摘要:

AbstractIn UMR‐106 osteosarcoma cells we found that PTH activated both the cAMP/protein kinase A and the Ca2+‐dependent phosphoinositide/protein kinase C (PKC) pathways, but prostaglandin E2(PGE2) activated only the cAMP pathway. Activation of PKC by the phorbol ester PMA had no effect on cAMP production but enhanced PTH‐stimulated cAMP production by 50% or more; the effect on PGE2‐induced cAMP was negligible. Inhibition of the α‐subunit of the inhibitory guanine nucleotide binding protein (Gi) by pertussis toxin pretreatment also enhanced PTH‐mediated cAMP production but had no effect on PGE2‐induced cAMP production. These results suggest that although PTH‐mediated adenylate cyclase activity is regulated via both the stimulatory (Gs) and inhibitory (Gi) guanine nucleotide binding proteins, only Gsregulates PGE2‐mediated adenylate cyclase activity in UMR‐106 cells. Costimulation with pertussis toxin and PMA did not increase PTH‐stimulated cAMP production above that obtained with PMA alone. This implies a similar target of action for pertussis toxin and PMA, that is, the α‐subunit of Gi. The α‐subunit of Giwas found to be a substrate for in vitro PKC phosphorylation of membrane fractions from UMR‐106 cells, seen as a ±40 kD band on SDS‐PAGE. Stimulation of in situ32P‐labeled cells with either PMA or PTH also enhanced incorporation of32P into the 40 kD band. Using the peptide antisera AS/7 and EC/2, we showed that pertussis toxin‐labeled subunits of both Gi1α/Gi2α and Gi3α could be immunoprecipitated, respectively, but immuinoprecipitation of membrane proteins after in situ phosphorylation and stimulation with PMA precipitated only Gi2α. We therefore conclude that modulation of adenylate cyclase activity by phorbol esters in UMR‐106 osteosarcoma cells can be ascribed, at least in part, to PKC‐mediated phosphorylation of the α‐subunit of the Gi2 compo

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071202

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 β (TGF‐β) has been proposed to have a role in bone remodeling by affecting the differentiation and activity of osteoblasts and osteoclasts and by inhibiting the production of proteinases, such as plasminogen activators (PAs). Studies on PAs have largely been based on data from nonhuman and fetal cell lines, however. The purpose of this study was to investigate the effect of TGF‐β on the PA activity of normal human osteoblast‐like cells and to compare this with its action on the human osteosarcoma cell line MG‐63. The action of interleukin‐1β (IL‐1β) was also assessed because it has been shown to increase PA activity in other connective tissue cell types. Normal osteoblast‐like cells had low to undetectable basal urokinase (uPA) and tissue plasminogen activator (tPA) activity, which was significantly stimulated by TGF‐β1. This action was shown to be dependent on transcription and new protein synthesis.TGY‐β2had a similar action. IL‐1β did not stimulate PA activity. In contrast, the MG‐63 cell line had high basal tPA and uPA activities. TGF‐β1decreased basal PA activity, the effect being most marked for uPA activity. IL‐1β stimulated uPA and tPA activity. TGF‐β1inhibited IL‐1β‐stimulated uPA activity, but the effect on tPA was more variable. This study has shown that TGF‐β has opposite effects on the PA activity of the two osteoblast‐like cell types studied. Care must therefore be used before extrapolating data from one cell type to another. It is suggested that under certain circumstances TGF‐β may be in

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071203

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:

AbstractInterunit variability among bone densitometers is due to different factors, including different calibration procedures and algorithms and variability in photon source energies and/or intensities. Other factors, such as the choice of scan parameters or the analysis procedures, can also introduce variability. The new generation of dual‐energy x‐ray absorptiometry (DXA) has partially improved this situation. The aim of this study was to investigate the operator‐dependent analysis procedures that can affect scan results and to evaluate the phantom and in vivo interunit variation of some DXA instruments. Four DXA instruments (QDR 1000 and 1000/W, Hologic, Inc.) were used. Potential sources of variability in the analysis procedures of anteroposterior lumbar spine and hip scans were considered: in most cases these procedures significantly influenced scan results. On lumbar spine, an enlargement of the scan window of less than 3 cm was responsible for an average increase in bone mineral density (BMD) of about 3%. On the hip, lowering the scan window by about 1 cm accounted for an increase in the whole‐segment BMD of about 4%. After standardization of analysis procedures, interunit and intraunit coefficients of variation and percentage differences among instruments were less than 1% for all the parameters considered (area and bone mineral content and density) with both an anatomic and a geometric phantom, and in nine subjects scanned by two different devices the percentage difference in BMD was greater than 2%. This study shows that present interunit variability allows comparisons among laboratories, but only if highly standardized analysis procedures a

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071204

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:

AbstractDual‐energy x‐ray absorptiometry and single‐photon absorptiometry were used to determine bone density at the lumbar spine and radial shaft in 62 patients with absorptive hypercalciuria, 27 patients with fasting hypercalciuria, and 31 nonhypercalciuric stone formers. Lumbar bone density was significantly lower in patients with absorptive (‐10%) as well as in those with fasting hypercalciuria (‐12%), with 74 and 92% of patients displaying values below the normal mean, whereas only 48% of the nonhypercalciuric stone formers had bone density values below the normal mean. In contrast, radial bone density was similar in all three groups of renal stone formers investigated. The comparison of urinary chemistry in patients with absorptive hypercalciuria and low normal bone density compared to those with high normal bone density showed a significantly increased 24 h urinary calcium excretion on random diet and a trend toward a higher 24 h urinary uric acid excretion and a higher body mass index in patients with low normal bone density. Moreover, among the patients with absorptive hypercalciuria we found a statistically significant correlation between the spinal bone density and the 24 h sodium and sulfate excretion and the urinary pH. These results gave evidence for an additional role of environmental factors (sodium and animal proteins) in the pathogenesis of bone loss in absorptive hypercalciuria. In conclusion, our data suggest an osteopenia of trabecular‐rich bone tissues in patients with fasting and absorptive hype

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071205

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 developed a sensitive and specific two‐site radioimmunoassay (IRMA) for human osteocalcin using human osteocalcin as a standard and two monoclonal antibodies raised against human osteocalcin purified from human cortical bone, a solid‐phase anti‐25–37 region and a tracer anti‐5–13 sequence of the molecule. A wide range of osteocalcin levels (up to 300 ng/ml) can be measured with a sensitivity of 0.4 ng/ml. The intra‐ and interassay coefficients of variation are less than 4 and 6%, respectively. The recovery of human osteocalcin from serum samples ranges from 96 to 103%. IRMA was linear for serial sample dilutions in a wide range of serum osteocalcin levels, even in patients with chronic renal failure on hemodialysis. Depletion of serum in intact osteocalcin demonstrated that IRMA detects, in addition to the intact peptide, a large N‐terminal midregion fragment that represents about 50% of total osteocalcin levels in normals and patients with Paget's disease and up to 75% in patients with chronic renal failure. This large fragment, previously unrecognized because it cannot be distinguished from intact osteocalcin with gel filtration chromatography, is not generated in vitro by incubation of the serum up to 26 h. We measured osteocalcin in the serum of 309 healthy adults (180 men and 129 women, age range 20–95 years), 36 patients with Paget's disease, 12 patients with primary hyperparathyroidism, 70 patients with chronic renal failure on hemodialysis, and 10 patients on corticosteroid therapy, simultaneously with human IRMA and with a conventional radioimmunoassay (RIA) based on bovine reagents. A tight correlation (r= 0.889) was observed between the two assays in the normal population, but the values obtained with IRMA were about threefold higher (mean 23.3 ± 10.5 versus 7.5 ± 3.4 ng/ml) than those obtained with RIA. Reported asZscores, that is, number of standard deviations from the predicted normal mean adjusted for sex and age, these two assays (IRMA and RIA) gave concordant results in patients with Paget's disease (4.05 ± 6.21 versus 2.41 ± 2.53), primary hyperparathyroidism (4.14 ± 7.17 versus 2.13 ± 2.28), chronic renal failure (25.32 ± 24.49 versus 6.93 ± 5.48), and glucocorticoid treatment (‐1.48 ± 0.78 versus −1.11 ± 0.57). However, IRMA was more discriminant from controls for all these metabolic bone diseases because the absolute values of meanZscores with IRMA were significantly higher than those obtained with the RIA (p<0.05–0.0001). We conclude that this new human‐specific IRMA of osteocalcin may be more sensitive than bovine RIA for the clinical inves

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071206

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:

AbstractBisphosphonates inhibit osteoclast‐mediated bone resorption, but their effects on the mechanical behavior of bone remain uncertain. This study investigated the effects of 4‐amino‐1‐hydroxybutylidene bisphosphonate (AHBuBP) on the biomechanical and morphologic properties of bone in ovariectomized rats. Sprague‐Dawley rats (four groups,n= 6) were ovariectomized at 3 months of age. From 7 to 13 months, the groups received vehicle or 0.28, 2.8, or 28 μg/kg of AHBuBP twice weekly through subcutaneous injection. An additional group of control animals (n= 6) received neither surgery nor drug. We determined the stiffness, yield, and ultimate loads of the femoral midshaft, the sixth lumbar (L6) vertebra, and the femoral neck. Geometric properties of the cortical bone were measured from digitized images of the tibial diaphysis at the level of the synostosis. The area fraction of trabecular bone was determined through the midsagittal plane of the fifth lumbar (L5) vertebra. There were no significant differences in the structural properties of the femoral neck and midshaft, with the exception that the medium‐dose group had a greater ultimate load than the vehicle group for the femoral midshaft in bending. Cross‐sectional analysis of the tibia did not show significant differences in the inertial properties or area. Ovariectomy caused a significant reduction in the stiffness and ultimate load of L6 and in the area fraction of trabecular bone of L5. Using the Tukey method of pairwise comparisons, the mean ultimate load ± standard deviation of the L6 vertebrae from the ovariectomized vehicle group was 30% lower than that from the nonovariectomized control group (276 ± 35 versus 396 ± 63 N,p<0.05). The mean stiffness was 46% lower (723 ± 206 versus 1327 ± 336 N/mm,p<0.05), and the area fraction was 41% lower (17.8 ± 5.2 versus 30.0 ± 7.9%,p<0.05). The high dose of AHBuBP maintained the structural properties of the vertebral bone; the ultimate load (424 ± 55 N), stiffness (1149 ± 145 N/mm), and area fraction (33.7 ± 5.5%) in the high‐dose group were all significantly greater than those in the vehicle group but did not differ significantly from those in the control group. We conclude that long‐term administration of AHBuBP in rats (started 4 months after ovariectomy) preserves the strength and area fraction of trabecular bone in the lumbar spine in a dose‐dependent manner, without evidence of detrimental effects upon the biomechanical or morpholog

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071207

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 effects of androgen and estrogen deficiency on skeletal homeostasis were studied in the guinea pig. Male and female adult (7 months old) guinea pigs were either sham operated (9 females and 7 males) or gonadec‐tomized [9 ovariectomized (OVX) females and 6 orchidectomized (ORX) males] and sacrificed 4 months later for evaluation of bone mass, bone turnover, and serum calcium homeostasis. Parameters of bone turnover, calcium homeostasis, and vitamin D metabolites were similar in all groups except for increased serum IGF‐I concentrations (+30%) in males compared to females. Gonadectomy resulted in a 50% decrease in serum IGF‐I concentrations in males only (p<0.001). Volume, total calcium content, and cortical density of the tibia were significant higher in males than in females. Estrogen deficiency had no effect on bone volume or calcium content. Androgen deficiency resulted in a significant lower volume and calcium content of the tibia and in a lower calcium content of the distal lumbar vertebrae. Single‐photon absorptiometry of the tibia showed that only cortical, not trabecular bone density of the tibia was decreased after ORX. Histomorphometric studies of the tibial metaphysis also did not show significant differences in trabecular bone volume between sham‐operated and ORX males. We conclude that in adult male guinea pigs androgen deficiency results in a decrease in (cortical) bone volume and content concomitant with decreased IGF‐I levels. In female guinea pigs of the same age, estrogen deficiency did not affect total or regiona

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071208

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:

AbstractOrthogonal intercepts from random test lines (OIr), uniformly distributed intercepts (OIu), and areas and lengths (Ar/Le) were measured manually and by computer to determine individual profile and sample widths of artificial profiles and human osteoid seams. Individual widths were equal by Ar/Le and OIu methods. The means of individual profile widths (all methods), of all orthogonal intercepts (OIu and OIr), or of profile widths weighted in proportion to their lengths (Ar/Le) were also equal. Ar/Le and OIu had smaller variance than OIr. Discrepant individual OIu and Ar/Le widths in digitized images were corrected by thresholding and did not significantly affect sample means. Unweighted Ar/Le sample means were 15–44% lower than weighted means. Distributions of osteoid seam widths were not normal, and all but one had more than one mode. We conclude that the Ar/Le method is comparable to direct orthogonal intercept methods and suitable for automated histomorphometr

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071209

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:

AbstractBone mineral density (BMD) and clinical status of 42 patients treated surgically 10–11 years earlier for an acute knee ligament injury were determined. The BMD was measured at the spine (L2–4) and the femoral neck, distal femur, patella, proximal tibia, and calcaneus of both lower extremities using dual‐energy x‐ray absorptiometric (DXA) scanner. The relative BMD results of the injured knee were correlated with sex, age, activity level, knee stability, and functional scores of the patient. In the 11 patients with moderate injury (isolated rupture of the medial collateral ligament), the BMDs of the injured and uninjured extremities were equal. In the 31 patients with severe injury (cruciate ligament rupture), the BMDs were significantly lower in the injured knee: distal femur, −6.0% (p= 0.0000); patella, −9.0% (p= 0.0000); and proximal tibia, −3.3% (p= 0.0012). Neither the femoral neck nor the calcaneus showed any differences. There were no significant differences either between men and women or between patients with different activity levels. The relative BMDs of the injured knee did not correlate with age or static knee stability but correlated significantly (r= 0.42–0.78,p<0.01–0.001) with the functional scores of the same knee: the better the knee function in comparison with the healthy knee, the higher the relative BMD. The spinal BMDs corresponded with the age‐adjusted reference values of the used densitometry. In conclusion, a severe knee ligament injury results in permanently decreased BMD in the injured knee. Other parts of the same extremity and lumbar spine are not affected. The observed decrease (3–9%) is clinically important with respect to age‐related bone loss of 1%/year after the age of 35 years. Additional follow‐up is needed to determine any increased risk of o

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071210

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 report that neural cell adhesion molecules (NCAM) are expressed transiently in developing chicken osteoblasts during osteogenesis using immunostaining on cryostat sections. NCAM is strongly expressed in most osteoblasts along bone trabeculae that coincide with the presence of collagen I and alkaline phosphatase activity. In endochondral ossification, NCAM is highly expressed in osteogenic buds as seen in the epiphysis and diaphysis of tibia and vertebrae. In intramembranous ossification, NCAM is seen in osteogenic condensation of calvaria and in the periosteum of tibial diaphysis. The expression is transient because NCAM is not expressed in mesenchymal cells before osteogenic condensation and NCAM expression is lost in osteocytes in later stages. The staining pattern suggests that NCAM is present on the cell membrane of osteoblasts. Using a specific monoclonal antibody, the osteoblast NCAM is shown to contain polysialic acid, which is enriched in embryonic brain. Northern blot analysis using chicken brain NCAM cDNA as probes showed two major sizes of mRNA at 6.4 and 4.2 kb in calvarial mRNA as opposed to bands at 7.2, 6.4, and 4.2 kb in the brain. An immunoblot showed major proteins at Mr 165 and 110 kd, unlike brain NCAM, which are 180, 140, and 120 kD. That NCAM is involved in bone morphogenesis is consistent with the general hypothesis that NCAM plays pivotal roles in mesenchymal condensation, as shown in the formation of muscle, kidney, skin, and cartilage. The results establish NCAM as a cell surface molecule expressed transiently during osteoblast lineage. The implication that NCAM may mediate osteoblast interaction and regulate skeletal morphogenesis is discussed.

ISSN:0884-0431    

DOI:10.1002/jbmr.5650071211

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