Long-term effects of withdrawal of bisphosphonate incadronate disodium (YM175) on bone mineral density, mass, structure, and turnover in the lumbar vertebrae of ovariectomized rats.

This study was designed to evaluate the long-term effects of incadronate disodium (YM175) after its withdrawal on cancellous bone mass in ovariectomized (OVX) rats. Thirteen-week-old female SD rats were randomized into four groups: sham-operated, OVX, low-YM, and high-YM (0.01 mg/kg or 0.1 mg/kg subcutaneously [sc], three times a week after OVX) groups. After 4 weeks of treatment with vehicle or YM175, rats from each group were killed at time points of 0 (baseline), 3, 6, 9, and 12 months after withdrawal of the agent. Bone mineral density (BMD) of the lumbar vertebrae was measured by dual-energy X-ray absorptiometry (DXA). Bone volume (BV/TV), trabecular number and trabecular separation (Tb.N and Tb.Sp), eroded surface (ES/BS), osteoclast number and osteoclast surface (N.Oc/BS and Oc.S/BS), osteoid surface (OS/BS), and bone formation rate (BFR/BS) were measured as histomorphometric parameters of the fifth lumbar vertebra. BMD, BV/TV, Tb.N, and Tb.Sp in YM175-treated groups were maintained at the same level as in the sham group until 12 months after withdrawal in the high-YM group and until 3 months after withdrawal in the low-YM group. YM175 decreased both bone formative and resorptive parameters in histomorphometry. Serum bone-specific alkaline phosphatase (ALP) and urinary deoxypyridinoline at both doses of YM175 also showed a suppressive effect of this agent on bone turnover. These results indicate that YM175, after withdrawal, still maintains bone volume dose dependently by depressing bone resorption and formation in OVX rats. Intermittent YM175 treatment with a long interval may be sufficient to maintain the bone volume and structure in OVX rats.     

Clodronate Prevents Bone Loss in Aged Ovariectomized Rats

The purpose of this study was to investigate the ability of clodronate to prevent ovariectomy (OVX)-induced osteopenia in aged rats. Fourteen-month-old female Sprague-Dawley rats (n = 166) were randomized into six groups. One group was sacrificed at the start of the study, four groups were ovariectomized, and one group was sham-operated (Sham). The OVX rats were given subcutaneously either vehicle (veh) or clodronate at doses of 3, 7, or 25 mg/kg once a week for 3 months, and the Sham rats were given the vehicle. At all dose levels clodronate inhibited trabecular bone loss in the distal femur and in the fourth lumbar vertebral body (L4), and decreased bone resorption as evidenced by urinary deoxypyridinoline excretion. The lowest dose of clodronate preserved serum osteocalcin and endosteal bone formation of secondary spongiosa in L4 at the level of the Sham/veh group. The OVX-induced increase in periosteal bone formation of femoral diaphysis was unaffected by two smaller doses of clodronate, but was decreased to the level of Sham rats after the highest dose. After 3 mg/kg clodronate, the percentage of femoral cortical bone area and the mean relative cortical thickness were higher compared with the OVX/veh group. There was a good positive correlation between the maximum load in three-point bending of the tibia and tibial ash weight. Normal lamellar pattern of newly formed cancellous and cortical bone was found after clodronate treatment. No signs of adverse accumulation of osteoid or any deleterious effect on mechanical strength of long bones and lumbar vertebrae were found. Received: 28 August 1996 / Accepted: 5 March 1997     

A Comparison of the Effect of Risedronate and Etidronate on Lumbar Bone Mineral Density in Japanese Patients with Osteoporosis: A Randomized Controlled Trial

To demonstrate the clinical benefit of 2.5 mg daily risedronate in the treatment of involutional osteoporosis, the effect of risedronate on bone mineral density (BMD) of the lumbar spine was compared with that of etidronate, selected as a representative of the bisphosphonates currently marketed in Japan. In this multicenter, randomized, double-masked, active (etidronate) controlled comparative study, a total of 235 Japanese patients with involutional osteoporosis were randomized to receive either treatment with 2.5 mg/day of risedronate for 48 weeks or intermittent treatment with etidronate (4 cycles of 2 weeks of treatment with 200 mg/day followed by 10-week medication-free periods). All patients received 200 mg of calcium supplement daily in the form of the calcium lactate. Bone mineral density of the lumbar spine (L2–L4 BMD) was determined at 12, 24, 36 and 48 weeks by dual-energy X-ray absorptiometry. The primary endpoint was the percent change in L2–L4 BMD from baseline to the time of final evaluation. Changes in biochemical markers of bone turnover and safety profiles were also compared. A significant increase in L2–L4 BMD was observed at 12 weeks after initiation of therapy in both the risedronate (2.8%) and etidronate (1.8%) groups. The increase in L2–L4 BMD at the time of final evaluation in the risedronate group (4.9%) was significantly greater (p = 0.002) than that in the etidronate group (3.1%). The changes in bone resorption markers (urinary total deoxypyridinoline and N-terminal telopeptide of type I collagen) from baseline to 48 weeks were −37.6% and −41.3% for risedronate and −22.5% and −26.6% for etidronate, respectively. New vertebral fractures or deterioration of existing fractures were observed in 2.8% (3/106) of the patients in the etidronate group, while no such cases (0/101) were observed in the risedronate group. No significant difference in the incidence of adverse events was found between two treatments. Daily oral risedronate (2.5 mg) exhibited efficacy superior to that of intermittent cyclical etidronate (200 mg) in increasing L2–L4 BMD, and was well tolerated by Japanese patients with involutional osteoporosis. Received: 7 February 2002 / Accepted: 18 July 2002     

双侧卵巢切除山羊不同时间段骨形态计量学与骨密度的变化

采用骨病理形态学、骨计量学与骨密度测定法,动态观察了山羊假手术组（Ｓｈａｍ）和双侧卵巢切除组（ＯＶＸ）术前、术后３月、６月、１２月和１８月各不同时间段髂骨骨病理形态学、骨小梁体积百分比（Ｖｖ％）,骨小梁宽度和腰椎（Ｌ２－Ｌ４）骨密度的变化。结果显示：Ｓｈａｍ组术后各时间段髂骨骨病理形态学、骨小梁体积百分比和骨小当宽度无明显变化（Ｐ〉０．０５）。Ｌ２￣Ｌ４骨密度呈缓慢上升趋势,至术后１８月比术前增高     

Mice lacking the plasminogen activator inhibitor 1 are protected from trabecular bone loss induced by estrogen deficiency.

Bone turnover requires the interaction of several proteases during the resorption phase. Indirect evidence suggests that the plasminogen activator/plasmin pathway is involved in bone resorption and turnover, and recently we have shown that this cascade plays a role in the degradation of nonmineralized bone matrix in vitro. To elucidate the role of the plasminogen activator inhibitor 1 (PAI-1) in bone turnover in vivo, bone metabolism was analyzed in mice deficient in the expression of PAI-1 gene (PAI-1-/-) at baseline (8-week-old mice) and 4 weeks after ovariectomy (OVX) or sham operation (Sham) and compared with wild-type (WT) mice. PAI-1 inactivation was without any effect on bone metabolism at baseline or in Sham mice. However, significant differences were observed in the response of WT and PAI-1-/- mice to ovariectomy. The OVX WT mice showed, as expected, decreased trabecular bone volume (BV/TV) and increased osteoid surface (OS/BS) and bone formation rate (BFR), as assessed by histomorphometric analysis of the proximal tibial metaphysis. In contrast, no significant change in any of the histomorphometric variables studied was detected in PAI-1-/- mice after ovariectomy. As a result, the OVX PAI-1-/- had a significantly higher BV/TV, lower OS/BS, lower mineral apposition rate (MAR) and BFR when compared with the OVX WT mice. However, a comparable decrease in the cortical thickness was observed in OVX PAI-1-/- and WT mice. In addition, the cortical mineral content and density assessed in the distal femoral metaphysis by peripheral quantitative computed tomography (pQCT), decreased significantly after ovariectomy, without difference between PAI-1-/- mice and WT mice. In conclusion, basal bone turnover and bone mass are only minimally affected by PAI-1 inactivation. In conditions of estrogen deficiency, PAI-1 inactivation protects against trabecular bone loss but does not affect cortical bone loss, suggesting a site-specific role for PAI-1 in bone turnover.     

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.     

Bone histomorphometry in celiac disease.

BACKGROUND AND AIMS: Bone metabolism in celiac patients is not well understood and there are no previous histomorphometric studies on bone of patients with celiac disease. The aim of the study was to investigate bone metabolism in celiac patients using bone histomorphometry, measurement of bone mineral density (BMD) and biochemical parameters. MATERIAL AND METHODS: The patient groups included 19 men and 23 women with previously diagnosed celiac disease in remission (group I), 7 women not in remission (group II) and 19 women and 9 men with newly diagnosed celiac disease (group III). Static and dynamic parameters of bone structure, formation and resorption were measured using undecalcified sections. The following parameters were obtained: bone volume (BV/TV; %), osteoid volume (OV/BV; %), osteoid surface (OS/BS; %), resoprtion surface (ES/BS; %), osteoid thickness(Oth; microm), mineral apposition rate (MAR; microm/day), and mineralization lag time (MinLag; day). BMD was measured at the spine (L2-4) and left femoral neck, trochanter and Ward's triangle. Serum calcium (S-Ca; mmol/L), alkaline phosphatase (AP; U/L), intact parathyroid hormone (S-PTH; ng/L), 25-hydroxyvitamin D (S-25(OH)D; nmol/L), cross-linked carboxyterminal telopeptide of human type I collagen (S-ICTP; microg/L) and C-terminal extension peptide of type I procollagen (S-PICP; microg/L) were analysed. RESULTS: In the histomorphometric analysis there were no significant differences in static or dynamic parameters between the groups. Very low bone mass (< 10%) was found in one patient in every group. OV/BV was increased (> 3.5%) in all groups (31% in group I, 13% in group II and 29% in group III). OS/BS was increased (> 36.5%) in groups I and III (17% and 29% respectively). In group II OS/BS was normal in all patients. Mineralization defect was found in four patients in group I and in two patients in group III with otherwise normal histomorphometric results. ES/BS was increased in three patients in group I and III and in one patient in group II. One female patient in group III had increased osteoid parameters, resorption surface, S-PTH and low concentration of vitamin D reflecting hyperparathyroid changes in bone. S-PTH was increased (66-87 ng/L) in four patients in group III and one of these patients had hyperparathyroid histological changes in bone. CONCLUSIONS: Static and dynamic histomorphometry of iliac crest bone biopsy are useful tools to evaluate bone metabolism in celiac disease especially if hyperparathyroidism or mineralization defect are suspected. Hyperparathyroidism may be a problem in the patients before introducing gluten-free diet. Mineralization defect and osteomalacic changes are common later on irrespective of whether the patients are in remission or not.     

Characterization of a new experimental model of osteoporosis in rabbits

To characterize an experimental model of osteoporosis in rabbits induced either by ovariectomy (OVX), glucocorticoids, or by a combination of both. Thirty-five rabbits were randomly allocated into five groups: bilateral OVX, daily methylprednisolone hemisuccinate (MPH) injections at a 1.5 mg/kg/day dose for 4 consecutive weeks (MPH group), or variable dose of MPH between 0.5 and 2 mg/kg/day in combination with OVX (OVX + MPH at low, medium, and high dose). Twenty-two animals were killed 6 weeks after OVX, and 13 were killed 16 weeks later. Dual-energy X-ray absorptiometry was obtained at baseline and 6 and 16 weeks after OVX. High-resolution magnetic resonance imaging (MRI) was carried out at 0 and 6 weeks after OVX. Glucose, total cholesterol, triglyceride, and oestradiol blood levels before and 16 weeks after OVX were determined. Bone mineral density (BMD) decreased significantly at lumbar spine in MPH and OVX + MPH medium-dose groups, and at global knee and subchondral bone of the knee in MPH, OVX + MPH low- and medium-dosage groups (P < 0.05). BMD variations in OVX rabbits were not significant in any of the three anatomical locations analyzed. BMD variation 16 weeks after OVX was significant at lumbar spine and global knee in the OVX + MPH medium-dose group and only at global knee in the OVX + MPH low-dose group (P < 0.05). MRI did not show bone or cartilage changes. Osteoporosis can be induced experimentally in rabbits through isolated MPH or by a combination of OVX and medium dose corticosteroid for 4 weeks. OVX alone was not sufficient to induce osteoporosis. Part of this study was presented at the 28th annual meeting of the American Society for Bone and Mineral Research, Philadelphia, PA, USA, September 15–19, 2006 [J Bone Miner Res 2006;21(suppl 1):S178]     

Effects of Treatment with Parathyroid Hormone 1–84 on Quantity and Biomechanical Properties of Thoracic Vertebral Trabecular Bone in Ovariectomized Rhesus Monkeys

Osteoporosis is characterized by impaired bone quality leading to increased susceptibility to fracture, particularly of the thoracic spine. However, the lumbar spine is studied most commonly. We investigated the effects of 16 months of treatment with full-length parathyroid hormone (PTH) 1–84 (5, 10, or 25 μg/kg) on bone mineral density (BMD) and on architecture and biomechanical properties of trabecular bone at the thoracic spine of ovariectomized (OVX) adult rhesus monkeys and compared the results with those from the lumbar spine. At baseline, 9 months after surgery, dual-energy X-ray absorptiometric BMD at T9–T12 was 7% lower in OVX than in sham animals. All PTH(1–84) doses increased BMD to sham levels within 7 months. Micro-computed tomography of T10 vertebrae showed that trabecular bone volume and connectivity were higher in PTH(1–84)-treated animals than in sham controls, primarily through a significantly greater trabecular number. Peripheral quantitative computed tomography of trabecular bone cores from T11 and T12 confirmed that PTH(1–84) increased BMD. Compression testing of the cores showed that PTH(1–84) treatment increased stiffness, modulus, yield load, and yield stress to levels significantly greater than in sham animals, with the largest effect in the 10 μg/kg group (35–54% greater than in OVX controls). Thus, PTH(1–84) treatment increased BMD and the biomechanical properties of trabecular bone at the thoracic spine of OVX rhesus monkeys. The 10 μg/kg dose produced the greatest effect on trabecular strength, possibly because the highest dose stimulated bone remodeling excessively. Importantly, the changes observed were similar to those in lumbar vertebrae, thereby validating extrapolation of results from the lumbar to the thoracic spine.     

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.     

Prevention of Bone Loss by Clodronate in Early Postmenopausal Women with Vertebral Osteopenia: A Dose-Finding Study

This double-masked, placebo-controlled study was undertaken to determine the efficacy and safety of oral clodronate in the prevention of bone loss in early postmenopausal women with vertebral osteopenia. Altogether 610 women with a mean age of 53 years were recruited for the study. They were 1–5 years postmenopausal and their lumbar spine bone mineral density (BMD) was at least 1 standard deviation below the mean of premenopausal women (T-score ≤−1). The subjects were randomized into five study groups to receive either placebo, clodronate 65 mg, 400 mg or 800 mg daily, or intermittent clodronate in 3 month cycles with 400 mg daily for 15 days followed with no treatment for 75 days for 3 years. One hundred and eighty-seven of 509 women who completed the primary study continued in the extension study of 2 years in which previous placebo users were switched to clodronate 800 mg daily, while previous users of 400 mg or 800 mg of clodronate used either placebo or 800 mg of clodronate daily. In the primary study clodronate was administered in the evening, and in the extension 1 h before breakfast on an empty stomach. In the primary study mean changes in lumbar spine BMD were −3.4% in the placebo group and +0.4% in 800 mg clodronate group [difference between groups at 3 years 3.8% (95% CI 2.7% to 4.9%, p<0.0001)], and in the trochanter area BMD −1.1% in the placebo group, and + 0.4% in the 800 mg clodronate group [difference between groups at 3 years 1.5% (95% CI 0.05% to 2.9%)]. During the extension study mean changes in lumbar spine BMD were +1.5% in the clodronate group and −0.2 % in the placebo group [difference between groups 1.7% (CI 0.4% to 3.0%, p = 0.010)] and in trochanter BMD were +2.5% in the clodronate group and no change in the placebo group [difference between groups 2.1% (CI 0.3% to 3.9%, p = 0.007)]. No statistically significant differences between the placebo and 800 mg clodronate groups were found in the femoral neck BMD. In the primary study the urinary excretion of type I collagen aminoterminal telopeptide (NTX) decreased by 44% (p<0.0001 compared with placebo) and that of deoxypyridinoline by 18% (p<0.0001) in the clodronate 800 mg group. In the extension study urinary NTX decreased by 51% (p<0.0001) in those who were switched to 800 mg of clodronate and increased by 67% (p<0.0001) in those who stopped using that dose. There was no difference in the frequency of gastrointestinal complaints between clodronate- and placebo-treated patients in the primary study, but they were more common among women who received clodronate in the extension phase. Clodronate in daily doses of 400–800 mg caused a slight elevation of aminotransferase levels, usually within the reference range. In bone biopsies no defect in mineralization was found. In conclusion, clodronate in a daily dose of 800 mg prevents early postmenopausal bone loss at the sites of the skeleton in which cancellous bone predominates. It effectively reduces bone resorption and bone turnover rate. Antifracture efficacy of clodronate remains to be established by prospective, placebo-controlled trials. Received: 4 March 2002 / Accepted: 9 July 2002     

Effects of Promethazine on Bone Mass and on Bone Remodeling in Ovariectomized Rats: A Morphometric, Densitometric, and Histomorphometric Experimental Study

The effect of promethazine on bone is debated. We studied the effect of promethazine on bone and the mechanism of action involved by densitometric and histomorphometric measurements in female Wistar rats (100 days old, mean weight 25 ± 20 g). A control group of 15 rats was not manipulated. An experimental group of 15 rats were ovariectomized (OVX) at 100 days of life and fed a diet supplemented with 4.8 mg/kg promethazine hydrochloride (OVX + Prom). The group that underwent OVX and a group of 15 rats that underwent sham ovariectomy (Sham-OVX) were not treated with promethazine. After 30 days, all the rats were killed. Their femur and 5th lumbar vertebra were dissected and cleaned of soft tissue. Femoral length and vertebral height were measured with a caliper and bones were weighed on a precision balance. The bone mineral content (BMC) and bone mineral density (BMD) of the whole right femurs and 5th lumbar vertebras were measured by dual-energy X-ray absorptiometry (DXA). Trabecular bone volume (Cn-BV-TV%), trabecular number (Tb-N mm⁻¹), trabecular thickness (Tb-Th μm), and trabecular separation (Tb-Sp μm) were measured in the femurs by histomorphometric study of nondecalcified bone. Our results showed that promethazine significantly inhibited postovariectomy loss of bone mass (P < 0.0001) by significantly reducing bone resorption, as shown by the smaller trabecular spaces observed in the treated OVX rats (P < 0.0001). Received: 1 June 1998 / Accepted: 17 February 1999     

MicroPET/CT评价去卵巢大鼠骨代谢变化的实验研究

目的探讨使用microPET/CT扫描仪对骨质疏松非创伤性探查的可行性,分析去卵巢大鼠雌激素缺乏引起骨质疏松的骨代谢变化。方法 12只6月龄雌性未孕Sprague-Dawley大鼠,体重290 g～310 g,随机分为两组(n=6):去卵巢组(OVX)和假手术组(SHAM)。用microPET/CT扫描仪进行骨显像,检测大鼠第4腰椎椎体、左股骨近端,左股骨干以及左胫骨中段对核素显像剂18F-F-的摄取。骨显像检测后腹主动脉放血法处死大鼠,取子宫称重,收集第4腰椎、左侧股骨和胫骨,用双能X线骨密度检测仪(DXA)测定第4腰椎椎体、左股骨近端,左股骨干以及左胫骨干的骨密度,取大鼠右侧胫骨制备成不脱钙硬组织切片进行形态学观察。结果 OVX组第4腰椎椎体、左股骨近端,左股骨干以及左胫骨干的骨密度比SHAM组有显著性下降。与SHAM组比较,microPET/CT骨显像可见OVX组第4腰椎椎体、左股骨近端,左股骨干以及左胫骨干核素浓聚,去卵巢大鼠骨骼兴趣区PET影像强度增加。两组大鼠胫骨干的骨几何结构参数比较无显著性差异,OVX组大鼠胫骨近端松质骨的结构参数与SHAM组比较有明显变化。OVX组大鼠胫骨近端的骨形成参数矿化表面、矿物质沉积率和骨形成率比SHAM组有显著性升高。两组大鼠胫骨第4腰椎椎体、左股骨近端,左股骨干以及左胫骨干PET影像强度与其骨密度之间呈显著性负相关。结论 MicroPET/CT可用于骨质疏松大鼠的骨代谢研究,为临床诊断骨质疏松新方法研究提供了新思路。     

骨碎补总黄酮对去卵巢大鼠下颌骨显微结构及最大载荷的影响

目的应用Micro-CT和骨生物力学技术,探讨骨碎补总黄酮对去卵巢大鼠的下颌骨显微结构及最大载荷的影响。方法40只3月龄雌性SD大鼠,随机分为5组:假手术组(sham operation group,Sham)、去卵巢模型组(ovariectomized group,OVX)、骨碎补总黄酮高剂量组(high-dose drynaria total flavonoids group,GB-H)、骨碎补总黄酮低剂量组(low-dose drynaria total flavonoids group,GB-L)和戊酸雌二醇组(Estradiol Valerate group,EV),建模成功后连续给药12 w。实验结束后,取下颌骨进行显微CT扫描及三维重建,然后进行最大载荷测量。结果与Sham组相比,OVX组大鼠下颌骨微结构:骨密度、相对骨体积、骨小梁厚度、骨小梁数目明显减小(P0.05),骨表面积体积比、骨小梁间隙明显增高(P0.05);下颌骨的最大载荷明显减少(P0.05)。与OVX组相比,EV组大鼠下颌骨微结构获得良好修复,最大载荷也明显修复。骨碎补总黄酮低剂量组相对骨体积、骨小梁数目较OVX组显著升高(P0.05),骨小梁间隙显著减小(P0.05)。骨碎补总黄酮高剂量组疗效优于低剂量组,大鼠下颌骨的相对骨体积、骨表面积体积比、骨小梁间隙、骨小梁数目以及下颌骨骨密度均得到一定程度修复,下颌骨最大载荷也较OVX组显著增加(P0.05)。结论骨碎补总黄酮能够修复去卵巢大鼠下颌骨微结构,提高下颌骨骨密度和最大载荷,这将可能为颌骨骨质疏松的防治提供一种新的途径。     

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.     

Effect of Raloxifene on Arthritis and Bone Mineral Density in Rats with Collagen-Induced Arthritis

We studied the effect of raloxifene (RAL) on arthritis and bone mineral density (BMD) in rats with collagen-induced arthritis (CIA). Seven-month-old female Sprague–Dawley rats were divided into five groups: rats without CIA (CNT), CIA rats that underwent ovariectomy (OVX) and were treated with RAL (CIA + OVX + RAL), CIA rats that underwent OVX and were treated with vehicle (CIA + OVX + Veh), CIA rats that had sham surgery and were treated with RAL (CIA + sham + RAL), and CIA rats that had sham surgery and were treated with vehicle (CIA + sham + Veh). RAL was orally administered at 10 mg/kg every day for 3 weeks, beginning 1 week after initial sensitization until death at 4 weeks. Every week until death, we evaluated hind paw thickness and arthritis score. BMD was measured by peripheral quantitative computed tomography at the distal metaphysis and the diaphysis of the femur; we also performed histomorphometry of the proximal tibia and histological evaluation of arthritis. RAL administration suppressed hind paw thickness and arthritis score and prevented decreases in BMD and cortical thickness. In the histomorphometric analysis, bone-resorption parameters were significantly lower in the RAL groups than in the Veh groups. RAL significantly inhibited synovial proliferation in CIA rats. RAL effects on arthritis and bone were apparent regardless of whether an animal had undergone OVX. RAL could suppress arthritis and bone loss in estrogen-replete or -depleted rats. These findings, using an animal model, indicate the potential usefulness of RAL as an effective treatment for premenopausal RA patients as well as postmenopausal ones.     

Correlation among geometric,densitometric, and mechanical properties in mandible and femur of osteoporotic rats

We have previously demonstrated bone loss of the mandible and femur in experimental osteoporotic rats and its prevention by medication, using peripheral quantitative computed tomography (pQCT). In the present study, the mechanical properties of the mandible and femur and the correlation to their geometric and densitometric properties were studied in ovariectomized rats with or without etidronate treatment. Fifty-four Wistar strain SPF female rats, 26 weeks old, were randomly assigned to four groups: (1) Basal group (12 rats, 1.0% Ca diet); (2) Sham group (Sham-operated, 12 rats, 0.1% Ca diet); (3) OVX group (ovariectomized, 15 rats, 0.1% Ca diet); (4) Treated group (OVX + etidronate, 15 rats, 0.1% Ca diet). Total bone mineral density (BMD), cortical BMD, cross-sectional cortical bone area, cross-sectional cortical bone thickness, crosssectional moment of inertia (CSMI), and polar strength index (SSI) of the mandible and femur were measured by pQCT. The failure load of mandible and femur was evaluated by three-point bending. The failure load of both bones was significantly lower in the Sham group compared with the Basal group. The OVX group further had a 8% and 7% decrease in the failure load for mandible and femur, respectively, compared to the Sham group. Treatment with etidronate led to an increase in the failure load compared with the OVX group. The failure load was related to the pQCT-assessed variables, especially with cortical bone area and total BMD. Moreover, the geometric and densitometric properties and failure load in the mandible showed a correlation to those in the femur.     

Influence of bone osteocalcin levels on bone loss induced by ovariectomy in rats

To investigate the role of osteocalcin (OC) in bones, bone parameters in warfarin (WF)-treated rats after ovariectomy (OVX) were compared with those in intact rats. Rats were divided into an intact group and WF-treated group. Warfarin was orally given to rats for 16 weeks, and then OVX was performed and rats in the WF-treated groups continued receiving WF. Twelve weeks after OVX, bone properties were observed. The diaphysial bone OC level in the WF group was 10%–14% of the normal level at the preoperative point and 12 weeks after surgery. On comparison of the intact and WF groups before surgery, no significant differences were noted in bone mass parameters or mechanical properties, but 12 weeks after surgery, the diaphysial bone mineral content (BMC), bone area, and cortical thickness (Cth) were significantly higher in the WF-sham group than in the intact-sham group. Ovariectomy significantly decreased the diaphysial BMC, bone mineral density (BMD), Cth, and maximum load, and increased the endosteal perimeter in the WF group. In the intact group, no such OVX-induced changes were noted, and the metaphysial bone area and the endosteal and periosteal perimeters were increased by OVX. The CO₃/PO₄ ratio in the femur measured by Fourier-transform infrared imaging using reflection preparations was higher in the WF-sham group than the intact-sham group, and higher in the intact-OVX group than the intact-sham group, but no significant difference was noted between the WF-sham and WF-OVX groups. It has been reported that CO₃ ⁻ is contained in new bone and decreases with mineral maturation. These data suggest that long-term reduction in bone OC levels may induce the formation of immature bone, which is easily resorbed with changes in bone metabolism such as OVX, and that OC may be one of the factors affecting bone turnover.     

Daily Treatment of Aged Ovariectomized Rats with Human Parathyroid Hormone (1-84) for 12 Months Reverses Bone Loss and Enhances Trabecular and Cortical Bone Strength

Most studies that have investigated the anabolic effects of parathyroid hormone (1-84) (PTH) or PTH fragments on the skeleton of ovariectomized (OVX) rats have evaluated the short-term effects of high-dose PTH(1-34) in young animals. This study used densitometry, histomorphometry, and biomechanical testing to evaluate the effects of 12-month daily treatment with low-dose PTH (15 or 30 μg/kg) in rats that were 10 months old at baseline, 4 months after OVX. Bone mineral density (BMD) and bone strength were reduced substantially in control OVX rats. The 15 μg/kg dose of PTH restored BMD to levels similar to those in sham animals within 6 months at the lumbar spine, distal and central femur, and whole body and maintained the BMD gain from 6 to 12 months. The 30 μg/kg dose produced greater effects. Both PTH doses normalized the trabecular bone volume-to-total volume ratio (BV/TV) at lumbar vertebra 3 but not at the proximal tibia (where baseline BV/TV was very low), solely by increasing trabecular thickness. PTH dose-dependently increased bone formation by increasing the mineralizing surface, but only the 30 μg/kg dose increased resorption. PTH increased cortical BMD, area, and thickness, primarily by increasing endocortical bone formation, and restored all measures of bone strength to levels similar to those in sham animals at all skeletal sites. PTH increased bone mass safely; there was no osteoid accumulation, mineralization defect, or marrow fibrosis and there were no abnormal cells. Thus, long-term PTH therapy normalized bone strength in the aged OVX rat, a model of postmenopausal osteoporosis, through increased bone turnover and enhanced formation of both trabecular and cortical bone.     

Inhibition of bone resorption by alendronate and risedronate does not require osteoclast apoptosis

Bisphosphonate inhibition of bone resorption was proposed to be due to osteoclast apoptosis. We tested this hypothesis for both the N-containing bisphosphonates alendronate and risedronate, which inhibit farnesyldiphosphate synthase and thus protein isoprenylation, and for clodronate and etidronate, which are metabolized to adenosine triphosphate (ATP) analogs. We found, in dose-response studies, that alendronate and risedronate inhibit bone resorption (in pit assays) at doses tenfold lower than those reducing osteoclast number. At an N-bisphosphonate dose that inhibited resorption and induced apoptosis, the antiapoptotic caspase inhibitor, Z-VAD-FMK, maintained osteoclast (Oc) number but did not prevent inhibition of resorption. Furthermore, when cells were treated with either alendronate alone or in combination with Z-VAD-FMK for 24 or 48 h, subsequent addition of geranylgeraniol, which restores geranylgeranylation, returned bone resorption to control levels. On the other hand, Z-VAD-FMK did block etidronate and clodronate inhibition of resorption. Moreover, in cells treated with etidronate, but not alendronate or risedronate, Z-VAD-FMK also prevented actin disruption, an early sign of osteoclast inhibition by bisphosphonates. These observations indicate that, whereas induction of apoptosis plays a major role in etidronate and clodronate inhibition of resorption, alendronate and risedronate suppression of bone resorption is independent of their effects on apoptosis.