Periosteal Region Research Articles

Vertebral Development in Paleozoic and Mesozoic Tetrapods Revealed by Paleohistological Data.

Basal tetrapods display a wide spectrum of vertebral centrum morphologies that can be used to distinguish different tetrapod groups. The vertebral types range from multipartite centra in stem-tetrapods, temnospondyls, and seymouriamorphs up to monospondylous centra in lepospondyls and have been drawn upon for reconstructing major evolutionary trends in tetrapods that are now considered textbook knowledge. Two modes of vertebral formation have been postulated: the multipartite vertebrae formed first as cartilaginous elements with subsequent ossification. The monospondylous centrum, in contrast, was formed by direct ossification without a cartilaginous precursor. This study describes centrum morphogenesis in basal tetrapods for the first time, based on bone histology. Our results show that the intercentra of the investigated stem-tetrapods consist of a small band of periosteal bone and a dense network of endochondral bone. In stereospondyl temnospondyls, high amounts of calcified cartilage are preserved in the endochondral trabeculae. Notably, the periosteal region is thickened and highly vascularized in the plagiosaurid stereospondyls. Among “microsaur” lepospondyls, the thickened periosteal region is composed of compact bone and the notochordal canal is surrounded by large cell lacunae. In nectridean lepospondyls, the periosteal region has a spongy structure with large intertrabecular spaces, whereas the endochondral region has a highly cancellous structure. Our observations indicate that regardless of whether multipartite or monospondylous, the centra of basal tetrapods display first endochondral and subsequently periosteal ossification. A high interspecific variability is observed in growth rate, organization, and initiation of periosteal ossification. Moreover, vertebral development and structure reflect different lifestyles. The bottom-dwelling Plagiosauridae increase their skeletal mass by hyperplasy of the periosteal region. In nectrideans, the skeletal mass decreases, as the microstructure is spongy and lightly built. Additionally, we observed that vertebral structure is influenced by miniaturization in some groups. The phylogenetic information that can be drawn from vertebral development, however, is limited.

JUVENILE IDIOPATHIC ARTHRITIS ASSOCIATED WITH INCOMPLETE PRIMARY HYPERTROPHIC OSTEOARTHROPATHY (PACHYDERMOPERIOSTOSIS)

Introduction. Hypertrophic osteoarthropathy (HOA) is a rare, hereditary disease characterized by clubbing and new bone formation in the periosteal region that may be associated with joint pain, cutaneous abnormalities, seborrhea or hyperhydrosis. Juvenile idiopathic arthritis (JIA) is one of the most frequent chronic diseases with childhood onset, patients develop inflammatory joint pain and extra-articular manifestations with imunologic substrate. The association of the two diseases is very rare. 24 years old patient related disease onset at the age of 14 with arthritis of proximal interphalangeal joints (PIP), knees and right ankle. He is diagnosed with oligoarticular form of juvenile idiopathic arthritis. He receives Methotrexate, Suphasalazyne and association of the two, but after 5 years biological therapy with Etanercept is started. Later on, repeated physical examination revealed digital clubbing, non-painful enlargment of hands and feet with sweaty teguments. Radiological examination with subperiosteal new bone formation of the distal tibia, the fibula, the radius, the ulna, the metacarpals and the phalanges confirms the suspicion of HOA – pachydermoperiostosis (PDP). Conclusion. Final diagnosis considers both entities. The incomplete form of PDP included hands and feet enlargement with extensive periostitis, palmoplantar hyperhydrosis, but no significant facial changes. The specific treatment for JIA did not influence the evolution of PDP.

Establishment of a xenograft model to explore the mechanism of bone destruction by human oral cancers and its application to analysis of role of RANKL.

The molecular mechanism underlying bone invasion caused by oral squamous cell carcinoma (OSCC) is not well understood. To elucidate the molecular mechanism, the development of more suitable xenograft models mimicking human mandibular bone destruction by OSCC has been required. Human OSCC cell lines, HSC3, HSC3-C1, and HSC3-R2, were injected in the periosteal region of the mandible in athymic mice, and the bone destruction was analyzed. Receptor activators of nuclear factor κ-B ligand (RANKL) mRNA and protein expression levels were measured in the OSCC cell lines. Antibody that specifically neutralizes mouse RANKL and human RANKL, respectively, was injected into HSC3-cell-transplanted mice. Transplantation of HSC3 cells induced mandibular bone destruction. Histological examination revealed numerous osteoclasts on the bone destruction surface. Fibroblastic cell intervention between the cancer nests and resorbing bone surface was observed in a similar fashion to those observed in human OSCC cases. The number of osteoclasts and fibroblasts was significantly correlated. Bone destruction induced by the transplantation of HSC3 cells was reduced by injection of an antibody that specifically neutralizes mouse RANKL. Transplantation of HSC3-R2 cells, which overexpresses RANKL, induced advanced bone destruction compared to that of HSC3-C1 cells, which only overexpress the empty vector. We established a useful xenograft model for investigating the molecular mechanism underlying the bone destruction induced by OSCC in the jaw. This model will be used to investigate the precise roles of several cytokines synthesized by both cancer cells and fibroblastic cells in OSCC-associated bone destruction in the jaw.

Abcès sous périosté chez un enfant : la théorie de l’ostéomyélite selon Trueta remise en cause ?

Subperiosteal abscess formation is almost exclusively seen secondary to underlying hematogenous infected osteomyelitis or secondary as a result of a contagious focus. We present an unusual case of a 9-year-old girl with progressive ankle pain due to an isolated subperiosteal abscess of the distal fibula without concomitant osteomyelitis. The subperiosteal abscess was most likely caused by hematogenous spread to the periosteal region of the distal fibula located above the highly vascularized metaphysis. Remarkably, there were no signs of osteomyelitis on either MRI or during surgical inspection. She was successfully treated with debridement and antibiotic therapy. We hypothesize that subperiosteal abscess formation near the metaphysis originates in the periosteal region and not from outward extension from the sinusoidal veins in the intrametaphyseal area to the cortex and subperiosteal region.

Subperiosteal abscess in a child. Trueta's osteomyelitis hypothesis undermined?

Subperiosteal abscess formation is almost exclusively seen secondary to underlying hematogenous infected osteomyelitis or secondary as a result of a contagious focus. We present an unusual case of a 9-year-old girl with progressive ankle pain due to an isolated subperiosteal abscess of the distal fibula without concomitant osteomyelitis. The subperiosteal abscess was most likely caused by hematogenous spread to the periosteal region of the distal fibula located above the highly vascularized metaphysis. Remarkably, there were no signs of osteomyelitis on either MRI or during surgical inspection. She was successfully treated with debridement and antibiotic therapy. We hypothesize that subperiosteal abscess formation near the metaphysis originates in the periosteal region and not from outward extension from the sinusoidal veins in the intrametaphyseal area to the cortex and subperiosteal region.

Skeletal maturation substantially affects elastic tissue properties in the endosteal and periosteal regions of loaded mice tibiae

Although it is well known that the bone adapts to changes in the mechanical environment by forming and resorbing the bone matrix, little is known about the influence of mechanical loading on tissue material properties of the pre-existing and newly formed bone. In this study, we analyzed the newly formed and pre-existing tissue after two weeks of controlled in vivo axial compressive loading in tibia of young (10week-old) and adult (26week-old) female mice and compared to the control contralateral limb, by means of scanning acoustic microscopy. Additionally, we used quantitative backscattered electron imaging to determine the bone mineral density distribution within the newly formed and pre-existing bone of young mice. No significant differences were found in tissue stiffness or mineral density in the pre-existing bone tissue as a result of external loading. In the endosteal region, 10 and 26week loaded animals showed a 9% reduction in bone tissue stiffness compared to control animals. An increase of 200% in the mineral apposition rate in this region was observed in both age groups. In the periosteal region, the reduction in bone tissue stiffness and the increase in bone mineral apposition rate as a result of loading were two times higher in the 10 compared to the 26week old animals. These data suggest that, during growth and skeletal maturation, the response of bone to mechanical loading is a deposition of new bone matrix, where the tissue amount but not its mineral or elastic properties are influenced by animal age.

Effects of strain artefacts arising from a pre-defined callus domain in models of bone healing mechanobiology.

Iterative computational models have been used to investigate the regulation of bone fracture healing by local mechanical conditions. Although their predictions replicate some mechanical responses and histological features, they do not typically reproduce the predominantly radial hard callus growth pattern observed in larger mammals. We hypothesised that this discrepancy results from an artefact of the models' initial geometry. Using axisymmetric finite element models, we demonstrated that pre-defining a field of soft tissue in which callus may develop introduces high deviatoric strains in the periosteal region adjacent to the fracture. These bone-inhibiting strains are not present when the initial soft tissue is confined to a thin periosteal layer. As observed in previous healing models, tissue differentiation algorithms regulated by deviatoric strain predicted hard callus forming remotely and growing towards the fracture. While dilatational strain regulation allowed early bone formation closer to the fracture, hard callus still formed initially over a broad area, rather than expanding over time. Modelling callus growth from a thin periosteal layer successfully predicted the initiation of hard callus growth close to the fracture site. However, these models were still susceptible to elevated deviatoric strains in the soft tissues at the edge of the hard callus. Our study highlights the importance of the initial soft tissue geometry used for finite element models of fracture healing. If this cannot be defined accurately, alternative mechanisms for the prediction of early callus development should be investigated.

Haversian system of compact bone and comparison between endosteal and periosteal sides using three-dimensional reconstruction in rat.

The current model of compact bone is that of a system of Haversian (longitudinal) canals connected by Volkmann's (transverse) canals. Models based on either histology or microcomputed tomography do not accurately represent the morphologic detail and microstructure of this system, especially that of the canal networks and their spatial relationships. The aim of the present study was to demonstrate the morphologic pattern and network of the Haversian system and to compare endosteal and periosteal sides in rats using three-dimensional (3D) reconstruction. Ten Sprague-Dawley rats aged 8-10 weeks were used. The femurs were harvested from each rat and fixed, decalcified with 10% EDTA-2Na, serially sectioned at a thickness of 5 µm, and then stained with hematoxylin and eosin. The serial sections were reconstructed three-dimensionally using Reconstruct software. The Haversian canals in the endosteal region were found to be large, highly interconnected, irregular, and close to neighboring canals. In contrast, the canals in the periosteal region were straight and small. This combined application of 3D reconstruction and histology examinations to the Haversian system has confirmed its microstructure, showing a branched network pattern on the endosteal side but not on the periosteal side.

Study of fossil bones by synchrotron radiation micro-spectroscopic techniques and scanning electron microscopy

Earlymost Villafranchian fossil bones of an artiodactyl and a perissodactyl from the Milia excavation site in Grevena, Greece, were studied in order to evaluate diagenetic effects. Optical microscopy revealed the different bone types (fibro-lamellar and Haversian, respectively) of the two fragments and their good preservation state. The spatial distribution of bone apatite and soil-originating elements was studied using micro-X-ray fluorescence (µ-XRF) mapping and scanning electron microscopy. The approximate value of the Ca/P ratio was 2.2, as determined from scanning electron microscopy measurements. Bacterial boring was detected close to the periosteal region and Fe bearing oxides were found to fill bone cavities, e.g. Haversian canals and osteocyte lacunae. In the perissodactyl bone considerable amounts of Mn were detected close to cracks (the Mn/Fe weight ratio takes values up to 3.5). Goethite and pyrite were detected in both samples by means of metallographic microscopy. The local Ca/P ratio determined with µ-XRF varied significantly in metal-poor spots indicating spatial inhomogeneities in the ionic substitutions. XRF line scans that span the bone cross sections revealed that Fe and Mn contaminate the bones from both the periosteum and medullar cavity and aggregate around local maxima. The formation of goethite, irrespective of the local Fe concentration, was verified by the Fe K-edge X-ray absorption fine structure (XAFS) spectra. Finally, Sr K-edge extended XAFS (EXAFS) revealed that Sr substitutes for Ca in bone apatite without obvious preference to the Ca1 or Ca2 unit-cell site occupation.

Radial Anatomic Variation of Ultrasonic Velocity in Human Cortical Bone

Quantitative ultrasound techniques can be used to retrieve cortical bone quality. The aim of this study was to investigate the anatomic variations in speed of sound (SOS) in the radial direction of cortical bone tissue. SOS measurements were realized in 17 human cortical bone samples with a 3.5-MHz transverse transmission device. The radial dependence of SOS was investigated in a direction perpendicular to the periosteum. For each sample, bone porosity was measured using an X-ray micro-computed tomography device. The mean SOS was 3586±255m/s. For 16 of 17 specimens, similar radial variations in SOS were observed. In the periosteal region, SOS first decreased in the direction of the endosteum and reached a minimum value approximately in the middle of the cortical bone. SOS then increased, moving to the endosteal region. A significant negative correlation was obtained between SOS and porosity (R=-0.54, p=0.02).

Unremodeled endochondral bone is a major architectural component of the cortical bone of the rat (Rattus norvegicus)

The laboratory rat is one of the most frequently-used animal models for studying bone biology and skeletal diseases. Here we show that a substantial portion of the cortical bone of mature rats is primary endochondral bone, consisting of a disorganized arrangement of mineralized collagen fibers. We characterize the structure and mechanical properties of the cortical bone of the rat. We show that the cortical bone consists of two architecturally distinct regions. One region, consisting of well-organized circumferential lamellae (CLB), is located in the endosteal and/or the periosteal regions while another, disorganized region, is located in the more central region of the cortex. Unexpectedly, we found that the disorganized region contains many islands of highly mineralized cartilage. Micro tomography showed different structural and compositional properties of the two primary structural elements; the CLB region has lower mineral density, lower porosity, larger but fewer blood vessels and fewer lacunae. However, no difference was found in the average lacunar volume. Additionally the mean indentation modulus of the CLB region was lower than that of the disorganized region. The islands of calcified cartilage were found to be extremely stiff, with an indentation modulus of 33.4 ± 3.5GPa. We conclude that though the cortical bone of rats is in part lamellar, its architecture is markedly different from that of the cortical bone of humans, a fact that must be borne in mind when using the rat as a model animal for studies of human bone biology and disease.

Induced Biochemical osteoporosis: Effects of 1-month calcium–deprived diet on rat bone remodelling with/ without contemporary administration of PTH(1-34)

It is known that rats fed calcium-deprived diet develop osteoporosis due to enhanced bone resorption secondary to parathyroid overactivity resulting from nutritional hypocalcemia. Therefore, rats provide a good experimental animal model for studying bone remodelling alterations during biochemical osteoporosis. This preliminary study is performed in 3 month-old Sprague Dawley male rats, divided into 4 groups (5 rats each): 1) base line, 2) normal diet for 4 weeks, 3) calcium-deprived diet for 4 weeks; 4) calcium-deprived diet for 4 weeks plus contemporary administration of PTH(1-34) 40µg/kg/day. Three labels of osteogenesis were performed at 1st , 20th and 27th day of experimental period in order to evaluate bone formation during animal treatment. Histomorphometrical analyses were performed on cortical bone of femoral diaphyses, as well as on trabecular bone of distal femoral metaphyses, both transversely sectioned. The preliminary results showed that at femur mid-diaphyseal level the diet induced a reduction of cortical bone area (even if not significant) with enlargement of the medullary canal due to endosteal resorption, while periosteal neo-deposition is similar in all groups and particularly abundant in those periosteal regions mainly devoted in answering the mechanical demands. PTH(1-34) treatment seems to reduce endosteal resorption only in those surfaces where periosteal mechanical loading are less consistent. Conversely, PTH(1-34) treatment doesn’t seem to affect osteoblast activity. Moreover, in distal femoral metaphyses, diet induced osteoclast activity, with a decrease in the amount of trabecular bone volume, confirming that this architecture is mainly devoted in answering the metabolic demands. The novelty of the proposed model Is the contemporary administration of PTH(1-34) together with calcium deprived diet to evaluate induced-biochemical osteoporosis. This model seems a good starting point for successive studies in order to study bone alterations during unbalanced calcium metabolism frequently occurring in aging and to define time and manner of bone mass recovery.

In vivo tracking of segmental bone defect healing reveals that callus patterning is related to early mechanical stimuli

This study addresses the hypothesis that callus formation, patterning, and mineralisation are impaired during the early phase of critical sized bone defect healing, and may relate to inter-fragmentary tissue strains within the bone defect area. Twenty four 12 week old Sprague Dawley rats were used for this study. They were divided into two groups defined by the femur bone defect size: (i) 1mm resulting in normal healing (NH), and (ii) a large sized 5mm defect resulting in critical healing (CH). Callus formation, patterning, and mineralisation kinetics in both groups were examined in the periosteal and osteotomy gap regions using a novel longitudinal study setup. Finite element analyses on µCT generated tomograms were used to determine inter-fragmentary tissue strain patterns and compared to callus formation and patterning over the course of time. Using a novel longitudinal study technique with µCT, in vivo tracking and computer simulation approaches, this study demonstrates that: (i) periosteal bone formation and patterning are significantly influenced by bone defect size as early as 2 weeks; (ii) osteotomy gap callus formation and patterning are influenced by bone defect size, and adapt towards a non-union in critical cases by deviating into a medullary formation route as early as 2 weeks after osteotomy; (iii) the new bone formation in the osteotomy gap enclosing the medullary cavity in the CH group is highly mineralised; (iv) inter-fragmentary strain patterns predicted during the very early soft callus tissue phase (less than 2 weeks) are concurrent with callus formation and patterning at later stages. In conclusion, bone defect size influences early onset of critical healing patterns.

Life History and Skeletal Adaptations in the Galapagos Marine Iguana (Amblyrhynchus cristatus) as Reconstructed with Bone Histological Data—A Comparative Study of Iguanines

The skeletal adaptations of Amblyrhynchus cristatus (Marine Iguanas) are of particular interest based on their amphibious lifestyle, which is unique among living lacertilian squamates. The well-known ecological data are applied to new bone histological findings, which revealed expected and unexpected congruencies. The cortical bone matrix consists of avascular lamellar-zonal bone tissue type. The geometrical disposition of the growth marks (i.e., their spacing) shows an unusual pattern for lizards: the growth cycles maintain a constant thickness until the growth is terminated, which is marked by the development of the external fundamental system (efs). Minor resorption processes within the inner periosteal cortical region and the occurrence of these thick growth cycles in A. cristatus result in high mean bone compactness values. The reported life-history data from ecological studies and the hypothesized annuality of the growth cycles indicate that this first decline in annual bone deposition rate is not congruent with the attainment of sexual maturity. In contrast, this event might be indicated by other histological changes in the growth record of A. cristatus, which they share exclusively with their sister group, Conolophus subcristatus (Land Iguana). The bone matrix of the growth zones and annuli differ in their thickness, their color in polarized light, and vary slightly in the amount and shape of osteocyte lacunae in both A. cristatus and C. subcristatus. These well-recognizable growth zones and annuli of the growth cycles change their thickness abruptly within the reported time frame of the attainment of sexual maturity in A. cristatus.

CXCL2 synthesized by oral squamous cell carcinoma is involved in cancer-associated bone destruction

To explore the mechanism of bone destruction associated with oral cancer, we identified factors that stimulate osteoclastic bone resorption in oral squamous cell carcinoma. Two clonal cell lines, HSC3-C13 and HSC3-C17, were isolated from the maternal oral cancer cell line, HSC3. The conditioned medium from HSC3-C13 cells showed the highest induction of Rankl expression in the mouse stromal cell lines ST2 and UAMS-32 as compared to that in maternal HSC3 cells and HSC3-C17 cells, which showed similar activity. The conditioned medium from HSC3-C13 cells significantly increased the number of osteoclasts in a co-culture with mouse bone marrow cells and UAMS-32 cells. Xenograft tumors generated from these clonal cell lines into the periosteal region of the parietal bone in athymic mice showed that HSC3-C13 cells caused extensive bone destruction and a significant increase in osteoclast numbers as compared to HSC3-C17 cells. Gene expression was compared between HSC3-C13 and HSC3-C17 cells by using microarray analysis, which showed that CXCL2 gene was highly expressed in HSC3-C13 cells as compared to HSC3-C17 cells. Immunohistochemical staining revealed the localization of CXCL2 in human oral squamous cell carcinomas. The increase in osteoclast numbers induced by the HSC3-C13-conditioned medium was dose-dependently inhibited by addition of anti-human CXCL2-neutralizing antibody in a co-culture system. Recombinant CXCL2 increased the expression of Rankl in UAMS-32 cells. These results indicate that CXCL2 is involved in bone destruction induced by oral cancer. This is the first report showing the role of CXCL2 in cancer-associated bone destruction.

Vascular endothelial cells as a source of multipotent stem‐like cells

During development, cartilage‐ and bone‐forming stem cells collaborate with vascular, nerve and hematopoietic cells to produce skeletal elements and the joints that link them together. When activated during postnatal repair, stem cells in the periosteal regions or recruited from bone marrow via the circulation are fully capable of healing most fractures. However, regeneration of skeletal tissues fails in patients with loss of articular cartilage (osteoarthritis), large, localized defects in bone (when large tumors or necrotic bones are removed) or generalized bone loss (osteoporosis). Bone marrow‐derived mesenchymal stem cells, long thought of as promising for cartilage, bone and skeletal muscle repair/regeneration, appear to be more important for the growth factors they produce than for their in vivo ability to differentiate into specific cell lineages. This talk will discuss the discovery that differentiated vascular endothelial cells can undergo a conversion into mesenchymal stem‐like cells (endothelial‐mesenchymal transition; EndMT); these stem‐like cells can in turn differentiate into chondrocytes, osteoblasts or adipocytes in vitro and in vivo. Thus, blood vessels may not only bring blood supply and oxygen to tissues undergoing repair but may also provide mesenchymal stem cells for cell differentiation. EndMT is regulated via TGF_/BMP receptor signaling. In patients with Fibrodysplasia ossificans progressiva (FOP), where skeletal muscle and associated soft tissues progressively turn into bone, endothelial cells contribute significantly to the chondrocytic and osteoblastic cells that form the ectopic bone. FOP patients are heterozygous for activating mutations in the BMP type I receptor ALK2. However, EndMT can also be induced in endothelial cells from individuals without FOP when BMP receptor signaling is activated in the absence of inhibitors of EndMT. One of these inhibitors, Vascular Endothelial Growth Factor‐A (VEGF), is critical at skeletal tissue repair sites by stimulating angiogenesis and osteoblast differentiation. New insights into mechanisms by which VEGF regulates stem cell differentiation suggest that it may be possible to block the negative effect of VEGF on EndMT without affecting its positive effect on osteoblast differentiation.

Anatomical distribution of the degree of mineralization of bone tissue in human femoral neck: Impact on biomechanical properties

Osteoporotic hip fractures represent a major public health problem associated with high human and economic costs. The anatomical variation of the tissue mineral density (TMD) and of the elastic constants in femoral neck cortical bone specimens is an important determinant of bone fragility. The purpose of this study was to show that a Synchrotron radiation microcomputed tomography system coupled with a multiscale biomechanical model allows the determination of the 3-D anatomical dependence of TMD and of the elastic constants (i.e. the mechanical properties of an anisotropic material) in human femoral neck.Bone specimens from the inferior femoral neck were obtained from 18 patients undergoing standard hemiarthroplasty. The specimens were imaged using 3-D synchrotron micro-computed tomography with a voxel size of 10.13μm, leading to the determination of the anatomical distributions of porosity and TMD. The elastic properties of bone tissue were computed using a multiscale model. The model uses the experimental data obtained at the scale of several micrometers to estimate the components of the elastic tensor of bone at the scale of the organ.Statistical analysis (ANOVA) revealed a significant effect of the radial position on porosity and TMD and a significant effect of axial position on TMD only. Porosity was found to increase in the radial direction moving from the periosteum inwards (p<10−5). At any given distance from the periosteum, porosity does not vary noticeably along the bone axis. TMD was found to be significantly higher (p<10−5) in the periosteal region than in other bone locations and decreases from the periosteal to the endosteal region with an average slope of 10.05g.cm−3.m−1, the decrease being faster in the porous part of the samples (average slope equal of 30.04g.cm−3.m−1) than in dense cortical bone. TMD was found to decrease from the distal to the proximal part of the femur neck (average slope of 6.5g.cm−3.m−1). Considering TMD variations in the radial direction induces weak changes of bone properties compared to constant TMD. TMD variations in the axial direction are responsible for a significant variation of elastic constants.These results demonstrate that the anatomical variations of TMD affect the bone elastic properties, which could be explained by the complex stress field in bone affecting bone remodeling. TMD spatial variations should be taken into account to properly describe the spatial heterogeneity of elastic coefficients of bone tissue at the organ scale.

Changes in intracortical microporosities induced by pharmaceutical treatment of osteoporosis as detected by high resolution micro-CT

Bone's microporosities play important biologic and mechanical roles. Here, we quantified 3D changes in cortical osteocyte-lacunae and other small porosities induced by estrogen withdrawal and two different osteoporosis treatments. Unlike 2D measurements, these data collected via synchrotron radiation-based μCT describe the size and 3D spatial distribution of a large number of porous structures. Six-month old female Sprague–Dawley rats were separated into four groups of age-matched controls, untreated OVX, OVX treated with PTH, and OVX treated with Alendronate (ALN). Intracortical microporosity of the medial quadrant of the femoral diaphysis was quantified at endosteal, intracortical, and periosteal regions of the samples, allowing the quantification of osteocyte lacunae that were formed primarily before versus after the start of treatment. Across the overall thickness of the medial cortex, lacunar volume fraction (Lc.V/TV) was significantly lower in ALN treated rats compared to PTH. In the endosteal region, average osteocyte lacunar volume (<Lc.V>) of untreated OVX rats was significantly lower than in age-matched controls, indicating a decrease in osteocyte lacunar size in bone formed on the endosteal surface after estrogen withdrawal. The effect of treatment (OVX, ALN, PTH) on the number of lacunae per tissue volume (Lc.N/TV) was dependent on the specific location within the cortex (endosteal, intracortical, periosteal). In both the endosteal and intracortical regions, Lc.N/TV was significantly lower in ALN than in untreated OVX, suggesting a site-specific effect in osteocyte lacuna density with ALN treatment. There also were a significantly greater number of small pores (5–100μm3 in volume) in the endosteal region for PTH compared to ALN. The mechanical impact of this altered microporosity structure is unknown, but might serve to enhance, rather than deteriorate bone strength with PTH treatment, as smaller osteocyte lacunae may be better able to absorb shear forces than larger lacunae. Together, these data demonstrate that current treatments of osteoporosis can alter the number, size, and distribution of microporosities in cortical rat lamellar bone.

Type VI collagen deficiency induces osteopenia with distortion of osteoblastic cell morphology

Bone consists of type I collagen as a major protein with minor various matrix proteins. Type VI collagen is one of bone matrix proteins but its function is not known. We therefore examined the effects of type VI collagen deficiency on bone. 3D-μCT analysis revealed that type VI collagen deficiency reduced cancellous bone mass. Cortical bone mass was not affected. Type VI collagen deficiency distorted the shape of osteoblasts both in the cancellous bone and in the cambium layer of periosteal region. Furthermore, type VI collagen deficiency disorganized collagen arrangement. These data indicate that type VI collagen contributes to maintain bone mass.

Decrease in the osteocyte lacunar density accompanied by hypermineralized lacunar occlusion reveals failure and delay of remodeling in aged human bone

Aging decreases the human femur's fatigue resistance, impact energy absorption, and the ability to withstand load. Changes in the osteocyte distribution and in their elemental composition might be involved in age-related bone impairment. To address this question, we carried out a histomorphometric assessment of the osteocyte lacunar distribution in the periosteal and endosteal human femoral cortexes of 16 female and 16 male donors with regard to age- and sex-related bone remodeling. Measurements of the bone mineral density distribution by quantitative backscattered electron imaging and energy dispersive X-ray analysis were taken to evaluate the osteocyte lacunar mineral composition and characteristics. Age-dependent decreases in the total osteocyte lacunar number were measured in all of the cases. This change signifies a risk for the bone's safety. Cortical subdivision into periosteal and endosteal regions of interest emphasized that, in both sexes, primarily the endosteal cortex is affected by age-dependent reduction in number of osteocyte lacunae, whereas the periosteal compartment showed a less pronounced osteocyte lacunar deficiency. In aged bone, osteocyte lacunae showed an increased amount of hypermineralized calcium phosphate occlusions in comparison with younger cases. With respect to Frost's early delineation of micropetrosis, our microanalyses revealed that the osteocyte lacunae are subject to hypermineralization. Intralacunar hypermineralization accompanied by a decrease in total osteocyte lacunar density may contribute to failure or delayed bone repair in aging bone. A decreased osteocyte lacunar density may cause deteriorations in the canalicular fluid flow and reduce the detection of microdamage, which counteracts the bone's structural integrity, while hypermineralized osteocyte lacunae may increase bone brittleness and render the bone fragile.