bluuebunny/crossref_metadata_2025 huggingface dataset:
Aluminum intoxication in vitamin D-deficient rats: Studies of bone aluminum localization and histomorphometry before and after vitamin D repletion
Authors: Dr. Hodsman A.B. , Steer B.M. , Arsenault A.L.
Published: February 2010

Aluminum accumulation by both dialysis patients and nonuremic patients, requiring chronic total parenteral nutrition, may be an etiological factor in the development of severe osteomalacia. To study the role of aluminum toxicity in bone, further experiments have been conducted in the nonuremic, vitamin D-deficient rat. Weanling rats were raised on vitamin D-deficient diets, and half received parenteral aluminum (5 mg/wk), for 30 days. In the first experiment low doses of 25-OH cholecalciferol (500 ng/week) were given subcutaneously for a further 30 days. Control rats were maintained on a similar protocol, but were supplemented with cholecalciferol (5 μg/week) from the outset until sacrifice at 60 days. In the second experiment a single bolus of cholecalciferol (5 μ/g) was given to study short-term changes in serum biochemistry and bone histology at 96 hr. Quantitative bone histomorphometric analyses of the proximal tibial metaphysis were made in all experimental groups. In the experimental vitamin D-deficient group, with the highest bone aluminum content (as assessed by extraction of whole bone aluminum), X-ray microanalysis was performed to determine the distribution of aluminum in bone tissue and bone cell organelles. The results showed that control rats treated with prolonged aluminum therapy (30 mg over 60 days) had evidence of both reduced osteoid matrix synthesis and mineralization. However, in vitamin D-deficient rats, there was no evidence that aluminum exacerbated the osteomalacic lesion, even though there was histochemical evidence of aluminum deposition at the bone-osteoid interface. X-ray microanalysis confirmed the presence of aluminum at this site, but did not reveal significant peaks of aluminum either in mineralized bone or within osteoblasts. In both experiments, vitamin D-induced osteomalacia was rapidly reversed by both low-dose 25-OH cholecalciferol and bolus cholecalciferol, suggesting that aluminum accumulation alone does not lead to impaired mineralization if other conditions favoring bone metabolism are present. In these experiments, secondary hyperparathyroidism may have been one such factor in explaining the failure of bone aluminum accumulation to impair bone mineralization in vitamin D-deficient rats when compared to vitamin D-replete controls. Although several animal models of aluminum toxicity have been explored, clearly the experimental conditions, dose, and duration of aluminum administration, and the degree of secondary hyperparathyroidism, are critical factors in determining whether aluminum accumulation in bone results in significant pathology.
Indomethacin inhibition of tenotomy-induced bone resorption in rats
Authors: Dr. Thompson D.D. , Rodan G.A.
Published: February 2010

Loss of biomechanical function results in rapid bone loss. This study assesses the role of arachidonic acid metabolites in immobilization-related osteopenia. A hind limb of the rat was immobilized by knee tenotomy and bone resorption and formation parameters were quantitated by histological methods in indomethacin-treated (0.5 mg/kg per day) and vehicle-treated animals. Control animals sacrificed 30, 72, and 240 hr post-tenotomy revealed a significant increase in osteoclast number (30 hr) and resorption surfaces (72 hr) and a decrease in trabecular bone volume (240 hr) in the tenotomized tibiae. In the indomethacin-treated tibial metaphysis, no significant differences were noted for these parameters by comparison to the nontenotomized leg. Bone formation parameters remained reduced in the tenotomized legs of both the indomethacin and vehicle-treated groups compared to the control legs. Indomethacin inhibited bone resorption, but did not prevent the decrease in bone formation produced by immobilization over the 10 days of these experiments.
Bone status of senescent female rats: Chemical, morphometric, and biomechanical analyses
Authors: Kiebzak Gary M. , Smith Ronald , Howe Juliette C. , Gundberg Caren M. , Dr. Sacktor Bertram
Published: February 2010

The bone status of female rats, 6, 12, and 24 months of age was examined. Femur Ca, Pi, and osteocalcin contents, as well as biomechanical properties, were measured and correlated to physical indices and serum chemistry. Diaphyseal Ca, Pi, and osteocalcin did not change significantly with increasing age. Serum Ca and Pi concentrations were not altered in the aged rat. Immunoreactive parathyroid hormone (PTH) levels increased significantly with age, when analyzed by linear regression. Serum osteocalcin decreased progressively from 6 to 12 months (-21%) and from 12 to 24 months (-23%). Maximum breaking force required to fracture femurs at midshaft did not change with senescence. Hence, the strength of the femurs as an intact organ was not compromised in aging. However, ultimate stress, a parameter that normalizes for differences in bone geometry and size, decreased 14% from 12 to 24 months. Changes in other biomechanical parameters, including yield and ultimate deformation, strain, and modulus of elasticity, were relatively small, but statistically significant, or were negligible. Morphometric measurements indicated a progressive age-related increase in second moment of area and cortical area. Medullary area did not change with age. Therefore, strength of the intact femur was maintained by architectural compensations, although normalized tissue strength decreased in senescence. The bone status and Ca/Pi homeostasis of the female rat were compared to similar findings, reported previously, for the male animal. The results suggest that bone status and mineral metabolism were compromised in the aged female rat, but the magnitude of change was less than that found for the senescent male rat.
Evidence of a probable role for 25-hydroxyvitamin D in the regulation of human calcium metabolism
Authors: Dr. Bell Norman H. , Epstein Sol , Shary Judith , Greene Virginia , Oexmann Mary Joan , Shaw Sheryl
Published: February 2010

1,25-Dihydroxyvitamin D [1,25-(OH)2D] is the principal mediator of the biologic effects of vitamin D. We showed previously that obese white subjects have low serum vitamin D and 25-hydroxyvitamin D (25-OHD) with increased serum-immunoreactive parathyroid hormone (PTH) and 1,25-(OH)2D, low urinary calcium, and increased urinary cyclic adenosine 3′,5′-monophosphate (cyclic AMP) compared with nonobese white individuals. To determine whether 25-OHD modulates calcium metabolism, the effects of 25-OHD3, 40–100 μg/day for 9 days, were compared in seven obese and seven nonobese white subjects who were between the ages of 20 and 34 years. Each of them was hospitalized on a metabolic ward and given a constant daily diet that contained 400 mg calcium, 900 mg phosphate, and 18 mEq magnesium. Whereas 25-OHD3 increased mean serum 25-OHD from 7 ± 1 to 37 ± 5 ng/ml (P < 0.01) and urinary calcium from 102 ± 18 to 146 ± 17 mg/day (P < 0.001) and decreased mean serum 1,25-(OH)2D from 40 ± 2 to 28 ± 2 pg/ml (P < 0.01) and urinary cyclic AMP from 3.23 ± 0.57 to 2.00 ± 0.17 nM/dl GF (P < 0.05), it did not change mean serum calcium, ionized calcium, phosphate, magnesium, immunoreactive PTH or urinary phosphate, or creatinine clearance in the obese subjects. In contrast, 25-OHD3 increased mean serum 25-OHD from 16 ± 1 to 46 ± 4 pg/ml (P < 0.001) but did not alter mean serum 1,25-(OH)2D or urinary calcium or cyclic AMP in the nonobese subjects. Metabolic balance studies carried out in two of the obese subjects showed that 25-OHD3 increased serum 25-OHD and urinary calcium and lowered serum 1,25-(OH)2D. The findings are interpreted to mean that elevated serum 1,25-(OH)2D and urinary cyclic AMP and low urinary calcium in obese subjects result from a deficiency of vitamin D and 25-OHD, that these changes are reversed by 25-OHD3, and that in humans 25-OHD has a modest role in the regulation of calcium metabolism that is independent of the regulation by 1,25-(OH)2D.
The relationship between long bone growth rate and duodenal calcium transport in female rats
Authors: Dr. Thomas Mary L. , Hope William G. , Ibarra Michael J.
Published: February 2010

Studies were carried out to determine the relationship between long bone growth rate and duodenal calcium (Ca) transport in female rats and the regulation of these two parameters by ovarian hormones. Female rats were ovariectomized (ovx) at 6 weeks of age. Some animals were implanted with silastic implants containing either estradiol or progesterone at the time of ovx. Studies were carried out 3 weeks later when the rats were 9 weeks old. Ovx resulted in an increase in long bone growth rate and duodenal Ca transport without any alteration in circulating levels of parathyroid hormone or 1,25-dihydroxyvitamin D [1,25-(OH)2D]. Animals receiving estradiol exhibited decreased long bone growth rate and duodenal Ca transport relative to ovx animals. These animals were mildly hypercalcemic and had lower levels of 1,25-(OH)2D than ovx or intact animals. The results of these studies suggest that the effects of ovarian hormone status on duodenal Ca transport are more closely related to long bone growth rate than to circulating levels of 1,25-(OH)2D. Further studies are required to determine whether the two parameters are coregulated by some as-yet-unidentified factor or whether bone growth is able to emit some signal, directly or indirectly, to increase duodenal Ca transport.
The distribution of podosomes in osteoclasts cultured on bone laminae: Effect of retinol
Authors: Zambonin-Zallone Alberta , Teti Anna , Carano Aldo , Marchisio Pier Carlo
Published: February 2010

Osteoclasts, isolated and purified from the medullary bone of calcium-deficient egg-laying hens, adhere to glass coverslips in vitro by means of specialized protrusions of the ventral membrane, denoted podosomes. These structures represent dotlike close-contact adhesion sites in which most cytoskeletal proteins involved in linking the plasma membrane to microfilaments are organized according to a specific and previously described pattern also shared by many oncogene-transformed cells. We show now that podosomes are not only a feature of osteoclasts adhering to artificial glass surfaces but are also present in the ventral membrane of osteoclasts adhering to bone laminae. Moreover, the quantity and the topography of podosomes may be modulated by retinol, which increases bone-resorbing activity of osteoclasts both in vivo and in vitro. A comparative transmission electron microscopy study of osteoclasts adhering on bone laminae in vitro or in vivo indicates that podosomes with identical features are present in the clear zone of the osteoclasts in either condition. Since podosomes are the sealing structures of the clear zone, podosome formation may represent one of the modifications involved in the reorganization process of the osteoclast that precedes bone resorption.
Effects of cisplatin on parathyroid hormone- and human lung tumor-induced bone resorption
Authors: Abramson Edith C. , Chang Jeffry , Mayer Marjorie , Kukla Lawrence J. , Shevrin Daniel H. , Lad Thomas E. , Buschman Robert , Kukreja Subhash C.
Published: February 2010

We have previously shown that dichlorodiamine platinum (DDP), or cisplatin, a cancer chemotherapeutic agent, is effective in the treatment of malignancy-associated hypercalcemia. In the present studies, we evaluated its effects on bovine parathyroid hormone (PTH)- or tumor-induced bone resorption in vitro in the neonatal mouse calvarial bone resorption assay. PTH alone or tumor extract (TE) of a human squamous cell lung cancer alone caused a significant increase in the bone resorption and in the number of osteoclasts in the calvaria. The addition of 3 and 10 μg/ml DDP inhibited the PTH- or TE-induced bone resorption. Lower doses of 1 and 2 μg/ml DDP, although not effective in inhibiting the PTH-induced bone resorption, were effective in lowering the TE-induced bone resorption. The number of osteoclasts was also reduced by DDP treatment. We therefore conclude that DDP is effective in the treatment of malignancy-associated hypercalcemia by virtue of its inhibitory effects on osteoclast numbers and on bone resorption.
Skeletal contribution of cyclic adenosine monophosphate in response to parathyroid hormone and calcitonin in vivo in the rat
Authors: Dr. Morrison Nancy E. , Ramanathan Subramanyam , Suh Se Mo
Published: February 2010

Cyclic adenosine monophosphate (cAMP) is thought to be a second messenger for the actions of both parathyroid hormone (PTH) and calcitonin (CT). We examined the release of cAMP from rat bone in vivo after administration of synthetic rat PTH-(1–34) (rPTH), synthetic human PTH-(1–34) (hPTH), or synthetic human CT (hCT). Blood from the venous effluent of the femoral bone of rats (bone blood) was drawn at 5 and 10 minutes after the administration of hormones. The cAMP content of the bone blood was then compared to the cAMP content of arterial blood. In both kidney-clamped and non-kidney-clamped rats, hCT led to a significantly greater concentration of cAMP in the bone blood than in the arterial blood. We interpret this to be due to bone production and release of cAMP. Neither hPTH nor rPTH produced a significantly greater amount of cAMP in the bone blood than in arterial blood. These data do not preclude the possibility that there was a production of cAMP within the bone tissue itself after PTH but suggest that there was no release of cAMP from the bone into the bone blood.
Direct transformation from quiescence to bone formation in the adult periosteum following a single brief period of bone loading
Authors: Pead Matthew J. , Skerry Timothy M. , Dr. Lanyon Lance E.
Published: February 2010

The concept of resorption preceding formation in a coupled response is well established as the normal sequence of remodeling in adult bone. So prevalent is this concept, however, that the idea of the direct activation of osteogenic modeling in normal adult bone is often ignored. This experiment documents the direct transformation of the normal, quiescent, adult periosteum to active bone formation. The osteogenic stimulus was provided by a single short period of dynamic loading. Periosteal activation and the production of new bone within 5 days of loading was unaccompanied by resorption or the presence of osteoclasts. We therefore conclude that an adult resting periosteum can become directly converted to formation as a physiologic response to an appropriate osteogenic stimulus without the need for resorption. To distinguish this process from remodeling we suggest it be called renewed modeling. It is notable that a single short exposure to an “osteogenic” loading regime can influence the full cascade of cellular events between quiescence and active bone formation.
Elevated calcium, phosphorus, and magnesium retention in pregnant rats prior to the onset of fetal skeletal mineralization
Authors: Brommage Robert , Baxter Daniel C.
Published: February 2010

Intestinal absorption and renal excretion rates of Ca, P, and Mg were compared in pregnant and control virgin rats fed a purified diet containing 0.55% Ca, 0.54% P, and 0.067% Mg. Four consecutive balance intervals of 5 days each were examined, beginning on day 1 of gestation. During days 6–10 of gestation, pregnant rats had elevated fractional intestinal absorptions of Ca (29.6 ± 1.7 versus 20.6 ± 1.5%), P (65.4 ± 2.0 versus 59.9 ± 0.9%), and Mg (54.3 ± 1.5 versus 42.4 ± 2.6%). Since urinary excretions of these elements did not change, the pregnant rats retained more Ca, P, and Mg than virgin rats. Fractional intestinal absorptions of these elements during pregnancy were similar to control values during days 1–5 and 11–15 of gestation and then rose for each element during days 16–20 of gestation. Presumably because of an increase in glomerular filtration rate, urinary excretions of Ca and Mg were elevated by 40 and 26% during days 16–20 of gestation. In contrast, urinary P excretion was decreased by 30% during days 11–20 of gestation. Analyses of uterine mineral contents indicated the increased maternal Ca and Mg retentions during pregnancy were balanced by the transfers of these elements to the fetuses. In contrast, pregnancy was associated with a net maternal retention of P. These data are consistent with previous observations of increased maternal skeletal mineralization during early pregnancy before the onset of fetal osteogenesis and subsequent enhanced maternal Ca intestinal absorption concurrent with fetal skeletal mineralization.
Determinants of peak trabecular bone density in women: The role of androgens, estrogen, and exercise
Authors: Buchanan James R. , Myers Cathleen , Lloyd Tom , Leuenberger Paula , Demers Laurence M.
Published: February 2010

To elucidate determinants of peak trabecular bone density, we studied the role of androgens, estrogen, and aerobic exercise in 30 women from 18 to 22 years old. The women were divided into three groups: Sedentary, 11 normal women who did not exercise regularly; eumenorrheic, 10 athletes with normal menstrual function; and oligomenorrheic, 9 athletes with exercise-induced oligomenorrhea. All athletes participated in aerobic sports that did not involve selective resistance loading of the back. Serum free and albumin-bound testosterone (fab T), androstenedione (A), and estradiol (E2) were measured on four separate occasions at consecutive 7 day intervals and averaged. Trabecular density was measured by quantitative computed tomography of the lumbar spine.

Peak trabecular bone density was related to fab T (r = 0.48, p = 0.007), A (r = 0.40, p = 0.03), and E2 (r = 0.40, p = 0.04). When taken in combination, androgens and estrogen each accounted independently for significant portions of the variance in bone density [fab T and E2 (R2 = 0.38, p = 0.002) and A and E2 (R2 = 0.27, p = 0.01)]. Bone density (mg/ml, mean ± standard error of the mean, SEM) in the sedentary group (174 ± 6) was not significantly different from that in the eumenorrheic (183 ± 12, p = 0.47) or oligomenorrheic (161 ± 11, p = 0.32) subjects.

We conclude that androgens and estrogen function as independent and additive determinants of peak trabecular bone density in young women. The quantitative impact of aerobic exercise (without resistance loading) and exercise-induced menstrual dysfunction appears to be less important than that of the hormones.
Tetrapyrroles as inhibitors of normal cartilage metabolism: Relative potency of different compounds
Authors: Vassilopoulou-Sellin Rena , Oyedeji Caroline O.
Published: February 2010

The present study was designed to explore the role of different tetrapyrroles as inhibitors of cartilage metabolism. We studied the effects of tetrapyrroles on the incorporation of [35S]sulfate into proteoglycans, [14C]-leucine into protein, and [3H]uridine into the RNA of normal cartilage from two different vertebrate classes using the embryonic chicken pelvic rudiment bioassy and the hypophysectomized rat costal cartilage bioassay, both very sensitive to cartilage growth factors and growth inhibitors. We compared the relative potencies of the following compounds: both metalloporphyrins (heme and chlorophyllin), linear tetrapyrroles (bilirubin), and heme proteins (hemoglobin, myoglobin, and cytochrome c). Hemoglobin and heme were the most potent inhibitors of rat cartilage metabolism, and bilirubin was a far more potent inhibitor of embryonic chick cartilage metabolism. Chlorophyllin had moderate inhibitory activity, especially on chick cartilage, whereas cytochrome c was inactive in these bioassays. Surprisingly, myoglobin was relatively ineffective despite its close similarity to heme and hemoglobin. The bilirubin-induced inhibition of sulfate incorporation into chick cartilage was partially prevented when glutathione was included in the incubation medium, suggesting that a free-radical mechanism may be involved. There were significant differences in the sensitivity of the two cartilages studied, indicating there may be species-dependent sensitivity to different tetrapyrroles.
Glucocorticoids increase parathyroid hormone receptors in rat osteoblastic osteosarcoma cells (ROS 17/2)
Authors: Yamamoto Itsuo , Potts John T. , Segre Gino V.
Published: February 2010

The effects of glucocorticoids on parathyroid hormone (PTH) receptors was studied using rat osteosarcoma-derived cells (ROS 17/2), which have an osteoblastic phenotype, and [125I][Nle8, Nle18, Tyr34]bovine(b)PTH-(1–34)amide as the radioligand. Treatment of cells with physiologic concentrations of hydrocortisone resulted in a time and dose-dependent increase in PTH binding. The increase in PTH binding could be observed by 10 h of exposure to hydrocortisone (2 × 10−7 M), was maximally enhanced by 48 h, and was maintained for the subsequent 7 days of continuous exposure to the steroid. With removal of hydrocortisone, PTH receptor binding promptly returned toward control levels. The increase in PTH binding was attributed to an increase in the availability of receptor binding sites, not to altered receptor binding affinity, and was blocked by cycloheximide. PTH-stimulated adenylate cyclase was also enhanced by glucocorticoids, and a close correlation was observed between PTH binding and PTH-stimulated adenylate cyclase. However, hydrocortisone not only increased PTH binding but also enhanced the efficiency of postreceptor signaling: 5′-guanylimidodiphosphate [Gpp(NH)p]- and forskolin-stimulated adenylate cyclase activities were also increased. Thus, enhanced PTH stimulation of adenylate cyclase by glucocorticoids resulted from at least two effects—increased receptor availability and enhanced postreceptor efficiency of transmembranous signaling.
Characterization of insulin binding in the UMR-106 rat osteoblastic osteosarcoma cell
Authors: Ituarte Eloy A. , Ituarte Hector G. , Iida-Klein Akiko , Hahn Theodore J.
Published: February 2010

The correlation of insulin receptor occupancy with classic insulin effects, such as stimulation of glucose uptake, have not been examined in osteoblastlike cells. Accordingly, we characterized insulin binding and examined its relationship to stimulation of glucose analog transport in the UMR-106 rat osteoblastic osteosarcoma cell line. Insulin binding in UMR-106 cells was found to be pH sensitive, temperature dependent, saturable, and specific. Proinsulin was 100-fold less effective than insulin in displacing specific [125I]insulin binding in these cells, whereas IGF-I at concentrations between 0.1 and 10 nM produced no displacement of [125I]insulin but did produce significant displacement of insulin binding at 100 and 1000 nM. Insulin receptor downregulation was observed after exposure to 100 nM insulin for 6 h at 37°C and was temperature dependent. Insulin binding was reversible after 24 h at 4°C. Insulin binding correlated directly with stimulation of 2-deoxyglucose uptake at insulin concentrations between 0.1 and 100 nM, with a half-maximal concentration (ED50) of 0.9 nM for both [125I]insulin binding displacement and stimulation of 2-deoxyglucose uptake. Hence, there was no evidence for spare insulin receptors with regard to stimulation of glucose analog transport. Scatchard analysis of insulin binding kinetics yielded a curvilinear plot, suggesting negative cooperativity. Analysis of insulin binding kinetics using a two-site model yielded a KD of 0.9 nM for the apparent high-affinity binding site and an estimated 80,000 high-affinity binding sites per cell. These findings demonstrate that osteoblastlike cells exhibit a relationship between insulin binding and glucose transport stimulation that is similar to that in liver cells and other insulin-sensitive tissues. The UMR-106 cell line therefore appears to represent a useful system for studies of the regulation of insulin binding and action in an osteoblastic cell.
Interleukin-1 and tumor necrosis factor stimulate the formation of human osteoclastlike cells in vitro
Authors: Pfeilschifter Johannes , Chenu Chantal , Bird Andrew , Mundy Gregory R. , Roodman David G.
Published: February 2010

Interleukin-1 (IL-1) α and β and tumor necrosis factor (TNF) α and β are potent stimulators of bone resorption in vitro and in vivo. However, the mechanisms underlying this increased bone resorption have not been clearly defined. Increased bone resorption can result from increased activity of individual osteoclasts, increased numbers of osteoclasts, or both. Therefore, we have used a long-term human marrow culture system that forms multinucleated cells (MNC) with the characteristics of osteoclasts to examine the effects of IL-1 and TNF on osteoclast formation. Human recombinant IL-1α and IL-1β and human recombinant TNF-α and TNF-β stimulated MNC formation from 4- to 60-fold. IL-1α, IL-1β, TNF-α, and TNF-β significantly increased MNC formation at very low concentrations: 2.5 × 10−13 M for IL-1α and IL-1β, 10−11 M for TNF-α, and 10−10 M for TNF-β In addition, these cytokines enhanced MNC formation in the presence of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], a potent osteotropic factor that stimulates MNC formation by stimulating fusion of mononuclear precursor cells. Simultaneous addition of IL-1 and TNF to the cultures resulted in a synergistic stimulation of MNC formation. These results suggest that: (1) IL-1 and TNF stimulate bone resorption in part by increasing osteoclast formation and (2) an extremely low concentration of these factors can synergistically increase osteoclast formation in the absence of other factors, such as 1,25-(OH)2D3. These data suggest that synergistic interactions among cytokines play an important role in maintaining bone cell activity in normal and pathologic states.
Cyclic therapy of osteoporosis with neutral phosphate and brief, high-dose pulses of etidronate
Authors: Mallette Lawrence E. , Leblanc Adrian D. , Pool James L. , Mechanick Jeffrey I.
Published: February 2010

We have designed a cyclic regimen for the treatment of osteoporosis based on the activate, depress, free, and repeat (ADFR) concept. Osteoclastic bone resorption is activated by 7 days of oral neutral phosphate and inhibited with a brief pulse (5 days) of etidronate disodium at a high dose (20 mg/kg body weight). Patients next take calcium supplements for 48 days before resuming phosphate to enter the next cycle. Osteoporotic women increased the bone mineral density of the lumbar spine at 6 months by 7.2 ± 5.2% (mean ± SD, N = 14) and at 12 months by 8.2 ± 4.0% (N = 8). Control observations in regularly exercising postmenopausal women (N = 30) showed no significant change in spine mineral density after 20 months (0.5 ± 3.2%), confirming the stability of the measurement technique. The two patients who responded poorly to the cyclic regimen each showed a blunted rise in serum PTH during oral phosphate administration, suggesting that the rise in PTH induced by oral phosphate may be an important component of this cyclic regimen. This preliminary study does not identify which component or components of the regimen are responsible for the increase in bone mass but provides positive encouragement for randomized studies designed to determine the optimum dosage, duration, and timing of each component of the regimen.
Transforming growth factor β modulates proliferation of osteoblastic cells: Relation to its effect on receptor levels for epidermal growth factor
Authors: Uneno Susumu , Dr. Yamamoto Itsuo , Yamamuro Takao , Okumura Hideo , Ohta Shuichi , Lee Kaechoong , Kasai Ryuichi , Konishi Junji
Published: February 2010

The effect of transforming growth factor β (TGF-β) on cellular proliferation of osteoblastic MC3T3-E1 cells was studied with particular emphasis on its effect on modulation of epidermal growth factor (EGF) receptors. In other cells, TGF-β has been reported to augment EGF receptors. Exposure of MC3T3-E1 cells to TGF-β initially increased cell surface EGF receptor levels and decreased the rate of DNA synthesis. The initial elevation of EGF receptor levels was due to increased receptor number per cell, not to changes in binding affinity. On the contrary, prolonged exposure (longer than 40 h) resulted in a decrease in EGF receptor and an increase in the rate of DNA synthesis. Thus, the effects of TGF-β on these cells appears to be biphasic, reflecting complex mechanisms of action; the early effects of TGF-β may be consistent with cellular differentiation to the osteoblastic phenotype with decreased cellular proliferation, whereas chronic exposure of these cells to TGF-β stimulated cellular proliferation and inhibited osteoblastic phenotype expression. It is not likely that stimulation of cellular proliferation was through elevation of EGF receptor levels, because TGF-β did not enhance the stimulatory effect of EGF on cellular proliferation. Thus, we conclude that TGF-β possesses a stimulatory effect on the cellular proliferation of osteoblastic MC3T3-E1 cells independent of its modulative effect on EGF receptor level.
Quantification of bone mineral content using dual-photon absorptiometry in a normal Japanese population
Authors: Hagiwara Satoshi , Miki Takami , Nishizawa Yoshiki , Ochi Hironobu , Onoyama Yasuto , Morii Hirotoshi
Published: February 2010

The bone mineral content of lumbar spine and/or total body were quantified in 217 healthy Japanese (86 males and 131 females) using a Dichromatic Bone Densitometer (Norland Corp., Model 2600). The bone mineral density of the third lumbar vertebra (L3 BMD) decreased significantly after age 20 in males (r = -0.417, p < 0.0002), with acceleration of the decrease after age 50 (r = -0.621, p < 0.00002). A significant correlation was found between L3 BMD and age after age 40 (r = -0.747, p < 0.00001) in females. L3 BMD correlated with both the body height (r = 0.335, p < 0.0001) and the body weight (r = 0.340, p < 0.0001). Total bone mineral content from the second to fourth lumbar vertebrae correlated significantly with total body bone mineral (r = 0.880, p < 0.00001) in these normal subjects. Lumbar spine bone mineral as measured by dual-photon absorptiometry is lower in Japan than the bone mass in the United States, although not lower than in other parts of the world.
Plasma carriers influence the uptake of cholecalciferol by human hepatoma-derived cells
Authors: Haddad John G. , Aden David P. , Aw T. Choon
Published: February 2010

The uptake of [3H]cholecalciferol by the human hepatoma-derived cell lines Hep G2 and Hep 3B was examined as a function of the sterol's presentation on various plasma proteins at their native concentrations. Control cultures utilized devitalized cells cross-linked with glutaraldehyde and estimated nonspecific sterol adherence to cells. With both cell lines, neither albumin nor plasma vitamin D binding protein permitted cholecalciferol uptake above control values. With Hep G2 cells, only low-density lipoprotein presentation of the sterol resulted in significant cellular uptake that had features resembling a receptor-mediated process. With Hep 3B, only high-density lipoprotein presentation of the sterol resulted in a significant uptake that was cell, carrier, and time dependent. These results support the hypothesis that lipoprotein carriers could account for the efficient hepatic accumulation of cholecalciferol in vivo.
Bone resorption by isolated human osteoclasts in vitro: Effects of calcitonin
Authors: Murrills Richard J. , Shane Elizabeth , Lindsay Robert , Dr. Dempster David W.
Published: February 2010

Human osteoclasts were isolated from 12- to 17-week-old fetal tissue and from transiliac crest bone biopsies for an in vitro study of their biology. A hypodermic needle was used to flush either the fetal long bones or the trabeculae of the iliac crest bone biopsy with tissue culture medium and the resulting cell suspension sedimented briefly either onto the surface of plastic tissue culture dishes, for time-lapse microcinematography, or onto slices of devitalized bovine cortical bone for quantitative assay of bone resorption. The osteoclasts were motile, tartrate-resistant acid phosphatase positive and capable of excavating pits in slices of devitalized bovine cortical bone. Human calcitonin, at doses of 1 ng/ml and 1 μg/ml, caused a 70% inhibition of bone resorption by human fetal osteoclasts over a 24 h period but had no apparent effect on the morphology or motility of either fetal or adult osteoclasts.
Effects of calcitonin gene-related peptide on cyclic AMP formation in chicken, rat, and mouse bone cells
Authors: Michelangeli V. P. , Fletcher A. E. , Allan E. H. , Nicholson G. C. , Martin T. J.
Published: February 2010

Mixed bone cell cultures obtained by sequential collagenase-trypsin digestion of newborn chick, rat, and mouse calvaria responded to calcitonin gene-related peptide (CGRP) with a dose-dependent increase in cyclic AMP formation. The amplitude of response to CGRP in each species was less than that to parathyroid hormone (PTH). The CGRP effect was not the result of an action as a weak calcitonin agonist, since in most instances a calcitonin effect was not observed. Only in early digests of mouse calvarial cells were consistent stimulatory effects of calcitonin on cyclic AMP noted, and these were always considerably less in amplitude than those to CGRP. It is concluded that chick, rat, and mouse bones contain cells in osteoblast-rich populations that respond specifically to CGRP with a rise in cyclic AMP.
Bone-resorbing activity of thyroid hormones is related to prostaglandin production in cultured neonatal mouse calvaria
Authors: Klaushofer Klaus , Hoffmann Oskar , Gleispach Helmut , Leis Hans-Jörg , Czerwenka Erwin , Koller Kristian , Peterlik Meinrad
Published: February 2010

The bone-resorbing activity of thyroid hormones was evaluated in neonatal mouse calvaria maintained in organ culture for 96 h. Thyroxine (T4) between 10−8 and 10−5 mol/liter and triiodothyronine (T3) between 10−8 and 10−7 mol/liter caused a dose-dependent release of calcium from cultured bone. The thyroid hormone effect was delayed in onset for at least 24 h, and after 96 h of culture amounted to 50–90% of the bone-resorbing activity of 10−8 mol/liter parathyroid hormone (PTH). The bone-resorbing action of T4 as well as of T3 was completely blocked by 100 U/ml interferon-γ (IF-γ) or 20 mU/ml salmon calcitonin (CT). “Escape” from CT inhibition, which is a well-known phenomenon in the action of PTH, was not observed with thyroid hormone-mediated bone resorption. Thyroid hormone treatment of cultured calvaria resulted in a gradual increase between 48 and 96 h of medium concentrations of prostaglandin (PG) E2 and particularly of 6-keto-PGF1α, the inactive metabolite of prostacyclin (PGI2). The release of PGF2α in general was not significantly affected. Although the effect of thyroid hormones on PG release from cultured calvaria was completely abolished by 5 × 10−7 mol/liter indomethacin, in some experiments indomethacin reduced thyroid hormone-mediated bone resorption by only 50%. This indicates that thyroid hormone action on bone is also mediated by a PG-independent mechanism.
Comparison of cross-sectional and longitudinal measurements of age-related changes in bone mineral content
Authors: Davis James W. , Ross Philip D. , Wasnich Richard D. , Maclean Charles J. , Vogel John M.
Published: February 2010

Age-related, postmenopausal bone loss was examined among a cohort of Japanese-American women living in Hawaii. None of the women were using estrogens or thiazides. Cross-sectional and longitudinal measurements of bone mineral content were compared at the calcaneus, the proximal radius, and the distal radius. Cross-sectional measurements were also available for the lumbar spine. The longitudinal data showed a slowing rate of bone loss with increasing age at the radius sites. By contrast, the cross-sectional data suggested constant rates of bone loss for all ages at both radius sites and the spine. The calcaneus demonstrated a complex pattern of bone loss in both cross-sectional and longitudinal analyses. The loss rates among women in their fifties were greater than for those in their sixties. From the middle sixties onward calcaneal bone loss remained essentially constant. Because of the sustained bone loss, however, women in their seventies were actually losing greater percentages of their calcaneal bone mineral than they had in their sixties.
Accelerated bone resorption in senescence-accelerated mouse (SAM-P/6)
Authors: Kawase Masahiro , Tsuda Masao , Matsuo Takao
Published: February 2010

Age-associated changes in the femoral bone and in urine and serum composition were studied to understand the low bone mass in a substrain of senescence-accelerated mouse (SAM), SAM-P/6. Age-matched normal SAM-R/1 mice were used as controls. After 13 weeks of age, the concentration of hydroxyproline in the femur of SAM-P/6 was slightly but significantly lower. The urinary excretion rates of cAMP and calcium were higher in SAM-P/6 throughout the whole experimental period, and those of hydroxyproline and phosphorus were higher after 8 weeks. The serum concentrations of calcium and inorganic phosphorus of SAM-P/6 were higher at 13 and 20 weeks. At 5 weeks, the serum alkaline phosphatase and tartrate-resistant acid phosphatase (TRAP) activities of SAM-P/6 were significantly higher. Furthermore, the serum of SAM-P/6 significantly stimulated calcium release from cultured fetal rat ulna. Since urinary cAMP and TRAP in the serum reflect the circulating level of parathyroid hormone and osteoclastic function, respectively, the present results suggest that, in SAM-P/6, accelerated bone resorption produced by a putative hyperparathyroid state causes the decrease in bone mass.
Renal cell carcinoma in tissue culture secretes nondialyzable product that stimulates bone resorption in organ-cultured mouse calvaria
Authors: Dr. Lerner Ulf H. , Ljungberg Börje
Published: February 2010

The bone-resorbing capacity of human renal cell carcinomas in vitro has been examined. Bone resorption in cultures of mouse calvarial bones was assessed by the release of 45Ca from bones prelabeled in vivo and the mobilization of stable calcium and inorganic phosphate from nonlabeled bones. In addition, bone organic matrix degradation was determined either by the release of 3H from [3H]proline-labeled bones or by the loss of hydroxyproline from bone explants during culture. Tumor tissue-conditioned media (TCM) from 13 of 13 renal cell carcinomas stimulated bone resorption in a dose-dependent manner. From 5 of 13 kidneys with renal cell carcinoma, normal kidney cortex tissue was cultured and 4 of these 5 also produced bone-resorbing activity, but the amount was much less compared with the tumor tissue. The stimulatory effect of TCM on 45Ca release could be observed first after 12–24 h of culture. The effect could be inhibited by calcitonin but not by inhibitors of prostaglandin synthesis. The production of bone-resorbing activity by tumor cells could be inhibited by indomethacin and meclofenamic acid. In some tumors, the inhibition by indomethacin was total, whereas in other tumors only partial inhibition could be obtained. In 3 of 4, TCM bone-resorbing activity could be found in the retentate after dialysis. The results show that fresh human renal cell carcinoma tissue can elaborate prostanoid as well as nonprostanoid products that can stimulate bone resorption.
Acetate inhibition of chick bone cell proliferation and bone growth in vitro
Authors: Saitta Joseph C. , Lipkin Edward W. , Howard Guy A.
Published: February 2010

A hypothesis has been advanced that parenteral solutions as commonly formulated for use in clinical practice have a toxic effect on cell metabolism. A specific component of these solutions, sodium acetate, has been suggested to disrupt normal bone turnover and therefore to contribute to the osteopenia observed in patients receiving hemodialysis and parenteral nutrition (PN). We developed an in vitro model to test the hypothesis that sodium acetate at concentrations that are infused in PN solutions has a deleterious effect on bone metabolism. Osteoblasts and preosteoblasts from 16- to 17-day-old embryonic chick calvaria, and tibiae and femora from 10-day-old embryonic chicks were grown in BGJb medium (control) or in BGJb medium plus sodium acetate (5, 10, or 20 mM). Calvarial cell proliferation was quantified by direct cell counts as well as by incorporation of [3H]TdR into DNA as an index of cell proliferation. Calvarial cell alkaline phosphatase activity was quantified by the ability of extracts of the cultured cells to hydrolyze p-nitrophenyl phosphate to p-nitrophenol, and bone growth was determined by measuring final dry weight. Calvarial cell counts as well as DNA synthesis showed a dose-dependent decrease in the presence of sodium acetate (5–20 mM) compared with controls. [3H]TdR incorporation was decreased a mean 19% with 5 mM, 38% with 10 mM, and 63% with 20 mM acetate. Alkaline phosphatase activity per cell increased 48% with 5 mM, 140% with 10 mM, and 355% with 20 mM acetate. Cell viability as assessed by trypan blue exclusion was identical for test and control media (>95%). Long bone dry weight was decreased 6% with 5 mM acetate, 15% with 10 mM, and 33% with 20 mM acetate compared with controls. These effects were independent of the presence or absence of fetal bovine serum in the culture medium and could not be accounted for by pH differences between the test and control media. We conclude that sodium acetate inhibits the proliferation of chick osteoblasts and preosteoblasts as well as the growth of chick long bones in vitro and that the potential role of sodium acetate in parenteral solutions as a contributor to abnormal bone metabolism merits further investigation.
Cyclosporin A in the oophorectomized rat: Unexpected severe bone resorption
Authors: Movsowitz Colin , Epstein Sol , Ismail Firhaad , Fallon Michael , Thomas Steven
Published: February 2010

Local factors, such as interleukin-1, may mediate the accelerated bone remodeling in the acute estrogen-deficient rat. Cyclosporin A (CsA), which in vitro inhibits some of these local factors, was administered to oophorectomized (OX) rats in an attempt ot modify this high turnover state. Three groups of 15 rats were studied. Group A was sham operated, group B was OX, and group C was OX and received CsA (15 mg/kg per day) by gavage commencing 4 days postoophorectomy for 28 days. Estradiol levels were determined to confirm oophorectomy. Blood was sampled on days −7, 0, 7, 14, 21, and 28 for ionized calcium (Ca2+), 1,25-(OH)2-vitamin D, PTH, and bone gla protein (BGP). Rats received tetracycline hydrochloride for bone histomorphometric labeling. All results were compared to group A. Body weight was increased in group B (p < 0.003) but not in group C. There was no difference in Ca2+ or PTH between the groups. BGP levels were higher in group B by day 28 (p < 0.05); BGP levels were increased in group C from days 7–28 (p < 0.002). 1,25-(OH)2-vitamin D was significantly increased in group C (p < 0.0001) but not in group B. Tibial bone histomorphometry revealed increased measurements of bone formation and osteoclast number without a loss of bone volume (BV/TV) in group B. Group C showed a dramatic increase in bone turnover with significant loss of BV/TV (p < 0.001). In conclusion, CsA in the OX rat resulted in unexpected enhanced bone remodeling with high BGP levels and severe bone resorption. These changes may be due to CsA alone or may be modified by oophorectomy.
Relationship of body composition, muscle strength, and aerobic capacity to bone mineral density in older men and women
Authors: Bevier Wendy C. , Wiswell Robert A. , Pyka Gisela , Kozak Kathryn C. , Newhall Katherine M. , Marcus Robert
Published: February 2010

We evaluated the relationship of body composition, maximal aerobic capacity (VO2max), and muscle strength to bone mineral density in 91 healthy men and women, age 61–84 years. Lean body mass was estimated from two independent measures of fat mass, bioelectrical impedance and skinfold thickness. VO2max was determined by treadmill ergometry with direct measurement of oxygen consumption. Grip and back strength were measured by isometric dynamometry. Mineral density of lumbar spine and midradius were measured by dual- and single-photon absorptiometry. Men had significantly greater lean mass, muscle strength, aerobic capacity, and bone density than women. In women, grip strength correlated with forearm and spine density (r = 0.37, r = 0.28, p < 0.05). In men, grip strength correlated with forearm density (r = 0.47, p < 0.05), and back strength was significantly correlated with both spine (r = 0.46, p < 0.01) and forearm density (r = 0.46, p < 0.01). In women, neither forearm nor spine density correlated significantly with aerobic capacity. In men, midradius density did not correlate significantly with oxygen consumption, but the simple correlation between spine density and VO2max was significant (r = 0.41, p < 0.05). Back strength and VO2max were significantly related in men (r = 0.47, p < 0.01). By stepwise multiple regression, back strength emerged as the most robust predictor of spine mineral, accounting for 19% of the variation in bone density. Addition of VO2max to the regression did not add significant predictive value. However, when VO2max was expressed per kilogram lean body mass, both back strength and VO2max contributed significantly to the prediction of spine density in men, and the coefficient of determination R2 increased to 0.30. We conclude that body mass and grip strength, but not aerobic capacity, significantly predict bone density in elderly women. In elderly men, back strength is a more robust predictor of axial bone density than traditional expressions of aerobic capacity, but VO2max per kilogram lean mass and back strength both make significant contributions to the prediction of spine mineral density. The applicability of these results to younger men and women is uncertain.
Calcium absorption in women: Relationships to Calcium intake, Estrogen status, and age
Authors: Heaney Robert P. , Recker Robert R. , Stegman Mary Ruth , Moy Alan J.
Published: February 2010

A total of 526 absorption studies, using both the double-isotope and balance-based methods, were performed in 189 middle-aged women in good general health. The study extended over 17 years of observation, with most subjects studied from two to four times at 5 year intervals. Each study was done on the woman's own self-selected calcium intake and was carried out under inpatient, metabolic balance controls.

There was a highly significant inverse correlation between calcium intake and absorption fraction, with the best fit provided by an hyperbola in which absorption fraction is approximately inversely proportional to the square root of intake. The range of absorptive performance was very broad at all intake levels. Mean absorption fraction declined from a value of 0.45 at very low intakes (approximately 200 mg Ca per day) to approximately 0.15 at intakes above 2000 mg/day. There was a highly significant fall in absorption efficiency with age, amounting to approximately 0.0021 per year and a one-time decrease, amounting to approximately 0.022 at the time of menopausal estrogen loss.