ISSN:0884-0431    

DOI:10.1002/jbmr.5650070502

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractWe examined the effect of intermittent administration of bovine parathyroid hormone (1–34) (bPTH) on spinal bone mineral content (BMC) and bone mineral density (BMD), serum 1,25‐dihydroxyvitamin D concentrations, and serum markers of osteoblast function in senile male and female rats (23 and 24 months of age, respectively). Sexually mature young (3 month) male rats were similarly treated for comparison. bPTH administration increased serum osteocalcin concentrations without changing serum inorganic phosphate or calcium concentrations in either group of old animals. In young animals, PTH administration increased the serum calcium and inorganic phosphate concentrations significantly (p<0.05), although values remained within the normal range. In the vehicle‐treated male rats, serum 1,25‐dihydroxyvitamin D concentrations were lower in the senile than in the young animals (18 ± 5 versus 47 ± 6 pg/ml,p<0.05). PTH administration resulted in significantly increased serum 1,25‐dihydroxyvitamin D concentrations in the senile and young male animals (both,p<0.05) and the final mean serum 1,25‐dihydroxy vitamin D concentrations were not statistically different (68 ± 9 versus 85 ± 6 pg/ml respectively;p= NS). Serum 1,25‐dihydroxyvitamin D concentrations were significantly (p<0.05) higher in the PTH‐treated senile female rats than the sex‐matched, vehicle‐treated controls. The pretreatment spinal BMC and BMD as assessed by dual‐energy x‐ray absorptiometry (DEXA) were significantly higher in the senile male animals than in the young animals. Spinal BMC and BMD decreased in the vehicle‐treated senile male rats (p<0.05) over the 3 weeks of the study despite a gain in weight. bPTH administration prevented this fall in spinal BMC and increased spinal BMD (p<0.05). Spinal BMC and BMD increased significantly (p<0.05) in the young vehicle‐treated rats, and PTH administration caused a further significant increase in both parameters of bone mass. Spinal BMD was significantly (p<0.05) higher in the PTH‐treated senile female rats than in vehicle‐treated controls. These findings demonstrate that intermittent PTH administration increases serum 1,25‐dihydroxyvitamin D concentrations of senile animals to the levels of PTH‐treated, young, sexually mature animals. In addition, PTH administration arrests bone loss

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070503

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractATP released from damaged cells or by controlled secretion could be an important factor in the formation or remodeling of bone. In a variety of other tissues ATP has been shown to control cellular processes by acting on P2‐purinoceptors and activating the calcium signaling pathway. Here we demonstrate for the first time that extracellular ATP increases the intracellular free calcium [Ca2+]iconcentration in normal human osteoblasts and in SaOS‐2 cells, a human osteosarcoma‐derived cell line, but not in ROS 17/2.8 cells. The ATP‐induced increase in [Ca2+]iwas dose dependent, and the concentrations of ATP required were similar to those reported to regulate cellular functions in other cell types. Although ATP is metabolized rapidly by bone cells, the effects on [Ca2+]iappeared to be mediated directly by ATP rather than one of its metabolites. Adenosine 3‐thiotriphosphate, a nonhydrolyzable analog of ATP, induced similar changes in [Ca2+]i. This indicates that P2‐purinoceptors are present on osteoblast‐like cells and that extracellular ATP from various sources might be an important factor in the regulation of osteobl

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070504

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractDuring the process of endochondral bone formation, the maturing chondrocyte exhibits profound changes in energy metabolism. To explore the mechanism of energy conservation in cartilage we examined the expression of creatine kinase, an enzyme that catalyzes the formation of ATP in tissues under oxygen stress. Measurement of creatine kinase activity and cytochemical assessment of enzyme distribution clearly showed that the level of enzyme activity was related to chondrocyte maturation. Thus, as the cells hypertrophied, there was a progressive increase in creatine kinase activity. Similarly, an elevation in creatine kinase activity was noted in chondrocyte cultures as the cells assumed an hypertrophic state. When cartilage calcification was disturbed by rickets, there was a decrease in enzyme activity in the hypertrophic region. Studies were performed to examine the creatine kinase isozyme profile of cells of the epiphysis. In resting and proliferating cartilage, the isoform was MM. In hypertrophic cartilage, the predominant isoforms were MB and BB. In terms of the creatine phosphate content, the highest values were seen in the proliferative region; lower amounts were present in hypertrophic and resting cartilage; and no creatine phosphate was detected in calcified cartilage. These data suggest that turnover of creatine phosphate is greatest in the mineralized region of the epiphysis. The results of these investigations point to creatine kinase as being under developmental control. The activity of the enzyme in cartilage cells should serve as a marker of developmental events associated with chondrocyte proliferation, hypertrophy, and mineralization.

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070505

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractTo better understand the biochemistry of matrix‐forming cells, we developed a simple and reproducible procedure for the isolation and identification by N‐terminal sequencing of proteins secreted by cells into culture medium and applied this procedure to the analysis of the major Coomassie blue‐staining proteins under 100 kD that are secreted from three different human osteoblastic cell cultures. The major proteins secreted by normal human osteoblasts from adult trabecular bone were identified by N‐terminal sequencing to be gelatinase, osteonectin, the C‐terminal propeptides of the α1and α2chains of type I collagen, tissue inhibitor of metalloproteinase 1 (TIMP‐1), and β2‐microglobulin. The amounts of each of these proteins secreted into medium over a 24 h interval did not change over the 7 consecutive days of culture under serum‐free conditions, which indicates that this pattern of protein secretion is not significantly affected by the serum‐free conditions needed for protein identification by this method. In addition, radioimmunoassay for bone gla protein (BGP), a marker for osteoblast phenotype, revealed that BGP secretion remained high over 7 days of culture under serum‐free conditions and was comparable to the rate of BGP secretion in control cultures with 10% serum. The major proteins secreted by MG‐63 cells were identified by N‐terminal sequencing to be gelatinase, a novel 40 kD human bone protein we termed YKL‐40, TIMP‐1, the recently discovered TIMP‐2, and β2‐microglobulin. Further studies revealed that YKL‐40 is the only protein detectable by Coomassie staining of SDS gels of MG‐63 media proteins that is induced by extended time at confluence or by treatment with 1,25‐(OH)2D3. The apparent absence of detectable Coomassie‐stained bands corresponding to the C‐terminal propeptides of collagen in the medium of MG‐63 cells suggests that these transformed cells may not be a good model for bone matrix formation. The major proteins secreted by normal fetal osteoblastic cells were identified by N‐terminal sequencing to be osteonectin and the C‐terminal propeptides of the α1and α2chains of type I collagen. Gelatinase and TIMP could not be detected among the conditioned medium proteins by these methods. These observations indicate that fetal osteoblasts primarily express proteins that are matrix constituents and adult human osteoblasts secrete, in addition to t

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070506

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractThe retention of radioactivity in human, rat, and dog following a single injected dose of radiolabeled etidronate disodium (EHDP) is shown to follow power‐law decay curves with similar slopes for times up to 4, 60, and 80 days, respectively. During this period retention declines with time according to a weak inverse power of the time since dosing, with an exponent ranging from −0.05 (dog) to −0.09 (human and rat). Direct analyses of dog bones either 90 days after a single dose or 365 days after cessation of chronic dosing indicate a more rapid bone clearance of EHDP than predicted by the initial power law. Direct skeletal analysis also shows a more rapid loss of radioactivity in the rat between 60 and 365 days, indicative of either a second power law or a terminal exponential phase in the retention function occurring after 60 days. These data are used to estimate the minimum and maximum amounts of drug that would remain in the body following long‐term treatment in humans. For the intermittent cyclic EHDP treatment (ICT) regimen for osteoporosis (repeated cycles of 14 daily doses of 400 mg orally followed by 76 days drug free), the projected retention of EHDP after 3 years of treatment is 25–50 times the daily absorbed dose. Thus, for a 60 kg woman with a daily absorbed dose of 12 mg, the retained mass of EHDP would be about 300–600 mg. The surface area available for diphosphonate adsorption combined with measured bone‐remodeling rates in ICT‐treated subjects dictates that this mass would be distributed into a sufficiently large volume of mineral such that the fractional occupancy of active bone surfaces is only a few percent. The predicted retained mass of diphosphonate is well below that known to inhibit mineralization. This analysis is consistent with the clinical histopathology of bone from ICT‐treated subjects, which has indicated that no mineralization defect ensues from this treatment

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070507

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractThe hypophosphatemic (Hyp) mouse is the murine homolog for human hypophosphatemic vitamin D‐resistant rickets. We previously reported that bone cells isolated from normal andHypmice produced abnormal bone when transplanted intramuscularly into mutant mice. To assess the role of hypophosphatemia on bone formation in transplants, normal andHypmouse periostea were pair transplanted into control or phosphate (P)‐supplementedHypmice and into control or P‐deprived normal mice. The bone nodules formed in transplants after 2 weeks were characterized by measuring the thickness of the surrounding osteoid seams and the relative osteoid volume. P restriction in normal recipient mice impaired bone formation by transplanted normal cells and aggravated the defective bone formation byHypcells. The osteoid thickness and volume remained significantly higher inHyptransplants than in normal cotransplants, however. P supplementation ofHyprecipient mice normalized bone formation by transplanted normal cells but not byHypcells. However, a marked decrease in osteoid thickness and volume was observed inHyptransplants down to values observed in normal recipient mice. These results indicate that hypophosphatemia is not the only cause of abnormal bone formation in theHypmouse but that an osteoblast dysfunction contributes to the bone disease. These observations further support the concept that the osteoblast may be an important target for theHypmut

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070508

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractWe show that osteoclasts bind parathyroid hormone (PTH) in a manner that displays the properties of receptor‐dependent hormone binding, that is, saturability, time dependence, temperature dependence, and hormone specificity. Osteoclasts were isolated from the endosteum of 2 to 3 week chick tibiae and maintained in culture for 4–6 days. Bovine PTH‐(1–84) was biotinylated withN‐hydroxysuccinimidobiotin. Biotinyl‐PTH (btPTH, 10−5‐10−11M) was added to the cultured osteoclasts for 2–20 minutes. After rinsing away unbound btPTH, fluorescein isothiocyanate‐labeled avidin (FITC‐avidin) at a concentration of 66 μg/ml was applied. Receptor binding characteristics were assessed: (1) saturation occurred at around 10−6M btPTH; (2) competition of excess unlabeled PTH was found, namely, a 10‐fold excess abolished fluorescence; (3) specificity was shown by adding other polypeptide hormones (insulin, glucagon, and calcitonin) in 10‐ to 100‐fold excess—no effect on PTH binding was observed; and (4) affinity of btPTH for its binding site was indicated by half‐maximal binding ≅10−7M for both osteoclasts and osteoblasts. Biotin (10−5M) or FITC‐avidin (66 μg/ml) alone did not cause fluorescence. The time course of btPTH on the cell exterior was short: at 2 and 5 minutes dots of fluorescence were randomly dispersed over the cell surface, by 10 minutes most of the fluorescence was clustered in one region of the membrane, and by 20 minutes most of the hormone was no longer present on the surface of the cells. This sequence of events and the finding that maintaining the cells at 4°C blocked the clustering process indicates that occupied receptors were rapidly internalized by endocytosis. Fibroblasts and osteoblasts were more intensely stained by btPTH‐FITC‐avidin than osteoclasts, and no internalization of label was observed over the time period studied. This study shows direct, short‐term binding of PTH to osteoclasts through specific receptors. The binding of PTH to both osteoclasts and

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070509

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractTransforming growth factor β (TGF‐β) is now recognized as an important growth regulator and modulator in bone, where it apparently acts in an autocrine or paracrine fashion. In an effort to help elucidate how TGF‐β may interact with parathyroid hormone (PTH) to influence bone turnover, we examined the idea that TGF‐β might alter the number or affinity of PTH receptors in osteoblastic bone cells, PTH receptor binding was assessed in cultured ROS 17/2.8 cells using [125I]PTHrP‐(1–34) as labeled ligand. Specific binding to intact cells was measured in the presence of up to 1 μM unlabeled rPTH‐(1–34), and cAMP in cell extracts was determined by RIA. Incubation of ROS cells with 2 ng/ml of TGF‐β for the maximally effective time of 3 days increased the number of PTH binding sites (Bmax) by 47 ± 13%, with no change in theKp(3 nM). TGF‐β also increased the intracellular cAMP response to 0.3 nM rPTH‐(1–34) (ED50) by 53 ± 22%. Both effects were dose dependent, with 1–4 ng/ml of TGF‐β producing maximal effects, and both effects were blocked by the protein synthesis inhibitor cycloheximide (2–5 μM). Since TGF‐β induced comparable increases in both PTH binding and cAMP formation, the findings suggest that TGF‐β can increase the number of functional PTH receptors in cultured ROS 17/2.8 cells. This effect may reflect an action of TGF‐β to slow replication and

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070510

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY

摘要:

AbstractOur study investigated bone mineral density of the proximal femur and ultradistal and proximal radius in a population of elderly men and women. The Framingham study started in 1948, following a population‐based sample for evaluation of cardiovascular risk factors and events. During the 20th biennial Framingham examination (1988–89) we conducted the Framingham osteoporosis study, measuring bone mineral density in the proximal femur and distal and proximal radius for 1154 study participants. Ages ranged from 68 to 98 years, with a mean age of 76 years. Bone mineral density was measured using Lunar SP2 and DP3 densitometers. This cross‐sectional study evaluates mean bone mineral density measurements at each site by 5 year age intervals for men and women, testing for trends in bone density with age. Analyses were repeated adjusting for weight and height. Among the 446 and 708 women, bone mineral density of the femur and bone mineral content of the proximal radius were inversely and significantly related to age in both sexes and were considerably higher in men than women at all sites. The linear decline with age group in our cross‐sectional study remained after multivariate adjustment for height and weight. The ultradistal radius showed no significant correlation with age for either sex. There were significant correlations between the bone measurements made at different sites for both men and women (range inr= 0.27–0.89). Cross‐sectional curves of bone mineral density with age showed no significant differences in slope between males and females. In this cross‐sectional study, we found that increased age was significantly associated with decreased bone mass in a linear and equivalent fashion for both men and women through the elderly years in the proximal femur and pr

ISSN:0884-0431    

DOI:10.1002/jbmr.5650070511

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1992

数据来源: WILEY