bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression and Regulation of RAB3 Proteins in Osteoclasts and Their Precursors
Authors: Abu-Amer Yousef , Teitelbaum Steven L. , Chappel Jean C. , Schlesinger Paul , Ross F. Patrick 
Published: April 2006

Abstract
The ruffled membrane, the resorptive organelle of the osteoclast, is generated by fusion of intracytoplasmic acidifying vesicles with the plasma membrane, an event analogous to regulated exocytosis. While the ruffled membrane is essential to the bone resorptive process, the mechanisms governing its generation are unknown. However, regulated exocytosis is mediated, in part, by isoforms of the Rab3 subset of Rab GTPases. Because of similarities between exocytosis and ruffled membrane formation, we asked if Rab3 proteins are expressed by osteoclasts or their precursors, and if so, are these molecules regulated by agents known to prompt the osteoclast phenotype? We find murine osteoclast precursors, in the form of bone marrow macrophages (BMMs), express at least two Rab3 isoforms, namely A and B/C, which are individually enhanced by a variety of hematopoietic cytokines. Consistent with the osteoclastogenic properties of a number of these cytokines, differentiation of BMMs into osteoclasts, in vitro, is associated with increased expression of both isoforms, particularly Rab3B/C. Finally, Rab3B/C localizes with the avian osteoclast H+ATPase (vacuolar proton pump) and pp60c–src, both intracellularly and within acidifying vesicles derived largely from the ruffled membrane. Thus, expression of specific rab3 proteins, an event which may control formation of the osteoclast ruffled membrane, is modulated by cytokines during osteoclastogenesis.
Oligodeoxynucleotide Targeted to the αv Gene Inhibits αv Integrin Synthesis, Impairs Osteoclast Function, and Activates Intracellular Signals to Apoptosis
Authors: Villanova Ida , Townsend Paul A. , Uhlmann Eugen , Knolle Jochen , Peyman Anusch , Amling Michael , Baron Roland , Horton Michael A. , Teti Anna 
Published: April 2006

Abstract
The αv integrin subunit is highly expressed in osteoclasts where it dimerizes with β1 and β3 subunits to form receptors for vitronectin and bone sialoproteins. Inhibition of osteoclast adhesion and function has previously been achieved by αvβ3 antibodies or Arg-Gly-Asp–containing peptides which have the disadvantages of blocking a single receptor type, or of being rather nonspecific, respectively. Here we show that αv integrin expression in rabbit osteoclasts can be inhibited by partially phosphorothioated antisense oligodeoxynucleotide (ODN) spanning the adenine-uracil-guanine (AUG) translational start site of the human/rabbit αv gene, a procedure which offers the advantage of affecting all the αv receptors with high efficiency. The αv antisense ODN caused a dose-dependent, substrate-specific reduction of osteoclast adhesion and bone resorption. Control ODNs, such as sense, inverted, and mismatch, were without effect, providing evidence of specificity of the antisense reagent. It is likely as a consequence of loss of substrate interaction, the antisense ODN induced osteoclast retraction and apoptosis, increase of the cyclin/cyclin-dependent kinase complex inhibitor p21WAF1/CIP1, and inhibition of the cell survival gene, bcl-2. Although the expression of the cell death–promoting gene, bax, remained unchanged, a reduction of the bcl-2/bax ratio, known to underlie the intracellular signal to apoptosis, was observed. This finding led us to hypothesize that these changes could provide a link between reduction of αv synthesis and osteoclast programmed death. In conclusion, this study provides novel insights into the use of αv antisense ODN as an efficacious mechanism for blocking osteoclast function and underscores for the first time the involvement of integrins in bone cell apoptosis. In vivo studies may verify potential application of this ODN as alternative therapy for bone diseases.
Determination of Bone Markers in Pycnodysostosis: Effects of Cathepsin K Deficiency on Bone Matrix Degradation
Authors: Nishi Yoshikazu , Atley Lynn , Eyre David E. , Edelson Jacob G. , Superti-Furga Andrea , Yasuda Toshiyuki , Desnick Robert J. , Gelb Bruce D. 
Published: April 2006

Abstract
Pycnodysostosis (Pycno) is an autosomal recessive osteosclerotic skeletal dysplasia that is caused by the markedly deficient activity of cathepsin K. This lysosomal cysteine protease has substantial collagenase activity, is present at high levels in osteoclasts, and is secreted into the subosteoclastic space where bone matrix is degraded. In vitro studies revealed that mutant cathepsin K proteins causing Pycno did not degrade type I collagen, the protein that constitutes 95% of organic bone matrix. To determine the in vivo effects of cathepsin K mutations on bone metabolism in general and osteoclast-mediated bone resorption specifically, several bone metabolism markers were assayed in serum and urine from seven Pycno patients. Two markers of bone synthesis, type I collagen carboxy-terminal propeptide and osteocalcin, were normal in all Pycno patients. Tartrate-resistent acid phosphatase, an osteoclast marker, was also normal in these patients. Two markers that detect type I collagen telopeptide cross-links from the N and C termini, NTX and CTX, respectively, were low in Pycno. A third marker which detects a more proximal portion of the C terminus of type I collagen in serum, ICTP, was elevated in Pycno, a seemingly paradoxical result. The finding of decreased osteoclast-mediated type I collagen degradation as well as the use of alternative collagen cleavage sites by other proteases, and the accumulation of larger C-terminal fragments containing the ICTP epitope, established a unique biochemical phenotype for Pycno.
Expression Profiles of mRNAs for Osteoblast and Osteoclast Proteins as Indicators of Bone Loss in Mouse Immobilization Osteopenia Model
Authors: Rantakokko Juho , Uusitalo Hannele , Jämsä Timo , Tuukkanen Juha , Aro Hannu T. , Vuorio Eero 
Published: April 2006

Abstract
An experimental mouse model for disuse osteopenia was developed using unilateral cast immobilization. Analysis of the distal femurs and proximal tibias by quantitative histomorphometry revealed significant osteopenia within 10–21 days of immobilization. At 3 weeks, bone loss was also demonstrated with peripheral quantitative computed tomography as diminished bone mineral content and as concomitant reduction in the cross-sectional moment of inertia. These structural and geometrical alterations resulted in decreased strength of the distal femurs tested by cantilever bending. Analysis of the underlying cellular and molecular mechanisms of bone loss revealed a rapid increase in bone resorption within 3 days of immobilization. The mRNA levels for cathepsin K, matrix metalloproteinase-9, and tartrate resistant acid phosphatase were all significantly increased during the 21-day immobilization period, but with different expression profiles. These increases were paralleled by an increased number of osteoclasts as measured by histomorphometry. By day 6 of immobilization, the balance of bone turnover was further shifted toward net bone loss as the mRNA levels for major bone components (type I collagen and osteocalcin) were decreased. In histomorphometric analysis this was observed as reduced rates of mineral apposition and bone formation after 10 days of immobilization. The results of this study demonstrate that immobilization has a dual negative effect on bone turnover involving both depressed bone formation and enhanced bone resorption.
Cross-Sectional Analysis of Renal Transplantation Osteoporosis
Authors: Parker Cornelle R. , Freemont Anthony J. , Blackwell Penny J. , Grainge Matthew J. , Hosking David J. 
Published: April 2006

Abstract
We report a cross-sectional study of 54 adult female renal transplant recipients. We measured bone mineral density (BMD) of the lumbar spine, femoral neck, total hip, and mid- and total radius, and 38 patients underwent transiliac crest bone biopsy. Osteopenia was widespread with 31/54 (57%) of patients osteoporotic at one or more sites. Seventeen out of 54 (32%) of the patients had a prevalent low-trauma fracture. There was a clear trend in BMD reduction across spine, hip and midradius, with the predominantly cortical midradial site showing the greatest loss. We found no relationship between BMD and body mass index, parathyroid hormone (PTH), dose of immunosuppressant, years since transplantation, age at menopause, or years since menopause. Histologically, abnormal biopsies could be classified into three categories: hyperparathyroid (n = 20), adynamic (n = 14), and osteomalacic (n = 2). Mean PTH was lower (p = NS) and mean cumulative prednisolone dose was higher (p = 0.04) in the adynamic group compared with the hyperparathyroid group, but because of overlap between groups neither was an effective discriminator of histology. We suggest that bone biopsy is indicated in these patients to direct appropriate treatment. At the cellular level, there were significant negative correlations between osteoclast function (eroded surface, r = 0.47, p = 0.003) and osteoblast numbers (osteoblast surface, r = –0.40, p = 0.01) and cumulative exposure to prednisolone. We postulate that suppression of osteoblast function by prednisolone with unopposed bone resorption may result in relative hypercalcaemia and low PTH. This progressive reduction in bone turnover may promote or prolong the adynamic state.
Monitoring Skeletal Changes by Radiological Techniques
Authors: Glüer Claus-C. 
Published: April 2006

Abstract
The longitudinal sensitivity of a technique, i.e., its ability to monitor skeletal changes, is affected by two parameters: the long-term precision error (PElt) and the subject group-specific response rate (i.e., annual rates of change). Both need to be considered to avoid misinterpretation of measured changes. A new concept to aid clinical decision making for longitudinal measurements is proposed which is based on three types of measures: criteria for detecting changes—the “least significant change” (LSC) is the smallest change to be considered statistically significant, but for certain clinical questions a smaller margin, the “trend assessment margin” (TAM), can be sufficient for decision making; follow-up time intervals—for follow-up exams the patient should be called in at about the time interval specified by the (population specific) “monitoring time interval” (MTI) or, about one-third of the time earlier, after the “trend assessment interval” (TAI), depending on whether the decision can be based on the LSC or the TAM; and the standard precision error (stdPE)—the smaller stdPE, the more sensitive the technique to monitor skeletal changes. Together, these three measures yield a good characterization of a technique's ability to monitor skeletal changes. Compared with previous concepts, the proposed standardization by a response ratio instead of measures of spread or response rates makes the stdPE substantially less subject group dependent. It allows comparison of stdPE across different studies and could replace the misleading concept of expressing precision as a coefficient of variation. Application of this concept should facilitate the interpretation of measured skeletal changes.
Longitudinal Changes of Bone Density and Bone Resorption in Hyperthyroid Girls During Treatment
Authors: Mora Stefano , Weber Giovanna , Marenzi Karen , Signorini Elisabetta , Rovelli Rosanna , Proverbio Maria Carla , Chiumello Giuseppe 
Published: April 2006

Abstract
Low bone mineral density (BMD) and increased bone turnover are common features of untreated hyperthyroidism in adult patients. The effect of treatment on BMD is still controversial. BMD and bone metabolism in hyperthyroid children have not been thoroughly investigated. In the present study, we measured spinal and whole body BMD by dual-energy X-ray absorptiometry in a group of 13 girls (aged 5.0–14.9 years) at diagnosis of hyperthyroidism. The bone resorption rate was assessed by urine measurement of N-terminal telopeptide of type I collagen (NTX). Hyperthyroid patients have been studied longitudinally during treatment. BMD values and NTX urine concentrations have been also determined in 155 healthy Caucasian girls (aged 2.4–24.2 years). Spinal and whole body bone density measurements were significantly lower compared with healthy controls in untreated hyperthyroid girls, after correction for differences in age and anthropometric measurements (p ≤ 0.033). Bone density measurements obtained after 12 and 24 months of treatment were no longer different from those of healthy girls. NTX urine levels at diagnosis of thyrotoxicosis were significantly higher than those found in healthy controls (p &lt; 0.0001); 6 months after treatment, the urine levels did not show significant differences, and they remained stable after 12 and 24 months of therapy. Inverse correlations at diagnosis were found between serum-free thyroxine (FT4) serum levels and spinal (r = –0.42) and whole body bone density (r = –0.41); FT4 and free triiodothyronine serum levels directly correlated with the NTX concentration (r = 0.77, and r = 0.71, respectively). In conclusion, the results of the present study demonstrate that low bone density values and high bone resorption rates are found in hyperthyroid children and adolescents at diagnosis of the disease. Our data also demonstrate that antithyroid treatment is able to reduce dramatically the bone resorption and to increase significantly both spinal and total body BMD, granting physiologic conditions for the achievement of the best obtainable peak bone mass.
Coordinated Maturational Regulation of PHEX and Renal Phosphate Transport Inhibitory Activity: Evidence for the Pathophysiological Role of PHEX in X-Linked Hypophosphatemia
Authors: Nesbitt Teresa , Fujiwara Ikuma , Thomas Ronald , Xiao Zhou-Sheng , Quarles L. Darryl , Drezner Marc K. 
Published: April 2006

Abstract
The mechanism by which inactivating mutations of PHEX (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) cause X-linked hypophosphatemia remains unknown. However, recent reports suggest errant PHEX activity in osteoblasts may fail to inactivate a phosphaturic factor produced by these cells. To test this possibility, we examined coordinated maturational expression of PHEX and production of phosphate transport inhibitory activity in osteoblasts from normal and hyp-mice. We assessed the inhibitory activity in conditioned medium by examining the effects on opossum kidney cell phosphate transport and osteoblast PHEX expression by reverse transcriptase-polymerase chain reaction during a 17-day maturational period. Inhibitory activity increased as a function of osteoblast maturational stage, with no activity after 3 days and persistent activity by 6 days of culture. More significantly, equal phosphate transport inhibitory activity in conditioned medium from normal and hyp-mouse osteoblasts (control 1.90 ± 0.12, normal 1.48 ± 0.10, hyp 1.45 ± 0.04 nmol/mg of protein/minute) was observed at 6 days. However, by 10 days hyp-mouse osteoblasts exhibited greater inhibitory activity than controls, and by 17 days the difference in phosphate transport inhibition maximized (control 2.08 ± 0.09, normal 1.88 ± 0.06, hyp 1.58 ± 0.06 nmol/mg of protein/minute). Concurrently, we observed absent PHEX expression in normal osteoblasts after 3 days, limited production at 6 days, and significant production by day 10 of culture, while hyp-mouse osteoblasts exhibited limited PHEX activity secondary to an inactivating mutation. The data suggest that the presence of inactivating PHEX mutations results in the enhanced renal phosphate transport inhibitory activity exhibited by hyp-mouse osteoblasts.
Atherogenic Diet and Minimally Oxidized Low Density Lipoprotein Inhibit Osteogenic and Promote Adipogenic Differentiation of Marrow Stromal Cells
Authors: Parhami Farhad , Jackson Simon M. , Tintut Yin , Le Vien , Balucan Jennifer P. , Territo Mary , Demer Linda L. 
Published: April 2006

Abstract
In osteoporosis, the bone marrow stroma osteogenic cell population declines and adipocyte numbers increase. We recently showed that oxidized lipids inhibit differentiation of preosteoblasts. In this report, we assess the effect of minimally oxidized low density lipoprotein (MM-LDL) on osteoblastic differentiation of murine marrow stromal cells, M2–10B4. MM-LDL, but not native LDL, inhibited stromal cell osteoblastic differentiation as demonstrated by inhibition of alkaline phosphatase activity, collagen I processing, and mineralization, through a mitogen-activated protein kinase–dependent pathway. In addition, marrow stromal cells from C57BL/6 mice fed a high fat, atherogenic diet failed to undergo osteogenic differentiation in vitro. The ability of MM-LDL to regulate adipogenesis was also assessed. Treatment of M2–10B4 as well as 3T3-L1 preadipocytes with MM-LDL, but not native LDL, promoted adipogenic differentiation in the presence of peroxisome proliferator-activated receptor (PPAR) γ agonist thiazolidinediones, BRL49653 and ciglitizone. Based on promoter-reporter construct experiments, MM-LDL may be acting in part through activating PPARα. These observations suggest that LDL oxidation products promote osteoporotic loss of bone by directing progenitor marrow stromal cells to undergo adipogenic instead of osteogenic differentiation. These data lend support to the “lipid hypothesis of osteoporosis.”
Mechanisms of Osteoclast Dysfunction in Human Osteopetrosis: Abnormal Osteoclastogenesis and Lack of Osteoclast-Specific Adhesion Structures
Authors: Teti Anna , Migliaccio Silvia , Taranta Anna , Bernardini Silvia , De Rossi Giulio , Luciani Matteo , Iacobini Metello , De Felice Lidia , Boldrini Renata , Bosman Cesare , Corsi Alessandro , Bianco Paolo 
Published: April 2006

Abstract
Osteoclasts from a patient affected by osteopetrosis were examined in vivo and in vitro. Iliac crest biopsy revealed an osteosclerotic pattern, with prominent numbers of osteoclasts noted for hypernuclearity and incomplete adherence to the bone surface. A population comprising tartrate-resistant acid phosphatase (TRAP)-positive, multinucleated and mononuclear cells, and alkaline phosphatase-positive stromal fibroblasts was obtained in vitro from bone marrow. Mononuclear TRAP-positive precursors spontaneously fused in culture to form giant osteoclast-like cells. These cells expressed the osteoclast marker MMP-9 and calcitonin receptor, and lacked the macrophage marker, Fc receptor. Expression and distribution of c-src, c-fms, and CD68, and response to steroid hormones relevant to osteoclast differentiation and function were apparently normal, whereas cell retraction in response to calcitonin was impaired. TRAP-positive multinucleated cells did not form osteoclast-specific adhesion structures (clear zone, podosomes, or actin rings). Bone resorption rate was severely reduced in vitro. Focal adhesions and stress fibers were observed en lieu of podosomes and actin rings. Adhesion structures contained low levels of immunoreactive vitronectin receptor, most of this integrin being retained in cytoplasmic vesicles. These data provide the first characterization of abnormal differentiation and function of human osteopetrotic osteoclast-like cells.
Vacuolar H+-ATPase Activity and Expression in Mouse Bone Marrow Cultures
Authors: Lee Beth S. , Holliday L. Shannon , Krits Irina , Gluck Stephen L. 
Published: April 2006

Abstract
We examined vacuolar H+-ATPase (V-ATPase) structure, enzymatic properties, and protein and mRNA expression from mouse marrow cultured in the presence or absence of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), which stimulates formation of bone-resorptive osteoclasts. V-ATPases from osteoclast-containing cultures were similar in ion and inhibitor sensitivities to the enzyme from kidney-derived sources. Immunopurified V-ATPase from 1,25(OH)2D3-stimulated cultures exhibited 20-fold greater ATPase activity than the enzyme from unstimulated cultures, which do not contain osteoclasts. In contrast, 1,25(OH)2D3-treated cultures contained only 2-fold more assembled V-ATPase, as determined by immunoprecipitation. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunoblot analysis similarly showed ∼2-fold increases of V-ATPase mRNA and protein levels in 1,25(OH)2D3-treated cultures. The bulk of the relative difference in V-ATPase activity between the two cultures was due to a 10-fold difference in enzyme specific activity. Quantitative RT-PCR also revealed that expression levels of V-ATPase mRNAs reflected the stoichiometry of enzyme subunits in the assembled complex. These data indicate that in mouse bone marrow cultures, V-ATPase expression is controlled at the level of mRNA, and that increases in subunit expression and assembly cannot account for the 20-fold difference in enzyme activity in osteoclast-containing cultures. Therefore, osteoclast V-ATPase activity may be regulated by subtle alterations in enzyme structure or associated factors.
Down-Regulation of the Serum Stimulatory Components of the Insulin-like Growth Factor (IGF) System (IGF-I, IGF-II, IGF Binding Protein [BP]-3, and IGFBP-5) in Age-Related (Type II) Femoral Neck Osteoporosis
Authors: Boonen Steven , Mohan Subburaman , Dequeker Jan , Aerssens Jeroen , Vanderschueren Dirk , Verbeke Geert , Broos Paul , Bouillon Roger , Baylink David J. 
Published: April 2006

Abstract
Both a decrease in bone formation and an increase in bone resorption have been implicated in the pathogenesis of age-related (type II) femoral neck osteoporosis. While the increase in the bone resorption rate has been shown to be partially related to secondary hyperparathyroidism, the mechanisms underlying the decline in bone formation have not yet been identified. The aim of the present study was to test the hypothesis that the bone formation deficit associated with type II osteoporosis might be due to secondary hyperparathyroidism and/or to a deficiency of the insulin-like growth factor (IGF) system. Circulating concentrations of IGF-I, IGF-II, IGF binding protein (IGFBP)-3, IGFBP-4, IGFBP-5, 25-hydroxycholecalciferol (25(OH)D3), and intact parathyroid hormone (PTH) were measured in 50 elderly women after sustaining a hip fracture and in 50 healthy age-matched controls. In addition, serum levels of osteocalcin (OC), skeletal alkaline phosphatase, and N-terminal procollagen peptide and urinary pyridinium cross-links were determined as markers of bone remodeling, and bone mineral density (BMD) was assessed at the proximal femur. In the patient group, serum was drawn within 18 h of the fracture and prior to surgery. Circulating protein concentrations did not change over this time frame. No difference was found between mean IGFBP-4 serum levels in the two groups studied, while mean levels of IGF-I, IGF-II, IGFBP-3, IGFBP-5, 25(OH)D3, and markers of bone formation were significantly lower (p &lt; 0.006) in patients as compared with healthy subjects. Serum PTH and urinary pyridinium cross-links, however, were markedly increased (p &lt; 0.001) in the osteoporotic group. In pooled data from the normal and osteoporotic populations, age-adjusted multiple regression models based on IGF-I, IGF-II, IGFBP-3, and IGFBP-5 were found to be highly predictive of serum OC (R2 = 19%, p &lt; 0.001) and BMD of femoral neck (R2 = 49%, p &lt; 0.0001), consistent with an effect of the anabolic IGF components on overall bone formation rate. Similar models based on 25(OH)D3 and PTH, however, were statistically unrelated to OC. To address further the potential impact of trauma on circulating IGF system components, we measured IGF system component levels in 10 male patients within 18 h following tibial fracture and in 10 age-matched normal male subjects. There was no significant difference in serum level of any of the IGF system components between the two groups. Although limited by its cross-sectional design, the present study suggests that, in addition to bone resorption resulting from secondary hyperparathyroidism, impaired bone formation associated with deficiency of the IGF system might predispose elderly women to fragility fracture of the proximal femur.