bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth Hormone Treatment in Adults with Adult-Onset Growth Hormone Deficiency Increases Iliac Crest Trabecular Bone Turnover: A 1-Year, Double-Blind, Randomized, Placebo-Controlled Study
Authors: Brixen Kim , Hansen Tony Bill , Hauge Ellen , Vahl Nina , Jørgensen Jens Otto Lunde , Christiansen Jens Sandahl , Mosekilde Leif , Hagen Claus , Melsen Flemming
Published: April 2006

The effects of growth hormone (GH) substitution on bone metabolism were evaluated by dynamic histomorphometry on iliac crest bone biopsies. Twenty-nine patients, aged 21–61 years (mean 45.5 years), with adult-onset GH deficiency (GHD) were randomized to receive subcutaneous injections with GH (2 IU/m2/day = 0.67 mg/m2/day) or placebo for 12 months. Serum insulin-linke growth factor I (IGF-I) levels increased 263 ± 98% (mean ± SD) during GH treatment (p < 0.0001). In the GH group, osteoid surface increased during treatment from 11% (3–15%) (median [25–75 percentiles]) to 21% (10–27%) (p = 0.01) and mineralizing surface from 4% (1–8%) to 11% (7–16%) (p = 0.04). Moreover, erosion surface tended to increase in the GH group from 2% (1–3%) to 4% (3–5%) (p = 0.07). The quiescent surface decreased in the GH group from 87% (83–96%) to 74% (68–87%) (p = 0.01). The adjusted appositional rate, mineral apposition rate, bone formation rate, bone erosion rate, mineralization lag time, and osteoid thickness remained unchanged during treatment Erosion depth showed a trend toward increase in the GH group (p = 0.09), whereas wall thickness was unchanged. Bone balance at the remodeling unit level and activation frequency were unchanged. At the tissue level, bone erosion rate increased significantly from 26% (17–36%)/year to 39% (23–72%)/year (p = 0.03). Similarly, the bone formation rate at the tissue level tended to increase, from 24% (15–31%)/year to 36% (17%-63%)%/year (p = 0.06). Finally, bone balance at the tissue level decreased significantly from 1% (–2–2%)/year to −5% (−13–1%)/year (p = 0.01). No significant difference in change was seen in the cancellous bone volume. We conclude that 12 months of GH substitution therapy increases trabecular bone turnover. Moreover, our data suggest that bone balance at the bone multicellular unit level is not changed to positive. (J Bone Miner Res 2000;15:293–300)
Study of the Nonresorptive Phenotype of Osteoclast-like Cells from Patients with Malignant Osteopetrosis: A New Approach to Investigating Pathogenesis
Authors: Flanagan Adrienne M. , Sarma Usha , Steward Colin G. , Vellodi Ashok , Horton Michael A.
Published: April 2006

Osteopetrosis manifests as failure of osteoclastic bone resorption. The cause of the disease lies either in the hematopoietic lineage or in the bone marrow stromal microenvironment. It has not been possible to define the cell type involved in the various forms of the human disease because of the inability to form human osteoclasts in vitro. Using the recently described method for generating human osteoclasts from peripheral blood in coculture with rat osteoblastic UMR 106 cells, we demonstrate that a defect lies in the mature osteoclast-like cells in four cases of this disease. Control and osteopetrotic cocultures generated large numbers of osteoclast-like cells (calcitonin and vitronectin receptor positive, and F-actin ring–positive cells) with similar morphology. Bone resorption did not occur in three of the four osteopetrotic cultures. In case 1, in which bone resorption was identified, the area of resorption was negligible compared with the number of osteoclast-like cells in the culture and was detected only by scanning electron microscopy. In contrast, up to 20% of the bone surface in controls was resorbed. The normal and osteopetrotic osteoclast-like cells had a similar phenotype except that two of the osteopetrotic cases did not express CD44 and two expressed CD44 weakly, whereas CD44 was strongly expressed in the controls. This study shows that it is possible to reproduce in vitro the pathological features of human osteopetrosis, and the assay provides a means of acquiring a greater understanding of the pathogenesis of human osteopetrosis. (J Bone Miner Res 2000;15:352–360)
Genetic Epidemiological Approaches to the Search for Osteoporosis Genes
Authors: Nguyen T. V. , Blangero J. , Eisman J. A.
Published: April 2006

Important progress has been made in the identification of specific environmental factors and estimation of hereditary components in bone density, quantitative ultrasound (QUS), and bone turnover indices. By contrast, the search for specific genes that regulate bone mass has progressed rather slowly, and the results are more difficult to interpret and reproduce. This article reviews the genetics of osteoporosis and problems plaguing genetic research. It is argued that the search for genes involved in the expression of osteoporotic phenotypes should be based on linkage studies in relatively homogeneous populations. Strategies for increasing the power of studies, such as making use of information from extended pedigrees and multivariate analysis, are discussed. With the advent of a comprehensive human genetic linkage map, a complete identification of genes for osteoporosis appears feasible. Understanding the genetic mechanisms and their interactions with environmental factors should allow more focused and cost-effective osteoporosis prevention and treatment strategies.
Prevalence of Pelvic Paget's Disease of Bone in the United States
Authors: Altman R. D. , Bloch D. A. , Hochberg M. C. , Murphy W. A.
Published: April 2006

The objective of this article was to estimate the prevalence of Paget's disease of bone in the United States from a statistically derived sample of the general population. Pelvic radiographs obtained in the First National Health and Nutrition Examination Survey (NHANES-I) were reviewed for the presence of Paget's disease. Age, sex, and geographic distribution of Paget's disease of the pelvic region were determined. The overall prevalence of Paget's disease in the United States was estimated. Pelvic Paget's disease is estimated to be present in 0.71 + 0.18% of the radiographs of the general population. The disease was higher in frequency in people who were in the older decades of life with the highest prevalence of 2.32 + 0.54% in the 65- to 74-year-old people. There is a slight male predominance in the 45- to 74-year age group. The regional distribution suggests the highest prevalence in the Northeast (1.48 + 0.52%) with the lowest prevalence in the South (0.26 + 0.25%). The prevalence was equal in white people and black people. An estimate of the overall prevalence of Paget's disease in the United States was at least 1% and perhaps as much as 2% of the general population with near equal sex distribution and the highest prevalence in the northeastern United States.
Decreased Nitric Oxide Levels Stimulate Osteoclastogenesis and Bone Resorption Both in Vitro and in Vivo on the Chick Chorioallantoic Membrane in Association with Neoangiogenesis
Authors: Collin-Osdoby Patricia , Rothe Linda , Bekker Simon , Anderson Fred , Osdoby Philip
Published: April 2006

High nitric oxide (NO) levels inhibit osteoclast (OC)-mediated bone resorption in vivo and in vitro, and nitrate donors protect against estrogen-deficient bone loss in postmenopausal women. Conversely, decreased NO production potentiates OC bone resorption in vitro and is associated with in vivo bone loss in rats and humans. Previously, we reported that bone sections from rats administered aminoguanidine (AG), a selective inhibitor of NO production via inducible NO synthase, exhibited both increased OC resorptive activity as well as greater numbers of OC. Here, we investigated further whether AG promoted osteoclastogenesis, in addition to stimulating mature OC function, using a modified in vivo chick chorioallantoic membrane (CAM) system and an in vitro chick bone marrow OC-like cell developmental model. AG, focally administered in small agarose plugs placed directly adjacent to a bone chip implanted on the CAM, dose-dependently elicited neoangiogenesis while stimulating the number, size, and bone pit resorptive activity of individual OC ectopically formed in vivo. In addition to enhancing OC precursor recruitment via neoangiogenesis, AG also exerted other vascular-independent effects on osteoclastogenesis. Thus, AG promoted the in vitro fusion and formation from bone marrow precursor cells of larger OC-like cells that contained more nuclei per cell and exhibited multiple OC differentiation markers. AG stimulated development was inversely correlated with declining medium nitrite levels. In contrast, three different NO donors each dose-dependently inhibited in vitro OC-like cell development while raising medium nitrite levels. Therefore, NO sensitively regulates OC-mediated bone resorption through affecting OC recruitment (angiogenesis), formation (fusion and differentiation), and bone resorptive activity in vitro and in vivo. Possibly, the stimulation of neoangiogenesis and OC-mediated bone remodeling via AG or other pro-angiogenic agents may find clinical applications in reconstructive surgery, fracture repair, or the treatment of avascular necrosis.
Vascular Endothelial Growth Factor Is Expressed in Human Fetal Growth Cartilage
Authors: Garcia-Ramirez Marta , Toran Nuria , Andaluz Pilar , Carrascosa Antonio , Audi Laura
Published: April 2006

Angiogenesis is a crucial event in endochondral ossification. Chemoattractants and mitogens for endothelial cells (such as basic fibroblast growth factor [bFGF] and transforming growth factor β [TGF-β]), which act as local regulators of the process, are synthesized by chondrocytes under several stimuli and in relation to the differentiation stage of the cartilage. Vascular endothelial growth factor (VEGF) is a 44-kDa protein well known as a potent angiogenic molecule owing to its mitogenic and permeability-causing properties. In this work, VEGF was located by immunohistochemistry in growth plate cartilage of human fetuses (20–22 weeks old) and its expression was demonstrated by reverse-transcription polymerase chain reaction (RT-PCR). Primary culture of human fetal epiphyseal chondrocytes (HFEC) maintained VEGF expression at protein and messenger RNA (mRNA) levels and this expression was stimulated by cartilage-promoting growth factors incorporated into the culture media (rFGF-b, rTGF-β1, and insulin-like growth factor [rIGF-I] at 50 ng/ml). The conditioned medium (CM) of HFEC stimulated the proliferation of endothelial cells, and this was partially blocked by anti-VEGF antibody. These studies showed VEGF production by chondrocytes of the epiphyseal growth cartilage and suggested a role of this factor in cartilage physiology and the angiogenic process.
A Detailed Assessment of Alterations in Bone Turnover, Calcium Homeostasis, and Bone Density in Normal Pregnancy
Authors: Black A. J. , Topping J. , Durham B. , Farquharson R. G. , Dr. Fraser W. D.
Published: April 2006

The effects of pregnancy on bone turnover and the potential risk of developing an osteoporotic fracture in pregnancy are controversial. Utilizing biochemical markers of bone formation and resorption and dual-energy X-ray absorptiometry (DEXA), bone turnover before, during, and after pregnancy was studied in detail. Ten women (mean age 30 years; range 23–40) were recruited. Prepregnancy data were obtained and then a review was performed at 2-week intervals, once pregnancy was confirmed, until 14 weeks of gestation and thereafter monthly until term. Bone mineral density (BMD) was estimated by DEXA scanning of hip, spine, and forearm preconception and postpartum. In addition, BMD of the forearm at 14 weeks and 28 weeks gestation was obtained. All pregnancies had a successful outcome. Urinary free pyridinium cross-links, free pyridinoline (fPyr) and free deoxypyridinoline (fDPyr), were normal prepregnancy (mean [±SD]) 14.6 nmol/mmol (1.8) and 5.0 nmol/mmol (1.0) creat, respectively. By 14 weeks, they had increased to 20.8 nmol/mmol (4.3) and 6.1 nmol mmol (1.4) (both p < 0.02) and by 28 weeks to 26.3 nmol/mmol (5.6) and 7.4 nmol/mmol (1.6) (both p < 0.01). The ratio of fPyr to fDPyr remained constant. A similar significant increase was observed in N-telopeptide (NTx). Bone formation was assessed by measurement of carboxy-terminal propeptide of type 1 collagen (P1CP) and bone-specific alkaline phosphatase (BSAP). Neither were altered significantly before 28 weeks, but subsequently mean P1CP increased from 110 μg/liter (23) to 235 μg/liter (84) at 38 weeks and mean BSAP increased from 11.1 U/liter (5.0) to 28.6 U/liter (11.1) (p < 0.01 for both variables). Lumbar spine (L1–L4) BMD decreased from a prepregnancy mean of 1.075 g/cm (0.115) to 1.054 g/cm2 (0.150) postpartum (p < 0.05). Total hip BMD decreased from a prepregnancy mean of 0.976 g/cm2 (0.089) to 0.941 g/cm2 (0.097) (p < 0.05). Forearm BMD at midradius, one-third distal and ultradistal decreased but did not reach statistical significance. As assessed by these bone markers, in the first 2 trimesters of pregnancy, bone remodeling is uncoupled with a marked increase in bone resorption. A corresponding increase in formation markers is not observed until the third trimester. Spinal BMD exhibits a significant decrease from prepregnancy to the immediate postpartum period with a mean reduction in BMD of 3.5% in 9 months.
The Effect of Alendronate on Bone Mass After Distal Forearm Fracture
Authors: Van Der Poest Clement E. , Patka P. , Vandormael K. , Haarman H. , Lips P.
Published: April 2006

Fracture and immobilization of an extremity lead to bone loss at the fracture and at adjacent sites. We conducted a 1-year, single-center, prospective, randomized, double-blind study to determine whether bone loss would occur in the distal radius after a Colles' fracture and whether this loss could be prevented using an antiresorptive drug (alendronate). Thirty-seven women with a recent fracture of the distal forearm and low bone mineral density (BMD) of the lumbar spine were randomized to receive either 10 mg alendronate daily or placebo. BMD of both forearms was measured at baseline and after 3, 6, and 12 months. The results of four women who developed reflex sympathetic dystrophy were not included in the analysis. In the placebo group, there was a significant reduction at 3 months and 6 months in BMD of total radius (p < 0.01), one-third distal radius (p < 0.01), middistal radius (p < 0.05), and ultradistal radius (p < 0.01) on the fractured side. The loss in BMD at one-third distal radius remained significant at month 12 (p ≤ 0.001). In the alendronate group BMD of total distal radius, one-third distal radius, and middistal radius at the fractured side remained unchanged. BMD of ultradistal radius increased significantly at months 3, 6, and 12, compared with baseline (p < 0.05). The difference between the two treatment groups was significant at 3 months and 6 months and borderline significant (p = 0.054) after 1 year in total distal radius. In ultradistal radius the differences were significant at all time points. We conclude that BMD of the distal radius of a recently fractured forearm decreases significantly in the 6 months after fracture and the resulting deficit remains evident at least 1 year after fracture. This bone loss can be prevented by alendronate.
Fractal Dimension of Trabecular Bone Projection Texture Is Related to Three-Dimensional Microarchitecture
Authors: Pothuaud L. , Benhamou C. L. , Porion P. , Lespessailles E. , Harba R. , Levitz P.
Published: April 2006

The purpose of this work was to understand how fractal dimension of two-dimensional (2D) trabecular bone projection images could be related to three-dimensional (3D) trabecular bone properties such as porosity or connectivity. Two alteration processes were applied to trabecular bone images obtained by magnetic resonance imaging: a trabeculae dilation process and a trabeculae removal process. The trabeculae dilation process was applied from the 3D skeleton graph to the 3D initial structure with constant connectivity. The trabeculae removal process was applied from the initial structure to an altered structure having 99% of porosity, in which both porosity and connectivity were modified during this second process. Gray-level projection images of each of the altered structures were simply obtained by summation of voxels, and fractal dimension (Df) was calculated. Porosity (φ) and connectivity per unit volume (Cv) were calculated from the 3D structure. Significant relationships were found between Df, φ, and Cv. Df values increased when porosity increased (dilation and removal processes) and when connectivity decreased (only removal process). These variations were in accordance with all previous clinical studies, suggesting that fractal evaluation of trabecular bone projection has real meaning in terms of porosity and connectivity of the 3D architecture. Furthermore, there was a statistically significant linear dependence between Df and Cv when φ remained constant. Porosity is directly related to bone mineral density and fractal dimension can be easily evaluated in clinical routine. These two parameters could be associated to evaluate the connectivity of the structure.
Diet- or Warfarin-Induced Vitamin K Insufficiency Elevates Circulating Undercarboxylated Osteocalcin Without Altering Skeletal Status in Growing Female Rats
Authors: Haffa A. , Krueger D. , Bruner J. , Engelke J. , Gundberg C. , Akhter M. , Binkley N.
Published: April 2006

To further characterize the skeletal role of vitamin K (K), markers of bone turnover, density, and strength were evaluated in rats with diet- or warfarin (W)-induced K insufficiency. One hundred two, 7-week-old, female rats were randomly assigned to low K (phylloquinone [K1], 20 μg/kg diet), control K (K1, 1300 μg/kg diet), low-dose W (W, 1.5 mg/kg control diet), or high-dose W plus K (W/K1, 10/100 mg/kg diet). Femur bone mineral content (BMC) and bone mineral density (BMD), plasma prothrombin time (PT) and prothrombin concentration (PC), and serum total alkaline phosphatase (ALP) and skeletal alkaline phosphatase (sALP) were measured at baseline and days 20, 40, 60, and 80. Serum total osteocalcin (OC) and undercarboxylated osteocalcin (ucOC) and femur length (FL) were measured at baseline and day 80. Left femur OC was measured and biomechanical testing of the right femur and third lumbar vertebral body was performed at day 80. Low dietary K elevated circulating ucOC (17% higher than control; p < 0.0001) at day 80. Furthermore, in both W groups, essentially all circulating OC was undercarboxylated and femur OC was lower than control (p < 0.0001). However, there was no change in femur percent ucOC, suggesting deposition of less newly synthesized OC. No between group differences were observed in PT, ALP, sALP, FL, BMC, BMD, or bone strength. In conclusion, skeletal K insufficiency can be induced by W or diet manipulation. This does not hinder peak bone mass attainment in female rats; however, W causes less newly synthesized OC to be deposited in bone. (J Bone Miner Res 2000;15:872–878)
Heterotopic Ossification of Degenerating Rat Skeletal Muscle Induced by Adenovirus-Mediated Transfer of Bone Morphogenetic Protein-2 Gene
Authors: Dr. Gonda K. , Nakaoka T. , Yoshimura K. , Otawara-Hamamoto Y. , Harrii K.
Published: April 2006

In vivo gene transfer is a recently developed device for efficient delivery of a therapeutic recombinant protein. We formulated the hypothesis that a high level of expression of bone morphogenetic protein 2 (BMP-2) could be a future therapeutic modality in terms of inducing substantial bone formation in vivo. First, to test this hypothesis, adenoviruses carrying BMP-2 gene were directly injected into the soleus muscle of adult rat. The BMP-2 gene was successfully overexpressed in the target muscle by adenovirus-mediated transfer, whereas bone formation in and around the muscle failed to occur in this case. Second, to recruit putative osteoprogenitor cells, we then induced ischemic degeneration of the target muscle by orthotopically grafting it simultaneously with the gene transfer. The combination of BMP-2 gene transfer and orthotopic muscle grafting resulted in successful ossification of almost the whole grafted muscle, whereas neither muscle grafting alone nor the combination of muscle grafting and adenovirus-mediated transfer of reporter gene LacZ induced any bone formation in the muscle. The ossification process was evident by positive von Kossa staining of the histological sections and roentgenographical radio-opacity of the region. It was also found that the BMP-2 transgene overexpressed in grafted muscles inhibited muscle regeneration, which should otherwise follow the muscle degeneration. We further demonstrated an up-regulation of BMP receptor type IA in grafted muscles, suggesting its involvement in the bone-formation process. In conclusion, overexpression of BMP-2 gene induced massive heterotopic ossification in skeletal muscles under graft-induced ischemic degeneration, which possibly up-regulates osteoprogenitor cells in situ.
Androgen Deficiency Induces High Turnover Osteopenia in Aged Male Rats: A Sequential Histomorphometric Study
Authors: Erben Reinhold G. , Eberle Johannes , Stahr Kerstin , Goldberg Michel
Published: April 2006

Hypogonadism is considered to be one of the major risk factors for osteoporosis in men. However, the mechanisms of bone loss caused by androgen deficiency are still unclear. In the present study, we sequentially investigated the skeletal and hormonal effects of androgen deficiency in aged orchiectomized (ORX) rats over a time period of 9 months. One hundred seventy 13-month-old male Fischer-344 rats were either ORX or sham-operated (SHAM). Eight rats served as baseline controls. After in vivo fluorochrome labeling, groups of 8–15 SHAM and ORX rats each were killed at 2 weeks and at 1, 2, 3, 4, 6, and 9 months postsurgery. As expected, ORX induced a fall in serum total and free testosterone levels, but also reduced serum estradiol concentrations. Cancellous bone area (BAr) in the proximal tibia but not in the first lumbar vertebral body showed an age-dependent decline in SHAM rats. Relative to SHAM controls, ORX rats had significantly reduced cancellous BAr after 2 weeks post-ORX in the tibia and after 2 months post-ORX in the vertebral body. Thereafter, vertebral and tibial cancellous BAr continued to decline in ORX animals throughout the study. Osteoclast number (NOc), osteoblast surface, bone formation rate (BFR), and activation frequency were increased in ORX animals from 1 month postsurgery until the end of the trial. Moreover, in close temporal association with the histomorphometric findings, serum osteocalcin and urinary excretion of collagen cross-links and calcium were elevated in ORX rats. In a stepwise model of multiple regression analysis using estradiol and free and total testosterone as independent variables, estradiol was the only significant predictor of histomorphometric indices of bone formation and bone resorption in SHAM and ORX rats. These data show that androgen deficiency induces substantial loss of cancellous bone in the axial and appendicular skeleton of aged male rats and that this osteopenia is associated with a sustained increase in bone turnover. Thus, the skeletal effects of androgen withdrawal in aged male rats appear to resemble those induced by estrogen withdrawal in female rats. Furthermore, our study suggests that estradiol may act as a physiological suppressor of bone remodeling in aged male rats.
Genetic Regulation of Cortical and Trabecular Bone Strength and Microstructure in Inbred Strains of Mice
Authors: Turner Charles H. , Hsieh Yeou-Fang , Müller Ralph , Bouxsein Mary L. , Baylink David J. , Rosen Clifford J. , Grynpas Marc D. , Donahue Leah Rae , Beamer Wesley G.
Published: April 2006

The inbred strains of mice C57BL/6J (B6) and C3H/HeJ (C3H) have very different femoral peak bone densities and may serve as models for studying the genetic regulation of bone mass. Our objective was to further define the bone biomechanics and microstructure of these two inbred strains. Microarchitecture of the proximal femur, femoral midshaft, and lumbar vertebrae were evaluated in three dimensions using microcomputed tomography (μCT) with an isotropic voxel size of 17 μm. Mineralization of the distal femur was determined using quantitative back-scatter electron (BSE) imaging. μCT images suggested that C3H mice had thicker femoral and vertebral cortices compared with B6. The C3H bone tissue also was more highly mineralized. However, C3H mice had few trabeculae in the vertebral bodies, femoral neck, and greater trochanter. The trabecular number (Tb.N) in the C3H vertebral bodies was about half of that in B6 vertebrae (2.8−1 ± 0.1 mm−1 vs. 5.1−1 ± 0.2 mm−1; p < 0.0001). The thick, more highly mineralized femoral cortex of C3H mice resulted in greater bending strength of the femoral diaphysis (62.1 ± 1.2N vs. 27.4 ± 0.5N, p < 0.0001). In contrast, strengths of the lumbar vertebra were not significantly different between inbred strains (p = 0.5), presumably because the thicker cortices were combined with inferior trabecular structure in the vertebrae of C3H mice. These results indicate that C3H mice benefit from alleles that enhance femoral strength but paradoxically are deficient in trabecular bone structure in the lumbar vertebrae.
Identification and Characterization of the New Osteoclast Progenitor with Macrophage Phenotypes Being Able to Differentiate into Mature Osteoclasts
Authors: Takeshita Sunao , Kaji Keisuke , Kudo Akira
Published: April 2006

Osteoclasts are thought to belong to a macrophage lineage. However, the nature of common precursors of osteoclasts and macrophages remains to be investigated. We have characterized the differentiation potential of mouse bone marrow macrophages into mature osteoclasts. Monocyte macrophage-colony-stimulating factor (M-CSF) stimulated the proliferation of bone marrow macrophages in a dose-dependent manner and these M-CSF-dependent bone marrow macrophage (MDBM) cells efficiently differentiated into the tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts in the presence of soluble RANKL (sRANKL) and M-CSF in the in vitro culture. The macrophage-like cell line TMC16 was established from tsA58 (temperature-sensitive SV40 large T-antigen) transgenic mice in the same manner to the preparation of MDBM cells and also differentiated into mature osteoclasts. During this differentiation in vitro, the morphology of the cells changed from spindle to round and smaller (termed pOC) on day 2 and to multinuclear (termed multinucleated cells [MNCs]) on day 4. The surface expression of macrophage marker CD14 was down-regulated and that of CD43 was up-regulated on pOC, analyzed by flow cytometry. RNA analysis revealed that osteoclast marker genes such as calcitonin receptor (CTR), carbonic anhydrase II (CAII), cathepsin K (cath K), MMP9, and TRAP were strongly expressed in MNCs and weakly in pOC whereas MDBM cells did not express these genes. However, the osteopontin (OPN) gene was strongly expressed in MDBM cells and this expression became weakened after differentiation into pOC. The TMC16 cell line weakly expressed cath K, TRAP, and OPN, suggesting that the TMC16 cell line is immortalized at a stage slightly differentiated from MDBM cells. Furthermore, cell sorting analysis revealed that osteoclast early progenitors in bone marrow cells are preferentially present in the Mac-1− F4/80dull population, which differentiated into MDBM cells (the osteoclast progenitor) expressing Mac-1+ F4/80int, suggesting that M-CSF plays roles of a differentiation factor as well as a growth factor for osteoclast early progenitors. These results showed the transition of morphology, surface markers, and gene expression from the early to mature stage in osteoclast differentiation. We propose three differentiation stages in the osteoclast lineage: the pro-osteoclast (spindle-shaped macrophage cells), the pre-osteoclast (small round mononucleated TRAP-positive cells), and the mature osteoclast (multinucleated TRAP-positive cells) stage.
Biochemical Markers of Bone Turnover, Endogenous Hormones and the Risk of Fractures in Postmenopausal Women: The OFELY Study
Authors: Garnero Patrick , Sornay-Rendu Elisabeth , Claustrat Bruno , Delmas Pierre D.
Published: April 2006

The mechanisms leading to increased bone loss and skeletal fragility in women with postmenopausal osteoporosis are still poorly understood. Increased bone resorption, low serum estradiol and high serum sex-hormone-binding globulin (SHBG) recently have been reported as predictors of vertebral and hip fractures in elderly women. In a cohort of healthy untreated younger postmenopausal women aged 50–89 years (mean, 64 years), we compared baseline levels of bone markers and endogenous hormones in 55 women who subsequently had a fracture (20 vertebral and 35 peripheral fractures) with levels in the 380 women who did not fracture during a mean 5 years of follow-up. Women with levels in the highest quartile of four bone resorption markers including urinary-free deoxypyridinoline (D-Pyr), urinary type I collagen N-telopeptides (NTX), and urinary and serum type I collagen C-telopeptides (CTX) had about a 2-fold increased risk of fractures compared with women with levels in the three lowest quartiles with relative risk (RR) and 95% CI of 1.8 (1.0-3.4) for free D-Pyr, 1.7 (0.9-3.2) for urinary NTX, 2.3 (1.3-4.1) for urinary CTX, and 2.1 (1.2-3.8) for serum CTX. Serum levels of bone alkaline phosphatase (BAP) in the highest quartile were associated with an RR of fracture of 2.4 (1.3-4.2). Women with serum levels of estradiol and dehydroepiandrosterone (DHEA) sulfate in the lowest quartile had an RR of fracture of 2.2 (1.2-4.0) and 2.1 (1.2-3.8), respectively. Increased levels of SHBG and intact parathyroid hormone (PTH) were moderately associated with an increased risk of fracture. Similar results were obtained when the analysis was restricted to symptomatic vertebral and nonvertebral fractures. Adjustment of biochemical markers by hormone levels did not significantly alter the results. Women with both high bone resorption markers and low estradiol (or low DHEA sulfate) had a higher risk of fracture with RRs of 3.0-3.3 (p < 0.001). After adjustment for bone mineral density (BMD) of the hip, spine, radius, or total body, bone markers and hormones were still predictive of fracture risk with similar RRs. We conclude that high levels of some biochemical markers of bone turnover, low serum estradiol, low DHEA sulfate, high SHBG, and high PTH are associated with increased risk of osteoporotic fracture in postmenopausal women, independently of each other and of BMD. The mechanism by which some postmenopausal women have an increased rate of bone turnover leading to an increased risk of fracture remains to be elucidated.
Dietary Protein Deficiency Induces Osteoporosis in Aged Male Rats
Authors: Bourrin S. , Toromanoff A. , Ammann P. , Bonjour J. P. , Rizzoli R.
Published: April 2006

Low dietary intake is common in elderly males with low femoral neck areal bone mineral density (BMD). To evaluate the selective influence of a low-protein diet in the pathogenesis of osteoporosis in males and to uncover early and late adaptation of bone cells to protein deficiency, 8-month-old male rats were pair-fed a control (15% casein) or isocaloric low-protein (2.5% casein) diet for 1 or 7 months. BMD, bone ultimate strength, stiffness, and absorbed energy were measured in tibia proximal metaphysis and diaphysis. After double-labeling, histomorphometric analysis was performed at the same sites. Serum osteocalcin, insulin-like growth factor I (IGF-I), and urinary deoxypyridinoline excretion were measured. In proximal tibia, isocaloric low-protein diet significantly decreases BMD (12%), cancellous bone mass (71%), and trabecular thickness (Tb.Th; 30%), resulting in a significant reduction in ultimate strength (27%). In cortical middiaphysis, a low-protein diet decreases BMD (9%) and enlarges the medullary cavity (36%), leading to cortical thinning and lower mechanical strength (20%). In cancellous bone, protein deficiency transiently depresses the bone formation rate (BFR; 60%), osteoid seam thickness (15%), and mineral apposition rate (MAR; 20%), indicating a decrease in osteoblast recruitment and activity. Cortical loss (15%) results from an imbalance between endosteal modeling drifts with impaired BFR (70%). From the first week of protein deficiency, osteocalcin and IGF-I levels drop significantly. Bone resorption activity and urinary deoxypyridinoline remain unchanged throughout the experiment. Protein deficiency in aged male rats induces cortical and trabecular thinning, and decreases bone strength, in association with a remodeling imbalance with a bone formation impairment and a decrease in IGF-I levels.
Consequences of Vertebral Deformities in Older Men and Women
Authors: Pluijm S. M. F. , Tromp A. M. , Smit J. H. , Deeg D. J. H. , Lips P.
Published: April 2006

The objectives of this study were to ascertain the prevalence of the number and severity of vertebral deformities in elderly people and determine the extent to which these are associated with several aspects of functioning. The study was conducted in a subsample of the Longitudinal Aging Study Amsterdam (LASA) consisting of 527 participants (260 men and 267 women), aged 65 years or over. Lateral radiographs of the spine (T4-L5) were made of each participant and a semiquantitative method was used to assess the number and degree of vertebral deformities. The prevalence of having at least one vertebral deformity was 39% in both men and women. Six percent of the men and 5% of the women had at least three vertebral deformities. For severe deformities, the prevalence was 8% in men and 12% in women. The number of vertebral deformities was significantly associated with a height loss of more than 5 cm, difficulties in activities of daily living, poor performance, more than 3 days in bed and more than 3 days with limited activities because of health problems in the past month, and poor self-perceived health. For most of these outcome measures, associations were strongest when three or more deformities were present. The presence of a severe deformity was associated with a height loss of more than 5 cm, poor performance, more than 3 days with limited activities in the past month, and poor self-perceived health. None of the associations between number and severity of vertebral deformities and the level of functioning was modified by sex. We can conclude that vertebral deformities are very common in both older men and older women and that vertebral deformities, even if they are not clinically manifest, have a substantial impact on the level of functioning and well-being of older people.
Low-Level Cadmium Exposure and Osteoporosis
Authors: Alfvén Tobias , Elinder Carl-Gustaf , Carlsson Margareta Dea , Grubb Anders , Hellström Lennart , Persson Bodil , Pettersson Conny , Spång Gunnar , Schütz Andrejs , Järup Lars
Published: April 2006

Osteoporosis is a major cause of morbidity worldwide. A number of risk factors, such as age and gender, are well established. High cadmium exposure causes renal damage and in severe cases also causes osteoporosis and osteomalacia. We have examined whether long-term low-level cadmium exposure increases the risk of osteoporosis. Bone mineral density (BMD) in the forearm was measured in 520 men and 544 women, aged 16–81 years, environmentally or occupationally exposed to cadmium, using dual-energy X-ray absorptiometry (DXA) technique. Cadmium in urine was used as the dose estimate and protein HC was used as a marker of renal tubular damage. There was a clear dose-response relation between cadmium dose and the prevalence of tubular proteinuria. Inverse relations were found between cadmium dose, tubular proteinuria, and BMD, particularly apparent in persons over 60 years of age. There was a dose-response relation between cadmium dose and osteoporosis. The odds ratios (ORs) for men were 2.2 (95% CI, 1.0-4.8) in the dose group 0.5-3 nmol Cd/mmol creatinine and 5.3 (2.0-14) in the highest dose category (≥3 nmol/mmol creatinine) compared with the lowest dose group (<0.5 nmol Cd/mmol creatinine). For women, the OR was 1.8 (0.65-5.3) in the dose group 0.5-3 nmol Cd/mmol creatinine. We conclude that exposure to low levels of cadmium is associated with an increased risk of osteoporosis.
Osteogenesis from Cultured Chick Periostea Has a Specific Requirement for Chloride
Authors: Lovitch Dinah , Christianson Michael L.
Published: April 2006

Bone development, like embryonic development in general, depends on a particular internal electrical milieu. Ions are the carriers of currents that maintain this internal environment. In embryonic bone, chloride is a major carrier of such current. To explore the role chloride plays in embryonic bone development we performed several ion-removal experiments, using the chick periosteal osteogenesis (CPO) system as our model. We found that if chloride is reduced in the medium and replaced with a nontoxic anion, alkaline phosphatase (ALP) activity does not rise, nor does osteogenic development occur. However, acid phosphatase (AP) activity is not affected by level of chloride. Experiments using metabolic inhibitors showed that explants cultured in low chloride medium remain viable. Dose-response studies revealed that the response of ALP activity to chloride concentration is sigmoidal, with a [Cl−]0.5 of 45.9 mM. Reciprocal transfers of explants between complete and low chloride medium show that the rise in ALP activity depends on the length of time explants are cultured with chloride. In contrast, such transfer experiments show that osteogenesis requires chloride only during days 2–3 of culture.
Particulate Wear Debris Activates Protein Tyrosine Kinases and Nuclear Factor κB, Which Down-Regulates Type I Collagen Synthesis in Human Osteoblasts
Authors: Vermes Csaba , Roebuck Kenneth A. , Chandrasekaran Raman , Dobai József G. , Jacobs Joshua J. , Glant Tibor T.
Published: April 2006

Particulate wear debris generated mechanically from prosthetic materials is phagocytosed by a variety of cell types within the periprosthetic space including osteoblasts, which cells with an altered function may contribute to periprosthetic osteolysis. Exposure of osteoblast-like osteosarcoma cells or bone marrow-derived primary osteoblasts to either metallic or polymeric particles of phagocytosable sizes resulted in a marked decrease in the steady-state messenger RNA (mRNA) levels of procollagen α1[I] and procollagen α1[III]. In contrast, no significant effect was observed for the osteoblast-specific genes, such as osteonectin and osteocalcin (OC). In kinetic studies, particles once phagocytosed, maintained a significant suppressive effect on collagen gene expression and type I collagen synthesis for up to five passages. Large particles of a size that cannot be phagocytosed also down-regulated collagen gene expression suggesting that an initial contact between cells and particles can generate gene responsive signals independently of the phagocytosis process. Concerning such signaling, titanium particles rapidly increased protein tyrosine phosphorylation and nuclear transcription factor κB (NF-κB) binding activity before the phagocytosis of particles. Protein tyrosine kinase (PTK) inhibitors such as genistein and the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) significantly reduced the suppressive effect of titanium on collagen gene expression suggesting particles suppress collagen gene expression through the NF-κB signaling pathway. These results provide a mechanism by which particulate wear debris can antagonize the transcription of the procollagen α1[I] gene in osteoblasts, which may contribute to reduced bone formation and progressive periprosthetic osteolysis.
Importance of Membrane- or Matrix-Associated Forms of M-CSF and RANKL/ODF in Osteoclastogenesis Supported by SaOS-4/3 Cells Expressing Recombinant PTH/PTHrP Receptors
Authors: Itoh Kanami , Udagawa Nobuyuki , Matsuzaki Kenichiro , Takami Masamichi , Amano Hitoshi , Shinki Toshimasa , Ueno Yutaka , Takahashi Naoyuki , Suda Tatsuo
Published: April 2006

SaOS-4/3, a subclone of the human osteosarcoma cell line SaOS-2, established by transfecting the human parathyroid hormone/parathyroid hormone-related protein (PTH/PTHrP) receptor complementary DNA (cDNA), supported osteoclast formation in response to PTH in coculture with mouse bone marrow cells. Osteoclast formation supported by SaOS-4/3 cells was completely inhibited by adding either osteoprotegerin (OPG) or antibodies against human macrophage colony-stimulating factor (M-CSF). Expression of messenger RNAs (mRNAs) for receptor activator of NF-κB ligand/osteoclast differentiation factor (RANKL/ODF) and both membrane-associated and secreted forms of M-CSF by SaOS-4/3 cells was up-regulated in response to PTH. SaOS-4/3 cells constitutively expressed OPG mRNA, expression of which was down-regulated by PTH. To elucidate the mechanism of PTH-induced osteoclastogenesis, SaOS-4/3 cells were spot-cultured for 2 h in the center of a culture well and then mouse bone marrow cells were uniformly plated over the well. When the spot coculture was treated for 6 days with both PTH and M-CSF, osteoclasts were induced exclusively inside the colony of SaOS-4/3 cells. Osteoclasts were formed both inside and outside the colony of SaOS-4/3 cells in coculture treated with a soluble form of RANKL/ODF (sRANKL/sODF) in the presence of M-CSF. When the spot coculture was treated with sRANKL/sODF, osteoclasts were formed only inside the colony of SaOS-4/3 cells. Adding M-CSF alone failed to support osteoclast formation in the spot coculture. PTH-induced osteoclast formation occurring inside the colony of SaOS-4/3 cells was not affected by the concentration of M-CSF in the culture medium. Mouse primary osteoblasts supported osteoclast formation in a similar fashion to SaOS-4/3 cells. These findings suggest that the up-regulation of RANKL/ODF expression is an essential step for PTH-induced osteoclastogenesis, and membrane- or matrix-associated forms of both M-CSF and RANKL/ODF are essentially involved in osteoclast formation supported by osteoblasts/stromal cells.
Retracted: Effect of Deconditioning on Cortical and Cancellous Bone Growth in the Exercise Trained Young Rats
Authors: Iwamoto Jun , Yeh James K. , Aloia John F.
Published: April 2006

Exercise enhances bone growth and increases peak bone mass. The aim of this study was to determine whether or not 4 weeks of deconditioning after 8 weeks of exercise in growing rats would result in a decrease in bone gain or reverse the benefits of exercise. Fifty 4-week-old female Sprague-Dawley rats were randomized by a stratified weight method into 5 groups with 10 rats in each group: 8 weeks exercise (8EX), 8 weeks sedentary control (8S), 12 weeks exercise (12EX), 8 weeks exercise followed by 4 weeks sedentary (8EX4S), and 12 weeks sedentary control (12S). The exercise consisted of running on a treadmill with a 5° slope at 24 m/minute for 1 h/day and 5 days/week. After each period of exercise, cancellous and cortical bone histomorphometry were performed on double fluorescent labeled 5-μm-thick sections of the proximal tibia and 40-μm-thick sections of the tibial shaft, respectively. Eight and 12 weeks of exercise resulted in a significant increase in the body weight and gastrocnemius muscle weight by two-way analysis of variance (ANOVA). The femoral wet weight (mg; mean ± SD; 8EX, 781 ± 45.1 vs. 8S, 713 ± 40.5; p < 0.05; 12EX, 892 ± 41.6 vs. 12S, 807 ± 19.8; p < 0.05) was significantly higher in the exercise group than that in the respective control groups. The femoral wet weight and bone volume (BV) of the 8EX4S group (818 ± 46.2 mg and 531 ± 31.2 μl, respectively) were significantly lower than those of the 12EX group (p < 0.05) and did not differ significantly from those of the 12S groups. The cancellous BV was significantly higher in the 8EX and 12EX groups than that in the respective sedentary groups (p < 0.05). The cortical bone area of the tibial shaft was also significantly higher in the 12EX than that in the 12S group (p < 0.05). The increase in the cancellous BV or cortical bone area was caused by an increase in the mineral apposition rate (MAR), without a significant effect in the labeled perimeter. The bone formation rate (BFR; μm3/μm2 per day) in the cancellous bone (12EX, 27.9 ± 7.74 vs. 12S, 15.4 ± 4.56; p < 0.05) or periosteal surface (12EX, 127.6 ± 27.7 vs. 12S, 79.5 ± 18.6; p < 0.05) was significantly higher in the exercised groups than that in the respective control group (p < 0.05). Again, deconditioning resulted in a decrease in the cancellous BFR, BV, periosteal BFR, and cortical bone area to levels not significantly different from the 12S group. In conclusion, our findings showed that exercised growing rats, when deconditioned, lost the benefits gained through exercise and their bone parameters were reduced to levels not different from the sedentary control. Thus, continued exercise is required to maintain high bone mass.
Reconciling Quantitative Ultrasound of the Calcaneus with X-Ray-Based Measurements of the Central Skeleton
Authors: Ayers Michael , Prince Michelle , Ahmadi Sadri , Baran Daniel T.
Published: April 2006

Osteoporosis is frequently undiagnosed before fracture because of the lack of availability of instruments to quantitate bone mass. To evaluate the utility of quantitative ultrasound (QUS) of the calcaneus to diagnose osteoporosis, we determined bone mineral density (BMD) of the posterior-arterior spine, total hip, and femoral neck by dual-energy X-ray absorptiometry (DXA) and QUS in 312 women aged 50 years and older. A risk factor assessment (simple calculated osteoporosis risk estimation [SCORE]) also was quantitated in all women. Ninety-four of the 312 women were diagnosed as osteoporotic based on T scores ≤ −2.5 at the spine, total hip, and/or femoral neck. The sensitivity of the individual central sites for the diagnosis of osteoporosis was 49% at the spine (46 of 94 women), 32% at the total hip (30 of 94 women), and 81% at the femoral neck (76 of 94 women). At a QUS T score ≤ −1, the peripheral technique had a sensitivity of 62% and a specificity of 72%. Combining a QUS T score of ≤ −1 followed by a risk factor assessment of women with a QUS T score ≥ −0.99 using a cut point of 11 increased sensitivity to 81% (comparable with femoral neck DXA) but decreased specificity to 58%. If peripheral QUS measurements and risk factor assessment are the only tools employed before initiation of therapy, the benefits of increased ease of diagnosis will need to be balanced against potentially unnecessary treatment in some normal patients and lack of treatment in some osteoporotic patients.