Full Text - Hydrogen peroxide (H2O2) induces oxidative injury to human osteoblasts. The expression and potential function of circular RNA HIPK3 (circHIPK3) in H2O2-treated human osteoblasts were tested. We show that H2O2 significantly downregulated circHIPK3 in OB-6 cells and primary human osteoblasts. Furthermore, circHIPK3 levels were decreased in the necrotic femoral head tissues of dexamethasone-treated patients. In OB-6 osteoblastic cells and primary human osteoblasts, forced overexpression of circHIPK3 by a lentiviral construct alleviated H2O2-induced viability reduction, cell death and apoptosis. Contrarily, circHIPK3 silencing by targeted shRNA potentiated H2O2-induced cytotoxicity in OB-6 cells and primary human osteoblasts. Moreover, circHIPK3 downregulation by H2O2 induced miR-124 accumulation in OB-6 cells and primary human osteoblasts. On the contrary, miR-124
Bisphosphonates are therapeutically applied to treat metabolic bone diseases, such as osteoporosis or metastasis to the bone. Clinical studies have shown their potency to increase bone density over an extended period of time [25-28]. This effect is not only caused by a positive bone turnover, but also by a direct stimulation of osteoblast and osteoblast precursor cells by applying nitrogen-containing bisphosphonates [15, 29]. An anabolic effect to the bone could be caused by proliferation and by extracellular matrix production, mainly of collagen type I. With respect to osteoblast proliferation, we examined cyclin D1, an important regulator of the cell cycle and a surrogate of cell proliferation. Our results did not show a significant impact on osteoblast proliferation during the first 6 days. However, after day 6 zoledronate led to a reduced Cyclin D1 gene expression. As shown in other in vitro studies, pamidronate, a nitrogen-containing bisphosphonate, decreased osteoblast proliferation in a ...
Citation. Wei J, Shi Y, Zheng L, Zhou B, Inose H, Wang J, Guo XE, Grosschedl R, Karsenty G. miR-34s inhibit osteoblast proliferation and differentiation in the mouse by targeting SATB2.. The Journal of cell biology. 2012 May 07;. External Citation. Abstract. A screen of microRNAs preferentially expressed in osteoblasts identified members of the miR-34 family as regulators of osteoblast proliferation and/or differentiation. Osteoblast-specific gain- and loss-of-function experiments performed in vivo revealed that miR-34b and -c affected skeletogenesis during embryonic development, as well as bone mass accrual after birth, through two complementary cellular and molecular mechanisms. First, they inhibited osteoblast proliferation by suppressing Cyclin D1, CDK4, and CDK6 accumulation. Second, they inhibited terminal differentiation of osteoblasts, at least in part through the inhibition of SATB2, a nuclear matrix protein that is a critical determinant of osteoblast differentiation. Genetic evidence ...
The extracellular matrix glycoprotein, tenascin, is associated in vivo with mesenchyme undergoing osteogenesis and chondrogenesis, but is absent from mature bone and cartilage matrix. The expression of tenascin by osteoblastic cells in vitro has been investigated by immunoblotting and immunocytochemistry. Tenascin was secreted into the medium and deposited in the matrix by human and rat osteoblast-like cell lines, as well as by primary osteoblast-enriched cultures from chick embryo calvarial bones. In primary osteoblast-enriched cultures, extracellular tenascin was found only in cell aggregates expressing the osteoblast marker alkaline phosphatase. Chicken osteoblast cultures synthesized almost exclusively the largest tenascin subunit, whereas fibroblast cultures from periostea of chicken calvariae synthesized approximately equal amounts of all three subunits. In situ hybridization studies of developing chicken bones, using a cDNA probe that hybridizes to all chicken tenascin splice variants, ...
Current treatments for the prevention of thromboembolism include heparin and low-molecular weight heparins (LMWHs). A number of studies have suggested that long term administration of these drugs may adversely affect osteoblasts and therefore, bone metabolism. Xarelto™ (Rivaroxaban) is a new anti-thrombotic drug for the prevention of venous thromboembolism in adult patients undergoing elective hip and knee replacement surgery. The aim of this in vitro study was to investigate the possible effects of rivaroxaban on osteoblast viability, function and gene expression compared to enoxaparin, a commonly used LMWH. Primary human osteoblast cultures were treated with varying concentrations of rivaroxaban (0.013, 0.13, 1.3 and 13 μg/ml) or enoxaparin (1, 10 and 100 μg/ml). The effect of each drug on osteoblast function was evaluated by measuring alkaline phosphatase activity. The MTS assay was used to assess the effect of drug treatments on cell proliferation. Changes in osteocalcin, Runx2 and BMP-2
Osteoblasts are a key component in the regulation of the hematopoietic stem cell (HSC) niche. Manipulating osteoblast numbers results in a parallel change in HSC numbers. We tested the activity of strontium (Sr), a bone anabolic agent that enhances osteoblast function and inhibits osteoclast activity, on hematopoiesis. In vitro treatment of primary murine osteoblasts with Sr increased their ability to form bone nodules, and in vivo it increased osteoblast number, bone volume, and trabecular thickness and decreased trabecular pattern factor. However, the administration of Sr had no influence on primitive HSCs, although the number of hematopoietic progenitors was higher than in control cells. When Sr-treated mice were used as donors for HSC transplantation, no difference in the engraftment ability was observed, whereas hematopoietic recovery was delayed when they were used as recipients. Despite the changes in osteoblast numbers, no increment in the number of N-cadherin(+) osteoblasts and ...
Osteoblasts are a key component in the regulation of the hematopoietic stem cell (HSC) niche. Manipulating osteoblast numbers results in a parallel change in HSC numbers. We tested the activity of strontium (Sr), a bone anabolic agent that enhances osteoblast function and inhibits osteoclast activity, on hematopoiesis. In vitro treatment of primary murine osteoblasts with Sr increased their ability to form bone nodules, and in vivo it increased osteoblast number, bone volume, and trabecular thickness and decreased trabecular pattern factor. However, the administration of Sr had no influence on primitive HSCs, although the number of hematopoietic progenitors was higher than in control cells. When Sr-treated mice were used as donors for HSC transplantation, no difference in the engraftment ability was observed, whereas hematopoietic recovery was delayed when they were used as recipients. Despite the changes in osteoblast numbers, no increment in the number of N-cadherin(+) osteoblasts and N-cadherin
TY - JOUR. T1 - Fibroblasts expressing Sonic hedgehog induce osteoblast differentiation and ectopic bone formation. AU - Kinto, Naoki. AU - Iwamoto, Masahiro. AU - Enomoto-Iwamoto, Motomi. AU - Noji, Sumihare. AU - Ohuchi, Hideyo. AU - Yoshioka, Hidefumi. AU - Kataoka, Hiroko. AU - Wada, Yasuhiro. AU - Yuhao, Gao. AU - Takahashi, Hideaki E.. AU - Yoshiki, Shusaku. AU - Yamaguchi, Akira. PY - 1997/3/10. Y1 - 1997/3/10. N2 - We investigated the role of Sonic hedgehog (SHH) in osteoblast differentiation and bone formation. The numbers of ALP-positive cells in the mouse fibroblastic cell line C3H10T1/2 and the mouse osteoblastic cell line MC3T3-E1 were increased by co-culture with chicken fibroblasts transfected with chicken Shh cDNA encoding amino-terminal peptide (Shh-N). The conditioned medium of Shh-N-RCAS-transfected chicken fibroblast cultures also significantly increased ALP activity in both C3H10T1/2 and MC3T3-E1 cells. Intramuscular transplantation of Shh-N-RCAS-transfected chicken ...
Results IL-6 significantly reduced ALP activity, and expression of osteoblastic gene, Runx2 and osteocalcin, and also inhibited mineralization in a dose-dependent manner, which indicates a negative effect of IL-6 on osteoblast differentiation. Signal transduction analyses by Western blotting demonstrated that IL-6 significantly promoted phosphorylation of ERK and STAT3. The negative effect of IL-6 on ALP activity was restored by inhibition of ERK in a dose-dependent manner. On the other hand, the negative effect of IL-6 on ALP activity was enhanced by inhibition of STAT3. These results indicate that ERK has negative effect on osteoblast differentiation, whereas STAT3 has positive one. Moreover, ERK inhibitor enhanced IL-6-triggered STAT3 phosphorylation, and STAT3 inhibitor enhanced IL-6-triggered ERK phosphorylation, suggesting that ERK and STAT3 signaling pathway negatively regulates each other.. ...
DOI: 10.11607/jomi.4247 Purpose: To assess and compare topographic features and preosteoblastic cell responses of a new hydrothermally treated, calcium-incorporated surface against other commercially available implant surfaces. Materials and Methods: Four different surfaces were the subject of comparison in this study: machined (MC), resorbable blast media (RBM), sandblasted/large-grit/acid-etched (SLA), and calcium-incorporated SLA (Ca-SLA). Surface morphology and roughness were first characterized by scanning electron microscope (SEM) and white light interferometer, respectively. Preosteoblastic MC3T3-E1 cells were then cultured on the titanium surfaces. Cell morphology was observed at 24 hours, 48 hours, 7 days, and 15 days by SEM; differentiation was assessed at 7, 11, and 15 days by assaying alkaline phosphatase (ALP) activity and osteocalcin (OCN) levels. Results: Surface characterization revealed nanotopographic features on Ca-SLA. At topographic analysis, SLA and Ca-SLA showed similar ...
Exogenous bone morphogenetic proteins (Bmp) are well known to induce ectopic bone formation, but the physiological effect of Bmp signaling on normal bone is not completely understood. By deleting the receptor Bmpr1a in osteoblast-lineage cells with Dmp1-Cre, we observed a dramatic increase in trabecular bone mass in postnatal mice, due to a marked increase in osteoblast number likely driven by hyperproliferation of Sp7+ preosteoblasts. Similarly, inducible deletion of Bmpr1a in Sp7-positive cells specifically in postnatal mice increased trabecular bone mass. However, deletion of Smad4 by the same approaches had only a minor effect, indicating that Bmpr1a signaling suppresses trabecular bone formation through effectors beyond Smad4. Besides increasing osteoblast number in the trabecular bone, deletion of Bmpr1a by Dmp1-Cre also notably reduced osteoblast activity, resulting in attenuation of periosteal growth. The impairment in osteoblast activity correlated with reduced mTORC1 signaling in vivo, ...
Genetic studies show that Msx2 and Dlx5 homeodomain (HD) proteins support skeletal development, but null mutation of the closely related Dlx3 gene results in early embryonic lethality. Here we find that expression of Dlx3 in the mouse embryo is associated with new bone formation and regulation of osteoblast differentiation. Dlx3 is expressed in osteoblasts, and overexpression of Dlx3 in osteoprogenitor cells promotes, while specific knock-down of Dlx3 by RNA interference inhibits, induction of osteogenic markers. We characterized gene regulation by Dlx3 in relation to that of Msx2 and Dlx5 during osteoblast differentiation. Chromatin immunoprecipitation assays revealed a molecular switch in HD protein association with the bone-specific osteocalcin (OC) gene. The transcriptionally repressed OC gene was occupied by Msx2 in proliferating osteoblasts, while Dlx3, Dlx5, and Runx2 were recruited postproliferatively to initiate transcription. Dlx5 occupancy increased over Dlx3 in mature osteoblasts at the
Adhesion of Human Osteoblasts Cell on TiN Thin Film Deposited by Cathodic Arc Plasma Deposition - Human osteoblast;TiN film;Cell adhesion;Surface modification;Cytoskeleton;
Glucocorticoids are known to increase the cyclic AMP response to parathyroid hormone (PTH) in cultured bone organs or bone cells. Using the osteoblast-like cell line ROS 17/2.8, which possesses receptors for both PTH and glucocorticoids, we investigated which component of the complex hormone receptor-guanine nucleotide regulatory unit-adenylate cyclase was affected by dexamethasone treatment. In response to PTH, isoproterenol or forskolin, a compound that is supposed to act directly on the catalytic unit, cyclic AMP production by intact cells and adenylate cyclase activity in purified plasma membrane were markedly increased by dexamethasone. Whereas NaF, guanosine 5′-[beta gamma-imido]triphosphate and Mn/ stimulated adenylate cyclase activity were similarly enhanced in membranes isolated from glucocorticoid-treated cells, the activity of the stimulatory guanine nucleotide regulatory unit, as assessed by reconstitution into membranes from the CYC- clone, which is genetically devoid of this ...
Accumulating evidence suggests that extracellular nucleotides, signaling through P2 receptors, play a role in modulating bone cell function. ATP and ADP stimulate osteoclastic resorption, while ATP and UTP are powerful inhibitors of bone formation by osteoblasts. We investigated changes in the expression of P2 receptors with cell differentiation in primary osteoblast cultures. Rat calvarial osteoblasts, cultured for up to 10 days, were loaded with the intracellular Ca(2+)-sensing fluorophore, Fluo-4 AM, and a fluorescence imaging plate reader was used to measure responses to nucleotide agonists. Peak responses occurred within 20 s and were evoked by ATP or UTP at concentrations as low as 2 microM. Osteoblast number doubled between day 4 and 10 of culture, but the peak intracellular Ca(2+) response to ATP or UTP increased up to 6-fold over the same period, indicating that osteoblast responsiveness to nucleotides increases as cell differentiation proceeds. The approximate order of potency for the ...
Pereira, B.P., Aung, K.Z., Pho, R.W.H., Cool, S.M., Nathan, S.S., Zhou, Y., Gupta, A., Leong, D.T., Salto-Tellez, M., Van, Wijnen A.J., Ling, L., Galindo, M., Stein, G.S. (2009). Runx2, p53, and pRB status as diagnostic parameters for deregulation of osteoblast growth and differentiation in a new pre-chemotherapeutic osteosarcoma cell line (OS1). Journal of Cellular Physiology 221 (3) : 778-788. [email protected] Repository. https://doi.org/10.1002/jcp. ...
1|i|α|/i|,25-Dihydroxyvitamin D|sub|3|/sub| (1,25(OH)|sub|2|/sub|D|sub|3|/sub|), the active metabolite of vitamin D (Vit D), increases intestinal absorption of calcium and phosphate, maintaining a correct balance of bone remodeling. Vit D has an anabolic effect on the skeletal system and is key in promoting osteoblastic differentiation of human Mesenchymal Stem Cells (hMSCs) from bone marrow. MSCs can be also isolated from the immature form of the tooth, the dental bud: Dental Bud Stem Cells (DBSCs) are adult stem cells that can effectively undergo osteoblastic differentiation. In this work we investigated the effect of Vit D on DBSCs differentiation into osteoblasts. Our data demonstrate that DBSCs, cultured in an opportune osteogenic medium, differentiate into osteoblast-like cells; Vit D treatment stimulates their osteoblastic features, increasing the expression of typical markers of osteoblastogenesis like RUNX2 and Collagen I (Coll I) and, in a more important way, determining a higher
Growth and differentiation of osteoblasts are often studied in cell cultures. In vivo, however, osteoblasts are embedded within a complex three-dimensional (3D) microenvironment, which bears little relation to standard culture flasks. Our study characterizes osteoblast-like cells cultured in 3D collagen gels and compares them with cells in two-dimensional (2D) cultures. Primary rat osteoblasts and MC3T3-E1 cells were seeded within type I collagen gels, and differentiation was determined by mineral staining and gene expression analysis. Cells growing in 3D gels showed positive mineral staining and induction of osteoblast marker genes earlier than cells growing in 2D. A number of genes, including osteocalcin, bone sialoprotein, alkaline phosphatase and dentin matrix protein 1, were already highly upregulated in 3D cultures 24 h after seeding. The early expression of osteoblast genes was dependent on the 3D structure and was not induced in cells growing on collagen-coated dishes in 2D. Comparison of
The relationship of proliferation to the developmental sequence associated with bone cell differentiation was examined in primary osteoblast cultures derived from fetal rat and embryonic chick calvaria. A reciprocal and functional relationship exists between the decline in proliferative activity which occurs during the initial stages of the developmental sequence and the induction of genes encoding osteoblast phenotype proteins associated with matrix maturation and mineralization. This relationship is supported by 1) a temporal sequence of events in which there is an enhanced expression of alkaline phosphatase (AP) and osteopontin (OP) genes immediately following the proliferative period and expression of osteocalcin with the onset of mineralization, and 2) increases in AP and OP when DNA synthesis is inhibited. By determining cellular mRNA levels and rates of mRNA synthesis in isolated nuclei, we found that the down-regulation of cell growth-related genes is modified at both the levels of transcription
More than 95% of TAF gets eliminated from plasma 2 hours after dosing. To mimic that process, the researchers pulsed TAF into PBMCs and primary human osteoblasts for 2 hours, followed by a washout. They measured TFV-DP in cells collected at multiple points after dosing. The investigators conducted PBMC loading experiments with multiple TAF concentrations to find the concentration that results in intracellular TFV-DP levels similar to those seen in vivo (677 nM). They then evaluated similar TAF concentrations in primary osteoblasts. Next the Gilead team developed a primary osteoblast cell growth assay and evaluated TFV-DP levels after single and multiple TAF pulses. They assessed cell viability after treating primary osteoblasts with TAF for 3 days ...
Alkaline phosphatase activity on osteoblast - posted in Tissue and Cell Culture: HelloI have cultured human fetal osteoblast (hFOB 1.19) and I need to check for the alkaline phosphatase activity (ALP) for that cells.I use the plant extract as for the treatment for differentiation and proliferation of the osteoblast.I never did ALP test before. Does everybody can help regarding the protocol for ALP assay?I will use ALP kit from RANDOX.Thank you for help.
TY - JOUR. T1 - The effects of cytokines and growth factors on osteoblastic cells. AU - Mundy, G. R.. AU - Boyce, B.. AU - Hughes, D.. AU - Wright, K.. AU - Bonewald, L.. AU - Dallas, S.. AU - Harris, S.. AU - Ghosh-Choudhury, N.. AU - Chen, D.. AU - Dunstan, C.. AU - Izbicka, E.. AU - Yoneda, T.. PY - 1995/8. Y1 - 1995/8. N2 - In this short review, some regulatory mechanisms that are involved in the control of normal bone formation are proposed, based on several in vivo and in vitro models our group has utilized recently to study osteoblast differentiation and mineralized bone matrix formation. Of course, these proposals must be assessed in the light of the limitations of the models, which probably represent a simplification of the complex and different ways in which normal mammalian bone is formed at different sites. Nevertheless, it is likely that the same general types of control mechanisms are active in each of the different types of bone formation. In adult humans, bone formation ...
The exposure of human osteoblastic cells to EMD, TGF-β1 and EMD+TGF-β1 resulted in early increased cell proliferation, and reduced ALP activity and matrix mineralization. The present results are corroborated by several works that observed EMD stimulation of the proliferative capacity of both osteoblastic cells[14, 16, 36] and PDL fibroblasts[10, 12, 13, 20, 22]. In contrast to PDL fibroblast response to EMD, which shows signs of matrix mineralization when EMD are used even at earlier time points[13], osteoblastic cell cultures seem to be inhibited in terms of osteogenic differentiation. Interestingly, the association of EMD and exogenous TGF-β1 did not alter the osteogenic potential of the cultures.. Although the results of the present study point toward the development of a less differentiated osteoblastic phenotype when cells were exposed to EMD, TGF-β1 or EMD+TGF-β1, no morphologic differences were observed among the groups. Cell morphology was considered within the typical features of ...
To examine the effect of fluoride on the survival of osteoblasts, media containing various concentrations of fluoride, 0〗5.0×10,sup,-2,/sup, mol/L, were prepared and mouse osteoblast MC3T3-E1 cells were cultured. At the same time, media containing 0〗5.0×10,sup,-2,/sup, mol/L of chloride as an indicator of pH were also adjusted. Although osteoblasts grew well at fluoride levels below 1.0×10,sup,-3,/sup, mol/L, osteoblast survival was dramatically inhibited over 5.0×10,sup,-3,/sup, mol/L of fluoride despite the pH of the medium scarcely decreasing when compared with the control pH 7.81, and osteoblasts showing favorable survival with 2.0×10,sup,-2,/sup, mol/L of chloride even at pH 6.81. Fluorospectrophotometric observation showed quite different features between control cells and at higher concentrations of fluoride. These results suggested that fluoride dramatically inhibits osteoblast growth at concentrations above 5.0×10,sup,-3,/sup, mol/L in the medium solution.. ...
Staphylococcus aureus is one of the most frequently involved pathogens in bacterial infections such as skin abscess, pneumonia, endocarditis, osteomyelitis and implant-associated infection. As for bone homeostasis, it is partly altered during infections by Staphylococcus aureus by the induction of various responses from osteoblasts, which are the bone-forming cells responsible for extracellular matrix synthesis and its mineralization. Nevertheless, bone-forming cells are a heterogeneous population with different stages of maturation and the impact of the latter on their responses toward bacteria remains unclear. We describe the impact of Staphylococcus aureus on two populations of human primary bone-forming cells which have distinct maturation characteristics in both acute and persistent models of interaction. Cell maturation did not influence the internalization and survival of Staphylococcus aureus inside bone-forming cells or the cell death related to the infection. By studying the expression of
Primary osteoblast cultures from calvaria of newborn, wild-type, and Lrp5−/− mice were established and mineralized in vitro as previously described (Ducy et al., 1999). Transfections were performed in 24-well plates (20,000 cells/well; Fugene6; Roche) in triplicate and repeated at least three times; cells were harvested 40 h after transfection. Empty vector was added to keep the total amount of DNA constant at 225 ng/well. The FOPtkluc reporter, containing mutated Lef1 binding sites, was used in control transfections. All luciferase values were corrected for β-galactosidase activity as a control for transfection efficiency. For Western blot analysis, primary osteoblast lysates were collected in physiological buffer with protease inhibitor cocktail (Roche), homogenized, fractionated into cytosolic and membrane fractions by ultracentrifugation at 100,000 g for 90 min, and separated by SDS-PAGE. The epitope tags were detected by the anti-FLAG M2 antibody (Sigma-Aldrich) and anti-HA antibody ...
Dentin matrix protein 1 (DMP1) is an acidic extracellular matrix phosphoprotein that can bind calcium. DMP1 is required for bone and dentin mineralization and is expressed in the cells of bone and teeth. It is thought to play a role in regulating expression of osteoblast-specific genes during osteoblast cell differentiation and is localized to the nucleus in osteoblast precursor cells. In mature osteoblasts, the DMP1 protein is phosphorylated and localized to the extracellular matrix, where it plays a role in forming mineralized matrix. While other constitutively expressed proteins in the extracellular matrix, such as osteopontin and osteocalcin, are expressed in osteoblast cells, DMP1 is expressed in osteocytes, making it a candidate biomarker for osteocyte activity.. ...
Dentin matrix protein 1 (DMP1) is an acidic extracellular matrix phosphoprotein that can bind calcium. DMP1 is required for bone and dentin mineralization and is expressed in the cells of bone and teeth. It is thought to play a role in regulating expression of osteoblast-specific genes during osteoblast cell differentiation and is localized to the nucleus in osteoblast precursor cells. In mature osteoblasts, the DMP1 protein is phosphorylated and localized to the extracellular matrix, where it plays a role in forming mineralized matrix. While other constitutively expressed proteins in the extracellular matrix, such as osteopontin and osteocalcin, are expressed in osteoblast cells, DMP1 is expressed in osteocytes, making it a candidate biomarker for osteocyte activity.. ...
TY - JOUR. T1 - Effects of nano-engineered surfaces on osteoblast adhesion, growth, differentiation, and apoptosis. AU - Miralami, Raheleh. AU - Sharp, John G.. AU - Namavar, Fereydoon. AU - Hartman, Curtis W.. AU - Garvin, Kevin L.. AU - Thiele, Geoffrey M.. PY - 2019/1/1. Y1 - 2019/1/1. N2 - Modifying implant surfaces to improve their biocompatibility by enhancing osteoblast activation, growth, differentiation, and induction of greater bone formation with stronger attachments should result in improved outcomes for total joint replacement surgeries. This study tested the hypothesis that nano-structured surfaces, produced by the ion beam-assisted deposition method, enhance osteoblast adhesion, growth, differentiation, bone formation, and maturation. The ion beam-assisted deposition technique was employed to deposit zirconium oxide films on glass substrates. The effects of the ion beam-assisted deposition technique on cellular functions were investigated by comparing adhesion, proliferation, ...
We have previously shown that targeted expression of a dominant-negative truncated form of N-cadherin (Cdh2) delays acquisition of peak bone mass in mice and retards osteoblast differentiation; whereas deletion of cadherin 11 (Cdh11), another osteoblast cadherin, leads to only modest osteopenia. To determine the specific roles of these two cadherins in the adult skeleton, we generated mice with an osteoblast/osteocyte specific Cdh2 ablation (cKO) and double Cdh2+/−;Cdh11−/− germline mutant mice. Age-dependent osteopenia and smaller diaphyses with decreased bone strength characterize cKO bones. By contrast, Cdh2+/−;Cdh11−/− exhibit severely reduced trabecular bone mass, decreased in vivo bone formation rate, smaller diaphyses and impaired bone strength relative to single Cdh11 null mice. The number of bone marrow immature precursors and osteoprogenitor cells is reduced in both cKO and Cdh2+/−;Cdh11−/− mice, suggesting that N-cadherin is involved in maintenance of the stromal ...
Recent studies have suggested the existence of osteoblastic cells in the circulation, but the origin and role of these cells in vivo are not clear. Here, we examined how these cells contribute to osteogenesis in a bone morphogenetic protein (BMP)-induced model of ectopic bone formation. Following lethal dose-irradiation and subsequent green fluorescent protein-transgenic bone marrow cell-transplantation (GFP-BMT) in mice, a BMP-2-containing collagen pellet was implanted into muscle. Three weeks later, a significant number of GFP-positive osteoblastic cells were present in the newly generated ectopic bone. Moreover, peripheral blood mononuclear cells (PBMNCs) from the BMP-2-implanted mouse were then shown to include osteoblast progenitor cells (OPCs) in culture. Passive transfer of the PBMNCs isolated from the BMP-2-implanted GFP-mouse to the BMP-2-implanted nude mouse led to GFP-positive osteoblast accumulation in the ectopic bone. These data provide new insight into the mechanism of ectopic ...
A cDNA library prepared from the mouse osteoblastic cell line MC3T3-E1 was screened for the presence of specifically expressed genes by employing a combined subtraction hybridization/differential screening approach. A cDNA was identified and sequenced which encodes a protein designated osteoblast-specific factor 2 (OSF-2) comprising 811 amino acids. OSF-2 has a typical signal sequence, followed by a cysteine-rich domain, a fourfold repeated domain and a C-terminal domain. The protein lacks a typical transmembrane region. The fourfold repeated domain of OSF-2 shows homology with the insect protein fasciclin I. RNA analyses revealed that OSF-2 is expressed in bone and to a lesser extent in lung, but not in other tissues. Mouse OSF-2 cDNA was subsequently used as a probe to clone the human counterpart. Mouse and human OSF-2 show a high amino acid sequence conservation except for the signal sequence and two regions in the C-terminal domain in which in-frame insertions or deletions are observed, ...
TY - JOUR. T1 - Glucocorticoid enhances the expression of dickkopf-1 in human osteoblasts. T2 - Novel mechanism of glucocorticoid-induced osteoporosis. AU - Ohnaka, Keizo. AU - Taniguchi, Hiroshi. AU - Kawate, Hisaya. AU - Nawata, Hajime. AU - Takayanagi, Ryoichi. PY - 2004/5/21. Y1 - 2004/5/21. N2 - To clarify the underlying mechanism of glucocorticoid-induced osteoporosis, we investigated the effect of glucocorticoid on the expression of dickkopf-1 (Dkk-1), an antagonist of Wnt signaling, in primary cultured human osteoblasts. Dexamethasone markedly induced the expression of mRNA for Dkk-1 in a dose- and time-dependent manner. The expression of Kremen1, a receptor for Dkk, did not change by the treatment with dexamethasone, while that of low-density lipoprotein receptor-related protein 5 (LRP5), a Wnt coreceptor, slightly decreased by the treatment with dexamethasone. Dexamethasone increased the transcriptional activity of the Dkk-1 gene promoter in human osteoblasts. Serial deletion and ...
Betaine (BET), a component of many foods, is an essential osmolyte and a source of methyl groups; it also shows an antioxidant activity. Moreover, BET stimulates muscle differentiation via insulin like growth factor I (IGF-I). The processes of myogenesis and osteogenesis involve common mechanisms with skeletal muscle cells and osteoblasts sharing the same precursor. Therefore, we have hypothesized that BET might be effective on osteoblast cell differentiation. The effect of BET was tested in human osteoblasts (hObs) derived from trabecular bone samples obtained from waste material of orthopedic surgery. Cells were treated with 10 mM BET at 5, 15, 60 min and 3, 6 and 24 h. The possible effects of BET on hObs differentiation were evaluated by real time PCR, western blot and immunofluorescence analysis. Calcium imaging was used to monitor intracellular calcium changes. Real time PCR results showed that BET stimulated significantly the expression of RUNX2, osterix, bone sialoprotein and osteopontin. Western
Levesque, Jean-Pierre, Bendall, Linda J., Pettit, Allison R., Raggatt, Liza, Jacobsen, Rebecca, Barbier, Valarie, Nowlan, Bianca, Shen, Yi Shen, Sims, Natalie A. and Winkler, Ingrid G. (2011). Mobilizing agents G-CSF, cyclophosphamide or AMD3100 (Plerixafor) Have distinct effects on osteoblasts, hematopoietic stem cell niches, and B-Lymphopoiesis. In: Abstracts of the American Society of Hematology 53rd Annual Meeting. 53rd Annual Meeting and Exposition of the American Society of Hematology (ASH), San Diego CA, United States, (1713-1713). 10-13 December 2011. ...
The Runx2 transcription factor is critical for commitment to the osteoblast lineage. However, its role in committed osteoblasts and its functions during postnatal skeletogenesis remain unclear. We established a Runx2-floxed line with insertion of loxP sites around exon 8 of the Runx2 gene. Runx2 protein lacking the region encoded by exon 8 is imported into the nucleus and binds target DNA, but exhibits diminished transcriptional activity. We specifically deleted the Runx2 gene in committed osteoblasts using 2.3kb col1a-Cre transgenic mice. Surprisingly, the homozygous Runx2 mutant mice were born alive. The Runx2 heterozygous and homozygous null were grossly indistinguishable from wild-type littermates at birth. Runx2 deficiency did not alter proliferative capacity of osteoblasts during embryonic development (E18). Chondrocyte differentiation and cartilage growth in mutants was similar to wild-type mice from birth to 3 months of age. Analysis of the embryonic skeleton revealed poor calcification ...
Currently, no model is available to study the cellular and molecular events associated with bone metastases of prostate cancer. This study shows that MDA PCa 2a and MDA PCa 2b cells induce a specific and reproducible increase in osteoblast differentiation and proliferation when the cells share the medium during coculturing. Osteoblast differentiation in this system was associated with up-regulation of the osteoblast-specific transcriptor factor Cbfa1. Moreover, up-regulation of Cbfa1 and Osteocalcin expression was also induced in PMOs by CM produced by MDA PCa 2b cells, suggesting that soluble factors produced by prostate cancer cells promote osteoblast differentiation and that Cbfa1 mediates this effect. To our knowledge, this is the first in vitro model of bone metastasis from prostate cancer that recapitulates the osteoblastic phenotype typical of the disease. These results confirmed in vivo and at the molecular level, suggest that the pathophysiology of osteoblastic bone metastases from ...
Osteoblasts are derived from mesenchymal tissue along a tightly regulated pathway. Mesenchymal cells can form connective tissue fibroblasts, adipocytes, and bone. Differentiation of mesenchymal cell into osteoblasts occurs along a pathway involving regulation by autocrine, paracrine, and endocrine factors. Endocrine factors including parathyroid hormone, growth hormone, and insulin-like growth factor stimulate proliferation and in certain instances differentiation for pre-osteoblastic cells.. Critically, RUNX2, a nuclear transcription factor, must be expressed for a mesenchymal cell to differentiate along an osteoblastic lineage. The mechanisms by which expression of RUNX2 lead eventually to an osteoblast phenotype is beyond the scope of this chapter. However, it should be noted that bone morphogenetic proteins (BMPs) are critical for the induction of RUNX2 expression. BMPs are a member of the transforming growth factor family of proteins, and to date 30 have been identified. BMPs are present in ...
Ca2+ is one of the most important second messengers and regulates many cellular processes (Berridge et al., 2000b), and the CaM pathway is critical for osteoblast differentiation (Zayzafoon, 2006). However, the mechanism for controlling [Ca2+]i during osteoblast differentiation is not yet clear. We found that Panx3 functions as a Ca2+ channel in the ER, through which it regulates [Ca2+]i. C2C12 and calvarial cells express IP3Rs. Both Panx3 and IP3Rs function as ER Ca2+ channels; however, their activation mechanisms are different. We found that the Panx3 ER Ca2+ channel was activated through Akt signaling (Fig. 7 A), distinct from IP3-mediated activation of ubiquitous IP3R ER Ca2+ channels (Mikoshiba, 2007). In addition, siRNA for IP3R3 and 2-APB, which is an inhibitor of IP3R-mediated Ca2+ release, inhibited IP3R ER Ca2+ channel activity, but did not inhibit the Panx3 channel (Fig. 5 A). The role of IP3Rs in osteogenic differentiation is also not yet clear. The inhibition of endogenous Panx3 by ...
Osteoblasts are responsible for deposition of the mineralized bone matrix. Osteoblasts attach to the surface of the bone matrix, proliferate, and secrete type I collagen and bone-specific non-collagenous proteins such as osteocalcin. These proteins are crucial for mineralization of the bone matrix ...
A specific oligodeoxynucleotide (ODN), ODN MT01, was found to have positive effects on the proliferation and activation of the osteoblast-like cell line MG 63. In this study, the detailed signaling pathways in which ODN MT01 promoted the differentiation of osteoblasts were systematically examined. ODN MT01 enhanced the expression of osteogenic marker genes, such as osteocalcin and type I collagen. Furthermore, ODN MT01 activated Runx2 phosphorylation via ERK1/2 mitogen-activated protein kinase (MAPK) and p38 MAPK. Consistently, ODN MT01 induced up-regulation of osteocalcin, alkaline phosphatase (ALP) and type I collagen, which was inhibited by pre-treatment with the ERK1/2 inhibitor U0126 and the p38 inhibitor SB203580. These results suggest that the ERK1/2 and p38 MAPK pathways, as well as Runx2 activation, are involved in ODN MT01-induced up-regulation of osteocalcin, type I collagen and the activity of ALP in MG 63 cells.
c-Src and IL-6 inhibit osteoblast differentiation and integrate IGFBP5 signalling.: Interleukin-6 (IL-6) and c-Src impair osteoblast maturation in vitro and in
A regulated expression of 394 known genes and 295 ESTs was tedected. The sensitivity and realiability of detection by microarrays was shown by confirming the expression pattern for 20 genes by radioactive quantitative RT-PCR. Extensive functional classification of regulated genes was performed. The most interesting finding was concomitant activation of TGF-β, Wnt and Notch signaling pathways, confirmed by strong upregulation of their target genes by PCR. The TGF-β pathway is activated by stimulated production of the growth factor itself, while the exact mechanism of Wnt and Notch activation remains elusive. We showed BMP-2 stimulated expression of Hey1, a direct Notch target gene, in mouse MC3T3 and C2C12 cells, in human mesenchymal cells and in mouse calvaria. Small interfering RNA-mediated inhibition of Hey1 induction led to an increase in osteoblast matrix mineralization, suggesting that Hey1 is a negative regulator of osteoblast maturation. This negative regulation is apparently achieved ...
The molecular mechanisms that transduce the osteoblast response to physical forces in the bone microenvironment are poorly understood. Here, we used genetic and pharmacological experiments to determine whether the polycystins PC1 and PC2 (encoded by Pkd1 and Pkd2) and the transcriptional coactivator TAZ form a mechanosensing complex in osteoblasts. Compound-heterozygous mice lacking 1 copy of Pkd1 and Taz exhibited additive decrements in bone mass, impaired osteoblast-mediated bone formation, and enhanced bone marrow fat accumulation. Bone marrow stromal cells and osteoblasts derived from these mice showed impaired osteoblastogenesis and enhanced adipogenesis. Increased extracellular matrix stiffness and application of mechanical stretch to multipotent mesenchymal cells stimulated the nuclear translocation of the PC1 C-terminal tail/TAZ (PC1-CTT/TAZ) complex, leading to increased runt-related transcription factor 2-mediated (Runx2-mediated) osteogenic and decreased PPARγ-dependent adipogenic ...
Definition of osteoblast in the Definitions.net dictionary. Meaning of osteoblast. What does osteoblast mean? Information and translations of osteoblast in the most comprehensive dictionary definitions resource on the web.
Ion irradiation is a very promising tool to modify the chemical structure and physical properities of polymers. This study was aimed to evaluate the cellular adhesion to ion beam-irradiated surface of biodegradable poly-l-lactide(PLLA) membrane. The PLLA membrane samples were irradiated by using 35 KeV to fluence of , and . Water contact angles to control and each dose of ion beam-irradiated PLLA membranes were measured. Cultured fetal rat calvarial osteoblasts were seeded onto control and each dose of ion beam-irradiated PLLA membranes and cultured. After 24 hours, each PLLA membranes onto which osteoblasts attached were examined by scanning electron microscopy(SEM). Osteoblasts were removed from each PLLA membrane and then, the vitality and the number of cells were calibrated. Alkaline phosphatase of detached cells from each PLLA membranes were measured. Ion beam-irradiated PLLA membranes showed no significantly morphological change from control PLLA membranes. In the measurement of water ...
TY - JOUR. T1 - Function of Fos proteins in bone cell differentiation. AU - Matsuo, K.. AU - Jochum, W.. AU - Owens, J. M.. AU - Chambers, T. J.. AU - Wagner, E. F.. PY - 1999/7/1. Y1 - 1999/7/1. UR - http://www.scopus.com/inward/record.url?scp=0344867028&partnerID=8YFLogxK. UR - http://www.scopus.com/inward/citedby.url?scp=0344867028&partnerID=8YFLogxK. U2 - 10.1016/S8756-3282(99)00120-9. DO - 10.1016/S8756-3282(99)00120-9. M3 - Article. C2 - 10423040. AN - SCOPUS:0344867028. VL - 25. JO - Bone. JF - Bone. SN - 8756-3282. IS - 1. ER - ...
The small GTP-binding protein Rad (RRAD, Ras associated with diabetes) is the founding member of the RGK (Rad, Rem, Rem2, and Gem/Kir) family that regulates cardiac voltage-gated Ca(2+) channel function. However, its cellular and physiological functions outside of the heart remain to be elucidated. Here we report that Rad GTPase function is required for normal bone homeostasis in mice, as Rad deletion results in significantly lower bone mass and higher bone marrow adipose tissue (BMAT) levels. Dynamic histomorphometry in vivo and primary calvarial osteoblast assays in vitro demonstrate that bone formation and osteoblast mineralization rates are depressed, while in vitro osteoclast differentiation is increased, in the absence of Rad ...
osteoblast cell for scanning electron microscope - posted in Tissue and Cell Culture: i grow osteoblast cell and now it is in incubator. how i need to prepare that to view the image under scanning electron microscope? what steps i do?
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