bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tumor necrosis factors α and β can stimulate bone resorption in cultured mouse calvariae by a Prostaglandin-independent mechanism
Authors: Lerner Ulf H. , Ohlin Acke 
Published: February 2010

Abstract
Human recombinant tumor necrosis factors α and β (TNF-α and TNF-β), at and above 1 ng/ml (≅ 70 pM), caused a dose- and time-dependent enhancement of 45Ca release from neonatal mouse calvarial bones in vitro. In addition, TNF-α and TNF-β (3–100 ng/ml) caused a dose-dependent stimulation of prostaglandin E2 (PGE2) formation in the calvarial bones. TNF-α also enhanced the biosynthesis of PGI2, as assessed by analysis of the stable breakdown product 6-keto-PGF1α. The stimulatory actions of TNF-α and TNF-β on PGE2 formation was maximal at 12 h. Indomethacin, flurbiprofen, and meclofenamic acid, three structurally unrelated nonsteroidal antiinflammatory drugs, abolished PGE2 biosynthesis induced by TNF-α and TNF-β (100 ng/ml). The 45Ca release stimulated by TNF-α and TNF-β (100 ng/ml), however, was only slightly reduced by indomethacin, flurbiprofen, and meclofenamic acid. The partial inhibitory effect of indomethacin on 45Ca release was seen over a wide range of TNF-α concentrations, without affecting the concentration producing half-maximal stimulatory response. TNF-α and TNF-β (100 ng/ml) stimulated bone matrix breakdown, as assessed by analysis of the release of 3H from bone prelabeled with [3H]proline. Also, the stimulatory effect of TNF-α and TNF-β on bone matrix degradation was partially reduced by indomethacin. Hydrocortisone (1 μM) and dexamethasone (0.1 μM) abolished TNF-α- and TNF-β-induced production of PGE2. In contrast to the cyclooxygenase inhibitors, the corticosteroids did not affect the stimulatory action by the cytokines on 45Ca release. These observations suggest that TNF-α and TNF-β can stimulate bone resorption in vitro by prostaglandin-independent mechanisms.
Modulation of vitronectin receptor-mediated osteoclast adhesion by Arg-Gly-Asp peptide analogs: A structure-function analysis
Authors: Horton Michael A. , Dorey Elaine L. , Nesbitt Stephen A. , Samanen James , Ali Fadia E. , Stadel Jeffrey M. , Nichols Andrew , Greig Russel , Helfrich Miep H. 
Published: February 2010

Abstract
This study details the investigation of induction of retractile shape change in the osteoclast through inhibition of adhesion between osteoclasts and matrix with (1) peptide analogs bearing an Arg-Gly-Asp (RGD) sequence, (2) antibodies to the integrin αvβ3 vitronectin receptor, and (3) the RGD-containing snake venom peptide echistatin. Osteoclast retraction on dentin has been demonstrated for GRGDSP peptide, in contrast to the inactivity of the analog containing the conservative RGE sequence modification. An osteoclast adhesion assay employing rat or chick bone cells and serum-coated glass coverslips as substrate was developed for routine evaluation of inhibition of adhesion. Antibodies F4 and F11 to the β3 chain of rat vitronectin receptor were effective at submicromolar concentrations in rat osteoclasts (IC50 0.29 and 0.05 μM, respectively), whereas MAb 23C6 to human/chick vitronectin receptor was somewhat less effective against chick osteoclasts (ICso 1.6 μM). A rank order of RGD analog activity (mean IC50, μM) in the serum-coated glass adhesion assay was derived for the linear peptides GRGDSP (201 μM), GRGDTP (180 μM), Ac-RGDS-NH2 (84 μM), Ac-RGDV-NH2 (68 μM), RGDV (43 μM), GRGDS (38 μM), and RGDS (26 μM). The two most potent short peptides were the cyclic analog SK&F 106760 Ac-S,S-cyclo-(Cys-(NαMe)Arg-Gly-Asp-Pen)-NH2 (IC50 7.0 μM), and the Telios peptide H-Gly-S,S-cyclo-(Pen-Gly-Arg-Gly-Asp-Ser-Pro-Cys)-Ala-OH (IC50 6.6 μM). The snake venom peptide echistatin was the most potent substance evaluated in the serum-coated glass assay (IC50 0.78 nM) employing either rat or chick osteoclasts. Specificity control peptides fibronectin CS1 (ligand for VLA-4), fibrinogen H12 (alternate ligand for gpIIb/IIIa), and Iaminin cell binding fragment YIGSR were inactive up to 800 μM. Comparison of SK&F 106760 and the Telios peptide as inhibitors of platelet aggregation (IC50 0.36 and 10.1 μM, respectively) and inhibitors of L8 skeletal muscle cell adhesion to vitronectin (IC50 67.2 and 12.3 μM, respectively) suggests that the Telios peptide is nonselective whereas SK&F 106760 may be selective with regard to β3 integrins. This study demonstrates that structural modification in RGD peptides and the use of antireceptor antibodies or the venom peptide echistatin yields potent inhibitors of vitronectin receptor-mediated adhesion in isolated rat and chick osteoclasts. It is hoped that further peptide modification will yield improved specificity and thus selective inhibitory effects upon bone resorption.
Subcellular localization and partial purification of the 25-hydroxyvitamin D3 1-hydroxylation reaction in the chick myelomonocytic cell line HD-11
Authors: Shany Shraga , Ren Song-Yang , Arbelle Jonathan E. , Clemens Thomas L. , Adams John S. 
Published: February 2010

Abstract
Hypercalcemia in human granuloma-forming diseases like sarcoidosis results from the endogenous overproduction of 1,25-dihydroxyvitamin D [1,25-(OH)2D] by disease-activated tissue macrophages. The recent identification of an immortalized chick myelomonocytic cell line, HD-11, that constitutively expresses a 25-hydroxyvitamin D (25-OHD) 1-hydroxylation reaction has alleviated dependence on studying primary macrophage cultures with no replicative potential in vitro. In these experiments we established conditions for the maximal expression of the HD-11 cell 25-OHD3-1-hydroxylation reaction and localized this activity to the mitochondrial fraction. On a per cell basis, the activity of HD-11 cell 25-OHD3 1-hydroxylation reaction was comparable to that in primary cultures of chick renal tubular epithelial cells, which express the authentic renal 25-OHD3 1-hydroxylase. Maximal product yield was achieved after incubation of HD-11 cells with 200 nM 25-OHD3 for 3 h. Although adherent monolayers possessed 3- to 4-fold more capacity for hormone production than cells in suspension, suspended cells exhibited easily detectable 25-OHD3 catalytic activity (0.58 ± 0.08 pmol per 106 cells per h; ± SEM), 50% of which remained solubilized in a sonicate of suspended cells cleared of nuclei and plasma membrane. Subcellular localization disclosed 91% of the residual activity to be concentrated in the mitochondrial subfraction. A detergent-solubilized extract of this mitochondrial subfraction contained 1.9 ± 0.3 pmol 1,25-(OH)2D3 synthetic capacity per mg protein. The catalytic activity (1-hydroxylase activity) was concentrated 20.2-fold after chromatography on octyl-amino agarose and was associated with 0.054 nmol cytochrome P450 per mg protein. These data suggest that HD-11 cell 25-OHD3 1-hydroxylation reaction may be similar to the well-characterized 25-OHD3 1-hydroxylase expressed in the kidney.
Rapid and sensitive method for quantitation of bone Gla protein mRNA using competitive polymerase chain reaction
Authors: Araki Nobuhito , Robinson Darlene F. , Nishimoto Satoru K. 
Published: February 2010

Abstract
A method for sensitive quantitation of bone gla protein (BGP, osteocalcin) mRNA has been developed using competitive polymerase chain reaction after reverse transcription (competitive RT-PCR). The complementary DNA (cDNA) reverse transcribed from sample RNA was coamplified in a PCR with a known amount of mutant BGP cDNA (competitor) using the identical oligonucleotide primers. The mutant cDNA with its unique restriction site allowed quantitation of sample and mutant PCR products after densitometric analysis of ethidium bromide-stained agarose gels. A linear relationship between initial sample BGP amount and the ratio of BGP to mutant BGP band intensity was obtained and used to make a standard curve to determine the initial BGP mRNA of unknown samples. These standard curves were made with known amounts of recombinant BGP cDNA. The competitive RT-PCR for BGP allows measurement of twofold differences in 1 and 2 μg total RNA and requires at least 10 times less sample RNA than usual Northern blotting. Moreover, heteroduplexes with one BGP strand and one mutant BGP strand formed as a result of high PCR cycles were quantifiable. This provided the advantages of rapid quantitation from ethidium bromide-stained gels without blotting, hybridization, or autoradiography. Multiple samples could be assayed for greater confidence in the results. The sensitivity, accuracy, and ease of the assay will facilitate analysis of BGP mRNA from a small amount of sample. The assay has been used to confirm the BGP mRNA changes with hormonal treatment in cultured cells and the age-related changes in whole tibia in vivo.
Ultrastructural immunolocalization of noncollagenous (osteopontin and osteocalcin) and plasma (albumin and α2HS-glycoprotein) proteins in rat bone
Authors: Dr. McKee Marc D. , Farach-Carson M.C. , Butler W.T. , Hauschka P.V. , Nanci A. 
Published: February 2010

Abstract
The high-resolution, postembedding protein A-gold immunocytochemical technique was used to visualize the distribution of two noncollagenous bone proteins, osteopontin (OPN) and osteocalcin (OC), and two plasma proteins, α2HS-glycoprotein (α2HS-GP) and albumin (ALB), in sections of Lowicryl K4M-embedded rat tibial and alveolar bone. In the primary spongiosa of the metaphysis, a seam of organic material (lamina limitans) that labeled intensely with OPN and OC antibodies was observed at the bone/calcified cartilage interface just below the zone of vascular invasion of the growth plate. With deposition of bone matrix proper by osteoblasts in this region and its subsequent mineralization, extensive areas of bone were heavily labeled with anti-OPN, anti-OC, and anti-α2HS-GP antibodies, where the majority of gold particles were associated with amorphous, electron-dense patches of organic material throughout the mineralized bone. In the unmineralized osteoid, substantially less labeling was observed, and where occasional mineralization loci were dispersed throughout the osteoid layer, these sometimes showed a concentration of gold particles. ALB labeling, on the other hand, was moderate and generally diffuse throughout the mineralized bone matrix and the osteoid. In alveolar bone, labeling patterns were generally similar to those found in tibial bone. Particularly striking in alveolar bone, however, was an intense anti-OPN labeling of (1) the lamina limitans at cell-lined bone surfaces, including that surrounding cell processes and osteocytes, (2) cement (reversal, resting) lines, and (3) the perilacumar matrix of some osteocytes. In summary, these data suggest that certain plasma proteins, such as α2HS-GP, interact with bone matrix proteins, such as OPN and OC, at sites of tissue mineralization and that the presence of OPN in mineralized bone and at bone surfaces (lamina limitans) and cement lines has a multifunctional role, including regulation of mineralization and mediation of cell dynamics during endochondral and intramembranous bone modeling and remodeling.
Ontogenesis of ultrastructural features during osteogenic differentiation in diffusion chamber cultures of marrow cells
Authors: Passi-Even L. , Gazit D. , Bab I. 
Published: February 2010

Abstract
Three stages of osteogenic differentiation can be identified in in vivo diffusion chamber cultures (DCC) of unselected marrow cells, namely, proliferation, differentiation, and maturation (mineralization). These stages were characterized correlatively by in situ differential cell counts, alkaline phosphatase activity, and mineral accumulation. In the present study, the ultrastructure of marrow cell DCC was examined after incubation for 3–21 days. Features characteristic of osteoblastic and chondroblastic differentiation were first noted in 12 day DCC. Sites of osteoblastic differentiation showed cell-cell contacts associated with an increased cell density. The osteoblastic cells had long processes and were embedded in matrix with prominent fiber bundles reminiscent of collagen type I. The chondroblastic cells appeared solitary in areas of lesser cell density. By contrast to the long osteoblastic cell processes, they had short plasmalemmal projections and the matrix surrounding them contained single, thin, short fibers reminiscent of collagen type II, as well as proteoglycan granules. Both cell types showed prominent cytoskeletal elements, rough endoplasmic reticulum, and Golgi. One finding, previously unnoted in differentiating osteogenic cells, was mitochondria with condensed cristae that represent an increased rate of energy metabolism. These mitochondria were particularly abundant in the differentiation stage and declined as the cultures matured. These findings, together with previous reports in the epiphyseal growth plate, suggest that mineralization is associated with an optimal level of energy metabolism rather than extreme hypo- or hyperoxia. The set of ultrastructural parameters defined here in the marrow cell DCC may serve as useful markers for cells undergoing osteogenic differentiation.
Immunoassay of pyridinoline crosslink excretion in normal adults and in paget's disease
Authors: Delmas Pierre D. , Gineyts Evelyne , Bertholin Annick , Garnero Patrick , Marchand François 
Published: February 2010

Abstract
A new immunoassay (ELISA) based on an antiserum that preferentially recognizes the free form of the pyridinoline (Pyr) crosslink was used to assess the urinary excretion of Pyr in a large normal sample of the population comprising 236 men and 193 women 30–90 years of age. Urinary Pyr increased significantly with age in both sexes. In women Pyr was higher than in men (57 ± 21 versus 46 ± 17 nmol/mmol creatinine, p &lt; 0.001), and the menopause was reflected by a mean 37% increase, from an average 43 to 59 nmol/mm creatinine (p &lt; 0.001). In 52 patients with active Paget's disease of bone, Pyr excretion was markedly increased (206 ± 160 nmol/mmol creatinine, p &lt; 0.001 versus controls) and decreased by 71% after 3 days of IV treatment with the bisphosphonate pamidronate. Free Pyr measured by the ELISA and the total urinary excretion measured by HPLC were highly correlated both in normals (n = 74, r = 0.83, p &lt; 0.001) and in Pagetic patients (n = 20, r = 0.93, p &lt; 0.001). The mean intra- and interassay coefficients of variation of the ELISA were below 9 and 15%, respectively. It is concluded that this new convenient immunoassay of Pyr should be valuable for the clinical investigation of patients with osteoporosis and other metabolic bone diseases.
Determinants of bone mineral density and risk factors for osteoporosis in healthy elderly women
Authors: Ooms Marcel E. , Lips Paul , van Lingen Arthur , Valkenburg Hans A. 
Published: February 2010

Abstract
Bone mineral density (BMD) and risk factors for osteoporosis were assessed in 348 apparently healthy women over 70 years of age (mean 82.3 years). BMD was measured at both hips and the dominant distal radius. With stepwise multiple regression the best determinants of BMD, selected from anthropometric measurements, age, and years since menopause, were body weight and years since menopause (R2 between 0.07 and 0.20, p &lt; 0.001). Risk and protective factors for osteoporosis were analyzed as indicator variables by multiple regression and corrected for confounding by age, years since menopause, and body weight. Significantly lower BMD at the hip was found in participants with impaired mobility (-5%) and users of loop diuretics (-5%). Use of thiazide(like) diuretics did not influence BMD significantly at any site. Users of oral corticosteroids had a significantly lower BMD at the hip and the distal radius (range -9.1 to -24.3%). Participants with a history of Colles' fracture (n = 56) had a significantly lower BMD at the other radius (-12.9%). The mean calcium intake from dairy products was high (mean 921 mg/day), only 11% having an intake below 500 mg. A relation of calcium intake with BMD could not be detected at any measurement site. We conclude that BMD cannot be adequately predicted in elderly women. Risk factors for low BMD in the elderly are low body weight, high number of years since menopause, impaired mobility, and use of loop diuretics and oral corticosteroids. Calcium intake was not a risk factor in this study, but the number of individuals on a low calcium intake was small.
Flutamide-mediated androgen blockade evokes osteopenia in the female rat
Authors: Dr. Goulding Ailsa , Gold E. 
Published: February 2010

Abstract
Androgens are believed to play a role in building and maintaining bone in the female, as well as in the male. The antiandrogen drug flutamide inhibits responses to androgens from both the gonads and the adrenals. Antiandrogens prevent androgens stimulating bone cell proliferation and differentiation in vitro, but effects of androgen blockade on bone metabolism in vivo have not been tested. The present study was undertaken to determine whether androgen blockade with flutamide (15 mg/kg body weight orally daily) would influence bone turnover or bone composition (1) in female rats with intact ovaries and (2) in rats made estrogen-deficient with the luteinizing hormone releasing hormone (LHRH) agonist, buserelin (25 μg/kg body weight per day SC). Four groups of rats with 45Ca-labeled skeletons were studied for 4 weeks: group A, placebo; group B, buserelin; group C, flutamide; group D, flutamide + buserelin. Total-body calcium values (mean ± SD) were (mg) 2007 ± 109, 1779 ± 138 (P &lt; 0.01 versus group A), 1818 ± 140 (P &lt; 0.01 versus group A), and 1690 ± 75 (P &lt; 0.01 versus group A) in groups A-D, respectively. Thus both buserelin and flutamide induced osteopenia. Skeletal 45Ca changes suggested buserelin-mediated estrogen deficiency bone loss was due to increased bone resorption, but flutamide-mediated androgen deficiency bone thinning was caused principally by reduced bone formation. These findings support the view that androgens play an important role in preserving bone mass in the female rat. Importantly, adequate estrogen status did not compensate for flutamide-mediated osteopenia. Although this work was done in an animal model, it seems possible that antiandrogen drugs cause osteopenia in people as well as in the rat. Further study of the likely bone-protective effects of androgen are also warranted in women.
Alkali absorption and citrate excretion in calcium nephrolithiasis
Authors: Sakhaee Khashayar , Williams Russel H. , Oh Man S. , Padalino Paulette , Adams-Huet Beverley , Whitson Peggy , Pak Charles Y.C. 
Published: February 2010

Abstract
The role of net gastrointestinal (GI) alkali absorption in the development of hypocitraturia was investigated. The net GI absorption of alkali was estimated from the difference between simple urinary cations (Ca, Mg, Na, and K) and anions (Cl and P). In 131 normal subjects, the 24 h urinary citrate was positively correlated with the net GI absorption of alkali (r = 0.49, p &lt; 0.001). In 11 patients with distal renal tubular acidosis (RTA), urinary citrate excretion was subnormal relative to net GI alkali absorption, with data from most patients residing outside the 95% confidence ellipse described for normal subjects. However, the normal relationship between urinary citrate and net absorbed alkali was maintained in 11 patients with chronic diarrheal syndrome (CDS) and in 124 stone-forming patients devoid of RTA or CDS, half of whom had “idiopathic” hypocitraturia. The 18 stone-forming patients without RTA or CDS received potassium citrate (30–60 mEq/day). Both urinary citrate and net GI alkali absorption increased, yielding a significantly positive correlation (r = 0.62, p &lt; 0.0001), with the slope indistinguishable from that of normal subjects. Thus, urinary citrate was normally dependent on the net GI absorption of alkali. This dependence was less marked in RTA, confirming the renal origin of hypocitraturia. However, the normal dependence was maintained in CDS and in idiopathic hypocitraturia, suggesting that reduced citrate excretion was largely dietary in origin as a result of low net alkali absorption (from a probable relative deficiency of vegetables and fruits or a relative excess of animal proteins).
Passive immunization with anti-parathyroid hormone-related protein monoclonal antibody markedly prolongs survival time of hypercalcemic nude mice bearing transplanted human PTHrP-producing tumors
Authors: Sato Kanji , Yamakawa Yukiko , Shizume Kazuo , Satoh Tomoko , Nohtomi Kyoko , Demura Hiroshi , Akatsu Takuhiko , Nagata Naokazu , Kasahara Tadashi , Ohkawa Hiroyuki , Ohsumi Kazuoki 
Published: February 2010

Abstract
Malignancy-associated hypercalcemia is mainly caused by excessive production of parathyroid hormone-related protein (PTHrP) by the tumor. Using anti-PTHrP-(1–34) monoclonal murine antibody (anti-PTHrP MoAb), we studied whether repeated injection of the homologous antibody would continuously decrease the serum calcium concentration in hypercalcemic nude mice bearing transplanted human PTHrP-producing tumors, leading to prolongation of their survival time. Daily SC injections of anti-PTHrP MoAb decreased the serum calcium concentration almost to within the normal range in nude mice bearing transplanted human PTHrP-producing tumors (T3M-1, EC-GI, PC-3, and FA-6) but not in a nude mouse bearing a transplanted parathyroid carcinoma. The antibody did not affect FA-6 tumor growth either in vitro or in vivo. Pancreatic carcinoma cells (FA-6), which caused the most severe hypercalcemia, were inoculated into 6-week-old nude mice. When severe hypercalcemia (˜19 mg/dl) had developed, daily SC injection of anti-PTHrP MoAb was started. Within 18 days of this time point, all untreated tumor-bearing mice (n = 10) died of hypercalcemia and cachexia, whereas all the treated mice (n = 10) showed an increase in body weight and survived for at least 25 days. Histologic examination of the treated mice revealed a marked decrease in osteoclastic bone resorption, without toxicologic findings in the kidney and liver. These results suggest that passive immunization against PTHrP can continuously ameliorate the hypercalcemia and markedly prolong the survival time of severely hypercalcemic, tumor-bearing mice. If a human monoclonal antibody against PTHrP-(1–34) could be developed, then passive immunization would be potentially one of the most effective therapies for patients with malignancy-associated hypercalcemia due to excessive production of PTHrP.
Effect of age and osteoarthritis on bone mineral in rhesus monkey vertebrae
Authors: Grynpas Marc D. , Huckell C.B. , Reichs K.J. , Derousseau C.J. , Greenwood C. , Kessler M.J. 
Published: February 2010

Abstract
We have used vertebrae of free-ranging rhesus macaques to study the effect of age and osteoarthritis on bone mineralization and bone density and to relate these findings to weight, sex, parity, and mineral chemistry. Bone mineralization was determined using the density fractionation technique and bone density using dual-photon absorptiometry. Arthritis was determined osteologically. We found a relationship between mineralization, age, and osteophytes, such that mineralization rose with age in nonarthritics and decreased with age in arthritics. This could also be seen when the females were examined separately. In males, only an increase in mineralization with age could be seen. In females mineralization decreases with parity. Also in females, DPA density decreases with age and increases with parity. No relationships with DPA density could be seen using males and females together or males alone. In conclusion, we have shown that normal skeletal aging in rhesus monkeys is accompanied by an increase in mineralization similar to that in other species, but this is not true in the presence of osteoarthritis. In the females parity has an important effect because it seems to build up bone mass even though the bone present may be undermineralized.