The benefit of combining non-mechanical agents with mechanical loading: a perspective based on the Utah Paradigm of Skeletal Physiology

The Utah Paradigm of Skeletal Physiology with its key component, the mechanostat hypothesis, suggest plausible explanations of some of the tissue-level changes occurring from combining selected non-mechanical agents (anabolic and anti-resorptive/( re)modeling agents) with mechanical loading (osteogenic exercise) to increase bone mass and strength. The evidence for combining selected anabolic agents like parathyroid hormone, prostaglandin E(2), growth hormone, etc. with mechanical loading can increase bone mass is strong. Anabolic agents influence loading-related bone formation changes in a permissive manner and modulate (increase) the responsiveness of bone tissue to mechanical loading by changing thresholds for bone formation and resorption. However, any beneficial effect of combining selected anti-resorptive/(re)modeling agents like estrogen with loading is marginal, especially in adult skeletons. Postulated changes in modeling and remodeling thresholds (set points) and known direct effects on bone cells by non-mechanical agents may explain the observed tissue-level changes associated with large and minor increases in bone mass. Although the pharmaceutical industry has avoided considering osteogenic loading in the treatment of osteoporosis, a methodical dose-response study of anabolic agents combined with loading should: (1) provide opportunities for therapeutic intervention to imitate or enhance the osteogenic response to loading in order to correct osteopenias; (2) provide the potential to diminish the dosage of drugs required to induce bone formation in ways that enhanced efficacy and reduced any side effects; and (3) improve the quality of life and reduce the risk of falls by improving balance, gait speed and muscle strength with a non-mechanical agent like GH that could improve both muscle and bone mass and strength. Lastly, more studies are needed which determine bone strength instead of only "mass" in aged skeletons so one can assess how effective such treatments would reduce the risk of fracture in the clinic.     

Increased intracortical bone remodeling during lactation in beagle dogs

There are substantial changes in skeletal and mineral metabolism during pregnancy and lactation. The purpose of this study was to determine the changes in intracortical bone remodeling and turnover during lactation in beagle dogs. A femur and rib were obtained from dogs near the end of lactation or soon after weaning and compared with nonlactating controls. Rib cortical bone had much higher bone turnover rates than did femoral diaphyseal cortical bone. The number of single-labeled osteons and the number of resorption spaces were significantly greater during lactation in both the rib and the femur. Additionally, the mineral apposition rate, basic multicellular unit activation frequency, and bone turnover rates were greater in the femoral cortical bone from the lactating dogs than from the controls. These data demonstrate that during lactation, intracortical bone remodeling increases, and this may provide a mechanism for the skeleton to be responsive to the calcium requirements of the mother. In addition, these data may help explain the transient decreases in cortical bone mineral density that are reported to occur during human lactation.     

Low bone mass may not be the only cause of skeletal fragility in osteoporosis

Skeletal fragility in postmenopausal osteoporosis is not due solely to reduction in bone mass. This fact explains some of the overlap in bone mineral measurements observed between patients who are fracturing and age- and sex-matched normals who are not. Changes in skeletal architecture and bone remodeling occur with age which can account for some of the fragility. These changes are exaggerated in patients with postmenopausal osteoporosis who are suffering spine fractures. Three abnormalities have been described by histomorphometric methods which can account for skeletal fragility out of proportion to the degree of bone loss. They are: (i) loss of trabecular connectivity such that vertical weight-bearing bars lose their cross-attachments with each other, thus becoming susceptible to buckling; (ii) inefficient and prolonged microdamage repair due to periods of pause in the formation phase of remodeling; and (iii) accumulation of unrepaired microdamage in unremodeled bone tissue in the central part of trabeculae due to reduced osteon wall thickness coupled with maintenance of trabecular thickness. Recognition of these abnormalities should broaden our approach to the study of skeletal fragility in the syndrome of postmenopausal osteoporosis.     

Cancellous and cortical bone mechanical properties and tissue dynamics during pregnancy, lactation, and postlactation in the rat

There are substantial changes in maternal skeletal dynamics during pregnancy, lactation, and after lactation. The purpose of this study was to correlate changes in cortical and cancellous bone mass, structure, and dynamics with mechanical properties during and after the first reproductive cycle in rats. Rats were mated and groups were taken at parturition, end of lactation and 8 wk after weaning, and were compared with age-matched, nulliparous controls. Measurements were taken on femoral cortical bone and lumbar vertebral body cancellous bone. At the end of pregnancy, there was an increase in cortical periosteal bone formation and an increase in cortical volume, but a suppression of turnover in cancellous bone with no change in cancellous or cortical mechanical properties. Lactation was associated with a decrease in cortical and cancellous bone strength with a decrease in bone volume, but an increase in turnover on cancellous and endocortical surfaces. After lactation, there was a partial or full restoration of mechanical properties. This study demonstrates substantial changes in bone mechanics that correlate with changes in bone structure and dynamics during the first reproductive cycle in rats. The greatest changes were observed during the lactation period with partial or full recovery in the postlactational period.     

Effects of reproduction on sexual dimorphisms in rat bone mechanics

Osteoporosis most commonly affects postmenopausal women. Although men are also affected, women over 65 are 6 times more likely to develop osteoporosis than men of the same age. This is largely due to accelerated bone remodeling after menopause; however, the peak bone mass attained during young adulthood also plays an important role in osteoporosis risk. Multiple studies have demonstrated sexual dimorphisms in peak bone mass, and additionally, the female skeleton is significantly altered during pregnancy/lactation. Although clinical studies suggest that a reproductive history does not increase the risk of developing postmenopausal osteoporosis, reproduction has been shown to induce long-lasting alterations in maternal bone structure and mechanics, and the effects of pregnancy and lactation on maternal peak bone quality are not well understood. This study compared the structural and mechanical properties of male, virgin female, and post-reproductive female rat bone at multiple skeletal sites and at three different ages. We found that virgin females had a larger quantity of trabecular bone with greater trabecular number and more plate-like morphology, and, relative to their body weight, had a greater cortical bone size and greater bone strength than males. Post-reproductive females had altered trabecular microarchitecture relative to virgins, which was highly similar to that of male rats, and showed similar cortical bone size and bone mechanics to virgin females. This suggests that, to compensate for future reproductive bone losses, females may start off with more trabecular bone than is mechanically necessary, which may explain the paradox that reproduction induces long-lasting changes in maternal bone without increasing postmenopausal fracture risk.     

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.     

Label‐free quantitative proteome analysis of skeletal tissues under mechanical load

Skeletal tissue has the capability to adapt its mass and structure in response to mechanical stress. However, the molecular mechanism of bone and cartilage to respond to mechanical stress are not fully understood. A label‐free quantitative proteome approach was used for the first time to obtain a global perspective of the response of skeletal tissue to mechanical stress. Label‐free quantitative analysis of 1D‐PAGE‐LC/MS/MS based proteomics was applied to identify differentially expressed proteins. Differential expression analysis in the experimental groups and control group showed significant changes for 248 proteins including proteins related to proliferation, differentiation, regulation of signal transduction and energy metabolic pathways. Fluorescence labeling by incorporation of alizarin/calcein in newly formed bone minerals qualitatively demonstrated new bone formation. Skeletal tissues under mechanical load evoked marked new bone formation in comparison with the control group. Bone material apposition was evident. Our data suggest that 39 proteins were assigned a role in anabolic process. Comparisons of anabolic versus catabolic features of the proteomes show that 42 proteins were related to catabolic. In addition, some proteins were related to regulation of signal transduction and energy pathways, such as tropomyosin 4, fibronectin 1, and laminin, might be new molecular targets that are responsive to mechanical force. Differentially expressed proteins identified in this model may offer a useful starting point for elucidating novel aspects of the effects of mechanical force on skeletal tissue. J. Cell. Biochem. 108: 600–611, 2009. © 2009 Wiley‐Liss, Inc.     

骨再生的力学生物学问题

生物力学主要探讨力学刺激与细胞的形态、结构和功能之间的关系。骨组织改变其形态和结构以适应力学刺激,表现为骨的适应性重建。骨的生长是骨塑形和骨重建两个过程协同作用的结果,以调整骨的形状、大小和组成,适应其所处的力学环境。骨组织工程的目的就是修复骨组织的正常生物力学功能。近年来,骨组织工程的研究主要集中于模拟骨生长的在体生理条件,从而刺激细胞形成有功能的骨组织。生物反应器能够模拟体内生理状态,为种子细胞在生物支架材料上生长提供一个适宜的力学环境。     

Parathyroid Hormone (PTH)/PTH-related Peptide Type 1 Receptor (PPR) Signaling in Osteocytes Regulates Anabolic and Catabolic Skeletal Responses to PTH

Parathyroid hormone (PTH) is the only Food and Drug Administration-approved anabolic agent to treat osteoporosis; however, the cellular targets of PTH action in bone remain controversial. PTH modulates bone turnover by binding to the PTH/PTH-related peptide (PTHrP) type 1 receptor (PPR), a G-protein-coupled receptor highly expressed in bone and kidneys. Osteocytes, the most abundant cells in adult bone, also express PPR. However, the physiological relevance of PPR signaling in osteocytes remains to be elucidated. Toward this goal, we generated mice with PPR deletion in osteocytes (Ocy-PPRKO). Skeletal analysis of these mice revealed a significant increase in bone mineral density and trabecular and cortical bone parameters. Osteoblast activities were reduced in these animals, as demonstrated by decreased collagen type I α1 mRNA and receptor activator of NF-κB ligand (RANKL) expression. Importantly, when subjected to an anabolic or catabolic PTH regimen, Ocy-PPRKO animals demonstrated blunted skeletal responses. PTH failed to suppress SOST/Sclerostin or induce RANKL expression in Ocy-PPRKO animals compared with controls. In vitro, osteoclastogenesis was significantly impaired in Ocy-PPRKO upon PTH administration, indicating that osteocytes control osteoclast formation through a PPR-mediated mechanism. Taken together, these data indicate that PPR signaling in osteocytes is required for bone remodeling, and receptor signaling in osteocytes is needed for anabolic and catabolic skeletal responses.     

Gap junctions in skeletal development and function

Gap junctions play a critical role in the coordinated function and activity of nearly all of the skeletal cells. This is not surprising, given the elaborate orchestration of skeletal patterning, bone modeling and subsequent remodeling, as well as the mechanical stresses, strains and adaptive responses that the skeleton must accommodate. Much remains to be learned regarding the role of gap junctions and hemichannels in these processes. A common theme is that without connexins none of the cells of bone function properly. Thus, connexins play an important role in skeletal form and function.     

Sympathectomy alters bone architecture in adult growing rats

Sympathetic nervous system (SNS) fibres and alpha- and beta-receptors are present in bone, indicating that the SNS may participate in bone metabolism. The importance of these observations is controversial because stimulation or inhibition of the SNS has had various effects upon both anabolic and catabolic activity in this tissue. In this study we evaluated the effects of pharmacological sympathectomy, using chronic treatment of maturing male rats with 40 mg of guanethidine/kg i.p., upon various parameters in bone. Double labelling with tetracycline injection was also performed 20 and 2 days before sacrifice. Bone mass, mineral content, density and histomorphometric characteristics in different skeletal regions were determined. Bone metabolic markers included urinary deoxypyridinoline and serum osteocalcin measurements. Guanethidine significantly reduced the accretion of lumbar vertebral bone and of mineral content and density, compared to controls. Femoral bone mineral content and density were also significantly reduced, compared to controls. Histomorphometric analyses indicated these effects were related to a reduction of cortical bone and mineral apposition rate at femoral diaphysials level. Both markers of bone metabolism were reduced in controls as they approached maturity. Guanethidine significantly decreased serum osteocalcin compared to controls, while urinary deoxypyridinoline was unchanged. These data indicate that guanethidine-induced sympathectomy caused a negative balance of bone metabolism, leading to decreased mass by regulating deposition rather than resorption during modeling and remodeling of bone.     

Physiological Factors Responsible for the Development of Osteopenia under Conditions of Mechanical Unloading

A summary of results of investigations by the author and a brief review of some literature data on human bone tissue deprived of mechanical loading (spaceflight, hypokinesia) is given. The direction and markedness of changes in bone mass—the bone mineral density and the bone mineral content—in different skeletal segments depend on their position relative to the gravity vector. A theoretically expected bone mass reduction was revealed in the trabecular structures of the bones of the lower part of the skeleton (local osteopenia). In the upper part of the skeleton, an increase in the bone mineral content is observed, which is considered as a secondary response and is due to redistribution of body fluids cephalad. The main cause of osteopenia is mechanical unloading. Arguments are presented that osteocyte osteolysis, delayed osteoblast histogenesis, and osteoclast resorption provoked by rearrangement in the hierarchy of the systems of volume regulation, ion regulation, and the endocrine regulation of calcium homeostasis are the main mechanisms of osteopenia.     

Thyroid and bone

The hypothalamic-pituitary-thyroid axis plays a key role in skeletal development, acquisition of peak bone mass and regulation of adult bone turnover. Euthyroid status is essential for maintenance of optimal bone mineralization and strength. In population studies, hypothyroidism and hyperthyroidism have both been associated with an increased risk of fracture. Furthermore, recent studies in healthy euthyroid post-menopausal women indicate that thyroid status in the upper normal range is also associated with low bone mineral density and an increased risk of non-vertebral fracture. Studies in mutant mice have demonstrated that thyroid hormone receptor α is the major mediator of T3 action in bone and that thyroid hormones exert anabolic actions during growth but have catabolic effects on the adult skeleton. Nevertheless, TSH has also been proposed to be a direct negative regulator of bone turnover, although the relative importance of T3 and TSH actions in the skeleton has yet to be clarified.     

The Effect of Locomotion on the Mobilization of Minerals from the Maternal Skeleton

Bone is a dynamic tissue from which minerals are deposited or withdrawn according to the body’s demand. During late pregnancy and lactation, female mammals mobilize mineral from bone to support the ossification of offspring skeleton(s). Conversely, in response to mechanical loading, minerals are deposited in bone enabling it to develop a stronger architecture. Despite their central importance to reproductive performance and skeletal integrity, the interactions between these potentially opposing forces remains poorly understood. It is possible that inter-individual differences in the loading imposed by different forms of locomotion may alter the amount of mineral mobilized during reproduction. Here, the impact of vertical versus horizontal locomotion on bone mobilization was examined during reproduction in the laboratory mouse. The vertical, or climbing, group had access to a 60-cm tower, increasing strain on their appendicular skeleton. The horizontal, or tunnel, group had access to a 100-cm tunnel, which encouraged movements within the horizontal plane. Form of locomotion did not impact the amount of bone females mobilized during reproduction or the amount of mineral females deposited in the litter, but maternal bone architecture differed between groups. The climbing group displayed more trabeculae than the tunnel group, whereas the tunnel group displayed greater cortical bone mineral density mid-shaft. Interestingly, pups born to mothers in the climbing group had a higher concentration of total body calcium at 16 days than pups of mothers in the tunnel group. As maternal total body calcium composition and the amount of calcium invested in the full litter were not different between groups, the difference in the relative calcium content of pups between groups is not suspected to reflect difference in mineral allocation. Future research should consider the impact of maternal activity on the efficiency of offspring skeletal ossification via hormones and other bioactive factors transferred in utero and in milk.     

Modern analysis of bone loss mechanisms in microgravity

A summary of results of investigations by the author and a brief review of some literature data on human bone tissue deprived of mechanical loading (spaceflight, hypokinesia) is given. The direction and markedness of changes in bone mass--the bone mineral density and the bone mineral content--in different skeletal segments depend on their position relative to the gravity vector. A theoretically expected bone mass reduction was revealed in the trabecular structures of the bones of the lower part of the skeleton (local osteopenia). In the upper part of the skeleton, an increase in the bone mineral content is observed, which is considered as a secondary response and is due to redistribution of body fluids cephalad. The main cause of osteopenia is mechanical unloading. Arguments are presented that osteocyte osteolysis, delayed osteoblast histogenesis, and osteoclast resorption provoked by rearrangement in the hierarchy of the systems of fluid volume and ion regulation, and the endocrine control of calcium homeostasis are the main mechanisms of osteopenia.     

Female reproductive system and bone

The female reproductive system plays a major role in regulating the acquisition and loss of bone by the skeleton from menarche through senescence. Onset of gonadal sex steroid secretion at puberty is the major factor responsible for skeletal longitudinal and radial growth, as well as significant gain in bone density, until peak bone density is achieved in third decade of life. Gonadal sex steroids then help maintain peak bone density until menopause, including during the transient changes in skeletal mineral content associated with pregnancy and lactation. At menopause, decreased gonadal sex steroid production normally leads to rapid bone loss. The most rapid bone loss associated with decreased estrogen levels occurs in the first 8-10 years after menopause, with slower age-related bone loss occurring during later life. Age-related bone loss in women after the early menopausal phase of bone loss is caused by ongoing gonadal sex steroid deficiency, vitamin D deficiency, and secondary hyperparathyroidism. Other factors also contribute to age-related bone loss, including intrinsic defects in osteoblast function, impairment of the GH/IGF axis, reduced peak bone mass, age-associated sarcopenia, and various sporadic secondary causes. Further understanding of the relative contributions of the female reproductive system and each of the other factors to development and maintenance of the female skeleton, bone loss, and fracture risk will lead to improved approaches for prevention and treatment of osteoporosis.     

The response of bone to mechanical loading and disuse: fundamental principles and influences on osteoblast/osteocyte homeostasis

Bone’s response to increased or reduced loading/disuse is a feature of many clinical circumstances, and our daily life, as habitual activities change. However, there are several misconceptions regarding what constitutes loading or disuse and why the skeleton gains or loses bone. The main purpose of this article is to discuss the fundamentals of the need for bone to experience the effects of loading and disuse, why bone loss due to disuse occurs, and how it is the target of skeletal physiology which drives pathological bone loss in conditions that may not be seen as being primarily due to disuse. Fundamentally, if we accept that hypertrophy of bone in response to increased loading is a desirable occurrence, then disuse is not a pathological process, but simply the corollary of adaptation to increased loads. If adaptive processes occur to increase bone mass in response to increased load, then the loss of bone in disuse is the only way that adaptation can fully tune the skeleton to prevailing functional demands when loading is reduced. The mechanisms by which loading and disuse cause bone formation or resorption are the same, although the direction of any changes is different. The osteocyte and osteoblast are the key cells involved in sensing and communicating the need for changes in mass or architecture as a result of changes in experienced loading. However, as those cells are affected by numerous other influences, the responses of bone to loading or disuse are not simple, and alter under different circumstances. Understanding the principles of disuse and loading and the mechanisms underlying them therefore represents an important feature of bone physiology and the search for targets for anabolic therapies for skeletal pathology.     

Size, structure and gender: lessons about fracture risk

The differences in age-related fracture risks among men and women must reflect gender differences in the relevant variables. We are concerned here with gender differences in structural variables that relate to the size and shape of bones. As children grow, their bones grow in diameter through periosteal modeling. Studies show that radial growth is driven by mechanical forces and is not just "genetically programmed". Moving bone mass farther from the center of the diaphysis makes it more effective in resisting bending and twisting forces, and disproportionately so in comparison to changes in bone mass. Gender differences in long bone structure appear to arise because the bone cells of males and females function in different hormonal environments which affect their responses to mechanical loading. In girls, bone formation on the metacarpal periosteal surface essentially stops at puberty, and is replaced by formation on the endosteal surface, reducing endosteal diameter until about age 20. Bone strength is 60% greater in male metacarpals than in those of females because bone is added periosteally in boys and endosteally in girls. At menopause endosteal resorption resumes, accompanied by slow periosteal apposition, weakening cortical structure. Similar phenomena occur in such critical regions as the femoral neck. Another fundamental gender difference in skeletal development is that whole body bone mineral content increases in linear proportion to lean body mass throughout skeletal maturation in boys, but in girls there is a distinct increase in the slope of this relationship at puberty, when estrogen rises. Frost's hypothesis is that this reflects an effect of estrogen on bone's mechanostat set point, and this is increasingly supported by data showing that estrogen and mechanical strain act through a common pathway in osteoblast-like cells. If Frost's hypothesis is correct, the mechanostat is set for maximal effect of mechanical loading on bone gain during the 2-3 years preceding menarche. During the childbearing years, the set point is at an intermediate level, and at menopause, it shifts again to place the skeleton into the metabolic equivalent of a disuse state. The most direct approach to resolving this problem would be to simulate the putative effect of estrogen on the set point itself.