bluuebunny/crossref_metadata_2025_split huggingface dataset:
Premenopausal Smoking and Bone Density in 2015 Perimenopausal Women
Authors: Dr. Hermann A. P. , Brot C. , Gram J. , Kolthoff N. , Mosekilde L. 
Published: April 2006

Abstract
The importance of cigarette smoking in relation to bone mass remains uncertain, especially in younger women. In a recent meta-analysis including 10 studies in premenopausal women no effect was seen in this age group. We used baseline data from a large national cohort study (Danish Osteoporosis Prevention Study [DOPS]) to study the cumulated effect of pre- and perimenopausal smoking on bone mineral density (BMD) measured shortly after the cessation of cyclic bleedings. Baseline observations on 2015 recently menopausal women were available. Eight hundred thirty-two women were current smokers and 285 were exsmokers. Significant negative associations of cigarette smoking coded as current, ex-, or never smoking were seen on bone mass in the lumbar spine (P = 0.012), femoral neck (P &lt; 0.001), and total body (P &lt; 0.001). Quantitatively, the differences between current smokers and never smokers were limited to 1.6, 2.9, and 1.9%, respectively. A statistical interaction was found between smoking and fat mass, indicating that women in the highest tertile of fat mass were unaffected by cigarette smoking. Serum vitamin D levels and osteocalcin were inversely related to the number of cigarettes smoked per day (r = 0.11 and P &lt; 0.001; r = 0.17 and P = 0.04), respectively. Bone alkaline phosphatase (BALP) and urinary hydroxyproline (U-OHP) were unaffected by current smoking. The average cumulated effect of premenopausal smoking on bone is small but biologically significant. Reduced body mass in smokers explains part of the negative effect on the skeleton and a complex interaction between smoking and fat mass on the skeleton is indicated. Serum levels of 25-hydroxyvitamin D (25-OHD) and osteocalcin are lower in smokers, which may effect rate of bone loss.
Parathyroid Hormone Regulates Transforming Growth Factor β1 and β2 Synthesis in Osteoblasts Via Divergent Signaling Pathways
Authors: Wu Yanhong , Kumar Rajiv 
Published: April 2006

Abstract
Parathyroid hormone 1–34 [PTH(1–34)] was shown to increase transforming growth factor β1 (TGF-β1) and TGF-β2 concentrations in supernatants of cultured human osteoblasts and to increase TGF-β1 and TGF-β2 messenger RNA (mRNA) concentrations and gene transcription in these cells. Because PTH(1–34) activates both protein kinase C (PKC) and protein kinase A (PKA) pathways in osteoblasts, we investigated the role of each kinase pathway in activation of TGF-β3 isoforms. PTH(29–32), which activates the PKC pathway in rat osteoblasts, increased TGF-β1 but not TGF-β2 concentrations in supernatants of osteoblasts. Phorbol myristate acetate (PMA), a PKC agonist, increased TGF-β1 but not TGF-β2 concentrations. Specific PKC antagonists safingol and Gö6976 attenuated PTH(1–34)-mediated increases in TGF-β1 but not TGF-β2 synthesis. PTH(1–31), which increases PKA activity in several cell culture systems, increased TGF-β2 but not TGF-β1 concentrations in human osteoblast supernatants. Forskolin, a PKA agonist, increased TGF-β2 but not TGF-β1 concentrations in supernatants of human osteoblasts. The PKA antagonist H-89 blunted PTH(1–34)-mediated increases in TGF-β2 but not TGF-β1 synthesis. Our results are consistent with the concept that PTH increases TGF-β1 expression and secretion by pathways that involve the PKC pathway, whereas it increases TGF-β2 expression and secretion via the PKA pathway. (J Bone Miner Res 2000;15:879–884)
The Inhibitory Effect of Interleukin-10 on Mouse Osteoclast Formation Involves Novel Tyrosine-Phosphorylated Proteins
Authors: Hong Mei Hua , Williams Hilde , Jin Cheng He , Pike J. Wesley 
Published: April 2006

Abstract
Interleukin-10 (IL-10) inhibits osteoclast (OC) formation in rat and mouse systems. However, little is known concerning the mechanism of this inhibitory effect. Using a coculture system of mouse bone marrow cells and primary osteoblastic cells (POB), we evaluated the potential target cells for IL-10 and components of the IL-10 activating pathway. In the coculture system, IL-10 treatment abolished OC differentiation in a dose-dependent manner. This inhibitory effect occurred regardless of the stage of cellular proliferation and differentiation, suggesting that IL-10 may act on a variety of genes participating in OC formation. IL-10 specifically abrogated the production of IL-6 by enriched bone marrow-derived mononuclear cells (BMM) but not by osteoblastic cells. IL-10 treatment also stimulated the binding of a protein in the BMM to an IL-10 response element, whereas no such activation was induced in osteoblastic cells. In contrast, interferon γ (IFN-γ), another inhibitory factor, stimulated tyrosine-phosphorylated proteins to bind to an IL-10 response element in both monocytes and osteoblastic cells. These data suggest that the BMM are the direct target of IL-10 action. Importantly, oligonucleotide-specific precipitation confirmed that IL-10 treatment strongly augmented 88,85, and 70 kDa tyrosine-phosphorylated proteins in BMM. Taken together, these data show that IL-10 inhibits mouse OC formation by acting directly on hemopoietic OC precursor, through a novel signal transduction and activation pathway. (J Bone Miner Res 2000;15:911–918)
Bone-Resorbing Osteoclasts Contain Gap-Junctional Connexin-43
Authors: Ilvesaro Joanna , Väänänen Kalervo , Tuukkanen Juha 
Published: April 2006

Abstract
Intercellular gap junctions have been previously described at contact sites between surface osteoblasts, between osteoblasts and underlying osteocytes, and between osteocyte cell processes in the canaliculi. The subunits of gap junction channels are assembled from a family of proteins called connexins. In the present work, we show that rat osteoclasts cultured on bovine bone slices show connexin-43 (C×43) staining localizing in the plasma membrane of the cells in cell-cell contacts and over the basolateral membrane of osteoclasts. The effect of heptanol, a known gap-junctional inhibitor, was studied using the well-characterized pit formation assay. Heptanol decreased the number and activity of osteoclasts. The proportion of mononuclear tartrate-resistant acid phosphatase (TRAP)–positive cells out of all TRAP-positive cells increased on heptanol treatment, suggesting a defect in the fusion of mononuclear osteoclast precursors to multinucleated mature osteoclasts. Furthermore, the total resorbed area and the number of resorption pits also decreased in the heptanol-treated cultures. These results suggest that gap-junctional C×43 plays a functional role in osteoclasts and that the blocking of gap junctions decreases both the number and the activity of osteoclasts. This can indicate both a direct communication between multinucleated osteoclasts and mononuclear cells through gap junctions or an indirect effect through gap junctions between osteoblasts. (J Bone Miner Res 2000;15:919–926)
Selective Drug Delivery System to Bone: Small Peptide (Asp)6 Conjugation
Authors: Kasugai Shohei , Fujisawa Ryuichi , Waki Yoshihiro , Miyamoto Ken-Ichi , Ohya Keiichi 
Published: April 2006

Abstract
Targeting a drug on hydroxyapatite (HA) could be a promising way for selective drug delivery to bone, because HA, an inorganic component in hard tissues (bone and teeth), does not exist in soft tissues. Several bone noncollagenous proteins, which bind to HA, have repeating sequences of acidic amino acids in their structures as possible HA-binding sites. Thus, we think that a small peptide of repetitive acidic amino acid could work as a carrier for selective drug delivery to the bone. To test this hypothesis, we conjugated (Asp)6 to fluorescein isothiocyanate (FITC), evaluated its affinity to HA in vitro, and examined its tissue distribution after injection into rats. Although fluorescein itself did not bind to HA, (Asp)6-FITC bound to HA as well as calceine and tetracycline. Twenty-four hours after intravenous injection of (Asp)6-FITC to rats, animals were killed, and ground sections of hard tissues and cryosections of soft tissues were made. Under a confocal laser scanning microscope, clear labeling lines were observed in bones and teeth, whereas no labeling was detected in soft tissues. In the rats administered with fluorescein alone, the fluorescent labeling was detected in neither hard nor soft tissues. Fluorescent analysis of blood, urine, and bones after (Asp)6-FITC administration revealed that biological half-life of FITC in blood was short (60 minutes) and that within 24 h, 95% of the administered FITC was excreted as urine whereas 2% of the FITC accumulated in bones. After subcutaneous administration of (Asp)6-FITC to mice, fluorescent intensity remaining in the femurs was measured periodically. In these mice the biological half-life of FITC in the femur was 14 days. Present results indicate that (Asp)6 is effective as a carrier for selective drug delivery to bone. (J Bone Miner Res 2000;15:936–943)
Lactam Formation Increases Receptor Binding, Adenylyl Cyclase Stimulation and Bone Growth Stimulation by Human Parathyroid Hormone (hPTH)(1–28)NH2
Authors: Whitfield J. F. , Morley P. , Willick G. E. , Isaacs R. J. , MacLean S. , Ross V. , Barbier J.-R. , Divieti P. , Bringhurst F. R. 
Published: April 2006

Abstract
Human parathyroid hormone (1–28)NH2 [hPTH(1–28)NH2] is the smallest of the PTH fragments that can fully stimulate adenylyl cyclase in ROS 17/2 rat osteoblast-like osteosarcoma cells. This fragment has an IC50 of 110 nM for displacing 125I-[Nle8,18, Tyr34]bovine PTH(1–34)NH2 from HKRK B7 porcine kidney cells, which stably express 950,000 human type 1 PTH/PTH-related protein (PTHrP) receptors (PTH1Rs) per cell. It also has an EC50 of 23.9 nM for stimulating adenylyl cyclase in ROS 17/2 cells. Increasing the amphiphilicity of the α-helix in the residue 17–28 region by replacing Lys27 with Leu and stabilizing the helix by forming a lactam between Glu22 and Lys26 to produce the [Leu27]cyclo(Glu22-Lys26)hPTH(1–28)NH2 analog dramatically reduced the IC50 for displacing 125I-[Nle8,18, Tyr34]bPTH(1–34)NH2 from hPTHIRs from 110 to 6 nM and dropped the EC50 for adenylyl cyclase stimulation in ROS 17/2 cells from 23.9 to 9.6 nM. These modifications also increased the osteogenic potency of hPTH(1–28)NH2. Thus, hPTH(1–28)NH2 did not significantly stimulate either femoral or vertebral trabecular bone growth in rats when injected daily at a dose of 5 nmol/100 g body weight for 6 weeks, beginning 2 weeks after ovariectomy (OVX), but it strongly stimulated the growth of trabeculae in the cancellous bone of the distal femurs and L5 vertebrae when injected at 25 nmol/100 g body weight. By contrast [Leu27]cyclo(Glu22-Lys26)hPTH(1–28)NH2 significantly stimulated trabecular bone growth when injected at 5 nmol/100 g of body weight. Thus, these modifications have brought the bone anabolic potency of hPTH(1–28)NH2 considerably closer to the potencies of the larger PTH peptides and analogs. (J Bone Miner Res 2000;15:964–970)
The Intracellular Target for the Antiresorptive Aminobisphosphonate Drugs in Dictyostelium discoideum Is the Enzyme Farnesyl Diphosphate Synthase
Authors: Grove Joanna E. , Brown Richard J. , Watts Donald J. 
Published: April 2006

Abstract
Aminobisphosphonate (aBP) drugs inhibit osteoclast-mediated bone resorption and also growth of amoebas of Dictyostelium discoideum apparently by interaction with the same intracellular target. Identification of the target in Dictyostelium therefore could also identify the target in osteoclasts. The aBPs (100 μM alendronate and 30 μM YM-175) inhibited conversion of [14C]mevalonate into sterols by cultures of Dictyostelium amoebas. One of three enzymes (isopentenyl diphosphate [IDP] isomerase, farnesyl diphosphate [FDP] synthase, and squalene synthase) appeared to be the target for this inhibition because conversion of [14C]IDP into squalene, the immediate precursor for sterol biosynthesis, was inhibited in extracts of wild-type amoebas by alendronate (IC50 = 75 nM) or risedronate (IC50 = 30 nM) whereas, when the extract had been prepared from amoebas of strains selected for having partial resistance to the growth-inhibitory effects of alendronate (strain MR102) or risedronate (strain RB101), the values of IC50 were increased to 700 nM for alendronate (MR102 extract) or 130 nM for risedronate (RB101 extract). Neither IDP isomerase nor squalene synthase was inhibited significantly by alendronate or risedronate but both of these aBP drugs, and all others tested, inhibited FDP synthase. Determination of the nucleotide sequences of complementary DNAs (cDNAs) encoding FDP synthase in the wild-type and aBP-resistant strains of Dictyostelium indicated that there had been no changes in the amino acid sequence of the enzyme in the mutant strains. However, both mutant strains overproduce FDP synthase. It is concluded that FDP synthase is the intracellular target for the aBP drugs. (J Bone Miner Res 2000;15:971–981)
Temporal Expression of the Chondrogenic and Angiogenic Growth Factor CYR61 During Fracture Repair
Authors: Hadjiargyrou Michael , Ahrens William , Rubin Clinton T. 
Published: April 2006

Abstract
The repair of a fractured bone is a complex biological event that essentially recapitulates embryonic development and requires the activity of a number of different cell types undergoing proliferation, migration, adhesion, and differentiation, while at the same time expressing a host of different genes. To identify such genes, we employed differential display and compared messenger RNA (mRNA) populations isolated from postfracture (PF) day 5 calluses to those of intact rat femurs. One such gene in which expression was up-regulated at PF day 5 is identified as CYR61, a member of the CCN family of secreted regulatory proteins. CYR61 is a growth factor that stimulates chondrogenesis and angiogenesis. We show that its mRNA expression during fracture repair is regulated temporally, with elevated levels seen as early as PF day 3 and day 5, rising dramatically at PF day 7 and day 10, and finally declining at PF day 14 and day 21. At the highest peak of expression (PF day 7 and day 10, which correlates with chondrogenesis), CYR61 mRNA levels are approximately 10-fold higher than those detected in intact femurs. Similarly, high protein levels are detected throughout the reparative phase of the callus, particularly in fibrous tissue and periosteum, and in proliferating chondrocytes, osteoblasts, and immature osteocytes. The secreted form of CYR61 also was detected within the newly made osteoid. No labeling was detected in hypertrophic chondrocytes or in mature cortical osteocytes. These results suggest that CYR61 plays a significant role in cartilage and bone formation and may serve as an important regulator of fracture healing.
Expression of Indian Hedgehog, Parathyroid Hormone-Related Protein, and Their Receptors in the Postnatal Growth Plate of the Rat: Evidence for a Locally Acting Growth Restraining Feedback Loop After Birth
Authors: Van Der Eerden Bram C. J. , Karperien Marcel , Gevers Evelien F. , Löwik Clemens W. G. M. , Wit Jan Maarten 
Published: April 2006

Abstract
A locally acting growth restraining feedback loop has been identified in the murine embryonic growth plate in which the level of parathyroid hormone-related peptide (PTHrP) expression regulates the pace of chondrocyte differentiation. To date, it is largely unknown whether this feedback loop also regulates the pace of chondrocyte differentiation in the growth plate after birth. We therefore characterized the spatio-temporal expression of Indian hedgehog (IHH), PTHrP, and their receptors in the postnatal growth plate from female and male rats of 1, 4, 7, and 12 weeks of age. These stages are representative for early life and puberty in rats. Using semiquantitative reverse-transcription polymerase chain reaction (RT-PCR) on growth plate tissue, IHH and components of its receptor complex, patched (PTC) and smoothened (SMO), PTHrP and the type I PTH/PTHrP receptor messenger RNA (mRNA) were shown at all ages studied irrespective of gender. Using in situ hybridization, IHH, PTHrP, and PTH/PTHrP receptor mRNA were detected in prehypertrophic and hypertrophic chondrocytes in both sexes during development. In addition, especially in the younger age groups, faint expression of PTH/PTHrP receptor mRNA also was shown in stem cells and proliferative chondrocytes. Immunohistochemistry confirmed the observations made with in situ hybridization, by showing the presence of IHH, PTC, PTHrP, and PTH/PTHrP receptor protein in prehypertrophic and hypertrophic chondrocytes. In addition, staining for hedgehog, PTC, and PTHrP also was observed in growth plate stem cells. No differences in staining patterns were observed between the sexes. Furthermore, no mRNA or protein expression of the mentioned factors was detected in the perichondrium. Our data suggest that in contrast to the proposed feedback loop in the early embryonic growth plate, which requires the presence of the perichondrium, a feedback loop in the postnatal growth plate can be confined to the growth plate itself. In fact, two loops might exist: (1) a loop confined to the transition zone and early hypertrophic chondrocytes, which might in part be autocrine and (2) a loop involving the growth plate stem cells.
Spaceflight Effects on Cultured Embryonic Chick Bone Cells
Authors: Dr. Landis William J. , Hodgens Karen J. , Block Diana , Toma Cyril D. , Gerstenfeld Louis C. 
Published: April 2006

Abstract
A model calcifying system of primary osteoblast cell cultures derived from normal embryonic chicken calvaria has been flown aboard the shuttle, Endeavour, during the National Aeronautics and Space Administration (NASA) mission STS-59 (April 9–20, 1994) to characterize unloading and other spaceflight effects on the bone cells. Aliquots of cells (∼7 × 106) grown in Dulbecco's modified Eagle's medium (DMEM) + 10% fetal bovine serum (FBS) were mixed with microcarrier beads, inoculated into cartridge culture units of artificial hollow fiber capillaries, and carried on the shuttle. To promote cell differentiation, cartridge media were supplemented with 12.5 μg/ml ascorbate and 10 mM β-glycerophosphate for varying time periods before and during flight. Four cartridges contained cells from 17-day-old embryos grown for 5 days in the presence of ascorbate prior to launch (defined as flight cells committed to the osteoblastic lineage) and four cartridges supported cells from 14-day-old embryos grown for 10 days with ascorbate before launch (uncommitted flight cells). Eight cartridges prepared in the same manner were maintained under normal gravity throughout the flight (control cells) and four additional identical cartridges under normal gravity were terminated on the day of launch (basal cells). From shuttle launch to landing, all cartridges were contained in closed hardware units maintaining 5% CO2, 37°C, and media delivery at a rate of ∼1.5 ml/6 h. During day 3 and day 5 of flight, duplicate aliquots of conditioned media and accumulated cell products were collected in both the flight and the control hardware units. At the mission end, comparisons among flight, basal, and control samples were made in cell metabolism, gene expression for type I collagen and osteocalcin, and ultrastructure. Both committed and uncommitted flight cells were metabolically active, as measured by glucose uptake and lactate production, at approximately the same statistical levels as control counterparts. Flight cells elaborated a less extensive extracellular matrix, evidenced by a reduced collagen gene expression and collagen protein appearance compared with controls. Osteocalcin was expressed by all cells, a result indicating progressive differentiation of both flight and control osteoblasts, but its message levels also were reduced in flight cells compared with ground samples. This finding suggested that osteoblasts subjected to flight followed a slower progression toward a differentiated function. The summary of data indicates that spaceflight, including microgravity exposure, demonstrably affects bone cells by down-regulating type I collagen and osteocalcin gene expression and thereby inhibiting expression of the osteogenic phenotype notably by committed osteoblasts. The information is important for insight into the response of bone cells to changes of gravity and of force in general.
Effects of a Short-Term Vitamin D and Calcium Supplementation on Body Sway and Secondary Hyperparathyroidism in Elderly Women
Authors: Pfeifer Michael , Begerow Bettina , Minne Helmut W. , Abrams Christine , Nachtigall Detlef , Hansen Corinna 
Published: April 2006

Abstract
Long-term vitamin D and calcium supplementation is effective in reducing nonvertebral fractures in elderly people. Increased bone fragility caused by secondary hyperparathyroidism (sHPT) and impaired balance are known risk factors for hip fractures. The hypothesis is that short-term therapy with calcium and vitamin D may improve body sway as well as sHPT more effectively than calcium monotherapy. The effects of 8 weeks of supplementation with vitamin D (cholecalciferol) and calcium on body sway and biochemical measures of bone metabolism were measured. The sample consisted of 148 women (mean [±SD] age, 74 ± 1 years) with a 25-hydroxycholecalciferol level below 50 nmol/liter. They received either 1200 mg of calcium plus 800 IU of vitamin D or 1200 mg of calcium per day. We measured intact parathyroid hormone (PTH), markers of bone turnover, and body sway before and after treatment. Falls and fractures among the participants were followed over a 1-year period. Compared with calcium mono, supplementation with vitamin D and calcium resulted in an increase in serum 25-hydroxyvitamin D of 72% (p &lt; 0.0001), a decrease in the serum PTH of 18% (p = 0.0432), and a decrease in body sway of 9% (p = 0.0435). The mean number of falls per subject during a 1-year follow-up period was 0.45 for the calcium mono group and 0.24 for the calcium and vitamin D group (p = 0.0346). Short-term supplementation with vitamin D and calcium improves sHPT and body sway and therefore may prevent falls and subsequent nonvertebral fractures in elderly women.