Temporal relationship between bone loss and increased bone turnover in ovariectomized rats

Summary To characterize osteopenic changes in ovariectomized (OVX) rats as a function of time, female Sprague Dawley rats (240 g body weight, 90 days old) were subjected to bilateral ovariectomy or sham surgery and killed at various times from 14–180 days postovariectomy. The proximal tibial metaphysis was processed undecalcified for quantitative bone histomorphometry. Osteopenia and increased indices of bone resorption and formation were detected in OVX rats as early as 14 days. Longitudinal bone growth was also significantly increased by ovariectomy at 14 days, but returned to control levels at all later times. In OVX rats, osteopenia became progressively more pronounced with time up to 100 days postovariectomy, after which trabecular bone volume appeared to stabilize at the markedly reduced level of 5%. Changes in osteoclast surface, osteoblast surface, and fluoro-chrome-based indices of bone formation in OVX rats followed a similar time course. The maximal increase in these parameters occurred during the first several months postovariectomy followed by a gradual decline toward control levels. Our results indicate that the initial rapid phase of bone loss in OVX rats is coincident with the maximal increase in bone turnover. At later times postovariectomy, bone loss and bone turnover both subside. These findings emphasize the close temporal association between the development of osteopenia and increased bone turnover in OVX rats.     

Effects of low-dose long-term sodium fluoride preventive treatment on rat bone mass and biomechanical properties

Effects of fluoride on bone strength and cortical bone mass remain controversial. We compared 9-month, low-dose sodium fluoride (NaF) treatment with estrogen replacement therapy. Female Wistar rats 4.5 months old were divided into baseline, sham-operated (sham), sham-treated with NaF at 0.5 mg NaF/kg/day in drinking water, and ovariectomy (OVX), OVX treated with NaF and with estrogen. Bone mass was measured by dual X-ray absorptiometry (DXA)in vitro. Dimensions of the first lumbar vertebral body (L1) were determined by radiogrammetry. The right femur was processed undecalcified to obtain a midshaft cross-section to determine cross-sectional moments of inertia (CSMIs). L1 compressive test and left femoral torsional test were performed. OVX induced significant bone loss in L1 and femoral midshaft. Bone mass was increased to a greater extent in NaF-treated rats than in rats receiving estrogen replacement therapy. Femoral CSMIs in OVX rars, both L1 sizes and femoral CSMIs in NaF-treated rats, were significantly increased. Estrogen treatment had the least dimension expansion. OVX significantly decreased L1 compressive variables. There was no statistical difference in compressive parameters between NaF-treated groups and controls. OVX significantly increased femoral torsional strength but NaF treatment did not. Bone fluoride content was significantly increased after treatment with NaF. No significant difference in bone mineralization degree (ash and calcium) was found between treated and control rats. The discrepancy that an increase in bone mass and geometric properties in both trabecular and cortical bones by low-dose, long-term NaF treatment did not increase vertebral strength nor proportionally improve femoral strength indicated that the bone intrinsic biomechanical properties could be changed by NaF treatment.     

A comparative study of the bone-restorative efficacy of anabolic agents in aged ovariectomized rats

Introduction The study was designed to compare the bone anabolic effects of basic fibroblast growth factor (bFGF), a selective agonist for prostaglandin E receptor subtype EP4, and parathyroid hormone (PTH) in aged ovariectomized (OVX) rats with severe cancellous osteopenia. Methods Groups of aged OVX rats were maintained untreated for 1 year postovariectomy (15 months of age) to develop severe tibial cancellous osteopenia. These animals were then treated with bFGF or the EP4 agonist (EP4) for 3 weeks. Other groups of aged OVX rats were treated with EP4 or PTH alone for 11 weeks, or sequentially with bFGF or EP4 for 3 weeks followed by PTH for 8 weeks. Cancellous and cortical bone histomorphometry were performed in the right proximal tibial metaphysis and tibial diaphysis respectively. Results Treatment with bFGF for 3 weeks markedly increased serum osteocalcin, osteoid volume, and osteoblast and osteoid surfaces to a greater extent than EP4. Basic FGF, but not EP4 or PTH, induced formation of osteoid islands within bone marrow. EP4 stimulated cancellous bone turnover, but failed to restore lost cancellous bone in the severely osteopenic proximal tibia after 11 weeks of treatment. In contrast, EP4, much like PTH, increased cortical bone mass in the tibial diaphysis by stimulating both periosteal and endocortical bone formation. Treatment of aged OVX rats with PTH alone tended to partially reverse the severe tibial cancellous osteopenia, whereas sequential treatment with bFGF and PTH increased tibial cancellous bone mass to near the level of vehicle-treated control rats. These findings indicate that bFGF had the strongest stimulatory effect on cancellous bone formation, and was the only anabolic agent to induce formation of osteoid islands within the bone marrow of the severely osteopenic proximal tibia. Therefore, bFGF may be more effective for the reversal of severe cancellous osteopenia. PTH and EP4 increased cortical bone mass to nearly the same extent, but cancellous bone mass was greater by two-fold in PTH-treated OVX rats than in EP4-treated OVX rats. Conclusion These findings in aged OVX rats suggest that PTH is more efficacious than EP4 for augmentation of cancellous bone in the severely osteopenic, estrogen-deplete skeleton.     

Short-Term Prophylaxis against Estrogen Depletion-Induced Bone Loss with Calcitriol does not Provide Long-Term Beneficial Effects on Cancellous Bone Mass or Structure in Ovariectomized Rats

It was the aim of the present study to investigate whether a 2-month prophylaxis of postovariectomy bone loss with low-dose calcitriol would have long-lasting beneficial effects on cancellous bone mass or structure after its withdrawal in rats. Six-month-old female Fischer 344 rats were either ovariectomized (OVX) or sham-operated (SHAM). Groups of SHAM and OVX rats were orally treated with either 0.05 μg calcitriol/kg per day or vehicle for 2 months postovariectomy, starting immediately after ovariectomy. Thereafter, the rats were maintained without treatment for another 4 months. Half the animals in each group were killed 2 months postovariectomy; the rest of the rats were killed 6 months postovariectomy. Cancellous bone histomorphometry was performed on the first lumbar vertebral body and on the proximal tibial metaphysis. Administration of low-dose calcitriol to SHAM and OVX rats resulted in hypercalciuria, but not hypercalcemia. By 2 months postovariectomy, calcitriol treatment of OVX rats had completely prevented tibial trabecular bone loss, and had increased vertebral cancellous bone mass in SHAM and OVX rats by about 30% over the level observed in SHAM vehicle controls. However, at the end of the experiment, i.e. 4 months after withdrawal of calcitriol, cancellous bone mass and structure in both the vertebrae and the tibiae of calcitriol-treated OVX rats were almost identical to those of vehicle-treated OVX rats. We conclude that prevention of bone loss with low-dose calcitriol during the phase of acute estrogen deficiency, when bone turnover is maximally increased, does not provide long-term beneficial effects on cancellous bone mass or structure in OVX rats. If extrapolated to postmenopausal women, this study would suggest that prophylaxis against postmenopausal bone loss with short-acting antiresorptive substances during only the first few years after menopause will probably not reduce the risk of postmenopausal osteoporosis later in life.     

Basic fibroblast growth factor has rapid bone anabolic effects in ovariectomized rats

Basic fibroblast growth factor (bFGF) has a strong bone anabolic effect in intact and ovariectomized (OVX) rats treated for 7–14 days. Other growth factors such as IGF-I and TGF- have been implicated as potential mediators for this effect. The purpose of this study was to examine the early effects of bFGF therapy, in vivo, on bone formation and gene expression in OVX rats in order to determine whether upregulation of gene expression for IGF-I and/or TGF- precedes or coincides with the stimulatory effects of bFGF on bone formation. At 3 months of age, Sprague Dawley rats were OVX or sham-operated (SHAM), then maintained untreated for 3 months. One group of baseline OVX rats (BSL OVX) and BSL SHAM rats were then killed. Additional OVX groups were treated IV with bFGF at a daily dose of 200 g/kg and killed at 1–7 and 10 days. Another group of OVX rats was treated IV with vehicle daily for 10 days, then killed. Lumbar vertebrae were processed for cancellous bone histomorphometry or RNA isolation. Ovariectomy induced increased cancellous bone turnover and a significant decrease in vertebral bone mass. Treatment of OVX rats with bFGF resulted in a significant increase in bone formation. As early as 24 h after bFGF treatment of OVX rats, osteoblast surface, osteoid surface, and osteoid volume were more than double those in BSL OVX rats and continued to increase with time. These variables were also significantly higher in bFGF-treated OVX rats at 10 days compared with vehicle-treated OVX rats. Gene expression for IGF-I was not different between BSL OVX rats and bFGF-treated OVX rats at 1 day, but was significantly higher by approximately 50% in OVX rats treated with bFGF for 2 and 7 days, and was also significantly higher by nearly 75% in OVX rats treated for 10 days compared with OVX rats treated with vehicle. Gene expression for TGF-₁ was unchanged at early times and only significantly upregulated by a relatively modest 30% in OVX rats treated with bFGF for 10 days. The results indicate that the bone anabolic effects of bFGF in OVX rats begin as early as 24 h following the initial treatment, and increase with time. These early stages of the strong stimulatory effect of bFGF on bone formation were not associated with a large upregulation of gene expression for IGF-I and TGF-. The rapid increase in osteoblast surface in bFGF-treated OVX rats suggests that the growth factor induces conversion of bone lining cells to osteoblasts.     

Short-term effects of organic silicon on trabecular bone in mature ovariectomized rats

Summary Silicon is known to ensure an essential role in the formation of cross-links between collagen and proteoglycans during bone growth. In this study, we have evaluated the short-term effects of a preventive treatment with silanol, a soluble organic silicon (Si), on trabecular bone in mature ovariectomized rats. Three-month-old rats were shamoperated (sham) or were ovariectomized (OVX) and treated with 10 g/kg/day of 170 estradiol (E2), or with 0.1 mg Si/kg/day or 1.0 mg Si/kg/day of silanol for 1 month. Plasma alkaline phosphatase and osteocalcin levels were increased by 50% in OVX rats compared with sham rats and were corrected by E2 but not by silanol treatment. The trabecular bone volume measured at the tibial metaphysis was decreased by 48%, and histomorphometric indices of bone resorption and formation were increased in OVX rats compared with sham, and these parameters were corrected by E2 treatment. Treatment of OVX rats with silanol decreased the osteoclast surface by 31% and the number of osteoclasts by 20%. The mineral apposition rate, the bone formation rate, and the osteoblast surface at the tibia metaphyseal area were increased by 30% at the higher dose of silanol compared with OVX rats. In contrast, silanol treatment had no effect on the periosteal apposition rate. The reduction of the metaphyseal bone resorption and the increased bone formation induced by silanol resulted in a slight improvement of the trabecular bone volume (+ 14%) compared with controls. The results indicate that a short-term preventive treatment with the organic silicon silanol partially prevented the trabecular bone loss in mature OVX rats by reducing bone resorption and increasing bone formation, possibly through stimulatory effects on the formation and/or the stability of the organic bone matrix.     

Sequential treatment with basic fibroblast growth factor and parathyroid hormone restores lost cancellous bone mass and strength in the proximal tibia of aged ovariectomized rats.

This study was designed to determine whether sequential treatment with basic fibroblast growth factor (bFGF) and parathyroid hormone (PTH) can restore lost cancellous bone mass and strength at a severely osteopenic skeletal site in aged ovariectomized (OVX) rats. Female Sprague-Dawley rats were subjected to sham surgery or ovariectomy at 3 months of age and maintained untreated for the first year after surgery. At 15 months of age, groups of baseline control and OVX rats were killed and catheters were inserted in the jugular veins of all remaining rats. Two groups of OVX rats were injected intravenously (iv) daily with bFGF for 14 days at a dose of 200 microg/kg body weight. At the end of bFGF treatment, one group was killed whereas the other group was subjected to 8 weeks of treatment with synthetic human PTH 1-34 [hPTH(1-34)] consisting of subcutaneous (sc) injections 5 days/week at a dose of 80 microg/kg. Another group of OVX rats was treated iv with vehicle for 2 weeks followed by treatment with PTH alone for 8 weeks. Other groups of sham-operated control rats and OVX rats were treated iv and sc with vehicle alone. The right proximal tibia from each rat was processed undecalcified for quantitative bone histomorphometry and the left proximal tibia was subjected to biomechanical testing. Baseline and vehicle-treated OVX rats were severely osteopenic because their tibial cancellous bone volumes were less than 5% compared with mean values of 20.3% and 15.0% in baseline and vehicle-treated control rats, respectively. Treatment of OVX rats for 2 weeks with bFGF alone did not significantly increase tibial cancellous bone volume but induced marked increases in osteoid volume, osteoblast surface, and osteoid surface. Sequential treatment of aged OVX rats with bFGF and PTH increased tibial cancellous bone volume (15.1%) and load to failure to at least the level of vehicle-treated control rats. Tibial cancellous bone volume (10.8%) and load to failure also were significantly increased by treatment with PTH alone, and these variables were not significantly different from those of OVX rats treated with bFGF + PTH. However, tibial ash density was significantly greater in OVX rats treated sequentially with bFGF and PTH compared with OVX rats treated with PTH alone. Our findings suggest that sequential treatment with bFGF and PTH may be useful for restoration of lost cancellous bone in the severely osteopenic, estrogen-deplete skeleton, but it cannot be concluded with certainty that this sequential treatment has a greater bone restorative effect than treatment with PTH alone.     

Fluoride-induced bone changes in lambs during and after exposure to sodium fluoride

The evolution of bone changes induced by fluoride after the end of exposure was investigated in lambs. Sodium fluoride (NaF) was given orally at a dose of 3.5 mg/kg per day to 14 animals for 120 days. A group of 7 control and 7 treated lambs was slaughtered at the end of NaF administration (T120) and another group 120 days after the end of NaF exposure (T240). At T120, the bone fluoride content (BFC) was very significantly increased in treated animals. The histomorphometric analysis confirmed that fluoride induces an increase in bone formation (the osteoid perimeter and area were 3-fold and 4.5-fold higher respectively in treated than in control animals). The number of osteoblasts was significantly augmented. Serum osteocalcin level was twice as high in treated animals compared with controls. The bone formation rate at the tissue level (BFR) doubled after treatment, but the apposition rate (Aj.AR) was half that in the control group. The mineralization lag time (Mlt) was 120 days in treated animals compared with 42 days in controls. At T240, BFC had decreased by 50% compared with the level at T120, but it was still significantly higher than in controls. The osteoid and osteoblastic parameters were 2 and 1.3 times higher than in control animals. BFR remained significantly increased in treated animals, but Aj.AR and Mlt were similar in control and treated animals. In conclusion, after 4 months of NaF exposure fluoride induced an increase in osteoblast natality and bone formation at the tissue level, associated with a toxic effect at the individual cell level. Four months after the end of NaF exposure, positive effects on bone formation were still present but the evidence of cellular toxicity had disappeared.     

Effects of progesterone on postovariectomy bone loss in aged rats

The effects of progesterone on oophorectomy-induced bone loss in aged rats were evaluated. Female rats aged 12 months were divided into three groups: (1) sham-operated controls (SHAM); (2) oophorectomized (OVX); (3) OVX rats treated with progesterone (OVX + PROG). After 20 weeks the dry weight, bone ash, and calcium content of femur, tibia, and fourth lumbar vertebra were significantly lower in OVX than in sham rats. These reductions did not occur in OVX rats treated with PROG. There was no difference in the bone composition between the control and progesterone-treated rats. Vertebral bone histomorphometry showed increased bone resorption as well as increased bone formation parameters in OVX rats. Progesterone treatment inhibited the increased resorption indices, but the bone formation remained elevated. The results indicate that progesterone therapy prevents the postovariectomy bone loss in aged rats. The protective effect of progesterone is mediated by inhibition of bone resorption while maintaining the increased bone formation. These findings suggest that progesterone alone may be a valuable agent for management of postmenopausal osteoporosis.     

The Effect of Tamoxifen on Bone Metabolism and Skeletal Growth is Different in Ovariectomized and Intact Rats

The effects of tamoxifen (TAM) treatment on bone metabolism and skeletal growth were studied in sexually mature intact or ovariectomized (OVX) rats. Experiment 1 was designed to observe the effects of TAM on bone metabolism and skeletal growth in intact rats and included two groups: (1) intact plus vehicle and (2) intact plus TAM. Experiment 2 was designed to investigate the effects of TAM on OVX rats and included the other two groups: (3) OVX plus vehicle and (4) OVX plus TAM. Serum calcium osteocalcin and urinary pyridinoline (Pyr) and deoxypyridinoline (Dpyr) were measured serially before and after TAM treatment for 6 weeks in order to monitor bone turnover. Bone mineral density (BMD) and bone mineral content (BMC) of excised right femora and lumbar vertebrae were determined by dual energy X-ray absorptiometry (DXA). To examine the effect of TAM on skeletal growth, the conventional parameters of femora and the histology of right tibiae were also measured. TAM did not induce significant change in the biochemical markers in intact rats during the 6-week experiment. Bone mass and skeletal growth were not changed by TAM treatment in intact rats. However, TAM treatment reduced the increase in serum osteocalcin and urinary pyridinium cross-links from 1 week to 6 weeks postovariectomy in the OVX rats. TAM inhibited the skeletal growth in OVX rats, because TAM treatment shortened femoral length and decreased the cell number in the growth plate in OVX rats in this study. Our findings indicate that TAM exerts an effect of estrogen agonist on bone metabolism and skeletal growth in OVX rats, however, it does not affect them in intact rats. Received: 1 September 1995 / Accepted: 20 February 1996     

重组人甲状旁腺素（1-34)与雌激素对去卵巢大鼠骨代谢的影响

目的观察重组人甲状旁腺素(1-34)与雌激素单用和联用对去卵巢大鼠骨代谢的影响。方法选用雌性4月龄SD大鼠45只,随机分为5组:①假手术(Sham)8只;②去卵巢(OVX)9只;③雌激素治疗组(OVX E)10只:OVX2个月后给予苯甲酸雌二醇治疗6w;④PTH治疗组(OVX PTH)9只:OVX2个月后给予rhPTH(1-34)治疗6w;⑤雌激素与PTH联合治疗组(OVX E PTH)9只:OVX2个月后给予rhPTH(1-34)和E2联合治疗6w。观察各组大鼠胫骨近端松质骨骨小梁形态计量学参数,椎体生物力学指标及部分血清骨生化代谢指标。结果雌激素治疗组和PTH治疗组的骨静态参数均表现为骨量增加;雌激素治疗组的骨形成参数和骨吸收参数降低;PTH治疗组的骨形成指标明显升高,而骨吸收指标虽较Sham高,但较OVX组有所降低;E2与PTH联合治疗组与单用E2组和单用PTH组比较,骨量明显提高,骨转换率参数介于单用E2和单用PTH组之间。3个治疗组的生物力学指标较OVX组均有明显提高,其中E2与PTH联合治疗组改善最明显。结论雌激素和PTH可使去卵巢大鼠松质骨骨量增加和改善生物力学性能,两者联合治疗有协同作用。     

Effect of parathyroid hormone (hPTH[1-84]) treatment on bone mass and strength in ovariectomized rats.

Skeletal fragility in osteoporotic patients is a prominent underlying cause of low-trauma fractures of most bone sites in humans. Clinical research is now focused on developing treatment strategies, including anabolic agents such as parathyroid hormone (PTH), to recover osteoporosis-related bone loss. Female Sprague-Dawley rats (4.5 mo old) were allowed to become osteopenic for 10 wk postovariectomy. Eight rats were killed at the time of ovariectomy (-10 wk) as a baseline control; sham and ovariectomized (OVX) groups were killed at wk 0. Eight rats per group (sham, OVX + vehicle, OVX + hPTH [5 d/wk], and OVX + hPTH [3 d/wk]) were killed after 4, 8, 14, and 20 wk of treatment with 50 microg/kg of human parathyroid hormone (hPTH[1-84]). Bone mineral content and density were measured only in the vertebral body. Bone strength was evaluated in the vertebral body, femoral diaphysis, femoral neck, and distal femur. Significant, lasting osteopenia developed in the vertebral body of OVX rats by 10 wk postovariectomy. Bone mineral density of the vertebral body partially recovered by 8 wk and fully recovered to that seen in sham animals only by 20 wk posttreatment with either a 5 or 3 d/wk dosing schedule of PTH[1-84]. Therefore, hPTH[1-84] (50 microg/kg) given either 3 or 5 d/wk fully restores vertebral and femoral bone strength in osteopenic OVX rats.     

虾青素对去卵巢糖尿病大鼠骨质流失的防治作用

目的探讨虾青素能否防治去卵巢糖尿病大鼠的骨质流失以及可能的机制。方法 3月龄雌性SD大鼠分为3组(每组6只):对照组CON(假手术),模型组OVX/T1DM(去卵巢糖尿病大鼠),药物组OVX/T1DM-ASX(去卵巢糖尿病大鼠,给予虾青素100 mg/(kg·d)。结果连续治疗60 d后,与OVX/T1DM组相比,OVX/T1DM-ASX组骨密度(BMD)明显升高(P0.01),血清I型胶原蛋白(CTX-1)、骨钙素、I型前胶原蛋白n端前肽(PINP)、抗酒石酸磷酸酶5b(TRACP 5b)水平均显著升高(P0.01)。虾青素治疗能抑制去卵巢糖尿病大鼠骨组织形态学的改变,减少骨髓脂肪细胞增加,提高OPG/RANKL的比值。结论虾青素对绝经后糖尿病骨质流失有保护作用,这种作用与调控OPG/RANKL轴有关。     

Prostaglandin E2 enhances cortical bone mass and activates intracortical bone remodeling in intact and ovariectomized female rats

To assess the efficacy of prostaglandin E2 (PGE2) in augmenting cortical bone mass, graded doses of PGE2 were subcutaneously administered for 30 days to seven-month old sham-ovariectomized (SHAM) and ovariectomized (OVX) rats. Both groups were operated at three months of age. Histomorphometric analyses of double fluorescent labeled tibial shafts were performed on basal control, OVX, and SHAM rats treated with 0, 0.3, 1, 3, and 6 mg PGE2/kg/d for 30 days. Baseline aging data showed increased cortical tissue and cortical bone area and reduced bone formation parameters at the periosteal and endocortical bone envelopes between three and eight months of age. The tibial shafts of OVX rats compared to SHAM controls showed elevated periosteal mineral apposition rate and endocortical bone formation parameters. PGE2 administration to OVX and SHAM rats increased cortical bone by the addition of new circumferential bone on the endocortical and periosteal surfaces, as well as woven cancellous bone in the marrow region. Stimulated osteoblastic recruitment and activity enhanced bone formation at all bone surfaces. The new bone was both lamellar and woven in nature. PGE2 treatment also activated intracortical bone remodeling (not seen in untreated eight-month old rats), creating a porous cortex. Thus, PGE2 administration activated cortical bone modeling in the formation mode (A----F), as well as intracortical bone remodeling (A----R----F). PGE2 administration to OVX rats resulted in more intracortical bone remodeling, periosteal bone formation, and new cancellous bone production than observed in PGE2 treated controls. The findings that PGE2 administration to OVX and intact female rats increases cortical bone mass, coupled with observations that mouse, rat, dog, and man respond similarly to PGE2, suggest that PGE2 administration may be useful in the prevention and treatment of postmenopausal osteoporosis.     

Captopril improves osteopenia in ovariectomized rats and promotes bone formation in osteoblasts

The present study was designed to investigate the effects of captopril, an angiotensin-converting enzyme inhibitor (ACEI), on bone loss in aged ovariectomized (OVX) rats and its impact on the differentiation of cultured primary osteoblasts. Ten-month-old female Sprague–Dawley rats were used for the study. After 2 months post ovariectomy (OVX), the rats were treated with captopril (1 or 5 mg/kg/day, respectively) for another 2 months. At endpoint, trabecular bone of the fourth lumbar vertebrae (L4) was undecalcified and examined by bone histomorphometry; the fifth lumbar vertebrae (L5) were examined by compression test. Primary osteoblasts were isolated from the calvaria of newborn rats and treated with different concentrations of captopril in a different durations. The content of secreted alkaline phosphatase (ALP) and mRNA expression of collagen I in osteoblasts were determined to demonstrate osteoblast bone formation. In aged rats with estrogen deficiency-induced osteopenia, captopril increased the trabecular area (%BV/TV) of L4 up to 33% and improved biomechanical properties by increasing L5 break stress and elastic modulus when compared to those in the OVX group (P < 0.01). Captopril showed dose-dependent effects on promoting the secretion of ALP and increased mRNA expression of collagen I in the cultured rat osteoblasts. In summary, captopril, one of the most widely used ACEIs, has the potential effects of improving lumbar vertebral bone strength in aged OVX rats and promoting osteoblast bone formation in vitro.     

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.     

复方仙贞汤对去卵巢大鼠模型骨密度和骨形态的影响

目的 研究复方仙贞汤对去卵巢大鼠的骨密度和骨形态的影响。方法 3月龄的雌性SD大鼠，普通饲料喂养1周后，随机分为假手术组（Sham）、模型组（OVX）、尼尔雌醇阳性药物对照组、复方仙贞汤小剂量组、复方仙贞汤中剂量组、复方仙贞汤大剂量组。除假手术组，其余都经背部切开后切除双侧卵巢，制造去卵巢模型（OVX），假手术组则背部切开后切除部分脂肪组织再分层缝合。造模完成后，假手术组及模型组用生理盐水灌胃，阳性药物组用尼尔雌醇混悬液灌胃，复方仙贞汤小、中、大剂量组分别给以6．5g／kg、13g／kg、26g／kg大鼠体重（按生药计算）的复方仙贞汤药液，每日固定上午九点灌胃1次。全部动物共持续喂养3个月后处死，测定股骨的骨密度和胫骨骨体积、胫骨类骨表面，类骨质宽、骨吸收表面。结果 复方仙贞汤组和尼尔雌醇组均较模型组的骨密度显著性增高（P〈0．01），而且复方仙贞汤中剂量组与尼尔雌醇阳性对照组接近。与模型组比较，中药组和尼尔雌醇组的骨体积显著上升（P〈0．01），而类骨表面、类骨质宽和骨吸收表面却显著下降（分别P〈0．01）。结论 复方仙贞汤能提高去卵巢大鼠的骨密度和骨体积，减少类骨质宽、类骨质表面、骨吸收表面，提示复方仙贞汤能起到和尼尔雌醇相似的作用，即降低绝经后骨质疏松的高转换率，从而减慢骨质的丢失。     

Simvastatin treatment partially prevents ovariectomy-induced bone loss while increasing cortical bone formation

Statins are commonly prescribed drugs that inhibit hepatic cholesterol synthesis and thereby reduce serum cholesterol concentrations. Some of the statins are thought to possess bone anabolic properties. Effects of statin on tibia, femur, and vertebral cortical and cancellous bone were studied in ovariectomized (OVX) rats. Sixty Wistar female rats, 4 months old, were allocated into four groups: baseline control, sham + placebo group, OVX + placebo, OVX + simvastatin. Simvastatin, 20 mg/kg, or placebo was given twice daily by a gastric tube for 3 months. The rats were labeled with tetracycline at day 11 and calcein at day 4 before sacrifice. Concerning cortical bone, the tibial diaphysis bending strength was increased by 8% and the periosteal bone formation rate (BFR) at the mid-diaphysis increased by twofold in the OVX + simvastatin group compared with the OVX + placebo group, in harmony with increased serum osteocalcin concentrations. Simvastatin did not affect the endocortical bone formation. Concerning cancellous bone, the cancellous bone volumes in the proximal tibia and vertebral body were reduced in both OVX groups, but the reduction was less in the OVX + simvastatin group compared with the OVX + placebo group. This reduction in cancellous bone loss is in agreement with the 36% decreased activity of serum tartrate-resistant-acid-phosphatase 5b (TRAP-5b), indicating decreased osteoclast activity in the OVX + simvastatin group compared with the OVX + placebo group. In conclusion, simvastatin induces a moderate increase in cortical bone formation at the periosteal bone surface. The new cortical bone exhibits a normal lamellar structure, and simvastatin seems to respect the regional pattern of bone formation, bone resorption, and drift; for example, no periosteal bone formation is observed in the vertebral canal. Furthermore, simvastatin reduces the loss of cancellous bone induced by ovariectomy.     

Calcitonin Provides Complete Protection Against Cancellous Bone Loss in the Femoral Neck of Ovariectomized Rats

Calcitonin (CT) has been found to partially prevent cancellous bone loss in the proximal tibia of ovariectomized (OVX) rats. The current study was designed to determine whether CT has similar bone protective effects in the femoral neck, a skeletal site with a slower rate of bone loss after ovariectomy than the proximal tibia. Female Sprague Dawley rats were sham-operated or ovariectomized at 3 months of age. Groups of OVX rats were injected s.c. with vehicle or CT at a dose of 16 U/kg body weight on alternate days for 30, 60, or 90 days. Sham-operated control rats were treated with vehicle alone on alternate days. The proximal femur from each rat was processed undecalcified for quantitative bone histomorphometry. Cancellous bone volume in the femoral neck of vehicle-treated OVX rats was significantly less than that of vehicle-treated control rats at all time points. This cancellous osteopenia induced by ovariectomy was associated with increased indices of bone turnover such as osteoclast surface, osteoblast surface, and bone formation rate (tissue level, total surface referent). In contrast, cancellous bone volume in the femoral neck of CT-treated OVX rats was nearly identical of that of vehicle-treated control rats throughout the study. In addition, CT treatment of OVX rats decreased all indices of bone turnover to near the level of vehicle-treated control rats. The results indicate that CT treatment depresses bone turnover and provides complete protection against moderate cancellous osteopenia in the femoral neck of OVX rats. Since previous studies have shown that CT only partially protects against more pronounced cancellous bone loss in the proximal tibia of OVX rats, our findings suggest that CT has a greater bone protective effect at a skeletal site with a slower rate of cancellous bone loss (femoral neck) than at a skeletal site with a rapid rate of cancellous bone loss (proximal tibia). Received: 21 August 1996 / Accepted: 10 December 1996     

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.