Estrogen receptor-ß regulates mechanical signaling in primary osteoblasts.

Mechanical loading is an important regulator in skeletal growth, maintenance, and aging. Estrogen receptors have a regulatory role in mechanically induced bone adaptation. Estrogen receptor-α (ERα) is known to enhance load-induced bone formation, whereas ERß negatively regulates this process. We hypothesized that ERß regulates mechanical signaling in osteoblasts. We tested this hypothesis by subjecting primary calvarial cells isolated from wild-type and ERß-knockout mice (BERKO) to oscillatory fluid flow in the absence or presence of estradiol (E2). We found that the known responses to fluid shear stress, i.e., phosphorylation of the mitogen-activated protein kinase ERK and upregulation of COX-2 expression, were inhibited in BERKO cells in the absence of E2. Flow-induced increase in prostaglandin E2 (PGE2) release was not altered in BERKO cells in the absence of E2, but was increased when E2 was present. Additionally, immunofluorescence analysis and estrogen response element luciferase assays revealed increased ERα expression and flow- and ligand-induced nuclear translocation as well as transcriptional activity in BERKO cells in both the presence and absence of E2. Taken together, these data suggest that ERß plays both ligand-dependent and ligand-independent roles in mechanical signaling in osteoblasts. Furthermore, our data suggest that one mechanism by which ERß regulates mechanotransduction in osteoblasts may result from its inhibitory effect on ERα expression and function. Targeting estrogen receptors (e.g., inhibiting ERß) may represent an effective approach for prevention and treatment of age-related bone loss.

Regulatory mechanisms in bone following mechanical loading.

Bone responds with increased bone formation to mechanical loading, and the time course of bone formation after initiating mechanical loading is well characterized. However, the regulatory activities governing the loading-dependent changes in gene expression are not well understood. The goal of this study was to identify the time-dependent regulatory mechanisms that governed mechanical loading-induced gene expression in bone using a predictive bioinformatics algorithm. A standard model for bone loading in rodents was employed in which the right forelimb was loaded axially for three minutes per day, while the left forearm served as a non-loaded, contralateral control. Animals were subjected to loading sessions every day, with 24 hours between sessions. Ulnas were sampled at 11 time points, from 4 hours to 32 days after beginning loading. Using a predictive bioinformatics algorithm, we created a linear model of gene expression and identified 44 transcription factor binding motifs and 29 microRNA binding sites that were predicted to regulate gene expression across the time course. Known and novel transcription factor binding motifs were identified throughout the time course, as were several novel microRNA binding sites. These time-dependent regulatory mechanisms may be important in controlling the loading-induced bone formation process.

Sost downregulation and local Wnt signaling are required for the osteogenic response to mechanical loading.

Sclerostin, the Wnt signaling antagonist encoded by the Sost gene, is secreted by osteocytes and inhibits bone formation by osteoblasts. Mechanical stimulation reduces sclerostin expression, suggesting that osteocytes might coordinate the osteogenic response to mechanical force by locally unleashing Wnt signaling. To investigate whether sclerostin downregulation is a pre-requisite for load-induced bone formation, we conducted experiments in transgenic mice (TG) engineered to maintain high levels of SOST expression during mechanical loading. This was accomplished by introducing a human SOST transgene driven by the 8 kb fragment of the DMP1 promoter that also provided osteocyte specificity of the transgene. Right ulnae were subjected to in vivo cyclic axial loading at equivalent strains for 1 min/day at 2 Hz; left ulnae served as internal controls. Endogenous murine Sost mRNA expression measured 24 h after 1 loading bout was decreased by about 50% in TG and wild type (WT) littermates. In contrast, human SOST, only expressed in TG mice, remained high after loading. Mice were loaded on 3 consecutive days and bone formation was quantified 16 days after initiation of loading. Periosteal bone formation in control ulnae was similar in WT and TG mice. Loading induced the expected strain-dependent increase in bone formation in WT mice, resulting from increases in both mineralizing surface (MS/BS) and mineral apposition rate (MAR). In contrast, load-induced bone formation was reduced by 70-85% in TG mice, due to lower MS/BS and complete inhibition of MAR. Moreover, Wnt target gene expression induced by loading in WT mice was absent in TG mice. Thus, downregulation of Sost/sclerostin in osteocytes is an obligatory step in the mechanotransduction cascade that activates Wnt signaling and directs osteogenesis to where bone is structurally needed.

High-bone-mass-producing mutations in the Wnt signaling pathway result in distinct skeletal phenotypes.

Mutations among genes that participate in the canonical Wnt signaling pathway can lead to drastically different skeletal phenotypes, ranging from severe osteoporosis to severe osteosclerosis. Many high-bone-mass (HBM) causing mutations that occur in the LRP5 gene appear to impart the HBM phenotype, in part, by increasing resistance to soluble Wnt signaling inhibitors, including sclerostin. Sost loss-of-function mutant mice (Sost knock-out) and Lrp5 gain-of-function mutant mice (Lrp5 HBM knock-in) have high bone mass. These mutants potentially would be predicted to be phenocopies of one another, because in both cases, the sclerostin-Lrp5 interaction is disrupted. We measured bone mass, size, geometry, architecture, and strength in bones from three different genetic mouse models (Sost knock-out, Lrp5 A214V knock-in, and Lrp5 G171V knock-in) of HBM. We found that all three mouse lines had significantly elevated bone mass in the appendicular skeleton and in the cranium. Sost mutants and Lrp5 A214V mutants were statistically indistinguishable from one another in most endpoints, whereas both were largely different from the Lrp5 G171V mutants. Lrp5 G171V mutants preferentially added bone endocortically, whereas Lrp5 A214V and Sost mutants preferentially added bone periosteally. Cranial thickness and cranial nerve openings were similarly altered in all three HBM models. We also assessed serum serotonin levels as a possible mechanism accounting for the observed changes in bone mass, but no differences in serum serotonin were found in any of the three HBM mouse lines. The skeletal dissimilarities of the Lrp5 G171V mutant to the other mutants suggest that other, non-sclerostin-associated mechanisms might account for the changes in bone mass resulting from this mutation.

Anabolic and catabolic regimens of human parathyroid hormone 1-34 elicit bone- and envelope-specific attenuation of skeletal effects in Sost-deficient mice.

PTH is a potent calcium-regulating factor that has skeletal anabolic effects when administered intermittently or catabolic effects when maintained at consistently high levels. Bone cells express PTH receptors, but the cellular responses to PTH in bone are incompletely understood. Wnt signaling has recently been implicated in the osteo-anabolic response to the hormone. Specifically, the Sost gene, a major antagonist of Wnt signaling, is down-regulated by PTH exposure. We investigated this mechanism by treating Sost-deficient mice and their wild-type littermates with anabolic and catabolic regimens of PTH and measuring the skeletal responses. Male Sost(+/+) and Sost(-/-) mice were injected daily with human PTH 1-34 (0, 30, or 90 µg/kg) for 6 wk. Female Sost(+/+) and Sost(-/-) mice were continuously infused with vehicle or high-dose PTH (40 µg/kg · d) for 3 wk. Dual energy x-ray absorptiometry-derived measures of intermittent PTH (iPTH)-induced bone gain were impaired in Sost(-/-) mice. Further probing revealed normal or enhanced iPTH-induced cortical bone formation rates but concomitant increases in cortical porosity among Sost(-/-) mice. Distal femur trabecular bone was highly responsive to iPTH in Sost(-/-) mice. Continuous PTH (cPTH) infusion resulted in equal bone loss in Sost(+/+) and Sost(-/-) mice as measured by dual energy x-ray absorptiometry. However, distal femur trabecular bone, but not lumbar spine trabecular bone, was spared the bone-wasting effects of cPTH in Sost(-/-) mice. These results suggest that changes in Sost expression are not required for iPTH-induced anabolism. iPTH-induced resorption of cortical bone might be overstimulated in Sost-deficient environments. Furthermore, Sost deletion protects some trabecular compartments, but not cortical compartments, from bone loss induced by high-dose PTH infusion.

Non-overlapping functions for Pyk2 and FAK in osteoblasts during fluid shear stress-induced mechanotransduction.

Mechanotransduction, the process by which cells convert external mechanical stimuli such as fluid shear stress (FSS) into biochemical changes, plays a critical role in maintenance of the skeleton. We have proposed that mechanical stimulation by FSS across the surfaces of bone cells results in formation of unique signaling complexes called mechanosomes that are launched from sites of adhesion with the extracellular matrix and with other bone cells [1]. Deformation of adhesion complexes at the cell membrane ultimately results in alteration of target gene expression. Recently, we reported that focal adhesion kinase (FAK) functions as a part of a mechanosome complex that is required for FSS-induced mechanotransduction in bone cells. This study extends this work to examine the role of a second member of the FAK family of non-receptor protein tyrosine kinases, proline-rich tyrosine kinase 2 (Pyk2), and determine its role during osteoblast mechanotransduction. We use osteoblasts harvested from mice as our model system in this study and compared the contributions of Pyk2 and FAK during FSS induced mechanotransduction in osteoblasts. We exposed Pyk2(+/+) and Pyk2(-/-) primary calvarial osteoblasts to short period of oscillatory fluid flow and analyzed downstream activation of ERK1/2, and expression of c-fos, cyclooxygenase-2 and osteopontin. Unlike FAK, Pyk2 was not required for fluid flow-induced mechanotransduction as there was no significant difference in the response of Pyk2(+/+) and Pyk2(-/-) osteoblasts to short periods of fluid flow (FF). In contrast, and as predicted, FAK(-/-) osteoblasts were unable to respond to FF. These data indicate that FAK and Pyk2 have distinct, non-redundant functions in launching mechanical signals during osteoblast mechanotransduction. Additionally, we compared two methods of generating FF in both cell types, oscillatory pump method and another orbital platform method. We determined that both methods of generating FF induced similar responses in both primary calvarial osteoblasts and immortalized calvarial osteoblasts.

Heterogeneous stock rat: a unique animal model for mapping genes influencing bone fragility.

Previously, we demonstrated that skeletal mass, structure and biomechanical properties vary considerably among 11 different inbred rat strains. Subsequently, we performed quantitative trait loci (QTL) analysis in four inbred rat strains (F344, LEW, COP and DA) for different bone phenotypes and identified several candidate genes influencing various bone traits. The standard approach to narrowing QTL intervals down to a few candidate genes typically employs the generation of congenic lines, which is time consuming and often not successful. A potential alternative approach is to use a highly genetically informative animal model resource capable of delivering very high resolution gene mapping such as Heterogeneous stock (HS) rat. HS rat was derived from eight inbred progenitors: ACI/N, BN/SsN, BUF/N, F344/N, M520/N, MR/N, WKY/N and WN/N. The genetic recombination pattern generated across 50 generations in these rats has been shown to deliver ultra-high even gene-level resolution for complex genetic studies. The purpose of this study is to investigate the usefulness of the HS rat model for fine mapping and identification of genes underlying bone fragility phenotypes. We compared bone geometry, density and strength phenotypes at multiple skeletal sites in HS rats with those obtained from five of the eight progenitor inbred strains. In addition, we estimated the heritability for different bone phenotypes in these rats and employed principal component analysis to explore relationships among bone phenotypes in the HS rats. Our study demonstrates that significant variability exists for different skeletal phenotypes in HS rats compared with their inbred progenitors. In addition, we estimated high heritability for several bone phenotypes and biologically interpretable factors explaining significant overall variability, suggesting that the HS rat model could be a unique genetic resource for rapid and efficient discovery of the genetic determinants of bone fragility.

Alternative splicing in bone following mechanical loading.

It is estimated that more than 90% of human genes express multiple mRNA transcripts due to alternative splicing. Consequently, the proteins produced by different splice variants will likely have different functions and expression levels. Several genes with splice variants are known in bone, with functions that affect osteoblast function and bone formation. The primary goal of this study was to evaluate the extent of alternative splicing in a bone subjected to mechanical loading and subsequent bone formation. We used the rat forelimb loading model, in which the right forelimb was loaded axially for 3 min, while the left forearm served as a non-loaded control. Animals were subjected to loading sessions every day, with 24 h between sessions. Ulnae were sampled at 11 time points, from 4 h to 32days after beginning loading. RNA was isolated and mRNA abundance was measured at each time point using Affymetrix exon arrays (GeneChip® Rat Exon 1.0 ST Arrays). An ANOVA model was used to identify potential alternatively spliced genes across the time course, and five alternatively spliced genes were validated with qPCR: Akap12, Fn1, Pcolce, Sfrp4, and Tpm1. The number of alternatively spliced genes varied with time, ranging from a low of 68 at 12h to a high of 992 at 16d. We identified genes across the time course that encoded proteins with known functions in bone formation, including collagens, matrix proteins, and components of the Wnt/ß-catenin and TGF-ß signaling pathways. We also identified alternatively spliced genes encoding cytokines, ion channels, muscle-related genes, and solute carriers that do not have a known function in bone formation and represent potentially novel findings. In addition, a functional characterization was performed to categorize the global functions of the alternatively spliced genes in our data set. In conclusion, mechanical loading induces alternative splicing in bone, which may play an important role in the response of bone to mechanical loading.

Gene expression patterns in bone following mechanical loading.

The advent of high-throughput measurements of gene expression and bioinformatics analysis methods offers new ways to study gene expression patterns. The primary goal of this study was to determine the time sequence for gene expression in a bone subjected to mechanical loading during key periods of the bone-formation process, including expression of matrix-related genes, the appearance of active osteoblasts, and bone desensitization. A standard model for bone loading was employed in which the right forelimb was loaded axially for 3 minutes per day, whereas the left forearm served as a nonloaded contralateral control. We evaluated loading-induced gene expression over a time course of 4 hours to 32 days after the first loading session. Six distinct time-dependent patterns of gene expression were identified over the time course and were categorized into three primary clusters: genes upregulated early in the time course, genes upregulated during matrix formation, and genes downregulated during matrix formation. Genes then were grouped based on function and/or signaling pathways. Many gene groups known to be important in loading-induced bone formation were identified within the clusters, including AP-1-related genes in the early-response cluster, matrix-related genes in the upregulated gene clusters, and Wnt/ß-catenin signaling pathway inhibitors in the downregulated gene clusters. Several novel gene groups were identified as well, including chemokine-related genes, which were upregulated early but downregulated later in the time course; solute carrier genes, which were both upregulated and downregulated; and muscle-related genes, which were primarily downregulated.

A longitudinal study of the effect of genistein on bone in two different murine models of diminished estrogen-producing capacity.

This experiment was designed to assess the capacity of dietary genistein (GEN), to attenuate bone loss in ovariectomized (OVX) and ovary-intact VCD-treated mice. Pretreatment of mice with 4-vinylcyclohexene diepoxide (VCD) gradually and selectively destroys ovarian follicles whilst leaving ovarian androgen-producing cells largely intact. VCD induces a perimenopause-like condition prior to the onset of reproductive acyclicity. Sixteen-week-old C57BL/6J mice were randomized to five treatment groups: sham(SHM), OVX, SHM + VCD, OVX + GEN, and SHM + VCD + GEN. In vivo, blood samples were drawn for hormone and isoflavone analyses, estrous cycles were monitored, and X-ray imaging was performed to assess changes in bone parameters. Following sacrifice, ovaries were assessed histologically, bone microarchitecture was evaluated via microcomputed tomography, and bone mechanical properties were measured. Some effects of GEN were observed in OVX mice, but GEN effects were not able to be evaluated in VCD-treated mice due to the subtle diminution of bone during the 4 months of this experiment.

Identification of genes influencing skeletal phenotypes in congenic P/NP rats.

We previously showed that alcohol-preferring (P) rats have higher bone density than alcohol-nonpreferring (NP) rats. Genetic mapping in P and NP rats identified a major quantitative trait locus (QTL) between 4q22 and 4q34 for alcohol preference. At the same location, several QTLs linked to bone density and structure were detected in Fischer 344 (F344) and Lewis (LEW) rats, suggesting that bone mass and strength genes might cosegregate with genes that regulate alcohol preference. The aim of this study was to identify the genes segregating for skeletal phenotypes in congenic P and NP rats. Transfer of the NP chromosome 4 QTL into the P background (P.NP) significantly decreased areal bone mineral density (aBMD) and volumetric bone mineral density (vBMD) at several skeletal sites, whereas transfer of the P chromosome 4 QTL into the NP background (NP.P) significantly increased bone mineral content (BMC) and aBMD in the same skeletal sites. Microarray analysis from the femurs using Affymetrix Rat Genome arrays revealed 53 genes that were differentially expressed among the rat strains with a false discovery rate (FDR) of less than 10%. Nine candidate genes were found to be strongly correlated (r(2) > 0.50) with bone mass at multiple skeletal sites. The top three candidate genes, neuropeptide Y (Npy), alpha synuclein (Snca), and sepiapterin reductase (Spr), were confirmed using real-time quantitative PCR (qPCR). Ingenuity pathway analysis revealed relationships among the candidate genes related to bone metabolism involving beta-estradiol, interferon-gamma, and a voltage-gated calcium channel. We identified several candidate genes, including some novel genes on chromosome 4 segregating for skeletal phenotypes in reciprocal congenic P and NP rats.

Modulation of Wnt signaling influences fracture repair.

While the importance of Wnt signaling in skeletal development and homeostasis is well documented, little is known regarding its function in fracture repair. We hypothesized that activation and inactivation of Wnt signaling would enhance and impair fracture repair, respectively. Femoral fractures were generated in Lrp5 knockout mice (Lrp5-/-) and wild-type littermates (Lrp5+/+), as well as C57BL/6 mice. Lrp5-/- and Lrp5+/+ mice were untreated, while C57BL/6 mice were treated 2x/week with vehicle or anti-Dkk1 antibodies (Dkk1 Ab) initiated immediately postoperatively (Day 0) or 4 days postoperatively (Day 4). Fractures were radiographed weekly until sacrifice at day 28, followed by DXA, pQCT, and biomechanical analyses. Lrp5-/- mice showed impaired repair compared to Lrp5+/+ mice, as evidenced by reduced callus area, BMC, BMD, and biomechanical properties. The effects of Dkk1 Ab treatment depended on the timing of initiation. Day 0 initiation enhanced repair, with significant gains seen for callus area, BMC, BMD, and biomechanical properties, whereas Day 4 initiation had no effect. These results validated our hypothesis that Wnt signaling influences fracture repair, with prompt activation enhancing repair and inactivation impairing it. Furthermore, these data suggest that activation of Wnt signaling during fracture repair may have clinical utility in facilitating fracture repair.

Psychotropic drugs have contrasting skeletal effects that are independent of their effects on physical activity levels.

Popular psychotropic drugs, like the antidepressant selective serotonin reuptake inhibitors (SSRIs) and tricyclic antidepressants (TCAs), and the mood stabilizer lithium, may have skeletal effects. In particular, preclinical observations suggest a direct negative effect of SSRIs on the skeleton. A potential caveat in studies of the skeletal effects of psychotropic drugs is the hypoactive (skeletal unloading) phenotype they induce. The aim of this study was to investigate the contribution of physical inactivity to the skeletal effects of psychotropic drugs by studying bone changes in cage control and tail suspended mice treated with either vehicle, SSRI, TCA or lithium. Tail suspension was used to control for drug differences on physical activity levels by normalizing skeletal loading between groups. The psychotropic drugs were found to have contrasting skeletal effects which were independent of drug effects on animal physical activity levels. The latter was evident by an absence of statistical interactions between the activity and drug groups. Pharmacological inhibition of the 5-hydroxytryptamine (5-HT) transporter (5-HTT) using a SSRI reduced in vivo gains in lower extremity BMD, and negatively altered ex vivo measures of femoral and spinal bone density, architecture and mechanical properties. These effects were mediated by a decrease in bone formation without a change in bone resorption suggesting that the SSRI had anti-anabolic skeletal effects. In contrast, glycogen synthase kinase-3[beta] (GSK-3[beta]) inhibition using lithium had anabolic effects improving in vivo gains in BMD via an increase in bone formation, while TCA-mediated inhibition of the norepinephrine transporter had minimal skeletal effect. The observed negative skeletal effect of 5-HTT inhibition, combined with recent findings of direct and indirect effects of 5-HT on bone formation, are of interest given the frequent prescription of SSRIs for the treatment of depression and other affective disorders. Likewise, the anabolic effect of GSK-3[beta] inhibition using lithium reconfirms the importance of Wnt/beta-catenin signaling in the skeleton and it's targeting by recent drug discovery efforts. In conclusion, the current study demonstrates that different psychotropic drugs with differing underlying mechanisms of action have contrasting skeletal effects and that these effects do not result indirectly via the generation of animal physical inactivity.

Lengthening of mouse hindlimbs with joint loading.

For devising clinical approaches to treating limb length discrepancies, strategies that will generate differential longitudinal growth need to be improved. This report addresses the following question: does knee loading increase bone length of the loaded hindlimb? Knee loading has been shown to induce anabolic responses on the periosteal and endosteal surfaces, but its effects on longitudinal bone growth have not yet been examined. In the present studies, loads were applied to the left hindlimb (5-min bouts at 0.5 N) of C57/BL/6 mice (21 mice, ~8 weeks old). Compared to the contralateral and age-matched control groups, knee loading increased the length of the femur by 2.3 and 3.5%, together with the tibia by 2.3 and 3.7% (all P < 0.001), respectively. In accordance with the length measurements, knee loading elevated BMD and BMC in both the femur and the tibia. Histological analysis of the proximal tibia revealed that the loaded growth plate elevated its height by 19.5% (P < 0.001) and the cross-sectional area by 30.7% (P < 0.05). Particularly in the hypertrophic zone, knee loading increased the number of chondrocytes (P < 0.01) as well as their cellular height (P < 0.001) along the length of the tibia. Taken together, this study demonstrates for the first time the potential effectiveness of knee loading in adjusting limb length discrepancy.

Genes influencing spinal bone mineral density in inbred F344, LEW, COP, and DA rats.

Previously, we identified the regions of chromosomes 10q12-q31 and 15p16-q21 harbor quantitative trait loci (QTLs) for lumbar volumetric bone mineral density (vBMD) in female F2 rats derived from Fischer 344 (F344) x Lewis (LEW) and Copenhagen 2331 (COP) x Dark Agouti (DA) crosses. The purpose of this study is to identify the candidate genes within these QTL regions contributing to the variation in lumbar vBMD. RNA was extracted from bone tissue of F344, LEW, COP, and DA rats. Microarray analysis was performed using Affymetrix Rat Genome 230 2.0 Arrays. Genes differentially expressed among the rat strains were then ranked based on the strength of the correlation with lumbar vBMD in F2 animals derived from these rats. Quantitative PCR (qPCR) analysis was performed to confirm the prioritized candidate genes. A total of 285 genes were differentially expressed among all strains of rats with a false discovery rate less than 10%. Among these genes, 18 candidate genes were prioritized based on their strong correlation (r (2) > 0.90) with lumbar vBMD. Of these, 14 genes (Akap1, Asgr2, Esd, Fam101b, Irf1, Lcp1, Ltc4s, Mdp-1, Pdhb, Plxdc1, Rabep1, Rhot1, Slc2a4, Xpo4) were confirmed by qPCR. We identified several novel candidate genes influencing spinal vBMD in rats.

The emerging role of serotonin (5-hydroxytryptamine) in the skeleton and its mediation of the skeletal effects of low-density lipoprotein receptor-related protein 5 (LRP5).

Novel molecular pathways obligatory for bone health are being rapidly identified. One pathway recently revealed involves gut-derived 5-hydroxytryptamine (5-HT) mediation of the complete skeletal effects of low-density lipoprotein receptor-related protein 5 (LRP5). Mounting evidence supports 5-HT as an important regulatory compound in bone with previous evidence demonstrating that bone cells possess functional pathways for responding to 5-HT. In addition, there is growing evidence that potentiation of 5-HT signaling via inhibition of the 5-HT transporter (5-HTT) has significant skeletal effects. The later is clinically significant as the 5-HTT is a popular target of pharmaceutical agents, such as selective serotonin reuptake inhibitors (SSRIs), used for the management of major depressive disorder and other affective conditions. The observation that 5-HT mediates the complete skeletal effects of LRP5 represents a significant paradigm shift from the traditional view that LRP5 located on the cell surface membrane of osteoblasts exerts direct skeletal effects via Wnt/beta-catenin signaling. This paper discusses the mounting evidence for skeletal effects of 5-HT and the ability of gut-derived 5-HT to satisfactorily explain the skeletal effects of LRP5.

Numerical modeling of long bone adaptation due to mechanical loading: correlation with experiments.

The process of external bone adaptation in cortical bone is modeled mathematically using finite element (FE) stress analysis coupled with an evolution model, in which adaptation response is triggered by mechanical stimulus represented by strain energy density. The model is applied to experiments in which a rat ulna is subjected to cyclic loading, and the results demonstrate the ability of the model to predict the bone adaptation response. The FE mesh is generated from micro-computed tomography (microCT) images of the rat ulna, and the stress analysis is carried out using boundary and loading conditions on the rat ulna obtained from the experiments [Robling, A. G., F. M. Hinant, D. B. Burr, and C. H. Turner. J. Bone Miner. Res. 17:1545-1554, 2002]. The external adaptation process is implemented in the model by moving the surface nodes of the FE mesh based on an evolution law characterized by two parameters: one that captures the rate of the adaptation process (referred to as gain); and the other characterizing the threshold value of the mechanical stimulus required for adaptation (referred to as threshold-sensitivity). A parametric study is carried out to evaluate the effect of these two parameters on the adaptation response. We show, following comparison of results from the simulations to the experimental observations of Robling et al. (J. Bone Miner. Res. 17:1545-1554, 2002), that splitting the loading cycles into different number of bouts affects the threshold-sensitivity but not the rate of adaptation. We also show that the threshold-sensitivity parameter can quantify the mechanosensitivity of the osteocytes.

Bone loss or lost bone: rationale and recommendations for the diagnosis and treatment of early postmenopausal bone loss.

Recent reports suggest that bone loss begins during late perimenopause at a dramatic rate, even before estrogen levels plummet. During the ensuing 5 years, there is evidence of the beginnings of microarchitectural deterioration, which impacts bone strength and ultimately enhances its propensity to fracture. The diagnosis of osteoporosis based on T-scores alone, or through stratification for a high fracture risk by FRAX, excludes these women who are rapidly losing bone. Because all antiosteoporosis therapies, in particular bisphosphonates, reduce bone loss, we propose aggressive, likely short-term therapy with a goal to reduce bone loss, stabilize bone density, and prevent microarchitectural deterioration.

Mechanical signaling for bone modeling and remodeling.

Proper development of the skeleton in utero and during growth requires mechanical stimulation. Loading results in adaptive changes in bone that strengthen bone structure. Bone's adaptive response is regulated by the ability of resident bone cells to perceive and translate mechanical energy into a cascade of structural and biochemical changes within the cells a process known as mechanotransduction. Mechanotransduction pathways are among the most anabolic in bone, and consequently, there is great interest in elucidating how mechanical loading produces its observed effects, including increased bone formation, reduced bone loss, changes in bone cell differentiation and lifespan, among others. A molecular understanding of these processes is developing, and with it comes a profound new insight into the biology of bone. In this article, we review the nature of the physical stimulus to which bone cells mount an adaptive response, including the identity of the sensor cells, their attributes and physical environment, and putative mechanoreceptors they express. Particular attention is allotted to the focal adhesion and Wnt signaling, in light of their emerging role in bone mechanotransduction. Te cellular mechanisms for increased bone loss during disuse, and reduced bone loss during loading are considered. Finally, we summarize the published data on bone cell accommodation, whereby bone cells stop responding to mechanical signaling events. Collectively, these data highlight the complex yet finely orchestrated process of mechanically regulated bone homeostasis.