Effect of incadronate on corticosteroid-induced osteopenia in rats

The effect of incadronate, a third-generation bisphosphonate, was evaluated in rats with corticosteroid-induced osteopenia. Male Wistar rats were treated with methylprednisolone acetate (1 mg/kg, s.c.) once daily, 3 days a week for 12 weeks. Other groups received simultaneous treatment with methylprednisolone acetate and incadronate (0.03, 0.3 or 3 mg/kg, p.o.); incadronate was given once daily, 6 days a week for 12 weeks. Bone mineral densities (BMDs) of the second lumbar (L2) vertebra as well as the ultimate compressive strength of the fifth lumbar (L5) vertebra decreased. Incadronate dose-dependently inhibited the loss of L2 BMDs and the decrease in strength of the L5 vertebrae. These results suggest that incadronate may be effective in treating osteopenia accompanying corticosteroid therapy.     

Pharmacokinetic and pharmacodynamic analysis of the antihypercalcemic effect of incadronate disodium in rats.

Incadronate concentrates into the bone as a target organ after intravenous administration of incadronate disodium. Mature osteoclasts has take up incadronate from the bone surface and convert it from an active to an inactive form. As a result, incadronate decreases the plasma calcium concentration by suppressing bone resorption. In this study, the pharmacokinetic and pharmacodynamic (PK/PD) analysis model for ascertaining the antihypercalcemic effects of incadronate disodium was developed in rats. Data on both the concentration of incadronate in bone and that of free calcium in blood after intravenous administration from our previous study were used for analysis. To estimate the concentration in the surface layer of bone, data on the concentration of incadronate in bone after single intravenous administration were analyzed based on the PK model considering three-compartments. The estimated concentrations in the surface layer in bone were applied to the PD model as an input function. The PD model was developed to analyze the changes in the plasma calcium concentration after a single intravenous administration considering an irreversible inhibition of osteoclast activity. The obtained fitted curves were in good agreement with the observed data. The model could explain the long duration of the antihypercalcemic effect of incadronate disodium and should be useful for planning rational dose regimens for effective antihypercalcemic therapy.     

Comparison of incadronate and alfacalcidol on increased bone turnover caused by ovariectomy in rats

Mineral density of trabecular bone at the metaphyses of right tibiae was measured by peripheral quantitative computed tomography (pQCT) in ovariectomized rats. Bone mineral density (BMD) decreased dramatically in the 4 weeks following ovariectomy, suggesting that the method is sensitive enough to detect decreased bone mineral density within a short period. Orally administered incadronate dose dependently inhibited the decrease in trabecular bone mineral density induced by ovariectomy, as assessed 4 weeks after surgery. Significant inhibition was observed at doses of more than 0.3 mg/kg/day. Moreover, incadronate at doses of 1 mg/kg or more inhibited the increase in urinary deoxypyridinoline levels induced by ovariectomy, and although slightly increased serum intact parathyroid hormone (PTH) levels were observed, no significant alteration in serum calcium ion levels or urinary calcium excretion occurred. In contrast, while alfacalcidol inhibited the decrease in bone mineral density and the increase in urinary deoxypyridinoline levels at a dose of 300 ng/kg, it significantly lowered serum intact PTH levels and elevated serum free calcium levels as well as urinary calcium excretion. These results suggest that incadronate exerts its pharmacological effect (inhibition of bone resorption and increase in bone mass) by a mechanism different from that of alfacacidol.     

Effects of XT-44, a phosphodiesterase 4 inhibitor, in osteoblastgenesis and osteoclastgenesis in culture and its therapeutic effects in rat osteopenia models.

We have reported that denbufylline, a phosphodiesterase 4 (PDE4) inhibitor, inhibits bone loss in Walker256/S tumor-bearing rats, suggesting therapeutic potentiality of a PDE4 inhibitor in osteopenia. In the present study, effects of a new PDE4 inhibitor, 1-n-butyl-3-n-propylxanthine (XT-44), in bone were evaluated in cell cultures and animal experiments. In rat bone marrow culture, XT-44 stimulated mineralized-nodule formation, whereas it inhibited osteoclast-like cell formation in mouse bone marrow culture. In Walker256/S-bearing rats (6-week-old female Wistar Imamichi rats), rapid decrease in bone mineral density (BMD) was prominent, and oral administration of XT-44 (0.3 mg/kg, every 2 days) inhibited the decrease in BMD. In the second animal experiment, female Wistar rats (6-week-old) were sciatic neurectomized, and XT-44 was orally administered to these rats every 2 days for 4 weeks. XT-44 administration (0.3 mg/kg) recovered BMD in these neurectomized animals. Furthermore, 19-week-old, female Wistar rats were ovariectomized (OVX), and 15 weeks after surgery, these rats were orally administered XT-44 every 2 days for 8 weeks. XT-44 treatment (1 mg/kg) increased the BMD of OVX rats. These results indicate that XT-44 could be a candidate as a therapeutic drug for treating osteopenia including osteoporosis.     

Effect of KW-8232 on bone turnover in ovariectomized rats

The effects of 3-[bis(4-hydroxyphenyl)methyl]-2-[4-(2-chlorophenyl)- piperazin-1-ylcarbonyl]-1-(2-dimethylamino ethyl) indole methanesulfonate (KW-8232) on the bone turnover in ovariectomized (OVX) rats were studied by the oral administration for 6 weeks. KW-8232 inhibited the decreased bone mineral densities of the femur and tibia in OVX rats. OVX induced the increase of serum alkaline phosphatase and osteocalcin levels, markers of high bone turnover, and also increased urinary hydroxyproline, pyridinoline and deoxypyridinoline excretion, markers of bone resorption. KW-8232 significantly inhibited the increase of serum alkaline phosphatase level at doses of 10 and 30 mg/kg and the increase of osteocalcin level at a dose of 3 mg/kg. KW-8232 (1 mg/kg) markedly suppressed the OVX-induced increase of urinary hydroxyproline, pyridinoline and deoxypyridinoline excretion. KW-8232 didn't affect serum calcium level, but significantly decreased urinary calcium excretion at doses of 10 and 30 mg/kg. These results suggest that KW-8232 decreased bone loss in this model by suppressing bone resorption.     

Effects of methylprednisolone on bone formation and resorption in rats

Excessive glucocorticoids induce osteoporosis. However, there is some controversy regarding the mechanism of action, and even the endpoint result. The present study was carried out to obtain further insight into the action of glucocorticoids on bone formation and resorption in rats. Growing rats were injected subcutaneously with methylprednisolone (mPSL) at doses of 0, 2.5, 5, 10 or 20 mg/kg per day for 4 weeks. Bone mineral density (BMD), enchondral and periosteal bone formation, collagen synthetic activities of osteoblasts, numbers of osteoblasts and osteoclasts, and serum markers to assess bone turnover were determined. Administration of mPSL dose-dependently increased the BMD in the tibial metaphysis, while it dose-dependently decreased the BMD in the diaphysis. Both enchondral and periosteal bone formation were decreased in a dose-dependent fashion. The incorporation and secretion of (3)H-proline by osteoblasts were both decreased in trabecular and cortical bones. The number of osteoclasts, together with the number of osteoblasts, in the tibial metaphysis was drastically decreased. Serum alkaline phosphatase and osteocalcin were decreased at higher doses. These results support the recent notion that glucocorticoids inhibit both bone formation and resorption. In addition, BMD as an endpoint result might differ from site to site in bone due to a different balance between bone formation and resorption.     

A combination of subtherapeutic doses of chemically modified doxycycline (CMT-8) and a bisphosphonate (clodronate) inhibits bone loss in the ovariectomized rat: a dynamic histomorphometric and gene expression study

Recent studies have demonstrated that tetracyclines can reduce bone loss in the ovariectomized (OVX) rat model of osteoporosis. In the current study, a non-antimicrobial, chemically modified doxycycline (CMT-8), alone or in combination with a bisphosphonate (Clodronate), was evaluated in this model. Forty-two, 6month old, female rats were randomly assigned to the following groups, (6/ group): a) sham/vehicle, b) OVX/vehicle; c) OVX/1 mg/day CMT-8; d) OVX/2 mg/day CMT-8, e) OVX/1 mg/week Clodronate; and f) OVX/1 mg/day CMT-8 + 1 mg/week Clodronate, CMT-8 was administered by oral gavage, Clodronate injected S/C. Following sham surgery or OVX, the rats were treated for 90 days with CMT-8 or vehicle alone, injected at three different times with fluorochrome labels, the rats were sacrificed, and the tibiae excised for analysis by dynamic bone histomorphometry. Femurs were aseptically removed and analyzed for collagen, collagenase and osteopontin mRNAs by Northern and dot blot analysis. As expected, OVX decreased trabecular bone volume (BV/TV by 73.8% vs. sham p<.01), and also reduced trabecular thickness, numbers, and increased spacing. Bone loss in the OVX animals was partially prevented with either 2 mg/day CMT-8 or 1 mg/wk Clodronate (p<.01), while the 1 mg/day CMT-8 had no effect. Interestingly, the efficacy of the combination therapy of CMT-8 and Clodronate was significantly better than either treatment by itself, maintaining bone mass and structural indices at levels identical to sham values. OVX rats mRNA for collagen, collagenase and osteopontin were elevated indicating high-turnover bone loss. Only COMBO therapy significantly reduced the collagenase and osteopontin mRNA. In summary, CMT-8 mono-therapy (2 mg) alone partially inhibited bone loss in this animal model of osteoporosis. However, 1 mg/day (CMT-8) monotherapy had no effect on bone loss or bone mRNA levels and when combined with Clodronate, interacted to increase efficacy. Thus, a combination of a suboptimal dose of CMT-8 and a bisphosphonate appears to increase the amount of bone by suppressing resorption in a model of osteoporosis.     

Effect of Dose,Sex and Age on the Drug Disposition of Incadronate,a New Bisphosphonate,in Rat Bone

Abstract: We investigated the effects of dose, sex and age on the uptake and elimination of incadronate, a new biosphos-phonate, in rat bone after intravenous administration. Following administration of 0.03-3 mg/kg to young male rats (age 7 weeks), intact drug concentration in humerus at 24 hr after administration (C_{24 hr}) increased in a dose-dependent manner, indicating the linear uptake of the drug into bone. The estimated bone uptake clearance of 0.13 ml/min./g bone is comparable to the estimated plasma flow rate in bone, suggesting that uptake into bone is plasma flow-limited. Concentration from 24 hr after administration had declined bi-exponentially. Although t_1/2β (350-444 days) was little altered among doses, t_1/2α was prolonged with increasing dose from 13.4 to 16.2 days. This effect seemed to be due to inhibition of bone resorption at higher doses resulting in the suppression of drug release from bone. No sex difference was seen on C_{24 hr} in young rats, while the value in senescent (age 12 months) rats was 24% greater in females than in males. When comparing between ages it is seen that C_{24 hr} values in senescent rats decreased to 50-66% of those in young rats. As for elimination from bone, t_1/2β values in senescent rats were shortened to 76-79% of those in young rats. In contrast, little difference in t_1/2α was observed between ages or in either t_1/2α and t_1/2β between sexes.     

JTT-305, an orally active calcium-sensing receptor antagonist, stimulates transient parathyroid hormone release and bone formation in ovariectomized rats

Intermittent administration of parathyroid hormone (PTH) has a potent anabolic effect on bone in humans and animals. Calcium-sensing receptor (CaSR) antagonists stimulate endogenous PTH secretion through CaSR on the surface of parathyroid cells and thereby may be anabolic agents for osteoporosis. JTT-305 is a potent oral short-acting CaSR antagonist and transiently stimulates endogenous PTH secretion. The objective of the present study was to investigate the effects of JTT-305 on PTH secretion and bone in ovariectomized rats. Female rats, immediately after ovariectomy (OVX), were orally administered vehicle or JTT-305 (0.3, 1, or 3 mg/kg) for 12 weeks. The serum PTH concentrations were transiently elevated with increasing doses of JTT-305. In the proximal tibia, JTT-305 prevented OVX-induced decreases in both the cancellous and total bone mineral density (BMD) except for the 0.3mg/kg dose. At the 3mg/kg dose, JTT-305 increased the mineralizing surface and bone formation rate in histomorphometry. The efficacy of JTT-305 at the 3mg/kg dose on the BMD corresponded to that of exogenous rat PTH1-84 injection at doses between 3 and 10 μg/kg. In conclusion, JTT-305 stimulated endogenous transient PTH secretion and bone formation, and consequently prevented bone loss in OVX rats. These results suggest that JTT-305 is orally active and has the potential to be an anabolic agent for the treatment of osteoporosis.     

In vivo effects of high-dose methotrexate on bone remodeling in rats

Methotrexatae (MTX) is a folate antagonist. MTX osteopathy is well recognized to accompany a high-dose therapy with this drug for the treatment of childhood malignancy. Clinical tests also show that low-dose MTX used in the treatment of rheumatoid arthritis may impair bone formation in a population already predisposed to osteoporosis. However, results of clinical tests are hard to interpret, as it is necessary to take into account malignancy-induced changes in the osseous tissue, long-term immobility and concurrent administration of glucocorticosteroids. We conducted in vivo tests to evaluate the effects of oral and intramuscular administration of high dose of MTX on bone remodeling processes in rats. Effects of MTX on the processes of bone remodeling were evaluated by assessing macrometric and histomorphometric parameters as well as mechanical properties of the femur. The tests were carried out on male Wistar rats. Animals were divided into four groups, composed of 7 animals each: Control group (0.9% NaCl solution), MTX-1 po group (MTX at the dose of 1 mg/kg po daily for 10 days: every day for the first five days, and after an 18-day interval, every day for five days), MTX-1 im group (MTX at the dose of 1 mg/kg im daily for 10 days: every day for the first five days, and after an 18-day interval, every day for five days), MTX-5 im group (MTX at the dose 5 mg/kg im daily for 2 days a week for the period of four weeks). Changes in bone remodeling were examined 4 weeks after the first MTX administration. These results show that MTX administered intramuscularly at high doses inhibited the formation and mineralization of new osseous matrix and impaired mechanical properties of the femoral bone, whereas its oral administration had no effect on bone remodeling in rats.     

Oral administration of quercetin inhibits bone loss in rat model of diabetic osteopenia

Diabetic osteopenia can result in an increased incidence of bone fracture and a delay in fracture healing. Quercetin, one of the most widely distributed flavonoids in plants, possesses antioxidant property and beneficial effect on osteoporosis in ovariectomized mice. All these properties make quercetin a potential candidate for controlling the development of diabetic osteopenia. Therefore, the present study was designed to investigate the putative beneficial effect of quercetin on diabetic osteopenia in rats. Diabetes mellitus was induced by streptozotocin. The diabetic rats received daily oral administration of quercetin (5 mg/kg, 30 mg/kg and 50 mg/kg) for 8 weeks, which was started at 4 weeks after streptozotocin injection. Quercetin at 5 mg/kg showed little effect on diabetic osteopenia, while quercetin at 30 mg/kg and 50 mg/kg could increase the decreased serum osteocalcin, serum alkaline phosphatase activity, and urinary deoxypyridinoline in diabetic rats. In addition, quercetin (30 mg/kg and 50 mg/kg) could partially reverse the decreased biomechanical quality and the impaired micro-architecture of the femurs in diabetic rats. Histomorphometric analysis showed that both decreased bone formation and resorption were observed in diabetic rats, which was partially restored by quercetin (30 mg/kg and 50 mg/kg). Further investigations showed that quercetin significantly lowered the oxidative DNA damage level, up-regulated the total serum antioxidant capability and the activity of serum antioxidants in diabetic rats. All those findings indicate the beneficial effect of quercetin on diabetic osteopenia in rats, and raise the possibility of developing quercetin as potential drugs or an ingredient in diet for controlling diabetic osteopenia.     

女贞子治疗去卵巢大鼠骨质疏松的实验研究

目的观察女贞子对去卵巢(ovariectomy,OVX)大鼠骨质疏松(osteoporosis,OP)的治疗作用,并探讨其作用机制。方法大鼠骨质疏松模型,女贞子(9、4.5、2.25 g.kg-1.d-1)灌胃给药,实验过程称体重,连续给药26周后测定血钙(S-Ca)、血磷(S-P)、尿钙(U-Ca)、尿磷(U-P)的含量,ELISA试剂盒双抗体夹心法测定血清碱性磷酸酶(ALP)、血清骨特异性碱性磷酸酶(BALP)含量、骨钙素(OCN)含量、尿液中脱氧吡啶啉(DPD)含量,DEXA型骨密度仪分析大鼠的股骨、胫骨、第4椎骨的BMD(bone mineral density),动物处死后完整取下心、肝、脾、肺、肾、胸腺、脑、子宫进行称重,采用HE染色法进行子宫病理学检查。结果女贞子能有效抑制去卵巢所致的大鼠体重增加、升高S-Ca、S-P、降低U-Ca、U-P、降低ALP、BALP、OCN、DPD的含量,增加大鼠股骨、胫骨、第4椎骨的BMD,并且长期用药对子宫无明显刺激作用。结论女贞子对OVX大鼠所致的骨质疏松症有良好的治疗效果,其作用机制可能与其能降低高的骨转换率有关。     

Development of polyethylene glycol-conjugated alendronate, a novel nitrogen-containing bisphosphonate derivative: evaluation of absorption, safety, and effects after intrapulmonary administration in rats

Bisphosphonates are widely used for the treatment of bone diseases, including hypercalcemia and osteoporosis. However, the bioavailability (BA) of orally administered bisphosphonates is low, at approximately 0.9%-1.8%. In addition, the oral administration of bisphosphonates is associated with mucosal damage, including gastritis, gastric ulcer, and erosive esophagitis. Here, to develop a new delivery system for bisphosphonates that improve their BA and safety, we developed polyethylene glycol (PEG)-conjugated alendronate, a novel nitrogen-containing bisphosphonate derivative. We evaluated the absorption and safety of PEG-alendronate in rats following intrapulmonary administration. The BA of PEG-alendronate after intrapulmonary administration was approximately 44 ± 10% in rats, which was similar to that of alendronate (54 ± 3.9%). Alendronate significantly increased total protein concentration and lactate dehydrogenase activity in bronchoalveolar lavage fluid, suggesting that pulmonary epithelium was locally damaged by intrapulmonary administration of alendronate. In marked contrast, PEG-alendronate did not significantly increase the markers following intrapulmonary administration. In an osteoporosis model in rats, intrapulmonary administration of PEG-alendronate effectively inhibited decreases in the width of the growth plate to a level similar to that achieved by intrapulmonary administration of alendronate. These results indicate that pulmonary delivery of PEG-alendronate is a promising approach for the treatment of bone diseases.     

Effects of the new antiallergic drug 11-oxo-11H-pyrido[2,1-b] quinazoline-2-carboxylic acid on immediate allergic reactions

The effects of 11-oxo-11H-pyrido[2,1-b]quinazoline-2-carboxylic acid (Sm 857) on immediate type, particularly type I allergic reactions were investigated and the following results were obtained. 48-h homologous passive cutaneous anaphylaxis (PCA) in rats was inhibited by oral administration of Sm 857 from the 4th h to the 30th min before antigen challenge and by intravenous administration from the 2nd h to just before antigen challenge. 48-h homologous PCA in rats was inhibited dose-dependently by oral administration of Sm 857 at 1-50 mg/kg 30 min before antigen challenge. On the other hand, the intravenous administration 10 min before antigen challenge showed similar dose-dependent effects on PCA and the effects were significant at doses of 0.5-2 mg/kg. The oral administration of Sm 857 for 1-4 weeks reduced the PCA inhibitory effect in comparison with single administration. Sm 857 injected intravenously twice at intervals of 15-120 min showed no change in inhibitory effect. The PCA inhibitory effect of Sm 857 in adrenalectomized rats was reduced markedly when compared with that in sham-operated rats. On the other hand, adrenalectomy had no influence on the inhibitory effect of tranilast. Sm 857, tranilast and ketotifen inhibited histamin-induced capillary permeability in rats and, in particular, the effect of ketotifen was conspicuous. Sm 857, tranilast and ketotifen significantly inhibited antigen-induced histamine release from the peritoneal cavity of passively sensitized rats. Release of slow reacting substance of anaphylaxis (SRS-A), on the other hand, was inhibited significantly by Sm 857 and ketotifen. Sm 857 showed a tendency to inhibit antigen-induced histamine release from the lung tissue of passively sensitized guinea pigs and significantly inhibited SRS-A release. Sm 857 inhibited histamine- and leukotriene D4 (LTD4)-induced contraction in the lung parenchyma and the tracheal muscle. However, the drug had no effect on contraction of the ileum. Sm 857 showed a tendency to inhibit the Schultz-Dale reaction in the lung parenchyma ana the ileum. Both oral and intravenous administration of Sm 857 showed an inhibitory effect on experimental asthma in guinea pigs, which was similar to that of tranilast.     

Calcitonin inhibits the increase in bone acid phosphatase activity by high dose of zinc in rats

The activity of femoral-epiphyseal acid phosphatase, a marker enzyme of bone resorption induced by parathyroid hormone, was increased by a single oral dose of Zn 100 mg/kg to rats. This increase was completely inhibited by calcitonin which has antagonistic effect on the action of parathyroid hormone on bone, thus supporting the hypothesis that the effect of Zn administration on bone may be mediated by parathyroid hormone.     

Effects of dopamine d2 receptor agonists in a pituitary transplantation-induced hyperprolactinaemia/anovulation model in rats

1. In the present study, we investigated the effects of hyperprolactinaemia, induced by transplantation of anterior pituitary glands under the kidney capsule in female rats, on the relationship between serum and pituitary concentrations of the gonadotropins and on the oestrous cycle. 2. Rats with pituitary transplants showed increased serum prolactin concentrations and decreased serum concentrations of gonadotropins and increased pituitary concentrations of gonadotropins. Moreover, these rats showed persistent dioestrous and anovulation from 3 to 6 days after transplantation. 3. A single oral administration of cabergoline (at doses between 0.001 and 0.1 mg/kg) dose-dependently inhibited the elevated serum prolactin concentrations in hyperprolactinaemic rats. At 0.1 mg/kg, cabergoline induced a continuous reduction in serum prolactin concentrations for 5 days after administration. Terguride (0.1 mg/kg) and bromocriptine (10 mg/kg) also reduced serum prolactin concentrations at 1 and 3 days after administration. All three dopamine D2 receptor agonists increased serum gonadotropin concentrations and ovarian weight at 3 days after administration. 4. In rats exhibiting anovulation, a single oral administration of any one of the three dopamine D2 receptor agonists dose-dependently restored ovulation and a normal oestrous cycle appeared. Oral administration of cabergoline (0.03 mg/kg) or terguride (0.1 mg/kg) restored ovarian function and abolished the anovulation following a reduction in serum prolactin concentrations. However, bromocriptine (10 mg/kg) did not completely abolish anovulation. Following administration of terguride (0.3 mg/kg) or bromocriptine (30 mg/kg), only one normal oestrous cycle appeared; however, following cabergoline (0.1 mg/kg), two normal oestrous cycles appeared. 5. These results suggest that cabergoline has a potent and long-lasting action as a dopamine D2 receptor agonist and, thus, should be a useful drug for the treatment of galactorrhoea and hyperprolactinaemic amenorrhoea and/or anovulation in humans.     

Phenobarbital inhibits osteoclast differentiation and function through NF-κB and MAPKs signaling pathway

The purpose of this study was to determine the direct effects of phenobarbital (PB) on receptor activator of nuclear factor kappa-B ligand (RANKL) induced osteoclast differentiation and function in vitro and in vivo. Here, PB significantly inhibited osteoclast formation and bone resorption ability induced by RANKL in vitro. Meanwhile, intracellular signaling transduction analysis revealed PB specifically decreasing the phosphorylation level of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and mitogen-activated protein kinase (MAPK), respectively. Besides, oral administration of PB at the dose of 60 mg/kg/day for 6 weeks led to improve the bone loss and to decrease the activity on both osteoblast and osteoclast. This suppression effect is more obvious in osteoblast-induced bone formation than that on osteoclast-induced bone resorption. Taken together, our findings demonstrated that PB down-regulate osteoclast differentiation and activity through modulation of NF-κB and MAPKs signaling pathway. The direct suppression effect on osteoclast can induce bone loss after long term oral administration. This bone loss is due to reducing bone turnover rate on both sides of bone formation and bone resorption.     

Comparison of the effects of D-003, a mixture of high-molecular-weight aliphatic acids from sugarcane wax, and pravastatin on bones and osteoclast apoptosis of ovariectomized rats

The mevalonate pathway is relevant in bone cells. Statins inhibit 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the key enzyme of this route, stimulating osteoblast differentiation and activity. Pravastatin increases bone formation markers in postmenopausal women and bone density in diabetics. D-003 is a mixture of high-molecular-weight acids purified from sugarcane wax which inhibits cholesterol biosynthesis through HMG-CoA reductase regulation, preventing bone loss in osteoporosis induced with ovariectomy and prednisolone in rats. We investigated the effects of D-003 (50 mg/kg) and pravastatin (20 mg/kg) orally administered for 12 weeks to ovariectomized rats. Female rats were randomized in four groups (10 rats/group): two control groups treated with the vehicle, one false-operated (sham) and another ovariectomized (positive control), while two other groups received D-003 or pravastatin. Bone resorption and formation was studied through histomorphometry and apoptosis through immunohistochemistry. D-003 and pravastatin significantly (p < 0.001) prevented the changes of trabecular bone versus ovariectomized rats and (p < 0.001) the increase of the surface and number of osteoclasts versus ovariectomized controls. D-003 and pravastatin, however, did not modify osteoblast surfaces, a bone formation marker D-003 and pravastatin increased osteoclast apoptosis and reduced (p < 0.05) osteoblast and osteocyte apoptosis versus ovariectomized controls; D-003 was more effective (p < 0.05) than pravastatin. In conclusion, D-003 (50 mg/kg) orally administered for 12 weeks prevented bone loss and bone resorption in ovariectomized rats, increasing osteoclast apoptosis. The preventive effects of D-003 on bone loss and resorption in ovariectomized rats are comparable to those of pravastatin. Both drugs inhibited osteoblast apoptosis but failed to change osteoblast surface. The effects of D-003 on bone cell apoptosis were greater than those of pravastatin. Therefore, D-003 could be used to prevent or treat bone loss in postmenopausal women, but further animal studies and clinical trials are required to confirm the clinical relevance of this potential effect.     

废用性骨质疏松动物模型发生骨质疏松时间段的研究

目的建立废用性骨质疏松动物模型,探讨发生废用性骨质疏松的最佳时间段。方法选取3个月龄SD大鼠78只,其中的72只大鼠随机均分为假手术组(S组)和模型组(M组),每组根据术后检测时间(1、2、4、6、8、12周)的时间段分为6个亚组,每组6只。另6只为空白对照组(BC组)。检测血清骨钙蛋白(BGP)、碱性磷酸酶(ALP)、抗酒石酸酸性磷酸酶(Strap)及尿羟脯氨酸与尿肌酐比值(HOP/Cr);进行左胫骨近侧干骺段HE染色和Masson染色,观察骨组织形态学。结果随着去神经时间的延长,M组BGP和尿HOP/Cr逐渐降低,而ALP和Strap逐渐升高,4、6、8、12周的水平与1、2周差异明显(P<0.05),而术后4、6、8、12周之间的差异无统计学意义(P>0.05)。M组骨皮质变薄、孔隙增多、骨小梁稀疏、纤细并有多处断裂,小梁间吻合减少,骺板下区空旷,髓腔扩大,破骨细胞数增多。Masson染色见随着废用时间的延长,新生胶原的量也在减少。结论坐骨神经切断术是建立废用性骨质疏松动物模型的有效方法。在废用性骨质疏松形成过程中,骨形成显著持续降低,但速度趋缓,观察的最佳时间为造模后4-6周。     

Oral delivery of biologically active parathyroid hormone

Purpose. Parathyroid hormone (PTH), the only drug known to stimulate bone formation, is a peptide therapeutic indicated in the treatment of osteoporosis. Unfortunately, PTH is only effective when dosed by injection because it has no oral bioavailability. Herein we report the oral absorption of PTH in rats and monkeys facilitated by the novel delivery agent, N-[8-(2-hydroxy-4-methoxy)bensoyl]amino caprylic acid (4-MOAC). Methods. 4-MOAC was selected from a group of 100 delivery agents based on in vitro chromotography studies and in vivo screening studies in rats. The PTH/4-MOAC combination was then tested in monkeys. The interaction of 4-MOAC and PTH was evaluated by nuclear magnetic resonance (NMR) spectroscopy. Results. Monkeys were administered an aqueous solution containing 4-MOAC and PTH and mean peak serum PTH concentrations of about 3000 pg/mL were obtained. The relative bioavailability of oral PTH was 2.1% relative to subcutaneous administration. The biological activity of the orally-delivered PTH was further evaluated in a rat model of osteoporosis. These studies showed that the bone formed following oral PTH/4-MOAC administration was comparable to that formed following PTH injections. The 4-MOAC mediated absorption of PTH is hypothesized to be the result of a noncovalent interaction between 4-MOAC and PTH. The preliminary evaluation of this interaction by NMR is described. Conclusions. 4-MOAC facilitates the absorption of PTH following oral administration to both rats and monkeys. The orally-absorbed PTH is biologically active as demonstrated in a rat model of osteoporosis.