bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bone marrow from mechanically unloaded rat bones expresses reduced osteogenic capacity in vitro
Authors: Keila Senia , Pitaru Sandu , Grosskopf Ayala , Dr. Weinreb Miron 
Published: February 2010

Abstract
Bone formation during mechanical unloading is reduced, mainly as a result of osteoblastic hypofunction. At the same time, the total number of osteoblasts per long bone is also markedly reduced. We tested the hypothesis that the number of osteogenic precursors present in the bone marrow stroma was concomitantly diminished by using an in vitro cell culture system in which femoral adherent bone marrow cells differentiate into active osteoblasts and produce bone-like nodules. Hindlimbs of 32-day-old male rats were either immobilized (unloaded) by sciatic neurectomy (immo) or sham operated (sham) and animals were killed after 11 days. Femora were either ashed to determine bone mass or used to generate bone marrow cultures. Adherent marrow cells were cultured in the presence of ascorbic acid, β-glycerophosphate, and dexamethasone. Bone mass was significantly reduced in unloaded femora (by 16%) and tibiae (by 18%). The number of adherent cells (determined on day 6) was reduced by 50% in the immo group. Reduced cell number did not result from slower proliferation in culture since [3H]thymidine incorporation on days 4 and 6 was similar in the two groups. The osteogenic potential in vitro of marrow from unloaded bones was diminished compared with that from loaded ones as evidenced by (1) lower alkaline phosphatase (ALP) activity per mg protein (by 25–40%, examined on days 6 and 12), and (2) reduced nodule formation (by 70%, expressed as percentage of the dish area stained with Alizarin Red S on day 21). None of these changes occurred in the contralateral limb of operated (immobilized) animals. In an additional experiment, adherent cells from both groups were subcultured on day 6 and seeded at an equal density of 40,000 cells per 35 mm dish. The osteogenic potential in cultures from unloaded bones was reduced as evidenced by lower ALP activity (by 25–40%) and fewer nodules formed (by 40%). These experiments show that unloading for 11 days causes a deficit in bone mass and reduces the number of femoral adherent marrow stromal cells and their osteogenic potential in vitro in subsequent (ex vivo) cultures. They also suggest that unloading specifically reduces the number of osteogenic precursors present within the marrow stroma.
Two-site enzyme immunoassay for measuring intact human osteocalcin in serum
Authors: Parviainen Markku , Kuronen Ilpo , Kokko Harri , Lakaniemi Mari , Savolainen Kari , Mononen Ilkka 
Published: February 2010

Abstract
We developed a sensitive two-site sandwich ELISA for quantitative analysis of human osteocalcin in serum or plasma. Our method is based on two different highly specific antibodies recognizing epitopes at different ends of the protein so that only intact osteocalcin is detected. The method is fast (total analysis time less than 6 h/96 wells), precise (intraassay variation less than 2.3% at four different levels; n = 10, and interassay variation less than 2.5%, n = 5, respectively), and accurate, with a mean recovery of 105%. The detection limit in serum is approximately 0.1 μg/liter. The mean concentration of osteocalcin in normal serum with this assay is 3.3 μg/liter (SD 3.7 μg/liter; range 0.1–13.1 μg/liter; n = 41), and the reference range is 0.28–10.1 μg/liter (10 and 90% confidence limits). The method shows a reasonable positive linear correlation with other osteocalcin assays (Incstar, r = 0.55, p &lt; 0.05, n = 13; Henning Oscatest, r = 0.52, p &lt; 0.005, n = 34). A good correlation (r = 0.70, p &lt; 0.001) between individual osteocalcin and bone-specific alkaline phosphatase serum concentrations was observed in normal subjects. We found a low or undetectable concentration of intact osteocalcin in serum of all four of our patients with acute primary hyperparathyroidism, and in all five patients with hypocalcemic secondary hyperparathyroidism, which suggests that PTH effectively inhibited the synthesis of osteocalcin in osteoblasts. The serum concentration of intact osteocalcin was elevated in two of three patients with chronic primary hyperparathyroidism. In five patients with normocalcemic secondary hyperparathyroidism, the serum concentration of intact osteocalcin was low: three had a normal and two had an elevated value. An age-related decrease in intact osteocalcin in serum of females younger than 50 years and an increase in older women were observed.
Evidence for a diminished maturation of preosteoblasts into osteoblasts during aging in rats: An ultrastructural analysis
Authors: Roholl P.J.M. , Blauw E. , Zurcher C. , Dormans J.A. M. A. , Dr. Theuns H.M. 
Published: February 2010

Abstract
Bone is subject to continuous remodeling throughout life. The age-related loss of (trabecular) bone, leading to senile osteopenia, is mainly due to impaired bone formation. Osteoblasts (OB) and osteoclasts (OC) have been identified as playing a crucial role in the process of bone turnover, but the contribution made by their precursors is not well documented. We analyzed the cells of the osteoblast and osteoclast cell lineage along the trabecular bone of tibiae and the stromal cells in the marrow of aging BN/Bi Rij rats using electron microscopy. It appeared possible to distinguish preosteoblasts (pre-OB), OB, preosteoclasts (pre-OC), OC, and inactive bone-lining cells. Periods of increase, the maximal peak, and the decrease in trabecular bone volume were defined by means of morphometric measurements of trabecular bone volume. We found a decrease of more than 10-fold in the number of OB with age, but the numbers of pre-OB, pre-OC, and OC expressed per unit bone length, although variable, were age independent. The relative bone resorption and formation surface, expressed as a percentage of the total bone surface, decreased 2- and 15-fold, respectively. In 2-year-old animals the total volume of stromal cells, part of which constitutes the stem cell compartment of the osteogenic lineage, was a quarter of that found in 1-month-old animals and a third of that found in 6-month-old animals. The loss of trabecular bone is concomitant with a sharp increase in the ratio of pre-OB/OB, the ratio of OC/OB, and in the ratio of resorption to formation surfaces. There was no relation between the ratio of pre-OC/OC with age. These data lead to the conclusion that the main factor causing bone loss with age is a diminished maturation of pre-OB into OB.
Deposition of cement at reversal lines in rat femoral bone
Authors: Zhou Hong , Chernecky Robert , Dr. Davies John E. 
Published: February 2010

Abstract
Femora from young adult Wistar rats were prepared for both light and electron microscopy. Routinely processed wax sections showed the appearance of cement lines immediately proximal to the resorption surface formed by active osteoclasts and distal to the onset of lamella formation in femoral bone tissue. This early stage of extracellular matrix elaboration at reversal lines was then studied by scanning electron microscopy (SEM) of actively remodeling sites, mainly on trabecular and endosteal surfaces. The resorption surface was shown to comprise a decalcified collagenous mat with individual fibers running either parallel or perpendicular to the surface plane. By examining different, neighboring, areas of resorption lacunae, a temporal sequence of new extracellular matrix production could be established. Before the deposition of new collagen, globular accretions were deposited onto the resorption surface. In areas where individual collagen fibers were oriented perpendicular to the surface plane, this globular matrix was initially deposited on the exposed fiber tips. The globules increased in size and fused laterally to form a continuous cement layer, which not only interdigitated with the collagen mesh of the resorption surface but also provided anchorage for new collagen fibers, which themselves became mineralized. These morphologic results provide a mechanistic explanation of coupling at reversal lines.
Expression of bone morphogenetic protein messenger RNA in prolonged cultures of fetal rat calvarial cells
Authors: Harris S.E. , Sabatini M. , Harris M.A. , Feng J.Q. , Wozney J. , Mundy G.R. 
Published: February 2010

Abstract
In addition to structural proteins of bone, such as type I collagen, bone cells synthesize a number of growth regulatory peptides that are also stored in the bone matrix, presumably as a consequence of local production by osteoblasts. Among the bone growth regulatory peptides found in the bone matrix are the recently described bone morphogenetic proteins (BMPs). These factors were purified from bone matrix by their capacity to stimulate ectopic bone formation, but it is not known whether they are produced by normal bone cells and influence normal bone formation. To determine whether they are expressed by normal osteoblasts during differentiation, we used the technique of prolonged primary culture of fetal rat calvarial osteoblasts. These cultures have been shown to be an informative model for studying expression of bone-related genes by cultured osteoblasts, since specific genes are expressed as the cells undergo proliferation and differentiation. We found that the bone morphogenetic proteins 1, 2, 4, and 6 are expressed by cultures of fetal rat calvarial osteoblasts before they form mineralized bone nodules and as they express alkaline phosphatase, osteocalcin, and osteopontin. This model can be used for study of regulation of expression of bone morphogenetic proteins by osteoblasts.
Long-term exercise of young and adult female rats: Effect on femoral neck biomechanical competence and bone structure
Authors: Søgaard Charlotte H. , Danielsen Carl Christian , Thorling Eivind B. , Mosekilde Lis 
Published: February 2010

Abstract
The present study was designed to examine the effect of exercise on femoral neck bone structure and strength in female rats after 4 and 10 months of exercise. Female Fischer rats aged 2 months were exercised for 4 h a day, 5 days a week on a motor-driven treadmill the speed of which was gradually increased until a daily distance of 2 km was reached. The training level was maintained for 4 months (n = 15) and 10 months (n = 15). Sedentary age-matched rats served as controls (n = 30). At death the proximal third of both femora was obtained from each rat. The left femoral neck was tested in a materials-testing machine, and the right was used for either trabecular bone mass measurement (BV/TV) or scanning electron microscopy (SEM). Biomechanical testing revealed a significant training-induced increase in femoral neck bone strength after 4 months of exercise, which although not accelerated was preserved after 10 months of exercise. Histologic investigation revealed a significant training-induced increase in BV/TV, accounted for by a significantly smaller proportion of marrow space. No difference in cortical area was found. Both histology and SEM revealed a tendency to an earlier closure of the growth line in the exercised animals. However, the exercised animals aged 6 months had a significantly increased total femoral length compared with the sedentary rats. No difference was found in total femoral length between sedentary and exercised rats aged 12 months. On the basis of this study, it is concluded that exercise has a positive effect on femoral neck bone strength. The gain of bone strength may be due partly to an increased bone mass and partly to an earlier maturation of the skeleton. The method presented could form an excellent basis for evaluating potential therapeutic regimens for the treatment of osteoporosis.
Osteoporosis in men with a history of tibial fracture
Authors: Kannus Pekka , Järvinen Markku , Sievänen Harri , Oja Pekka , Vuori Ilkka 
Published: February 2010

Abstract
The long-term effects of bone fractures on bone mineral density (BMD) at various skeletal sites are poorly established, although a serious fracture, such as a tibia fracture, followed by long immobility and disuse may lead to permanently decreased BMD and, through this mechanism, may be a risk factor for osteoporotic fractures in later life. To determine whether such an injury leads to osteoporosis, we measured the areal BMD (g/cm2) from the lumbar spine (L2–4), right distal radius and ulna, and the femoral neck, distal femur, patella, proximal tibia, distal tibia, and calcaneus of both extremities in 14 men with a history of primarily nonunited (finally bone-grafted) shaft fracture of the tibia and 20 men with a history of primary union. For evaluation of the patients' BMD in the spine and distal forearm, 22 age-, weight-, and height-matched normal men were also measured. The average time of immobilization in a long plaster cast was 27 weeks in the former group of patients and 16 weeks in the latter. The measurements were performed an average of 9 years after the fracture using a dual-energy x-ray absorptiometric scanner. Compared with normal men (mean ± SD = 1.116 ± 0.160), the spinal BMDs were significantly lower in men with a history of a primary nonunion (0.979 ± 0.100, −12.3%) and union (1.010 ± 0.124, −9.5%). In distal radius and distal ulna, there were no significant differences between the three groups. In the 14 patients with a primary nonunion, the mean BMD of the injured extremity (compared with the uninjured side) was significantly lower in the distal femur (-10.0%), patella (-11.2%), proximal tibia (-9.2%), distal tibia (-7.9%), and calcaneus (-5.6%). In the 20 patients with a primary union, the side-to-side difference was significant in the femoral neck (-2.4%), distal femur (-4.0%), patella (-3.7%), and proximal tibia (-5.1%). The relative BMD of the injured extremity did not correlate with the fracture type, fracture location, or patients' age but did show significant (r = 0.41–0.79) positive correlation with short immobilization time, low pain assessment, good muscle strength, and high functional scores of the injured extremity. In conclusion, the tibial shaft fracture may be associated with reduced bone density in the lumbar spine and injured extremity. The observed decrease (10–12% in spine and 4–11% in the knee region of the injured limb) is clinically important with respect to age-related bone loss of 1% per year after the age of peak bone mass. Additional follow-up is needed to determine any increased risk of osteoporotic fractures in spine and injured extremity.
Carboxyl-terminal parathyroid hormone peptide (53–84) elevates alkaline phosphatase and osteocalcin mRNA levels in SaOS-2 cells
Authors: Dr. Sutherland M. Kung , Rao L.G. , Wylie J.N. , Gupta A. , Ly H. , Sodek J. , Murray T.M. 
Published: February 2010

Abstract
Previous findings in our laboratory have shown that hPTH-(53–84) stimulates alkaline phosphatase activity in dexamethasone-treated ROS 17/2.8 cells. In the present study, we examined the effects of hPTH-(53–84) and hPTH-(1–34) on the expressions of alkaline phosphatase, osteocalcin, and collagen type I mRNA levels in the human osteosarcoma cell line SaOS-2. The effect of hPTH-(53–84) on alkaline phosphatase and osteocalcin message levels was dose dependent (ANOVA, p &lt; 0.005 and p &lt; 0.001, respectively), with significant stimulation observed at 10 nM. Treatment with 10 nM hPTH-(53–84) for 24 h resulted in significant 2- and 1.4-fold increases in mRNA levels for alkaline phosphatase and osteocalcin, respectively (p &lt; 0.05), but had no effect on collagen type I expression. The increased alkaline phosphatase mRNA levels was associated with a 1.5-fold increase in enzyme activity (p &lt; 0.05). In contrast, under similar incubation conditions, hPTH-(1–34) had no significant effects on alkaline phosphatase or osteocalcin mRNA levels. On the other hand, hPTH-(1–34) had dose-dependent stimulatory effects on collagen type I mRNA levels (ANOVA, p &lt; 0.001), 10 nM hPTH-(1–34) stimulating collagen type I expression 1.6-fold (p &lt; 0.05). The results indicate that carboxyl-terminal hPTH-(53–84) has direct and unique biologic effects in human osteoblast-like cells in culture.
Bone matrix RGD glycoproteins: Immunolocalization and interaction with human primary osteoblastic bone cells in vitro
Authors: Grzesik Wojciech J. , Robey Pamela Gehron 
Published: February 2010

Abstract
The interaction of cells with extracellular matrix is essential for their anchorage, proliferation, migration, and differentiation. In bone matrix there are multiple glycoproteins that contain the integrin-binding RGD sequence: fibronectin (FN), thrombospondin (TSP), osteopontin (OPN), bone sialoprotein (BSP), type I collagen (COLL I), and vitronectin (VN). In this study, the localization of TSP, FN, VN, and several integrins within developing human long bone using immunohistochemical methods was examined, as was the effect of all bone RGD proteins on the adhesion of human osteoblastic cells. Thrombospondin, fibronectin, and vitronectin showed distinct localization patterns within bone tissue. TSP was found mainly in osteoid and the periosteum; VN appeared to be present mainly in mature bone matrix. FN was present in the periosteum as well as within both mature and immature bone matrix. Using a panel of antiintegrin antibodies we found that bone cells in vivo and in vitro express α4, αv, α5β1, αvβ3, and β3/β5 integrins, and these receptors are for the most part expressed on all bone cells at different stages of maturation with quantitative rather than qualitative variations, with the exception of α4, which is expressed mainly by osteoblasts. Cell attachment assays were performed using primary human cells of the osteoblastic lineage under serum-free conditions. COLL I, TSP, VN, FN, OPN, and BSP promoted bone cell attachment in a dose-dependent manner and were equivalent in action when used in equimolar concentrations. In the presence of GRGDS peptide in the medium, the adhesion to BSP, OPN, and VN was almost completely blocked (10, 10, and 15% of control, respectively), and attachment to FN, COLL I, and TSP was only slightly decreased (80, 75, and 55%, respectively). These results suggest that human bone cells may use RGD-independent mechanisms for attachment to the latter glycoproteins.
Carboxyl-terminal parathyroid hormone-related protein inhibits bone resorption by isolated chicken osteoclasts
Authors: Dr. Fenton A.J. , Martin T.J. , Nicholson G.C. 
Published: February 2010

Abstract
Carboxyl-terminal peptides from parathyroid hormone-related protein (PTHrP) have been studied for their effect on bone resorption by osteoclasts isolated from 15 day embryonic chickens. Basal bone resorption by chicken osteoclasts was directly inhibited by chicken and human PTHrP-(107–139) and the pentapeptide PTHrP-(107–111). The chicken and human analogs were equipotent. Both the number of resorption pits and the total area resorbed per bone slice were reduced by PTHrP-(107–139), but it did not alter the size of individual resorption pits. Resorption stimulated by hPTH-(1–34) in cocultures of chicken osteoclasts with osteoblasts was also inhibited by cPTHrP-(107–139) but required a concentration three orders of magnitude greater than that required to inhibit basal resorption in cocultures or cultures of isolated osteoclasts. The finding of resorption inhibitory activity by PTHrP-(107–139) in avian as well as mammalian species strengthens the hypothesis that carboxyl-terminal PTHrP may act as a paracrine regulator of bone cell activity.
Effects of treatment of malignancy-associated hypercalcemia on serum parathyroid hormone-related protein
Authors: Budayr A.A. , Zysset E. , Jenzer A. , Thiébaud D. , Ammann P. , Rizzoli R. , Jaquet-Müller F. , Bonjour J.P. , Gertz B. , Burckhardt P. , Halloran B.P. , Nissenson R.A. , Strewler Gordon J. 
Published: February 2010

Abstract
Parathyroid hormone-related protein (PTHrP) is the primary mediator of hypercalcemia in patients with malignancy-associated hypercalcemia. We conducted this study to examine the effects of treatment with a bisphosphonate on serum PTHrP. We analyzed 41 episodes of hypercalcemia occurring in 38 patients: 22 patients received alendronate, and 16 were treated with pamidronate. At baseline, 29 patients had an increased serum PTHrP (group I) and 9 had low or undetectable levels (group II). The two groups did not differ significantly in baseline hypercalcemia (3.26 versus 3.41 mM) or the response of serum calcium to therapy. Serum calcium was normalized in 88% of group I and 70% of group II patients. Lowering of the mean calcium level was not associated with a change in the level of PTHrP in group I patients (40.2 versus 36.7 pgEq/ml) or group II patients. We also analyzed data on serum PTH and 1,25-(OH)2D in 20 of the patients. Serum PTH rose with treatment in group I patients (9.7–40.2 pg/ml, p &lt; 0.05), as did the serum 1,25-(OH)2D (19.1–32.4 pg/ml, p &lt; 0.001). Similarly, treatment of group II patients was associated with an increase in serum PTH (9.8–37.2 pg/ml) and serum 1,25-(OH)2D (22.9–40.2 pg/ml). The individual increases in 1,25-(OH)2D levels associated with therapy could not be predicted from the level of PTHrP or the changes in levels of serum calcium or PTH. Our data show that effective treatment of malignancy-associated hypercalcemia is not associated with a consistent change in serum levels of PTHrP. Therapy is associated with a variable increase in the serum levels of PTH and 1,25-(OH)2D.
Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts
Authors: Wucherpfennig Anne L. , Li Yi-Ping , Stetler-Stevenson William G. , Rosenberg Andrew E. , Stashenko Philip 
Published: February 2010

Abstract
The digestion of type I collagen is an essential step in bone resorption. It is well established that osteoclasts solubilize the mineral phase of bone during the resorptive process, but the mechanism by which they degrade type I collagen, the major proteinaceous component of bone, is controversial. Differential screening of a human osteoclastoma cDNA library was performed to characterize genes specifically expressed in osteoclasts. A large number of cDNA clones obtained by this procedure were found to represent 92 kD type IV collagenase (gelatinase B; MMP-9, EC 3.4.24.35), as well as tartrate-resistant acid phosphatase. In situ hybridization localized mRNA for gelatinase B to multinucleated giant cells in human osteoclastomas. Gelatinase B immunoreactivity was demonstrated in giant cells from eight of eight osteoclastomas, osteoclasts in normal bone, and osteoclasts of Paget's disease by use of a polyclonal antiserum raised against a synthetic gelatinase B peptide. In contrast, no immunoreactivity for 72 kD type IV collagenase (gelatinase A; MMP-2, EC 3.4.24.24), which is the product of a separate gene, was detected in osteoclastomas or normal osteoclasts. We propose that the 92 kD type IV collagenase/gelatinase B plays an important role in the resorption of collagen during bone remodeling.