Bone Biomechanical Properties Research Articles

Mechanical competence of ovariectomy-induced compromised bone after single or combined treatment with high-frequency loading and bisphosphonates.

Osteoporosis leads to increased bone fragility, thus effective approaches enhancing bone strength are needed. Hence, this study investigated the effect of single or combined application of high-frequency (HF) loading through whole body vibration (WBV) and alendronate (ALN) on the mechanical competence of ovariectomy-induced osteoporotic bone. Thirty-four female Wistar rats were ovariectomized (OVX) or sham-operated (shOVX) and divided into five groups: shOVX, OVX-shWBV, OVX-WBV, ALN-shWBV and ALN-WBV. (Sham)WBV loading was applied for 10 min/day (130 to 150 Hz at 0.3g) for 14 days and ALN at 2 mg/kg/dose was administered 3x/week. Finite element analysis based on micro-CT was employed to assess bone biomechanical properties, relative to bone micro-structural parameters. HF loading application to OVX resulted in an enlarged cortex, but it was not able to improve the biomechanical properties. ALN prevented trabecular bone deterioration and increased bone stiffness and bone strength of OVX bone. Finally, the combination of ALN with HF resulted in an increased cortical thickness in OVX rats when compared to single treatments. Compared to HF loading, ALN treatment is preferred for improving the compromised mechanical competence of OVX bone. In addition, the association of ALN with HF loading results in an additive effect on the cortical thickness.

Obesity-related changes in bone structural and material properties in hyperphagic OLETF rats and protection by voluntary wheel running

PurposeTo examine how the development of obesity and the associated insulin resistance affect bone structural and material properties, and bone formation and resorption markers in the Otsuka Long-Evans Tokushima Fatty (OLETF) rat model. MethodsThis was a 36-week study of sedentary, hyperphagic, male OLETF rats (OLETF-SED), exercise-treated OLETF rats (OLETF-EX) and sedentary non-hyperphagic controls (LETO-SED) with data collection at 13, 20, and 40weeks of age (n=5–8 animals per group per timepoint). ResultsBody mass and fat (%) were significantly greater in OLETF-SED versus controls. OLETF-SED were insulin resistant at 13 and 20weeks, with overt diabetes by 40weeks. At 13weeks, OLETF-SED had lower total body BMC and BMD and serum P1NP compared with LETO-SED. Differences in total body BMC and BMD between OLETF-SED and LETO-SED persisted at 20weeks, with reductions in total and cortical BMD of the tibia. OLETF-SED also had lesser femur diameter, cross-sectional area, polar moment of area, and torque at fracture than LETO-SED. By 40weeks, OLETF-SED had elevated bone resorption and reduced intrinsic bone strength. OLETF-EX did not show the excessive weight gain, obesity, insulin resistance or diabetes observed in OLETF-SED. OLETF-EX had greater BMD than OLETF-SED, and structural and material properties of the femur were significantly increased in OLETF-EX relative to OLETF-SED and LETO-SED. ConclusionsThe negative skeletal effects of excessive adiposity and insulin resistance were evident early in the progressive obesity with lasting negative impacts on intrinsic and extrinsic bone strength. Exercise protected against obesity-associated skeletal changes with marked benefits on the biomechanical properties of bone.

Abnormal Skeletal Strength and Microarchitecture in Women With Celiac Disease.

Osteoporosis is often a presenting sign of celiac disease (CD). Whether skeletal fragility in CD is associated with microarchitectural abnormalities is not known. The objective of the study was to evaluate microarchitecture and biomechanical properties of bone in CD. This was a case-control study. The study was conducted at a university hospital outpatient facility. Patients included premenopausal women with newly diagnosed CD (n = 33) and healthy controls (n = 33). Areal bone mineral density by dual-energy x-ray absorptiometry was measured as was trabecular and cortical volumetric bone mineral density (vBMD) and microarchitecture by high-resolution peripheral computed tomography of the distal radius and tibia. Whole-bone stiffness estimated by finite element analysis. PTH, 25-hydroxyvitamin D, and bone turnover markers were also measured. Groups had similar age, race, and body mass index. Both groups had sufficient 25-hydroxyvitamin D and normal calcium and PTH. Areal bone mineral density was lower in CD. By high-resolution peripheral computed tomography, CD had lower trabecular vBMD, fewer, more widely, and irregularly spaced trabeculae at both the radius and tibia (8%-33%). At the tibia, they also had lower total density (8%) and thinner cortices (10%). Whole-bone stiffness and failure load were lower (11%-21%) in CD at both sites. Biomechanical deficits were associated with trabecular abnormalities. Women with CD had abnormal vBMD and microarchitecture at both the radius and tibia. Trabecular bone was preferentially affected. These deficits were associated with lower estimates of skeletal strength. These findings suggest a potential structural mechanism for skeletal fragility in CD and support further research into the pathogenesis of fracture in this population.

Effects of the combined extracts of Herba Epimedii and Fructus Ligustri Lucidi on bone mineral content and bone turnover in osteoporotic rats.

BackgroundThe decoction combination of Herba Epimedii and Fructus Ligustri Lucidi has been used to treat osteoporosis for almost 50 years by practitioners of traditional Chinese medicine. However, it is unclear what specific effects this combination of herbs has on the skeleton. The aim of this study was to assess the effects of the combined extracts from Herba Epimedii and Fructus Ligustri Lucidi on the bone turnover and bone mineral content in a rat model of osteoporosis induced by retinoic acid.MethodsFifty male Wistar rats were randomly assigned to the normal control group, osteoporosis model group, or treatment groups in which osteoporosis was induced and then the combined extracts of Herba Epimedii and Fructus Ligustri Lucidi were administered at 50, 100, or 200 mg/kg/day for 3 weeks via oral gavage. The rat osteoporosis model was induced by intragastric administration of 70 mg/kg/day of retinoic acid for 2 weeks. Bone turnover markers, bone biomechanical properties, and the calcium and phosphorus content of the right tibia and serum were measured.ResultsThe retinoic acid administration decreased the bone mass and the contents of calcium and phosphorus in the bone mineral, weakened the biomechanical properties, and increased bone turnover by stimulating bone resorption and collagen metabolism. Treatment with the combined extracts of Herba Epimedii and Fructus Ligustri Lucidi significantly mitigated the effects of osteoporosis on the rats by decreasing bone metabolism, improving the bone mineral content, and increasing the biomechanical properties.ConclusionsThe results of this study highlight the anti-osteoporosis effects of the combined extracts of Herba Epimedii and Fructus Ligustri Lucidi. These findings may contribute to the development of natural anti-osteoporosis herbal medicines.

Mechanism of enhanced antiosteoporosis effect of circinal-icaritin by self-assembled nanomicelles in vivo with suet oil and sodium deoxycholate.

BackgroundCircinal–icaritin (CIT), one new active aglycone of Epimedium, can exert a beneficial effect on osteoporotic bone. However, its low bioavailability limits its clinical efficacy for the treatment of osteoporosis.Materials and methodsIn this paper, suet oil (SO) was used to improve the oral bioavailability of CIT and enhance its antiosteoporosis effect and absorption. After oral administration of CIT together with SO, the CIT and SO self-assembled into nanomicelles under the action of sodium deoxycholate (DOC) by bile secretion. The antiosteoporosis effects of the CIT-SO-DOC nanomicelles were evaluated in osteoporotic rats by bone mineral density, serum biochemical markers, bone microarchitecture, bone biomechanical properties, and related protein and gene expressions. We examined the bioavailability of CIT and its nanomicelles in vivo, and subsequently the nanomicelles were verified using transmission electron microscopy. Finally, we evaluated absorption across a rat intestinal perfusion model.ResultsCompared with CIT, in the CIT-SO groups, protein and messenger ribonucleic acid expressions of osteoprotegerin were increased, while expressions of receptor activator of nuclear factor-κB ligand in bone tissue were decreased; bone-turnover markers in serum of hydroxyproline, alkaline phosphatase, tartrate-resistant acid phosphatase 5b, and receptor activator of nuclear factor-κB ligand levels were decreased, while osteoprotegerin and osteocalcin levels were increased; and trabecular bone mass, microarchitecture, and bone biomechanical strength were enhanced. The relative bioavailabilities of CIT-SO high dosage, CIT-SO medium dosage, and CIT-SO low dosage (area under concentration–time curve [AUC]0–∞) compared with that of raw CIT high dosage, CIT medium dosage, and CIT low dosage (AUC0–∞) were 127%, 121%, and 134%, respectively. The average particle size of CIT-DOC was significantly decreased after adding SO (P<0.01), and the intestinal permeability coefficients of CIT-SO-DOC nanomicelles in the duodenum, jejunum, ileum, and colon were all significantly improved (P<0.01).ConclusionThe increased antiosteoporosis effects and bioavailability of CIT-SO-DOC self-assembled nanomicelles were due to an increase in absorption of CIT by reducing the particle sizes of CIT. SO may be a practical oral carrier for antiosteoporosis drugs with low bioavailability.

Inhibition of CaMKK2 reverses age-associated decline in bone mass

Decline in bone formation is a major contributing factor to the loss of bone mass associated with aging. We previously showed that the genetic ablation of the tissue-restricted and multifunctional Ca2+/calmodulin (CaM)-dependent protein kinase kinase 2 (CaMKK2) stimulates trabecular bone mass accrual, mainly by promoting anabolic pathways and inhibiting catabolic pathways of bone remodeling. In this study, we investigated whether inhibition of this kinase using its selective cell-permeable inhibitor STO-609 will stimulate bone formation in 32week old male WT mice and reverse age-associated of decline in bone volume and strength. Tri-weekly intraperitoneal injections of saline or STO-609 (10μM) were performed for six weeks followed by metabolic labeling with calcein and alizarin red. New bone formation was assessed by dynamic histomorphometry whereas micro-computed tomography was employed to measure trabecular bone volume, microarchitecture and femoral mid-shaft geometry. Cortical and trabecular bone biomechanical properties were assessed using three-point bending and punch compression methods respectively. Our results reveal that as they progress from 12 to 32weeks of age, WT mice sustain a significant decline in trabecular bone volume, microarchitecture and strength as well as cortical bone strength. However, treatment of the 32week old WT mice with STO-609 stimulated apposition of new bone and completely reversed the age-associated decrease in bone volume, quality, as well as trabecular and cortical bone strength. We also observed that regardless of age, male Camkk2−/− mice possessed significantly elevated trabecular bone volume, microarchitecture and compressive strength as well as cortical bone strength compared to age-matched WT mice, implying that the chronic loss of this kinase attenuates age-associated decline in bone mass. Further, whereas STO-609 treatment and/or the absence of CaMKK2 significantly enhanced the femoral mid-shaft geometry, the mid-shaft cortical wall thickness and material bending stress remained similar among the cohorts, implying that regardless of treatment, the material properties of the bone remain similar. Thus, our cumulative results provide evidence for the pharmacological inhibition of CaMKK2 as a bone anabolic strategy in combating age-associated osteoporosis.

Finite element simulation of ultrasonic wave propagation in a dental implant for biomechanical stability assessment.

Dental implant stability, which is an important parameter for the surgical outcome, can now be assessed using quantitative ultrasound. However, the acoustical propagation in dental implants remains poorly understood. The objective of this numerical study was to understand the propagation phenomena of ultrasonic waves in cylindrically shaped prototype dental implants and to investigate the sensitivity of the ultrasonic response to the surrounding bone quantity and quality. The 10-MHz ultrasonic response of the implant was calculated using an axisymetric 3D finite element model, which was validated by comparison with results obtained experimentally and using a 2D finite difference numerical model. The results show that the implant ultrasonic response changes significantly when a liquid layer is located at the implant interface compared to the case of an interface fully bounded with bone tissue. A dedicated model based on experimental measurements was developed in order to account for the evolution of the bone biomechanical properties at the implant interface. The effect of a gradient of material properties on the implant ultrasonic response is determined. Based on the reproducibility of the measurement, the results indicate that the device should be sensitive to the effects of a healing duration of less than one week. In all cases, the amplitude of the implant response is shown to decrease when the dental implant primary and secondary stability increase, which is consistent with the experimental results. This study paves the way for the development of a quantitative ultrasound method to evaluate dental implant stability.

Arthritis induces early bone high turnover, structural degradation and mechanical weakness.

BackgroundWe have previously found in the chronic SKG mouse model of arthritis that long standing (5 and 8 months) inflammation directly leads to high collagen bone turnover, disorganization of the collagen network, disturbed bone microstructure and degradation of bone biomechanical properties. The main goal of the present work was to study the effects of the first days of the inflammatory process on the microarchitecture and mechanical properties of bone.MethodsTwenty eight Wistar adjuvant-induced arthritis (AIA) rats were monitored during 22 days after disease induction for the inflammatory score, ankle perimeter and body weight. Healthy non-arthritic rats were used as controls for compar-ison. After 22 days of disease progression rats were sacrificed and bone samples were collected for histomorphometrical, energy dispersive X-ray spectroscopical analysis and 3-point bending. Blood samples were also collected for bone turnover markers.ResultsAIA rats had an increased bone turnover (as inferred from increased P1NP and CTX1, p = 0.0010 and p = 0.0002, respectively) and this was paralleled by a decreased mineral content (calcium p = 0.0046 and phos-phorus p = 0.0046). Histomorphometry showed a lower trabecular thickness (p = 0.0002) and bone volume (p = 0.0003) and higher trabecular sepa-ration (p = 0.0009) in the arthritic group as compared with controls. In addition, bone mechanical tests showed evidence of fragility as depicted by diminished values of yield stress and ultimate fracture point (p = 0.0061 and p = 0.0279, re-spectively) in the arthritic group.ConclusionsWe have shown in an AIA rat model that arthritis induc-es early bone high turnover, structural degradation, mineral loss and mechanical weak-ness.

In vitro BMP2 stimulation of osteoblast citrate production in concert with mineralized bone nodule formation.

BackgroundThat citrate is a major indispensible component of bone in humans and in all osteovertebrates has been known for about seventy-five years. Yet, its role and importance in the structure and function of bone and bone formation have remained unknown. However, recent studies have identified that citrate is a major and essential component of the apatite/collagen structure of bone; and that the biomechanical properties of bone (e.g., stability, strength, resistance to fracture) depend on the appropriate incorporation of citrate in the structure of bone. The osteoblasts have recently been identified as citrate-producing cells that provide the citrate that is incorporated in the apatite/collagen structure during osteogenesis. Little is known regarding the factors and mechanisms involved in the regulation of citrate that is incorporated along with mineralization during the process of bone formation. Because of the importance of BMP2 in the initiation of osteogenesis and the development of the osteoblasts, it is essential to determine its possible implication in the development of the citrate-producing capability of the osteoblasts (i.e., “citration”) during the formation of mineralized bone nodules.MethodsThe goal of this study was to determine if BMP2 promotes the development of citrate-producing osteoblasts for increased citrate incorporation in the formation of mineralized bone nodules. The study employed MC3T3 mesenchyme stem cell osteogenic differentiation in the presence and absence of BMP2.ResultsThe results showed that BMP2 treatment increased the osteogenic development of mineralized bone nodules. In addition, BMP2 increased osteoblast citrate production and incorporation in the mineralized bone nodule. This was accompanied by increased ZIP1 transporter, which is an essential genetic/metabolic event for citrate-producing cells.ConclusionsThe results demonstrate, for the first time, that BMP2 facilitates the osteoblast “citration” process in concert with mineralization during bone formation; and provide confirmation of the important role of osteoblasts as specialized citrate-producing cells in the process of bone formation. However, it is essential to determine if these in vitro effects will occur in vivo in BMP2-implant induction of bone formation. “Citration” is essential for osteoinductive bone to represent the chemical, structural, and biomechanical properties of “normal” bone.

Effects of long-term doxycycline on bone quality and strength in diabetic male DBA/2J mice

In type 1 diabetes, diabetic bone disease (DBD) is characterized by decreased bone mineral density, a state of low bone turnover and an increased risk of fracture. Animal models of DBD demonstrate that acquired alterations in trabecular and cortical bone microarchitecture contribute to decreased bone strength in diabetes. With anti-collagenolytic and anti-inflammatory properties, tetracycline derivatives may prevent diabetes-related decreases in bone strength. To determine if doxycycline, a tetracycline class antibiotic, can prevent the development of DBD in a model of long-term diabetes, male DBA/2J mice, with or without diabetes, were treated with doxycycline-containing chow for 10weeks (dose range, 28–92mg/kg/day). Long-term doxycycline exposure was not deleterious to the microarchitecture or biomechanical properties of healthy bones in male DBA/2J mice. Doxycycline treatment also did not prevent or alleviate the deleterious changes in trabecular microarchitecture, cortical structure, and biomechanical properties of bone induced by chronic diabetes.

OS19-1 Changes in the Biomechanical Properties of Cortical Bone in Stroke-Prone Spontaneously Hypertensive Rats Due to the Exposure of Undernutrition During Pregnancy(Bone Mechanics &amp; Rehabilitation,OS19 Biomechanics and its applications,BIOMECHANICS)

OS19-1 Changes in the Biomechanical Properties of Cortical Bone in Stroke-Prone Spontaneously Hypertensive Rats Due to the Exposure of Undernutrition During Pregnancy(Bone Mechanics &amp; Rehabilitation,OS19 Biomechanics and its applications,BIOMECHANICS)

Evidence that Osteoblasts are Specialized Citrate-producing Cells that Provide the Citrate for Incorporation into the Structure of Bone.

Citrate is a major component of bone in all vertebrates, but its implications in bone have remained largely unknown. Recent studies identified that citrate is incorporated into the structure of the hydroxyapatite nanocrystal/collagen complex; and is essential for the important biomechanical properties of bone. This raises the important question, “What is the source of citrate for incorporation into bone?”; A question that heretofore had remained unresolved. Studies in this report were designed to determine the plausibility of our concept that the osteoblasts are specialized citrate-producing cells, which provide the citrate that is incorporated into the structure of bone; and that osteogenic differentiation of mesenchyme cells leads to the development of the citrate-producing osteoblasts. The results demonstrated that primary human osteoblasts exhibit the capability of citrate-production. Undifferentiated mesenchyme cells do not exhibit the capability of citrate production; and osteogenic differentiation results in citrate-producing osteoblasts. The up-regulation of zinc uptake transporter ZIP1 is essential for the manifestation of the citrate-producing capability of the osteoblasts. We determined that osteoblast transport of citrate from plasma is not a likely source of citrate in bone. Thus, this study establishes for the first time that the osteoblasts are specialized citrate-producing cells that provide the citrate for incorporation into the structure of bone; and that mesenchyme cell osteogenesis leads to differentiated citrate-producing osteoblasts. This is a new understanding; which must include the osteogenic development of citrate-producing osteoblasts, and the process of “citration” in concert with mineralization during bone formation. It also provides a new understanding of the role of bone in the homeostatic maintenance of plasma citrate concentration.

Relationship between serum uric Acid and bone mineral density in the general population and in rats with experimental hyperuricemia.

Higher serum uric acid concentrations have been associated with higher bone mineral density (BMD) in observational studies of older men and perimenopausal or postmenopausal women, prompting speculation of a potential protective effect of uric acid on bone. Whether this relationship is present in the general population has not been examined and there is no data to support causality. We conducted a cross-sectional analysis of a probability sample of the U.S. Demographic data, dietary intake, lifestyle risk factors and physical activity assessment data, serum biochemistry including serum uric acid, and BMD were obtained from 6759 National Health and Nutrition Examination Survey (NHANES; 2005-2010) participants over 30 years of age. In unadjusted analyses, higher serum uric acid levels were associated with higher BMD at the femoral neck, total hip, and lumbar spine in men, premenopausal women, and postmenopausal women not treated with estrogen. However, these associations were no longer statistically significant after adjustment for potential confounders, including age, body mass index (BMI), black race, alcohol consumption, estimated glomerular filtration rate (eGFR), serum alkaline phosphatase, and C-reactive protein (CRP). This is in contradistinction to some prevailing conclusions in the literature. To further examine the causal effect of higher serum uric acid on skeletal health, including biomechanical properties that are not measurable in humans, we used an established rat model of inducible mild hyperuricemia. There were no differences in BMD, bone volume density, and bone biomechanical properties between hyperuricemic rats and normouricemic control animals. Taken together, our data do not support the hypothesis that higher serum uric acid has protective effects on bone health.

Osteoblast-specific deletion of Pkd2 leads to low-turnover osteopenia and reduced bone marrow adiposity.

Polycystin-1 (Pkd1) interacts with polycystin-2 (Pkd2) to form an interdependent signaling complex. Selective deletion of Pkd1 in the osteoblast lineage reciprocally regulates osteoblastogenesis and adipogenesis. The role of Pkd2 in skeletal development has not been defined. To this end, we conditionally inactivated Pkd2 in mature osteoblasts by crossing Osteocalcin (Oc)-Cre;Pkd2 +/null mice with floxed Pkd2 (Pkd2 flox/flox) mice. Oc-Cre;Pkd2 flox/null (Pkd2 Oc-cKO) mice exhibited decreased bone mineral density, trabecular bone volume, cortical thickness, mineral apposition rate and impaired biomechanical properties of bone. Pkd2 deficiency resulted in diminished Runt-related transcription factor 2 (Runx2) expressions in bone and impaired osteoblastic differentiation ex vivo. Expression of osteoblast-related genes, including, Osteocalcin, Osteopontin, Bone sialoprotein (Bsp), Phosphate-regulating gene with homologies to endopeptidases on the X chromosome (Phex), Dentin matrix protein 1 (Dmp1), Sclerostin (Sost), and Fibroblast growth factor 23 (FGF23) were reduced proportionate to the reduction of Pkd2 gene dose in bone of Oc-Cre;Pkd2 flox/+ and Oc-Cre;Pkd2 flox/null mice. Loss of Pkd2 also resulted in diminished peroxisome proliferator-activated receptor γ (PPARγ) expression and reduced bone marrow fat in vivo and reduced adipogenesis in osteoblast culture ex vivo. Transcriptional co-activator with PDZ-binding motif (TAZ) and Yes-associated protein (YAP), reciprocally acting as co-activators and co-repressors of Runx2 and PPARγ, were decreased in bone of Oc-Cre;Pkd2 flox/null mice. Thus, Pkd1 and Pkd2 have coordinate effects on osteoblast differentiation and opposite effects on adipogenesis, suggesting that Pkd1 and Pkd2 signaling pathways can have independent effects on mesenchymal lineage commitment in bone.

The effect of locally delivered recombinant human bone morphogenetic protein-2 with hydroxyapatite/tri-calcium phosphate on the biomechanical properties of bone in diabetes-related osteoporosis.

BackgroundRecombinant human bone morphogenetic protein-2 (rhBMP-2) is particularly effective in improving osteogenesis in patients with diminished bone healing capabilities, such as individuals with type 1 diabetes mellitus (T1DM) who have impaired bone healing capabilities and increased risk of developing osteoporosis. This study measured the effects of rhBMP-2 treatment on osteogenesis by observing the dose-dependent effect of localized delivery of rhBMP-2 on biomechanical parameters of bone using a hydroxyapatite/tri-calcium phosphate (HA/TCP) carrier in a T1DM-related osteoporosis animal model.Materials and methodsTwo different doses of rhBMP-2 (LD low dose, HD high dose) with a HA/TCP carrier were injected into the femoral intramedullary canal of rats with T1DM-related osteoporosis. Two more diabetic rat groups were injected with saline alone and with HA/TCP carrier alone. Radiographs and micro-computed tomography were utilized for qualitative assessment of bone mineral density (BMD). Biomechanical testing occurred at 4- and 8-week time points; parameters tested included torque to failure, torsional rigidity, shear stress, and shear modulus.ResultsAt the 4-week time point, the LD and HD groups both exhibited significantly higher BMD than controls; at the 8-week time point, the HD group exhibited significantly higher BMD than controls. Biomechanical testing revealed dose-dependent, higher trends in all parameters tested at the 4- and 8-week time points, with minimal significant differences.ConclusionsGroups treated with rhBMP-2 demonstrated improved bone mineral density at both 4 and 8 weeks compared to control saline groups, in addition to strong trends towards improvement of intrinsic and extrinsic biomechanical properties when compared to control groups. Data revealed trends toward dose-dependent increases in peak torque, torsional rigidity, shear stress, and shear modulus 4 weeks after rhBMP-2 treatment.Level of evidenceNot applicable.

The antiosteoporotic activity of central-icaritin (CIT) on bone metabolism of ovariectomized rats.

Central-icaritin (CIT) is a flavonoid aglycone first discovered in our laboratory, which is an isomeric aglycone of icaritin (IT). We wanted to know whether CIT also had anti-osteoporosis activity. In this study, CIT was investigated in an ovariectomized rat (OVX) model. Fifty-six 6-month old female Sprague-Dawley rats were randomly assigned to sham operated group (Sham) and six OVX subgroups (n = 8 each). The OVX rats were then subdivided into six groups treated with vehicle (OVX), icaritin (IT, 40 mg/kg body weight/day), estradiol valerate (EV, 100 μg/kg body weight/day) or CIT (10, 20, and 40 mg/kg body weight/day) for 12 weeks, respectively. Then, the serum biochemical parameters, bone mineral density (BMD), bone biomechanical properties, bone microarchitecture, bone immunohistochemistry and related protein and gene expressions were evaluated. In OVX rats, the increases of body weight, HOP, AKP, and TRACP5b levels, and the decreases of uterus wet weight, femurs weight, BMD, serum OPG/RANKL and OCN were significantly inhibited by CIT treatment. Micro-CT analysis results showed that CIT apparently enhanced trabecular bone compared with the OVX group (p < 0.05). Total femur BMD and biomechanical strength of tibia were significantly improved (p < 0.05) after 12 weeks of CIT administration. In addition, the CIT administration also significantly enhanced the OPG expression, whereas reduced the RANKL expression in femurs according to RT-PCR, western blot assays and immunohistochemical evaluation. CIT had the antiosteoporotic activity, and its antiosteoporotic effects in OVX rats may be stronger than that of IT.

Biomechanical properties of bone in a mouse model of Rett syndrome

Rett syndrome (RTT) is an X-linked genetic disorder and a major cause of intellectual disability in girls. Mutations in the methyl-CpG binding protein 2 (MECP2) gene are the primary cause of the disorder. Despite the dominant neurological phenotypes, MECP2 is expressed ubiquitously throughout the body and a number of peripheral phenotypes such as scoliosis, reduced bone mineral density and skeletal fractures are also common and important clinical features of the disorder. In order to explore whether MeCP2 protein deficiency results in altered structural and functional properties of bone and to test the potential reversibility of any defects, we have conducted a series of histological, imaging and biomechanical tests of bone in a functional knockout mouse model of RTT. Both hemizygous Mecp2stop/y male mice in which Mecp2 is silenced in all cells and female Mecp2stop/+ mice in which Mecp2 is silenced in ~50% of cells as a consequence of random X-chromosome inactivation, revealed significant reductions in cortical bone stiffness, microhardness and tensile modulus. Microstructural analysis also revealed alterations in both cortical and cancellous femoral bone between wild-type and MeCP2-deficient mice. Furthermore, unsilencing of Mecp2 in adult mice cre-mediated stop cassette deletion resulted in a restoration of biomechanical properties (stiffness, microhardness) towards wild-type levels. These results show that MeCP2-deficiency results in overt, but potentially reversible, alterations in the biomechanical integrity of bone and highlights the importance of targeting skeletal phenotypes in considering the development of pharmacological and gene-based therapies.

Raman spectroscopy delineates radiation-induced injury and partial rescue by amifostine in bone: a murine mandibular model.

Despite its therapeutic role in head and neck cancer, radiation administration degrades the biomechanical properties of bone and can lead to pathologic fracture and osteoradionecrosis. Our laboratories have previously demonstrated that prophylactic amifostine administration preserves the biomechanical properties of irradiated bone and that Raman spectroscopy accurately evaluates bone composition ex vivo. As such, we hypothesize that Raman spectroscopy can offer insight into the temporal and mechanical effects of both irradiation and amifostine administration on bone to potentially predict and even prevent radiation-induced injury. Male Sprague-Dawley rats (350-400 g) were randomized into control, radiation exposure (XRT), and amifostine pre-treatment/radiation exposure groups (AMF-XRT). Irradiated animals received fractionated 70 Gy radiation to the left hemi-mandible, while AMF-XRT animals received amifostine just prior to radiation. Hemi-mandibles were harvested at 18 weeks after radiation, analyzed via Raman spectroscopy, and compared with specimens previously harvested at 8 weeks after radiation. Mineral (ρ958) and collagen (ρ1665) depolarization ratios were significantly lower in XRT specimens than in AMF-XRT and control specimens at both 8 and 18 weeks. amifostine administration resulted in a full return of mineral and collagen depolarization ratios to normal levels at 18 weeks. Raman spectroscopy demonstrates radiation-induced damage to the chemical composition and ultrastructure of bone while amifostine prophylaxis results in a recovery towards normal, native mineral and collagen composition and orientation. These findings have the potential to impact on clinical evaluations and interventions by preventing or detecting radiation-induced injury in patients requiring radiotherapy as part of a treatment regimen.

Abnormalities in cortical bone, trabecular plates, and stiffness in postmenopausal women treated with glucocorticoids.

The mechanisms by which glucocorticoids (GCs) increase skeletal fragility are not well understood. The objective of the study was to evaluate the microarchitecture, trabecular morphology, and biomechanical properties of bone in postmenopausal women treated with GCs. This was a case-control study. The study was conducted at a university hospital outpatient facility. Postmenopausal women treated with oral GCs for longer than 3 months (n = 30) and age/race-matched controls (n = 60) participated in the study. Areal bone mineral density aBMD (BMD) by dual-energy x-ray absorptiometry (DXA) was measured. Trabecular and cortical volumetric BMD (vBMD) and microarchitecture by high-resolution peripheral computed tomography of the distal radius and tibia were also measured. Whole-bone stiffness was estimated by finite element analysis. A novel technique, individual trabecula segmentation, was used to evaluate trabecular type (as plate or rod), orientation, and connectivity. DXA T-scores did not differ significantly at any site. GC subjects had significantly lower total, cortical, and trabecular vBMD and thinner cortices, fewer, thinner, more widely, and irregularly spaced trabeculae. They had fewer trabecular plates, fewer axially aligned trabeculae, and lower trabecular connectivity. Differences ranged from 4% to 65% for these trabecular measures and 5% to 17% for the cortical measures. Whole-bone stiffness was significantly lower (11%-16%) in GC subjects. Markers of bone formation (osteocalcin and amino-terminal propeptide of type I procollagen) and resorption (C-telopeptide) were lower in the GC subjects. Despite similar areal BMD by DXA, GC-treated women had abnormal cortical and trabecular vBMD and microarchitecture at both the radius and tibia, including fewer trabecular plates, a less axially aligned trabecular network, lower trabecular connectivity, thinner cortices, and lower whole-bone stiffness. Further research into these abnormalities as mechanisms for fracture in GC-treated women is warranted.

Partial agonist, telmisartan, maintains PPARγ serine 112 phosphorylation, and does not affect osteoblast differentiation and bone mass.

Peroxisome proliferator activated receptor gamma (PPARγ) controls both glucose metabolism and an allocation of marrow mesenchymal stem cells (MSCs) toward osteoblast and adipocyte lineages. Its activity is determined by interaction with a ligand which directs posttranscriptional modifications of PPARγ protein including dephosphorylation of Ser112 and Ser273, which results in acquiring of pro-adipocytic and insulin-sensitizing activities, respectively. PPARγ full agonist TZD rosiglitazone (ROSI) decreases phosphorylation of both Ser112 and Ser273 and its prolonged use causes bone loss in part due to diversion of MSCs differentiation from osteoblastic toward adipocytic lineage. Telmisartan (TEL), an anti-hypertensive drug from the class of angiotensin receptor blockers, also acts as a partial PPARγ agonist with insulin-sensitizing and a weak pro-adipocytic activity. TEL decreased S273pPPARγ and did not affect S112pPPARγ levels in a model of marrow MSC differentiation, U-33/γ2 cells. In contrast to ROSI, TEL did not affect osteoblast phenotype and actively blocked ROSI-induced anti-osteoblastic activity and dephosphorylation of S112pPPARγ. The effect of TEL on bone was tested side-by-side with ROSI. In contrast to ROSI, TEL administration did not affect bone mass and bone biomechanical properties measured by micro-indentation method and did not induce fat accumulation in bone, and it partially protected from ROSI-induced bone loss. In addition, TEL induced “browning” of epididymal white adipose tissue marked by increased expression of UCP1, FoxC2, Wnt10b and IGFBP2 and increased overall energy expenditure. These studies point to the complexity of mechanisms by which PPARγ acquires anti-osteoblastic and pro-adipocytic activities and suggest an importance of Ser112 phosphorylation status as being a part of the mechanism regulating this process. These studies showed that TEL acts as a full PPARγ agonist for insulin-sensitizing activity and as a partial agonist/partial antagonist for pro-adipocytic and anti-osteoblastic activities. They also suggest a relationship between PPARγ fat “browning” activity and a lack of anti-osteoblastic activity.