Increased osteogenic response to exercise in metaphyseal versus diaphyseal cortical bone

Recent experimental data suggest that the anabolic response of bone to changes in physical activity and mechanical loading may vary among different skeletal elements, and even within different regions of the same bone. In order to better understand site-specific variation in bone modeling we used an experimental protocol in which locomotor activity was increased in laboratory mice with regular treadmill exercise for only 30 min/day. We predicted that the regular muscle contractions that occur during exercise would significantly increase cortical bone formation in these animals, and that the increase in cortical bone mass would vary between metaphyseal and diaphyseal regions. Cortical bone mass, density, and bone geometry were compared between these two regions using pQCT technology. Results indicate that exercise increases bone mineral content (BMC) in the mid-diaphysis by approximately 20%, whereas bone mass in the metaphyseal region is increased by approximately 35%. Endosteal and periosteal circumference at the midshaft are increased with exercise, whereas increased periosteal circumference is accompanied by marked endosteal contraction at the metaphysis, resulting in an increase in cortical area of more than 50%. These findings suggest that the osteogenic response of cortical bone to exercise varies significantly along the length of a bone, and more distal regions appear most likely to exhibit morphologic changes when loading conditions are altered.     

Blood flow in different regions of bone marrow after short-term exercise

Total and regional blood flow was studied in femoral bone marrow of rats exposed to a short-term exhaustive exercise. The 10 micron microspheres labeled with 58Co were injected into femoral artery and the subsequent radioactivity yield was measured in 3 regions of the bone marrow (proximal metaphysis, diaphysis and distal metaphysis). Following exercise a rise of blood flow in the metaphyseal regions was accompanied by a fall in the diaphyseal area. The changes did not alter, however, the total blood flow rate in bone marrow which remained constant irrespective of the heavy exercise load.     

Damage and recovery of the bone growth mechanism in young rats following 5-fluorouracil acute chemotherapy

Chemotherapy-induced bone growth arrest and osteoporosis are significant problems in paediatric cancer patients, and yet how chemotherapy affects bone growth remains unclear. This study characterised development and resolution of damage caused by acute chemotherapy with antimetabolite 5-fluorouracil (5-FU) in young rats in the growth plate cartilage and metaphyseal bone, two important tissues responsible for bone lengthening. In metaphysis, 5-FU induced apoptosis among osteoblasts and preosteoblasts on days 1-2. In growth plate, chondrocyte apoptosis appeared on days 5-10. Interestingly, Bax was induced prior to apoptosis and Bcl-2 was upregulated during recovery. 5-FU also suppressed cell proliferation on days 1-2. While proliferation returned to normal by day 3 in metaphysis, it recovered partially on day 3, overshot on days 5-7 and normalised by day 10 in growth plate. Histologically, growth plate heights decreased by days 4-5 and returned normal by day 10. In metaphysis, primary spongiosa height was also reduced, mirroring changes in growth plate thickness. In metaphyseal secondary spongiosa, a reduced bone volume was observed on days 7-10 as there were fewer but more separated trabeculae. Starting from day 4, expression of some cartilage/bone matrix proteins and growth factors (TGF-beta1 and IGF-I) was increased. By day 14, cellular activity, histological structure and gene expression had returned normal in both tissues. Therefore, 5-FU chemotherapy affects bone growth directly by inducing apoptosis and inhibiting proliferation at growth plate cartilage and metaphyseal bone; after the acute damage, bone growth mechanism can recover, which is associated with upregulated expression of matrix proteins and growth factors.     

Morphology of epiphyseal apparatus of a ranid frog (Rana Esculenta)

Morphological, histochemical and ultrastructural investigations on epiphyseal apparatus of Rana Esculenta were made. The most important findings were the following: 1) metaphyseal cartilage is localized inside proximal diaphyseal compact bone as a plug; 2) metaphyseal cartilage do not reduce in thickness during ageing; 3) metaphyseal cartilage do not show vascular invasion and do not mineralize in degenerative zone; 4) trabecular bone was not at all evident in this animal; 5) external periosteum is well vascularized and proliferates in correspondence to marginal epiphyseal end of the diaphyseal. From these results the hypothesis that the ranid frog bone growth is not due to metaphyseal metabolism (as in avian and mammals) but to bone periosteal marginal mineralization is reached.     

Declining tibial curvature parallels ∼6150 years of decreasing mobility in central european agriculturalists

Long bones respond to mechanical loading through functional adaptation in a suite of morphological characteristics that together ensure structural competence to in vivo loading. As such, adult bone structure is often used to make inferences about past behavior from archaeological remains. However, such biomechanical approaches often investigate change in just one aspect of morphology, typically cross‐sectional morphology or trabecular structure. The relationship between longitudinal bone curvature and mobility patterns is less well understood, particularly in the tibia, and it is unknown how tibial curvature and diaphyseal cross‐sectional geometry interact to meet the structural requirements of loading. This study examines tibial curvature and its relationship with diaphyseal cross‐sectional geometry (CSG) and body size in preindustrial Central Europeans spanning ～6150 years following the introduction of agriculture in the region. Anteroposterior centroid displacement from the proximo‐distal longitudinal axis was quantified at nine diaphyseal section locations (collectively representative of diaphyseal curvature) in 216 tibial three‐dimensional laser scans. Results documented significant and corresponding temporal declines in midshaft centroid displacement and CSG properties. Significant correlations were found between mid‐diaphyseal centroid displacement and all mobility‐related CSG properties, while the relationship weakened toward the diaphyseal ends. No significant relationship was found between centroid displacement and body size variables with the exception of the most distal section location. Results support a relationship between tibial curvature and cross‐sectional geometry among prehistoric Central European agricultural populations, and suggest that changes in mechanical loading may have influenced a suite of morphological features related to bone adaptation in the lower limb. Am J Phys Anthropol 157:260–275, 2015. © 2015 Wiley Periodicals, Inc.     

The physical and mechanical effects of suspension-induced osteopenia on mouse long bones.

The present investigation addresses the extent of tail-suspension effects on the long bones of mice. The effects are explored in both sexes, in both forelimb and hindlimb bones, and in both diaphyseal and metaphyseal/epiphyseal bones. Two weeks of suspension provided unloading of the femora and tibiae and an altered loading of the humeri. Whole-bone effects included lower mass (approximately 10%) and length (approximately 4%) in the bones of suspended mice compared to controls. The geometric and material properties of the femora were considered along the entire length of the diaphysis and in the metaphysis/epiphysis portions as a unit. Geometric effects included lower cross-sectional cortical area (16%), cortical thickness (25%) and moment of inertia (21%) in the femora of suspended mice; these differences were observed in both distal and proximal portions of the femur diaphysis. The relative amount of bone comprising the middle 8 mm of the diaphysis was greater (3%) in the control mice than in the suspended mice. Significant mass differences between the group in the metaphysis/epiphysis were not observed. Material effects included lower %ash (approximately 2%) in the femora and tibiae as well as in the humeri of suspended mice compared to controls. With respect to the measured physical and material properties, suspension produced similar bone responses in male and female mice. The effects of suspension are manifested largely through geometric rather than through material changes.     

Ontogenetic adaptation to bipedalism: age changes in femoral to humeral length and strength proportions in humans, with a comparison to baboons

The increase in lower/upper limb bone length and strength proportions in adult humans compared to most other anthropoid primates is commonly viewed as an adaptation to bipedalism. The ontogenetic development of femoral to humeral proportions is examined here using a longitudinal sample of 20 individuals measured radiographically at semiannual or annual intervals from 6 months of age to late adolescence (a subset of the Denver Growth Study sample). Anthropometric data (body weights, muscle breadths) were also available at each examination age. Results show that while femoral/humeral length proportions close to those of adults are already present in human infants, characteristically human femoral/humeral diaphyseal strength proportions only develop after the adoption of bipedalism at about 1 year of age. A rapid increase in femoral/humeral strength occurs between 1 and 3 years, followed by a slow increase until mid-late adolescence, when adult proportions are reached. When age changes in material properties are factored in, femoral strength shows an almost constant relationship to body size (body mass.bone length) after 5 years of age, while humeral strength shows a progressive decline relative to body size. Femoral/humeral length proportions increase slightly throughout growth, with no apparent change in growth trajectory at the initiation of walking, and with a small decline in late adolescence due to later growth in length of the humerus. A sex difference in femoral/humeral strength proportions (females greater) but not length proportions, develops early in childhood. Thus, growth trajectories in length and strength proportions are largely independent, with strength proportions more responsive to actual changes in mechanical loading. A cross-sectional ontogenetic sample of baboons (n=30) illustrates contrasting patterns of growth, with much smaller age changes in proportions, particularly strength proportions, although there is some indication of an adaptation to altered limb loadings early in baboon development.     

Dating and rating of Harris's lines

Well-preserved skeletons of 50 Dutch whalers from the 17th and 18th centuries, buried on the island of Zeeusche Uytkyck , were excavated during the Spitsbergen Expedition 1980. In order to estimate the biological age of these skeletons at the time that Harris's lines were formed, a method which was developed from routine X-ray data on tibial growth was tested. Formation ages of diaphyseal lines and strong metaphyseal lines appear to correspond well with hypoplastic enamel deformations within the same individuals. Because of the involvement of both the bone and tooth developmental systems, and of the resistance of these lines to bone remodeling, it can be assumed that they generally are the result of serious health insults.     

Mechanical properties of metaphyseal bone in the proximal femur

We used a three-point bending test to investigate the structural behavior of 123 rectangular flat plate specimens harvested from the metaphyseal shell of the cervical and intertrochanteric regions of five fresh/frozen human proximal femora. For comparison purposes, 36 specimens of similar geometry were also fabricated from bone of the femoral diaphysis. All specimens were oriented in either the local longitudinal or transverse directions. The mean longitudinal elastic modulus was 9650 +/- 2410 (SD) MPa and demonstrated a 24% decrease from that measured for the diaphysis (12500 +/- 2140 MPa) using the same testing technique. However, the transverse elastic moduli did not differ significantly between the proximal (5470 +/- 1720 MPa) and diaphyseal (5990 +/- 1520 MPa) specimens. The globally averaged values for the ultimate tensile strengths of the metaphyseal shell were 101 +/- 26 MPa in the longitudinal and 50 +/- 12 MPa in the transverse directions. These compared with diaphyseal values of 128 +/- 16 MPa and 47 +/- 12 MPa, respectively. While these differences were largely due to the reduced density of the proximal specimens, a slight decrease in transverse anisotropy for the proximal specimens was also noted by comparing the ratio of longitudinal to transverse moduli (1.76) and tensile strength (2.02) to the diaphyseal values (2.09 and 2.71, respectively). Use of these data should lead to improved performance of analytical models for the proximal femur, and thus help focus increased attention on the structural contribution of trabecular bone to the strength and rigidity of the proximal femur.     

Comparison of diaphyseal growth between the Libben population and the Hamann-Todd chimpanzee sample

The differences in limb lengths and proportions between humans and chimpanzees are widely known. Humans have relatively shorter forelimbs and longer hind limbs than chimpanzees. Humans have a longer period of long bone formation than chimpanzees. Recent advances in estimating age-at-death in chimpanzees from their dentition have allowed us to reexamine long bone growth in chimpanzees using their skeletal remains and compare it with similar data for humans. A chronological normalization procedure allowing direct interspecific comparison of long bone growth is presented. The preadult chimpanzee sample (n = 43) is from the Hamann-Todd Osteological Collection from the Cleveland Museum of Natural History. All human specimens (n = 202) are from the late Woodland Libben Population currently housed at Kent State University. Relying on these cross-sectional data, we conclude that both species elongate their femora at similar absolute (length per unit time) but different relative (length relative to normalized dental age) rates. The species differ in the absolute growth rate of the humerus but share a common normalized rate of growth. Forelimb segment proportion differences between species are due to differential elongation rates of the segments. Hind limb diaphyseal proportions are the same in both species, which suggests that changes in segment length are proportional. Therefore, alternative developmental mechanisms exist in these closely related species which can produce changes in limb length. © 1996 Wiley-Liss, Inc.     

The effects of socioeconomic status on endochondral and appositional bone growth,and acquisition of cortical bone in children from 19th century Birmingham,England

Endochondral growth, appositional growth, and acquisition of cortical bone thickness in the femur are investigated in subadult skeletons (N = 43, dental age range birth to 12 years) from the 19^th‐century AD burial site of St. Martin's churchyard, Birmingham, England. Endochondral growth is monitored using diaphyseal femoral length. Appositional growth is monitored using radiographic midshaft mediolateral width and acquisition of cortical bone using combined mediolateral cortical thickness measured at the midshaft from radiographs. The methodology involves plotting these variables against dental age. Growth is compared in children of differing socioeconomic status. Higher and lower status individuals are identified in the assemblage by their burial in brick vaults in the case of the former and in earth‐cut graves in the case of the latter. The relationships between bone dimensions and dental age are described using a polynomial regression procedure, and analysis of regression residuals is used to evaluate differences in bone dimension‐for‐dental age between the two status groups. Results show that lower socioeconomic status individuals had lower cortical thickness‐for‐dental age than those of higher status. This was interpreted as likely reflecting poorer nutrition in the children of lower socioeconomic backgrounds. There was no patterning with respect to socioeconomic status in femur diaphyseal length or midshaft width. The results support the idea that, for skeletal populations, growth in cortical thickness may be a more sensitive indicator of adverse conditions in childhood than growth in bone length or width. Am J Phys Anthropol, 2009. © 2009 Wiley‐Liss, Inc.     

Adaptive changes in micromechanical environments of cancellous and cortical bone in response to in vivo loading and disuse

The skeleton accommodates changes in mechanical environments by increasing bone mass under increased loads and decreasing bone mass under disuse. However, little is known about the adaptive changes in micromechanical behavior of cancellous and cortical tissues resulting from loading or disuse. To address this issue, in vivo tibial loading and hindlimb unloading experiments were conducted on 16-week-old female C57BL/6J mice. Changes in bone mass and tissue-level strains in the metaphyseal cancellous and midshaft cortical bone of the tibiae, resulting from loading or unloading, were determined using microCT and finite element (FE) analysis, respectively. We found that loading- and unloading-induced changes in bone mass were more pronounced in the cancellous than cortical bone. Simulated FE-loading showed that a greater proportion of elements experienced relatively lower longitudinal strains following load-induced bone adaptation, while the opposite was true in the disuse model. While the magnitudes of maximum or minimum principal strains in the metaphyseal cancellous and midshaft cortical bone were not affected by loading, strains oriented with the long axis were reduced in the load-adapted tibia suggesting that loading-induced micromechanical benefits were aligned primarily in the loading direction. Regression analyses demonstrated that bone mass was a good predictor of bone tissue strains for the cortical bone but not for the cancellous bone, which has complex microarchitecture and spatially-variant strain environments. In summary, loading-induced micromechanical benefits for cancellous and cortical tissues are received primarily in the direction of force application and cancellous bone mass may not be related to the micromechanics of cancellous bone.     

Bone modeling response to voluntary exercise in the hindlimb of mice

The functional adaptation of juvenile mammalian limb bone to mechanical loading is necessary to maintain bone strength. Diaphyseal size and shape are modified during growth through the process of bone modeling. Although bone modeling is a well-documented response to increased mechanical stress on growing diaphyseal bone, the effect of proximodistal location on bone modeling remains unclear. Distal limb elements in cursorial mammals are longer and thinner, most likely to conserve energy during locomotion because they require less energy to move. Therefore, distal elements are hypothesized to experience greater mechanical loading during locomotion and may be expected to exhibit a greater modeling response to exercise. In this study, histomorphometric comparisons are made between femora and tibiae of mice treated with voluntary exercise and a control group (N = 20). We find that femora of exercised mice exhibit both greater bone growth rates and growth areas than do controls (P < 0.05). The femora of exercised mice also have significantly greater cortical area, bending rigidity, and torsional rigidity (P < 0.05), although bending and torsional rigidity are comparable when standardized by bone length. Histomorphometric and cross-section geometric properties of the tibial midshaft of exercised and control mice did not differ significantly, although tibial length was significantly greater in exercised mice (P < 0.05). Femora of exercised mice were able to adapt to increased mechanical loading through increases in compressive, bending, and torsional rigidity. No such adaptations were found in the tibia. It is unclear if this is a biomechanical adaptation to greater stress in proximal elements or if distal elements are ontogenetically constrained in a tradeoff of bone strength of distal elements for bioenergetic efficiency during locomotion.     

Estrogen, exercise, and the skeleton

Patterns of variation in bone size and shape provide crucial data for reconstructing hominin paleobiology, including ecogeographic adaptation, life history, and functional morphology. Measures of bone strength, including robusticity (diaphyseal thickness relative to length) and cross-sectional geometric properties such as moments of area, are particularly useful for inferring behavior because bone tissue adapts to its mechanical environment. Particularly during skeletal growth, exercise-induced strains can stimulate periosteal modeling so that, to some extent, bone thickness reflects individual behavior. Thus, patterns of skeletal robusticity have been used to identify gender-based activity differences, temporal shifts in mobility, and changing subsistence strategies. Although there is no doubt that mechanical loading leaves its mark on the skeleton, less is known about whether individuals differ in their skeletal responses to exercise. For example, the potential effects of hormones or growth factors on bone-strain interactions are largely unexplored. If the hormonal background can increase or decrease the effects of exercise on skeletal robusticity, then the same mechanical loads might cause different degrees of bone response in different individuals. Here I focus on the role of the hormone estrogen in modulating exercise-induced changes in human bone thickness.     

Parathyroid hormone (1-34) regulates integrin expression in vivo in rat osteoblasts.

Intermittent administration of parathyroid hormone (PTH) activates new sites of bone formation by stimulating osteoblast differentiation and function resulting in an increase in bone mass. Because integrins have been shown to play a crucial role in osteoblast differentiation and bone formation, in the present study, we evaluated whether human PTH (1-34) upon administration to rats, influenced integrin expression in osteoblastic cells isolated from the metaphysis and the diaphysis of rat long bones. Initial immunohistochemical evaluation of bone sections demonstrated that the osteoblasts expressed at least alphav, alpha2, alpha3, and alpha5beta1 integrins. Immunocolocalization studies for integrins and vinculin established that alphav, alpha2, and alpha5beta1, but not alpha3 integrins were present in the focal adhesion sites of osteoblasts attached to FN coated surfaces. Osteoprogenitor cells isolated from metaphyseal (but not diaphyseal) marrow of rats injected with intermittent PTH (1-34) exhibited greater alphav and reduced alpha2 levels, with no apparent changes in alpha3, and alpha5beta1 integrin levels, as assessed by immunohistochemistry, Northern, and Western blot analyses. However, these changes were not observed on the same cells treated with PTH in vitro. These observations suggest that integrin modulation by PTH is likely to be indirect and that selective phenotypic expression of integrin subtypes is part of the cascade of events that lead to PTH (1-34) mediated osteoblast differentiation.     

Effects of intermittent parathyroid hormone (PTH) administration on SOST mRNA and protein in rat bone

Summary Sclerostin, the secreted protein product of the SOST gene, which is mainly expressed by osteocytes, has recently been proposed as a negative regulator of bone osteoblastogenesis. Chronic elevation of PTH reduces SOST expression by osteocytes, while controversial results have been obtained by intermittent PTH administration. We have investigated the effects of intermittently administered PTH on SOST expression and sclerostin localization, comparing them with those of controls, as they appeared in three different bone segments of rat tibia: secondary trabecular metaphyseal and epiphyseal bone, and cortical diaphyseal bone. The histomorphometric results demonstrate that PTH enhances bone turnover through anabolic effects, as shown by the association of increased bone resorption variables with a significant rise in BV/TV, Tb.Th and Tb.N and a fall in Tb.Sp. PTH induces a SOST mRNA and protein fall in secondary metaphyseal trabeculae, diaphyseal bone and in epiphyseal trabeculae. Numbers of sclerostin immunopositive osteocytes/mm² show no change, compared with controls; there are fewer sclerostin-positive osteocytes in secondary metaphyseal trabeculae than in the other two bone areas, both in the control and PTH groups. The low numbers of sclerostin-positive osteocytes in the metaphyseal trabecular bone seem to be directly related to the fact that this area displays a high remodeling rate. The anabolic effects of PTH are in line with the fall of SOST mRNA and protein in all the three bone segments examined; the rise of bone turnover supports a negative role of SOST in bone formation.     

Determination of crown-rump length from fetal long bones: humerus and femur

Lengths of the ossified diaphysis of the humerus and femur were measured in 50 fetuses ranging from 65 to 290 mm Crown-Rump length. A significant correlation was found between the diaphyseal length and the CR length. No appreciable difference was noted in diaphyseal lengths of the humerus and femur in fetuses of 85 mm CR length or below. Various growth phases were observed. The difference in length of the femur and humerus is due to a comparative slow growth of the humerus during the later period of prenatal life. Diaphyseal growth rates of the humerus and femur are 0.18 mm and 0.21 mm respectively for every 1 mm increase in CR length. Regression coefficients for the lengths of the ossified shaft of the humerus and femur are 5.35 and 5.00 respectively. With the help of these coefficients, CR length of the fetus was estimated within a range of ± 15 mm. From the estimated CR length, age of the fetus was determined with the help of a standard age and size curve with reasonable accuracy.     

Development and growth of long bones in European water frogs (Amphibia: Anura: Ranidae), with remarks on age determination

Differentiation and development of long bones were studied in European water frogs: Rana lessonae, R. ridibunda, and R. esculenta. The study included premetamorphic larvae (Gosner Stage 40) to frogs that were 5 years old. Femora, metatarsal bones, and proximal phalanges of the hindlimb exhibit the same pattern of periosteal bone differentiation and the same pattern of growth. Longitudinal and radial growth of these bones was studied by examination of the diaphyses and epiphyses, particularly where the edge of periosteal bone is inserted into the epiphysis. The periosteum seems to be responsible for both longitudinal and radial growth. Investigation of the formation, length, and arrangement of lines of arrested growth reveals that the first line is present only in the middle 25-35% of the length of the diaphysis of an adult bone; therefore, only the central portion of the diaphysis should be used for age estimation in skeletochronological studies. Comparison of the shapes and histological structures of epiphyses in the femur, metatarsal bones, and phalanges revealed that epiphyseal cartilages are composed of an inner and outer part. The inner metaphyseal cartilage has distinct zones and plugs the end of the periosteal bone cylinder; its role in longitudinal growth is questioned. The outer epiphyseal cartilage is composed of articular cartilages proper, in addition to lateral articular cartilages. Differences in the symmetry of the lateral articular cartilages of distal epiphyses of the femur and toes may reflect adaptations to different kinds of movements at the knee and in the foot.     

Relationship between formation of the femoral bicondylar angle and trochlear shape: independence of diaphyseal and epiphyseal growth

During hominin evolution, an increase in the femoral bicondylar angle was the initial change that led to selection for protuberance of the lateral trochlear lip and the elliptical profile of the lateral condyle. No correlation is found during ontogeny between the degree of femoral obliquity and of the prominence of the lateral trochlear lip. Might there be a relationship with the elliptical profile of the lateral condyle? On intact femoral diaphyses of juvenile humans and great apes, we compared the anteroposterior length of the lateral and medial sides of the distal metaphysis. The two diaphyseal pillars remain equal during postnatal growth in great apes, while the growth of the lateral pillar far exceeds that of the medial pillar in humans. Increase in bicondylar angle is correlated with disproportionate anteroposterior lengthening of the lateral pillar. The increased anteroposterior length of the lateral side of the metaphysis would contribute to increasing the radius of the curvature of the lateral condyle, but not to the projection of the lateral trochlear lip. The similar neonatal and adult femoro‐patellar joint shape in humans prompted an assessment of the similarity during growth of the entire neonatal and adult epiphyses. We showed that the entire epiphysis undergoes drastic changes in proportions during postnatal growth. Finally, we emphasize the need to distinguish the cartilaginous phenotype and the ossified phenotype of the distal femoral epiphysis (and of any epiphysis) during postnatal growth. This crucial distinction applies to most postcranial bones, for they almost all develop following the process of endochondral ossification. Am J Phys Anthropol, 2006. © 2006 Wiley‐Liss, Inc.