bluuebunny/crossref_metadata_2025_split huggingface dataset:
Growth hormone-dependent insulin-like growth factor binding protein is a major determinant of bone mineral density in healthy men
Authors: Johansson Anna G. , Forslund Anders , Hambraeus Leif , Blum Werner F. , Ljunghall Sverker 
Published: February 2010

Abstract
To establish the major determinants of bone mass, we assessed relationships between bone mineral density (BMD) and height, weight, body mass index (BMI), muscle strength, physical capacity (Vo2max), body composition, serum concentrations of insulin-like growth factor I (IGF-I), growth hormone (GH), the GH-dependent IGF binding protein (IGFBP-3), testosterone, sex hormone binding globulin (SHBG), osteocalcin, and parathyroid hormone (PTH) in 38 healthy men between 25 and 59 years of age. Values of BMD at all sites (total body, lumbar spine, and hip) were strongly correlated with IGFBP-3 (r = 0.51–0.64, p &lt; 0.001 at all sites), and total-body BMD was also significantly correlated with IGF-I (r = 0.43, p = 0.01). BMD measurements of the total body and of the different sites of the hip were negatively correlated with age and positively with weight, BMI, muscle strength, Vo2max, and fat-free weight. IGF-I and IGFBP-3 were both positively related to muscle strength and Vo2max. In a stepwise forward multiple-regression analysis, the best model was obtained for the femoral neck, where IGFBP-3, GH, PTH, age, IGF-I, and BMI explained 77% of the variation in BMD. The partial regression coefficients of IGFBP-3, PTH, and BMI were all positive, whereas age, GH, and IGF-I were negatively correlated with BMD. In summary, IGFBP-3 correlated better with BMD than any other study parameter. The findings indicate that GH is of importance for bone mass and suggest that IGFBP-3 not only reflects the integrated GH secretion but also has a direct role in the endocrine regulation of bone metabolism.
Mutation analysis of coding sequences for type I procollagen in individuals with low bone density
Authors: Spotila Loretta D. , Colige Alain , Sereda Larisa , Constantinou-Deltas Constantinos D. , Whyte Michael P. , Riggs Lawrence B. , Shaker Joseph L. , Spector Timothy D. , Hume Eric , Olsen Nancy , Attie Maurice , Tenenhouse Alan , Shane Elizabeth , Briney Walter , Prockop Darwin J. 
Published: February 2010

Abstract
Mutations in one of the two genes encoding type I procollagen (COL1A1 and COL1A2) are frequently the cause of osteogenesis imperfecta (OI), a disorder characterized by brittle bones. Here we tested whether patients with low bone density also have mutations in these genes. The 26 patients studied had no apparent metabolic bone disease, but most had a positive family history of osteopenia or osteoporosis. Although a diagnosis of OI was considered by the clinician in some cases, the clinical criteria for OI were not satisfied. Our strategy for mutation analysis consisted of PCR amplification of cDNA made to fibroblast mRNA using primers specific for the coding regions of COL1A1 and COL1A2. The PCR products were then sequenced directly with primers located within each PCR product. We found that 3 of 26 patients had mutations that altered the encoded amino acid. One mutation, at position α2(I)-661 has been reported (Spotila et al. 1991 Proc Natl Acad Sci USA PNAS 88: 5423). The other 2 patients, who were not related to each other, had a mutation that altered the proline codon at α1(I)-27 to alanine. This mutation was not found in 81 normal individuals or in 37 additional osteopenic individuals. However, its effect on the biologic function of type I collagen, as well as its role in osteopenia, is uncertain. In addition to the two mutations, we found a polymorphism in codon α2(I)-459. Although this polymorphism involved an amino acid substitution, it was present with equal frequency in the patient and the normal population. By analyzing this and previously reported neutral sequence variants in the COL1A2 gene, we determined that all patients expressed both alleles of the COL1A2 gene. The 12 patients who were heterozygous for a COL1A1 neutral sequence variant also expressed both alleles. Here we present all PCR primer and sequencing primer information. The results suggest that surveying a larger group of similarly selected individuals may reveal additional mutations in the COL1A1 or COL1A2 genes.
Stimulation of signal transduction pathways in osteoblasts by mechanical strain potentiated by parathyroid hormone
Authors: Carvalho Roberto S. , Scott J. Elliot , Suga Dolores M. , DR. Yen Edwin H. K. 
Published: February 2010

AbstractSecond-messenger systems have been implicated to transmit mechanical stimulation into cellular signals; however, there is no information on how mechanical stimulation is affected by such systemic factors as parathyroid hormone (PTH). Regulation of adenylyl cyclase and phosphatidylinositol pathways in rat dentoalveolar bone cells by mechanical strain and PTH was investigated. Two different cell populations were isolated after sequential enzyme digestions from dentoalveolar bone (group I and group II) to study potential differences in response. Mechanical strain was applied with 20 kPa of vacuum intermittently at 0.05 Hz for periods of 0.5, 1, 5, 10, and 30 minutes and 1, 3, and 7 days using the Flexercell system. Levels of cAMP, measured by RIA, and levels of inositol 1,4,5-triphosphate (IP3) and protein kinase C activity (PKC), measured by assay systems, increased with mechanical strain. When PTH was added to the cells, there was a significant increase in levels of all the intracellular signals, which appeared to potentiate the response to mechanical strain. IP3 levels (0.5 minute) peaked before those of PKC activity (5 minutes), which in turn peaked before those of cAMP (10 minutes). Group II cells showed higher levels of cAMP and IP3 than the group I cells. This suggests that the former may ultimately play the predominant roles in skeletal remodeling in response to strain. Immunolocalization of the cytoskeleton proteins vimentin and α-actinin, focal contact protein vinculin, and PKC showed a marked difference between strained and nonstrained cells. However, the addition of PTH did not cause any significant effect in cytoskeleton reorganization. Staining of PKC and vimentin, α-actinin, and vinculin suggests that PKC participates actively in the transduction of mechanical signals to the cell through focal adhesions and the cytoskeleton, although only PKC seemed to change with short time periods of strain. In conclusion, dentoalveolar osteoblasts responded to mechanical strain initially through increases in levels of IP3, PKC activity, and later cAMP, and this response was potentiated when PTH was applied together with mechanical strain.
Integrins and osteoclastic resorption in three bone organ cultures: Differential sensitivity to synthetic arg-gly-asp peptides during osteoclast formation
Authors: van der Pluijm Gabri , Mouthaan Henk , Baas Corafien , de Groot Henny , Papapoulos Socrates , Löwik Clemens 
Published: February 2010

Abstract
We investigated possible inhibitory effects of five synthetic Arg-Gly-Asp (RGD)-containing peptides on osteoclastic resorption in three distinct in vitro resorption assays (17-day-old fetal mouse bone organ cultures) that differ in stages of osteoclast differentiation. RGD peptides, which can bind the adhesion receptors called integrins, inhibited osteoclastic resorption (45Ca release) in fetal mouse bone explants in which osteoclast precursors have yet to adhere to the mineralized matrix and develop into mature osteoclasts (metacarpals and coculture system). Treatment of metacarpals with RGD peptides inhibited the formation of multinucleated TRAP+ osteoclasts in the mineralized matrix because their mononuclear TRAP+ osteoclast precursors remained localized in the periosteum. In particular, echistatin, a viper venom protein with known affinity for αvβ3 integrin, and GdRGDSP inhibited osteoclastic resorption dose dependently in these systems (ED50 10−9 and 10−4 M, respectively) but did not alter the activity of mature resorbing osteoclasts in radii. In addition, 45Ca release was significantly inhibited by the cyclic peptide GPenGRGDSPCA, which has a relatively higher affinity for the vitronectin than fibronectin receptor(s). In contrast, GRDGdSP, which has a much higher affinity for the fibronectin receptor (than the vitronectin receptors), had no effect on resorption at similar concentrations in any resorption system used. In summary, the data presented in this paper show that peptides with RGD motifs are capable of inhibiting osteoclastic resorption in bone organ cultures. Our studies not only support the hypothesis concerning the importance of αvβ3 in osteoclastic resorption but also suggest an important role of integrin(s) in events preceding the actual resorption of calcified matrix by osteoclasts.
Functional properties of a synthetic chicken parathyroid hormone-related protein 1–36 fragment
Authors: Schermer D.T. , Bradley M.S. , Bambino T.H. , Nissenson R.A. , Strewler G.J. 
Published: February 2010

Abstract
The biologic activities of human parathyroid hormone-related protein [hPTHrP(1–34] and bovine PTH [bPTH(1-34)] are remarkably similar despite marked sequence divergence in their primary binding domain, residues 25–34. Chicken PTHrP (cPTHrP) is identical to hPTHrP through residue 21. However, in the 25–34 region, cPTHrP displays three fewer basic residues than hPTHrP and contains five residues not present in any other member of the PTH/PTHrP family. To assess the biologic consequences of these structural differences, we compared the activities of synthetic [36Tyr]cPTHrP(1–36)NH2 and hPTHrP(1–34)NH2 with those of bPTH(1–34) in avian systems (chicken renal plasma membranes and 19 day chick embryonic bone cells) and mammalian systems [canine renal plasma membranes and rat osteosarcoma cells (UMR-106-H5)]. In both avian and mammalian systems the binding affinity of [36Tyr]cPTHrP(1–36)NH2 (0.8–3.4 nM) was approximately one-half that of hPTHrP(1–34)NH2 (0.4–1.1 nM). The potencies of [36Tyr]cPTHrP(1–36)NH2 and hPTHrP(1–34)NH2 for activation of adenylate cyclase were similar in canine renal membranes (5.2 and 6.7 nM) and chick bone cells (1.0 nM). In UMR-106 cells and chicken renal membranes the potency of [36Tyr[cPTHrP(1–36)NH2 for activation of adenylate cyclase was about one-half that of [36Tyr]hPTHrP(1–36)NH2. Binding of 125I-[36Tyr]cPTHrP(1–36)NH2 to chick bone cells and chicken renal membranes was completely displaced by bPTH(1–34) and hPTHrP(1–34)NH2: thus there was no evidence for a distinct chicken PTHrP receptor. In general, [36Tyr]cPTHrP(1–36)NH2 and hPTHrP(1–34)NH2 activated adenylate cyclase similarly despite their sequence differences in the 25–32 region. This suggests that basic residues in the 25–32 region are not required for the peptide to assume a biologically active conformation at the receptor. In cross-linking studies, both 125I-hPTHrP(1–34)NH2 and 125I-[36Tyr]cPTHrP(1–36)NH2 labeled a major 85 kD PTH/PTHrP receptor component in canine renal plasma membranes. 125I-hPTHrP(1–34)NH2 also labeled a ≤14 kD receptor fragment, whereas 125I-[36Tyr]cPTHrP(1–36)NH2 did not. The present results suggest that retained sequence features in the 25–32 region may be critical determinants of receptor binding and that sequence differences in this region alter the sites of interaction of PTHrP peptides with the PTH/PTHrP receptor.
Automated evaluation of hip axis length for predicting hip fracture
Authors: Faulkner Kenneth G. , Mcclung Michael , Cummings Steven R. 
Published: February 2010

Abstract
The hip axis length has been shown in previous studies to be predictive of hip fracture independent of age and femoral bone density. The first studies of hip axis length were performed by manual measurement of dual x-ray absorptiometry (DXA) scan printouts. In this study, an automated analysis procedure is defined using software tools provided by the DXA manufacturer. Manual and automatic hip axis length measurements in 198 women were highly correlated (r = 0.98). Because of scaling factors of the printout, the automatic measurement was 58% longer than the manual value. Precision of the automatic measurement, based upon triplicate DXA scans of 33 women, was 0.07 cm or 0.68%. To define normative data, the hip axis length was measured from femoral DXA scans of 471 female volunteers aged 40–92 scanned on 14 different Hologic QDR-1000 systems. Mean hip axis length was 10.5 cm, with a standard deviation of 0.62 cm. No significant relationship between hip axis length and age was found (r = 0.07, P = 0.15). Based on previously reported odds ratios corrected for femoral bone density, age, height, and weight, an automatic hip axis length measurement of 11.0 cm is associated with a twofold increase in hip fracture risk compared with a woman with an average hip axis length. A hip axis length value of 11.6 cm increases hip fracture risk by a factor of 4 compared with a woman with a normal hip dimension. We conclude that the hip axis length can be easily incorporated into existing DXA hip analysis software in combination with a bone density measurement. With further research, the hip axis length may become a clinically useful tool for the evaluation of hip fracture risk.
Carbonic anhydrase II mRNA expression in individual osteoclasts under “resorbing” and “nonresorbing” conditions
Authors: Asotra S. , Gupta A.K. , Sodek J. , Aubin J.E. , Heersche J. N. M. 
Published: February 2010

Abstract
Rabbit osteoclasts can be transformed from a nonresorbing state to a resorbing state by transferring them from culture medium at pH 7.5 to one at pH 6.5. We evaluated whether expression of mRNA for carbonic anhydrase (CA-II) could be used as an indicator of the state of activity of individual osteoclasts. A cDNA probe to rabbit carbonic anhydrase II (CA-II) was prepared and used for in situ hybridization analysis of osteoclasts isolated from neonatal rabbit long bones. Quantitation by grain counting revealed heterogeneity within the osteoclast population: osteoclasts with a “compact” (rounded, less spread) morphology expressed higher levels of CA-II mRNA than “spread” osteoclasts with similar numbers of nuclei. When maintained at pH 6.5 for 6 h, the level of CA-II mRNA was increased significantly in osteoclasts of both morphologies compared with those in parallel cultures maintained at pH 7.5. These results were confirmed by quantitating CA-II mRNA using the polymerase chain reaction (PCR). Oligonucleotide primers specific for rabbit CA-II were synthesized and used to amplify CA-II cDNA transcribed from mRNA prepared from single or small numbers (one to eight cells) of osteoclasts that were collected with a micromanipulator. This generated a ∼510 bp PCR product, corresponding to the predicted size of the CA-II fragment encompassed by the primers. For quantitation, CA-II mRNA levels were compared with the levels of a ∼900 bp actin fragment that was coamplified in the same reaction mixture or amplified separately in a duplicate sample of the reaction mixture. The ratio of CA-II mRNA expression to actin mRNA expression was significantly increased in osteoclasts cultured at pH 6.5 for 6 h compared with osteoclasts maintained at pH 7.5 (1.89 + 0.12 versus 0.98 + 0.06, n = 39, mean + SEM, of all assays combined; P &lt; 0.001). Our results demonstrate that CA-II mRNA expression is upregulated in osteoclasts in the resorptive state. The methods used provide a novel molecular approach for analyzing osteoclast activity with assays that are applicable to single cells and obviate the problem of osteoclast impurity, allowing investigation of osteoclast heterogeneity.
Periosteal bone formation stimulated by externally induced bending strains
Authors: Raab-Cullen D.M. , Akhter M.P. , Kimmel D.B. , Recker R.R. 
Published: February 2010

Abstract
The rat tibia four-point bending model is a new mechanical loading model in which force is applied through external pads to the rat lower limb. The advantages of the model are controlled force application to a well-defined bone, noninvasive external loading, and the addition of loads to normal daily activity. A disadvantage of the model is that the pads create local pressure on the leg at the contact sites. This study examined the differences in tibial response to bending strains and to local pressure under the pads. A total of 30 adult Sprague-Dawley rats were randomized into three external loading groups: bending, cyclic pressure, and static pressure. The right leg of each rat was externally loaded to create either bending or local pressure without bending; the left leg served as a control. Strains on the lateral surface averaged 1200 μϵ in compression during bending load application and &lt;200 μϵ in compression during pressure loading. Histomorphometric data were collected from three regions: the maximal bending region, under the loading pads, and outside the maximal bending region. In the maximal bending region, bending loads created greater mineral apposition rate (MAR) on the lateral surface and greater MAR and formation surface on the medial surface of loaded than control tibiae. The region under the bending pad was exposed to similar bending strains and showed the same pattern of increased MAR as sections from the maximal bending region. Cyclic pressure had no effect on periosteal MAR or formation surface. Static pressure increased MAR only on the lateral tibial surface. Bending stimulates bone formation in regions with the highest bending strains. Similar forces applied only in the form of pressure loading do not stimulate tibial formation either at the contact site or between loading pads. These results suggest that externally applied forces of moderate magnitude stimulate bone formation primarily as a result of increased bending strains, not local pressure at the contact site.
Tartrate-resistant acid phosphatase accumulated in the matrix of developing medullary bone induced by estrogen treatment of male Japanese quail
Authors: DR. Yamamoto Toshio , Nagai Hiroshi 
Published: February 2010

Abstract
Tartrate-resistant acid phosphatase (TRAP) in medullary bone matrix and bone cells was histochemically examined employing estrogen-induced medullary bone of male Japanese quail. Within 2 days after estrogen administration, medullary bone matrix was identified and the matrix and osteoblasts were negative for TRAP activity. Osteoclasts showing TRAP activity were occasionally seen. By 4 days, the projection of medullary bone trabeculae was seen and TRAP became positive in the deeper part of the bone matrix of trabeculae as well as the matrix located near cortical bone, but the surface areas and extending margins of medullary bone trabeculae were negative in activity. Osteoblasts were also negative for TRAP activity. By 7 days, TRAP-positive bone matrix conspicuously increased accompanying increment of bone volume, but TRAP-negative bone matrix was still seen at the surface areas or extending areas of the medullary bone trabeculae. Osteoblasts showed no reaction product. Osteoclasts showing TRAP activity increased in number. The results demonstrate that TRAP accumulates in medullary bone matrix with bone formation. Circumstantial evidence suggests that medullary bone osteoblasts may be involved in TRAP accumulation in the matrix despite their negative activity.
Influence of osteoclasts and osteoclast-like cells on osteoblast alkaline phosphatase activity and collagen synthesis
Authors: Galvin Rachelle J. Sells , Cullison James W. , Avioli Louis V. , Osdoby Philip A. 
Published: February 2010

Abstract
Osteoblasts have been shown to modulate osteoclast activity, but the reverse process has not been investigated. In the current study conditioned medium (CM) was collected from osteoclasts and osteoclast-like cells and its effects on osteoblast alkaline phosphatase (ALPase) activity and collagen synthesis ([3H]proline hydroxylation) were determined. In primary chick osteoblasts, cultured chick embryo frontal bones, and UMR-106-01 cells, collagen synthesis and ALPase activity, but not [3H]thymidine incorporation, were inhibited by CM from chick marrow-derived giant cells, which possess some of the phenotypic characteristics of osteoclasts. However, collagen synthesis in chick embryo fibroblasts was not affected by giant cell CM. CM collected from cultures of chicken osteoclasts and human osteoclastoma cells and marrow-derived giant cells inhibited collagen synthesis in UMR-106-01 cells, but the effects on ALPase activity varied with the cell type. In contrast, mononuclear cell and fibroblast CM did not alter collagen synthesis. Initial characterization studies demonstrate that the inhibitor is a heat-labile factor with a molecular weight greater than 3500. In summary, authentic osteoclasts, tumor osteoclast-like cells, and chicken and human multinucleated giant cells produce a soluble factor that alters osteoblast collagen synthesis, suggesting that osteoclasts play a role in the modulation of osteoblast activity.
A number of osteocalcin antisera recognize epitopes on proteins other than osteocalcin in cultured skin fibroblasts: Implications for the identification of cells of the osteoblastic lineage in vitro
Authors: Bradbeer J.N. , Virdi A.S. , Serre C.M. , Beresford J.N. , Delmas P.D. , Reeve J. , Triffitt J.T. 
Published: February 2010

Abstract
Rabbit antisera to bovine osteocalcin were produced independently in two laboratories and their specificities established by western blot analysis. By immunohistochemistry each of the five polyclonal antisera produced an intense cytoplasmic staining in human bone-derived cells. Staining intensity was strongly attenuated by preabsorption of the antisera with osteocalcin. No staining was observed using nonimmune rabbit serum. However, the choice of skin cells as negative controls for osteocalcin synthesis yielded an unexpected positive staining pattern similar to that seen with the bone-derived cells over a range of antiserum dilutions. This was not caused by the uptake of exogenous osteocalcin from the culture medium because a similar pattern of staining was observed when medium was supplemented with osteocalcin-depleted fetal calf serum. Treatment with 1,25-dihydroxyvitamin D3 induced osteocalcin mRNA expression and osteocalcin secretion in cultures of bone-derived cells but not in skin fibroblasts. The results demonstrate that these polyclonal antisera also recognize epitopes shared with other proteins synthesized in culture by skin fibroblasts. Furthermore, three mouse monoclonal antibodies to distinct regions of the osteocalcin molecule show differential staining of human bone-derived cells, skin cells, and osteosarcoma cells (MG63). These observations indicate that the shared epitope resides in the central region of osteocalcin and are consistent with the specific synthesis of osteocalcin by bone cells alone. The observed nonspecificity of many osteocalcin antisera may compromise immunocytochemical studies of the osteoblast phenotype in studies in vitro when based solely on reactivity with inadequately characterized osteocalcin antisera.