bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of basic fibroblast growth factor on the growth and differentiation of adult stromal bone marrow cells: Enhanced development of mineralized bone-like tissue in culture
Authors: Pitaru Sandu , Kotev-Emeth Shlomo , Noff Dina , Kaffuler Shoshana , Savion Naphtali 
Published: February 2010

Abstract
Rat stromal bone marrow cells (SBMC) were shown to produce mineralized bone-like tissue in culture in the presence of dexamethasone, ascorbic acid, and β-glycerophosphate. The addition of 3 ng/ml of basic fibroblast growth factor (bFGF) resulted in a significant increase in formation of mineralized tissue. The present study was aimed at assessing the effect of bFGF on the proliferation and differentiation of SBMC and on the sequential development of mineralized bone-like tissue in culture. Transmission electron microscopy of bFGF-treated cultures demonstrated the development of a multilayered structure resembling mineralized bone tissue consisting of cell layers embedded within a heavy extracellular matrix. The matrix was rich in bundles of collagen fibers associated with extensive mineral deposits consisting of hydroxyapatite as determined by infrared spectrophotometry. The addition of 3 ng/ml of bFGF resulted in significant enhancement of [3H]thymidine and [3H]proline incorporation and protein accumulation by 12-, 2.5-, and 2.5-fold, respectively. bFGF treatment increased cAMP responsiveness, alkaline phosphatase activity, osteocalcin level, 45Ca2+ deposition, and mineralized-like tissue formation and induced the earlier expression of these markers in the treated culture. A biphasic sequence of events was observed during the development of mineralized bone-like tissue in bFGF-treated and control cultures. The first phase is characterized by cell proliferation and matrix accumulation and is reflected by a progressive increase in [3H]thymidine and [3H]proline incorporation until day 11. The second phase, which follows, is characterized by a sharp decline in cell proliferation and matrix accumulation and a concomitant expression of osteoblast differentiation as reflected by the progressive increase in alkaline phosphatase activity, mineral deposition, and osteocalcin expression. Treatment of cultures with bFGF accentuated this biphasic sequence of events. These results indicate that bFGF has the capacity to stimulate both the growth and the biochemical functions of SBMC obtained from a young adult animal.
New resorption assay with mouse osteoclast-like multinucleated cells formed in vitro
Authors: Tamura Tatsuya , Takahashi Naoyuki , Akatsu Takuhiko , Sasaki Takahisa , Udagawa Nobuyuki , Tanaka Sakae , Suda Tatsuo 
Published: February 2010

Abstract
We previously reported a procedure to obtain a preparation containing a large number of mouse osteoclast (OCL)-like multinucleated cells (MNCs) formed in cocultures of mouse osteoblastic and bone marrow cells in the presence of 1α,25-dihydroxyvitamin D3 [1α,25-(OH)2D3]. The MNCs satisfied major criteria of OCLs, such as tartrate-resistant acid phosphatase (TRAP) activity, acid production, calcitonin (CT) receptors, and the ability to form resorption pits on bone slices. In this report, we describe a simple resorption assay system using MNC preparations. After culturing MNC preparations or disaggregated rat OCL preparations on dentin slices, they were stained with Mayer's hematoxylin. The stained area corresponded exactly with the resorption pits visualized by scanning electron microscopy and were measured using an image analysis system attached to a light microscope. Pit formation by MNCs was gradually enhanced by reducing the medium pH (pH 7.5 &lt; 7.2 &lt; 6.9). The plan area resorbed by MNCs increased linearly for up to 72 h. These results are very similar to those obtained with OCL preparations. In multiple standard assays with MNC preparations, more than 250 MNCs could be placed on a dentin slice, and the total area resorbed to a level of up to 9% of the whole surface within 48 h. In contrast, in multiple assays with OCL preparations, it was not easy to place more than 50 OCLs on a slice and the resorbed area was only 0.7% of the surface. The resorbing activity of the MNC preparation expressed by the resorbed area per a TRAP-positive MNC (4.4 ± 0.4 × 10−3 μm2) was greater than that of the OCL preparation (1.9 ± 0.4 × 10−3 μm2). Pit formation by MNCs was dose dependently inhibited by salmon and human CT (ED50: salmon CT, 10−14 M; human CT, 3 × 10−13 M). Bafilomycin A1, a specific inhibitor of vacuolar type H+-ATPase, also inhibited pit formation by MNCs at concentrations of 10−9 M and above. In contrast, estrogen at 10−12–10−6 M had no significant inhibitory effect on pit formation in this assay system. The assay system presented here is sensitive and reproducible and will be useful for examining the effects of various natural factors and synthetic compounds on osteoclastic bone resorption. We have previously established a culture system for examining MNC formation. Therefore, it is now possible to examine the effects of various test compounds on the recruitment of OCLs and pit formation by the same OCLs in a series of experiments.
Does weight-bearing exercise affect non-weight-bearing bone?
Authors: Tommerup Lorri J. , Raab Diane M. , Crenshaw Thomas D. , Smith Everett L. 
Published: February 2010

Abstract
It was previously reported that weight-bearing exercise increases femur periosteal formation in 3-year-old, 230 kg cross-bred sows. To determine if this weight-bearing exercise also stimulated non-weight-bearing bone, bone formation in the seventh rib from these same sows was measured histomorphometrically on the periosteal, endocortical, and osteonal envelopes. The sows were randomly assigned by body weight to basal (B, n = 6), control (C, n = 7), or trained (T, n = 7) groups. After 3 weeks of exercise adaptation, T walked on a treadmill for 17 weeks at 5 km/h, 20 minutes/day, 5 days/week, at 5% grade. Groups were sacrificed initially (B) or after 20 weeks (C and T). Periosteal mineral apposition rate (MAR) was calculated over 136 days. Osteonal and endocortical MAR were calculated over the 14 days prior to sacrifice. There were no effects of exercise on the periosteal or endocortical percentage double-labeled surface (dLS/BS), osteonal remodeling frequency (N.dL.On/B.Ar), or MAR in any bone envelope of the rib. In conclusion, weight-bearing exercise sufficient to stimulate periosteal formation in the femur did not activate formation in the rib. Bone response to weight-bearing exercise appears to be specific to the loaded bones.
In vivo effect of 1,25-dihydroxycholecalciferol on the proliferation and differentiation of avian chondrocytes
Authors: Dr. Farquharson Colin , Whitehead C.C. , Rennie J.S. , Loveridge N. 
Published: February 2010

Abstract
A combination of immunocytochemistry and in situ biochemistry has been used to determine the in vivo effects of 1,25-dihydroxycholecalciferol [1,25-(OH)2D3] on the proliferation and differentiation of chondrocytes. Chicks were fed a diet supplemented with 1,25-(OH)2D3 (2.5, 5, or 10 μg/kg diet) for 3 weeks, and measurements were made in sections of growth plate of chondrocyte proliferation and rate of maturation through the growth plate [using bromodeoxyuridine (BrdUrd) labeling] and also chondrocyte differentiation [assessed by alkaline phosphatase (ALP) activity]. The labeling indices of the control and supplemented chicks were similar (23.1 ± 1.3 versus 23.2 ± 1.6%); however, within a 21 h period the BrdUrd-positive cells of the supplemented chicks had moved down the growth plate significantly farther than in the control chicks (71.0 ± 2.8 versus 52.6 ± 1.8%). Greater ALP (mean integrated absorbance) activity higher up the growth plate of the supplemented chicks indicated a more differentiated phenotype in cells closer to the epiphyseal junction. Within individual transitional chondrocytes ALP activity in the 10 μg/kg supplemented chicks was 26.6 ± 0.85, which was significantly higher (p &lt; 0.01) than that of the control chicks (19.2 ± 0.9). These results suggest that 1,25-(OH)2D3 in vivo does not increase the rate of chondrocyte prolferation but accelerates the onset of maturation.
Immunohistochemical detection and northern blot analysis of estrogen receptor in osteoblastic cells
Authors: Ikegami Akira , Inoue Satoshi , Hosoi Takayuki , Mizuno Yuzo , Nakamura Tetsuro , Ouchi Yasuyoshi , Orimo Hajime 
Published: February 2010

Abstract
Expression of estrogen receptor (ER) was studied in MC3T3-E1 cells (mouse osteoblastic cell line), HOS TE85 cells (human osteosarcoma cell line), and primary osteoblastic cells derived from mouse calvaria with immunohistochemical techniques. The staining of ER was readily detectable in MC3T3-E1 cells, HOS TE85 cells, and primary osteoblastic cells by using a monoclonal anti-ER antibody that recognizes the DNA binding domain of ER. The immunoreactivity was distributed in the cytoplasm as well as in the nuclei. 17β-Estradiol (10−8 M) did not alter this staining pattern. The expression of ER was confirmed by Northern blot analysis using rat ER cDNA probe, which revealed a 6.5 kb band in MC3T3-E1 cells and a 6.2 kb band in HOS TE85 cells. The mRNA level of ER was not altered by 17β-estradiol (10−8 M). The immunohistochemical studies showed that ER was not detectable in all cells but in a small population of each cell type. This study is the first report to demonstrate the presence of ER immunohistochemically, and our results suggest the heterogeneity of ER expression among osteoblastic cells.
Expression of annexin I, II, V, and VI by rat osteoblasts in primary culture: Stimulation of annexin I expression by dexamethasone
Authors: Suarez Françoise , Rothhut Bernard , Comera Christine , Touqui Lhousseine , Marie Françoise Russo , Silve Caroline 
Published: February 2010

Abstract
To determine whether rat osteoblasts synthesize proteins of the annexin family and to evaluate the extent to which glucocorticoids modulate the expression of annexins by these cells, osteoblasts were grown in primary cultures in the absence or presence of dexamethasone, and the expression of annexins was evaluated by immunoblotting using polyclonal antibodies against human annexins. Four different annexins (I, II, V, and VI) were found to be expressed by rat osteoblasts. The expression of annexin I, but not the other annexins studied, was increased in osteoblasts cultured in the presence of dexamethasone (173 ± 33% increase comparing untreated cells and cells treated for 10 days with 5 × 10−7 M dexamethasone). Increased expression of annexin I was observed after the third day of exposure to dexamethasone and rose thereafter until day 10; annexin I expression increased with dexamethasone concentrations above 10−10 M throughout the range of concentrations studied. The increase in annexin I protein was associated with an increase in annexin I mRNA and was completely blocked by the concomitant addition of the glucocorticoid receptor antagonist RU 38486. The increase in annexin I content following dexamethasone treatment was associated with an increase in alkaline phosphatase activity and PTH-induced cAMP stimulation, whereas phospholipase A2 activity in the culture medium was reduced to undetectable levels. The finding that four annexins are expressed in rat osteoblasts in primary culture raises the possibility that these proteins could play an important role in bone formation by virtue of their ability to bind calcium and phospholipids, serve as Ca2+ channels, interact with cytoskeletal elements, and/or regulate phospholipase A2 activity. In addition, the dexamethasone-induced increase in annexin I may represent a mechanism by which glucocorticoids modify osteoblast function.
Osteoblasts express the PMCA1b isoform of the plasma membrane Ca2+-ATPase
Authors: Meszaros J. Gary , Karin Norman J. 
Published: February 2010

Abstract
We report here that osteoblasts and osteoblast-like osteosarcoma cells express PMCA1b, an alternatively spliced transcript of plasma membrane Ca2+-ATPase. Synthetic oligonucleotide pairs were designed based upon unique regions of the cDNA encoding known PMCA isoforms (PMCA1–3) and used as primers in PCR-mediated amplification of cDNA synthesized from ROS 17/2.8 osteosarcoma cell RNA. A product was observed only when PMCA1-specific primers were present; no products were seen with PMCA2 or PMCA3 primers unless cDNA synthesized from rat brain RNA was present. Examination of the cDNA encoding the C terminus of PMCA1 from ROS 17/2.8 cells revealed that the mRNA is spliced to yield the PMCA1b isoform, a Ca2+-ATPase containing a consensus phosphorylation site for cAMP-dependent protein kinase A and a modified calmodulin binding domain. PMCA1b was also detected in UMR-106–01 osteosarcoma cells and unpassaged primary rat calvarial osteoblasts. These results suggest that the regulation of osteoblast function by agents that act via cAMP-mediated pathways may involve alterations in the activity of the plasma membrane Ca2+-ATPase.
Circulating levels of vitamins K1 and K2 decreased in elderly women with hip fracture
Authors: Hodges S.J. , Akesson K. , Vergnaud P. , Obrant K. , Prof. Delmas P.D. 
Published: February 2010

Abstract
We measured the serum levels of phylloquinone (vitamin K1) and of the menaquinones, MK-7 and MK-8, in a group of 51 women with a mean age of 81 years who were studied within a few hours after a hip fracture. A group of 38 healthy age-matched women randomly chosen from the same population served as controls. Patients with hip fracture had a marked reduction in serum vitamin K1 (336 ± 302 versus 585 ± 490 pg/ml, p &lt; 0.01), MK-7 (120 ± 84 versus 226 ± 178 pg/ml, p &lt; 0.001), and MK-8 (89 ± 113 versus 161 ± 145 pg/ml, p &lt; 0.01), and a large number had undetectable levels, especially of MK-8. Vitamin K levels were not correlated with the time elapsed after fracture or with serum cortisol or other biochemical variables. These data suggest that patients with hip fracture have vitamin K deficiency, an abnormality that could affect bone metabolism through an impairment of the gamma carboxylation of the gla-containing proteins of bone.
Reactive oxygen species activate and tetracyclines inhibit rat osteoblast collagenase
Authors: Ramamurthy Nungavaram S. , Vernillo Anthony T. , Greenwald Robert A. , Lee H.-M. , Sorsa Timo , Golub Lorne M. , Rifkin Barry R. 
Published: February 2010

Abstract
Recent studies have demonstrated that tetracyclines (TCs) scavenge reactive oxygen species (ROS). Hypochlorous acid (HOCI), an ROS produced by neutrophils, has been shown to activate neutrophil procollagenase. The objective of the present study was to determine whether (1) HOCI also activated osteoblast procollagenase and (2) TCs inhibited this enzyme in the presence of HOCI. HOCI (5 μM) activated the proenzyme approximately sixfold (P &lt; 0.01) from the medium of PTH-treated UMR-106–01 osteoblastic osteosarcoma cells as determined by functional collagenase assay (3H-methyl-labeled collagen substrate). Doxycycline (50–400 μM) and chemically modified tetracycline, CMT-1 (100–400 μM), significantly inhibited collagenase activity 50–90% and 40–80%, respectively, in the presence of 5 μM HOCI. Concentrations of 6–25 μM doxycycline and 10–50 μM CMT-1 had no significant effect. Furthermore, an excess concentration of cation (50 mM CaCI2 or 50 μM ZnCI2) added to the incubation mixtures containing either doxycycline or CMT-1 did not restore collagenase activity, as demonstrated by SDS-PAGE-fluorography. These data suggested that TCs reduced available HOCI and thus prevented the hypochlorous acid conversion of the osteoblast proenzyme to active collagenase. TCs may have therapeutic potential in the treatment of periodontitis and other diseases by several mechanisms that inhibit pathologic collagen breakdown.
Intestinal malabsorption of 45calcium in young Gy mice, a second model for X-linked hypophosphatemia
Authors: Woodward Jennifer E. , Meyer Martha H. , Gray Richard W. , Dr. Meyer Ralph A. 
Published: February 2010

Abstract
X-linked hypophosphatemia, a common metabolic bone disease in humans and mice (the Hyp and Gy mutations), is characterized by decreased plasma phosphate, decreased renal tubular reabsorption of phosphate, rickets, and osteomalacia. The question of whether intestinal malabsorption of calcium contributes to the bone disease is controversial. Intestinal absorption of 45Ca was studied in three different mouse colonies: Gy on B6C3H background, Hyp on B6C3H background, and Hyp on C57BL/6J background, all at 4 weeks of age. The duodenum was isolated by sutures, and 45Ca in a 150 mM NaCl and 2 mM CaCl2 solution at pH 7.2 was injected into the lumen. Absorption was measured by the amount of 45Ca remaining in the lumen and by the plasma isotope level. The Gy and Hyp mice of both sexes significantly malabsorbed 45Ca at 4 weeks of age compared to normal littermates. Following the 4 week study, intestinal absorption was measured at 2, 7–8, and 12 weeks of age in normal and Gy mice on the B6C3H background. At 2 and 7–8 weeks of age, the Gy males significantly malabsorbed 45Ca compared to their normal littermates. Serum 1,25-dihydroxyvitamin D was not significantly altered in Gy males at 4 weeks of age. This suggests the possibility of resistance of the intestine to stimulation. Malabsorption of calcium in young Gy and Hyp mice may exacerbate the low mineralization in their rachitic bone disease.
Effects of bone-seeking hormones on DNA synthesis, cyclic AMP level, and alkaline phosphatase activity in cultured cells from human posterior longitudinal ligament of the spine
Authors: Ishida Yoichiro , Kawai Shinya 
Published: February 2010

Abstract
In a study of the osteogenesis capability of the human posterior longitudinal ligament of the spine, ligament cells were isolated and cultured. The effect of bone-seeking hormones, such as parathyroid hormone (PTH), calcitonin (CT), prostaglandin E2 (PGE2), and 1,25-dihydroxycholecalciferol [1,25-(OH)2D3], on the ligament cells was investigated with respect to DNA synthesis, adenosine-3′,5′-cyclic monophosphate (cAMP) levels, alkaline phosphatase (ALP) activity, and acid phosphatase (ACP) activity. Cell lines obtained from nonossified sites in patients with ossification of the posterior longitudinal ligament of the spine (OPLL) were found to have several different phenotypic characteristics for osteoblasts: high ALP activity, PTH- and PGE2-stimulated increases in cAMP, and responses to both CT and 1,25-(OH)2D3. It is clear that proliferation and differentiation in such ligament cells are controlled by various types of bone-seeking hormones, and it was suggested that many cells with osteoblast-like characteristics are present. These results are considered important with respect to the etiology of OPLL, and an experimental system using cultured ligament cells appears to be useful in research on OPLL.
A new bisphosphonate, BM 21.0955, prevents bone loss associated with cessation of ovarian function in experimental dogs
Authors: Monier-Faugere Marie-Claude , Friedler Robert M. , Bauss Frieder , Malluche Hartmut H. 
Published: February 2010

AbstractWe previously found that bone loss occurs as soon as 1 month after ovariohysterectomy (OHX) in beagle dogs. Indirect evidence pointed to an early dramatic increase in bone resorption. To verify this hypothesis and evaluate the effects of a newly developed bisphosphonate, BM 21.0955 (Boehringer Mannheim), 36 beagle dogs were subjected to OHX and 12 dogs were sham operated (Sham). OHX dogs were divided into six groups (n = 6 each) and received subcutaneous injections of vehicle or BM 21.0955 at various doses (0.1, 0.3, 1, 10, and 100 μg/kg/day) for 1 month. Sham dogs were given vehicle (n = 6) or BM 21.0955 (1 μg/kg/day, n = 6). Iliac crest biopsies and blood drawings were done at baseline and at month 1. OHX dogs given vehicle exhibited a decrease in cancellous bone volume associated with an increase in erosion depth and a decrease in serum levels of 1,25-dihydroxyvitamin D. BM 21.0955 prevented the bone loss at a dose ≥ 1 μg/kg and the increase in erosion depth and the decrease in serum levels of 1,25-(OH)2D at a dose ≥ 0.3 μg/kg. No osteomalacia was observed at any dose of BM 21.0955. Bone turnover was reduced only when BM 21.0955 was administered at doses of 10 or 100 μg/kg. There were no changes in body weight or serum levels of calcium, phosphorus, creatinine, parathyroid hormone, or osteocalcin in all groups. The increase in erosion depth in OHX dogs given vehicle proves that the early rapid bone loss after cessation of ovarian function is related to an increase in osteoclastic activity. The antiosteoclastic activity of BM 21.0955 at a dose ≥ 1 μg/kg prevents this increase and preserves bone volume. The absence of any signs of osteomalacia at any dose confers a relatively wide therapeutic margin to BM 21.0955. BM 21.0955 at a dose ≥ 10 μg/kg also acts as an inhibitor of bone turnover. This is not observed at a dose of 1 μg/kg, at least after 1 month of administration.
Differential effects of fluoride during initiation and progression of mineralization of osteoid nodules formed in vitro
Authors: Dr. Bellows C.G. , Aubin J.E. , Heersche J.N.M. 
Published: February 2010

Abstract
Osteoid nodules form in cultures of fetal rat calvarial (RC) cells grown in medium containing 10% FBS and 50 μg/ml of ascorbic acid. When 10 mM β-glycerophosphate (β-GP) is added, osteoid nodules mineralize in two phases: an initiation phase, which is dependent upon alkaline phosphatase activity for conversion of β-GP to Pi, and a progression phase that proceeds independently of alkaline phosphatase activity and does not require exogenous phosphate. We have now used this system to investigate the effects of fluoride (F−) on mineralization. In cultures in which osteoid was formed and mineralization initiated in the presence of F−, a dose-dependent inhibition of the initiation of mineralization occurred over a concentration range of 25–500 μM F− (p &lt; 0.001 in all cases). The initiation of mineralization was not inhibited if F− was removed from the cultures at the time when mineralization was initiated with β-GP. In osteoid nodules grown in the absence of F−, addition of F− resulted in a dose-dependent inhibition of the initiation of mineralization, with significant decreases in 45Ca uptake occurring at F− concentrations of 3 μM (p &lt; 0.01) and higher. However, if F− was added to cultures after mineralization was initiated in the absence of F−, a stimulation of 45Ca uptake was observed at F− concentrations of 250 μM and above (p &lt; 0.001). F− (1–1000 μM) did not affect the conversion of β-GP to Pi or alkaline phosphatase activity in the cultures. The data show that mineralization of osteoid formed in the presence of F− is unaffected when F− is removed before initiation, that F− inhibits the initiation phase of mineralization at concentrations as low as 3 μM, and that F− stimulates mineralization during the progression phase. These data agree with clinical observations of hypermineralization of existing bone and hypomineralization of newly formed osteoid in association with F− administration.