04 mm²) were 1.4× to 18.7× greater in number and area in the third metacarpal bone from rested than exercised animals (p = 0.005–0.008), but were similar in number and area in ribs from rested and exercised animals (p = 0.575–0.688). An intermediate relationship was present in the tibia, and when large canals and smaller canals that result from partial bony infilling (Ca.Ar >0.002 mm²) were considered together. The mechanostat may override targeted remodelling during periods of high mechanical load by enhancing bone formation, reducing resorption and suppressing turnover. Both systems may work synergistically in rest periods to remove excess and damaged tissue.     

健骨颗粒对去卵巢骨质疏松模鼠骨组织结构的影响

目的 :观察健骨颗粒对去卵巢骨质疏松模鼠股骨颈超微结构的影响以及同期骨密度 (BMD)的改变 ,探讨该药药效及作用机制。材料和方法 :SD雌性大鼠去卵巢造模 ,随机分成健骨颗粒预防组和治疗组 ,并设生理盐水和骨松宝对照组。术后 2 4周处死 ,分别行骨密度检测及股骨颈骨粒透射电镜观察。结果 :去势 1 2周 ,骨细胞以吸收相和退变相为主 ,BMD明显降低。生理盐水组骨细胞多表现为退变相 ,骨细胞性溶骨更加明显 ,且出现破骨细胞性溶骨现象。健骨颗粒治疗组和骨松宝组多见成骨相骨细胞 ,成骨细胞活性加强 ,BMD明显升高。健骨颗粒预防组活跃的成骨细胞数量明显增加 ,可见成骨细胞成骨过程 ,BMD接近假手术组。结论 :健骨颗粒能有效防治骨质疏松的发生和发展。本文还就其作用机制进行了讨论。     

Subchondral osteopenia and accelerated bone remodelling post‐ovariectomy – a possible mechanism for subchondral microfractures in the aetiology of spontaneous osteonecrosis of the knee?

Osteopenia and subchondral microfractures are implicated in the aetiology of spontaneous osteonecrosis of the knee (SPONK). The ovine tibia shows significant alterations of the trabecular architecture within the subchondral bone of the medial tibial plateau post‐ovariectomy (OVX), including reduced trabecular bone volume fraction. We hypothesise that accelerated subchondral bone resorption may also play a role in increasing microfracture risk at this site. Twenty‐two sheep were examined in this study; 10 of the sheep underwent OVX, while the remainder (n = 13) were kept as controls (CON). Five fluorochrome dyes were administered intravenously at 12‐week intervals via the jugular vein to both groups, to label sites of bone turnover. These animals were then killed at 12 months post‐operatively. Bone turnover was significantly increased in the OVX group in both trabecular bone (2.024 vs. 1.047 no. mm^?2; P = 0.05) and within the subchondral bone plate (4.68 vs. 0.69 no. mm^?2; P < 0.001). In addition to the classically described turnover visible along trabecular surfaces, we also found visual evidence of intra‐trabecular osteonal remodelling. In conclusion, this study shows significant alterations in bone turnover in both trabecular bone and within the subchondral bone plate at 1 year post‐OVX. Remodelling of trabecular bone was due to both classically described hemi‐osteonal and intra‐trabecular osteonal remodelling. The presence of both localised osteopenia and accelerated bone remodelling within the medial tibial plateau provide a possible mechanism for subchondral microfractures in the aetiology of SPONK. Further utilisation of the OVX ewe may be useful for further study in this field.     

Osteoclast induction in periodontal tissue during experimental movement of incisors in osteoprotegerin‐deficient mice

Osteoprotegerin (OPG) is a novel secreted member of the tumor necrosis factor (TNF) receptor superfamily that negatively regulates osteoclastogenesis. The receptor activator of the NFKB ligand (RANKL) is one of the key regulatory molecules in osteoclast formation and binds to OPG. In this study, it was suggested that OPG and RANKL are involved in alveolar bone remodeling during orthodontic tooth movement. We examined RANKL localization and osteoclast induction in periodontal tissues during experimental movement of incisors in OPG‐deficient mice. To produce orthodontic force, an elastic band was inserted between the upper right and left incisors for 2 or 5 days, and the dissected maxillae were examined for cytochemical and immunocytochemical localization of tartrate‐resistant acid phosphatase (TRAP), vacuolar‐type H⁺‐ATPase, and RANKL. Compared to wild‐type OPG (+/+) littermates, TRAP‐positive multinucleated cells were markedly induced in the periodontal ligament (PDL) on the compressed side and in the adjacent alveolar bone of OPG‐deficient mice. These multinucleated cells exhibited intense vacuolar‐type H⁺‐ATPase along the ruffled border membranes. Because of accelerated osteoclastic resorption in OPG‐deficient mice, alveolar bone was severely destroyed and partially perforated at 2 and 5 days after force application. In both wild‐type and OPG‐deficient mice, RANKL expression became stronger at 2 and 5 days after force application than before force application. There was no apparent difference in intensity of RANKL expression between OPG (+/+) littermates and OPG‐deficient mice. In both wild‐type and OPG‐deficient mice, expression of RANKL protein was detected in osteoblasts, fibroblasts, and osteoclasts mostly located in resorption lacunae. These results suggest that during orthodontic tooth movement, RANKL and OPG in the periodontal tissues are important determinants regulating balanced alveolar bone resorption. Anat Rec 266:218–225, 2002. © 2002 Wiley‐Liss, Inc.