The New Selective Estrogen Receptor Modulator MDL 103,323 Increases Bone Mineral Density and Bone Strength in Adult Ovariectomized Rats

Selective estrogen receptor modulators (SERMs) can prevent the bone loss induced by ovariectomy (OVX), but it is not established whether they can increase bone mass and strength in a curative protocol in ovariectomized osteopenic animals. We investigated the influence of a SERM of the new generation, MDL 103,323, on areal bone mineral density (BMD), as measured by dual-energy X-ray absorptiometry, bone strength and remodeling in OVX osteopenic rats. Nine weeks after OVX, 8-month-old rats were divided into six groups of 10 animals. MDL 103,323 was given by gavage at doses of 0.01, 0.1 or 0.6 mg/kg body weight, 5 days a week. The effect of MDL 103,323 was compared with that of the bisphosphonate pamidronate (APD), which was injected subcutaneously at a dose of 1.6 mmol/kg body weight for 5 days every 4 weeks. Lumbar spine (LS), femoral neck (FN), proximal tibia (PT) and midshaft tibia (MT) BMD, bone strength, and proximal tibia histomorphometry, serum osteocalcin, urinary total deoxypyridinoline and serum insulin-like growth factor I (IGF-I) were measured. After 16 weeks of treatment, BMD changes (means ± SEM) were −11.4 ± 2.2, +4.0 ± 2.1 and +6.4 ± 1.0% respectively in OVX controls, in rats treated with 0.1 mg/kg MDL 103,323 (p<0.05) and in APD-treated rats (p<0.02) at the level of LS; −0.4 ± 1.1, +6.7 ± 1.4, +7.2 ± 1.8% (p<0.01 and NS) at the level of FN; and −2.6 ± 1.2%, +5.8 ± 1.2, +6.9 ± 1.4% (p<0.03 and 0.01) at the level of PT. MDL 103,323-treated animals had a higher trabecular bone volume, a higher number of trabeculae and smaller intertrabecular spaces compared with OVX controls. Vertebral body ultimate strength was 186 ± 13, 292 ± 16, 249 ± 23 N (p<0.05) in OVX controls, MDL 103,323-treated rats and APD-treated rats, respectively. The administration of 0.6 mg/kg of MDL 103,323 did not further increase BMD or bone strength, indicating a bell-shaped dose–response curve. MDL 103,323 lowered plasma osteocalcin concentration and urinary deoxypyridinoline excretion. In rats treated with 0.1 mg/kg MDL 103,323, plasma IGF-I was increased as compared with OVX controls (664 ± 36 ng/ml vs 527 ± 39 ng/ml, p<0.05). In conclusion, these results indicate that this new SERM positively influences BMD and lumbar spine bone strength in estrogen-deficient rats. Received: 30 October 1998 / Accepted: 12 April 1999     

Skeletal effects of bazedoxifene paired with conjugated estrogens in ovariectomized rats

A novel approach to menopausal therapy is the tissue selective estrogen complex (TSEC) that partners bazedoxifene (BZA) with conjugated estrogens (CE). We examined the effects of daily treatment with BZA 0.3mg/kg, CE 2.5mg/kg, or combined BZA/CE (BZA 0.1, 0.3, or 1.0mg/kg with CE 2.5mg/kg) over 12months on bone mass, bone architecture and strength, and biochemical markers of bone turnover in ovariectomized (OVX) female Sprague-Dawley rats vs OVX control rats. Total cholesterol and uterine weights were also evaluated. All BZA/CE dose combinations prevented ovariectomy-induced increases in bone turnover and significantly increased bone mineral density (BMD) at the lumbar spine, proximal femur, and tibia compared with OVX controls. All BZA/CE doses evaluated also prevented many of the ovariectomy-induced changes of the static and dynamic parameters of the cortical compartment of the tibia and the cancellous compartment of the L1 and L2 vertebrae. All BZA/CE doses increased biomechanical strength at the lumbar spine (L4) compared with OVX animals. The co-administration of BZA 0.3 and 1.0mg/kg/day with CE 2.5mg/kg/day showed a dose-dependent reduction in uterine wet weight compared with administration of CE alone. All BZA/CE doses significantly lowered total cholesterol levels compared with OVX controls. In conclusion, 12months of treatment with BZA/CE in OVX rats effectively maintained BMD, bone microstructure, and bone quality; and the pairing of BZA with CE prevented CE-induced uterine stimulation.     

Long-term dosing of arzoxifene lowers cholesterol, reduces bone turnover, and preserves bone quality in ovariectomized rats.

Long-term effects of a new selective estrogen receptor modulator (SERM) arzoxifene were examined in ovariectomized (OVX) rats. Arzoxifene was administered postoperatively (po) at 0.1 mg/kg per day or 0.5 mg/kg per day to 4-month-old rats, starting 1 week after OVX for 12 months. At study termination, body weights for arzoxifene groups were 16-17% lower than OVX control, which was caused by mainly reduced gain of fat mass. Longitudinal analysis of the proximal tibial metaphysis (PTM) by computed tomography (CT) at 0, 2, 4, 6,9, and 12 months showed that OVX induced a 22% reduction in bone mineral density (BMD) at 2 months, which narrowed to a 12% difference between sham-operated (sham) and OVX rats by 12 months. Both doses of arzoxifene prevented the OVX-induced decline in BMD. Histomorphometry of the PTM showed that arzoxifene prevented bone loss by reducing osteoclast number in OVX rats. Arzoxifene maintained bone formation indices at sham levels and preserved trabecular number above OVX controls. Micro-CT analysis of lumbar vertebrae showed similar preservation of BMD compared with OVX, which were not different from sham. Compression testing of the vertebra and three-point bending testing of femoral shaft showed that strength and toughness were higher for arzoxifene-treated animals compared with OVX animals. Arzoxifene reduced serum cholesterol by 44-59% compared with OVX. Uteri wet weight from arzoxifene animals was 38-40% of sham compared with OVX rats, which were 29% of sham. Histology of the uterine endometrium showed that cell heights from both doses of arzoxifene were not significantly different from OVX controls. In summary, treatment of OVX rats with arzoxifene for nearly one-half of a lifetime maintained beneficial effects on cholesterol and the skeleton. These data suggest that arzoxifene may be a useful therapeutic agent for osteoporosis in postmenopausal women.     

Long-term minodronic acid (ONO-5920/YM529) treatment suppresses increased bone turnover, plus prevents reduction in bone mass and bone strength in ovariectomized rats with established osteopenia

The present study examined the effect of the highly potent nitrogen-containing bisphosphonate, minodronic acid (ONO-5920/YM529), on bone mineral density (BMD), bone turnover, bone histomorphometry and bone strength in ovariectomized (OVX) rats. Female F344/DuCrj rats, aged 14 weeks, were OVX or sham operated. After 3 months, the OVX rats showed an increase in bone turnover, and a decrease in bone mass and bone strength. Minodronic acid was administered orally once a day for 12 months at doses of 0, 0.006, 0.03 and 0.15 mg/kg from 3 months after OVX. Minodronic acid dose-dependently inhibited the decrease in BMD of lumbar vertebrae and femur. In the femur, treatment with 0.15 mg/kg minodronic acid increased the BMD of distal and mid sites to sham levels. Minodronic acid dose-dependently suppressed OVX-induced increase in urinary deoxypyridinoline, a bone resorption marker, after a month of treatment and these effects were maintained for 12 months of treatment. Minodronic acid also decreased serum osteocalcin, a bone formation marker. In bone histomorphometric analysis after 12 months of treatment, OVX rats showed an increase in bone resorption (Oc.S/BS and N.Oc/BS) and bone formation (MS/BS and BFR/BV) at lumbar vertebral bodies. Minodronic acid suppressed the OVX-induced increase in bone turnover at tissue level. Trabecular bone volume, trabecular thickness and trabecular number of lumbar vertebral bodies were decreased after OVX. Minodronic acid increased these structural indices, indicating that it prevented the deterioration in trabecular architecture. In a mechanical test at 12 months of treatment, ultimate load of lumbar vertebral bodies and mid femur in the OVX-control group was decreased compared to the sham group. Minodronic acid prevented the reduction in bone strength at both sites. In particular, in the mid femur, treatment with 0.03 and 0.15 mg/kg minodronic acid increased bone strength to sham levels or greater. In conclusion, minodronic acid suppressed increased bone turnover, plus prevented the decrease in BMD, deterioration of bone microarchitecture and reduction in bone strength in OVX rats with established osteopenia. These results suggest that minodronic acid may be clinically useful for treatment of osteoporosis.     

Effect of weight manipulation on bone loss due to ovariectomy and the protective effects of estrogen in the rat

Summary While characterizing the effects of estrogen on an ovariectomized (OVX) rat model of bone loss, we examined several weight-matching regimens e.g.,ad libitum (feed bins continually full), weight matched (rate of weight gain for OVX and Sham-OVX groups was equalized), and weight restricted (weight gain rates for all groups were equalized to that of estrogen-treated OVX rats) for possible effects. Bone loss following ovariectomy is primarily the result of an increase in bone resorption and is extremely sensitive to the effects of estrogens. Thus, in all of our analyses, treatment with 170-estradiol served as a positive control for the prevention of bone loss. Each weight-matching study had three groups: control (Sham-OVX), OVX, and OVX + 170 estradiol (0.1 mg/kg/day), and lasted for either 2, 4, or 6 weeks. Throughout the study, each Sprague Dawley rat was weighed every other day, and following sacrifice, a femur was removed for bone mineral density (BMD) analysis at the distal metaphysis by single photon absorptiometry. Following 2 weeks of dietary modifications, no significant differences were detected in BMD among thead lib or weight matched groups. However, an estradiol-preventable reduction in BMD in restricted OVX rats was detected at 2 weeks postovariectomy. Additionally, OVX rats in all three dietary regimens displayed an estrogen-preventable reduction in proximal femur BMD at 4 and 6 weeks postovariectomy. These results indicate that a 4-week rat ovariectomized model of bone loss, under conditions of ad libitum feeding, shows great potential for pharmacologic manipulation.     

Lycopene treatment against loss of bone mass,microarchitecture and strength in relation to regulatory mechanisms in a postmenopausal osteoporosis model

Lycopene supplementation decreases oxidative stress and exhibits beneficial effects on bone health, but the mechanisms through which it alters bone metabolism in vivo remain unclear. The present study aims to evaluate the effects of lycopene treatment on postmenopausal osteoporosis. Six-month-old female Wistar rats (n = 264) were sham-operated (SHAM) or ovariectomized (OVX). The SHAM group received oral vehicle only and the OVX rats were randomized into five groups receiving oral daily lycopene treatment (mg/kg body weight per day): 0 OVX (control), 15 OVX, 30 OVX, and 45 OVX, and one group receiving alendronate (ALN) (2 μg/kg body weight per day), for 12 weeks. Bone densitometry measurements, bone turnover markers, biomechanical testing, and histomorphometric analysis were conducted. Micro computed tomography was also used to evaluate changes in microarchitecture. Lycopene treatment suppressed the OVX-induced increase in bone turnover, as indicated by changes in biomarkers of bone metabolism: serum osteocalcin (s-OC), serum N-terminal propeptide of type 1 collagen (s-PINP), serum crosslinked carboxyterminal telopeptides (s-CTX-1), and urinary deoxypyridinoline (u-DPD). Significant improvement in OVX-induced loss of bone mass, bone strength, and microarchitectural deterioration was observed in lycopene-treated OVX animals. These effects were observed mainly at sites rich in trabecular bone, with less effect in cortical bone. Lycopene treatment down-regulated osteoclast differentiation concurrent with up-regulating osteoblast together with glutathione peroxidase (GPx) catalase (CAT) and superoxide dismutase (SOD) activities. These findings demonstrate that lycopene treatment in OVX rats primarily suppressed bone turnover to restore bone strength and microarchitecture.     

The Less Potent Estrogenic Effect of Tamoxifen on Bone in Ovariectomized Rats With Established Osteopenia

The longitudinal effects of tamoxifen (TAM) treatment on bone metabolism, spinal bone mineral density (BMD), and bone mineral content (BMC) were compared with those of estrogen in ovariectomized (OVX) rats with established osteopenia. The 6-month-old rats were divided into Sham (n = 8) and OVX (n = 24) groups. First, the OVX rats were allowed to lose bone for 6 weeks. Six weeks after ovariectomy they were divided into three groups: (1) OVX rats treated with solvent vehicle (OVX+Vehicle), (2) OVX rats injected with TAM subcutaneously six times a week at a dosage of 1.0 mg/kg body weight (OVX+TAM), (3) OVX rats injected with 17-β estradiol subcutaneously six times a week at a dosage of 0.1 mg/kg body weight (OVX+ET). The longitudinal effects of TAM and estrogen on bone were studied by dual energy X-ray absorptiometry (DXA) and biochemical markers including urinary pyridinoline (Pyr) and deoxypyridinoline (Dpyr). Ovariectomy resulted in a significant increase in urinary Pyr, Dpyr, and a significant decrease in spine BMD and BMC. TAM treatment completely inhibited the further bone loss in OVX rats with established osteopenia, however, estrogen increased spine BMD and BMC significantly compared with OVX+Vehicle, OVX+TAM, and baseline of treatment. Both TAM and estrogen treatment decreased urinary Pyr and Dpyr significantly in OVX rats. Our findings indicate that TAM acts as an estrogen agonist with respect to effects on spine BMD, BMC, and bone resorption in OVX rats with established osteopenia, but fails to restore spine BMD and BMC to the extent observed with estrogen in this study.     

Lasofoxifene (CP-336,156) protects against the age-related changes in bone mass, bone strength, and total serum cholesterol in intact aged male rats.

The purpose of this study was to evaluate if long-term (6 months) treatment with lasofoxifene (LAS), a new selective estrogen receptor modulator (SERM), can protect against age-related changes in bone mass and bone strength in intact aged male rats. Sprague-Dawley male rats at 15 months of age were treated (daily oral gavage) with either vehicle (n = 12) or LAS at 0.01 mg/kg per day (n = 12) or 0.1 mg/kg per day (n = 11) for 6 months. A group of 15 rats was necropsied at 15 months of age and served as basal controls. No significant change was found in body weight between basal and vehicle controls. However, an age-related increase in fat body mass (+42%) and decrease in lean body mass (-8.5%) was observed in controls. Compared with vehicle controls, LAS at both doses significantly decreased body weight and fat body mass but did not affect lean body mass. No significant difference was found in prostate wet weight among all groups. Total serum cholesterol was significantly decreased in all LAS-treated rats compared with both the basal and the vehicle controls. Both doses of LAS treatment completely prevented the age-related increase in serum osteocalcin. Peripheral quantitative computerized tomography (pQCT) analysis at the distal femoral metaphysis indicated that the age-related decrease in total density, trabecular density, and cortical thickness was completely prevented by treatment with LAS at 0.01 mg/kg per day or 0.1 mg/kg per day. Histomorphometric analysis of proximal tibial cancellous bone showed an age-related decrease in trabecular bone volume (TBV; -46%), trabecular number (Tb.N), wall thickness (W.Th), mineral apposition rate, and bone formation rate-tissue area referent. Moreover, an age-related increase in trabecular separation (Tb.Sp) and eroded surface was observed. LAS at 0.01 mg/kg per day or 0.1 mg/kg per day completely prevented these age-related changes in bone mass, bone structure, and bone turnover. Similarly, the age-related decrease in TBV and trabecular thickness (Tb.Th) and the age-related increase in osteoclast number (Oc.N) and osteoclast surface (Oc.S) in the third lumbar vertebral cancellous bone were completely prevented by treatment with LAS at both doses. Further, LAS at both doses completely prevented the age-related decrease in ultimate strength (-47%) and stiffness (-37%) of the fifth lumbar vertebral body. These results show that treatment with LAS for 6 months in male rats completely prevents the age-related decreases in bone mass and bone strength by inhibiting the increased bone resorption and bone turnover associated with aging. Further, LAS reduced total serum cholesterol and did not affect the prostate weight in these rats. Our data support the potential use of a SERM for protecting against the age-related changes in bone and serum cholesterol in elderly men.     

Du-Zhong (Eucommia ulmoides Oliv.) cortex extract prevent OVX-induced osteoporosis in rats

Du-Zhong, rich in polyphenolic compounds such as lignans, phenolic acid, and flavonoids, is a kidney-tonifying herbal medicine with a long history of safe use for treatment of bone fractures and joint diseases in China. In the present study, we examined whether Du-Zhong cortex extract (DZCE) with graded doses exerted its preventive effects on estrogen deficiency-induced osteoporosis. Eighty 3-month-old female Sprague-Dawley rats were used and randomly assigned into sham-operated group (Sham) and five ovariectomy (OVX) subgroups, i.e. OVX with vehicle (OVX); OVX with 17alpha-ethinylestradiol (E(2), 25 microg/kg/day); OVX with DZCE of graded doses (100, 300, or 500 mg/kg/day). Daily oral administration of DZCE or E(2) started on week 4 after OVX for 16 weeks. Treatment with DZCE at higher doses (300 or 500 mg/kg/day) was found to be able to significantly prevent OVX-induced decrease in biomechanical quality of femur such as maximum stress and Young's modulus. The mechanical changes were associated with the prevention of a further bone mineral density (BMD) decrease or even with some improvements in microarchitecture. DZCE dose-dependently inhibited total BMD decrease in the femur caused by OVX, which was accompanied by a significant decrease in skeletal remodeling, as was evidenced by the decreased levels of the bone turnover markers osteocalcin (OC), alkaline phosphatese (ALP), deoxypyridinoline (DPD), and urinary Ca and P excretions. muCT analysis of the femoral metaphysis showed that DZCE at the highest doses (500 mg/kg/day) significantly prevents decrease in bone volume/tissue volume (BV/TV), connect density (Conn.D), trabecula number (Tb.N) and trabecula thickness (Tb.Th), and increase in trabecula separation (Tb.Sp) and structure model index (SMI) in OVX rats. We conclude that 16 weeks of DZCE treatment improves bone biomechanical quality through modifications of BMD, and trabecular microarchitecture without hyperplastic effect on uterus, and it might be a potential alternative medicine for treatment of postmenopausal osteoporosis.     

Relationship between age,skeletal site,and time post‐ovariectomy on bone mineral and trabecular microarchitecture in rats

The ovariectomized (OVX) rat is widely used in osteoporosis research, but no standard model exists. The individual effects of rat age, skeletal site, and time post‐ovariectomy (post‐OVX) on bone have been examined. However, the relationship between them is not yet fully explored. This study examined how various combinations of rat age, skeletal site, and time post‐OVX affect bone mineral and microarchitecture. The rats used were 12 (n = 28), 24 (n = 28), and 44 (n = 31) weeks old. In each age group, approximately half underwent OVX and other half underwent Sham surgeries. Bone mineral (content and density) and trabecular morphology was assessed at 2, 5, 10, 15, 20, 25, and 30 weeks post‐surgery. Sites examined included the proximal tibia, spine, distal femur, and proximal femur. Overall, the proximal tibia showed the earliest and greatest differences between OVX and Sham groups. The 24‐week‐old group showed the best osteoporotic response. The 12‐week‐old group showed growth effects, whilst the 44‐week‐old group showed aging effects. The response of certain sites to OVX was also found to depend on the rat age used. These findings may aid in explaining discrepancies reported in the literature as well as synergistic combinations that may signify advanced conditions. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:189–196, 2011     

Do Different Aminobisphosphonates Have Similar Preventive Effect on Experimental Thyroid Hormone-Induced Osteopenia in Rats?

We compared similar doses of three different aminobisphosphonates (BP): olpadronate (OPD), pamidronate (APD), and alendronate (ALE) on osteopenia induced by thyroxine (T4)-treatment in OVX and SHAM adult rats. Female Sprague Dawley rats (259 +/- 8 g) were treated with vehicle (SHAM+Vh and OVX+Vh), 250 microg T4/kg/day (SHAM+T4 and OVX+T4), 0.3 mg OPD/kg/day (SHAM+OPD and OVX+OPD), 0.2 mg ALE/kg/day (SHAM+ALE and OVX+ALE), 1.5 mg APD/kg/day (SHAM+APD and OVX+APD), T4+OPD (SHAM+T4+OPD and OVX+T4+OPD), T4+ALE (SHAM+T4+ALE and OVX+T4+ALE), and T4 +APD (SHAM+T4+APD and OVX+T4+APD) during a 5-week period. At the onset and at the end of the experiment, total skeleton bone mineral density (BMD) was assessed in vivo by DXA. Lumbar spine and proximal tibia BMDs were evaluated. T4 treatment to SHAM rats did not modify BGP levels significantly: neither did ovariectomy. T4 treatment to OVX rats significantly increased bone-gla-protein (BGP) levels compared with the other studied groups (P < 0.05). BP treatment reduced BGP levels to values significantly lower than SHAM rats (P < 0.05) and reduced bone alkaline phosphatase in SHAM groups (P < 0.05) but no changes were found in OVX groups. The increased D-Pyr excretion observed in SHAM+T4 rats (P = 0.056), OVX+Vh (P < 0.05), and OVX+T4 group (P < 0.001) compared with the SHAM+Vh rats was prevented by the BP treatment. OVX+Vh rats had total skeleton and proximal tibia BMD, and OVX+T4 group had total skeleton, spine, and proximal tibia BMD significantly lower than the SHAM+Vh group. BP treatment was also found to prevent this reduction. The reduced bone resorption and the prevention of bone loss showed no differences among very close, potentially equivalent doses of the three aminoBPs used. Consequently, treatment with very close similar doses of APD, ODP, and ALE prevented bone resorption and bone changes with the same efficacy.     

Thyroid-stimulating hormone restores bone volume, microarchitecture, and strength in aged ovariectomized rats.

We show the systemic administration of low levels of TSH increases bone volume and improves bone microarchitecture and strength in aged OVX rats. TSH's actions are mediated by its inhibitory effects on RANKL-induced osteoclast formation and bone resorption coupled with stimulatory effects on osteoblast differentiation and bone formation, suggesting TSH directly affects bone remodeling in vivo. INTRODUCTION: Thyroid-stimulating hormone (TSH) receptor haploinsufficient mice with normal circulating thyroid hormone levels have reduced bone mass, suggesting that TSH directly affects bone remodeling. We examined whether systemic TSH administration restored bone volume in aged ovariectomized (OVX) rats and influenced osteoclast formation and osteoblast differentiation in vitro. MATERIALS AND METHODS: Sprague-Dawley rats were OVX at 6 months, and TSH therapy was started immediately after surgery (prevention mode; n = 80) or 7 mo later (restoration mode; n = 152). Hind limbs and lumbar spine BMD was measured at 2- or 4-wk intervals in vivo and ex vivo on termination at 8-16 wk. Long bones were subjected to microCT, histomorphometric, and biomechanical analyses. The direct effect of TSH was examined in osteoclast and osteoblast progenitor cultures and established rat osteosarcoma-derived osteoblastic cells. Data were analyzed by ANOVA Dunnett test. RESULTS: In the prevention mode, low doses (0.1 and 0.3 microg) of native rat TSH prevented the progressive bone loss, and importantly, did not increase serum triiodothyroxine (T3) and thyroxine (T4) levels in aged OVX rats. In restoration mode, animals receiving 0.1 and 0.3 microg TSH had increased BMD (10-11%), trabecular bone volume (100-130%), trabecular number (25-40%), trabecular thickness (45-60%), cortical thickness (5-16%), mineral apposition and bone formation rate (200-300%), and enhanced mechanical strength of the femur (51-60%) compared with control OVX rats. In vitro studies suggest that TSH's action is mediated by its inhibitory effects on RANKL-induced osteoclast formation, as shown in hematopoietic stem cells cultivated from TSH-treated OVX rats. TSH also stimulates osteoblast differentiation, as shown by effects on alkaline phosphatase activity, osteocalcin expression, and mineralization rate. CONCLUSIONS: These results show for the first time that systemically administered TSH prevents bone loss and restores bone mass in aged OVX rats through both antiresorptive and anabolic effects on bone remodeling.     

Effects of a new selective estrogen receptor modulator (MDL 103,323) on cancellous and cortical bone in ovariectomized ewes: a biochemical, histomorphometric, and densitometric study.

The aims of this study performed in ewes were: (1) to confirm in this animal model the effects on bone of ovariectomy (OVX) alone or associated with Lentaron (L), a potent peripheral aromatase inhibitor, used to amplify the effects of OVX and (2) to evaluate the effects of a new selective estrogen receptor modulator (SERM; MDL 103,323) on bone remodeling. Thirty-nine old ewes were divided into five groups: sham (n = 7); OVX (n = 8); OVX + L (n = 8); OVX + L + MDL; 0.1 mg/kg per day (n = 8); and OVX + L + MDL 1 mg/kg per day (n = 8). The animals were treated for 6 months. Biochemical markers of bone turnover (urinary excretion of type 1 collagen C-telopeptide [CTX], serum osteocalcin [OC], and bone alkaline phosphatase [BAP]) were measured each month. Bone biopsy specimens were taken at the beginning and after death at the end of the experiment. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry (DXA) on the lumbar spine and femur. OVX induced a significant increase in biochemical markers. This effect was the highest after 3 months for CTX (+156% vs. sham) and after 4 months for OC and BAP (+74% and +53% vs. sham, respectively). L tended to amplify the effect of OVX on OC and BAP. OVX induced significant increases in the porosity, eroded, and osteoid surfaces in cortical bone but no effect was observed in cancellous bone. MDL treatment reduced the bone turnover as assessed by bone markers, which returned to sham levels as well as histomorphometry both in cortical and in cancellous bone. Cancellous osteoid thickness decreased by 27% (p < 0.05), mineralizing perimeter by 81% (p < 0.05), and activation frequency by 84% (p < 0.02) versus OVX + L. Femoral and spinal BMD were increased by MDL and tended to return to the sham values. The effects of OVX on bone turnover were different on cortical and cancellous bone. These effects on cortical bone were reflected by changes in biochemical markers. MDL markedly reduces bone turnover and increases BMD suggesting that this new agent may prevent postmenopausal bone loss.     

Prolonged Treatments With Antiresorptive Agents and PTH Have Different Effects on Bone Strength and the Degree of Mineralization in Old Estrogen‐Deficient Osteoporotic Rats

Current approved medical treatments for osteoporosis reduce fracture risk to a greater degree than predicted from change in BMD in women with postmenopausal osteoporosis. We hypothesize that bone active agents improve bone strength in osteoporotic bone by altering different material properties of the bone. Eighteen‐month‐old female Fischer rats were ovariectomized (OVX) or sham‐operated and left untreated for 60 days to induce osteopenia before they were treated with single doses of either risedronate (500 μg/kg, IV), zoledronic acid (100 μg/kg, IV), raloxifene (2 mg/kg, PO, three times per week), hPTH(1–34) (25 μg/kg, SC, three times per week), or vehicle (NS; 1 ml/kg, three times per week). Groups of animals were killed after days 60 and 180 of treatment, and either the proximal tibial metaphysis or lumbar vertebral body were studied. Bone volume and architecture were assessed by μCT and histomorphometry. Measurements of bone quality included the degree of bone mineralization (DBM), localized elastic modulus, bone turnover by histomorphometry, compression testing of the LVB, and three‐point bending testing of the femur. The trabecular bone volume, DBM, elastic modulus, and compressive bone strength were all significantly lower at day 60 post‐OVX (pretreatment, day 0 study) than at baseline. After 60 days of all of the bone active treatments, bone mass and material measurements agent were restored. However, after 180 days of treatment, the OVX + PTH group further increased BV/TV (+30% from day 60, p < 0.05 within group and between groups). In addition, after 180 days of treatment, there was more highly mineralized cortical and trabecular bone and increased cortical bone size and whole bone strength in OVX + PTH compared with other OVX + antiresorptives. Treatment of estrogen‐deficient aged rats with either antiresorptive agents or PTH rapidly improved many aspects of bone quality including microarchitecture, bone mineralization, turnover, and bone strength. However, prolonged treatment for 180 days with PTH resulted in additional gains in bone quality and bone strength, suggesting that the maximal gains in bone strength in cortical and trabecular bone sites may require a longer treatment period with PTH.     

Comparative effects of 17β-estradiol,raloxifene and genistein on bone 3D microarchitecture and volumetric bone mineral density in the ovariectomized mice

Summary This study assessed the effect of estradiol, raloxifene and genistein on the preservation of bone 3D-microarchitecture and volumetric bone mineral density (vBMD) in the ovariectomized mouse model. Our results indicated that raloxifene was more effective in preserving bone ovariectomized-induced changes, the advantage being concentrated in both bone microarchitecture and vBMD. Introduction This study assessed the effect of different estrogen receptor (ER) agonists on the preservation of bone 3D-microarchitecture and volumetric bone mineral density (vBMD) in the ovariectomized (OVX) mouse model. Methods Twelve-week-old female C57BL/6 mice were randomly assigned to one of five groups: (1) SHAM-operated + vehicle; (2) OVX + vehicle; (3) OVX + 17β-estradiol (5 μg/kg); (4) OVX + raloxifene (1 mg/kg); (5) OVX + genistein (25 mg/kg), during 4-weeks. Bone microarchitecture and trabecular, cortical and total vBMD of distal femur were imaged by ex vivo microcomputed tomography (micro-CT). Results Ovariectomy produced a global deterioration involving both trabecular and cortical 3D-microarchitecture and vBMD. Raloxifene maintained both microarchitecture and vBMD, whereas estradiol prevented deterioration of some microstructural parameters, such as trabecular thickness (Tb.Th), trabecular bone pattern factor (Tb.Pf), and cortical periosteal perimeter (Ct.Pe.Pm), but did not completely block the loss in vBMD. Mice treated with genistein exhibited the less favourable profile in both vBMD and microstructural parameters preserving only cross-sectional bone area (B.Ar) and Ct.Pe.Pm in cortical bone. Conclusion Our data indicate that, at the selected doses, raloxifene was more effective in preserving bone OVX-induced changes than either estradiol or genistein, the advantage being concentrated in both bone microarchitecture and vBMD.     

Effect of silicon supplement on osteopenia induced by ovariectomy in rats

The effect of silicon (Si) supplement on preventing bone mass loss induced by ovariectomy (OVX) in rats was investigated. Three groups of 15, 100-day-old female Wistar rats each, with a mean initial weight of ∼260 g per animal, were selected for the present study. One of the experimental group consisting of 15 OVX rats was fed a diet supplemented with 500 mg of Si per kg of feed (Si + OVX). The other two groups consisting of 15 OVX and 15 sham-OVX rats did not receive these supplements. Morphometric (weight and length) and densitometric studies with dual-energy X-ray absorptiometry were performed on the whole femur and 5th lumbar vertebra of each animal 30 days after the experiment. The Si + OVX rats did not show a loss of bone mass induced by OVX at axial level (5th lumbar vertebra) or periphery (femur). Nonetheless, a significant increase (ANOVA with Bonferroni/Dunn post hocs test) of longitudinal development of the femur (P < 0.0001) was patent. These results, obtained through the measurements of axial and peripheral bones, warrant closer scrutiny in connection with the Si inhibitory effect on bone mass loss as well as the stimulatory effect on bone formation. Both actions, namely, inhibition of resorption and stimulation of formation, infer that Si may have a potential therapeutic application in the treatment of involutive osteoporosis. Received: 15 February 1999 / Accepted: 25 June 1999     

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 17β-estradiol (10 μg/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.     

The effects of nandrolone decanoate on bone mass and metabolism in ovariectomized rats with osteopenia

The effects of nandrolone decanoate (ND) treatment on bone mass and metabolism were studied in ovariectomized (OVX) rats with osteopenia. The 6-month-old rats were divided into Sham (n = 12) and OVX (n = 24). The OVX rats were allowed to lose bone for 6 weeks. At 6 weeks post ovariectomy, the OVX rats were divided into two groups: (1) OVX + Vehicle and (2) OVX + ND. The effects of ND on bone mineral density (BMD), bone mineral content (BMC), and bone metabolism were studied by dual-energy X-ray absorptiometry (DXA) and biochemical markers including urinary pyridinoline (Pyr), deoxypyridinoline (Dpyr), and serum osteocalcin. After 24 weeks of treatment, histomorphometry of the right tibiae and the wet weight of the gastrocnemius and soleus skeletal muscles were also examined. Ovariectomy resulted in a significant increase in biochemical markers and a significant decrease in spine BMD (0.221 ± 0.016 g/cm² in OVX group vs 0.239 ± 0.008 g/cm² in Sham group) and BMC (0.550 ± 0.055 g in OVX group vs 0.605 ± 0.042 g in Sham group) at 6 weeks post ovariectomy. Spine BMD (0.227 ± 0.017 g/cm²), femoral BMD (0.263 ± 0.012 g/cm²), and bone density of femur (1.035 ± 0.036 g/cm³) in the OVX + ND group were significantly greater than those in the OVX + Vehicle group (0.204 ± 0.013 g/cm² for spine BMD, 0.243 ± 0.009 g/cm² for femoral BMD, 0.938 ± 0.06 g/cm³ for bone density of femur) after 24 weeks of treatment. ND treatment decreased urinary Pyr and Dpyr significantly in OVX rats. Histomorphometric findings indicated that ND-treated rats had greater cancellous bone volume, greater trabecular number, greater trabecular thickness, and less trabecular separation than vehicle-treated OVX rats. OVX rats had greater wet weight of the gastrocnemius and soleus muscles than rats treated with ND. The data suggest that the effect of ND on bone mass is not influenced by the condition of the muscles in OVX rats. Our findings indicate that ND blocks further bone loss by inhibition of bone resorption in OVX rats with osteopenia. Received: August 25, 1999 / Accepted: January 28, 2000     

Romosozumab Improves Bone Mass and Strength While Maintaining Bone Quality in Ovariectomized Cynomolgus Monkeys

Romosozumab (Romo), a humanized sclerostin antibody, is a bone‐forming agent under development for treatment of osteoporosis. To examine the effects of Romo on bone quality, mature cynomolgus monkeys (cynos) were treated 4 months post‐ ovariectomy (OVX) with vehicle, 3 mg/kg, or 30 mg/kg Romo for 12 months, or with 30 mg/kg Romo for 6 months followed by vehicle for 6 months (30/0). Serum bone formation markers were increased by Romo during the first 6 months, corresponding to increased cancellous, endocortical, and periosteal bone formation in rib and iliac biopsies at months 3 and 6. Dual‐energy X‐ray absorptiometry (DXA) bone mineral density (BMD) was increased by 14% to 26% at the lumbar spine and proximal femur at month 12, corresponding to significant increases in bone strength at 3 and 30 mg/kg in lumbar vertebral bodies and cancellous cores, and at 30 mg/kg in the femur diaphysis and neck. Bone mass remained positively correlated with strength at these sites, with no changes in calculated material properties at cortical sites. These bone‐quality measures were also maintained in the 30/0 group, despite a gradual loss of accrued bone mass. Normal bone mineralization was confirmed by histomorphometry and ash analyses. At the radial diaphysis, a transient, reversible 2% reduction in cortical BMD was observed with Romo at month 6, despite relative improvements in bone mineral content (BMC). High‐resolution pQCT confirmed this decline in cortical BMD at the radial diaphysis and metaphysis in a second set of OVX cynos administered 3 mg/kg Romo for 6 months. Radial diaphyseal strength was maintained and metaphyseal strength improved with Romo as estimated by finite element modeling. Decreased radial cortical BMD was a consequence of increased intracortical remodeling, with no increase in cortical porosity. Romo resulted in marked improvements in bone mass, architecture, and bone strength, while maintaining bone quality in OVX cynos, supporting its bone efficacy and safety profile. © 2016 American Society for Bone and Mineral Research.     

Changes in proximal femur bone properties following ovariectomy and their association with resistance to fracture

Bone strength depends on several material and structural properties, but findings concerning the best predictors of bone mechanical performance are conflicting. The aim of this study was to investigate how a broad set of bone properties in the proximal femur are influenced by age and hormonal status, and how these properties together determine bone strength. Twenty-five Wistar rats were ovariectomized (OVX, n = 13) or sham operated (SHAM, n = 12) at 5 months of age, and killed after 9 months. Another group of rats was killed at 5 months as baseline control (BSL, n = 7). At sacrifice, serum 17β-estradiol and bone turnover marker concentrations were determined in the serum. Both femurs were collected for assessment of trabecular microarchitecture, femoral neck geometry, radiographic absorptiometry, calcium and phosphate content, and biomechanical properties. While stiffness was mostly associated with proximal femur trabecular microarchitecture and mineralization degree, bone strength was mostly linked to bone size and femoral neck geometry, which predicted almost 50% of its variance. Despite the decrease in cortical and trabecular bone as well as in mineralization degree following estrogen loss, bone strength was not reduced in OVX animals compared to BSL or sham-operated rats. This was due to a change in femoral neck geometry as well as to an increase in femur size in OVX, which apparently compensated their lower bone volume and mineral content, thereby preserving bone strength. Estrogen loss leads to a deterioration of bone tissue quality, but bone strength was preserved at the expense of geometric adaptations.