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The study of skeletochronology and bone tissue as a record of information on ontogenetic stages and events is widely used for improving the knowledge about life histories (LHs) of extinct and extant vertebrates. Compared with dinosaurs and extant reptiles, mammalian bone histology has received little attention. Here, we calibrate for the first time bone and dental age with histological bone characteristics and LH stages in ontogenetic series of red deer. We rely on known LHs of different aged individuals of captive Cervus elaphus hippelaphus from Austria to correlate epiphyseal closure, dental eruption pattern, bone growth marks and bone tissue patterns in femora and tibiae, and of wild Cervus elaphus hispanicus from Spain. Our data show that females (of both subspecies) attain skeletal maturity earlier than males. At this moment, epiphyseal closure (in femora and tibiae) and dental eruption are complete and long bones start to deposit an external fundamental system. The results also show that the attainment of reproductive maturity in red deer occurs slightly before skeletal maturity., (© 2019 Anatomical Society.)

Endochondral ossification in the diaphysis of long bones has been studied in-depth during fetal development but not postnatally in the epiphysis. Immunohistochemical studies revealed that Sox9 and Col2 expressing immature chondrocytes in the epiphysis transition into prehypertrophic and hypetrophic chondrocytes and finally into osteoblasts expressing Col1 and BSP during postnatal day 7-10, when serum levels of thyroid hormone (TH) rise. Lineage tracing using Rosa-td tomato Col2-Cre-ERT2 mice treated with tamoxifen indicated that the same Col2 expressing chondrocytes expressed prehypertrophic, hypertrophic, and subsequently bone formation markers in a sequential manner in euthyroid but not hypothyroid mice, thus providing evidence that chondrocyte to osteoblast transdifferentiation is TH-dependent. Vascular invasion was apparent at the time of bone formation but not earlier. In vitro studies revealed that TH acting via TRα1 promoted expression of SHH while TRβ1 activation increased IHH but inhibited SHH expression. SHH promoted expression of markers of immature chondrocytes but inhibited chondrocyte hypertrophy while IHH promoted chondrocyte hypertrophy. Based on our data, we propose a model in which TH acting through TRα1 and TRβ1, respectively, fine tune levels of SHH and IHH and, thereby control the transit of proliferating immature chondrocytes into mature hypertrophic chondrocytes to become osteoblasts at the epiphysis.

Objective: The role of Snorc, a novel cartilage specific transmembrane proteoglycan, was studied during skeletal development using two Snorc knockout mouse models. Hypothesizing that Snorc, like the other transmembrane proteoglycans, may be a coreceptor, we also studied its interaction with growth factors., Methods: Skeletal development was studied in wild type (WT) and Snorc knockout mice during postnatal development by whole mount staining, X-ray imaging, histomorphometry, immunohistochemistry and qRT-PCR. Snorc promoter activity was studied by applying the LacZ reporter expressed by the targeting construct. Slot blot binding and cell proliferation assays were used to study the interaction of Snorc with several growth factors., Results: Snorc expression was localized in the knee epiphyses especially to the prehypertrophic chondrocytes delineating the cartilage canals and secondary ossification center (SOC). Snorc was demonstrated to have a glycosaminoglycan independent affinity to FGF2 and it inhibited FGF2 dependent cell growth of C3H101/2 cells. In Snorc deficient mice, SOCs in knee epiphyses were smaller, and growth plate (GP) maturation was disturbed, but total bone length was not affected. Central proliferative and hypertrophic zones were enlarged with higher extracellular matrix (ECM) volume and rounded chondrocyte morphology at postnatal days P10 and P22. Increased levels of Ihh and Col10a1, and reduced Mmp13 mRNA expression were observed at P10., Conclusions: These findings suggest a role of Snorc in regulation of chondrocyte maturation and postnatal endochondral ossification. The interaction identified between recombinant Snorc core protein and FGF2 suggest functions related to FGF signaling., (Copyright © 2017 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.)

Objective: To compare regional proportions and spatial distributions of volumetric bone mineral density (BMDv) of the palmar aspect of the distal epiphysis of the third metacarpal bone (McIII) in limbs with or without a condylar fracture from Thoroughbred racehorses., Sample: McIIIs from cadavers of Thoroughbred racehorses with (n = 6 bones) and without (8) a condylar fracture., Procedures: BMDv and spatial distributions of BMDv in peripheral quantitative CT images of the distal epiphysis of McIIIs were quantitatively assessed with spatial analysis software. Relative proportions of voxels within 9 threshold categories of BMDv and spatial statistics for BMDv distribution were compared between fractured and nonfractured limbs., Results: No significant differences in BMDv characteristics were identified between fractured and nonfractured limbs, although fractured limbs had a lower proportion of voxels in the BMDv thresholds 700 to < 800 mg/cm(3) and 800 to < 900 mg/cm(3) but a higher proportion of voxels in the BMDv threshold 1,000 to < 1,100 mg/cm(3) for the central condylar region of the medial condyle. Results of spatial analysis reflected the response of bone to race training rather than differences between fractured and nonfractured limbs. In both limb groups, uniform clusters of low BMDv with areas of high BMDv were identified., Conclusions and Clinical Relevance: BMDv characteristics of the distal epiphysis of McIII reflected training load, and fracture characteristics were subtle. Serial imaging techniques in conjunction with detailed training data are required to elucidate the onset of the pathological response to load in horses.

Only two distal epiphyses of a radius and ulna are consensually attributed to the holotype skeleton of Proconsul heseloni, KNM-RU 2036. Here, we describe seven adult and immature distal antebrachial (radial and ulnar) epiphyses from two other individuals of P. heseloni from the Lower Miocene deposits of the Kaswanga Primate Site (KPS), Rusinga Island, Kenya. Because KNM-RU 2036 and KNM-KPS individuals III and VIII are conspecific and penecontemporaneous, their comparison provides the opportunity i) to characterize, for the first time, the morphological variation of the distal radioulnar joint in a Miocene ape, P. heseloni, and ii) to investigate the functional and evolutionary implications. Our results show that the distal antebrachial epiphyses of KNM-KPS III and VIII correspond to stages of bone maturation that are more advanced than those of KNM-RU 2036 (larger articulations and sharper articular borders and ligament attachments that are more developed). Accordingly, functional interpretations based solely on the skeleton of KNM-RU 2036 have involved an underestimation of the forearm rotation abilities of P. heseloni. In particular, the KPS fossils do not exhibit the primitive morphology of distal radioulnar syndesmosis, as those of KNM-RU 2036 and most nonhominoid primates, but rather the morphology of an incipient diarthrosis (as in extant lorisines and hominoids). The distal radioulnar diarthrosis permits more mobility and maintenance of the wrist during repeated and slow rotation of the forearms, which facilitates any form of quadrupedal locomotion on discontinuous and variably oriented supports. By providing the oldest evidence of a distal radioulnar joint in an early Miocene hominoid, the main conclusions of this study are consistent with the role of cautious climbing as a prerequisite step for the emergence of positional adaptations in apes., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Objectives: Preterm infants are born with low bone mineral. Neonatal stress further impedes bone mineralization. Clinical evidence suggests that tactile and kinesthetic stimulation (TKS) improves bone phenotype and decreases stress response. Clinical and translational studies indicate the IGF-1 axis, responsible for postnatal growth and bone mineralization, is a key player. We hypothesized that TKS would attenuate the negative impact of neonatal stress on bone phenotype and the IGF-1 axis in weanling rats., Methods: Neonatal stress (STRESS) or TKS (STRESS + 10min TKS) was administered from D6 to D10. Control animals received standard care. Tissue was harvested on D21. Dual energy x-ray absorptiometry (DXA) and bone morphometry were performed and serum osteocalcin, type I procollagen N-terminal propeptide (PINP), tartrate-resistant acid phosphatase (TRAP), and bone and liver mRNA levels of IGF-1, IGF-1 receptor (IGF-1R), and growth hormone receptor (GHR) were measured., Results: Neonatal stress increased bone mineral content (BMC), area (BA), growth plate width, liver IGF-1 mRNA, and serum IGF-1. TKS maintained areal bone mineral density (aBMD) and bone specific IGF-1 and IGF-1R mRNA while STRESS decreased compared to controls., Conclusions: Neonatal stress results in apparent accelerated growth response. TKS differed from STRESS with improved tibia aBMD and increased bone specific IGF-1 mRNA.

We model trabecular bone as a nanocomposite material with hierarchical structure and predict its elastic properties at different structural scales. The analysis involves a bottom-up multi-scale approach, starting with nanoscale (mineralized collagen fibril) and moving up the scales to sub-microscale (single lamella), microscale (single trabecula) and mesoscale (trabecular bone) levels. Continuum micromechanics methods, composite materials laminate theory and finite-element methods are used in the analysis. Good agreement is found between theoretical and experimental results.

Purpose: Fusionless growth modulation is an attractive alternative to conventional treatments of idiopathic scoliosis. To date, fusionless devices achieve unilateral growth modulation by compressing the intervertebral disc. This study explores a device to control spinal alignment and vertebral morphology via growth modulation while excluding the disc in a porcine model., Methods: A device that locally encloses the vertebral growth plate exclusive of the disc was introduced anteriorly over T5-T8 in four immature pigs (experimental) while three underwent surgery without instrumentation (sham) and two were selected as controls. Bi-weekly coronal and lateral radiographs were taken over the 12-week follow-up to document vertebral morphology and spinal alignment modifications via an inverse approach (creation of deformity)., Results: All animals completed the experiment with no postoperative complications. Control and sham groups showed no significant changes in spinal alignment. Experimental group achieved a final coronal Cobb angle of 6.5° ± 3.5° (constrained to the four instrumented levels) and no alteration to the sagittal profile was observed. Solely the experimental group ended with consistent vertebral wedging of 4.1° ± 3.6° amounting to a cumulative wedging of up to 25° and a concurring difference in left/right vertebral height of 1.24 ± 1.86 mm in the coronal plane., Conclusions: The proposed intravertebral epiphyseal device, for the early treatment of progressive idiopathic scoliosis, demonstrated its feasibility by manipulating spinal alignment through the realization of local growth modulation exclusive of the intervertebral disc.

Estrogens cause growth plate closure in both males and females, by decreasing proliferation and inducing apoptosis of postproliferative growth plate chondrocytes. In vitro studies using 17β-estradiol (E(2)) conjugated to bovine serum albumin (E(2)-BSA) show that rat costochondral growth plate resting zone chondrocytes also respond to E(2). Moreover, they are regulated by E(2)-BSA via a protein kinase C and ERK MAPK signaling pathway that is functional only in female cells. To better understand how E(2) regulates apoptosis of growth plate chondrocytes, rat resting zone chondrocytes cells were treated with E(2) or E(2)-BSA. E(2) caused apoptosis in male and female resting zone and growth zone chondrocytes in a dose-dependent manner, based on elevated DNA fragmentation, terminal deoxynucleotidyl transferase dUTP nick end labeling staining and caspase-3 activation. E(2) also up-regulated p53 and Bax protein (Bcl-2-associated X protein) levels and induced release of cytochrome C from the mitochondria, indicating a mitochondrial apoptotic pathway. The apoptotic effect of E(2) did not involve elevated nitric oxide production or MAPKs. It was reduced by ICI 182780, which is an estrogen receptor (ER) antagonist and blocked by antibodies to Erα36, a membrane-associated ER. E(2)-BSA reduced cell viability and increased caspase-3 activity; ICI 182780 had no effect, but anti-ERα36 antibodies blocked the effect. The results indicate that estrogen is able to directly affect the cell population kinetics of growth plate chondrocytes by regulating apoptosis, as well as proliferation and differentiation in both resting zone and growth zone cells. They also have provided further information about the physiological functions of estrogen on longitudinal bone growth.

The objective of this experiment was to determine the normal values of bone radiographic density (BRD) by using the optical densitometry in radiographic images and the biochemical values represented by serum calcium, ash percentage, and minerals (calcium, phosphorus, and magnesium) from tibia ash of Cobb broilers at 8, 22, and 43 d of age. A total of 14 broilers were used for densitometric analysis, and 15 were used for biochemical dosages. The BRD values increased (P < 0.05) with age and in all tibia regions (proximal epiphysis, diaphysis, and distal epiphysis), concluding that growth was a determinative factor for bone performance, demanding a higher BRD during broiler development. Tibia proximal epiphysis presented higher BRD values in relation to the other bone regions (P < 0.05), as a result of a possible biomechanical adaptation to ligaments and tension of the muscle tendons at this region, allowing the support of the muscle mass increase. The serum calcium values were kept constant, as a result of the appropriate nutritional levels of the diet that supported the animal homeostasis. The bone ash and mineral percentage increased (P < 0.05) at 22 d of age, due to the higher mineral requirement in this age. The correlation between bone densitometry and the invasive techniques showed that the bone densitometry can substitute the determination of mineral percentage in the ash. This experiment presented normal values of the noninvasive and invasive methods more used in aviculture, allowing us to compare, subsequently, pathological and physiological values or results of broilers fed with different diets.

Purpose: Cycling is very popular; however, it is often believed to be associated with below average bone mass. This study compared bone measures of sprint- and distance-trained cyclists competing at World Master Track Championships, along with sedentary controls (30-82 yr), and examined the associations of bone measures with age., Methods: Radius and tibia epiphyseal and shaft bone mineral density (BMD), bone mineral content (BMC), and cross-sectional area along with shaft polar moment of resistance (RPol) and endocortical/periosteal circumferences were assessed by peripheral quantitative computed tomography. Intergroup differences were assessed by ANOVA and age relationships by correlation analyses., Results: Sprint cyclists had the largest bone shafts and bone strength surrogates; the difference in diaphyseal BMC, area, and RPol compared with controls being >or=10% in the tibia and >or=8% in the radius (P < 0.01). Distance cyclists versus control group differences were smaller (tibia: approximately 4-10%; radius: <2%), reaching statistical significance only for tibial BMC and area (P < 0.05). Generally, epiphyseal bone measures showed no group differences. In the radius, age correlations were negative for both the sprinters' and the controls' diaphyseal and the sprinters' epiphyseal BMD; they were positive for the controls' diaphyseal endocortical and periosteal circumferences (P < 0.05). In the cyclists' tibiae, neither epiphyseal nor diaphyseal bone measures were correlated with age., Conclusions: Sprint cyclists and to a lesser extent distance cyclists had greater tibia and radius bone strength surrogates than the controls, with tibial bone measures being well preserved with age in all groups. This suggests that competition-based cycling and the associated training regime is beneficial in preserving average or above-average bone strength surrogates into old age in men.

Objective: Osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage and remodeling of the subchondral bone plate, comprising calcified cartilage and underlying subchondral bone. Calcified cartilage remodeling due to upward invasion by vascular canals or to calcified cartilage erosion may contribute to biomechanical alteration of the osteochondral tissue and its subchondral bone plate component. The study hypothesis was that hydraulic conductance of osteochondral tissue and subchondral bone plate increases with structural changes indicative of increasing stages of OA., Methods: Osteochondral cores were harvested from the knees of cadaveric tissue donors and from discarded fragments from patients with OA undergoing knee surgery. The osteochondral cores from tissue donors were macroscopically normal, and the cores from patients with OA had partial-thickness or full-thickness erosion to bone. The cores were perfusion-tested to determine the hydraulic conductance, or ease of fluid flow, in their native state and after enzymatic removal of cartilage. Adjacent portions were analyzed by 3-dimensional histology for calcified cartilage, subchondral bone, and subchondral bone plate thickness and vascular canal density., Results: Hydraulic conductance of native osteochondral tissue and subchondral bone plate was higher (2,700-fold and 3-fold, respectively) in fully eroded samples than in normal samples. The calcified cartilage layer was thicker (1.5-fold) in partially eroded samples than in normal samples but thinner and incomplete in fully eroded samples. Subchondral bone plate vascularity was altered with increasing stages of OA., Conclusion: During joint loading, increased hydraulic conductance of the osteochondral tissue and subchondral bone plate could have deleterious biomechanical consequences for cartilage. Increased fluid exudation from overlying and opposing cartilage, increased fluid depressurization, and increased cartilage tissue strains could lead to chondrocyte death and cartilage damage.

Our primary objective was to validate the Bone Strength Index for compression (BSIC) by determining the amount of variance in failure load and stiffness that was explained by BSIC and bone properties at two distal sites in human cadaveric tibiae when tested in axial compression. Our secondary objective was to assess the variance in failure moment and flexural rigidity that was explained by bone properties, geometry and strength indices in the tibial diaphysis when tested in 4-point bending. Twenty cadaver tibiae pairs from 5 female and 5 male donors (mean age 74 yrs, SD 6 yrs) were measured at the distal epiphysis (4 and 10% sites of the tibial length from the distal end) and diaphysis (50 and 66% sites) by peripheral Quantitative Computed Tomography (pQCT; XCT 2000, Stratec). After imaging, we conducted axial compression tests on the distal tibia and 4-point bending tests on the diaphysis. Total bone mineral content and BSIC (product of total area and squared density of the cross-section) at the 4% site predicted 75% and 85% of the variance in the failure load and 52% and 57% in stiffness, respectively. At the diaphyseal sites 80% or more of the variance in failure moment and/or flexural rigidity was predicted by total and cortical area and content, geometry and strength indices corresponding to the axes of bending.

The anatomy of the proximal femur was studied in 35 specimens using quantitative computed tomography (QCT) and compared with anatomical sections studied by plane radiography and gross dissection. We found the primary supporting structure of the femoral head to be the primary compressive strut, which is a dense column of trabecular bone projecting from the pressure buttress of the medial femoral neck to the epiphyseal scar. Trabecular bone mushroomed from the epiphyseal scar and terminated at right angles to the cortex of the femoral head. We believe the primary compressive strut is the predominant load-bearing structure connecting the femoral head to the femoral neck, as many specimens lacked continuity of the head cortex to the femoral neck. Based on the CT number, the primary compressive strut had similar bone density to cortical structures such as the lesser trochanter, calcar femorale and posterior lateral femoral cortex. Ward's triangle lacked structural integrity in many cases, and we doubt the significance of tensile trabculae for sharing load. Surgical techniques such as femoral fracture fixation, resurfacing hip arthroplasty and allograft transplantation may benefit from this knowledge.

Background: The past years have seen the development of many synthetic bone replacements. To test their biocompatibility and ability for osseointegration, osseoinduction and -conduction requires their placement within bone preferably in an animal experiment of a higher species., Methods: A suitable experimental animal model in sheep with drill holes of 8 mm diameter and 13 mm depth within the proximal and distal humerus and femur for testing biocompatibility issues is introduced., Results: This present sheep model allows the placing of up to 8 different test materials within one animal and because of the standardization of the bone defect, routine evaluation by means of histomorphometry is easily conducted. This method was used successfully in 66 White Alpine Sheep. When the drill holes were correctly placed no complications such as spontaneous fractures were encountered., Conclusion: This experimental animal model serves an excellent basis for testing the biocompatibility of novel biomaterials to be used as bone replacement or new bone formation enhancing materials.

Mechanical stimulation is critical for bone architecture and bone mass. The aim of this study was to examine the effects of mechanical loads applied to the knee. The specific question was whether loads applied to the tibial epiphysis would enhance bone formation in the tibial diaphysis. In C57/BL/6 mice, loads of 0.5 N were applied for 3 min per day for 3 days at 5, 10, or 15 Hz. Bone samples were harvested 13 days after the last loading. The strains were measured 13 +/- 2 microstrains at 5 Hz in the diaphysis. The histomorphometric data in the diaphysis clearly showed enhanced bone formation. First, compared with nonloaded control the cross-sectional cortical area was increased by 11% at 5 Hz and 8% at 10 Hz (both P < 0.05). Second, the cortical thickness was elevated by 12% at 5 Hz (P < 0.01) and 8% at 10 Hz (P < 0.05). Third, mineralizing surface (MS/BS), mineral apposition rate (MAR), and bone formation rate (BFR/BS) were increased at 5 Hz (P < 0.01 for MS/BS; P < 0.001 for MAR and BFR/BS) and at 10 Hz (P < 0.05 for MS/BS; P < 0.01 for MAR and BFR/BS). Bone formation was enhanced more extensively in the medial side than the lateral or the posterior side. The results reveal that knee loading is an effective means to enhance bone formation in the tibial diaphysis in a loading-frequency dependent manner without inducing significant in situ strain at the site of bone formation.

Estrogen is critical for epiphyseal fusion in both young men and women. In this study, we explored the cellular mechanisms by which estrogen causes this phenomenon. Juvenile ovariectomized female rabbits received either 70 microg/kg estradiol cypionate or vehicle i.m. once a week. Growth plates from the proximal tibia, distal tibia, and distal femur were analyzed after 2, 4, 6, or 8 weeks of treatment. In vehicle-treated animals, there was a gradual senescent decline in tibial growth rate, rate of chondrocyte proliferation, growth plate height, number of proliferative chondrocytes, number of hypertrophic chondrocytes, size of terminal hypertrophic chondrocytes, and column density. Estrogen treatment accelerated the senescent decline in all of these parameters. In senescent growth plates, epiphyseal fusion was observed to be an abrupt event in which all remaining chondrocytes were rapidly replaced by bone elements. Fusion occurred when the rate of chondrocyte proliferation approached zero. Estrogen caused this proliferative exhaustion and fusion to occur earlier. Our data suggest that (i) epiphyseal fusion is triggered when the proliferative potential of growth plate chondrocytes is exhausted; and (ii) estrogen does not induce growth plate ossification directly; instead, estrogen accelerates the programmed senescence of the growth plate, thus causing earlier proliferative exhaustion and consequently earlier fusion.

Ultrasound (US) waves are mechanical vibrations that are applied to a material--bone tissue--in order to study its properties, that is, density, elasticity, and structure. In this study we evaluated in which way density and elasticity of the spongy bone influenced the transmission of 1.25 MHz US pulses. Twelve cylindrical specimens (diameter, 8 mm; height, 5 mm) excised from phalanxes of pig were decalcified with 0.5 M EDTA for different times (0, 2, and 5 days). During these periods, the samples underwent the following investigations: US transmission, density, and elasticity measurements. To assess the homogeneity of decalcification, the cross-sections of some samples were microradiographed. A detailed analysis of the US signal received was performed using velocity, Fourier analysis, and some parameters typical of signal processing technique. A good correlation was found between US velocity and density (r2 = 0.70); a lower correlation was found between velocity and elasticity (r2 = 0.59). If density and elasticity are considered simultaneously, the correlation with the US velocity improves significantly (r2 = 0.84). Fourier analysis enabled us to observe a shift of the main frequency toward lower values as the decalcification process advanced. We also observed that in the regressions weighted for density, US velocity correlated poorly with elasticity (r2 = 0.16), whereas signal processing parameters maintain a good correlation with elasticity (ultrasound peak amplitude [UPA], r2 = 0.48; slope, r2 = 0.62). In this study, it has been observed that when using a signal processing technique to analyze US pulses, it is possible to identify some parameters that are related in different ways to density and to elastic properties of bone. Our results show the potentiality of US technique to separate information on bone density and elasticity that X-ray-based densitometric methods do not provide.

The aim of this work is to study the effects of chitosan on rat knee cartilages. 0.2 ml of 0.1% chitosan solution pH = 6.9 were injected inside rat knees articular cavity. One, three and six weeks after injection, histological and histomorphometric studies were performed on undecalcified samples embedded in polymethylmetacrylate. Results show that after 1 and 6 weeks: (i) chitosan slows significantly (P < 0.005) the decrease in epiphyseal cartilage thicknesses and (ii) increases significantly articular cartilage chondrocyte densities (P < 0.002). However chitosan solution induces a proliferation of fibrous tissue with abundant fibroblasts, fibrocytes and monocytes inside the joint and this proliferation is still present after 6 weeks. This study suggests that chitosan could act on the growth of epiphyseal cartilage and wound healing of articular cartilage.

Mechanical strain maintains bone architecture even under conditions of increased bone turnover such as occurs with ovarian hormone deficiency. The rat distal femur contains two sites that apparently experience different levels of mechanical strain and therefore the rat is a suitable model for investigating such effects. The femoral epiphysis experiences higher strain energy compared with the metaphysis and we report the effects of aging between 7 and 12 months and the postovariectomy effects over the same time period on cancellous bone variables measured at these two sites. Age-related bone loss in sham-operated (Sham) animals occurred in both regions, with a greater fall in the metaphysis than in the epiphysis (trabecular bone volume [BV/TV, %] Mean [SEM] Metaphysis: day 0, 25. 9 [2.4]; day 150, 8.8 [1.3]: Epiphysis: day 0, 44.8 [1.7]; day 150, 36.7 [1.4] [p < 0.0001]). With ovariectomy (OVX) there was a 73% reduction in cancellous bone at the metaphysis compared with no specific loss at the epiphysis (BV/TV [%] OVX: Metaphysis: day 150, 2.4 [0.4] [p < 0.01 compared with Sham]: Epiphysis: day 150 29.3 [2. 7] [NS]). Osteoblast cell activity and osteoclast surface were increased after ovariectomy in both regions. The mineral apposition rate decreased at 9.5 months of age in both regions (p < 0.0001), independent of ovariectomy, and was coincident with a reduction in trabecular number in the epiphyses of both operative groups and in the metaphysis of the ovary-intact group. These data suggest that local mechanical strain governs bone balance with aging and that architectural changes resulting from age-related bone loss may mirror those following estrogen deficiency but occur via a different cellular mechanism.

We examined 32 patients with 39 lengthenings (mean 8 cm) of the lower leg 3-14 years postoperatively. 23 distraction epiphyseolyses and 16 corticotomies with callus distraction had been carried out using the llizarov system. Both procedures yielded the same healing indices and the number of problems and complications was the same in the two groups. After distraction epiphyseolysis, serious complications of the knee and greater deviations from the normal leg axis were commoner than after callus distraction, which we now prefer.

Twelve-week-old female rats were ovariectomized (OVX) and compared with sham-operated control rats at 3, 5, 7, 10, 14, 30, and 60 days postoperation with respect to the expression of type I collagen and osteopontin mRNAs, as well as bone structure and the number of osteoclasts. The trabecular number and separation were significantly decreased and increased, respectively, in the metaphyseal trabecular bones of OVX rat femurs. The number of osteoclasts was significantly increased in the same region of OVX rats at 3 and 5 days postoperation. Type I collagen mRNA was expressed in osteoblasts, and osteopontin mRNA was expressed in some osteoclasts, in mononuclear cells on the bone resorption surface, and in osteocytes near the resorption surface. In the metaphyseal trabecular bone, type I collagen and osteopontin mRNA expression levels in individual cells was initially increased in OVX rats from 7 to 10 days postoperation, and this was sustained for 60 days. The number of osteopontin mRNA-expressing osteocytes was also significantly increased at 10 days postoperation, which lasted until 60 days. In the epiphysis, an increase in type I collagen mRNA expression was initially observed in OVX rats at 14 days postoperation, which lasted until 60 days. The number of osteopontin mRNA-expressing osteocytes was virtually identical until 30 days postoperation in the epiphysis. These findings indicated that the biological activities of osteoblasts and osteocytes are stimulated in bones of the OVX rat and that the response for OVX differs between the metaphysis and epiphysis. Furthermore, the number of osteopontin mRNA-expressing osteocytes was increased only in bones that tended to be resorbed after OVX. This indicates that some osteocytes were stimulated to express osteopontin mRNA by estrogen deficiency and suggests that these osteopontin mRNA-expressing osteocytes may be involved in regulation of bone metabolism.

Epiphyseal fusion is a finding frequently analysed in hand wrist radiographs. It is tested in the bone age assessment, the gonadal dysgenesis, etc. The computerized fusion analysis is performed on an automatically selected region of interest containing the lower edge of epiphysis and the upper edge of metaphysis. In the analysis a wavelets decomposition approach is employed. The wavelets decomposition components are first subjected to a preliminary test which rejects the overexposed images whose analysis would not give reasonable results. This increases the accuracy of the algorithm and a chance for an unsupervised application. Then, a quantitative measure is found. Its value decreases while the epiphyseal fusion proceeds. The analysis yields an assignment of fusion to one of four stages: no fusion, early stage of fusion, advanced stage of fusion, and fusion completed. The results show that wavelets decomposition components may efficiently be applied to a texture analysis.

A radiological examination of both elbows and wrists of 149 Northwest Indian schoolgirls between the ages of 11 and 19 years, of middle socio-economic status, was undertaken to determine the time of fusion of the epiphyses with the metaphyses of the medial epicondyle of the humerus, the proximal end of the radius and the distal ends of both radius and ulna. The data were subjected to discriminant function tests as well as sensitivity and specificity tests. If the epiphysis of the medial epicondyle of the humerus or that of the head of the radius has not fused with metaphysis completely, the age of the girl would be < 16 years. However, the age of the girl should be > 16 years if the distal epiphyses of the radius and the ulna show complete fusion with their respective metaphyses. Discriminant function tests show that 91.95% of girls can be correctly classified as being above or below the age of 16 years.

Skeletal maturity was studied in 82 healthy adolescent girls age between 12 and 19 who had reached the menarche from 0.5 to 5 years previously. 56 subjects (68.2%) had closed epiphyses while they were open in 26 (31.8%). The incidence of open epiphyses was 61% in the first year after the menarche and progressively decreased showing negative correlation with time elapsed since the menarche (r = 0.98, P < 0.0001). The mean +/- SE height increased from 158.7 +/- 1.3 cm in the first year after the menarche, to 162.7 +/- 1.3 cm in the fifth year after the menarche. No significant differences were seen in estradiol, estrone, testosterone, dihydrotestosterone and androstenedione levels between subjects with open or closed epiphyses while dehydroepiandrosterone and its sulfate levels were higher in the closed epiphyses group. Girls with open epiphyses at first evaluation grew about 3.5 cm in the next three years.

The affinity of bone for bioinert ceramics and stainless steel was compared using calcified bone specimens. We implanted cylinders of alumina ceramics (Al2O3), zirconia ceramics (ZrO2) and SUS-316 L stainless steel into the distal femoral epiphyses of dogs and then made observations from 4 to 96 wk post-operatively. Irregularities in the histological specimens suggested the presence of artefacts due to the insertion technique. We subsequently used screws inserted into holes tapped with a tap that had a diameter identical to the screws, and observed these implants from 4 to 96 wk after insertion. There was no detectable difference in the affinity index for all three materials from 4 to 96 wk after implantation. The affinity index was calculated as the ratio of the new bone directly adjoining the implant without any intervening fibrous membrane or bone marrow to the total length of the bone-implant interface x 100%.

The topographic variation of proximal tibial cancellous bone strength was investigated in 12 knees from routine autopsies. Samples from eight knees were tested to compressive failure in an Instron material testing machine, and four knees were tested with the osteopenetrometer, an instrument developed for intraoperative measurement of bone strength. Ultimate stress, elastic modulus and energy absorption of the bone were calculated from the Instron-curves. Mechanical properties varied considerably from knee to knee, but the topographic patterns were remarkably constant. The medial condyle showed the highest strength, the intercondylar area the lowest. On the medial side the bone was strongest at the front, while on the lateral side the reverse was true. The two horizontal levels tested did not differ significantly. The osteopenetrometer measurements closely modelled the pattern of ultimate stress.

The level of pyridine nucleotides (NADH and NAD+) in the mineralizing growth plate of the chick was ascertained by high-resolution scanning microfluorimetry and biochemical analysis. Scanning electron microscopy and light microscopy were used to relate the concentrations of NADH and NAD+ to stages of chondrocyte maturation. A dramatic increase was found in the relative concentration of reduced pyridine nucleotides in the hypertrophic zone. On either side of this zone, in proliferating and calcifying cartilage, there was a decrease in NADH fluorescence, and the NADH/NAD+ ratio was depressed. The finding that NADH accumulated in the tissue zone associated with the earliest deposition of bone mineral supports the hypothesis that a change in the redox state initiates tissue mineralization.

A modular apparatus to measure the bending and torsional properties of the rat femur is presented. Both intact femur diaphyses and diaphyseal fractures in different phases of healing can be tested. It is also possible to measure the bending-strength of the distal femur metaphysis and the epiphyseal plate. The apparatus can be used to investigate the effect of drugs and hormones on the remodelling of the rat femur.

The pineal gland plays an important role in the production of melatonin and in the synchronization of the reproduction process in seasonal breeding animals. Changes in the duration of day length are the most important stimulatory factors. In humans the pineal gland may not only have a physiological role in the seasonality of maturation, but also concerning the simultaneously observable changes of serum hormone and serum melatonin levels as well as in regard of changes in hair colour in prepuberal children. Therefore the pineal gland seems to play an important physiological role in the timing of human maturation.

Proteoglycans (PGs) are closely associated with cartilage calcification. We have examined the hypertrophic zone of rat epiphyseal cartilage, in which calcification is occurring, using the high-iron diamine-thiocarbohydrazide-silver proteinate (HID-TCH-SP) method for sulfated glycosaminoglycans, an immunoferritin method specific for chondroitin sulfate A, and the tannic acid-ferric chloride (TA-Fe) method to stain cartilage matrix granules (MGs) presumed to be PG monomers. HID-TCH-SP produced stain deposits with a diameter of 11.2 +/- 3.2 nm (mean +/- SD; n = 200) in the MGs. However, HID-TCH-SP staining was not discernible in membrane-limited matrix vesicles (MVs). In areas of advanced calcification, partially disrupted MVs and globular bodies (GBs), derived in part from disrupted and/or degenerated MVs, contained a few too many small HID-TCH-SP stain deposits. Further down the epiphyseal cartilage, intact MVs markedly decreased and the GBs, containing many small HID-TCH-SP stain deposits, significantly increased in number. These GBs were found exclusively in the longitudinal septa rather than in the transverse septa. After enzyme digestion with testicular hyaluronidase, small (7.2 +/- 1.2 nm in diameter) stain deposits remained in the MGs and GBs, presumably localized to keratan sulfate. Immunoferritin localizing chondroitin sulfate strongly stained MGs, whereas MVs and GBs lacked staining. TA-Fe staining of glycoconjugates in the GBs demonstrated a striking decrease in the diameter of MGs associated with calcification in the GBs as compared with those in the noncalcifying area around the GBs. These results indicate that the GBs containing needle-like apatite crystals in morphologic preparations represent sites of chondroitin sulfate degradation. Testicular hyaluronidase-resistant sulfated glycosaminoglycans presumed to be keratan sulfate and partially degraded PGs selectively remain within the GBs as a probable requisite for expansion of the initial calcification in MVs.

To study the influence of variations in mean arterial pressures (MAP) on long bone medullary pressures, seven anaesthetized dogs were investigated. The medullary pressures were measured in the epiphyseal, the metaphyseal and the diaphyseal regions and remained rather constant when MAP was above 80 mmHg. Below this level of MAP a statistically significant (P less than 0.01) reduction of the medullary pressures was seen. Comparing the mean medullary pressures obtained with ranges of MAP of 81--100 mmHg (the control medullary pressures) and of 61--80 mmHg, the greatest decline was seen in the epiphyseal and the diaphyseal regions, from 25.2 mmHg to 8.1 mmHg and from 26.7 mmHg to 8.3 mmHg, respectively. The corresponding decrease in the metaphyseal region was from 18.9 mmHg to 10.9 mmHg. The mean values of intraosseous pressure measured by our technique were between 20--30 mmHg and this is in accordance with measurements in normal humans found by other authors.

Nucleoside triphosphate pyrophosphohydrolase (EC 3.6.1.8) activity is associated with matrix vesicles purified from collagenase digests of fetal calf epiphyseal cartilage. This enzyme hydrolyzes nucleoside triphosphates to nucleotides and PPi, the latter inducing precipitation in the presence of Ca2+ and Pi. An assay for matrix vesicle nucleoside triphosphate pyrophosphohydrolase is developed using beta, gamma-methylene ATP as substrate. The assay is effective in the presence of matrix vesicle-associated ATPase, pyrophosphatase, and alkaline phosphatase activities. A soluble nucleoside triphosphate pyrophosphohydrolase is obtained from matrix vesicles by treatment with 5 mM sodium deoxycholate. The solubilized enzyme induced the precipitation of calcium phosphate in the presence of ATP, Ca2+, and Pi. Extraction of deoxycholate-solubilized enzymes from matrix vesicles with 1-butanol destroys nucleoside triphosphate pyrophosphohydrolase activity while enhancing the specific activities of ATPase, pyrophosphatase, and alkaline phosphatase. In solutions devoid of ATP and matrix vesicles, concentrations of PPi between 10 and 100 microM induce calcification in mixtures containing initial Ca2+ X P ion products of 3.5 to 7.9 mM2. This finding plus the discovery of nucleoside triphosphate pyrophosphohydrolase in matrix vesicles supports the view that these extracellular organelles induce calcium precipitation by the enzymatic production of PPi. Nucleoside triphosphate pyrophosphohydrolase is more active against pyrimidine nucleoside triphosphates than the corresponding purine derivatives. The pH optimum is 10.0 and the enzyme is neither activated nor inhibited by Mg2+ or Ca2+ ions or mixtures of the two. Vmax at pH 7.5 for beta, gamma-methylene ATP is 0.012 mumol of substrate hydrolyzed per min per mg of protein and Km is below 10 microM. The enzyme is irreversibly destroyed at pH 4 and is stable at pH 10.5.

The in vitro breaking forces of the distal femoral growth plates of young rabbits were measured as a background to the design of a bone lengthening method, using epiphyseal distraction. The mean breaking force in 16 femora was 12.98 +/- 3.48 kg and the mean strain was 0.91 +/- 0.33 mm. The mean stress in 10 femora was 14.51 +/- 3.88 kg/cm2. The procedure was repeated, after applying a 1.0 kg dead weight to 6 femora for 24 hours and the breaking force was then 15.01 +/- 4.70 kg, with a mean strain of 0.85 +/- 0.62 mm. A further 8 rabbits then underwent epiphyseal distraction for 2 days in vivo, with 1 or 2 kg forces delivered to two parallel K wires by a pair of spring devices, whereupon the femora were removed and tested as before. The breaking force on the distracted side was now only 8.91 +/- 3.71 kg, compared with 13.99 +/- 3.40 kg on the control side. Although not fractured, these plates had obviously been weakened. The clinical implication of this is discussed.

In nine children and adolescents, the motion of the distal fibula in relation to the tibia was measured by roentgen stereophotogrammetry. Between the extremes of ankle flexion, the fibular epiphysis moved on average 1.4 mm.

When cartilage is deformed, it becomes electrically polarized. At least two mechanisms seem to underlie this phenomenon, namely, a short-duration, high-amplitude, piezoelectric-like response and a longer-duration, lower-amplitude response secondary to streaming potentials. The polarity of articular cartilage during loading could hypothetically facilitate joint lubrication.