Endocortical Mineral Apposition Rate Research Articles

Short-term glucocorticoid excess blunts abaloparatide-induced increase in femoral bone mass and strength in mice

Glucocorticoids (GCs), such as prednisolone, are widely used to treat inflammatory diseases. Continuously long-term or high dose treatment with GCs is one of the most common causes of secondary osteoporosis and is associated with sarcopenia and increased risk of debilitating osteoporotic fragility fractures. Abaloparatide (ABL) is a potent parathyroid hormone-related peptide analog, which can increase bone mineral density (aBMD), improve trabecular microarchitecture, and increase bone strength. The present study aimed to investigate whether GC excess blunts the osteoanabolic effect of ABL. Sixty 12–13-week-old female RjOrl:SWISS mice were allocated to the following groups: Baseline, Control, ABL, GC, and GC + ABL. ABL was administered as subcutaneous injections (100 μg/kg), while GC was delivered by subcutaneous implantation of a 60-days slow-release prednisolone-pellet (10 mg). The study lasted four weeks. GC induced a substantial reduction in muscle mass, trabecular mineral apposition rate (MAR) and bone formation rate (BFR/BS), and endocortical MAR compared with Control, but did not alter the trabecular microarchitecture or bone strength. In mice not receiving GC, ABL increased aBMD, bone mineral content (BMC), cortical and trabecular microarchitecture, mineralizing surface (MS/BS), MAR, BFR/BS, and bone strength compared with Control. However, when administered concomitantly with GC, the osteoanabolic effect of ABL on BMC, cortical morphology, and cortical bone strength was blunted. In conclusion, at cortical bone sites, the osteoanabolic effect of ABL is generally blunted by short-term GC excess.

Osteoporosis Treatments Affect Bone Matrix Maturation in a Rat Model of Induced Cortical Remodeling.

ABSTRACTTo test how osteoporosis drugs affect bone matrix maturation during cortical bone remodeling, 72 pregnant rats were switched from a 0.4% to a 0.01% calcium diet at parturition for a 23‐day lactation period. At weaning, eight dams were sacrificed to establish baseline values, while the remaining dams were returned to 0.4% calcium and treated with vehicle (saline), sodium fluoride (NaF), zoledronic acid (ZA), or sclerostin antibody (Scl‐Ab) for either 7 or 28 days (eight animals per group per time point). Femora were examined by μCT, dynamic histomorphometry, Fourier transform infrared imaging, and three‐point bending of notched specimens. Cortical porosity decreased in all groups from baseline to day 28. Intracortical mineralizing surface (MS/BS) and mineral apposition rate (MAR), as well as the mineral‐to‐matrix ratio were unaffected by treatment, but intracortical crystallinity was increased in the ZA group at day 10 compared with vehicle. Cortical area increased in all groups over 28 days mainly because of an addition of bone at the endocortical surface. Endocortical MS/BS did not vary among the groups, but endocortical MAR was suppressed in the NaF group at day 2 and elevated in the Scl‐Ab group at day 4 compared with vehicle. Endocortical mineral‐to‐matrix ratio was increased at days 5 and 10 following NaF treatment and endocortical crystallinity was increased at day 5 following ZA treatment compared with vehicle. Fracture toughness did not differ among the groups. Thus, the treatments affected matrix maturation more strongly at the endocortical then intracortical envelope. In this model of induced remodeling, the bone formation phase is synchronized at multiple sites, facilitating study of the effects of drugs or other bone‐targeting agents on matrix maturation independent of their effects on the initiation of remodeling. © 2020 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Thyrotoxicosis Involves β2-Adrenoceptor Signaling to Negatively Affect Microarchitecture and Biomechanical Properties of the Femur.

Background: Thyrotoxicosis increases bone turnover, resulting in net bone loss. Sympathetic nervous system (SNS) activation, via β2-adrenoceptor (β2-AR) signaling, also has osteopenic effects. Because thyroid hormones (TH) interact with the SNS to regulate several physiological processes, we hypothesized that this interaction also occurs to regulate bone mass. Previous studies support this hypothesis, as α2-AR knockout (KO) mice are less susceptible to thyrotoxicosis-induced osteopenia. Here, we evaluated whether TH-SNS interactions in bone involve β2-AR signaling. Methods: Thyrotoxicosis was induced in 120-day-old female and male mice with β2-AR gene inactivation (β2-AR-/-) by daily treatment with supraphysiological doses of triiodothyronine (T3) for 12 weeks. The impact of thyrotoxicosis on femoral bone microarchitecture, remodeling, fracture risk, and gene expression of the receptor activator of nuclear factor-kappa-B (RANK)-RANK ligand (RANKL)-osteoprotegerin (OPG) pathway was evaluated. In addition, the effect of the β2-AR-specific agonist clenbuterol (CL) on cAMP accumulation was determined in osteoblastic (MC3T3-E1) cells treated with T3 and/or 17β-estradiol (E2). Results: Thyrotoxicosis negatively affected trabecular bone microarchitecture in wild-type (WT) females, but this effect was milder or nonexistent in β2-AR-/- animals, whereas the opposite was seen in males. T3 treatment increased the femoral RANKL/OPG mRNA ratio and the endosteal perimeter and medullary area of the diaphysis in WT females and males, but not in β2-AR-/- mice, suggesting that T3 promotes endosteal resorption in cortical bone, in a mechanism that involves β2-AR signaling. T3 treatment increased endocortical mineral apposition rate only in WT females but not in β2-AR-/- mice, suggesting that TH also induce bone formation in a β2-AR signaling-dependent mechanism. T3 treatment decreased femoral resistance to fracture only in WT females, but not in KO mice. E2 and CL similarly increased cAMP accumulation in MC3T3-E1 cells; whereas T3 alone had no effect, but it completely blocked E2-stimulated cAMP accumulation, suggesting that some T3 effects on bone may involve E2/cAMP signaling in osteoblasts. Conclusions: These findings sustain the hypothesis that T3 interacts with the SNS to regulate bone morphophysiology in a β2-AR signaling-dependent mechanism. The data also reveal sex as an important modifier of skeletal manifestations of thyrotoxicosis, as well as a modifier of the TH-SNS interactions to control bone microarchitecture, remodeling, and resistance to fracture.

Sustained Notch2 signaling in osteoblasts, but not in osteoclasts, is linked to osteopenia in a mouse model of Hajdu-Cheney syndrome

Individuals with Hajdu-Cheney syndrome (HCS) present with osteoporosis, and HCS is associated with NOTCH2 mutations causing deletions of the proline-, glutamic acid-, serine-, and threonine-rich (PEST) domain that are predicted to enhance NOTCH2 stability and cause gain-of-function. Previously, we demonstrated that mice harboring Notch2 mutations analogous to those in HCS (Notch2HCS) are severely osteopenic because of enhanced bone resorption. We attributed this phenotype to osteoclastic sensitization to the receptor activator of nuclear factor-κB ligand and increased osteoblastic tumor necrosis factor superfamily member 11 (Tnfsf11) expression. Here, to determine the individual contributions of osteoclasts and osteoblasts to HCS osteopenia, we created a conditional-by-inversion (Notch2COIN ) model in which Cre recombination generates a Notch2ΔPEST allele expressing a Notch2 mutant lacking the PEST domain. Germ line Notch2COIN inversion phenocopied the Notch2HCS mutant, validating the model. To activate Notch2 in osteoclasts or osteoblasts, Notch2COIN mice were bred with mice expressing Cre from the Lyz2 or the BGLAP promoter, respectively. These crosses created experimental mice harboring a Notch2ΔPEST allele in Cre-expressing cells and control littermates expressing a wild-type Notch2 transcript. Notch2COIN inversion in Lyz2-expressing cells had no skeletal consequences and did not affect the capacity of bone marrow macrophages to form osteoclasts in vitro In contrast, Notch2COIN inversion in osteoblasts led to generalized osteopenia associated with enhanced bone resorption in the cancellous bone compartment and with suppressed endocortical mineral apposition rate. Accordingly, Notch2 activation in osteoblast-enriched cultures from Notch2COIN mice induced Tnfsf11 expression. In conclusion, introduction of the HCS mutation in osteoblasts, but not in osteoclasts, causes osteopenia.

A comparative study of the bone metabolic response to dried plum supplementation and PTH treatment in adult, osteopenic ovariectomized rat

Dried plum has been reported to have potent effects on bone in osteopenic animal models, but the mechanisms through which bone metabolism is altered in vivo remain unclear. To address this issue, a study comparing the metabolic response of dried plum to the anabolic agent, parathyroid hormone (PTH), was undertaken. Six month-old female Sprague Dawley rats (n=84) were sham-operated (SHAM) or ovariectomized (OVX) and maintained on a control diet for 6wks until osteopenia was confirmed. Treatments were initiated consisting of a control diet (AIN-93M) supplemented with dried plum (0, 5, 15 or 25%; w/w) or a positive control group receiving PTH. At the end of 6wks of treatment, whole body and femoral bone mineral density (BMD) were restored by the two higher doses of dried plum to the level of the SHAM group. Trabecular bone volume and cortical thickness were also improved with these two doses of dried plum. Dried plum suppressed the OVX-induced increase in bone turnover as indicated by systemic biomarkers of bone metabolism, N-terminal procollagen type 1 (P1NP) and deoxypyridinoline (DPD). Dynamic bone histomorphometric analysis of the tibial metaphysis revealed that dried plum restored the OVX-induced increase in cancellous bone formation rate (BFR) and mineralizing surface (MS/BS) to the SHAM group, but some doses of dried plum increased endocortical mineral apposition rate (MAR). As expected, PTH significantly increased endocortical MAR and BFR, periosteal BFR, and trabecular MAR and BFR beyond that of the OVX and maintained the accelerated rate of bone resorption associated with OVX. Dried plum up-regulated bone morphogenetic protein 4 (Bmp4) and insulin-like growth factor 1 (Igf1) while down-regulating nuclear factor T cell activator 1 (Nfatc1). These findings demonstrate that in the adult osteopenic OVX animal, the effects of dried plum differ from that of PTH in that dried plum primarily suppressed bone turnover with the exception of the indices of bone formation at the endocortical surface.

Green Tea Polyphenols Improve Bone Microarchitecture in High-Fat-Diet–Induced Obese Female Rats Through Suppressing Bone Formation and Erosion

This study evaluates the effects of green tea polyphenols (GTPs) on bone microarchitecture in high-fat-diet (HFD)-induced obese female rats. Thirty-six 3-month-old female rats were fed either a control diet or a HFD for 4 months. Animals in the control group continued on the control diet for another 4 months. Animals in the HFD group were divided into two groups, with 0.5 g/100 mL GTP (the HFD+GTP group) or without GTP (the HFD group) in drinking water, in addition to the HFD for another 4 months. Compared to the control group, the HFD group increased bone formation and erosion rates at the tibia, decreased trabecular volume and thickness, but had no impact on bone mineral density (BMD), trabecular number (Tb.N), and separation. Compared to the control group, the HFD+GTP group demonstrates a greater Tb.N at the proximal tibia, and a greater trabecular thickness at the femur and the lumbar vertebrae, but a smaller trabecular separation (Tb.Sp) and mineralizing surface at the proximal tibia, and a reduced endocortical mineral apposition rate (MAR) at the tibia shaft. Relative to the HFD group, the HFD+GTP group demonstrates (1) a higher BMD at the femur, a greater trabecular volume, thickness, and number at the proximal tibia, a larger cortical area and thickness at the tibial shaft, and a greater trabecular volume and thickness at the femur and the lumbar vertebrae, (2) a smaller Tb.Sp, MAR, bone formation rate, and eroded surface at the tibia. We concluded that GTP supplementation in drinking water improves bone microarchitecture in the HFD-induced obese female rats, possibly through suppressing bone turnover, resulting in a larger net bone volume.

Association of renal failure with poor response to PTH treatment in osteoporosis

Association of renal failure with poor response to PTH treatment in osteoporosis

Quantitative Trait Locus That Determines the Cross-Sectional Shape of the Femur in SAMP6 and SAMP2 Mice

We segregated a QTL on chromosome 11 that affects femoral cross-sectional shape during growth by generating a congenic strain and an additional 16 subcongenic strains of the senescence-accelerated mouse strain, SAMP6. The QTL region was narrowed down to a 10.0-Mbp region. Genetic background is known to affect bone characteristics. However, little is known about how polymorphic genes modulate bone shape. In a previous study using SAMP2 and SAMP6 mice, we reported a quantitative trait locus (QTL) on chromosome (Chr) 11 that had significant linkage to peak relative bone mass in terms of cortical thickness index (CTI) in male mice. We named it Pbd1. Here we aimed to clarify the effects of Pbd1 on skeletal phenotype in male mice and to narrow down the QTL region. We generated a congenic strain named P6.P2-Pbd1(b), carrying a 39-cM SAMP2-derived Chr11 interval on a SAMP6 genetic background. Sixteen subcongenic strains with smaller overlapping intervals on the SAMP6 background were generated from P6.P2-Pbd1(b) to narrow the region of interest. The effects of Pbd1 on bone properties were determined. Gene expression analysis of all candidate genes in Pbd1 was performed using real-time RT-PCR. The CTI of strain P6.P2-Pbd1(b) at 16 wk was higher than that of SAMP6. This was not caused by differences in cortical thickness but by cross-sectional shape. Morphological analysis by microCT revealed that the femoral cross-sectional shape of P6.P2-Pbd1(b) (and the other subcongenic strains with higher CTI or bone area fraction [BA/TA]) was more compressed anteroposteriorly than that of SAMP6, which was associated with superior mechanical properties. This feature was formed during bone modeling up to 16 wk of age. Subcongenic strains with a higher CTI showed significant increases in endocortical mineral apposition rate and significant reductions in periosteal mineral apposition rate at 8 wk compared with those of the SAMP6. The Pbd1 locus was successfully narrowed down to a 10.0-Mbp region, and the expression analysis suggested a candidate gene, Cacng4. The Pbd1 affects femoral cross-sectional shape by regulating the rate of endocortical and periosteal bone formation of the femur during postnatal growth.

Perturbations in bone formation and resorption in insulin-like growth factor binding protein-3 transgenic mice.

IGF-I and their binding proteins are important in bone health. Examination of BMD, osteoblast proliferation, and markers of bone resorption in transgenic mice that constitutively overexpress IGFBP-3 indicates that overexpression of IGFBP-3 increases osteoclast number and bone resorption, impairs osteoblast proliferation, and has a significant negative effect on bone formation. Low serum insulin-like growth factor I (IGF-I) levels correlate with an increased risk of osteoporotic fractures. Serum IGF-I is largely bound to IGF-binding protein-3 (IGFBP-3), which can inhibit IGF-I action and enhance delivery of IGF-I to tissues. Its role in bone biology is unclear. Bone mineral density (BMD), osteoblast proliferation, and markers of bone resorption were examined in transgenic (Tg) mice that constitutively overexpressed human IGFBP-3 cDNA driven by either the cytomegalovirus (CMV) or phosphoglycerate kinase (PGK) promoter. Cultured calvarial osteoblasts from Tg mice expressed the transgene and grew more slowly than cells from wild-type (Wt) mice, and the mitogenic response to IGF-I was attenuated in osteoblasts from Tg mice. Total volumetric BMD and cortical BMD, measured in the femur using peripheral quantitative computed tomography (pQCT) were significantly reduced in both Tg mouse strains compared with Wt mice. PGKBP-3 Tg mice showed the most marked reduction in bone density. Osteocalcin levels were similar in Wt and CMVBP-3 Tg mice but were significantly reduced in PGKBP-3 Tg mice. Urinary deoxypyridinoline and osteoclast perimeter, markers of bone resorption, were significantly increased in both Tg mouse strains compared with Wt mice. Using double labeling with tetracycline, we demonstrated that pericortical and endocortical mineral apposition rate was significantly reduced in PGKBP-3 Tg mice compared with Wt mice. These data show that overexpression of IGFBP-3 increases osteoclast number and bone resorption, impairs osteoblast proliferation, and has a significant negative effect on bone formation.

B&W WINS US$105 MILLION CONSTRUCTION PROJECT

Dried plum has been reported to have potent effects on bone in osteopenic animal models, but the mechanisms through which bone metabolism is altered in vivo remain unclear. To address this issue, a study comparing the metabolic response of dried plum to the anabolic agent, parathyroid hormone (PTH), was undertaken. Six month-old female Sprague Dawley rats (n = 84) were sham-operated (SHAM) or ovariectomized (OVX) and maintained on a control diet for 6 wks until osteopenia was confirmed. Treatments were initiated consisting of a control diet (AIN-93M) supplemented with dried plum (0, 5, 15 or 25%; w/w) or a positive control group receiving PTH. At the end of 6 wks of treatment, whole body and femoral bone mineral density (BMD) were restored by the two higher doses of dried plum to the level of the SHAM group. Trabecular bone volume and cortical thickness were also improved with these two doses of dried plum. Dried plum suppressed the OVX-induced increase in bone turnover as indicated by systemic biomarkers of bone metabolism, N-terminal procollagen type 1 (P1NP) and deoxypyridinoline (DPD). Dynamic bone histomorphometric analysis of the tibial metaphysis revealed that dried plum restored the OVX-induced increase in cancellous bone formation rate (BFR) and mineralizing surface (MS/BS) to the SHAM group, but some doses of dried plum increased endocortical mineral apposition rate (MAR). As expected, PTH significantly increased endocortical MAR and BFR, periosteal BFR, and trabecular MAR and BFR beyond that of the OVX and maintained the accelerated rate of bone resorption associated with OVX. Dried plum up-regulated bone morphogenetic protein 4 (Bmp4) and insulin-like growth factor 1 (Igf1) while down-regulating nuclear factor T cell activator 1 (Nfatc1). These findings demonstrate that in the adult osteopenic OVX animal, the effects of dried plum differ from that of PTH in that dried plum primarily suppressed bone turnover with the exception of the indices of bone formation at the endocortical surface.

Time course for bone formation with long-term external mechanical loading.

Increased mechanical loading of bone with the rat tibia four-point bending device stimulates bone formation on periosteal and endocortical surfaces. With long-term loading cell activity diminishes, and it has been reported that early gains in bone size may reverse. This study examined the time course for bone cellular and structural response after 6, 12, and 18 wk of loading at 1,200-1, 700 microstrain (muepsilon). Bone formation rates, measured by histomorphometry, were compared within groups, between loaded and contralateral nonloaded tibiae, and between weeks. Formation surface, mineral apposition rate, and bone formation rate on periosteal and endocortical surfaces were elevated after 6 wk of loading. By 12 wk of loading, periosteal and endocortical formation surface and endocortical mineral apposition rates were elevated. By 18 wk of loading, periosteal adaptation appeared complete, whereas endocortical mineral apposition rate remained elevated. No periosteal resorption was observed. Average thickness of new bone formed, from baseline to collection, was greater in loaded than nonloaded tibiae by week 6 and was maintained through week 18. Early increases in bone formation result in periosteal apposition of new bone that persists after formation ceases.

Long‐Term Effects of Nicotine on Bone and Calciotropic Hormones in Adult Female Rats

This study determined the effects of nicotine on serum concentrations of several calciotropic hormones, and bone formation and resorption end-points in 7 month old, adult female rats. Animals were administered either saline (n= 9/group), low dose nicotine at 3.0 mg/kg/day (n=10/group) or high dose nicotine at 4.5 mg/kg/day (n=11/group) by subcutaneous osmotic minipumps. At the end of a three months treatment period, serum concentrations of calcium, phosphorus, parathyroid hormone, calcitonin, 25-hydroxyvitamin D and 1,25-dihydroxyvitamin D were determined. Femora, tibiae, and lumbar vertebrae (3-5) were collected and bone parameters evaluated included mineral density and content (femora and vertebrae), strength (femora and vertebrae) and histomorphometry (tibiae). Animals given nicotine had significantly lower levels of 25-hydroxyvitamin D than controls [20.8+/-1.4 ng/ml for the low dose group and 20.7+/-1.0 ng/ ml for the high dose group versus 27.6+/-1.3 ng/ml for the control group (mean+/-S.E.M.), P<0.01]. The high dose nicotine group had smaller vertebral areas (5.4+/-0.2 mm2 versus 6.2+/-0.2 mm2, P<0.05) and a lower bone mineral content than the controls (0.024+/-0.001 g versus 0.030+/-0.001 g, P<0.05). Tibial endocortical mineral apposition rate was also significantly lower in the high dose nicotine group than in the control group (1.06+/-0.13 microm/day versus 1.42+/-0.08 microm/day. P<0.05). No significant treatment differences were detected in bone density, cancellous bone histomorphometry, or bone strength. Results from the present study suggest that nicotine administration may adversely affect bone formation and decrease body storage of vitamin D.

Systemic injection of FGF-2 stimulates endocortical bone modelling in SAMP6, a murine model of low turnover osteopenia.

The effects of systemically administered fibroblast growth factor-2 (FGF-2) at doses of 0.1 and 0.3 mg/kg/day for 7 days were investigated 5-week-old male SAMP6 mice, a model of low turnover osteopenia. The bone histomorphometry in the distal epiphyseal growth plate of the femur showed that 0.3 mg/kg/day of FGF-2 decreased the longitudinal growth rate and cartilage cell production rate and increased the growth plate width. Growth plate chondrocytes showed the features of defective endochondral ossification at the same dosage level. In the distal one third of the femur, the marrow trabecular area, endocortical mineral apposition rate and/or bone formation rate were increased in both the SAMP6 mice given 0.1 and 0.3 mg of FGF-2/kg/day. In this region, the endocortical osteoblasts were hypertrophied with some layers of overlying proliferated fibroblastic mesenchymal cells. The presence of small foci of bone formation within the layers of these mesenchymal cells indicates their osteogenic potential. On the other hand, the periosteal bone formation rate in the mid-shaft of the femur was depressed in the 0.3 mg/kg/day group. These results suggest that systemically administered FGF-2 may have the possibility to increase the peak bone mass in SAMP6 by stimulating the osteoprogenitor cells to proliferate and differentiate into osteoblasts and enhancing endocortical bone modelling. The higher dose of FGF-2, however, inhibited both endochondral and periosteal bone formation.

Distribution of mineralization indices of modeling and remodeling over eight months in middiaphyseal cross sections of femurs from adult swine

Background This study assessed the distribution of active mineral formation sites within the middiaphyseal femoral cross section of swine and determined the extent to which various subsections represented bone formation activity in the entire cross section. Methods Twenty adult female swine (sows) were injected with two double (10-day intervals) labels 8 months apart. Labels involved fluorochrome markers of active mineral formation sites. Intact femoral middiaphyseal cross sections were embedded in polymethylmethacrylate, cut, and ground to 80 μm for analysis. Each specimen was subdivided into 16 anatomical and eight geometric subsections. Labeled mineralizing surface and mineral apposition rate were determined in the periosteal and endocortical envelopes. The number of labeled osteons per unit area of bone and osteonal mineral apposition rate were determined in the intracortical envelope. Results Periosteal mineralizing surface followed a bimodal distribution with highest surface activation on the anterior and posterior segments. Periosteal mineral apposition rate followed a modal distribution with highest apposition rates in the posterior portion of the cross section. The distribution of forming osteons was modal with highest frequencies of labeled osteons in the posterior segment. No significant regional differences were detected for osteonal mineral apposition rate, endocortical mineralizing surface, or endocortical mineral apposition rate. The location of either a single or a combination of two to four subsections that best predicted mineralizing surface and mineral apposition rate in the entire cross section differed with each trait and envelope. Fifty percent of the entire area as alternate anatomical subsections was required to predict >90% of variation in all traits evaluated. Overall, the predictability of mineralizing surface and mineral apposition rate was similar for geometrically defined subsections. Conclusion At least 50% of the cross-sectional area from alternate anatomical subsections must be measured to predict >90% of variation in periosteal, intracortical, and endocortical mineralizing surface and mineral apposition rate in the sow middiaphyseal femur. Anat. Rec. 250:136–145, 1998. © 1998 Wiley-Liss, Inc.

Distribution of mineralization indices of modeling and remodeling over eight months in middiaphyseal cross sections of femurs from adult swine.

This study assessed the distribution of active mineral formation sites within the middiaphyseal femoral cross section of swine and determined the extent to which various subsections represented bone formation activity in the entire cross section. Twenty adult female swine (sows) were injected with two double (10-day intervals) labels 8 months apart. Labels involved fluorochrome markers of active mineral formation sites. Intact femoral middiaphyseal cross sections were embedded in polymethylmethacrylate, cut, and ground to 80 microm for analysis. Each specimen was subdivided into 16 anatomical and eight geometric subsections. Labeled mineralizing surface and mineral apposition rate were determined in the periosteal and endocortical envelopes. The number of labeled osteons per unit area of bone and osteonal mineral apposition rate were determined in the intracortical envelope. Periosteal mineralizing surface followed a bimodal distribution with highest surface activation on the anterior and posterior segments. Periosteal mineral apposition rate followed a modal distribution with highest apposition rates in the posterior portion of the cross section. The distribution of forming osteons was modal with highest frequencies of labeled osteons in the posterior segment. No significant regional differences were detected for osteonal mineral apposition rate, endocortical mineralizing surface, or endocortical mineral apposition rate. The location of either a single or a combination of two to four subsections that best predicted mineralizing surface and mineral apposition rate in the entire cross section differed with each trait and envelope. Fifty percent of the entire area as alternate anatomical subsections was required to predict > 90% of variation in all traits evaluated. Overall, the predictability of mineralizing surface and mineral apposition rate was similar for geometrically defined subsections. At least 50% of the cross-sectional area from alternate anatomical subsections must be measured to predict > 90% of variation in periosteal, intracortical, and endocortical mineralizing surface and mineral apposition rate in the sow middiaphyseal femur.

The effects of clodronate on increased bone turnover and bone loss due to ovariectomy in rats.

The present study was carried out to investigate the ability of clodronate to inhibit ovariectomy-induced bone loss and increased bone turnover in rats. Estradiol was administered as a reference compound. Seventy Sprague-Dawley rats were ovariectomized (OVX) or sham-operated (Sham) at the age of 90 days and divided into seven groups. Two Sham and two OVX groups received subcutaneously either the vehicle of clodronate or the vehicle of estradiol. Other OVX groups were given s.c. either disodium clodronate at two dose levels (5 mg/kg or 12.5 mg/kg twice a week) or 17 beta-estradiol (10 micrograms/kg five times a week) for 8 weeks. Femur length, volume, dry weight, and ash weight were determined, and proximal ends of tibiae were used for bone histomorphometry. Markers of bone metabolism were measured from urine and serum. A significant loss of 54% of trabecular bone area of proximal tibial metaphysis was found at 8 weeks after ovariectomy. Clodronate and estradiol inhibited (p < 0.001) this osteopenia. Both drugs prevented the decrease in ash weight/volume of the femur. The inhibitory effect of clodronate and estradiol on bone resorption in OVX rats could be detected also in decreased urinary excretion of hydroxyproline and lysylpyridinoline (p < 0.001). Clodronate and estradiol decreased (p < 0.001) the ovariectomy-induced enhanced tibial endocortical mineral apposition rate (Ec.MAR) on the lateral cortex to the level of the Sham group. In contrast, periosteal MAR analyzed on the medial side of tibial cortical bone did not change significantly in the OVX/Veh group. Estradiol decreased periosteal MAR to below the level in the Sham group (p < 0.01). These results suggest that ovariectomy of growing rats resulted in tibial and femoral osteopenia two months later. Clodronate as well as estradiol can suppress bone resorption and turnover in ovariectomized rats, inhibiting the development of osteopenia. Both clodronate doses (5 and 12.5 mg/kg) had beneficial effects in ovariectomized animals.

Droloxifene inhibits cortical bone turnover associated with estrogen deficiency in rats.

Droloxifene (DRO), an estrogen antagonist/agonist, has been shown to possess estrogen-like effects in inhibiting bone turnover leading to cancellous bone loss in ovariectomized (OVX) rats. The purpose of this study was to determine the effects of DRO on cortical bone turnover in OVX rats. Sprague-Dawley female rats at 5 months of age were sham-operated (sham, n = 8) and orally treated with vehicle, or OVX (n = 56) and orally treated with either vehicle, DRO at 0.1, 1, 5, or 10 mg/kg/day, or 17 alpha-ethynyl estradiol (EE) at 3 or 30 micrograms/kg/day for 4 weeks. Static and dynamic cortical bone histomorphometry was performed on double fluorescent labeled, undecalcified cross sections of tibial diaphyses (proximal to the tibiofibular junction). There were no significant differences in tibial diaphyseal cross sectional area, marrow cavity area, and cortical bone area between groups after 4 weeks of administration. Periosteal mineralizing surface, mineral apposition rate, and bone formation rate-surface referent and endocortical eroded surface increased significantly, while endocortical mineral apposition rate and bone formation rate-surface referent increased nonsignificantly in OVX controls compared to sham controls. Treatment with DRO at doses of 0.1 to 10 mg/kg/day dose-dependently attenuated the OVX-induced higher bone formation indices in both the periosteal and endocortical surfaces and higher bone resorption index in the endocortical surface. At the highest dose (10 mg/kg/day), DRO completely inhibited the increases in bone formation and resorption indices in OVX rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Does weight-bearing exercise affect non-weight-bearing bone?

It was previously reported that weight-bearing exercise increases femur periosteal formation in 3-year-old, 230 kg cross-bred sows. To determine if this weight-bearing exercise also stimulated non-weight-bearing bone, bone formation in the seventh rib from these same sows was measured histomorphometrically on the periosteal, endocortical, and osteonal envelopes. The sows were randomly assigned by body weight to basal (B, n = 6), control (C, n = 7), or trained (T, n = 7) groups. After 3 weeks of exercise adaptation, T walked on a treadmill for 17 weeks at 5 km/h, 20 minutes/day, 5 days/week, at 5% grade. Groups were sacrificed initially (B) or after 20 weeks (C and T). Periosteal mineral apposition rate (MAR) was calculated over 136 days. Osteonal and endocortical MAR were calculated over the 14 days prior to sacrifice. There were no effects of exercise on the periosteal or endocortical percentage double-labeled surface (dLS/BS), osteonal remodeling frequency (N.dL.On/B.Ar), or MAR in any bone envelope of the rib. In conclusion, weight-bearing exercise sufficient to stimulate periosteal formation in the femur did not activate formation in the rib. Bone response to weight-bearing exercise appears to be specific to the loaded bones.

Mineral apposition rate in rat cortical bone: physiologic differences in different sites of the same tibia.

The mineral apposition rate (MAR) is a commonly used parameter for the characterization of bone formation and is often determined to test for experimental effects on cortical bone. We investigated whether there are physiologic variations in rat cortical MAR dependent on the side or site of measurement. In our experiment we used female rats. The animals were sacrificed on day 8, after double-fluorochrome labeling with calcein and tetracycline was performed. The MAR was calculated at 3.6, 5.4, 7.2, 9, and 10.8 mm from the epiphyseal growth plate of the lateral as well as of the medial endosteum of both right and left tibiae. We found a physiologic significant difference in the MAR between the lateral and the medial endosteal sites of the same tibia (p < 0.0001), especially near the epiphyseal growth plate. Regarding the same cortical side, there is a significant decrease (p < 0.0001) in the endosteal MAR with increasing distance from the epiphyseal growth plate. We conclude that the observed differences in endocortical MAR must be due to specific mechanical challenges. Because these differences are statistically significant, it is necessary to standardize the area of histomorphometric measurement not only with respect to the distance from the epiphyseal growth plate, but also with respect to the cortical side.