bluuebunny/crossref_metadata_2025 huggingface dataset:
Distraction Bone Healing Versus Osteotomy Healing: A Comparative Biochemical Analysis
Authors: Lammens Johan , Liu Zhendong , Aerssens Jeroen , Dequeker Jan , Fabry Guy
Published: April 2006

This study investigates the biochemical changes in a canine tibia lengthening model in comparison with a nonlengthened osteotomy model. The lengthened and the osteotomized callus and a contralateral corresponding segment were analyzed for their mineralization profile, collagen content, osteocalcin, insulin-like growth factor I (IGF-I), and transforming growth factor β1 (TGF-β1). Examinations of bone samples were performed using specimens excised at different time intervals (respectively at 3, 5, 7, 9, and 13 weeks postoperatively). Several serum parameters (alkaline phosphatase [ALP], osteocalcin, IGF-I, and TGF-β1) were also measured during the experimental period. A progressive increase in mineral parameters was noticed in both the lengthened and the osteotomized areas. A higher level of hydroxyproline and TGF-β1 was observed in the lengthened area compared with the osteotomized area. IGF-I showed a significant increase in both the lengthened and contralateral control area at the later stage of the experimental period in the lengthened group. In serum, a high level of TGF-β1 and a progressively increasing osteocalcin concentration were observed in the lengthened dogs in comparison with the osteotomized dogs. Serum ALP was significantly increased in both models during the experimental period. Serum IGF-I was increased in the lengthened models during the distraction period and decreased in the osteotomized models at the early stage of the experimental period. These results suggest that the mechanical strain induced by the Ilizarov distraction procedure stimulates osteoblast proliferation and promotes biosynthesis of bone extracellular matrix in distracted callus. Our data furthermore show that this process is different compared with normal fracture healing.
Rapid Bone Loss Is Associated with Increased Levels of Biochemical Markers
Authors: Dr. Ross Philip D. , Knowlton William
Published: April 2006

We examined associations of biochemical markers of bone turnover with rapid bone loss, as measured by changes in bone mineral density (BMD). To improve the precision of bone loss estimates, calcaneal BMD was measured up to eight times over a long interval (13 years) among postmenopausal women (mean age = 62 years at baseline). Women with fractures during the previous year, and users of corticosteroids, active vitamin D, bisphosphonates or calcitonin were excluded to avoid potential transient effects on marker levels. Among the remaining 354 women, markers were measured for 100 women with the fastest BMD loss (rapid loss group; mean = 2.2%/year) and 100 with the slowest loss (mean = 0.4%/year). Two markers of bone formation, serum bone alkaline phosphatase (Alkphase-B; BAP) and osteocalcin (NovoCalcin; OC), and two markers of bone resorption, urinary creatinine-corrected free deoxypyridinoline (Pyrilinks-D; DPD) and free pyridinolines (Pyrilinks; PYD), were measured. In separate logistic regression models, each of the markers was strongly associated with rapid loss: the odds of rapid loss increased by 1.8 to 2.0 times for each 1.0 standard deviation (SD) increase of the marker. For BAP levels 2 SD above the mean, the probability of rapid bone loss was 80%; in contrast, the probability was only 20% at 2 SD below the mean. The other markers yielded similar results. We conclude that these markers are associated with rapid bone loss; this relationship appears to be continuous, with progressively greater risk of rapid bone loss with increasing levels of biomarkers. Prospective studies that include the entire distribution of bone loss rates are warranted to confirm these findings.
The Long-Term Effect of Menopause on Postmenopausal Bone Loss in Japanese Women: Results from a Prospective Study
Authors: Okano Hiroya , Mizunuma Hideki , Soda Masayuki , Kagami Issei , Miyamoto Sumitaka , Ohsawa Minoru , Ibuki Yoshito , Shiraki Masataka , Suzuki Takao , Shibata Hiroshi
Published: April 2006

The aim of this study was to elucidate perimenopausal bone loss in relation to menstrual conditions and to investigate the long-term effect of menopause on bone loss in aged women. The rate of change in bone mineral density (BMD) was measured twice at an exact interval of 12 months by dual-energy X-ray absorptiometry (DXA) at the lumbar spine in 176 pre- and postmenopausal healthy women 41–65 years of age. Serum follicle-stimulating hormone, intact and N-fragment osteocalcin (OC), three types of vitamin D3, parathyroid hormone (PTH), and calcitonin were also determined. Women who exercised regularly or had anatomical changes at the lumbar spine were excluded from this study. The subjects were divided into eight groups based on their menstrual status and years since menopause. Annual bone loss at the lumbar spine of premenopausal women with regular menstruation was −0.2 ± 1.9% (95% confidence interval, −0.9∼–0.4%) and was not statistically different from zero, while that of women with irregular menstruation or at menopausal transition was −2.1 ± 3.4% (−3.4∼−0.8%), and −3.3 ± 2.3% (−5.2∼−0.3%), respectively, and was significantly different from zero. Serum OC levels of women at menopausal transition were significantly higher than those of women with regular menstruation, suggesting that bone loss had commenced in these women. The rate of annual change in BMD of women who were menopausal for 1–3, 4–6, 10–12, and more than 13 years was −3.1 ± 4.0% (−4.7∼−1.5%), −1.2 ± 2.6% (−2.2∼−0.2%), −1.0 ± 3.0% (−2.3∼−0.3%), and −2.3 ± 2.1% (−3.7∼−1.0%), respectively, and was significantly less than zero. But the annual bone loss of women who were menopausal for 7–9 years was −1.5 ± 2.6% (−3.0∼−0.1%) and was not statistically significant from zero. These results indicate that postmenopausal women lose BMD in two phases. The early bone loss is rapid and commences during irregular menstruation, then is attenuated within 6 years after the onset of menopause. The second bone loss commences after the attenuation of the first bone loss. Among bone metabolic hormones, intact PTH alone showed an age-related increase and was suggested as being a causal factor of bone loss in women who were menopausal for 13 years or more.
Bone and the Hematopoietic and Immune Systems: A Report of the Proceedings of a Scientific Workshop
Authors: Sharrock William J. ,   for the ORGANIZERS,   SPEAKERS
Published: April 2006

Recent observations underscore the linkage between endochondral bone formation and the establishment of hematopoietic marrow and suggest that interactions among bone, marrow, and the immune system persist in the mature skeleton. A workshop was held at the National Institutes of Health, Bethesda, Maryland, to discuss recent work on these interactions and to identify new areas of research. Marrow stromal cells include the precursors of the osteochondrogenic lineage, exert important influences on osteoclastogenesis and lymphopoiesis, and mediate the effects of some systemic factors on bone turnover. Recent evidence indicates that hematopoietic cells can influence the differentiation of osteogenic cells and suggests that mature lymphocytes can influence osteoclastic and osteoblastic functions. However, interpretation of experiments may be confounded by the potential for stage-specific responses within a cell lineage, the likelihood that divergent pathways compete for limited pools of precursor cells, and the possibility that important cells or factors are still unidentified. Further, in vitro models may be limited by species and anatomical site specificities, the absence of intermediary or accessory cells, and the absence of normal marrow spatial organization and cellular interactions with the extracellular matrix. Nevertheless, current approaches hold the potential for significant advances in our understanding of the relationships between bone and the hematopoietic and immune systems. Refinements of in vitro systems, the use of genetically manipulated mice, and the examination of clinical syndromes promise important insights. Collaborations among bone biologists, hematologists, and immunologists, and between basic scientists and clinical investigators, will be crucial for continued progress.
Coupling of Calcium Receptors to Inositol Phosphate and Cyclic AMP Generation in Mammalian Cells and Xenopus laevis Oocytes and Immunodetection of Receptor Protein by Region-Specific Antipeptide Antisera
Authors: Chang Wenhan , Pratt Stacy , Chen Tsui-Hua , Nemeth Edward , Huang Zhengmin , Dr. Shoback Dolores
Published: April 2006

Ca2+ and other divalent cations modulate parathyroid hormone secretion by interacting with cell-surface Ca2+-sensing receptors (CaRs). We assessed the ability of these receptors to couple to Ca2+ mobilization, inositol phosphate (InsP) accumulation, and cyclic AMP production in different expression systems. In Xenopus laevis oocytes injected with bovine parathyroid CaR cRNA, the addition of extracellular cations to 1.5 mM Ca2+, 5.5 mM Mg2+, or 10 μM Gd3+ significantly increased45Ca efflux (p < 0.01). InsP accumulation also increased dramatically when adding these cations to human embryonic kidney (HEK) 293 cells stably transfected with wild-type bovine parathyroid CaR cDNA. Raising the extracellular [Ca2+] ([Ca2+]o) from 0.1 to >1.4 mM in oocytes and to >1.0 mM in HEK 293 cells stimulated significant increments in45Ca efflux and InsP accumulation, respectively (p < 0.05). In contrast, Ca2+ and Mg2+ increased InsPs to a lesser extent in COS 7 cells transiently transfected with CaR cDNA. In HEK 293 cells stably expressing CaR cDNA, there were significant reductions in cAMP content when adding high Ca2+, Mg2+, Gd3+, or the CaR modulator NPS R-467. Three region-specific anti-CaR peptide antisera immunoblotted bands of ∼140 and 155 kDa in membranes from CaR-transfected HEK 293 cells and bovine parathyroid tissue. Immunocytochemistry demonstrated strong cell-surface staining in CaR-transfected HEK 293 cells and parathyroid tissue, which was absent when antisera were preabsorbed with CaR peptides. These results indicate that the activation of the recombinant CaR by extracellular Ca2+ can couple negatively to adenylate cyclase but positively to phospholipase C (PLC), the latter at physiological [Ca2+]o.
Nitrogen-Containing Bisphosphonates Inhibit the Mevalonate Pathway and Prevent Post-Translational Prenylation of GTP-Binding Proteins, Including Ras
Authors: Luckman Steven P. , Hughes David E. , Coxon Fraser P. , Russell R. Graham G. , Dr. Rogers Michael J.
Published: April 2006

Bisphosphonates are currently the most important class of antiresorptive drugs used for the treatment of metabolic bone diseases. Although the molecular targets of bisphosphonates have not been identified, these compounds inhibit bone resorption by mechanisms that can lead to osteoclast apoptosis. Bisphosphonates also induce apoptosis in mouse J774 macrophages in vitro, probably by the same mechanisms that lead to osteoclast apoptosis. We have found that, in J774 macrophages, nitrogen-containing bisphosphonates (such as alendronate, ibandronate, and risedronate) inhibit post-translational modification (prenylation) of proteins, including the GTP-binding protein Ras, with farnesyl or geranylgeranyl isoprenoid groups. Clodronate did not inhibit protein prenylation. Mevastatin, an inhibitor of 3-hydroxy-3-methylglutatyl (HMG)-CoA reductase and hence the biosynthetic pathway required for the production of farnesyl pyrophosphate and geranylgeranyl pyrophosphate, also caused apoptosis in J774 macrophages and murine osteoclasts in vitro. Furthermore, alendronate-induced apoptosis, like mevastatin-induced apoptosis, could be suppressed in J774 cells by the addition of farnesyl pyrophosphate or geranylgeranyl pyrophosphate, while the effect of alendronate on osteoclast number and bone resorption in murine calvariae in vitro could be overcome by the addition of mevalonic acid. These observations suggest that nitrogen-containing bisphosphonate drugs cause apoptosis following inhibition of post-translational prenylation of proteins such as Ras. It is likely that these potent antiresorptive bisphosphonates also inhibit bone resorption by preventing protein prenylation in osteoclasts and that enzymes of the mevalonate pathway or prenyl protein transferases are the molecular targets of the nitrogen-containing bisphosphonates. Furthermore, the data support the view that clodronate acts by a different mechanism.
Mesenchymal Stem Cell Surface Antigen SB-10 Corresponds to Activated Leukocyte Cell Adhesion Molecule and Is Involved in Osteogenic Differentiation
Authors: Bruder Scott P. , Ricalton Nancy S. , Boynton Raymond E. , Connolly Timothy J. , Jaiswal Neelam , Zaia Joseph , Barry Frank P.
Published: April 2006

Bone marrow contains a rare population of mesenchymal stem cells (MSCs) capable of giving rise to multiple mesodermal tissues including bone, cartilage, tendon, muscle, and fat. The cell surface antigen recognized by monoclonal antibody SB-10 is expressed on human MSCs but is lost during their developmental progression into differentiated phenotypes. Here we report on the immunopurification of the SB-10 antigen and its identification as activated leukocyte-cell adhesion molecule (ALCAM). Mass spectrometry establishes that the molecular mass of ALCAM is 80,303 ± 193 Da and that it possesses 17,763 ± 237 Da of N-linked oligosaccharide substituents. Molecular cloning of a full-length cDNA from a MSC expression library demonstrates nucleotide sequence identity with ALCAM. We also identified ALCAM homologs in rat, rabbit, and canine MSCs, each of which is over 90% identical to human ALCAM in their peptide sequence. The addition of antibody SB-10 Fab fragments to human MSCs undergoing osteogenic differentiation in vitro accelerated the process, thereby implicating a role for ALCAM during bone morphogenesis and adding ALCAM to the group of cell adhesion molecules involved in osteogenesis. Together, these results provide evidence that ALCAM plays a critical role in the differentiation of mesenchymal tissues in multiple species across the phylogenetic tree.
Role of Interleukin-6 in Uncoupling of Bone In Vivo in a Human Squamous Carcinoma Coproducing Parathyroid Hormone-Related Peptide and Interleukin-6
Authors: Nagai Yumiko , Yamato Hideyuki , Akaogi Keiko , Hirose Kunitaka , Ueyama Yoshito , Ikeda Kyoji , Matsumoto Toshio , Fujita Toshiro , Ogata Etsuro
Published: April 2006

OCC tumor has been established from a human squamous carcinoma associated with humoral hypercalcemia of malignancy (HHM) and shown to overproduce parathyroid hormone-related peptide (PTHrP) and cause aggressive hypercalcemia when implanted into nude rats. In the present study, we have demonstrated by reverse transcription-polymerase chain reaction and Northern blot analysis that OCC tumor also overexpressed interleukin 6 (IL-6) mRNA and that tumor-bearing animals exhibited a marked increase in plasma IL-6 as well as PTHrP concentrations. When a monoclonal antibody against human IL-6 was injected to block the activities of tumor-derived IL-6, bone loss in tumor-bearing animals was significantly prevented. Quantitative bone histomorphometric analysis revealed that treatment with anti–IL-6 antibody caused a substantial decrease in both osteoclast number and eroded surface (as parameters of bone resorption) and also a significant increase in the mineral apposition rate, but little effect on the osteoblastic surface. These results provide in vivo evidence suggesting that in tumors coproducing IL-6 and PTHrP, IL-6 is involved not only in the acceleration of osteoclastic bone resorption but also, at least in part, in the suppression of osteoblastic functions in HHM syndrome.
Characterization of Serum Tartrate-Resistant Acid Phosphatase and Development of a Direct Two-Site Immunoassay
Authors: Halleen Jussi M. , Hentunen Teuvo A. , Karp Matti , Käkönen Sanna-Maria , Pettersson Kim , Väänänen H. Kalervo
Published: April 2006

Osteoclasts secrete tartrate-resistant acid phosphatase (TRAP) to the circulation, where the amount of TRAP is expected to correlate with the bone resorption rate. We have developed two monoclonal antibodies, O1A and J1B, using purified human bone TRAP as antigen. The antibodies recognized different epitopes, allowing us to develop a two-site fluoroimmunoassay. The immunoreactivity in fresh serum specimens was less than 10% of the concentrations measured from the same specimens after 24 h of storage at 4°C, or after addition of 5 mM EDTA or EGTA to them. When fresh serum was gel filtrated using Sephacryl S-200 column, all of the enzyme eluted in the void volume as a complex with a molecular weight of more than 250 kDa. If the serum was treated with EDTA before the gel filtration, the complex was destroyed and the enzyme eluted in fractions corresponding to a molecular weight of 30 kDa, the size of monomeric purified human bone TRAP. The immunoassay was used to measure TRAP concentrations from serum samples that had been stored at 4°C for 24 h. According to the assay, premenopausal women had 13.1 ± 3.1, postmenopausal women 17.6 ± 4.2, and children 32.6 ± 12.2 μg TRAP/l of serum. We conclude that TRAP circulates in the serum as part of a complex, which also contains Ca2+, and that TRAP-immunoassay is a potentially useful method for determining bone resorption rates, as long as the complex is destroyed before the assay.
Osteoblast Programmed Cell Death (Apoptosis): Modulation by Growth Factors and Cytokines
Authors: Jilka Robert L. , Weinstein Robert S. , Bellido Teresita , Parfitt A. Michael , Manolagas Stavros C.
Published: April 2006

Once osteoblasts have completed their bone-forming function, they are either entrapped in bone matrix and become osteocytes or remain on the surface as lining cells. Nonetheless, 50–70% of the osteoblasts initially present at the remodeling site cannot be accounted for after enumeration of lining cells and osteocytes. We hypothesized that the missing osteoblasts die by apoptosis and that growth factors and cytokines produced in the bone microenvironment influence this process. We report that murine osteoblastic MC3T3-E1 cells underwent apoptosis following removal of serum, or addition of tumor necrosis factor (TNF), as indicated by terminal deoxynucleotidyl transferase–mediated dUTP-nick end labeling and DNA fragmentation studies. Transforming growth factor-β and interleukin-6 (IL-6)–type cytokines had antiapoptotic effects because they were able to counteract the effect of serum starvation or TNF. In addition, anti-Fas antibody stimulated apoptosis of human osteoblastic MG-63 cells and IL-6–type cytokines prevented these changes. The induction of apoptosis in MG-63 cells was associated with an increase in the ratio of the proapoptotic protein bax to the antiapoptotic protein bcl-2, and oncostatin M prevented this change. Examination of undecalcified sections of murine cancellous bone revealed the presence of apoptotic cells, identified as osteoblasts by their proximity to osteoid seams and their juxtaposition to cuboidal osteoblasts. Assuming an osteoblast life span of 300 h and a prevalence of apoptosis of 0.6%, we calculated that the fraction that undergo this process in vivo can indeed account for the missing osteoblasts. These findings establish that osteoblasts undergo apoptosis and strongly suggest that the process can be modulated by growth factors and cytokines produced in the bone microenvironment.
Inorganic Polyphosphate in Human Osteoblast-like Cells
Authors: Leyhausen Gabriele , Lorenz Bernd , Zhu Hua , Geurtsen Werner , Bohnensack Ralf , Müller Werner E. G. , Schröder Heinz C.
Published: April 2006

Significant amounts of inorganic polyphosphates and of polyphosphate-degrading exopolyphosphatase activity were detected in human mandibular-derived osteoblast-like cells. The amount of both soluble and insoluble long-chain polyphosphate in unstimulated osteoblast-like cells was higher than in human gingival cells, erythrocytes, peripheral blood mononuclear cells, and human blood plasma. The cellular content of polyphosphate in osteoblast-like cells strongly decreased after a combined treatment of the cells with the stimulators of osteoblast proliferation and differentation, dexamethasone, β-glycerophosphate, epidermal growth factor, and ascorbic acid. The amount of soluble long-chain polyphosphate, but not the amount of insoluble long-chain polyphosphate, further decreased after an additional treatment with 1α,25-dihydroxyvitamin D3(1,25(OH)2D3). The decrease in polyphosphate content during treatment with dexamethasone, β-glycerophosphate, epidermal growth factor, and ascorbic acid was accompanied by a decrease in exopolyphosphatase, pyrophosphatase, and alkaline phosphatase activity. However, additional treatment with 1,25(OH)2D3 resulted in an increase in these enzyme activities. Osteoblast-like cell exopolyphosphatase activity and exopolyphosphatase activity in yeast, rat tissues, and human leukemia cell line HL60 were inhibited by the bisphosphonates etidronate and, to a lesser extent, clodronate and pamidronate. From our results, we assume that inorganic polyphosphate may be involved in modulation of the mineralization process in bone tissue.
Expression of the Thrombin Receptor in Developing Bone and Associated Tissues
Authors: Abraham Linda A. , Jenkins Alison L. , Stone Stuart R. , Mackie Eleanor J.
Published: April 2006

Thrombin, a serine protease with a central role in thrombosis and hemostasis, is also a specific agonist for a variety of cellular responses in osteoblasts and stimulates bone resorption in organ culture. Cultured osteoblast-like cells express the proteolytically activated thrombin receptor, but the significance of this finding in vivo remains unknown. Immunohistochemistry was used to investigate the normal tissue distribution of the proteolytically activated thrombin receptor in developing rat bones and associated tissues. In hind limbs, the receptor was first observed on embryonic day 16 and became more abundant within the limb as gestation progressed. Thrombin receptor staining was detected on osteoblasts, macrophages, muscle cells, and endothelial cells, but not osteoclasts. Similarly, osteoblasts in developing calvariae stained positively for the thrombin receptor. The pattern of receptor expression by primary osteoblast cultures and freshly isolated macrophages and osteoclasts corresponded to that observed in vivo. The observed pattern of thrombin receptor expression in bone cells supports the hypothesis that cell-mediated thrombin-induced bone resorption is mediated by osteoblasts.
Estrogen via the Estrogen Receptor Blocks cAMP-Mediated Parathyroid Hormone (PTH)-Stimulated Osteoclast Formation
Authors: Kanatani Masanori , Sugimoto Toshitsugu , Takahashi Yasuyuki , Kaji Hiroshi , Kitazawa Riko , Chihara Kazuo
Published: April 2006

Several lines of evidence indicate that estrogen inhibits parathyroid hormone (PTH)-induced bone resorption in vivo and in vitro. However, its precise mechanism remains unknown. The present study was performed to investigate whether osteoclast precursor cells possess the receptors for PTH/PTH-related protein (PTHrP) and/or estrogen and to clarify the mechanism by which estrogen affects PTH-induced osteoclast-like cell (Ocl) formation. The polymerase chain reaction (PCR) product corresponding in size to the mouse PTH/PTHrP receptor cDNA was detected in mouse hemopoietic blast cells supported by granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as in osteoblastic MC3T3-E1 cells. The nucleotide sequence of the PTH/PTHrP receptor PCR product of hemopoietic blast cells was found to be 95.4% identical to that of PTH/PTHrP receptor cDNA of rat osteoblastic ROS cells. The PCR product corresponding in size to the mouse estrogen receptor cDNA was detected in mouse hemopoietic blast cells supported by GM-CSF as well as in MC3T3-E1 cells. The nucleotide sequence of the estrogen receptor PCR product of hemopoietic blast cells was completely identical to that of mouse estrogen receptor cDNA. 17β-estradiol (17β-E2) but not 17α-E2 dose dependently antagonized Ocl formation stimulated by human (h) PTH(1–34) at a minimal effective concentration of 10−10 M in the hemopoietic blast cell culture. 17β-E2 also significantly inhibited Ocl formation stimulated by 10−8 M hPTHrP(1–34), while it did not affect 1,25-dihydroxyvitamin D3–induced Ocl formation. However, 10−8 M 17β-E2 significantly inhibited Ocl formation stimulated by dibutyryladenosine cAMP (10−4 M) and Sp-cAMPS (10−4 M), an activator of cAMP-dependent protein kinase (PKA) as well as forskolin (10−5 M). In contrast, 17β-E2 did not affect Ocl formation by either phorbol myristate acetate (10−7 M), an activator of protein kinase C (PKC), or A23187 (10−7 M), a calcium ionophore. The pretreatment with 17β-E2 significantly inhibited Ocl formation induced by the combined treatment with PTH and PKC inhibitors (H7 or staurosporine), while it did not affect Ocl formation stimulated by the combined treatment with PTH and Rp-cAMPS, a PKA inhibitor. The present data indicate that estrogen inhibits PTH-stimulated Ocl formation by directly acting on hemopoietic blast cells, possibly through blocking a PKA pathway but not a calcium/PKC pathway.
Serum Levels of Insulin-like Growth Factor Binding Proteins (IGFBP)–4 and –5 Correlate with Bone Mineral Density in Growth Hormone (GH)–Deficient Adults and Increase with GH Replacement Therapy
Authors: Thorén Marja , Hilding Agneta , Brismar Torkel , Magnusson Per , Degerblad Marie , Larsson Lasse , Sääf Maria , Baylink David J. , Mohan Subburaman
Published: April 2006

Adults with growth hormone deficiency (GHD) exhibit low bone mineral density (BMD) which improves by growth hormone (GH) replacement therapy. The insulin-like growth factor (IGF) system has an established role in mediating the effects of GH on bone and IGF binding proteins (IGFBP)-4 and IGFBP-5 have been shown to modulate the effects of IGFs in bone. Therefore, we studied serum levels of IGFBP-4 and IGFBP-5 and their relationship to serum levels of bone biochemical markers and BMD in adults with GH deficiency (GHD) before and during GH therapy. Serum levels of IGFBP-5 and IGFBP-4 were measured on samples from 20 patients (11 males) 22–57 years of age. All had IGF-I serum values below –2 standard deviation score. The first 6 months were placebo controlled and all recieved 3 years of active treatment with the mean dose 0.23 ± 0.01 IU/kg/week divided into daily subcutaneous injections. Serum IGFBP-5 levels in GHD adults were low at baseline and positively related to total body, femoral neck, trochanter, and Ward's triangle BMD (r = 0.471, 0.549, 0.462, and 0.470, respectively, p < 0.05). The mean serum IGFBP-5 level increased by about 2-fold within 3 months after the initiation of GH therapy and was correlated with serum IGF-I (r = 0.719, 0.801, and 0.722 before and after 18 and 36 months, respectively, p < 0.001). A positive correlation between serum IGFBP-5 levels and lumbar spine BMD was found during GH treatment but not before. The percentage increase of serum IGFBP-5 after GH therapy showed a positive correlation with the percentage increase of total alkaline phosphate activity (r = 0.347 p < 0.05). In contrast to IGFBP-5, serum IGFBP-4 levels were positively related to body mass index (r = 0.607, p < 0.01). Baseline serum IGFBP-4 levels also correlated with total body, femoral neck, trochanter, and Ward's triangle BMD (r = 0.502, 0.590, 0.612, and 0.471, respectively, p < 0.05). The mean serum IGFBP-4 level was increased by 25% within 3 months after initiation of GH therapy and did not correlate with serum IGF-I levels. Although the above findings are consistent with the idea that GH-induced changes in serum IGFBP-5 and IGFBP-4 levels may in part mediate the anabolic effects of GH on bone tissue in adults with GHD, further studies are needed to establish the cause and effect relationship.
Paget's Disease of Bone
Authors: Siris Ethel S.
Published: April 2006

Paget's disease of bone is a localized disorder of bone remodeling. Increased numbers of larger than normal osteoclasts initiate the process at affected skeletal sites, and the increase in bone resorption is followed by an increase in new bone formation, altering bone architecture. The signs and symptoms of Paget's disease are varied, depending in part on the location of the involved sites and the degree of increased bone turnover. Recent progress in Paget's disease research includes new data regarding the etiology of this disorder and the ongoing development of more effective therapies. Although the cause of Paget's disease remains unproven, the creation of pagetic osteoclasts seems ever more likely to result from both genetic and environmental factors. Many studies indicate that in patients with Paget's disease, both osteoclasts and their precursors harbor evidence of a paramyxovirus infection, although not all studies confirm this finding. Very recent genetic investigations have identified one candidate gene on chromosome 18q, although genetic heterogeneity is almost certainly present. Advances in treatment have resulted from the availability of several potent bisphosphonate compounds (e.g., pamidronate, alendronate, and risedronate) that, unlike earlier treatments, produce normal or near normal bone turnover indices in a majority of patients. New bone formation after such treatment has a more normal, lamellar pattern, and mineralization abnormalities are rare to absent with the newer compounds. The availability of such agents has prompted a more aggressive management philosophy in which both symptomatic disease and also asymptomatic disease at sites with a risk of progression and future complications are viewed as clear indications for pharmacologic intervention.
Regulation of Prostaglandin G/H Synthase-2 Expression by Interleukin-1 in Human Osteoblast-like Cells
Authors: Min Yong-Ki , Rao Yue , Okada Yosuke , Raisz Lawrence G. , Pilbeam Carol C.
Published: April 2006

Interleukin-1 (IL-1) is an important factor in bone metabolism, and its actions may be mediated in part via prostaglandins. Prostaglandin G/H synthase (PGHS), a critical enzyme in the synthesis of prostaglandins, has two isoforms, PGHS-1, which is generally constitutively expressed, and PGHS-2, which is inducible. This study examines the effects of IL-1 on PGHS-2 mRNA expression in human osteosarcoma MG-63 cells, the human osteoblast-like initial transfectant (HOBIT) cell line, and primary human osteoblastic (HOB) cells. IL-1 induced PGHS-2 mRNA expression in MG-63 cells within 1 h, and expression was maintained for 24 h. There was a dose-related increase in PGHS-2 mRNA levels with 1–100 ng/ml of IL-1. Induction of PGHS-2 protein and media prostaglandin E2 (PGE2) paralleled induction of PGHS-2 mRNA levels. IL-1 similarly induced PGHS-2 mRNA expression and PGE2 production in HOBIT and HOB cells. Among other potential agonists, phorbol myristate acetate (PMA) was a potent inducer of PGHS-2 expression, while forskolin (FSK), serum, and prostaglandins had little effect. Cycloheximide enhanced effects of both IL-1 and PMA, suggesting that de novo protein synthesis is not required for induction of PGHS-2. Twenty-four hours of PMA pretreatment blocked the induction of PGHS-2 by PMA but not by IL-1, suggesting that IL-1 induction of PGHS-2 mRNA is not dependent on the protein kinase C pathway. Although FSK alone had little effect, it enhanced induction of PGHS-2 mRNA by IL-1. PGHS-1 was constitutively expressed and showed little change with treatment. In summary, we show that IL-1 is a potent inducer of PGHS-2 expression and PGE2 production in human osteosarcoma cells as well as in osteoblastic cells derived from normal human bone.
The Tromsø Study: Physical Activity and the Incidence of Fractures in a Middle-Aged Population
Authors: Joakimsen Ragnar M. , Fønnebø Vinjar , Magnus Jeanette H. , Størmer Jan , Tollan Anne , Søgaard Anne Johanne
Published: April 2006

We have studied the relation of occupational and recreational physical activity to fractures at different locations. All men born between 1925 and 1959 and all women born between 1930 and 1959 in the city of Tromsø were invited to participate in surveys in 1979–1980 and 1986–1987 (The Tromsø Study). Of 16,676 invited persons, 12,270 (73.6%) attended both surveys. All nonvertebral fractures (n = 1435) sustained from 1988 to 1995 were registered in the only hospital in the area. Average age in the middle of the follow-up period (December 31, 1991) was 47.3 years among men and 45.1 years among women, ranging from 32 to 66 years. Fracture incidence increased with age at all locations among women, but it decreased with or was independent of age among men. Low-energetic fractures constituted 74.4% of all fractures among women and 55.2% among men. When stratifying by fracture location, the most physically active persons among those 45 years or older suffered fewer fractures in the weight-bearing skeleton (relative risk [RR] 0.6, confidence interval [CI] 0.4–0.9, age-adjusted), but not in the non–weight-bearing skeleton (RR 1.0, CI 0.7–1.2, age-adjusted) compared with sedentary persons. The relative risk of a low-energetic fracture in the weight-bearing skeleton among the most physically active middle-aged was 0.3 (CI 0.1–0.7) among men and 0.9 (CI 0.4–1.8) among women compared with the sedentary when adjusted for age, body mass index, body height, tobacco smoking, and alcohol and milk consumption. It seems that the beneficial effect on the skeleton of weight-bearing activity is reflected also in the incidence of fractures at different sites.
Treatment with the Oral Growth Hormone Secretagogue MK-677 Increases Markers of Bone Formation and Bone Resorption in Obese Young Males
Authors: Svensson J. , Ohlsson C. , Jansson J.-O. , Murphy G. , Wyss D. , Krupa D. , Cerchio K. , Polvino W. , Gertz B. , Baylink D. , Mohan S. , Bengtsson B.-Å.
Published: April 2006

The effect of 2 months of treatment with the oral growth hormone (GH) secretagogue MK-677 on markers of bone metabolism was determined in healthy obese male subjects. This was a randomized, double-blind, parallel, placebo-controlled study. Twenty-four healthy obese males, 19–49 years of age, with body mass index > 30 kg/m2 were treated with MK-677 (25 mg/day; n = 12) or placebo (n = 12) for 8 weeks. MK-677 increased markers of bone formation; a 23% increase in the carboxy-terminal propeptide of type I procollagen levels and a 28% increase in procollagen III peptide levels were seen with as little as 2 weeks of MK-677 treatment (p < 0.01 and p = 0.001 vs. placebo, respectively) while a 15% increase in serum levels of osteocalcin was not detected until 8 weeks of treatment (p < 0.01 vs. placebo). Markers of bone resorption were induced within 2 weeks of treatment with MK-677; serum levels of the carboxy-terminal cross-linked telopeptide of type I collagen were increased 26% at 8 weeks (p = 0.001 vs. placebo), and urine hydroxyproline/creatinine and calcium/creatinine ratios at 8 weeks were increased by 23% (p < 0.05 vs. placebo) and 46% (p < 0.05 vs placebo), respectively. MK-677 increased serum insulin-like growth factor binding protein-5 (IGFBP-5) by 43–44% after 2–8 weeks of treatment (p < 0.01 vs. placebo). Serum IGFBP-4 was increased by 25% after 2 weeks of treatment (p < 0.001 vs. placebo) but no significant change from baseline was observed after 8 weeks of treatment. Plasma interleukin-6 was not significantly changed by active treatment. In conclusion, short-term treatment of healthy obese male volunteers with the GH secretagogue MK-677 increases markers of both bone resorption and formation. Large increases in serum levels of IGF-I and IGFBP-5 and a transient increase in serum IGFBP-4 were found. Future long-term studies are needed to investigate if prolonged treatment with MK-677 increases bone mass.
Hormonal Predictors of Bone Loss in Elderly Women: A Prospective Study
Authors: Stone Katie , Bauer Douglas C. , Black Dennis M. , Sklarin Peter , Ensrud Kristine E. , Cummings Steven R. ,   FOR THE STUDY OF OSTEOPOROTIC FRACTURES RESEARCH GROUP
Published: April 2006

To test the hypotheses that baseline concentrations of sex steroids, sex hormone binding globulin (SHBG), and calciotropic hormones predict rates of bone loss in elderly women, sera were stored at −190°C, and calcaneal bone mineral density (BMD) was measured in 9704 community-dwelling white women aged 65 and over (1986–1988). Hip BMD was measured 2 years later (1990). Repeat measurements of calcaneal and hip BMD were obtained in 1993–1994, after 5.7 and 3.5 years of follow-up, respectively. In 1994, sera were assayed for circulating hormone levels in random subcohorts of 231 and 218 women who did not report current use of hormone replacement therapy at baseline. Lower levels of endogenous estrogens and higher SHBG concentrations were associated with more rapid subsequent bone loss from both the calcaneus and hip. After adjusting for age and weight, women with high SHBG levels (highest quartile ≥ 2.3 μg/dl) experienced an average of 2.2% (95% confidence interval = 1.6%, 2.9%) calcaneal bone loss per year compared with 1.2% (0.7%, 1.2%) among women with low SHBG concentrations (lowest quartile < 1.1 μg/dl; p < 0.01). This association was independent of concentrations of other sex hormones. Women with estradiol levels ≥ 10 pg/ml averaged only 0.1% (−0.7%, 0.5%) annual hip bone loss while women with levels below 5 pg/ml averaged 0.8% (0.3, 1.2) hip bone loss per year. Lower 25-hydroxyvitamin D levels were associated with increased hip but not calcaneal bone loss. Levels of parathyroid hormone, 1,25-dihydroxyvitamin D, and calcium were not significantly associated with bone loss from the calcaneus or hip.
Urinary Ovarian and Gonadotropin Hormone Levels in Premenopausal Women with Low Bone Mass
Authors: Sowers Maryfran , Randolph John F. , Crutchfield Mary , Jannausch Mary L. , Shapiro Brahm , Zhang Bin , La Pietra Maggie
Published: April 2006

We hypothesized that lower ovarian and gonadotropin hormone concentrations would be associated with lower levels of peak bone mineral density (BMD) in apparently normally menstruating women who did not exercise intensively and did not report anorexia or bulimia. This hypothesis was evaluated using a case-with-control study design (n = 65) which was nested within a population-based longitudinal study of peak bone mass (Michigan Bone Health Study) with annual assessment in women aged 25–45 years (n = 582). Cases were 31 premenopausal women with BMD of the lumbar spine, femoral neck, and total body less than the 10th percentile of the distribution, where controls were 34 premenopausal women with BMD between the 50th and 75th percentile. BMD was measured by dual-energy X-ray absorptiometry. In addition to their annual measurements, these 65 participants collected first-voided morning urine specimens daily through two consecutive menstrual cycles. The urine from alternating days of this collection was analyzed for estrone-3-glucuronide (E1G), pregnanediol glucuronide (PdG), testosterone, and follicle-stimulating hormone by radioimmunoassay and these values adjusted for daily creatinine excretion levels. Additionally, analyses of daily urine specimens for luteinizing hormone (uLH) was undertaken to better characterize the possible uLH surge. Cases had significantly lower amounts of E1G (p = 0.009) and PdG (p = 0.002) than did controls, whether amounts were characterized by a mean value, the highest value, or the area under the curve, and after statistically controlling for body size. Further, when B-splines were used to fit lines to the E1G and PdG data across the menstrual cycle, the 95% confidence intervals (CIs) about the line for the controls consistently excluded and exceeded the 95% confidence bands for the cases in the time frame associated with the luteal phase in ovulatory cycles. Likewise, 95% CIs for the LH surge in controls exceeded the fitted line for cases around the time associated with the LH surge. The cases and controls were not different according to dietary intake (energy, protein, calcium), family history of osteoporosis, reproductive characteristics (parity, age at menarche, age of first pregnancy), follicular phase serum hormone levels, calciotropic hormone levels, or by evidence of perimenopause. We conclude that these healthy, menstruating women with BMD at the lowest 10th percentile from a population-based study had significantly lower urinary sex steroid hormone levels during the luteal phase of menstrual cycles as compared with hormone levels in premenopausal women with BMD between the 50th and 75th percentile of the same population-based study, even after considering the role of body size. These data suggest that subclinical decreases in circulating gonadal steroids may impair the attainment and/or maintenance of bone mass in otherwise reproductively normal women.
Effect of Combination Treatment with a Vitamin D Analog (OCT) and a Bisphosphonate (AHPrBP) in a Nude Mouse Model of Cancer-Associated Hypercalcemia
Authors: Endo Koichi , Katsumata Kyoko , Iguchi Haruo , Kubodera Noboru , Teramoto Tamio , Ikeda Kyoji , Fujita Toshiro , Ogata Etsuro
Published: April 2006

Hypercalcemia represents one of the important paraneoplastic syndromes affecting morbidity and mortality of cancer patients. We and others have demonstrated that vitamin D analogs with little calcemic activities suppress the transcription of the parathyroid hormone-related peptide (PTHrP) gene, a major humor responsible for cancer hypercalcemia, and thereby prevent the development of hypercalcemic syndrome. The present study was undertaken: to compare the therapeutic efficacy of a vitamin D analog, 22-oxa-1,25-dihydroxyvitamin D3 (OCT), and a bisphosphonate (disodium 3-amino-1-hydroxypropylidene-1,1-bisphosphonate pentahydrate [AHPrBP]), an inhibitor of osteoclastic bone resorption, on cancer-induced hypercalcemia; and to see if the effect could be enhanced by combination treatment, using a nude mouse model implanted with a human pancreas carcinoma (FA-6). After a single intravenous administration, OCT (5 μg/kg of body weight [BW]) was as effective as AHPrBP (10 mg/kg of BW) in lowering blood ionized calcium levels in tumor-bearing nude mice, and their combination further enhanced the therapeutic effect. Although AHPrBP lost its efficacy after repeated injections, OCT was still effective after the third administration. The therapeutic effect of OCT in cancer hypercalcemia was observed in four other human tumors, including another pancreas carcinoma (PAN-7), two squamous cell carcinomas of the lung (KCC-C1 and LC-6), and a squamous carcinoma of the pharynx (PHA-1), all of which elaborated PTHrP into the circulation. Treatment with OCT resulted in a decrease in circulating PTHrP levels by approximately 50% in two representative models. However, the mechanism underlying the antihypercalcemic effect of OCT seemed complex, involving inhibition of PTHrP production, suppression of excessive bone resorption, and an antitumor activity. OCT also markedly inhibited the body weight loss with tumor growth, while AHPrBP, which exhibited a similar antihypercalcemic effect, was less effective than OCT in preventing cachexia. The anticachectic activity of their combination did not exceed that of OCT alone, suggesting a hypercalcemia-dependent as well as an independent mechanism of cancer cachexia. It is concluded that OCT may be useful, either as a single agent or in combination with bisphosphonates, for the treatment of cancer-associated hypercalcemia and cachexia.
An Sp1 Binding Site Polymorphism in the COLIA1 Gene Predicts Osteoporotic Fractures in Both Men and Women
Authors: Langdahl Bente L. , Ralston Stuart H. , Grant Struan F. A. , Eriksen Erik F.
Published: April 2006

Genetic factors play an important role in the pathogenesis of osteoporosis, and recent studies have shown that a polymorphic Sp1 binding site in collagen type I α1 (COLIA1) gene is associated with bone mass and vertebral fractures in women from the U.K. Information on the predictive value of the COLIA1 Sp1 polymorphism in other populations is limited, however, and no studies have yet been performed in osteoporotic males. In view of this, we analyzed COLIA1 genotypes in relation to bone density and biochemical markers of bone turnover and the presence of osteoporotic fractures in a case-control study of Danish men and women. COLIA1 genotype was determined by polymerase chain reaction analysis of genomic DNA extracted from peripheral blood samples and related to bone mass, biochemical markers of bone turnover, and the presence of fracture in a study of 375 osteoporotic vertebral fracture patients and normal controls. There was no significant effect of COLIA1 genotype on bone mass or biochemical markers when data from the control group (n = 195) and fracture group (n = 180) were analyzed separately. However, the genotype distribution was significantly different in the fracture cases compared with age-matched controls (χ2 = 16.48, n = 249, p = 0.0003) due mainly to over-representation of the ss genotype in the fracture patients (14.3% vs. 1.4%), equivalent to an odds ratio for vertebral fracture of 11.83 (95% confidence interval 2.64–52.97) in those with the ss genotype. Similar differences in genotype distribution between osteoporotic patients and controls were observed in both men (χ2 = 11.52, n = 95, p = 0.0032, OR = 2.04) and women (χ2 = 6.90, n = 154, p = 0.032, OR = 1.37). In keeping with the above, logistic regression analysis showed that the ss genotype was an independent predictor of osteoporotic fracture (p = 0.028). This study confirms that the COLIA1 Sp1 polymorphism is significantly associated with osteoporotic vertebral fractures. The association is seen in both men and women, and the effect on fracture risk appears to be partly independent of bone mineral density. Our results raise the possibility that genotyping at the Sp1 site could be of clinical value in identifying individuals at risk of osteoporotic fractures in both genders.
Thyroid Hormone Inhibits Growth and Stimulates Terminal Differentiation of Epiphyseal Growth Plate Chondrocytes
Authors: Ishikawa Yoshinori , Genge Brian R. , Wuthier Roy E. , Wu Licia N. Y.
Published: April 2006

As a continuation of our studies on mineralization in epiphyseal growth plate (GP) chondrocyte cultures, the effects of tri-iodothyronine (T3) in both β-glycerophosphate-containing, serum-free (HL-1) and β-glycerophosphate-free, serum-containing medium (DATP5) were studied. The GP cells responded to T3 in a serum-, stage-, and dosage-dependent manner. Added at graded levels (0.1–10.0 nM) to preconfluent cultures (from day 7) in both HL-1 and DATP5, T3 caused progressive decreases in protein, collagen, and DNA synthesis but increased mineral deposition. In postconfluent cultures, these effects of T3 were generally muted. In preconfluent cultures, proteoglycan (PG) levels were not significantly affected in DATP5, although in HL-1 they were decreased by ∼50%. In postconfluent cultures, T3 increased PG levels in DATP5 but had no effect in HL-1. In HL-1, alkaline phosphatase (ALP) activity was progressively increased by 200–500% in both pre- and postconfluent cultures. In DATP5 in preconfluent cultures, T3 initially stimulated but later suppressed ALP; in postconfluent cultures, T3 also transiently increased ALP but did not suppress activity upon longer exposure. The inhibitory effects of T3 on protein, PG, and DNA levels of GP chondrocytes suggest that in vivo its effects on bone growth must occur primarily after cellular proliferation. Apparently by binding to the 50 kDa thyroxine-binding globulin, which cannot penetrate the PG barrier, accessibility of T3 to GP chondrocytes is limited until the time of vascular penetration when its stimulatory effects on ALP and mineral deposition become critical for continued bone development.