bluuebunny/crossref_metadata_2025_split huggingface dataset:
Localization and Regulation of the Growth Hormone Receptor and Growth Hormone-Binding Protein in the Rat Growth Plate
Authors: Gevers Evelien F. , Van Der Eerden Bram C. J. , Karperien Marcel , Raap Anton K. , Robinson Iain C. A. F. , Wit Jan-Maarten 
Published: April 2006

Abstract
Growth hormone (GH) has direct effects on the growth plate to stimulate longitudinal growth, but it is not clear which chondrocyte populations GH acts on. The dual effector theory suggests that GH would act primarily on the “stem cells.” However, staining with a GH receptor (GHR) antibody is found in all layers of the growth plate in rabbits and humans. We now have investigated the localization and regulation of GHR and the related GH binding protein (GHBP) in the rat growth plate using a sensitive immunohistochemical method involving tyramide signal amplification (TSA) and antibodies specific for GHR or GHBP. Both GHR and GHBP were shown in the germinal and proliferative chondrocytes, but most clearly in early maturing chondrocytes at the interface between proliferative and hypertrophic cells. Staining for GHR and GHBP was located in both the cytoplasm and the nucleus. Expression of GHR mRNA and GHBP mRNA in the growth plate was confirmed by reverse-transcription polymerase chain reaction (RT-PCR). Immunohistochemical staining for GHR and GHBP decreased with age; in 12-week-old normal rats, only the early maturing chondrocytes were stained. In GH-deficient dwarf rats, staining seemed less than in normal rats, and in hypophysectomized (Hx) rats, staining for GHBP was clearly reduced. Treatment of Hx rats with thyroid hormones (T3 + T4), via subcutaneously (sc) implanted osmotic minipumps, induced little growth and induced a small layer of GHR-positive and GHBP-positive early maturing chondrocytes. Treatment with GH and thyroid hormones (TH) resulted in greater growth and a broader layer of GHR-positive and GHBP-positive cells, indistinguishable from normal rats. In contrast, dexamethasone treatment of normal rats inhibited their growth and reduced GHR and GHBP staining in the growth plate. These results show that GHR and GHBP in the growth plate are under hormonal control. The localization of GHR/GHBP suggests that in addition to actions on germinal and proliferative cells in young rats, GH also has effects on early maturing chondrocytes and may be involved in their differentiation to a fully hypertrophic chondrocyte.
Bone Morphogenetic Protein 2 Induces Dental Follicle Cells to Differentiate Toward a Cementoblast/Osteoblast Phenotype
Authors: Zhao Ming , Xiao Guozhi , Berry Janice E. , Franceschi Renny T. , Reddi Anand , Somerman Martha J. 
Published: April 2006

Abstract
When triggered appropriately, dental follicle cells are considered to be able to differentiate toward a cementoblast/osteoblast phenotype. However, factors and mechanisms regulating follicle cell differentiation remain undefined. This study focused on determining the ability of bone morphogenetic protein (BMP) 2 to promote the differentiation of follicle cells and periodontal ligament (PDL) cells along a cementoblast/osteoblast pathway. Follicle cells and PDL cells were isolated from the first molar region of CD-1 mice and immortalized with SV40. Both cell types expressed BMP-4 and BMP receptors (BMPR) IA and II, but only follicle cells expressed BMP-2 mRNA. Cells were exposed to recombinant human BMP (rhBMP)-2 (0–100 ng/ml) and Northern blots were used to determine the expression of mineral-associated markers. BMP-2, in a dose- and time-dependent manner, induced cementoblast/osteoblast differentiation of follicle cells, as reflected by enhanced core binding factor α1 (Cbfa1), bone sialoprotein (BSP), and osteocalcin (OCN) mRNA expression and enhanced mineral formation. U0126, a specific inhibitor of MEK-1/2 members of the MAPK family, abolished BMP-2-mediated expression of BSP and OCN. In contrast, exposure of PDL cells to BMP-2 resulted in modest expression of OCN and minimal promotion of mineralization. These results suggest that BMP-2 triggers follicle cells to differentiate toward a cementoblast/osteoblast phenotype and that the MAPK pathway is involved.
Activation of Extracellular Signal–Regulated Kinase Is Involved in Mechanical Strain Inhibition of RANKL Expression in Bone Stromal Cells
Authors: Rubin Janet , Murphy Tamara C. , Fan Xian , Goldschmidt Mark , Taylor W. Robert 
Published: April 2006

Abstract
Mechanical input is known to regulate skeletal mass. In vitro, application of strain inhibits osteoclast formation by decreasing expression of the ligand RANKL in bone stromal cells, but the mechanism responsible for this down-regulation is unknown. In experiments here, application of 1.8% equibiaxial strain for 6 h reduced vitamin D-stimulated RANKL mRNA expression by nearly one-half in primary bone stromal cells. Application of strain caused a rapid activation of ERK1/2, which returned to baseline by 60 minutes. Adding the ERK1/2 inhibitor PD98059 30 minutes before strain delivery prevented the strain effect on RANKL mRNA expression, suggesting that activation of ERK1/2 was required for transduction of the mechanical force. Mechanical strain also activated N-terminal Jun kinase (JNK) that, in contrast, did not return to baseline during 24 h of continuous strain. This suggests that JNK may represent an accessory pathway for mechanical transduction in bone cells. Our data indicate that strain modulation of RANKL expression involves activation of MAPK pathways.
Bioavailability and Biological Efficacy of a New Oral Formulation of Salmon Calcitonin in Healthy Volunteers
Authors: Buclin Thierry , Rochat Monica Cosma , Burckhardt Peter , Azria Moïse , Attinger Martine 
Published: April 2006

Abstract
Salmon calcitonin (SCT) is a well-tolerated peptide drug with a wide therapeutic margin and is administered parenterally for long-term treatments of bone diseases. Its clinical usefulness would be enhanced by the development of an orally active formulation. In this randomized crossover double-blinded phase I trial, controlled by both a placebo and a parenteral verum, we have tested a new oral formulation of SCT associated with a caprylic acid derivative as carrier. Eight healthy volunteers received single doses of 400, 800, and 1200 μg of SCT orally, a placebo, and a 10-μg (50 IU) SCT intravenous infusion. SCT was reliably absorbed from the oral formulation, with an absolute bioavailability of 0.5–1.4%, depending on the dose. It induced a marked, dose-dependent drop in blood and urine C-terminal telopeptide of type I collagen (CTX), a sensitive and specific bone resorption marker, with the effects of 1200 μg exceeding those of 10 μg intravenously. It also decreased blood calcium and phosphate, and increased the circulating levels of parathyroid hormone (PTH) and, transiently, the urinary excretion of calcium. It was well-tolerated, with some subjects presenting mild and transient nausea, abdominal cramps, diarrheic stools, and headaches. This study shows that oral delivery of SCT is feasible with reproducible absorption and systemic biological efficacy. Such an oral formulation could facilitate the use of SCT in the treatment of osteoporosis and other bone diseases.
Alleles of RUNX2/CBFA1 Gene Are Associated With Differences in Bone Mineral Density and Risk of Fracture
Authors: Vaughan Tanya , Pasco Julie A. , Kotowicz Mark A. , Nicholson Geoff C. , Morrison Nigel A. 
Published: April 2006

Abstract
The aim of this study was to determine if DNA polymorphism within runt-related gene 2 (RUNX2)/core binding factor A1 (CBFA1) is related to bone mineral density (BMD). RUNX2 contains a glutamine-alanine repeat where mutations causing cleidocranial dysplasia (CCD) have been observed. Two common variants were detected within the alanine repeat: an 18-bp deletion and a synonymous alanine codon polymorphism with alleles GCA and GCG (noted as A and G alleles, respectively). In addition, rare mutations that may be related to low BMD were observed within the glutamine repeat. In 495 randomly selected women of the Geelong Osteoporosis Study (GOS), the A allele was associated with higher BMD at all sites tested. The effect was maximal at the ultradistal (UD) radius (p = 0.001). In a separate fracture study, the A allele was significantly protective against Colles' fracture in elderly women but not spine and hip fracture. The A allele was associated with increased BMD and was protective against a common form of osteoporotic fracture, suggesting that RUNX2 variants may be related to genetic effects on BMD and osteoporosis.
Two Polymorphisms in the Vitamin D Receptor Gene—Association With Bone Mass and 5-Year Change in Bone Mass With or Without Hormone-Replacement Therapy in Postmenopausal Women: The Danish Osteoporosis Prevention Study
Authors: Tofteng C. L. , Jensen J. E. B. , Abrahamsen B. , Odum L. , Brot C. 
Published: April 2006

Abstract
The significance of an interrelation between nongenetic factors and genotype effects in the regulation of bone mass is not clear. In this prospective study of 429 healthy early postmenopausal Danish women, we investigated the association between bone mineral density (BMD) and the FokI and BsmI polymorphisms in the vitamin D receptor (VDR) gene. Participants were allocated to either hormone-replacement therapy (HRT) or no treatment by randomization or personal choice. After 5 years, 332 women with unchanged treatment status were available for analyses, 98 of these women were still on HRT. No association with initial BMD or 5-year change in BMD was found for either polymorphism. In women with body mass index (BMI) &lt; 25 (n = 282), the f allele was associated with lower BMD of the hip (p &lt; 0.001) and forearm (p = 0.001), and the b allele was associated with lower spine BMD (p = 0.02). Comparing thin/normal weight women with overweight/obese women of the same genotype, FF women had similar BMD at all measured sites in contrast to Ff andff women in whom BMD, as expected, was higher in the overweight/obese women. Similar results were found for the BsmI polymorphism with no difference in BMD between BMI groups in BB women. Segregation into groups according to dietary calcium intake did not reveal any genotype association with BMD. These results provide some evidence of a modifying effect of nongenetic factors, specifically BMI, on the association between VDR genotype and BMD. High BMI may protect against lower BMD seen in association with the f or b alleles. In some genotypes (FF and BB), BMI had relatively little effect on BMD.
Improved Bone Structure and Strength After Long-Term Mechanical Loading Is Greatest if Loading Is Separated Into Short Bouts
Authors: Robling Alexander G. , Hinant Felicia M. , Burr David B. , Turner Charles H. 
Published: April 2006

Abstract
Mechanical loading presents a potent osteogenic stimulus to bone cells, but bone cells desensitize rapidly to mechanical stimulation. Resensitization must occur before the cells can transduce future mechanical signals effectively. Previous experiments show that mechanical loading protocols are more osteogenic if the load cycles are divided into several discrete bouts, separated by several hours, than if the cycles are applied in a single uninterrupted bout. We investigated the effect of discrete mechanical loading bouts on structure and biomechanical properties of the rat ulna after 16 weeks of loading. The right ulnas of 26 adult female rats were subjected to 360 load cycles/day, delivered in a haversine waveform at 17 N peak force, 3 days/week for 16 weeks. One-half of the animals (n = 13) were administered all 360 daily cycles in a single uninterrupted bout (360 × 1); the other half were administered 90 cycles four times per day (90 × 4), with 3 h between bouts. A nonloaded baseline control (BLC) group and an age-matched control (AMC) group (n = 9/group) were included in the experiment. The following measurements were collected after death: in situ mechanical strain at the ulna midshaft; ulnar length; maximum and minimum second moments of area (IMAX and IMIN) along the entire length of the ulnas (1-mm increments); and ultimate force, energy to failure, and stiffness of whole ulnas. Qualitative observations of bone morphology were made from whole bone images reconstructed from microcomputed tomography (μCT) slices. Loading according to the 360 × 1 and 90 × 4 schedules improved ultimate force by 64% and 87%, energy to failure by 94% and 165%, IMAX by 13% and 26% (in the middistal diaphysis), IMIN by 69% and 96% (in the middistal diaphysis), and reduced peak mechanical strain by 40% and 36%, respectively. The large increases in biomechanical properties occurred despite very low 5–12% gains in areal bone mineral density (aBMD) and bone mineral content (BMC). Mechanical loading is more effective in enhancing bone biomechanical and structural properties if the loads are applied in discrete bouts, separated by recovery periods (90 × 4 schedule), than if the loads are applied in a single session (360 × 1). Modest increases in aBMD and BMC can improve biomechanical properties substantially if the new bone formation is localized to the most biomechanically relevant sites, as occurs during load-induced bone formation.
Insulin‐Like Growth Factor I Is Required for the Anabolic Actions of Parathyroid Hormone on Mouse Bone
Authors: Bikle Daniel D. , Sakata Takeshi , Leary Colin , Elalieh Hashem , Ginzinger David , Rosen Clifford J. , Beamer Wesley , Majumdar Sharmila , Halloran Bernard P. 
Published: April 2006

AbstractParathyroid hormone (PTH) is a potent anabolic agent for bone, but the mechanism(s) by which it works remains imperfectly understood. Previous studies have indicated that PTH stimulates insulin‐like growth factor (IGF) I production, but it remains uncertain whether IGF‐I mediates some or all of the skeletal actions of PTH. To address this question, we examined the skeletal response to PTH in IGF‐I‐deficient (knockout [k/o]) mice. These mice and their normal littermates (NLMs) were given daily injections of PTH (80 μg/kg) or vehicle for 2 weeks after which their tibias were examined for fat‐free weight (FFW), bone mineral content, bone structure, and bone formation rate (BFR), and their femurs were assessed for mRNA levels of osteoblast differentiation markers. In wild‐type mice, PTH increased FFW, periosteal BFR, and cortical thickness (C.Th) of the proximal tibia while reducing trabecular bone volume (BV); these responses were not seen in the k/o mice. The k/o mice had normal mRNA levels of the PTH receptor and increased mRNA levels of the IGF‐I receptor but markedly reduced basal mRNA levels of the osteoblast markers. Surprisingly, these mRNAs in the k/o bones increased several‐fold more in response to PTH than the mRNAs in the bones from their wild‐type littermates. These results indicate that IGF‐I is required for the anabolic actions of PTH on bone formation, but the defect lies distal to the initial response of the osteoblast to PTH.
1α,25‐Dihydroxyvitamin D3 Promotes Vascularization of the Chondro‐osseous Junction by Stimulating Expression of Vascular Endothelial Growth Factor and Matrix Metalloproteinase 9
Authors: Lin Roberto , Amizuka Norio , Sasaki Tomoyo , Aarts Michelle M. , Ozawa Hideharo , Goltzman David , Henderson Janet E. , White John H. 
Published: April 2006

AbstractVitamin D deficiency results in defects in endochondral bone development characteristic of rickets, which include elongation of the cartilaginous growth plates and disorganization of the primary spongiosa. These defects are caused in part by impaired cartilage mineralization and vascularization of the chondro‐osseous junction. Blood vessel invasion of mineralized cartilage is an essential step in endochondral ossification, providing access for cells that degrade cartilage as well as those that form bone. Vascular endothelial growth factor (VEGF) was shown to be a key regulator of this process when infusion of a dominant negative VEGF receptor effectively blocked vascular invasion and endochondral ossification in the growth plates of juvenile mice. Here, we show that the active metabolite of vitamin D 1α,25‐dihydroxyvitamin D3 [1α,25(OH)2D3] directly stimulates transcription of mRNAs encoding VEGF121 and −165 isoforms in the CFK2 chondrogenic cell line. Enhanced VEGF expression also was evident in growth plate chondrocytes and osteoblasts in the tibia of juvenile mice treated systemically with 1α,25(OH)2D3. This was seen in conjunction with enhanced expression of matrix metalloproteinase (MMP) 9, which activates VEGF stored in the cartilage matrix, in osteoclastic cells adjacent to the chondro‐osseous junction. The alterations in VEGF and MMP‐9 expression were accompanied by enhanced vascular invasion of mineralized cartilage, as assessed by CD31 immunoreactivity. These results provide evidence that 1α,25(OH)2D3 signaling stimulates VEGF and MMP‐9 gene expression and promotes neovascularization of the epiphyseal growth plate in vivo through increased availability of active growth factor.
Mechanical Properties of Adult Vertebral Cancellous Bone: Correlation With Collagen Intermolecular Cross‐Links
Authors: Banse X. , Sims T. J. , Bailey A. J. 
Published: April 2006

AbstractAlthough the mechanical strength of cancellous bone is well known to depend on its apparent density, little is known about the influence of other structural or biochemical parameters. This study specifically investigates the cross‐linking of the collagen in human vertebral bone samples and its potential influence on their mechanical behavior. Multiple cylindrical samples were cored vertically in the vertebral bodies of nine subjects (aged 44–88 years). Three spinal levels (T9, T12 or L1, and L4) and three sample sites within a vertebral body (anterior, posterior, and lateral) were used, for a total of 68 samples. The density was measured with peripheral quantitative computed tomography (pQCT) and all cylinders were mechanically tested in compression. After mechanical testing, they were unmounted and used for biochemical analysis. The amount of collagen (wt/wt of bone) and its content in reduced immature cross‐links, that is, hydroxylysinonorleucine (HLNL, mol/mol of collagen) and dihydroxylysinornorleucine (DHLNL), as well as stable mature cross‐links, that is, hydroxylysyl‐pyridinoline (HP), lysyl‐pyridinoline (LP), and pyrrole cross‐link were determined for each cylinder. None of the biochemical parameters correlated to the density. On multiple linear regression, the prediction of the mechanical properties was improved by combining density data with direct collagen cross‐link assessment. The HP/LP ratio appeared as a significant predictor to the strength (r = 0.40; p = 0.001) and stiffness (r = 0.47; p < 0.001) samples with a high HP/LP ratio being stronger and stiffer. Additionally, the ultimate strain correlated to the HP or LP concentration (r = 0.38 or 0.49; p < 0.01). Different subjects had different HP/LP ratios and different HP or LP concentrations in their vertebral bone samples, and the location of origin within a subject had no influence on the concentration. These observations suggest that the nature of the organic matrix in adult vertebral bone is variable and that these variations influence its mechanical competence.
Regional Alterations of Type I Collagen in Rat Tibia Induced by Skeletal Unloading
Authors: Shiiba Masashi , Arnaud Sara B. , Tanzawa Hideki , Kitamura Eiji , Yamauchi Mitsuo 
Published: April 2006

AbstractSkeletal unloading induces loss of mineral density in weight‐bearing bones that leads to inferior bone mechanical strength. This appears to be caused by a failure of bone formation; however, its mechanisms still are not well understood. The objective of this study was to characterize collagen, the predominant matrix protein in bone, in various regions of tibia of rats that were subjected to skeletal unloading by 4 weeks tail suspension. Sixteen male Sprague‐Dawley rats (4 months old) were divided into tail suspension and ambulatory controls (eight rats each). After the tail suspension, tibias from each animal were collected and divided into five regions and collagen was analyzed. The collagen cross‐linking and the extent of lysine (Lys) hydroxylation in unloaded bones were significantly altered in proximal epiphysis, diaphysis, and, in particular, proximal metaphysis but not in distal regions. The pool of immature/nonmineralized collagen measured by its extractability with a chaotropic solvent was significantly increased in proximal metaphysis. These results suggest that skeletal unloading induced an accumulation of post‐translationally altered nonmineralized collagen and that these changes are bone region specific. These alterations might be caused by impaired osteoblastic function/differentiation resulting in a mineralization defect.
The Effect of In Vivo Mechanical Loading on Estrogen Receptor α Expression in Rat Ulnar Osteocytes
Authors: Ehrlich P. J. , Noble B. S. , Jessop H. L. , Stevens H. Y. , Mosley J. R. , Lanyon L. E. 
Published: April 2006

AbstractThe presence of estrogen receptor α (ERα) in osteocytes was identified immunocytochemically in transverse sections from 560 to 860 μm distal to the midshaft of normal neonatal and adult male and female rat ulnas (n = 3 of each) and from adult male rat ulnas that had been exposed to 10 days of in vivo daily 10‐minute periods of cyclic loading producing peak strains of either −3000 (n = 3) or −4000 microstrain (n = 5). Each animal ambulated normally between loading periods, and its contralateral ulna was used as a control. In animals in which limbs were subject to normal locomotor loading alone, 14 ±1.2% SEM of all osteocytes in each bone section were ERα positive. There was no influence of either gender (p = 0.725) or age (p = 0.577) and no interaction between them (p = 0.658). In bones in which normal locomotion was supplemented by short periods of artificial loading, fewer osteocytes expressed ERα (7.5 ± 0.91% SEM) than in contralateral control limbs, which received locomotor loading alone (14 ± 1.68% SEM; p = 0.01; median difference, 6.43; 95% CI, 2.60, 10.25). The distribution of osteocytes expressing ERα was uniform across all sections and thus did not reflect local peak strain magnitude. This suggests that osteocytes respond to strain as a population, rather than as individual strain‐responsive cells. These data are consistent with the hypothesis that ERα is involved in bone cells' responses to mechanical strain. High strains appear to decrease ERα expression. In osteoporotic bone, the high strains assumed to accompany postmenopausal bone loss may reduce ERα levels and therefore impair the capacity for appropriate adaptive remodeling.