Alfacalcidol increases cancellous bone in low turnover, fatty marrow sites in aged, orchidectomized rats.

The objectives of this study were to determine the responses of cancellous bone in the distal tibial metaphysis (DTM), a low turnover, fatty (yellow) marrow site, to sham-aged, orchidectomy (ORX) and alfacalcidol treatment in sham-aged and ORX rats. Eighteen-month-old male sham and ORX rats were treated with 0.1 and 0.2 microg/kg alfacalcidol 5 days/wk p.o. for 12 weeks, double fluorescent labeled, and the DTM were processed for bone histomorphometry analyses. The current study found the DTM in sham-aged male rats were resistant to age-related and ORX-induced cancellous bone loss and alfacalcidol-induced bone gain, findings that differ from that in the proximal tibial metaphysis (PTM) and lumbar vertebral body (LVB), two high turnover, red marrow bone sites. However, alfacalcidol treatment increased DTM bone mass in ORX rats where bone turnover was elevated by androgen deficiency. These results in concert with the previously positive findings in red marrow bone sites following alfacalcidol treatment suggest that alfacalcidol is more effective in increasing cancellous bone mass in the skeletal sites with higher bone turnover.

Greater efficacy of alfacalcidol in the red than in the yellow marrow skeletal sites in adult female rats.

The present study compared the bone anabolic effects of graded doses of alfacalcidol in proximal femurs (hematopoietic, red marrow skeletal site) and distal tibiae (fatty, yellow marrow skeletal site). One group of 8.5-month-old female Sprague-Dawley rats were killed at baseline and 4 groups were treated 5 days on/2 days off/week for 12 weeks with 0, 0.025, 0.05 and 0.1 microg alfacalcidol/kg by oral gavage. The proximal femur, bone site with hematopoietic marrow, as well as the distal tibia bone site with fatty marrow, were processed undecalcified for quantitative bone histomorphometry. In the red marrow site of the proximal femoral metaphysis (PFM), 0.1 microg alfacalcidol/kg induced increased cancellous bone mass, improved architecture (decreased trabecular separation, increased connectivity), and stimulated local bone formation of bone 'boutons' (localized bone formation) on trabecular surfaces. There was an imbalance in bone resorption and formation, in which the magnitude of depressed bone resorption greater than depressed bone formation resulted in a positive bone balance. In addition, bone 'bouton' formation contributed to an increase in bone mass. In contrast, the yellow marrow site of the distal tibial metaphysis (DTM), the 0.1 microg alfacalcidol/kg dose induced a non-significant increased cancellous bone mass. The treatment decreased bone resorption equal to the magnitude of decreased bone formation. No bone 'bouton' formation was observed. These findings indicate that the highest dose of 0.1 microg alfacalcidol/kg for 12 weeks increased bone mass (anabolic effect) at the skeletal site with hematopoietic marrow of the proximal femoral metaphysis, but the increased bone mass was greatly attenuated at the fatty marrow site of the distal tibial metaphysis. In addition, the magnitude of the bone gain induced by alfacalcidol treatment in red marrow cancellous bone sites of the proximal femoral metaphysis was half that of the lumbar vertebral body. The latter data were from a previous report from the same animal and protocol. These findings indicated that alfacalcidol as an osteoporosis therapy is less efficacious as a positive bone balance agent that increased trabecular bone mass in a non-vertebral skeletal site where bone marrow is less hematopoietic.

Continuous PGE2 leads to net bone loss while intermittent PGE2 leads to net bone gain in lumbar vertebral bodies of adult female rats.

The present study examined the effects of continuous and intermittent PGE2 administration on the cancellous and cortical bone of lumbar vertebral bodies (LVB) in female rats. Six-month-old Sprague-Dawley female rats were divided into 6 groups with 2 control groups and 1 or 3 mg PGE2/kg given either continuously or intermittently for 21 days. Histomorphometric analyses were performed on the cancellous and cortical bone of the fourth and fifth LVBs. Continuous PGE2 exposure led to bone catabolism while intermittent administration led to bone anabolism. Both routes of administration stimulated bone remodeling, but the continuous PGE2 stimulated more than the intermittent route to expose more basic multicellular units (BMUs) to the negative bone balance. The continuous PGE2 caused cancellous bone loss by stimulating bone resorption greater than formation (i.e., negative bone balance) and shortening the formation period. It caused more cortical bone loss than gain, the magnitude of the negative endocortical bone balance and increased intracortical porosity bone loss was greater than for periosteal bone gain. The anabolic effects of intermittent PGE2 resulted from cancellous bone gain by positive bone balance from stimulated bone formation and shortened resorption period; while cortical bone gain occurred from endocortical bone gain exceeding the decrease in periosteal bone and increased intracortical bone loss. Lastly, a scheme to take advantage of the marked PGE2 stimulation of lumbar periosteal apposition in strengthening bone by converting it to an anabolic agent was proposed.

Continuous infusion of PGE2 is catabolic with a negative bone balance on both cancellous and cortical bone in rats.

It is well documented that intermittent PGE(2) treatment increases both trabecular and cortical bone mass. However, the effects of continuous PGE(2) administration remain undocumented. The aim of the study was to investigate the effects of continuous prostaglandin E(2) (PGE(2)) on different bone sites in skeletally mature rats. Six-month-old Sprague Dawley rats were treated with PGE(2) at 1 or 3 mg/kg/d continuously via infusion pump for 21 days. Two other groups of rats received PGE(2) at the same doses by intermittent (daily) subcutaneous injections and served as positive controls. Histomorphometry was performed on cancellous bone of the proximal tibial metaphysis and cortical bone of the tibial shaft. As expected, intermittent PGE(2) treatment increased both cancellous and cortical bone mass by stimulating bone formation at the cancellous, periosteal and endocortical bone surfaces. In contrast, continuous PGE(2) treatment decreased cancellous bone mass with bone resorption exceeding bone formation. In addition, continuous PGE(2) treatment increased endocortical and intracortical bone remodeling, inducing bone loss which was partially offset by stimulating periosteal expansion. We conclude that continuous PGE(2) treatment induces overall catabolic effects on both cancellous and cortical bone envelopes, which differs from intermittent PGE(2) treatment that is anabolic. Lastly, we speculate that superior bone mass may be achieved by co-treatment of continuous PGE(2) in combination with an anti-catabolic agent.

Cancellous bone minimodeling-based formation: a Frost, Takahashi legacy.

The current dogma is that in adult human beings, remodeling creates nearly all the new cancellous bone tissue. However, Frost, Takahashi and colleagues hypothesized that minimodeling can go on in trabeculae throughout life. The current perspective will review the available reports on minimodeling-based formation to determine whether there is any support for his hypothesis. One: describe the methodology employed to characterize remodeling and minimodeling formation sites or packets, which restrict the analyses of these packets to a known age of the specimen. Two: present quantitative minimodeling data on cancellous bone of aging rats and transiliac bone biopsy of adult humans. Three: describe the occurrence and quantitation of mixed remodeling-minimodeling formation sites that could be misinterpreted as minimodeling sites. Fourth: present irrefutable evidence that bone anabolic agents initiate minimodeling-based formation sites. Fifth: discuss the mechanism of minimodeling-based formation may be the resumption of osteoblastic activity by bone lining cells to increase cancellous bone mass and trabecular connectivity. The findings of minimodeling is a rare activity in normal individuals, but may occur in a select population, and bone anabolic agents can initiate minimodeling-based formation are in support of Frost's hypothesis that minimodeling can continue throughout human life. Thus, another Frost, Takahashi legacy lives on.

Rolipram, a phosphodiesterase 4 inhibitor, prevented cancellous and cortical bone loss by inhibiting endosteal bone resorption and maintaining the elevated periosteal bone formation in adult ovariectomized rats.

Cyclic AMP (cAMP) is a continually produced nucleotide inactivated by hydrolysis to 5'AMP via phosphodiesterase (PDE) enzymes. Rolipram is a selective PDE4 inhibitor reported to have anti-inflammatory effects and used in the treatment of asthma and chronic obstructive pulmonary disease (COPD). The current study was designed to determine whether Rolipram could prevent and restore bone loss in ovariectomized (OVX) rats. Six-month-old Sprague Dawley rats underwent either sham-operated or bilateral ovariectomy, and were left untreated for 60 days to develop osteopenia. Then they were treated with vehicle, 6 mg/kg PGE(2), 3 microg/kg Alendronate or 0.1-1.0 mg/kg Rolipram for 60 days. At sacrifice, the right tibiae were processed for quantitative bone histomorphometric measurements. The right femurs were measured by dual energy A-ray absorptiometry and the 5th lumbar vertebrae were subjected to micro-computed tomography to access bone mass and architecture changes. Our results indicated that OVX induced negative bone balance in all five bone sites we tested, with bone resorption exceeding bone formation. Rolipram at 0.1-0.6 mg/kg dose levels prevented while at 1 mg/kg restored ovariectomy-induced cancellous and cortical bone loss in the tibia, femur and lumbar vertebra. Dynamic bone histomorphometry suggested that these beneficial effects were achieved by partially maintaining the elevated bone formation at the trabecular bone surface and increasing bone formation at the periosteal bone surface of the cortex. Furthermore, it reduced bone turnover at the trabecular and the endocortical bone surfaces. The prevention of further bone loss effects were comparable to those of an anti-resorption agent (Alendronate) but were not as great as those of an anabolic agent (PGE(2)). In addition, Rolipram treatment increased body and muscle weights compared to the vehicle-treated OVX rats. In conclusion, our study in an osteopenic rat model suggested that a selective PDE4 inhibitor may be used for the treatment of established osteoporosis.

Simvastatin did not prevent nor restore ovariectomy-induced bone loss in adult rats.

Current published results on whether statins have beneficial effects on bone metabolism have been conflicting so far. In order to further investigate if statins were promising candidates for the treatment for osteoporosis, we conducted a study in which rats were ovariectomized (OVX) at 6 months of age, allowed to lose bone for 60 days and followed by oral administration of simvastatin at the dose levels of 0.3-10 mg/kg/d for 60 days. PGE2 (6 mg/kg) was used as a positive control. Study endpoints included bone histomorphometry on the proximal tibial metaphysis (PTM) and the tibial diaphysis (TX), dual-energy X-ray absorptiometry on the right femur and micro computed tomography (ICT) on the 5th lumbar vertebra (LV). After 120 days of OVX, cancellous bone lost by 80% in the PTM and 18% in the LV accompanied by increased bone formation and resorption. Simvastatin at all dose levels did not affect bone volume, bone formation rate and bone erosion surface when compared to 120 day ovariectomized animals at all bone sites studied. By contrast, PGE2 restored cancellous and cortical bone area to sham control levels. In conclusion, this study demonstrated that unlike PGE2, oral administration of simvastatin did not have effects on cancellous or cortical bone formation and resorption; and consequently was not able to prevent further bone loss or restore bone mass in the osteopenic, OVX rats.

Estrogen and "exercise" have a synergistic effect in preventing bone loss in the lumbar vertebra and femoral neck of the ovariectomized rat.

This study was designed to study the individual or combined effects of estrogen and bipedal stance "exercise" on the lumbar vertebral body (LVB) and femoral neck (FN). At 6 months of age, six rats were sacrificed as baseline controls and all the others were either bilateral sham-ovariectomized or ovariectomized (OVX). Groups of OVX rats were housed in normal height cage (NC, 28 cm) or raised height cages (RC, 33 cm) and received biweekly s.c. injections of 10 microg/kg 17 beta estradiol (E2) or vehicle for 4 and 8 weeks. Histomorphometric measurements were performed on the undecalcified mid-transverse sections of the 4th LVB and FN. Ovariectomy alone induced cancellous bone loss by 21% and 39% in the LVB and FN, respectively; intracortical porosity area of the FN increased by 108% while total bone area did not change significantly because of the periosteal expansion following OVX. E2 alone partially prevented cancellous bone loss in the LVB and FN and prevented increased intracortical porosity area in the FN by reducing eroded surface and activation frequency. RC alone partially prevented the decrease of cancellous bone in the LVB and FN by reducing the bone-eroded surface but increased wall width. E2 plus RC completely preserved cancellous bone by having an additive effect on reducing eroded surface and activation frequency. RC helped to partially prevent decreased periosteal bone formation after estrogen administration. In conclusion, apart from inducing cancellous bone loss in the LVB and FN, OVX also increased intracortical remodeling in the FN. Estrogen prevented the overall activation of remodeling space induced by OVX. Apart from having similar effects as estrogen on remodeling space, RC induced positive bone balance within each remodeling unit. Combination treatment increased total bone mass beyond that of sham-control level by having an additive effect on lowering bone remodeling and increasing wall in both the LVB and FN.

Bipedal stance exercise and prostaglandin E2 (PGE2) and its synergistic effect in increasing bone mass and in lowering the PGE2 dose required to prevent ovariectomized-induced cancellous bone loss in aged rats.

Previous reports have shown that bone loss was partially prevented by bipedal stance "exercise" following ovariectomy (ovx), and it was well documented that prostaglandin E2 (PGE(2)) had an anabolic effect on the rat skeleton. The aim of this study was to determine whether lower doses of PGE(2) could prevent ovx-induced cancellous bone loss with the combination of bipedal stance exercise. Seventy-eight 10-month-old female Sprague-Dawley rats were either ovariectomized or sham-operated on day 0 and then treated with PGE(2) (0, 0.3, or 1 mg/kg per day) and/or housed in normal height cages (NC, 28 cm) or raised cages (RC, 33 cm) for 8 weeks. Bone histomorphometry was performed on the double-fluorescent-labeled proximal tibial metaphysis. In sham rats, 1 mg/kg PGE(2) + RC had synergistic effects in increasing trabecular bone area, width, and number by stimulating mineral apposition rate and bone formation rate. As expected, ovx induced cancellous bone loss, accompanied by elevated activation frequency. Without RC, PGE(2) monotherapy prevented ovx-induced bone loss at the 1 mg/kg per day dose, whereas this prevention effect was observed at the 0.3 mg/kg per day dose when combined with RC. Similar to their effects in sham rats, PGE(2) and RC had synergistic effects in augmenting cancellous bone mass and architecture and maintaining the elevated bone formation but depressing bone resorption and activation frequency. We conclude that bipedal stance exercise lowers the PGE(2) dose required to prevent ovx-induced cancellous bone loss in the proximal tibial metaphysis in aged rats.

Bipedal stance exercise enhances antiresorption effects of estrogen and counteracts its inhibitory effect on bone formation in sham and ovariectomized rats.

In this study we employed a raised cage model in combination with estrogen to observe their effects on the proximal tibial metaphysis (PTM) and tibial shaft (TX) in sham-operated or ovariectomized rats. A total of 105 6-month-old female Sprague-Dawley rats were used in the study. Bilateral sham ovariectomy or ovariectomy was performed at day 0 and the rats were housed in normal height or raised cages (RCs) and injected subcutaneously twice per week with 10 microg/kg of 17beta-estradiol (E2) or vehicle for 4 and 8 weeks. Because the time course of bone loss or bone gain distribution was not uniform in the metaphyses of the tibia, we subdivided the PTM into three zones (medial, central, and lateral) to observe the different bone loss or bone gain patterns after ovariectomy and/or raised cages. We found that: (1) E2 alone did not alter bone area or architecture in sham rats, whereas RC alone increased trabecular thickness and area of PTM, but had no effects on TX; (2) Ovx induced most bone loss from the central zone of the PTM and endocortical surface of TX, accompanied by decreased trabecular number and increased bone resorption; (3) E2 alone prevented ovx-induced bone loss by preserving trabecular number and depressing bone resorption; (4) RC alone partially compensated for bone loss following ovx by thickening the surviving trabeculae in lateral and medial zones, and tended to stimulate bone formation and decrease bone resorption; and (5) RC plus E2 increased trabecular bone area by having an additive effect on bone resorption and bone turnover. RCs helped to prevent the depressive effect of estrogen on periosteal bone formation. In conclusion, early and rapid bone loss occurred in the central zone of the metaphysis and endocortical surface after ovx. Estrogen replacement therapy prevented this loss. Raised cages partially compensated for bone loss following ovx by thickening the trabeculae in the lateral area of the metaphysis and decreased endocortical erosion. Combination treatment added bone to the PTM and prevented the decrease of periosteal bone formation after estrogen administration.

Lumbar vertebral cancellous bone is capable of responding to PGE2 treatment by stimulating both modeling and remodeling-dependent bone gain in aged male rats.

Previously we found that PGE2 3 mg/kg in 20-month-old male rats induced massive bone formation mainly by modeling dependent bone gain in cortical bone. It is not known whether cancellous bone will respond similarly; thus, we evaluated the effect of PGE2 on cancellous bone of the same aged rats. Thirty-four 20-month-old Wistar male rats were given PGE2 (3 mg/kg/day) or vehicle subcutaneously for 10 and 30 days. Double fluorescent labels were injected 9 and 2 days prior to the sacrifice. Histomorphometry was performed on 1% toluidine blue stained and unstained sagittal sections of lumbar vertebral bodies. The results demonstrated that 10-day PGE2 treatment increased osteoprogenitor cells, osteoblasts (x 2-fold), osteoid (x 4.5-fold), woven bone formation (0.04%), and 40% more trabecular area; it stimulated modeling (x 2-fold) and remodeling-dependent (x 1.5-fold) bone formation with increase of mineralization lag time (MLT, x 7.5-fold). Thirty-day treatment sustained increases in osteoblast numbers, modeling and remodeling-dependent bone formation and further stimulated woven bone formation (6.6%), turnover (x 3-fold), and trabecular area and number (x 2-fold). Osteoprogenitor cells were undetectable along with 70% less osteoid area compared with 10-day treatment but still was 1.5-fold higher than aging controls. MLT returned to aging control level. It was concluded that the aged cancellous bone of 20-month-old male Wistar rat retains a capability of responding to the anabolic effect of PGE2. Osteoblastogenesis and enhanced modeling and remodeling-dependent woven or lamellar formation contribute to this anabolic action. Bone formation differed in that the endocortical surface of cortical bone was stimulated mainly by modeling whereas both modeling and remodeling-dependent bone gain were equally stimulated at the trabecular surface of the lumbar vertebral body.

Cancellous bone of aged rats maintains its capacity to respond vigorously to the anabolic effects of prostaglandin E2 by modeling-dependent bone gain.

The present study examined the early effects of prostaglandin (PG)E2 on proximal tibial metaphyses of 20-month-old Wistar male rats. PGE, was given to intact rats for 10 and 30 days at 3mg/kg/day. After multiple in vivo fluorochrome labeling, undecalcified longitudinal sections were subjected to analysis of bone histomorphometry and classification of the contour of the cement line in bone formation units. The latter was used to classify bone formation units into modeling, remodeling and uncertain units. After 10 days of treatment, there was a 2% increase in woven bone formation with the appearance of osteoprogenitor cells and increases in the number of osteoblasts (649%) and osteoid (375%) surfaces. Remodeling and modeling units increased by 56% and 429%. respectively. After 30 days of treatment, there was an increase of 212% of total trabecular bone mass, 60% of which was woven bone. In addition, there were increases in labeling surface (147%), mineral apposition rate (760%), bone formation rates tissue area (BFR/T.Ar, 1920%; BFR/B.Pm, 343%), and bone turnover (BFR/B.Ar, 426%). Osteoblasts and osteoid production at 30 days were 29% and 58% less than at 10 days post-treatment. Modeling and remodeling activity did not differ from that seen at 10 days. In addition, PGE2 treatment tended to stimulate the closing of growth plates and decrease the fatty marrow area. We conclude that the aged skeleton was able to respond vigorously to PGE2 treatment. Massive osteoprogenitors cells, and osteoid and osteoblast formations were observed within 10 days. and dramatic woven and lamellar bone formation was seen at 30 days post-treatment. The anabolic effects were driven mainly by modeling.

Erect bipedal stance exercise partially prevents orchidectomy-induced bone loss in the lumbar vertebrae of rats.

This study investigates the responses of the fourth and fifth lumbar vertebral bodies of 6-month-old male Sprague-Dawley (SD) rats to orchidectomy (orx) and to erect bipedal stance for feeding for 12 weeks in specially designed raised cages (RC) for which the heights were raised from 20 cm to 35.5 cm. A total of 30 rats were divided into groups of: baseline; sham + housed in normal height cage (NC); orx + NC; sham + RC; and orx + RC. Bone histomorphometry was performed on the triple-labeled undecalcified fourth sagittal (LVL-4) and fifth transverse (LVX-5) sections. We found that orchidectomy induced high-turnover trabecular and cortical bone loss in the lumbar vertebrae. Forcing the rats to rise to erect stance for feeding reduced trabecular and cortical bone loss caused by orx. Apparently, depressing the elevated bone resorption next to the marrow induced by orx, and stimulating bone formation at the ventral periosteal surfaces, caused these effects. Orchidectomy and raised cage had similar effects on the two vertebrae except that the percentage of trabecular bone loss was greater in the LVL-4 than in LVX-5, and that bipedal stance exercise increased the total tissue area and mineral apposition rates (0-80 day interval) of ventral periosteal and dorsal endocortical surfaces of LVX-5 to a greater extent than it did in LVL-4. Such findings suggest that forcing rats to rise to an erect bipedal stance for feeding helps prevent loss of trabecular and cortical bone "mass," and presumably bone strength, in orchidectomized rats. This method also provides an inexpensive, noninvasive, reliable model to increase in vivo vertebral loading in rats that is similar in humans.

Making rats rise to erect bipedal stance for feeding partially prevented orchidectomy-induced bone loss and added bone to intact rats.

The objectives of this study were to investigate the different effects on muscle mass and cancellous (proximal tibial metaphysis [PTM]) and cortical (tibial shaft [TX]) bone mass of sham-operated and orchidectomized (ORX) male rats by making rats rise to erect bipedal stance for feeding. Specially designed raised cages (RC) were used so that the rats had to rise to erect bipedal stance to eat and drink for 12 weeks. Dual-energy X-ray absorptiometry (DEXA) and peripheral quantitative computerized tomography (pQCT) were used to estimate the lean leg mass and bone mineral. Static and dynamic histomorphometry were performed on the triple-labeled undecalcified sections. We found that making the intact rats rise to erect bipedal stance for feeding increased muscle mass, cortical bone volume, and periosteal bone formation. Orchidectomy increased net losses of bone next to the marrow by increasing bone turnover. Making the ORX rats rise to erect bipedal stance increased muscle mass, partially prevented cancellous bone loss in the PTM, and prevented net cortical bone loss in TX induced by ORX by depressing cancellous and endocortical high bone turnover and stimulating periosteal bone formation. The bone-anabolic effects were achieved mainly in the first 4 weeks in the PTM and by 8 weeks in the TX. These findings suggested that making the rats rise to erect bipedal stance for feeding helped to increase muscle mass and cortical bone mass in the tibias of intact rats, increase muscle mass, and partially prevented cancellous and net cortical bone loss in ORX rats.

Anabolic effect of prostaglandin E2 on cortical bone of aged male rats comes mainly from modeling-dependent bone gain.

In this study, prostaglandin E2 (3 mg/kg per day) was administered to 20-month-old male Wistar rats for 10 and 30 days. Histomorphometric analyses were performed on double-fluorescent-labeled undecalcified tibial shaft sections. Thirty days of prostaglandin E2 (PGE2) administration increased bone formation rate/total bone surface from undetectable levels to 0.6 microm/day at the periosteal surface and from 0.5 to 2.1 microm/day at the endocortical surface. Endocortical osteoid surface area increased from 2% to 67% at day 10 and decreased to 6% at day 30; woven and lamellar bone formation started at day 0, but was most obvious at day 30, resulting in a 12% increase of total bone mass. The red to yellow marrow ratio was 0.2 in pretreatment controls, and increased to 1.6 by day 10 and 2.4 by day 30 with PGE2 administration. Intracortical cavity number and area increased after 10 days of PGE2 treatment, but with forming osteon number and area far exceeding those of resorption cavities at day 30. Endocortical modeling surface/endocortical surface was only 1.5%, and remodeling was 11.1% in pretreatment controls. PGE2 treatment increased modeling to 24.5% in the 10 day group and 93.7% in the 30 day group, whereas remodeling remained unchanged at 10 days, and decreased to 6.2% at 30 days. Osteoprogenitor cells and osteoblasts could not be detected in pretreatment controls, but increased by day 10, and returned almost to control levels by 30 days. Our data indicate that PGE2 induced periosteal and endocortical bone formation mainly by modeling-dependent bone gain, accompanied by increases in intracortical remodeling and red bone marrow, and a transient increase in the osteoprogenitor cells adjacent to the endocortical surface. These findings suggest that 20-month-old male Wistar rats were very responsive to the anabolic action of PGE2 in the tibial shaft, a site consisting mainly of cortical bone and yellow marrow.

Effects of prostaglandin E2 and F2 alpha on the skeleton of osteopenic ovariectomized rats.

This article contains the histomorphometric evaluation of the effects of prostaglandin F2 alpha (PGF2 alpha) on cancellous bone from the lumbar vertebra and cortical bone from the tibial shaft of ovariectomized, osteopenic rats. These effects were then compared with those of prostaglandin E2 (PGE2). Three-month-old rats were either ovariectomized (ovx) or sham-ovx. Then, either PGF2 alpha or PGE2 in doses of 1 and 3 mg/kg/day was given subcutaneously for 21 days at 150 days post ovx. Histomorphometric analysis was performed separately on both the primary and secondary spongiosae of the fourth lumbar vertebral bodies (LVB) and on tibial shafts. The ovx rats exhibited osteopenia in both primary (-23% to -37%) and secondary (-20%) spongiosae of the LVB, but not in the tibial shafts at 150 and 171 days post ovx. In the LVB, PGE2 in doses of 1 or 3 mg/kg/day for 21 days restored trabecular bone volume to the levels of sham-ovx controls in the primary spongiosa. However, in the secondary spongiosa, the treatments only thickened the trabeculae. The effects of the PGF2 alpha treatment were similar to those of the PGE2 in both the primary and the secondary spongiosae. While both PGF2 alpha and PGE2 treatments stimulated bone formation in the LVB as indicated by the increases in labeled perimeter, tissue and bone area-based bone formation rates, PGE2 is about 10 times more potent than PGF2 alpha in these effects. The PGE2 treatment also elevated activation frequency in the LVB, while the PGF2 alpha treatment did not. The treatments differed in that PGE2 at these dose levels did not alter the eroded surface in the LVB while PGF2 alpha decreased it significantly. Thus, the increase of the ratio of labeled to eroded perimeter in the LVB in PGF2 alpha-treated animals was much more than that in PGE2-treated animals. In the tibial shafts, PGE2 in doses of 1 and 3 mg/kg/day produced new marrow trabeculae in 2 of 6 and 3 of 6 of the ovx rats. However, no new trabecula was found in PGF2 alpha-treated tibial shafts. Higher doses of PGE2 also increased periosteal labeled perimeter, MAR, and BFR/BS, while PGF2 alpha did not produce any significant change in these parameters. Both PGE2 and PGF2 alpha in doses of 1 and 3 mg/kg/day increased the labeled perimeter, MAR and BFR/BS and decreased the eroded perimeter in the endocortical surface. We concluded that both PGF2 alpha and PGE2 in doses of 1 and 3 mg/kg/day for 21 days exhibited anabolic bone effects. The effects were mostly confined to an increase in trabecular volume in the primary spongiosa of the LVB and in the endocortical surface of tibial shafts. The tissue level mechanism behind this appears to be that PGE2 and PGF2 alpha can both stimulate osteoblast recruitment and activity. Overall, we found PGE2 to be more potent than PGF2 alpha at the same dose level at the endocortical surface. Furthermore, new marrow trabecular bone formed only after PGE2 treatment. PGF2 alpha differed from PGE2 by significantly reducing the trabecular eroded surface in ovx rats.

Early effects of prostaglandin E2 on bone formation and resorption in different bone sites of rats.

The aim of this study was to determine early effects of prostaglandin E2 (PGE2) on bone mass, formation and resorption in a growing cancellous bone site (the proximal tibial metaphysis, PTM), non-growing cancellous bone site (the distal tibial metaphysis, DTM), and cortical bone site (the tibial shaft, TX) with histomorphometric analysis. Six mg PGE2/kg/d was given s.c. to 6-month-old Sprague-Dawley female rats for 5, 10 or 16 days. Double fluorescent labels were given to 0, 10- and 16-day PGE2 treatment and 16-day control groups. Significant increase in bone mass was found after 16 days treatment in cancellous bone sites but not in the cortical bone site. Stimulated bone formation, indicated by the increase in osteoid perimeter, was observed as early as 5 days post-treatment in all 3 bone sites. Bone formation indices were increased after 10 days of treatment, however, there was no difference in selected bone formation indices between 10 and 16 days PGE2 treatments at all 3 bone sites. Significant increase in eroded surface and eroded surface covered with osteoid was observed in cancellous bone sites after 5 days, but decreased after 10 days of treatment. Although the eroded surface was not elevated in TX at the 5th day, the eroded surface covered with osteoid was increased on endocortical surface which indicated that PGE2 stimulated bone resorption on this surface prior to day 5. We concluded that PGE2 stimulated the bone formation and resorption as early as 5 days post-treatment. The levels of stimulated bone formation was TX > DTM > PTM.(ABSTRACT TRUNCATED AT 250 WORDS)

Risedronate plus prostaglandin E2 is superior to prostaglandin E2 alone in maintaining the added bone after withdrawal in a non-growing bone site in ovariectomized rats.

Effects of risedronate and prostaglandin E2 (PGE2) alone or in combination on the distal tibia, a non-growing bone site with closed epiphysis at 3 months of age, were studied in ovariectomized (ovx) rats. Six-month-old Sprague-Dawley female rats were either ovx or sham-ovx. Rats were treated immediately after operation either with risedronate (5 micrograms/kg/2x/wk), PGE2 (6 mg/kg/d), or risedronate+PGE2 for 60 days (on-groups) and followed by 60 days without treatment (off-groups). Trabecular area, width and numbers were determined in metaphyseal cancellous bone of the distal tibia. No significant bone loss or structural changes were observed in the distal tibial metaphysis after 120 days of ovx. Risedronate alone did not produce any effect on bone mass during the treatment and the withdrawal periods. PGE2 alone increased the trabecular bone mass associated with thickened trabeculae and increased trabecular numbers. However, some of the newly formed bone was lost at the end of 60 days withdrawal. Combination of risedronate and PGE2 treatment added the same amount of bone mass as PGE2 alone, and the added new bone was maintained during the 60 days withdrawal. These results indicate that treatment with risedronate and PGE2 can preserve the anabolic effect of PGE2 on bone mass for at least 60 days after treatment.

Prostaglandin E2 administration prevents bone loss induced by orchidectomy in rats.

The objects of this study were to investigate whether prostaglandin E2 (PGE2) can prevent orchidectomy (ORX)-induced cancellous bone loss in growing male rats, and to determine the differential effects of PGE2 on sham-operated (sham) and ORX male rats. Fourteen-week-old Sprague-Dawley male rats were divided into groups of basal, vehicle-treated sham, PGE2-treated sham, vehicle-treated ORX, and PGE2-treated ORX rats for either 3 or 9 weeks. PGE2 was given at 6 mg/kg body weight daily by subcutaneous injection. Static and dynamic cancellous bone histomorphometry were performed on double-fluorescent labeled undecalcified proximal tibial metaphyseal sections. No effect was observed by ORX on body weight or longitudinal bone growth rate when compared with sham-operated controls. However, androgen deficiency caused significant increases in percent eroded perimeter, mineral apposition rate, and bone turnover (bone-volume-referent-bone formation rate), which resulted in a significant decrease in trabecular bone number, increase in trabecular separation, and a nonsignificant decrease in trabecular bone area by 3 weeks of ORX. After 9 weeks of ORX, trabecular bone area and number were significantly decreased, and trabecular separation, percent eroded perimeter, and the index of bone turnover (bone-volume-referent-bone formation rate) remained significantly increased while the index of bone formation (tissue-volume-referent-bone formation rate) was nonsignificantly decreased when compared with sham controls. When 6 mg PGE2/kg/day was given for 3 and 9 weeks, similar anabolic effects were observed in sham and ORX rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostaglandin E2 restores cancellous bone to immobilized limb and adds bone to overloaded limb in right hindlimb immobilization rats.

The purpose of this study was to determine whether prostaglandin E2 (PGE2) can restore cancellous bone mass and architecture to osteopenic, continuously immobilized (IM), proximal tibial metaphysis (PTM) in female rats. The right hindlimb of three and one-half-month-old Sprague-Dawley female rats were immobilized by right hindlimb immobilization (RHLI) in which the right hindlimb was underloaded and the contralateral left limb was overloaded during ambulation. After 4 or 12 weeks of RHLI, the rats were treated with 3 or 6 mg PGE2/kg/day and RHLI for 8 or 16 weeks. Bone histomorphometry was performed on microradiographs of PTM. Immobilization (IM) induced a transient cancellous bone loss and decreased trabecular thickness, number and node density, and increased free end density that established a new steady state after 4 weeks of IM. Three or 6 mg PGE2/kg/d for 8 weeks beginning at 4 or 12 weeks of IM completely restored cancellous bone mass (+127% to +188%) and structure to the age-related control levels in spite of continuous IM. Another 8 weeks of treatment maintained bone mass and architecture at these levels. No differences in cancellous bone mass and architecture were found between the overloaded PTM or RHLI rats and the age-related controls. However, 3 and 6 mg/kg/d of PGE2 treatment started at 4 or 12 weeks for 8 weeks significantly increased cancellous bone mass in the overloaded PTM (+45 to +74% of untreated controls), and another 8 weeks of treatment maintained bone mass at these levels.(ABSTRACT TRUNCATED AT 250 WORDS)