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Calcium Deposition (calcium + deposition)
Selected AbstractsThe solute budget of a forest catchment and solute fluxes within a Pinus radiata and a secondary native forest site, southern ChileHYDROLOGICAL PROCESSES, Issue 13 2002Geertrui Y. P. Uyttendaele Abstract Solute concentrations and fluxes in rainfall, throughfall and stemflow in two forest types, and stream flow in a 90 ha catchment in southern Chile (39°44,S, 73°10,W) were measured. Bulk precipitation pH was 6·1 and conductivity was low. Cation concentrations in rainfall were low (0·58 mg Ca2+ l,1, 0·13 mg K+ l,1, 0·11 mg Mg2+ l,1 and <0·08 mg NH4,N l,1), except for sodium (1·10 mg l,1). Unexpected high levels of nitrate deposition in rainfall (mean concentration 0·38 mg NO3,N l,1, total flux 6·3 kg NO3,N ha,1) were measured. Concentrations of soluble phosphorous in bulk precipitation and stream flow were below detection limits (<0·09 mg l,1) for all events. Stream-flow pH was 6·3 and conductivity was 28·3 ,s. Stream-water chemistry was also dominated by sodium (2·70 mg l,1) followed by Ca, Mg and K (1·31, 0·70 and 0·36 mg l,1). The solute budget indicated a net loss of 3·8 kg Na+ ha,1 year,1, 5·4 kg Mg2+ ha,1 year,1, 1·5 kg Ca2+ ha,1 year,1 and 0·9 kg K+ ha,1 year,1, while 4·9 kg NO3,N ha,1 year,1 was retained by the ecosystem. Stream water is not suitable for domestic use owing to high manganese and, especially, iron concentrations. Throughfall and stemflow chemistry at a pine stand (Pinus radiata D. Don) and a native forest site (Siempreverde type), both located within the catchment, were compared. Nitrate fluxes within both forest sites were similar (1·3 kg NO3,N ha,1 year,1 as throughfall). Cation fluxes in net rainfall (throughfall plus stemflow) at the pine stand generally were higher (34·8 kg Na+ ha,1 year,1, 21·5 kg K+ ha,1 year,1, 5·1 kg Mg2+ ha,1 year,1) compared with the secondary native forest site (24·7 kg Na+ ha,1 year,1, 18·9 kg K+ ha,1 year,1 and 4·4 kg Mg2+ ha,1 year,1). However, calcium deposition beneath the native forest stand was higher (15·9 kg Ca2+ ha,1 year,1) compared with the pine stand (12·6 kg Ca2+ ha,1 year,1). Copyright © 2002 John Wiley & Sons, Ltd. [source] Formation of three-dimensional cell/polymer constructs for bone tissue engineering in a spinner flask and a rotating wall vessel bioreactorJOURNAL OF BIOMEDICAL MATERIALS RESEARCH, Issue 1 2002Vassilios I. Sikavitsas Abstract The aim of this study is to investigate the effect of the cell culture conditions of three-dimensional polymer scaffolds seeded with rat marrow stromal cells (MSCs) cultured in different bioreactors concerning the ability of these cells to proliferate, differentiate towards the osteoblastic lineage, and generate mineralized extracellular matrix. MSCs harvested from male Sprague,Dawley rats were culture expanded, seeded on three-dimensional porous 75:25 poly(D,L -lactic- co -glycolic acid) biodegradable scaffolds, and cultured for 21 days under static conditions or in two model bioreactors (a spinner flask and a rotating wall vessel) that enhance mixing of the media and provide better nutrient transport to the seeded cells. The spinner flask culture demonstrated a 60% enhanced proliferation at the end of the first week when compared to static culture. On day 14, all cell/polymer constructs exhibited their maximum alkaline phosphatase activity (AP). Cell/polymer constructs cultured in the spinner flask had 2.4 times higher AP activity than constructs cultured under static conditions on day 14. The total osteocalcin (OC) secretion in the spinner flask culture was 3.5 times higher than the static culture, with a peak OC secretion occurring on day 18. No considerable AP activity and OC secretion were detected in the rotating wall vessel culture throughout the 21-day culture period. The spinner flask culture had the highest calcium content at day 14. On day 21, the calcium deposition in the spinner flask culture was 6.6 times higher than the static cultured constructs and over 30 times higher than the rotating wall vessel culture. Histological sections showed concentration of cells and mineralization at the exterior of the foams at day 21. This phenomenon may arise from the potential existence of nutrient concentration gradients at the interior of the scaffolds. The better mixing provided in the spinner flask, external to the outer surface of the scaffolds, may explain the accelerated proliferation and differentiation of marrow stromal osteoblasts, and the localization of the enhanced mineralization on the external surface of the scaffolds. © 2002 Wiley Periodicals, Inc. J Biomed Mater Res 62: 136,148, 2002 [source] Role of TNF alpha and PLF in bone remodeling in a rat model of repetitive reaching and grasping,JOURNAL OF CELLULAR PHYSIOLOGY, Issue 1 2010Shobha Rani We have previously developed a voluntary rat model of highly repetitive reaching that provides an opportunity to study effects of non-weight bearing muscular loads on bone and mechanisms of naturally occurring inflammation on upper limb tissues in vivo. In this study, we investigated the relationship between inflammatory cytokines and matricellular proteins (Periostin-like-factor, PLF, and connective tissue growth factor, CTGF) using our model. We also examined the relationship between inflammatory cytokines, PLF and bone formation processes. Rats underwent initial training for 5 weeks, and then performed a high repetition high force (HRHF) task (12,reaches/min, 60% maximum grip force, 2,h/day, 3 days/week) for 6 weeks. We then examined the effect of training or task performance with or without treatment with a rat specific TNF, antibody on inflammatory cytokines, osteocalcin (a bone formation marker), PLF, CTGF, and behavioral indicators of pain or discomfort. The HRHF task decreased grip strength and induced forepaw mechanical hypersensitivity in both trained control and 6-week HRHF animals. Two weeks of anti-TNF, treatment improved grip strength in both groups, but did not ameliorate forepaw hypersensitivity. Moreover, anti-TNF, treatment attenuated task-induced increases in inflammatory cytokines (TNF,, IL-1,, and MIP2 in serum; TNF, in forelimb bone and muscles) and serum osteocalcin in 6-week HRHF animals. PLF levels in forelimb bones and flexor digitorum muscles increased significantly in 6-week HRHF animals, increases attenuated by anti-TNF, treatment. CTGF levels were unaffected by task performance or anti-TNF, treatment in 6-week HRHF muscles. In primary osteoblast cultures, TNF,, MIP2 and MIP3a treatment increased PLF levels in a dose dependent manner. Also in primary osteoblast cultures, increased PLF promoted proliferation and differentiation, the latter assessed by measuring Runx2, alkaline phosphatase (ALP) and osteocalcin mRNA levels; ALP activity; as well as calcium deposition and mineralization. Increased PLF also promoted cell adhesion in MC3T3-E1 osteoblast-like cell cultures. Thus, tissue loading in vivo resulted in increased TNF,, which increased PLF, which then induced anabolic bone formation, the latter results confirmed in vitro. J. Cell. Physiol. 225: 152,167, 2010. © 2010 Wiley-Liss, Inc. [source] Constitutive expression of thrombospondin 1 in MC3T3-E1 osteoblastic cells inhibits mineralizationJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2006Akemichi Ueno Thrombospondin 1 (TSP1) is a multifunctional extracellular glycoprotein present mainly in the fetal and adult skeleton. Although an inhibitory effect of TSP1 against pathological mineralization in cultured vascular pericytes has been shown, its involvement in physiological mineralization by osteoblasts is still unknown. To determine the role of TSP1 in biomineralization, mouse osteoblastic MC3T3-E1 cells were cultured in the presence of antisense phosphorothioate oligodeoxynucleotides complementary to the TSP1 sequence. The 18- and 24-mer antisense oligonucleotides caused concentration-dependent increases in the number of mineralized nodules, acid-soluble calcium deposition in the cell/matrix layer, and alkaline phosphatase activity within 9 days, without affecting cell proliferation. The corresponding sense or scrambled oligonucleotides did not affect these parameters. In the antisense oligonucleotide-treated MC3T3-E1 cells, thickened extracellular matrix, well-developed cell processes, increased intracellular organelles, and collagen fibril bundles were observed. On the other hand, the addition of TSP1 to the culture decreased the production of a mineralized matrix by MC3T3-E1 cells. Furthermore, MC3T3-E1 clones overexpressing mouse TSP1 were established and assayed for TSP1 protein and their capacity to mineralize. TSP1 dose-dependently inhibited mineralization by these cells both in vitro and in vivo. These results indicate that TSP1 functions as an inhibitory regulator of bone mineralization and matrix production by osteoblasts to sustain bone homeostasis. J. Cell. Physiol. 209: 322,332, 2006. © 2006 Wiley-Liss, Inc. [source] Cobalt ions induce chemokine secretion in primary human osteoblasts,JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 7 2009J.M. Queally Abstract Chemokines are major regulators of the inflammatory response and have been shown to play an important role in periprosthetic osteolysis. Titanium particles have previously been shown to induce IL-8 and MCP-1 secretion in osteoblasts. These chemokines result in the chemotaxis and activation of neutrophils and macrophages, respectively. Despite a resurgence in the use of cobalt-chromium-molybdenum alloys in metal-on-metal arthroplasty, cobalt and chromium ion toxicity in the periprosthetic area has been insufficiently studied. In this study we investigate the in vitro effect of cobalt ions on primary human osteoblast activity. We demonstrate that cobalt ions rapidly induce the protein secretion of IL-8 and MCP-1 in primary human osteoblasts. This elevated chemokine secretion is preceded by an increase in the transcription of the corresponding chemokine gene. Using a Transwell migration chemotaxis assay we also demonstrate that the chemokines secreted are capable of inducing neutrophil and macrophage migration. Furthermore, cobalt ions significantly inhibit osteoblast function as demonstrated by reduced alkaline phosphatase activity and calcium deposition. In aggregate these data demonstrate that cobalt ions can activate transcription of the chemokine genes IL-8 and MCP-1 in primary human osteoblasts. Cobalt ions are not benign and may play an important role in the pathogenesis of osteolysis by suppressing osteoblast function and stimulating the production and secretion of chemokines that attract inflammatory and osteoclastic cells to the periprosthetic area. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 855,864, 2009 [source] Ethanol Alters the Osteogenic Differentiation of Amniotic Fluid-Derived Stem CellsALCOHOLISM, Issue 10 2010Jennifer A. Hipp Background:, Fetal alcohol spectrum disorder (FASD) is a set of developmental defects caused by prenatal alcohol exposure. Clinical manifestations of FASD are highly variable and include mental retardation and developmental defects of the heart, kidney, muscle, skeleton, and craniofacial structures. Specific effects of ethanol on fetal cells include induction of apoptosis as well as inhibition of proliferation, differentiation, and migration. This complex set of responses suggests that a bioinformatics approach could clarify some of the pathways involved in these responses. Methods:, In this study, the responses of fetal stem cells derived from the amniotic fluid (AFSCs) to treatment with ethanol have been examined. Large-scale transcriptome analysis of ethanol-treated AFSCs indicates that genes involved in skeletal development and ossification are up-regulated in these cells. Therefore, the effect of ethanol on osteogenic differentiation of AFSCs was studied. Results:, Exposure to ethanol during the first 48 hours of an osteogenic differentiation protocol increased in vitro calcium deposition by AFSCs and increased alkaline phosphatase activity. In contrast, ethanol treatment later in the differentiation protocol (day 8) had no significant effect on the activity of alkaline phosphatase. Conclusions:, These results suggest that transient exposure of AFSCs to ethanol during early differentiation enhances osteogenic differentiation of the cells. [source] Reduced gap junctional intercellular communication and altered biological effects in mouse osteoblast and rat liver oval cell lines transfected with dominant-negative connexin 43MOLECULAR CARCINOGENESIS, Issue 4 2003Brad L. Upham Abstract Gap junctional intercellular communication (GJIC) maintains normal growth and differentiation of cells in a tissue. The intercellular molecules traversing gap junctions are largely unknown, but the molecular weight (MW) cutoff is normally 1200 Da. No differences in dye transfer were observed in normal or vector controls of WB-F344 rat liver epithelial or mouse osteoblastic MC3T3-E1 cells with either Lucifer Yellow (LY) with a MW of 457 Da (LY-457) or LY with a MW of 649 Da (LY-649). Transfection of a dominant negative-connexin 43 (Cx43) gene decreased GJIC (>50%) when LY-649 was used, however, normal GJIC was observed in both cell lines when LY-457 was used. Therefore, the MW cut off in these clones was considerably less than the wild type. The dominant negative clones of the MC3T3-E1 cells exhibited over 90% less alkaline phosphatase (ALPase) activity and calcium deposition after the induction of differentiation. Similarly, dominant negative Cx43 inhibited gene expression of ALPase and bone sialoprotein but not osteocalcin in MC3T3-E1. WB-F344 cells normally exhibit a biphasic response to 12- O -tetradecanoylphorbol-13-acetate (TPA) where inhibition of GJIC recovers after 2 h, but the dominant negative clones showed no recovery from inhibition of GJIC by TPA. Dominant negative Cx43 also inhibited the formation of network-like structures by WB-F344 cells on Matrigel. These results demonstrate that the dominant negative gene transfected into cell types containing the wild-type connexins result in diminished channel sizes, thus allowing the determination of whether distinct biological endpoints, i.e., differentiation, are dependent upon either small or high MW intercellular signals. © 2003 Wiley-Liss, Inc. [source] Carthamus tinctorius flower extract prevents H2O2 -induced dysfunction and oxidative damage in osteoblastic MC3T3-E1 cellsPHYTOTHERAPY RESEARCH, Issue 7 2010Eun Mi Choi Abstract The flowers of Carthamus tinctorius L. (Compositae) have been widely used for enhancing blood circulation and postmenopausal disorder in women. In the present study, the potential protective effects of C. tinctorius flower extract (CFE) against reactive oxygen species (ROS) induced osteoblast dysfunction were investigated using osteoblastic MC3T3-E1 cells. The osteoblast function was assessed by measuring alkaline phosphatase activity, collagen content, calcium deposition, and RANKL production, and the oxidative status was assessed by measuring intracellular lipid peroxidation, and protein oxidation in osteoblastic MC3T3-E1 cells. A significant reduction in the alkaline phosphatase activity, collagen, and calcium deposition and an increase in the production of receptor activator of nuclear factor-kB ligand (RANKL) were observed after 0.3,mM H2O2 addition. The H2O2 -induced alterations were prevented by pre-incubating the osteoblasts with 2,10,,g/ml CFE for 48,h. When the oxidative stress was induced by H2O2, the increased production of protein carbonyl and malondialdehyde was also reduced at the same CFE concentration. These results demonstrate that C. tinctorius flower can act as a biological antioxidant in a cell culture experimental model and protect osteoblasts from oxidative stress-induced toxicity. Copyright © 2009 John Wiley & Sons, Ltd. [source] Stimulatory effect of undecylenic acid on mouse osteoblast differentiationPHYTOTHERAPY RESEARCH, Issue 4 2010Myung Hee Kim Abstract Natural compounds with bone-forming (or anabolic) activity have been recently focused on in bone research. The present study investigated the effect of undecylenic acid (UA) on osteoblast differentiation in mouse osteoblastic MC3T3-E1 subclone 4 cells and primary mouse calvarial cells. Low concentrations of UA (up to 5,,M) exhibited no cytotoxicity and significantly increased the expression and activity of alkaline phosphatase (early differentiation marker of osteoblast) and calcium deposition with the induction of expression of the osteocalcin gene in both cells. Interestingly, at low concentration of UA, the induction of NF-,B p65 translocation into nucleus and the up-regulation of AP-1 and NFATc1 transcript levels were also observed, suggesting that the stimulatory effect of UA on osteoblast differentiation could be mediated through the activation of transcription factors. Additionally, although the patterns of UA-induced activation of MAP kinases (JNK and p38) were not completely consistent with the increase of both ALP activity and calcium deposition by UA, MAP kinases might be partially involved in the biological function of UA during the early and late stages of osteoblast differentiation. Copyright © 2009 John Wiley & Sons, Ltd. [source] The effect of topical sodium thiosulfate in experimentally induced myringosclerosis,THE LARYNGOSCOPE, Issue 7 2010Yong Ho Park MD Abstract Objectives/Hypothesis: The purpose of this study was to investigate the effect of topical sodium thiosulfate (STS) in experimentally induced myringosclerosis (MS). Study Design: A prospective experimental animal study. Methods: Thirty Wistar albino rats were bilaterally myringotomized. The right ears were treated with STS or saline daily, and the left ears were left untreated and used as controls. The tympanic membranes were observed by otoendoscopy weekly, and tympanometric measurements were performed. All animals were histopathologically examined for myringosclerotic plaques. Results: Under otoendoscopy, myringosclerosis were observed around the handle of the malleus and near the annular region. The numbers of myringosclerotic ears were significantly more frequent in control and saline groups compared with the STS group (P < .05), and the formation of MS was more severe in control and saline groups compared with STS group (P < .05). Using tympanometric measurement, significantly reduced magnitudes of maximum admittance were observed in control and saline groups compared to normal and STS groups (P < .05). Under histopathologic examination, the tympanic membrane of the STS group appeared thinner than the control group (P < .05), with reduced calcium deposition than control and saline groups. Conclusions: Our results show that sodium thiosulfate has a preventive role in the development of myringosclerosis in the experimental animal model. Laryngoscope, 2010 [source] Matrix metalloproteinase 13 loss associated with impaired extracellular matrix remodeling disrupts chondrocyte differentiation by concerted effects on multiple regulatory factorsARTHRITIS & RHEUMATISM, Issue 8 2010Rosa Maria Borzí Objective To link matrix metalloproteinase 13 (MMP-13) activity and extracellular matrix (ECM) remodeling to alterations in regulatory factors leading to a disruption in chondrocyte homeostasis. Methods MMP-13 expression was ablated in primary human chondrocytes by stable retrotransduction of short hairpin RNA. The effects of MMP-13 knockdown on key regulators of chondrocyte differentiation (SOX9, runt-related transcription factor 2 [RUNX-2], and ,-catenin) and angiogenesis (vascular endothelial growth factor [VEGF]) were scored at the protein level (by immunohistochemical or Western blot analysis) and RNA level (by real-time polymerase chain reaction) in high-density monolayer and micromass cultures under mineralizing conditions. Effects on cellular viability in conjunction with chondrocyte progression toward a hypertrophic-like state were assessed in micromass cultures. Alterations in SOX9 subcellular distribution were assessed using confocal microscopy in micromass cultures and also in osteoarthritic cartilage. Results Differentiation of control chondrocyte micromasses progressed up to a terminal phase, with calcium deposition in conjunction with reduced cell viability and scant ECM. MMP-13 knockdown impaired ECM remodeling and suppressed differentiation in conjunction with reduced levels of RUNX-2, ,-catenin, and VEGF. MMP-13 levels in vitro and ECM remodeling in vitro and in vivo were linked to changes in SOX9 subcellular localization. SOX9 was largely excluded from the nuclei of chondrocytes with MMP-13,remodeled or ,degraded ECM, and exhibited an intranuclear staining pattern in chondrocytes with impaired MMP-13 activity in vitro or with more intact ECM in vivo. Conclusion MMP-13 loss leads to a breakdown in primary human articular chondrocyte differentiation by altering the expression of multiple regulatory factors. [source] Reduced chondrogenic and adipogenic activity of mesenchymal stem cells from patients with advanced osteoarthritisARTHRITIS & RHEUMATISM, Issue 3 2002J. Mary Murphy Objective Mesenchymal stem cells (MSCs) are resident in the bone marrow throughout normal adult life and have the capacity to differentiate along a number of connective tissue pathways, among them bone, cartilage, and fat. To determine whether functionally normal MSC populations may be isolated from patients with advanced osteoarthritis (OA), we have compared cells from patients undergoing joint replacement with cells from normal donors. Cell populations were compared in terms of yield, proliferation, and capacity to differentiate. Methods MSCs were prepared from bone marrow aspirates obtained from the iliac crest or from the tibia/femur during joint surgery. In vitro chondrogenic activity was measured as glycosaminoglycan and type II collagen deposition in pellet cultures. Adipogenic activity was measured as the accumulation of Nile Red O-positive lipid vacuoles, and osteogenic activity was measured as calcium deposition and by von Kossa staining. Results Patient-derived MSCs formed colonies in primary culture that were characteristically spindle-shaped with normal morphology. The primary cell yield in 36 of 38 cell cultures from OA donors fell within the range found in cultures from normal donors. However, the proliferative capacity of patient-derived MSCs was significantly reduced. There was a significant reduction in in vitro chondrogenic and adipogenic activity in cultures of patient-derived cells compared with that in normal cultures. There was no significant difference in in vitro osteogenic activity. There was no decline in chondrogenic potential with age in cells obtained from individuals with no evidence of OA. Conclusion These results raise the possibility that the increase in bone density and loss of cartilage that are characteristic of OA may result from changes in the differentiation profile of the progenitor cells that contribute to the homeostatic maintenance of these tissues. [source] | |||||||||||||