Histone H3 Lysine 36 Methyltransferase Whsc1 Promotes the Association of Runx2 and p300 in the Activation of Bone-Related Genes

PubMed Central

Lee, Yu Fei; Nimura, Keisuke; Lo, Wan Ning; Saga, Kotaro; Kaneda, Yasufumi

2014-01-01

The orchestration of histone modifiers is required to establish the epigenomic status that regulates gene expression during development. Whsc1 (Wolf-Hirschhorn Syndrome candidate 1), a histone H3 lysine 36 (H3K36) trimethyltransferase, is one of the major genes associated with Wolf-Hirshhorn syndrome, which is characterized by skeletal abnormalities. However, the role of Whsc1 in skeletal development remains unclear. Here, we show that Whsc1 regulates gene expression through Runt-related transcription factor (Runx) 2, a transcription factor central to bone development, and p300, a histone acetyltransferase, to promote bone differentiation. Whsc1 −/− embryos exhibited defects in ossification in the occipital bone and sternum. Whsc1 knockdown in pre-osteoblast cells perturbed histone modification patterns in bone-related genes and led to defects in bone differentiation. Whsc1 increased the association of p300 with Runx2, activating the bone-related genes Osteopontin (Opn) and Collagen type Ia (Col1a1), and Whsc1 suppressed the overactivation of these genes via H3K36 trimethylation. Our results suggest that Whsc1 fine-tunes the expression of bone-related genes by acting as a modulator in balancing H3K36 trimethylation and histone acetylation. Our results provide novel insight into the mechanisms by which this histone methyltransferase regulates gene expression. PMID:25188294

Molecular basis of differentiation therapy for soft tissue sarcomas

PubMed Central

Luther, Gaurav; Rames, Richard; Wagner, Eric R.; Zhu, Gaohui; Luo, Qing; Bi, Yang; Kim, Stephanie H.; Gao, Jian-Li; Huang, Enyi; Yang, Ke; Wang, Linyuan; Liu, Xing; Li, Mi; Hu, Ning; Su, Yuxi; Luo, Xiaoji; Chen, Liang; Luo, Jinyong; Haydon, Rex C.; Luu, Hue H.; Zhou, Lan; He, Tong-Chuan

2015-01-01

Stem cells are undifferentiated precursor cells with the capacity for proliferation or terminal differentiation. Progression down the differentiation cascade results in a loss of proliferative potential in exchange for the differentiated phenotype. This balance is tightly regulated in the physiologic state. Recent studies, however, have demonstrated that during tumorigenesis, disruptions preventing terminal differentiation allow cancer cells to maintain a proliferative, precursor cell phenotype. Current therapies (i.e., chemotherapy and radiation therapy) target the actively proliferating cells in tumor masses, which in many cases inevitably induce therapy-resistant cancer cells. It is conceivable that promising therapy regimens can be developed by treating human cancers by inducing terminal differentiation, thereby restoring the interrupted pathway and shifting the balance from proliferation to differentiation. For example, osteosarcoma (OS) is a primary bone cancer caused by differentiation defects in mesenchymal stem cells (MSCs) for which several differentiation therapies have shown great promise. In this review, we discuss the various differentiation therapies in the treatment of human sarcomas with a focus on OS. Such therapies hold great promise as they not only inhibit tumorigenesis, but also avoid the adverse effects associated with conventional chemotherapy regimens. Furthermore, it is conceivable that a combination of conventional therapies with differentiation therapy should significantly improve anticancer efficacy and reduce drug-resistance in the clinical management of human cancers, including sarcomas. PMID:26912947

Alpinia officinarum Stimulates Osteoblast Mineralization and Inhibits Osteoclast Differentiation.

PubMed

Shim, Ki-Shuk; Lee, Chung-Jo; Yim, Nam-Hui; Gu, Min Jung; Ma, Jin Yeul

2016-01-01

Alpinia officinarum rhizome has been used as a traditional herbal remedy to treat inflammatory and internal diseases. Based on the previously observed inhibitory effect of A. officinarum rhizome in an arthritis model, we evaluated whether a water extract of A. officinarum rhizome (WEAO) would enhance in vitro osteoblast mineralization using calvarial osteoblast precursor cells or would inhibit in vitro osteoclast differentiation and bone resorption using bone marrow derived macrophages. In osteoblasts, WEAO enhanced the mRNA levels of transcription factor (runt-related transcription factor 2, smad1, smad5, and junB) and marker (bone morphogenetic protein-2, collagen type 1alpha1, and osteocalcin) genes related to osteoblast mineralization, consistent with increased alizarin red S staining intensity. WEAO markedly inhibited osteoclast differentiation by suppressing the receptor activator for nuclear factor-[Formula: see text]B ligand-induced downregulation of inhibitor of DNA binding 2 and V-maf musculoaponeurotic fibrosarcoma oncogene homolog B and the phosphorylation of c-Jun N-terminal kinase, p38, nuclear factor-[Formula: see text]B, c-Src, and Bruton's tyrosine kinase to induce nuclear factor of activated T cells cytoplasmic 1 expression. WEAO also suppressed the resorbing activity of mature osteoclasts by altering actin ring formation. Therefore, the results of this study demonstrate that WEAO stimulates osteoblast mineralization and inhibits osteoclast differentiation. Thus, WEAO may be a promising herbal candidate to treat or prevent pathological bone diseases by regulating the balance between osteoclast and osteoblast activity.

Anabolic and Antiresorptive Modulation of Bone Homeostasis by the Epigenetic Modulator Sulforaphane, a Naturally Occurring Isothiocyanate*

PubMed Central

Thaler, Roman; Maurizi, Antonio; Roschger, Paul; Sturmlechner, Ines; Khani, Farzaneh; Spitzer, Silvia; Rumpler, Monika; Zwerina, Jochen; Karlic, Heidrun; Dudakovic, Amel; Klaushofer, Klaus; Teti, Anna; Rucci, Nadia; Varga, Franz; van Wijnen, Andre J.

2016-01-01

Bone degenerative pathologies like osteoporosis may be initiated by age-related shifts in anabolic and catabolic responses that control bone homeostasis. Here we show that sulforaphane (SFN), a naturally occurring isothiocyanate, promotes osteoblast differentiation by epigenetic mechanisms. SFN enhances active DNA demethylation via Tet1 and Tet2 and promotes preosteoblast differentiation by enhancing extracellular matrix mineralization and the expression of osteoblastic markers (Runx2, Col1a1, Bglap2, Sp7, Atf4, and Alpl). SFN decreases the expression of the osteoclast activator receptor activator of nuclear factor-κB ligand (RANKL) in osteocytes and mouse calvarial explants and preferentially induces apoptosis in preosteoclastic cells via up-regulation of the Tet1/Fas/Caspase 8 and Caspase 3/7 pathway. These mechanistic effects correlate with higher bone volume (∼20%) in both normal and ovariectomized mice treated with SFN for 5 weeks compared with untreated mice as determined by microcomputed tomography. This effect is due to a higher trabecular number in these mice. Importantly, no shifts in mineral density distribution are observed upon SFN treatment as measured by quantitative backscattered electron imaging. Our data indicate that the food-derived compound SFN epigenetically stimulates osteoblast activity and diminishes osteoclast bone resorption, shifting the balance of bone homeostasis and favoring bone acquisition and/or mitigation of bone resorption in vivo. Thus, SFN is a member of a new class of epigenetic compounds that could be considered for novel strategies to counteract osteoporosis. PMID:26757819

Angiocrine factors from Akt-activated endothelial cells balance self-renewal and differentiation of haematopoietic stem cells

PubMed Central

Kobayashi, Hideki; Butler, Jason M.; O'Donnell, Rebekah; Kobayashi, Mariko; Ding, Bi-Sen; Bonner, Bryant; Chiu, Vi K.; Nolan, Daniel J.; Shido, Koji; Benjamin, Laura; Rafii, Shahin

2010-01-01

Endothelial cells establish an instructive vascular niche that reconstitutes haematopoietic stem and progenitor cells (HSPCs) through release of specific paracrine growth factors, known as angiocrine factors. However, the mechanism by which endothelial cells balance the rate of proliferation and lineage-specific differentiation of HSPCs is unknown. Here, we demonstrate that Akt activation in endothelial cells, through recruitment of mTOR, but not the FoxO pathway, upregulates specific angiocrine factors that support expansion of CD34−Flt3− KLS HSPCs with long-term haematopoietic stem cell (LT-HSC) repopulation capacity. Conversely, co-activation of Akt-stimulated endothelial cells with p42/44 MAPK shifts the balance towards maintenance and differentiation of the HSPCs. Selective activation of Akt1 in the endothelial cells of adult mice increased the number of colony forming units in the spleen and CD34−Flt3− KLS HSPCs with LT-HSC activity in the bone marrow, accelerating haematopoietic recovery. Therefore, the activation state of endothelial cells modulates reconstitution of HSPCs through the upregulation of angiocrine factors, with Akt–mTOR-activated endothelial cells supporting the self-renewal of LT-HSCs and expansion of HSPCs, whereas MAPK co-activation favours maintenance and lineage-specific differentiation of HSPCs. PMID:20972423

A balance of FGF and BMP signals regulates cell cycle exit and Equarin expression in lens cells

PubMed Central

Jarrin, Miguel; Pandit, Tanushree; Gunhaga, Lena

2012-01-01

In embryonic and adult lenses, a balance of cell proliferation, cell cycle exit, and differentiation is necessary to maintain physical function. The molecular mechanisms regulating the transition of proliferating lens epithelial cells to differentiated primary lens fiber cells are poorly characterized. To investigate this question, we used gain- and loss-of-function analyses to modulate fibroblast growth factor (FGF) and/or bone morphogenetic protein (BMP) signals in chick lens/retina explants. Here we show that FGF activity plays a key role for proliferation independent of BMP signals. Moreover, a balance of FGF and BMP signals regulates cell cycle exit and the expression of Ccdc80 (also called Equarin), which is expressed at sites where differentiation of lens fiber cells occurs. BMP activity promotes cell cycle exit and induces Equarin expression in an FGF-dependent manner. In contrast, FGF activity is required but not sufficient to induce cell cycle exit or Equarin expression. Furthermore, our results show that in the absence of BMP activity, lens cells have increased cell cycle length or are arrested in the cell cycle, which leads to decreased cell cycle exit. Taken together, these findings suggest that proliferation, cell cycle exit, and early differentiation of primary lens fiber cells are regulated by counterbalancing BMP and FGF signals. PMID:22718906

The purinergic receptor P2X5 regulates inflammasome activity and hyper-multinucleation of murine osteoclasts

DOE PAGES

Kim, Hyunsoo; Walsh, Matthew C.; Takegahara, Noriko; ...

2017-03-15

Excessive bone resorption by osteoclasts (OCs) can result in serious clinical outcomes, including bone loss that may weaken skeletal or periodontal strength. Proper bone homeostasis and skeletal strength are maintained by balancing OC function with the bone-forming function of osteoblasts. Unfortunately, current treatments that broadly inhibit OC differentiation or function may also interfere with coupled bone formation. We therefore identified a factor, the purinergic receptor P2X5 that is highly expressed during the OC maturation phase, and which we show here plays no apparent role in early bone development and homeostasis, but which is required for osteoclast-mediated inflammatory bone loss andmore » hyper-multinucleation of OCs. We further demonstrate that P2X5 is required for ATP-mediated inflammasome activation and IL-1β production by OCs, and that P2X5-deficient OC maturation is rescued in vitro by addition of exogenous IL-1β. These findings identify a mechanism by which OCs react to inflammatory stimuli, and may identify purinergic signaling as a therapeutic target for bone loss related inflammatory conditions.« less

The purinergic receptor P2X5 regulates inflammasome activity and hyper-multinucleation of murine osteoclasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Hyunsoo; Walsh, Matthew C.; Takegahara, Noriko

Excessive bone resorption by osteoclasts (OCs) can result in serious clinical outcomes, including bone loss that may weaken skeletal or periodontal strength. Proper bone homeostasis and skeletal strength are maintained by balancing OC function with the bone-forming function of osteoblasts. Unfortunately, current treatments that broadly inhibit OC differentiation or function may also interfere with coupled bone formation. We therefore identified a factor, the purinergic receptor P2X5 that is highly expressed during the OC maturation phase, and which we show here plays no apparent role in early bone development and homeostasis, but which is required for osteoclast-mediated inflammatory bone loss andmore » hyper-multinucleation of OCs. We further demonstrate that P2X5 is required for ATP-mediated inflammasome activation and IL-1β production by OCs, and that P2X5-deficient OC maturation is rescued in vitro by addition of exogenous IL-1β. These findings identify a mechanism by which OCs react to inflammatory stimuli, and may identify purinergic signaling as a therapeutic target for bone loss related inflammatory conditions.« less

Simulating Bone Loss in Microgravity Using Mathematical Formulations of Bone Remodeling

NASA Technical Reports Server (NTRS)

Pennline, James A.

2009-01-01

Most mathematical models of bone remodeling are used to simulate a specific bone disease, by disrupting the steady state or balance in the normal remodeling process, and to simulate a therapeutic strategy. In this work, the ability of a mathematical model of bone remodeling to simulate bone loss as a function of time under the conditions of microgravity is investigated. The model is formed by combining a previously developed set of biochemical, cellular dynamics, and mechanical stimulus equations in the literature with two newly proposed equations; one governing the rate of change of the area of cortical bone tissue in a cross section of a cylindrical section of bone and one governing the rate of change of calcium in the bone fluid. The mechanical stimulus comes from a simple model of stress due to a compressive force on a cylindrical section of bone which can be reduced to zero to mimic the effects of skeletal unloading in microgravity. The complete set of equations formed is a system of first order ordinary differential equations. The results of selected simulations are displayed and discussed. Limitations and deficiencies of the model are also discussed as well as suggestions for further research.

LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance

PubMed Central

Lee, Sung-Uk; Maeda, Manami; Ishikawa, Yuichi; Li, Sierra Min; Wilson, Anne; Jubb, Adrian M.; Sakurai, Nagisa; Weng, Lihong; Fiorini, Emma; Radtke, Freddy; Yan, Minhong; MacDonald, H. Robson; Chen, Ching-Cheng

2013-01-01

Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in mouse embryos and are critical regulators of lymphoid lineage fate determination. However, it remains unclear how Notch regulates the balance between HSC self-renewal and differentiation in the adult bone marrow (BM). Here we report a novel mechanism that prevents HSCs from undergoing premature lymphoid differentiation in BM. Using a series of in vivo mouse models and functional HSC assays, we show that leukemia/lymphoma related factor (LRF) is necessary for HSC maintenance by functioning as an erythroid-specific repressor of Delta-like 4 (Dll4) expression. Lrf deletion in erythroblasts promoted up-regulation of Dll4 in erythroblasts, sensitizing HSCs to T-cell instructive signals in the BM. Our study reveals novel cross-talk between HSCs and erythroblasts, and sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and differentiation. PMID:23134786

Core binding factor beta (Cbfβ) controls the balance of chondrocyte proliferation and differentiation by upregulating Indian hedgehog (Ihh) expression and inhibiting parathyroid hormone-related protein receptor (PPR) expression in postnatal cartilage and bone formation.

PubMed

Tian, Fei; Wu, Mengrui; Deng, Lianfu; Zhu, Guochun; Ma, Junqing; Gao, Bo; Wang, Lin; Li, Yi-Ping; Chen, Wei

2014-07-01

Core binding factor beta (Cbfβ) is essential for embryonic bone morphogenesis. Yet the mechanisms by which Cbfβ regulates chondrocyte proliferation and differentiation as well as postnatal cartilage and bone formation remain unclear. Hence, using paired-related homeobox transcription factor 1-Cre (Prx1-Cre) mice, mesenchymal stem cell-specific Cbfβ-deficient (Cbfβ(f/f) Prx1-Cre) mice were generated to study the role of Cbfβ in postnatal cartilage and bone development. These mutant mice survived to adulthood but exhibited severe sternum and limb malformations. Sternum ossification was largely delayed in the Cbfβ(f/f) Prx1-Cre mice and the xiphoid process was noncalcified and enlarged. In newborn and 7-day-old Cbfβ(f/f) Prx1-Cre mice, the resting zone was dramatically elongated, the proliferation zone and hypertrophic zone of the growth plates were drastically shortened and disorganized, and trabecular bone formation was reduced. Moreover, in 1-month-old Cbfβ(f/f) Prx1-Cre mice, the growth plates were severely deformed and trabecular bone was almost absent. In addition, Cbfβ deficiency impaired intramembranous bone formation both in vivo and in vitro. Interestingly, although the expression of Indian hedgehog (Ihh) was largely reduced, the expression of parathyroid hormone-related protein (PTHrP) receptor (PPR) was dramatically increased in the Cbfβ(f/f) Prx1-Cre growth plate, indicating that that Cbfβ deficiency disrupted the Ihh-PTHrP negative regulatory loop. Chromatin immunoprecipitation (ChIP) analysis and promoter luciferase assay demonstrated that the Runx/Cbfβ complex binds putative Runx-binding sites of the Ihh promoter regions, and also the Runx/Cbfβ complex directly upregulates Ihh expression at the transcriptional level. Consistently, the expressions of Ihh target genes, including CyclinD1, Ptc, and Pthlh, were downregulated in Cbfβ-deficient chondrocytes. Taken together, our study reveals not only that Cbfβ is essential for chondrocyte proliferation and differentiation for the growth and maintenance of the skeleton in postnatal mice, but also that it functions in upregulating Ihh expression to promoter chondrocyte proliferation and osteoblast differentiation, and inhibiting PPR expression to enhance chondrocyte differentiation. © 2014 American Society for Bone and Mineral Research.

Substance P Promotes the Proliferation, but Inhibits Differentiation and Mineralization of Osteoblasts from Rats with Spinal Cord Injury via RANKL/OPG System.

PubMed

Liu, Hai-Juan; Yan, Hua; Yan, Jun; Li, Hao; Chen, Liang; Han, Li-Ren; Yang, Xiao-Fei

2016-01-01

Spinal cord injury (SCI) causes a significant amount of bone loss, which results in osteoporosis (OP). The neuropeptide substance P (SP) and SP receptors may play important roles in the pathogenesis of OP after SCI. To identify the roles of SP in the bone marrow mesenchymal stem cell derived osteoblasts (BMSC-OB) in SCI rats, we investigated the expression of neurokinin-1 receptors (NK1R) in BMSC-OB and the effects of SP on bone formation by development of BMSC-OB cultures. Sixty young male Sprague-Dawley rats were randomized into two groups: SHAM and SCI. The expression of NK1R protein in BMSC-OB was observed using immunohistochemistry and Western blot analysis. The dose- and time-dependent effects of SP on the proliferation, differentiation and mineralization of BMSC-OB and the expression of osteoblastic markers by in vitro experiments. The expression of NK1R in BMSC-OB was observed on plasma membranes and in cytoplasm. One week after osteogenic differentiation, the expression of NK1R was significantly increased after SCI at mRNA and protein levels. However, this difference was gradually attenuated at 2 or 3 weeks later. SP have the function to enhance cell proliferation, inhibite cell differentiation and mineralization at a proper concentration and incubation time, and this effect would be inhibited by adding SP or NK1R antagonist. The expression of RANKL/OPG was significantly increased in tibiae after SCI. Similarly, the RANKL/OPG expression in SCI rats was significantly increased when treating with 10-8 M SP. SP plays a very important role in the pathogenesis of OP after SCI. The direct effect of SP may lead to increased bone resorption through the RANKL/OPG axis after SCI. In addition, high expression of SP also results in the suppression of osteogenesis in SCI rats. Then, the balance between bone resorption and bone formation was broken and finally osteoporosis occurred.

Bone Balance within a Cortical BMU: Local Controls of Bone Resorption and Formation

PubMed Central

Smith, David W.; Gardiner, Bruce S.; Dunstan, Colin

2012-01-01

Maintaining bone volume during bone turnover by a BMU is known as bone balance. Balance is required to maintain structural integrity of the bone and is often dysregulated in disease. Consequently, understanding how a BMU controls bone balance is of considerable interest. This paper develops a methodology for identifying potential balance controls within a single cortical BMU. The theoretical framework developed offers the possibility of a directed search for biological processes compatible with the constraints of balance control. We first derive general control constraint equations and then introduce constitutive equations to identify potential control processes that link key variables that describe the state of the BMU. The paper describes specific local bone volume balance controls that may be associated with bone resorption and bone formation. Because bone resorption and formation both involve averaging over time, short-term fluctuations in the environment are removed, leaving the control systems to manage deviations in longer-term trends back towards their desired values. The length of time for averaging is much greater for bone formation than for bone resorption, which enables more filtering of variability in the bone formation environment. Remarkably, the duration for averaging of bone formation may also grow to control deviations in long-term trends of bone formation. Providing there is sufficient bone formation capacity by osteoblasts, this leads to an extraordinarily robust control mechanism that is independent of either osteoblast number or the cellular osteoid formation rate. A complex picture begins to emerge for the control of bone volume. Different control relationships may achieve the same objective, and the ‘integration of information’ occurring within a BMU may be interpreted as different sets of BMU control systems coming to the fore as different information is supplied to the BMU, which in turn leads to different observable BMU behaviors. PMID:22844401

A new role for HERPUD1 and ERAD activation in osteoblast differentiation and mineralization.

PubMed

Américo-Da-Silva, Luan; Diaz, Jheimmy; Bustamante, Mario; Mancilla, Georthan; Oyarzún, Ingrid; Verdejo, Hugo E; Quiroga, Clara

2018-03-23

Bone integrity depends on a finely tuned balance between bone synthesis by osteoblasts and resorption by osteoclasts. The secretion capacity of mature osteoblasts requires strict control of proteostasis. Endoplasmic reticulum-associated degradation (ERAD) prevents the accumulation of unfolded ER proteins via dislocation to the cytosol and degradation by the proteasome. The ER membrane protein, homocysteine-inducible endoplasmic reticulum protein with ubiquitin-like domain 1 (HERPUD1), is a key component of the ERAD multiprotein complex which helps to stabilize the complex and facilitate the efficient degradation of unfolded proteins. HERPUD1 expression is strongly up-regulated by the unfolded protein response and cellular stress. The aim of the current study was to establish whether HERPUD1 and ERAD play roles in osteoblast differentiation and maturation. We evaluated preosteoblastic MC3T3-E1 cell and primary rat osteoblast differentiation by measuring calcium deposit levels, alkaline phosphatase activity, and runt-related transcription factor 2 and osterix expression. We found that ERAD and proteasomal degradation were activated and that HERPUD1 expression was increased as osteoblast differentiation progressed. The absence of HERPUD1 blocked osteoblast mineralization in vitro and significantly reduced alkaline phosphatase activity. In contrast, HERPUD1 overexpression activated the osteoblast differentiation program. Our results demonstrate that HERPUD1 and ERAD are important for the activation of the osteoblast maturation program and may be useful new targets for elucidating bone physiology.-Américo-Da-Silva, L., Diaz, J., Bustamante, M., Mancilla, G., Oyarzún, I., Verdejo, H. E., Quiroga, C. A new role for HERPUD1 and ERAD activation in osteoblast differentiation and mineralization.

[Regulating effect of pineal gland peptides on development of T-lymphocytes in CBA aging mice: role of microenvironment of immune system organs and neuroendocrine factors].

PubMed

Labunets, I F; Butenko, G M; Khavinson, V Kh; Magdich, L V; Dragunova, V A; Pishel', I N; Azarskova, M V

2003-01-01

Studies were undertaken on the development of T-lymphocytes in adult and old CBA mice and its changes at aging after injections of pineal gland peptides. It was shown that in old mice the disturbances of T-cells differentiation are registered in bone marrow, thymus, spleen and characterized by the changes of lymphocyte markers expression, migration and proliferation of cells. In old mice FTS titer, melatonin and testosterone levels decreased, the balance of noradrenalin and serotonin in hypothalamus and the cell composition of microenvironment immune systems organs impaired. After chronic (18 mo) administration of the pineal gland preparation epithalamin the amount of stromal cells-precursors, CD4+ and Mac-1(+)-cells in old bone marrow increased, improved the migration of T-cell precursors from bone marrow to thymus and their proliferative potential. The proportion of CD3+, CD4+CD8-, CD4-CD8+, Mac-1(+)-cells in old thymus increased, while that of CD44(+)-cells decreased. The proportion of CD4-CD8(+)-cells in spleen increased. The most number of indices and their balance showed a pattern of adult mice. In old mice after epithalamin the balance of amines in hypothalamus improved, concentration of melatonin in pineal gland, testosterone and FTS titer in blood increased. Epithalon has also the possibility to increase of thymic endocrine function.

Variations in Urine Calcium Isotope: Composition Reflect Changes in Bone Mineral Balance in Humans

NASA Technical Reports Server (NTRS)

Skulan, Joseph; Anbar, Ariel; Bullen, Thomas; Puzas, J. Edward; Shackelford, Linda; Smith, Scott M.

2004-01-01

Changes in bone mineral balance cause rapid and systematic changes in the calcium isotope composition of human urine. Urine from subjects in a 17 week bed rest study was analyzed for calcium isotopic composition. Comparison of isotopic data with measurements of bone mineral density and metabolic markers of bone metabolism indicates the calcium isotope composition of urine reflects changes in bone mineral balance. Urine calcium isotope composition probably is affected by both bone metabolism and renal processes. Calcium isotope. analysis of urine and other tissues may provide information on bone mineral balance that is in important respects better than that available from other techniques, and illustrates the usefulness of applying geochemical techniques to biomedical problems.

Melatonin attenuates titanium particle-induced osteolysis via activation of Wnt/β-catenin signaling pathway.

PubMed

Ping, Zichuan; Hu, Xuanyang; Wang, Liangliang; Shi, Jiawei; Tao, Yunxia; Wu, Xiexing; Hou, Zhenyang; Guo, Xiaobin; Zhang, Wen; Yang, Huilin; Xu, Yaozeng; Wang, Zhirong; Geng, Dechun

2017-03-15

Wear debris-induced inhibition of bone regeneration and extensive bone resorption were common features in peri-prosthetic osteolysis (PPO). Here, we investigated the effect of melatonin on titanium particle-stimulated osteolysis in a murine calvariae model and mouse-mesenchymal-stem cells (mMSCs) culture system. Melatonin inhibited titanium particle-induced osteolysis and increased bone formation at osteolytic sites, confirmed by radiological and histomorphometric data. Furthermore, osteoclast numbers decreased dramatically in the low- and high-melatonin administration mice, as respectively, compared with the untreated animals. Melatonin alleviated titanium particle-induced depression of osteoblastic differentiation and mineralization in mMSCs. Mechanistically, melatonin was found to reduce the degradation of β-catenin, levels of which were decreased in presence of titanium particles both in vivo and in vitro. To further ensure whether the protective effect of melatonin was mediated by the Wnt/β-catenin signaling pathway, ICG-001, a selective β-catenin inhibitor, was added to the melatonin-treated groups and was found to attenuate the effect of melatonin on mMSC mineralization. We also demonstrated that melatonin modulated the balance between receptor activator of nuclear factor kappa-B ligand and osteoprotegerin via activation of Wnt/β-catenin signaling pathway. These findings strongly suggest that melatonin represents a promising candidate in the treatment of PPO. Peri-prosthetic osteolysis, initiated by wear debris-induced inhibition of bone regeneration and extensive bone resorption, is the leading cause for implant failure and reason for revision surgery. In the current study, we demonstrated for the first time that melatonin can induce bone regeneration and reduce bone resorption at osteolytic sites caused by titanium-particle stimulation. These effects might be mediated by activating Wnt/β-catenin signaling pathway and enhancing osteogenic differentiation. Meanwhile, the ability of melatonin to modulate the balance between receptor activator of nuclear factor kappa-B ligand and osteoprotegerin mediated by Wnt/β-catenin signaling pathway, thereby suppressing osteoclastogenesis, may be implicated in the protective effects of melatonin on titanium-particle-induced bone resorption. These results suggested that melatonin can be considered as a promising therapeutic agent for the prevention and treatment of peri-prosthetic osteolysis. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Rap1b Is an Effector of Axin2 Regulating Crosstalk of Signaling Pathways During Skeletal Development.

PubMed

Maruyama, Takamitsu; Jiang, Ming; Abbott, Alycia; Yu, H-M Ivy; Huang, Qirong; Chrzanowska-Wodnicka, Magdalena; Chen, Emily I; Hsu, Wei

2017-09-01

Recent identification and isolation of suture stem cells capable of long-term self-renewal, clonal expanding, and differentiating demonstrate their essential role in calvarial bone development, homeostasis, and injury repair. These bona fide stem cells express a high level of Axin2 and are able to mediate bone regeneration and repair in a cell autonomous fashion. The importance of Axin2 is further demonstrated by its genetic inactivation in mice causing skeletal deformities resembling craniosynostosis in humans. The fate determination and subsequent differentiation of Axin2+ stem cells are highly orchestrated by a variety of evolutionary conserved signaling pathways including Wnt, FGF, and BMP. These signals are often antagonistic of each other and possess differential effects on osteogenic and chondrogenic cell types. However, the mechanisms underlying the interplay of these signaling transductions remain largely elusive. Here we identify Rap1b acting downstream of Axin2 as a signaling interrogator for FGF and BMP. Genetic analysis reveals that Rap1b is essential for development of craniofacial and body skeletons. Axin2 regulates Rap1b through modulation of canonical BMP signaling. The BMP-mediated activation of Rap1b promotes chondrogenic fate and chondrogenesis. Furthermore, by inhibiting MAPK signaling, Rap1b mediates the antagonizing effect of BMP on FGF to repress osteoblast differentiation. Disruption of Rap1b in mice not only enhances osteoblast differentiation but also impairs chondrocyte differentiation during intramembranous and endochondral ossifications, respectively, leading to severe defects in craniofacial and body skeletons. Our findings reveal a dual role of Rap1b in development of the skeletogenic cell types. Rap1b is critical for balancing the signaling effects of BMP and FGF during skeletal development and disease. © 2017 American Society for Bone and Mineral Research. © 2017 American Society for Bone and Mineral Research.

Postnatal ablation of osteoblast Smad4 enhances proliferative responses to canonical Wnt signaling through interactions with β-catenin

PubMed Central

Salazar, Valerie S.; Zarkadis, Nicholas; Huang, Lisa; Watkins, Marcus; Kading, Jacqueline; Bonar, Sheri; Norris, Jin; Mbalaviele, Gabriel; Civitelli, Roberto

2013-01-01

Summary Canonical Wnt (cWnt) signaling through β-catenin regulates osteoblast proliferation and differentiation to enhance bone formation. We previously reported that osteogenic action of β-catenin is dependent on BMP signaling. Here, we further examined interactions between cWnt and BMP in bone. In osteoprogenitors stimulated with BMP2, β-catenin localizes to the nucleus, physically interacts with Smad4, and is recruited to DNA-binding transcription complexes containing Smad4, R-Smad1/5 and TCF4. Furthermore, Tcf/Lef-dependent transcription, Ccnd1 expression and proliferation all increase when Smad4, 1 or 5 levels are low, whereas TCF/Lef activities decrease when Smad4 expression is high. The ability of Smad4 to antagonize transcription of Ccnd1 is dependent on DNA-binding activity but Smad4-dependent transcription is not required. In mice, conditional deletion of Smad4 in osterix+ cells increases mitosis of cells on trabecular bone surfaces as well as in primary osteoblast cultures from adult bone marrow and neonatal calvaria. By contrast, ablation of Smad4 delays differentiation and matrix mineralization by primary osteoblasts in response to Wnt3a, indicating that loss of Smad4 perturbs the balance between proliferation and differentiation in osteoprogenitors. We propose that Smad4 and Tcf/Lef transcription complexes compete for β-catenin, thus restraining cWnt-dependent proliferative signals while favoring the matrix synthesizing activity of osteoblasts. PMID:24101723

Osteoporosis and body composition.

PubMed

Crepaldi, G; Romanato, G; Tonin, P; Maggi, S

2007-01-01

The Epidemiologic Study on the Prevalence of Osteoporosis in Italy showed that the prevalence of osteoporosis among women and men aged 60 yr and over is 22.8% and 14.5%, respectively, giving rise to about 80,000 new fractures a yr. Sarcopenia is considered to be one of the main features of the aging process. It is characterized by a reduction in muscle mass and muscle strength, and affects women more than men. It is associated with a increased risk of fractures consequent upon a greater predisposition to falls, but also to the lack of bone remodeling due to reduced muscle mechanical strength. Muscle strength determines quality bone modifications such as density, strength, and microarchitecture. Variations in the ratios of cortical and muscle areas give rise to various types of osteoporosis, with different risks of fracture. Bone mineral density increases with body fat mass, and obesity has a protective effect against osteoporosis. This protective effect is explained by a combination of hormonal (peripheral aromatization of androgens to estrogens in adipose tissue) and mechanical factors (on weight-bearing bone sites), but the hormone leptin also probably mediates fat and bone mass. Serum leptin levels are closely related to body fat mass, and some findings suggest the peripheral effect of leptin, which exerts estrogenic effects, enhancing osteoblastic differentiation and inhibiting late adipocytic differentiation. The overall effect of leptin on bone results from a balance between negative central effects and positive direct peripheral effects, according to serum leptin levels.

BMP-driven NRF2 activation in esophageal basal cell differentiation and eosinophilic esophagitis

PubMed Central

Jiang, Ming; Ku, Wei-Yao; Zhou, Zhongren; Dellon, Evan S.; Falk, Gary W.; Nakagawa, Hiroshi; Wang, Mei-Lun; Liu, Kuancan; Wang, Jun; Katzka, David A.; Peters, Jeffrey H.; Lan, Xiaopeng; Que, Jianwen

2015-01-01

Tissue homeostasis requires balanced self-renewal and differentiation of stem/progenitor cells, especially in tissues that are constantly replenished like the esophagus. Disruption of this balance is associated with pathological conditions, including eosinophilic esophagitis (EoE), in which basal progenitor cells become hyperplastic upon proinflammatory stimulation. However, how basal cells respond to the inflammatory environment at the molecular level remains undetermined. We previously reported that the bone morphogenetic protein (BMP) signaling pathway is critical for epithelial morphogenesis in the embryonic esophagus. Here, we address how this pathway regulates tissue homeostasis and EoE development in the adult esophagus. BMP signaling was specifically activated in differentiated squamous epithelium, but not in basal progenitor cells, which express the BMP antagonist follistatin. Previous reports indicate that increased BMP activity promotes Barrett’s intestinal differentiation; however, in mice, basal progenitor cell–specific expression of constitutively active BMP promoted squamous differentiation. Moreover, BMP activation increased intracellular ROS levels, initiating an NRF2-mediated oxidative response during basal progenitor cell differentiation. In both a mouse EoE model and human biopsies, reduced squamous differentiation was associated with high levels of follistatin and disrupted BMP/NRF2 pathways. We therefore propose a model in which normal squamous differentiation of basal progenitor cells is mediated by BMP-driven NRF2 activation and basal cell hyperplasia is promoted by disruption of BMP signaling in EoE. PMID:25774506

Confined Sandwichlike Microenvironments Tune Myogenic Differentiation.

PubMed

Ballester-Beltrán, José; Trujillo, Sara; Alakpa, Enateri V; Compañ, Vicente; Gavara, Rafael; Meek, Dominic; West, Christopher C; Péault, Bruno; Dalby, Matthew J; Salmerón-Sánchez, Manuel

2017-08-14

Sandwichlike (SW) cultures are engineered as a multilayer technology to simultaneously stimulate dorsal and ventral cell receptors, seeking to mimic cell adhesion in three-dimensional (3D) environments in a reductionist manner. The effect of this environment on cell differentiation was investigated for several cell types cultured in standard growth media, which promotes proliferation on two-dimensional (2D) surfaces and avoids any preferential differentiation. First, murine C2C12 myoblasts showed specific myogenic differentiation. Human mesenchymal stem cells (hMSCs) of adipose and bone marrow origin, which can differentiate toward a wider variety of lineages, showed again myodifferentiation. Overall, this study shows myogenic differentiation in normal growth media for several cell types under SW conditions, avoiding the use of growth factors and cytokines, i.e., solely by culturing cells within the SW environment. Mechanistically, it provides further insights into the balance between integrin adhesion to the dorsal substrate and the confinement imposed by the SW system.

Confined Sandwichlike Microenvironments Tune Myogenic Differentiation

PubMed Central

2017-01-01

Sandwichlike (SW) cultures are engineered as a multilayer technology to simultaneously stimulate dorsal and ventral cell receptors, seeking to mimic cell adhesion in three-dimensional (3D) environments in a reductionist manner. The effect of this environment on cell differentiation was investigated for several cell types cultured in standard growth media, which promotes proliferation on two-dimensional (2D) surfaces and avoids any preferential differentiation. First, murine C2C12 myoblasts showed specific myogenic differentiation. Human mesenchymal stem cells (hMSCs) of adipose and bone marrow origin, which can differentiate toward a wider variety of lineages, showed again myodifferentiation. Overall, this study shows myogenic differentiation in normal growth media for several cell types under SW conditions, avoiding the use of growth factors and cytokines, i.e., solely by culturing cells within the SW environment. Mechanistically, it provides further insights into the balance between integrin adhesion to the dorsal substrate and the confinement imposed by the SW system. PMID:28824958

Estrogen regulates the rate of bone turnover but bone balance in ovariectomized rats is modulated by prevailing mechanical strain

NASA Technical Reports Server (NTRS)

Westerlind, K. C.; Wronski, T. J.; Ritman, E. L.; Luo, Z. P.; An, K. N.; Bell, N. H.; Turner, R. T.

1997-01-01

Estrogen deficiency induced bone loss is associated with increased bone turnover in rats and humans. The respective roles of increased bone turnover and altered balance between bone formation and bone resorption in mediating estrogen deficiency-induced cancellous bone loss was investigated in ovariectomized rats. Ovariectomy resulted in increased bone turnover in the distal femur. However, cancellous bone was preferentially lost in the metaphysis, a site that normally experiences low strain energy. No bone loss was observed in the epiphysis, a site experiencing higher strain energy. The role of mechanical strain in maintaining bone balance was investigated by altering the strain history. Mechanical strain was increased and decreased in long bones of ovariectomized rats by treadmill exercise and functional unloading, respectively. Functional unloading was achieved during orbital spaceflight and following unilateral sciatic neurotomy. Increasing mechanical loading reduced bone loss in the metaphysis. In contrast, decreasing loading accentuated bone loss in the metaphysis and resulted in bone loss in the epiphysis. Finally, administration of estrogen to ovariectomized rats reduced bone loss in the unloaded and prevented loss in the loaded limb following unilateral sciatic neurotomy in part by reducing indices of bone turnover. These results suggest that estrogen regulates the rate of bone turnover, but the overall balance between bone formation and bone resorption is influenced by prevailing levels of mechanical strain.

BMP signaling balances proliferation and differentiation of muscle satellite cell descendants

PubMed Central

2011-01-01

Background The capacity of muscle to grow or to regenerate after damage is provided by adult stem cells, so called satellite cells, which are located under the basement lamina of each myofiber. Upon activation satellite cells enter the cell cycle, proliferate and differentiate into myoblasts, which fuse to injured myofibers or form new fibers. These processes are tightly controlled by many growth factors. Results Here we investigate the role of bone morphogenetic proteins (BMPs) during satellite cell differentiation. Unlike the myogenic C2C12 cell line, primary satellite cells do not differentiate into osteoblasts upon BMP signaling. Instead BMP signaling inhibits myogenic differentiation of primary satellite cells ex vivo. In contrast, inhibition of BMP signaling results in cell cycle exit, followed by enhanced myoblast differentiation and myotube formation. Using an in vivo trauma model we demonstrate that satellite cells respond to BMP signals during the regeneration process. Interestingly, we found the BMP inhibitor Chordin upregulated in primary satellite cell cultures and in regenerating muscles. In both systems Chordin expression follows that of Myogenin, a marker for cells committed to differentiation. Conclusion Our data indicate that BMP signaling plays a critical role in balancing proliferation and differentiation of activated satellite cells and their descendants. Initially, BMP signals maintain satellite cells descendants in a proliferating state thereby expanding cell numbers. After cells are committed to differentiate they upregulate the expression of the BMP inhibitor Chordin thereby supporting terminal differentiation and myotube formation in a negative feedback mechanism. PMID:21645366

Plasticity and Maintenance of Hematopoietic Stem Cells During Development

PubMed Central

Kanji, Suman; Pompili, Vincent J.; Das, Hiranmoy

2012-01-01

Maintenance of hematopoietic stem cells (HSCs) pool depends on fine balance between self-renewal and differentiation of HSCs. HSCs normally reside within the bone marrow niche of an adult mammal. The embryonic development of HSCs is a complex process that involves the migration of developing HSCs in multiple anatomical sites. Throughout the process, developing HSCs receive internal (transcriptional program) and external (HSC niche) signals, which direct them to maintain balance between self-renewal and differentiation, also to generate a pool of HSCs. In physiological condition HSCs differentiate into all mature cell types present in the blood. However, in pathological condition they may differentiate into non-hematological cells according to the need of the body. It was shown that HSCs can transdifferentiate into cell types that do not belong to the hematopoietic system suggests a complete paradigm shift of the hierarchical hematopoietic tree. This review describes the developmental origins and regulation of HSCs focusing on developmental signals that induce the adult hematopoietic stem cell program, as these informations are very critical for manipulating conditions for expansion of HSCs in ex vivo condition. This review also states clinical application and related patents using HSC. PMID:21517745

Transgenic medaka fish as models to analyze bone homeostasis under micro-gravity conditions in vivo

NASA Astrophysics Data System (ADS)

Winkler, C.; Wagner, T.; Renn, J.; Goerlich, R.; Schartl, M.

Long-term space flight and microgravity results in bone loss that can be explained by reduced activity of bone-forming osteoblast cells and/or an increase in activity of bone resorbing osteoclast cells. Osteoprotegerin (OPG), a secreted protein of 401 amino acids, has been shown to regulate the balance between osteoblast and osteoclast formation and thereby warrants constant bone mass under normal gravitational conditions. Consistent with this, earlier reports using transgenic mice have shown that increased activation of OPG leads to exc essive bone formation (osteopetrosis), while inactivation of OPG leads to bone loss (osteoporosis). Importantly, it has recently been reported that expression of murine OPG is regulated by vector averaged gravity (Kanematsu et al., 2002, Bone 30, p553). The small bony fish medaka (Oryzias latipes ) has attracted increasing attention as genetic model system to study developmental and pathological processes. To analyze the molecular mechanisms of bone formation in this small vertebrate, we have isolated two related genes, opr-1 and opr -2, from medaka. Our phylogenetic analysis revealed that both genes originated from a common ancestor by fish-specific gene duplication and represent the orthologs of the mammalian OPG gene. Both opr genes are differentially expressed during embryonic and larval development, in adult tissues and in cultured primary osteoblast cells. We have characterized their promoter regions and identified consensus binding sites for transcription factors of the bone-morphogenetic-protein (BMP) p thway and for core-binding-factor-1Aa (cbfa1). Cbfa1 has been shown to be the key regulator of OPG expression during several steps of osteoblast differentiation in mammals. This opens the possibility that the mechanisms controlling bone formation in teleost fish and higher vertebrates are regulated by related mechanisms. We are currently generating transgenic medakafish expressing a GFP reporter gene under control of the teleost OPG promoter in order to visualize osteoblast activity in a living organism under different gravity conditions. This work is supported by the German Aerospace Center, DLR.

Time-sequential changes of differentially expressed miRNAs during the process of anterior lumbar interbody fusion using equine bone protein extract, rhBMP-2 and autograft

NASA Astrophysics Data System (ADS)

Chen, Da-Fu; Zhou, Zhi-Yu; Dai, Xue-Jun; Gao, Man-Man; Huang, Bao-Ding; Liang, Tang-Zhao; Shi, Rui; Zou, Li-Jin; Li, Hai-Sheng; Bünger, Cody; Tian, Wei; Zou, Xue-Nong

2014-03-01

The precise mechanism of bone regeneration in different bone graft substitutes has been well studied in recent researches. However, miRNAs regulation of the bone formation has been always mysterious. We developed the anterior lumbar interbody fusion (ALIF) model in pigs using equine bone protein extract (BPE), recombinant human bone morphogenetic protein-2 (rhBMP-2) on an absorbable collagen sponge (ACS), and autograft as bone graft substitute, respectively. The miRNA and gene expression profiles of different bone graft materials were examined using microarray technology and data analysis, including self-organizing maps, KEGG pathway and Biological process GO analyses. We then jointly analyzed miRNA and mRNA profiles of the bone fusion tissue at different time points respectively. Results showed that miRNAs, including let-7, miR-129, miR-21, miR-133, miR-140, miR-146, miR-184, and miR-224, were involved in the regulation of the immune and inflammation response, which provided suitable inflammatory microenvironment for bone formation. At late stage, several miRNAs directly regulate SMAD4, Estrogen receptor 1 and 5-hydroxytryptamine (serotonin) receptor 2C for bone formation. It can be concluded that miRNAs play important roles in balancing the inflammation and bone formation.

Contribution of Vestibular-Evoked Myogenic Potential (VEMP) testing in the assessment and the differential diagnosis of otosclerosis.

PubMed

Tramontani, Ourania; Gkoritsa, Eleni; Ferekidis, Eleftherios; Korres, Stavros G

2014-02-07

The aim of this prospective clinical study was to evaluate the clinical importance of Vestibular-Evoked Myogenic Potentials (VEMPs) in the assessment and differential diagnosis of otosclerosis and otologic diseases characterized by "pseudo-conductive" components. We also investigated the clinical appearance of balance disorders in patients with otosclerosis by correlating VEMP results with the findings of caloric testing and pure tone audiometry(PTA). Air-conducted(AC) 4-PTA, bone-conducted(BC) 4-PTA, air-bone Gap(ABG), AC, BC tone burst evoked VEMP, and calorics were measured preoperatively in 126 otosclerotic ears. The response rate of the AC-VEMPs and BC-VEMPs was 29.36% and 44.03%, respectively. Statistical differences were found between the means of ABG, AC 4-PTA, and BC 4-PTA in the otosclerotic ears in relation to AC-VEMP elicitability. About one-third of patients presented with disequilibrium. A statistically significant interaction was found between calorics and dizziness in relation to PTA thresholds. No relationship was found between calorics and dizziness with VEMPs responses. AC and BC VEMPs can be elicited in ears with otosclerosis. AC-VEMP is more vulnerable to conductive hearing loss. Evaluation of AC-VEMP thresholds can be added in the diagnostic work-up of otosclerosis in case of doubt, enhancing differential diagnosis in patients with air-bone gaps. Otosclerosis is not a cause of canal paresis or vertigo.

Blockade of CD26 signaling inhibits human osteoclast development.

PubMed

Nishida, Hiroko; Suzuki, Hiroshi; Madokoro, Hiroko; Hayashi, Mutsumi; Morimoto, Chikao; Sakamoto, Michiie; Yamada, Taketo

2014-11-01

Bone remodeling is maintained by the delicate balance between osteoblasts (OBs) and osteoclasts (OCs). However, the role of CD26 in regulating bone remodeling has not yet been characterized. We herein show that CD26 is preferentially expressed on normal human OCs and is intensely expressed on activated human OCs in osteolytic bone alterations. Macrophage-colony stimulating factor (M-CSF) and soluble receptor activator of NF-κB ligand (sRANKL) induced human OC differentiation, in association with CD26 expression on monocyte-macrophage lineage cells. CD26 expression was accompanied by increased phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), which is crucial for early human OC differentiation. The humanized anti-CD26 monoclonal antibody, huCD26mAb, impaired the formation and function of tartrate-resistant acid phosphatase (TRAP)/CD26 positive multi-nucleated (nuclei > 3) OCs with maturation in the manner of dose-dependency. It was revealed that huCD26mAb inhibits early OC differentiation via the inactivation of MKK3/6, p38 MAPK and subsequent dephosphorylation of microphthalmia-associated transcription factor (mi/Mitf). These inhibitions occur immediately after RANKL binds to RANK on the human OC precursor cells and were demonstrated using the OC functional assays. huCD26mAb subsequently impaired OC maturation and bone resorption by suppressing the expression of TRAP and OC fusion proteins. In addition, p38 MAPK inhibitor also strongly inhibited OC formation and function. Our results suggest that the blockade of CD26 signaling impairs the development of human functional OCs by inhibiting p38 MAPK-mi/Mitf phosphorylation pathway and that targeting human OCs with huCD26mAb may have therapeutic potential for the treatment of osteolytic lesions following metastasis to alleviate bone destruction and reduce total skeletal-related events (SREs). © 2014 American Society for Bone and Mineral Research.

SCAPs Regulate Differentiation of DFSCs During Tooth Root Development in Swine

PubMed Central

Wu, Xiaoshan; Hu, Lei; Li, Yan; Li, Yang; Wang, Fu; Ma, Ping; Wang, Jinsong; Zhang, Chunmei; Jiang, Canhua; Wang, Songlin

2018-01-01

The tooth root transmits and balances occlusal forces through the periodontium to the alveolar bone. The periodontium, including the gingiva, the periodontal ligament, the cementum and the partial alveolar bone, derives from the dental follicle (DF), except for the gingiva. In the early developmental stages, the DF surrounds the tooth germ as a sphere and functions to promote tooth eruption. However, the morphological dynamics and factors regulating the differentiation of the DF during root elongation remain largely unknown. Miniature pigs are regarded as a useful experimental animal for modeling in craniofacial research because they are similar to humans with respect to dentition and mandible anatomy. In the present study, we used the third deciduous incisor of miniature pig as the model to investigate the factors influencing DF differentiation during root development. We found that the DF was shaped like a crescent and was located between the root apical and the alveolar bone. The expression levels of WNT5a, β-Catenin, and COL-I gradually increased from the center of the DF (beneath the apical foramen) to the lateral coronal corner, where the DF differentiates into the periodontium. To determine the potential regulatory role of the apical papilla on DF cell differentiation, we co-cultured dental follicle stem cells (DFSCs) with stem cells of the apical papilla (SCAPs). The osteogenesis and fibrogenesis abilities of DFSCs were inhibited when being co-cultured with SCAPs, suggesting that the fate of the DF can be regulated by signals from the apical papilla. The apical papilla may sustain the undifferentiated status of DFSCs before root development finishes. These data yield insight into the interaction between the root apex and surrounding DF tissues in root and periodontium development and shed light on the future study of root regeneration in large mammals. PMID:29511365

Bone loss from Wnt inhibition mitigated by concurrent alendronate therapy.

PubMed

Madan, Babita; McDonald, Mitchell J; Foxa, Gabrielle E; Diegel, Cassandra R; Williams, Bart O; Virshup, David M

2018-01-01

Dysregulated Wnt signaling is associated with the pathogenesis of cancers, fibrosis, and vascular diseases. Inhibition of Wnt signaling has shown efficacy in various pre-clinical models of these disorders. One of the key challenges in developing targeted anti-cancer drugs is to balance efficacy with on-target toxicity. Given the crucial role Wnts play in the differentiation of osteoblasts and osteoclasts, acute inhibition of Wnt signaling is likely to affect bone homeostasis. In this study, we evaluated the skeletal effect of small molecule inhibitor of an o-acyl transferase porcupine (PORCN) that prevents Wnt signaling by blocking the secretion of all Wnts. Micro-computed tomography and histomorphometric evaluation revealed that the bones of mice treated with two structurally distinct PORCN inhibitors LGK974 and ETC-1922159 (ETC-159) had loss-of-bone volume and density within 4 weeks of exposure. This decreased bone mass was associated with a significant increase in adipocytes within the bone marrow. Notably, simultaneous administration of a clinically approved anti-resorptive, alendronate, a member of the bisphosphonate family, mitigated loss-of-bone mass seen upon ETC-159 treatment by regulating activity of osteoclasts and blocking accumulation of bone marrow adipocytes. Our results support the addition of bone protective agents when treating patients with PORCN inhibitors. Mitigation of bone toxicity can extend the therapeutic utility of Wnt pathway inhibitors.

Bone remodelling: its local regulation and the emergence of bone fragility.

PubMed

Martin, T John; Seeman, Ego

2008-10-01

Bone modelling prevents the occurrence of damage by adapting bone structure - and hence bone strength - to its loading circumstances. Bone remodelling removes damage, when it inevitably occurs, in order to maintain bone strength. This cellular machinery is successful during growth, but fails during advancing age because of the development of a negative balance between the volumes of bone resorbed and formed during remodelling by the basic multicellular unit (BMU), high rates of remodelling during midlife in women and late in life in both sexes, and a decline in periosteal bone formation. together resulting in bone loss and structural decay each time a remodelling event occurs. The two steps in remodelling - resorption of a volume of bone by osteoclasts and formation of a comparable volume by osteoblasts - are sequential, but the regulatory events leading to these two fully differentiated functions are not. Reparative remodelling is initiated by damage producing osteocyte apoptosis, which signals the location of damage via the osteocyte canalicular system to endosteal lining cells which forms the canopy of a bone-remodelling compartment (BRC). Within the BRC, local recruitment of osteoblast precursors from the lining cells, the marrow and circulation, direct contact with osteoclast precursors, osteoclastogenesis and molecular cross-talk between precursors, mature cells, cells of the immune system, and products of the resorbed matrix, titrate the birth, work and lifespan of the cells of this multicellular remodelling machinery to either remove or form a net volume of bone appropriate to the mechanical requirements.

NF-κB as a Therapeutic Target in Inflammatory-Associated Bone Diseases.

PubMed

Lin, T-H; Pajarinen, J; Lu, L; Nabeshima, A; Cordova, L A; Yao, Z; Goodman, S B

Inflammation is a defensive mechanism for pathogen clearance and maintaining tissue homeostasis. In the skeletal system, inflammation is closely associated with many bone disorders including fractures, nonunions, periprosthetic osteolysis (bone loss around orthopedic implants), and osteoporosis. Acute inflammation is a critical step for proper bone-healing and bone-remodeling processes. On the other hand, chronic inflammation with excessive proinflammatory cytokines disrupts the balance of skeletal homeostasis involving osteoblastic (bone formation) and osteoclastic (bone resorption) activities. NF-κB is a transcriptional factor that regulates the inflammatory response and bone-remodeling processes in both bone-forming and bone-resorption cells. In vitro and in vivo evidences suggest that NF-κB is an important potential therapeutic target for inflammation-associated bone disorders by modulating inflammation and bone-remodeling process simultaneously. The challenges of NF-κB-targeting therapy in bone disorders include: (1) the complexity of canonical and noncanonical NF-κB pathways; (2) the fundamental roles of NF-κB-mediated signaling for bone regeneration at earlier phases of tissue damage and acute inflammation; and (3) the potential toxic effects on nontargeted cells such as lymphocytes. Recent developments of novel inhibitors with differential approaches to modulate NF-κB activity, and the controlled release (local) or bone-targeting drug delivery (systemic) strategies, have largely increased the translational application of NF-κB therapy in bone disorders. Taken together, temporal modulation of NF-κB pathways with the combination of recent advanced bone-targeting drug delivery techniques is a highly translational strategy to reestablish homeostasis in the skeletal system. © 2017 Elsevier Inc. All rights reserved.

NF-κB as a Therapeutic Target in Inflammatory-Associated Bone Diseases

PubMed Central

Lin, T.-h.; Pajarinen, J.; Lu, L.; Nabeshima, A.; Cordova, L.A.; Yao, Z.; Goodman, S.B.

2017-01-01

Inflammation is a defensive mechanism for pathogen clearance and maintaining tissue homeostasis. In the skeletal system, inflammation is closely associated with many bone disorders including fractures, nonunions, periprosthetic osteolysis (bone loss around orthopedic implants), and osteoporosis. Acute inflammation is a critical step for proper bone-healing and bone-remodeling processes. On the other hand, chronic inflammation with excessive proinflammatory cytokines disrupts the balance of skeletal homeostasis involving osteoblastic (bone formation) and osteoclastic (bone resorption) activities. NF-κB is a transcriptional factor that regulates the inflammatory response and bone-remodeling processes in both bone-forming and bone-resorption cells. In vitro and in vivo evidences suggest that NF-κB is an important potential therapeutic target for inflammation-associated bone disorders by modulating inflammation and bone-remodeling process simultaneously. The challenges of NF-κB-targeting therapy in bone disorders include: (1) the complexity of canonical and noncanonical NF-κB pathways; (2) the fundamental roles of NF-κB-mediated signaling for bone regeneration at earlier phases of tissue damage and acute inflammation; and (3) the potential toxic effects on nontargeted cells such as lymphocytes. Recent developments of novel inhibitors with differential approaches to modulate NF-κB activity, and the controlled release (local) or bone-targeting drug delivery (systemic) strategies, have largely increased the translational application of NF-κB therapy in bone disorders. Taken together, temporal modulation of NF-κB pathways with the combination of recent advanced bone-targeting drug delivery techniques is a highly translational strategy to reestablish homeostasis in the skeletal system. PMID:28215222

Mesenchymal stem cell proliferation and mineralization but not osteogenic differentiation are strongly affected by extracellular pH.

PubMed

Fliefel, Riham; Popov, Cvetan; Tröltzsch, Matthias; Kühnisch, Jan; Ehrenfeld, Michael; Otto, Sven

2016-06-01

Osteomyelitis is a serious complication in oral and maxillofacial surgery affecting bone healing. Bone remodeling is not only controlled by cellular components but also by ionic and molecular composition of the extracellular fluids in which calcium phosphate salts are precipitated in a pH dependent manner. To determine the effect of pH on self-renewal, osteogenic differentiation and matrix mineralization of mesenchymal stem cells (MSCs). We selected three different pH values; acidic (6.3, 6.7), physiological (7.0-8.0) and severe alkaline (8.5). MSCs were cultured at different pH ranges, cell viability measured by WST-1, apoptosis detected by JC-1, senescence was analyzed by β-galactosidase whereas mineralization was detected by Alizarin Red and osteogenic differentiation analyzed by Real-time PCR. Self-renewal was affected by pH as well as matrix mineralization in which pH other than physiologic inhibited the deposition of extracellular matrix but did not affect MSCs differentiation as osteoblast markers were upregulated. The expression of osteocalcin and alkaline phosphatase activity was upregulated whereas osteopontin was downregulated under acidic pH. pH affected MSCs self-renewal and mineralization without influencing osteogenic differentiation. Thus, future therapies, based on shifting acid-base balance toward the alkaline direction might be beneficial for prevention or treatment of osteomyelitis. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.

Grizzly bears (Ursus arctos horribilis) and black bears (Ursus americanus) prevent trabecular bone loss during disuse (hibernation).

PubMed

McGee-Lawrence, Meghan E; Wojda, Samantha J; Barlow, Lindsay N; Drummer, Thomas D; Castillo, Alesha B; Kennedy, Oran; Condon, Keith W; Auger, Janene; Black, Hal L; Nelson, O Lynne; Robbins, Charles T; Donahue, Seth W

2009-12-01

Disuse typically causes an imbalance in bone formation and bone resorption, leading to losses of cortical and trabecular bone. In contrast, bears maintain balanced intracortical remodeling and prevent cortical bone loss during disuse (hibernation). Trabecular bone, however, is more detrimentally affected than cortical bone in other animal models of disuse. Here we investigated the effects of hibernation on bone remodeling, architectural properties, and mineral density of grizzly bear (Ursus arctos horribilis) and black bear (Ursus americanus) trabecular bone in several skeletal locations. There were no differences in bone volume fraction or tissue mineral density between hibernating and active bears or between pre- and post-hibernation bears in the ilium, distal femur, or calcaneus. Though indices of cellular activity level (mineral apposition rate, osteoid thickness) decreased, trabecular bone resorption and formation indices remained balanced in hibernating grizzly bears. These data suggest that bears prevent bone loss during disuse by maintaining a balance between bone formation and bone resorption, which consequently preserves bone structure and strength. Further investigation of bone metabolism in hibernating bears may lead to the translation of mechanisms preventing disuse-induced bone loss in bears into novel treatments for osteoporosis.

Grizzly bears (Ursus arctos horribilis) and black bears (Ursus americanus) prevent trabecular bone loss during disuse (hibernation)

PubMed Central

McGee-Lawrence, Meghan E.; Wojda, Samantha J.; Barlow, Lindsay N.; Drummer, Thomas D.; Castillo, Alesha B.; Kennedy, Oran; Condon, Keith W.; Auger, Janene; Black, Hal L.; Nelson, O. Lynne; Robbins, Charles T.; Donahue, Seth W.

2009-01-01

Disuse typically causes an imbalance in bone formation and bone resorption, leading to losses of cortical and trabecular bone. In contrast, bears maintain balanced intracortical remodeling and prevent cortical bone loss during disuse (hibernation). Trabecular bone, however, is more detrimentally affected than cortical bone in other animal models of disuse. Here we investigated the effects of hibernation on bone remodeling, architectural properties, and mineral density of grizzly bear (Ursus arctos horribilis) and black bear (Ursus americanus) trabecular bone in several skeletal locations. There were no differences in bone volume fraction or tissue mineral density between hibernating and active bears or between pre- and post-hibernation bears in the ilium, distal femur, or calcaneus. Though indices of cellular activity level (mineral apposition rate, osteoid thickness) decreased, trabecular bone resorption and formation indices remained balanced in hibernating grizzly bears. These data suggest that bears prevent bone loss during disuse by maintaining a balance between bone formation and bone resorption, which consequently preserves bone structure and strength. Further investigation of bone metabolism in hibernating bears may lead to the translation of mechanisms preventing disuse induced bone loss in bears into novel treatments for osteoporosis. PMID:19703606

Contribution of Vestibular-Evoked Myogenic Potential (VEMP) testing in the assessment and the differential diagnosis of otosclerosis

PubMed Central

Tramontani, Ourania; Gkoritsa, Eleni; Ferekidis, Eleftherios; Korres, Stavros G.

2014-01-01

Background The aim of this prospective clinical study was to evaluate the clinical importance of Vestibular-Evoked Myogenic Potentials (VEMPs) in the assessment and differential diagnosis of otosclerosis and otologic diseases characterized by “pseudo-conductive” components. We also investigated the clinical appearance of balance disorders in patients with otosclerosis by correlating VEMP results with the findings of caloric testing and pure tone audiometry(PTA). Material/Methods Air-conducted(AC) 4-PTA, bone-conducted(BC) 4-PTA, air-bone Gap(ABG), AC, BC tone burst evoked VEMP, and calorics were measured preoperatively in 126 otosclerotic ears. Results The response rate of the AC-VEMPs and BC-VEMPs was 29.36% and 44.03%, respectively. Statistical differences were found between the means of ABG, AC 4-PTA, and BC 4-PTA in the otosclerotic ears in relation to AC-VEMP elicitability. About one-third of patients presented with disequilibrium. A statistically significant interaction was found between calorics and dizziness in relation to PTA thresholds. No relationship was found between calorics and dizziness with VEMPs responses. Conclusions AC and BC VEMPs can be elicited in ears with otosclerosis. AC-VEMP is more vulnerable to conductive hearing loss. Evaluation of AC-VEMP thresholds can be added in the diagnostic work-up of otosclerosis in case of doubt, enhancing differential diagnosis in patients with air-bone gaps. Otosclerosis is not a cause of canal paresis or vertigo. PMID:24509900

Macrolactin F inhibits RANKL-mediated osteoclastogenesis by suppressing Akt, MAPK and NFATc1 pathways and promotes osteoblastogenesis through a BMP-2/smad/Akt/Runx2 signaling pathway.

PubMed

Li, Liang; Sapkota, Mahesh; Gao, Ming; Choi, Hyukjae; Soh, Yunjo

2017-11-15

The balance between bone formation and bone resorption is maintained by osteoblasts and osteoclasts. In the current study, macrolactin F (MF) was investigated for novel biological activity on the receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclastogenesis in primary bone marrow-derived macrophages (BMMs). We found that RANKL-induced osteoclast formation and differentiation from BMMs was significantly inhibited by MF in a dose-dependent manner without cytotoxicity. RANKL-induced F-actin ring formation and bone resorption activity in BMMs which was attenuated by MF. In addition, MF suppressed the expression of osteoclast-related genes, including c-myc, RANK, tartrate-resistant acid phosphatase (TRAP), nuclear factor of activated T cells c1 (NFATc1), cathepsin K and matrix metalloproteinase 9 (MMP9). Furthermore, the protein expression NFATc1, c-Fos, MMP9, cathepsin K and phosphorylation of Jun N-terminal kinase (JNK), p38 and Akt were also down-regulated by MF treatment. Interestingly, MF promoted pre-osteoblast cell differentiation on Alizarin Red-mineralization activity, alkaline phosphatase (ALP) activity, and the expression of osteoblastogenic markers including Runx2, Osterix, Smad4, ALP, type I collagen alpha 1 (Col1α), osteopontin (OPN), and osteocalcin (OCN) via activation of the BMP-2/smad/Akt/Runx2 pathway on MC3T3-E1. Taken together, these results indicate that MF may be useful as a therapeutic agent to enhance bone health and treat osteoporosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Divergent Resorbability and Effects on Osteoclast Formation of Commonly Used Bone Substitutes in a Human In Vitro-Assay

PubMed Central

Busse, Björn; Schilling, Arndt F.; Schinke, Thorsten; Amling, Michael; Lange, Tobias

2012-01-01

Bioactive bone substitute materials are a valuable alternative to autologous bone transplantations in the repair of skeletal defects. However, clinical studies have reported varying success rates for many commonly used biomaterials. While osteoblasts have traditionally been regarded as key players mediating osseointegration, increasing evidence suggests that bone-resorbing osteoclasts are of crucial importance for the longevity of applied biomaterials. As no standardized data on the resorbability of biomaterials exists, we applied an in vitro-assay to compare ten commonly used bone substitutes. Human peripheral blood mononuclear cells (PBMCs) were differentiated into osteoclasts in the co-presence of dentin chips and biomaterials or dentin alone (control) for a period of 28 days. Osteoclast maturation was monitored on day 0 and 14 by light microscopy, and material-dependent changes in extracellular pH were assessed twice weekly. Mature osteoclasts were quantified using TRAP stainings on day 28 and their resorptive activity was determined on dentin (toluidin blue staining) and biomaterials (scanning electron microscopy, SEM). The analyzed biomaterials caused specific changes in the pH, which were correlated with osteoclast multinuclearity (r = 0.942; p = 0.034) and activity on biomaterials (r = 0.594; p = 0.041). Perossal led to a significant reduction of pH, nuclei per osteoclast and dentin resorption, whereas Tutogen bovine and Tutobone human strikingly increased all three parameters. Furthermore, natural biomaterials were resorbed more rapidly than synthetic biomaterials leading to differential relative resorption coefficients, which indicate whether bone substitutes lead to a balanced resorption or preferential resorption of either the biomaterial or the surrounding bone. Taken together, this study for the first time compares the effects of widely used biomaterials on osteoclast formation and resorbability in an unbiased approach that may now aid in improving the preclinical evaluation of bone substitute materials. PMID:23071629

Structurally-diverse, PPARγ-activating environmental toxicants induce adipogenesis and suppress osteogenesis in bone marrow mesenchymal stromal cells

PubMed Central

Watt, James; Schlezinger, Jennifer J.

2015-01-01

Environmental obesogens are a newly recognized category of endocrine disrupting chemicals that have been implicated in contributing to the rising rates of obesity in the United States. While obesity is typically regarded as an increase in visceral fat, adipocyte accumulation in the bone has been linked to increased fracture risk, lower bone density, and osteoporosis. Exposure to environmental toxicants that activate peroxisome proliferator activated receptor γ (PPARγ), a critical regulator of the balance of differentiation between adipogenesis and osteogenesis, may contribute to the increasing prevalence of osteoporosis. However, induction of adipogenesis and suppression of osteogenesis are separable activities of PPARγ, and ligands may selectively alter these activities. It currently is unknown whether suppression of osteogenesis is a common toxic endpoint of environmental PPARγ ligands. Using a primary mouse bone marrow culture model, we tested the hypothesis that environmental toxicants acting as PPARγ agonists divert the differentiation pathway of bone marrow-derived multipotent mesenchymal stromal cells towards adipogenesis and away from osteogenesis. The toxicants tested included the organotins tributyltin and triphenyltin, a ubiquitous phthalate metabolite (mono-(2-ethylhexyl) phthalate, MEHP), and two brominated flame retardants (tetrabromobisphenol-a, TBBPA, and mono-(2-ethylhexyl) tetrabromophthalate, METBP). All of the compounds activated PPARγ1 and 2. All compounds increased adipogenesis (lipid accumulation, Fabp4 expression) and suppressed osteogenesis (alkaline phosphatase activity, Osx expression) in mouse primary bone marrow cultures, but with different potencies and efficacies. Despite structural dissimilarities, there was a strong negative correlation between efficacies to induce adipogenesis and suppress osteogenesis, with the organotins being distinct in their exceptional ability to suppress osteogenesis. As human exposure to a mixture of toxicants is likely, albeit at low doses, the fact that multiple toxicants are capable of suppressing bone formation supports the hypothesis that environmental PPARγ ligands represent an emerging threat to human bone health. PMID:25777084

Formononetin, an isoflavone, activates AMP-activated protein kinase/β-catenin signalling to inhibit adipogenesis and rescues C57BL/6 mice from high-fat diet-induced obesity and bone loss.

PubMed

Gautam, Jyoti; Khedgikar, Vikram; Kushwaha, Priyanka; Choudhary, Dharmendra; Nagar, Geet Kumar; Dev, Kapil; Dixit, Preety; Singh, Divya; Maurya, Rakesh; Trivedi, Ritu

2017-03-01

Balance between adipocyte and osteoblast differentiation is the key link of disease progression in obesity and osteoporosis. We have previously reported that formononetin (FNT), an isoflavone extracted from Butea monosperma, stimulates osteoblast formation and protects against postmenopausal bone loss. The inverse relationship between osteoblasts and adipocytes prompted us to analyse the effect of FNT on adipogenesis and in vivo bone loss, triggered by high-fat diet (HFD)-induced obesity. The anti-obesity effect and mechanism of action of FNT was determined in 3T3-L1 cells and HFD-induced obese male mice. Our findings show that FNT suppresses the adipogenic differentiation of 3T3-L1 fibroblasts, through down-regulation of key adipogenic markers such as PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα) and sterol regulatory element-binding protein (SREBP) and inhibits intracellular TAG accumulation. Increased intracellular reactive oxygen species levels and AMP-activated protein kinase (AMPK) activation accompanied by stabilisation of β-catenin were attributed to the anti-adipogenic action of FNT. In vivo, 12 weeks of FNT treatment inhibited the development of obesity in mice by attenuating HFD-induced body weight gain and visceral fat accumulation. The anti-obesity effect of FNT results from increased energy expenditure. FNT also protects against HFD-induced dyslipidaemia and rescues deterioration of trabecular bone volume by increasing bone formation and decreasing bone resorbtion caused by HFD. FNT's rescuing action against obesity-induced osteoporosis commenced at the level of progenitors, as bone marrow progenitor cells, obtained from the HFD mice group supplemented with FNT, showed increased osteogenic and decreased adipogenic potentials. Our findings suggest that FNT inhibits adipogenesis through AMPK/β-catenin signal transduction pathways and protects against HFD-induced obesity and bone loss.

Role of RANKL in bone diseases.

PubMed

Anandarajah, Allen P

2009-03-01

Bone remodeling is a tightly regulated process of osteoclast-mediated bone resorption, balanced by osteoblast-mediated bone formation. Disruption of this balance can lead to increased bone turnover, resulting in excessive bone loss or extra bone formation and consequent skeletal disease. The receptor activator of nuclear factor kappaB ligand (RANKL) (along with its receptor), the receptor activator of nuclear factor kappaB and its natural decoy receptor, osteoprotegerin, are the final effector proteins of osteoclastic bone resorption. Here, I provide an overview of recent studies that highlight the key role of RANKL in the pathophysiology of several bone diseases and discuss the novel therapeutic approaches afforded by the modulation of RANKL.

MicroRNA-29a mitigates glucocorticoid induction of bone loss and fatty marrow by rescuing Runx2 acetylation.

PubMed

Ko, Jih-Yang; Chuang, Pei-Chin; Ke, Huei-Jin; Chen, Yu-Shan; Sun, Yi-Chih; Wang, Feng-Sheng

2015-12-01

Glucocorticoid treatment reportedly increases the morbidity of osteoporotic or osteonecrotic disorders. Exacerbated bone acquisition and escalated marrow adipogenesis are prominent pathological features of glucocorticoid-mediated skeletal disorders. MicroRNAs reportedly modulate tissue metabolism and remodeling. This study was undertaken to investigate the biological roles of microRNA-29a (miR-29a) in skeletal and fat metabolism in the pathogenesis of glucocorticoid-induced osteoporosis. Transgenic mice overexpressing miR-29a precursor or wild-type mice were given methylprednisolone. Bone mass, microarchitecture and histology were assessed by dual energy X-ray absorptiometry, μCT and histomorphometry. Differential gene expression and signaling components were delineated by quantitative RT-PCR and immunoblotting. Glucocorticoid treatment accelerated bone loss and marrow fat accumulation in association with decreased miR-29a expression. The miR-29a transgenic mice had high bone mineral density, trabecular microarchitecture and cortical thickness. miR-29a overexpression mitigated the glucocorticoid-induced impediment of bone mass, skeletal microstructure integrity and mineralization reaction and attenuated fatty marrow histopathology. Ex vivo, miR-29a increased osteogenic differentiation capacity and alleviated the glucocorticoid-induced promotion of adipocyte formation in primary bone-marrow mesenchymal progenitor cell cultures. Through inhibition of histone deacetylase 4 (HDAC4) expression, miR-29a restored acetylated Runx2 and β-catenin abundances and reduced RANKL, leptin and glucocorticoid receptor expression in glucocorticoid-mediated osteoporosis bone tissues. Taken together, glucocorticoid suppression of miR-29a signaling disturbed the balances between osteogenic and adipogenic activities, and thereby interrupted bone formation and skeletal homeostasis. miR-29a inhibition of HDAC4 stabilized the acetylation state of Runx2 and β-catenin that ameliorated the detrimental effects of glucocorticoid on mineralization and lipogenesis reactions in bone tissue microenvironments. This study highlighted emerging skeletal-anabolic actions of miR-29a signaling in the progression of glucocorticoid-induced bone tissue destruction. Sustaining miR-29a actions is beneficial in protecting against glucocorticoid-mediated osteoporosis. Copyright © 2015 Elsevier Inc. All rights reserved.

Canonical Wnt signaling in differentiated osteoblasts controls osteoclast differentiation.

PubMed

Glass, Donald A; Bialek, Peter; Ahn, Jong Deok; Starbuck, Michael; Patel, Millan S; Clevers, Hans; Taketo, Mark M; Long, Fanxin; McMahon, Andrew P; Lang, Richard A; Karsenty, Gerard

2005-05-01

Inactivation of beta-catenin in mesenchymal progenitors prevents osteoblast differentiation; inactivation of Lrp5, a gene encoding a likely Wnt coreceptor, results in low bone mass (osteopenia) by decreasing bone formation. These observations indicate that Wnt signaling controls osteoblast differentiation and suggest that it may regulate bone formation in differentiated osteoblasts. Here, we study later events and find that stabilization of beta-catenin in differentiated osteoblasts results in high bone mass, while its deletion from differentiated osteoblasts leads to osteopenia. Surprisingly, histological analysis showed that these mutations primarily affect bone resorption rather than bone formation. Cellular and molecular studies showed that beta-catenin together with TCF proteins regulates osteoblast expression of Osteoprotegerin, a major inhibitor of osteoclast differentiation. These findings demonstrate that beta-catenin, and presumably Wnt signaling, promote the ability of differentiated osteoblasts to inhibit osteoclast differentiation; thus, they broaden our knowledge of the functions Wnt proteins have at various stages of skeletogenesis.

Specific Biomimetic Hydroxyapatite Nanotopographies Enhance Osteoblastic Differentiation and Bone Graft Osteointegration

PubMed Central

Loiselle, Alayna E.; Wei, Lai; Faryad, Muhammad; Paul, Emmanuel M.; Lewis, Gregory S.; Gao, Jun; Lakhtakia, Akhlesh

2013-01-01

Impaired healing of cortical bone grafts represents a significant clinical problem. Cadaveric bone grafts undergo extensive chemical processing to decrease the risk of disease transmission; however, these processing techniques alter the bone surface and decrease the osteogenic potential of cells at the healing site. Extensive work has been done to optimize the surface of bone grafts, and hydroxyapatite (HAP) and nanotopography both increase osteoblastic differentiation. HAP is the main mineral component of bone and can enhance osteoblastic differentiation and bone implant healing in vivo, while nanotopography can enhance osteoblastic differentiation, adhesion, and proliferation. This is the first study to test the combined effects of HAP and nanotopographies on bone graft healing. With the goal of identifying the optimized surface features to improve bone graft healing, we tested the hypothesis that HAP-based nanotopographic resurfacing of bone grafts improves integration of cortical bone grafts by enhancing osteoblastic differentiation. Here we show that osteoblastic cells cultured on processed bones coated with specific-scale (50–60 nm) HAP nanotopographies display increased osteoblastic differentiation compared to cells on uncoated bone, bones coated with poly-l-lactic acid nanotopographies, or other HAP nanotopographies. Further, bone grafts coated with 50–60-nm HAP exhibited increased formation of new bone and improved healing, with mechanical properties equivalent to live autografts. These data indicate the potential for specific HAP nanotopographies to not only increase osteoblastic differentiation but also improve bone graft incorporation, which could significantly increase patient quality of life after traumatic bone injuries or resection of an osteosarcoma. PMID:23510012

The Rho-GEF Kalirin regulates bone mass and the function of osteoblasts and osteoclasts

PubMed Central

Huang, Su; Eleniste, Pierre P.; Wayakanon, Kornchanok; Mandela, Prashant; Eipper, Betty A.; Mains, Richard E.; Allen, Matthew R.; Bruzzaniti, Angela

2014-01-01

Bone homeostasis is maintained by the balance between bone resorption by osteoclasts and bone formation by osteoblasts. Dysregulation in the activity of the bone cells can lead to osteoporosis, a disease characterized by low bone mass and an increase in bone fragility and risk of fracture. Kalirin is a novel GTP-exchange factor protein that has been shown to play a role in cytoskeletal remodeling and dendritic spine formation in neurons. We examined Kalirin expression in skeletal tissue and found that it was expressed in osteoclasts and osteoblasts. Furthermore, micro-CT analyses of the distal femur of global Kalirin knockout (Kal-KO) mice revealed significantly reduced trabecular and cortical bone parameters in Kal-KO mice, compared to WT mice, with significantly reduced bone mass in 8, 14 and 36 week-old female Kal-KO mice. Male mice also exhibited a decrease in bone parameters but not to the level seen in female mice. Histomorphometric analyses also revealed decreased bone formation rate in 14 week-old female Kal-KO mice, as well as decreased osteoblast number/bone surface and increased osteoclast surface/bone surface. Consistent with our in vivo findings, the bone resorbing activity and differentiation of Kal-KO osteoclasts was increased in vitro. Although alkaline phosphatase activity by Kal-KO osteoblasts was increased in vitro, Kal-KO osteoblasts showed decreased mineralizing activity, as well as decreased secretion of OPG, which was inversely correlated with ERK activity. Taken together, our findings suggest that deletion of Kalirin directly affects osteoclast and osteoblast activity, leading to decreased OPG secretion by osteoblasts which is likely to alter the RANKL/OPG ratio and promote osteoclastogenesis. Therefore, Kalirin may play a role in paracrine and/or endocrine signaling events that control skeletal bone remodeling and the maintenance of bone mass. PMID:24380811

Calcium metabolism in health and disease.

PubMed

Peacock, Munro

2010-01-01

This brief review focuses on calcium balance and homeostasis and their relationship to dietary calcium intake and calcium supplementation in healthy subjects and patients with chronic kidney disease and mineral bone disorders (CKD-MBD). Calcium balance refers to the state of the calcium body stores, primarily in bone, which are largely a function of dietary intake, intestinal absorption, renal excretion, and bone remodeling. Bone calcium balance can be positive, neutral, or negative, depending on a number of factors, including growth, aging, and acquired or inherited disorders. Calcium homeostasis refers to the hormonal regulation of serum ionized calcium by parathyroid hormone, 1,25-dihydroxyvitamin D, and serum ionized calcium itself, which together regulate calcium transport at the gut, kidney, and bone. Hypercalcemia and hypocalcemia indicate serious disruption of calcium homeostasis but do not reflect calcium balance on their own. Calcium balance studies have determined the dietary and supplemental calcium requirements needed to optimize bone mass in healthy subjects. However, similar studies are needed in CKD-MBD, which disrupts both calcium balance and homeostasis, because these data in healthy subjects may not be generalizable to this patient group. Importantly, increasing evidence suggests that calcium supplementation may enhance soft tissue calcification and cardiovascular disease in CKD-MBD. Further research is needed to elucidate the risks and mechanisms of soft tissue calcification with calcium supplementation in both healthy subjects and CKD-MBD patients.

Impaired osteoblast function in osteoporosis: comparison between calcium balance and dynamic histomorphometry.

PubMed

Arlot, M; Edouard, C; Meunier, P J; Neer, R M; Reeve, J

1984-09-01

Osteoblast function was investigated in 27 patients with idiopathic osteoporosis. Transiliac bone biopsy specimens were taken after double labelling with tetracycline, and metabolic calcium balance was studied almost simultaneously. Many of the patients showed poor double labelling of their otherwise unremarkable trabecular osteoid, suggesting impaired formation of bone at many of these surfaces. This phenomenon was not accompanied by increased width of osteoid seams (as seen in osteomalacia), indicating that formation of the matrix and its mineralisation were in equilibrium. For the first time, highly significant positive correlations (p less than 0.01) were found between indices of bone formation, determined by labelling with tetracycline, and calcium balance. Thus some patients with osteoporosis who are rapidly losing bone have low rates of formation of trabecular bone both by individual osteoblasts and in relation to available bone surfaces. As histological indices of bone resorption also independently correlated strongly and inversely (p less than 0.01) with calcium balance the rate of initiation of new basic multicellular units by osteoclastic resorption of trabecular surfaces (or the depth of resorption at these surfaces) also appears to be an important determinant of mineral balance. The mechanisms that regulate the effective life span of mature osteoblasts require further investigation, particularly as some promising treatments that can increase trabecular bone volume in osteoporosis, such as parathyroid peptide hPTH (1-34) and sodium fluoride, must work through a reversal of osteoblastic depression.

Acute Ketamine Administration Corrects Abnormal Inflammatory Bone Markers in Major Depressive Disorder

PubMed Central

Kadriu, Bashkim; Gold, Philip W; Luckenbaugh, David A; Lener, Marc S; Ballard, Elizabeth D; Niciu, Mark J; Henter, Ioline D; Park, Lawrence T; De Sousa, Rafael Teixeira; Yuan, Peixiong; Machado-Vieira, Rodrigo; Zarate, Carlos A

2017-01-01

Patients with major depressive disorder (MDD) have clinically relevant, significant decreases in bone mineral density (BMD). We sought to determine if predictive markers of bone inflammation—the osteoprotegerin (OPG)-RANK-RANKL system or osteopontin (OPN)—play a role in the bone abnormalities associated with MDD and, if so, whether ketamine treatment corrected the abnormalities. The OPG-RANK-RANKL system plays the principal role in determining the balance between bone resorption and bone formation. RANKL is the osteoclast differentiating factor and diminishes BMD. OPG is a decoy receptor for RANKL, thereby increasing BMD. OPN is the bone glue that acts as a scaffold between bone tissues matrix composition to bind them together and is an important component of bone strength and fracture resistance. Twenty-eight medication-free inpatients with treatment-resistant MDD and 16 healthy controls (HCs) participated in the study. Peripheral bone marker levels and their responses to IV ketamine infusion in MDD patients and HCs were measured at four time points: at baseline, and post-infusion at 230 minutes, Day 1, and Day 3. Patients with MDD had significant decreases in baseline OPG/RANKL ratio and in plasma OPN levels. Ketamine significantly increased both the OPG/RANKL ratio and plasma OPN levels and significantly decreased RANKL levels. Bone marker levels in HCs remained unaltered. We conclude that the OPG-RANK-RANKL system and the OPN system play important roles in the serious bone abnormalities associated with MDD. These data suggest that in addition to its antidepressant effects, ketamine also has a salutary effect on a major medical complication of depressive illness. PMID:28555075

Asfotase-α improves bone growth, mineralization and strength in mouse models of neurofibromatosis type-1

PubMed Central

de la Croix Ndong, Jean; Makowski, Alexander James; Uppuganti, Sasidhar; Vignaux, Guillaume; Ono, Koichiro; Perrien, Daniel S.; Joubert, Simon; Baglio, Serena R.; Granchi, Donatella; Stevenson, David A.; Rios, Jonathan J.; Nyman, Jeffry S.; Elefteriou, Florent

2014-01-01

Mineralization of the skeleton depends on the balance between levels of pyrophosphate (PPi), an inhibitor of hydroxyapatite formation, and phosphate generated from PPi breakdown by alkaline phosphatase (ALP). We report here that ablation of Nf1, encoding the RAS/GTPase–activating protein neurofibromin, in bone–forming cells leads to supraphysiologic PPi accumulation, caused by a chronic ERK–dependent increase in genes promoting PPi synthesis and extracellular transport, namely Enpp1 and Ank. It also prevents BMP2–induced osteoprogenitor differentiation and, consequently, expression of ALP and PPi breakdown, further contributing to PPi accumulation. The short stature, impaired bone mineralization and strength in mice lacking Nf1 in osteochondroprogenitors or osteoblasts could be corrected by enzyme therapy aimed at reducing PPi concentration. These results establish neurofibromin as an essential regulator of bone mineralization, suggest that altered PPi homeostasis contributes to the skeletal dysplasiae associated with neurofibromatosis type-1 (NF1), and that some of the NF1 skeletal conditions might be preventable pharmacologically. PMID:24997609

Spaceflight-induced vertebral bone loss in ovariectomized rats is associated with increased bone marrow adiposity and no change in bone formation

PubMed Central

Keune, Jessica A; Philbrick, Kenneth A; Branscum, Adam J; Iwaniec, Urszula T; Turner, Russell T

2016-01-01

There is often a reciprocal relationship between bone marrow adipocytes and osteoblasts, suggesting that marrow adipose tissue (MAT) antagonizes osteoblast differentiation. MAT is increased in rodents during spaceflight but a causal relationship between MAT and bone loss remains unclear. In the present study, we evaluated the effects of a 14-day spaceflight on bone mass, bone resorption, bone formation, and MAT in lumbar vertebrae of ovariectomized (OVX) rats. Twelve-week-old OVX Fischer 344 rats were randomly assigned to a ground control or flight group. Following flight, histological sections of the second lumbar vertebrae (n=11/group) were stained using a technique that allowed simultaneous quantification of cells and preflight fluorochrome label. Compared with ground controls, rats flown in space had 32% lower cancellous bone area and 306% higher MAT. The increased adiposity was due to an increase in adipocyte number (224%) and size (26%). Mineral apposition rate and osteoblast turnover were unchanged during spaceflight. In contrast, resorption of a preflight fluorochrome and osteoclast-lined bone perimeter were increased (16% and 229%, respectively). The present findings indicate that cancellous bone loss in rat lumbar vertebrae during spaceflight is accompanied by increased bone resorption and MAT but no change in bone formation. These findings do not support the hypothesis that increased MAT during spaceflight reduces osteoblast activity or lifespan. However, in the context of ovarian hormone deficiency, bone formation during spaceflight was insufficient to balance increased resorption, indicating defective coupling. The results are therefore consistent with the hypothesis that during spaceflight mesenchymal stem cells are diverted to adipocytes at the expense of forming osteoblasts. PMID:28725730

Musculoskeletal strength, balance performance, and self-efficacy in elderly ving tsun chinese martial art practitioners: implications for fall prevention.

PubMed

Fong, Shirley S M; Ng, Shamay S M; Liu, Karen P Y; Pang, Marco Y C; Lee, H W; Chung, Joanne W Y; Lam, Priscillia L; Guo, X

2014-01-01

Objectives. To (1) compare the bone strength, lower limb muscular strength, functional balance performance, and balance self-efficacy between Ving Tsun (VT) martial art practitioners and nonpractitioners and (2) identify the associations between lower limb muscular strength, functional balance performance, and balance self-efficacy among the VT-trained participants. Methods. Thirty-five VT practitioners (mean age ± SD = 62.7 ± 13.3 years) and 49 nonpractitioners (mean age ± SD = 65.9 ± 10.5 years) participated in the study. The bone strength of the distal radius, lower limb muscular strength, functional balance performance, and balance self-efficacy were assessed using an ultrasound bone sonometer, the five times sit-to-stand test (FTSTS), the Berg balance scale (BBS), and the Chinese version of the activities-specific balance confidence scale, respectively. A multivariate analysis of covariance was performed to compare all the outcome variables between the two groups. Results. Elderly VT practitioners had higher radial bone strength on the dominant side (P < 0.05), greater lower limb muscular strength (P = 0.001), better functional balance performance (P = 0.003), and greater balance confidence (P < 0.001) than the nonpractitioners. Additionally, only the FTSTS time revealed a significant association with the BBS score (r = -0.575,  P = 0.013). Conclusions. VT may be a suitable health-maintenance exercise for the elderly. Our findings may inspire the development of VT fall-prevention exercises for the community-dwelling healthy elderly.

Musculoskeletal Strength, Balance Performance, and Self-Efficacy in Elderly Ving Tsun Chinese Martial Art Practitioners: Implications for Fall Prevention

PubMed Central

Fong, Shirley S. M.; Ng, Shamay S. M.; Liu, Karen P. Y.; Pang, Marco Y. C.; Lee, H. W.; Chung, Joanne W. Y.; Lam, Priscillia L.; Guo, X.

2014-01-01

Objectives. To (1) compare the bone strength, lower limb muscular strength, functional balance performance, and balance self-efficacy between Ving Tsun (VT) martial art practitioners and nonpractitioners and (2) identify the associations between lower limb muscular strength, functional balance performance, and balance self-efficacy among the VT-trained participants. Methods. Thirty-five VT practitioners (mean age ± SD = 62.7 ± 13.3 years) and 49 nonpractitioners (mean age ± SD = 65.9 ± 10.5 years) participated in the study. The bone strength of the distal radius, lower limb muscular strength, functional balance performance, and balance self-efficacy were assessed using an ultrasound bone sonometer, the five times sit-to-stand test (FTSTS), the Berg balance scale (BBS), and the Chinese version of the activities-specific balance confidence scale, respectively. A multivariate analysis of covariance was performed to compare all the outcome variables between the two groups. Results. Elderly VT practitioners had higher radial bone strength on the dominant side (P < 0.05), greater lower limb muscular strength (P = 0.001), better functional balance performance (P = 0.003), and greater balance confidence (P < 0.001) than the nonpractitioners. Additionally, only the FTSTS time revealed a significant association with the BBS score (r = −0.575,  P = 0.013). Conclusions. VT may be a suitable health-maintenance exercise for the elderly. Our findings may inspire the development of VT fall-prevention exercises for the community-dwelling healthy elderly. PMID:25530782

Water extract of the fruits of Alpinia oxyphylla inhibits osteoclast differentiation and bone loss.

PubMed

Ha, Hyunil; Shim, Ki-Shuk; Kim, Taesoo; Lee, Chung-Jo; Park, Ji Hyung; Kim, Han Sung; Ma, Jin Yeul

2014-09-23

Excessive bone resorption by osteoclasts causes pathological bone destruction, seen in various bone diseases. There is accumulating evidence that certain herbal extracts have beneficial effects on bone metabolism. The fruits of Alpinia oxyphylla has been traditionally used for the treatment of diarrhea and enuresis. In this study, we investigated the effects of water extract of the fruits of Alpinia oxyphylla (WEAO) on osteoclast differentiation and osteoclast-mediated bone destruction. For osteoclast differentiation assay, mouse bone marrow-derived macrophages (BMMs) were cultured in the presence of RANKL and M-CSF. RANKL signaling pathways and gene expression of transcription factors regulating osteoclast differentiation were investigated by real-time PCR and Western blotting. A constitutively active form of NFATc1 was retrovirally transduced into BMMs. Bone resorbing activity of mature osteoclast was examined on a plate coated with an inorganic crystalline calcium phosphate. The in vivo effect against bone destruction was assessed in a murine model of RANKL-induced osteoporosis by micro-computed tomography and bone metabolism marker analyses. WEAO dose-dependently inhibited RANKL-induced osteoclast differentiation from BMMs by targeting the early stages of osteoclast differentiation. WEAO inhibited RANKL-induced expression of NFATc1, the master regulator of osteoclast differentiation. Overexpression of a constitutively active form of NFATc1 blunted the inhibitory effect of WEAO on osteoclast differentiation, suggesting that NFATc1 is a critical target of the inhibitory action of WEAO. WEAO inhibited RANKL-induced expression of c-Fos, an upstream activator of NFATc1, by suppressing the classical NF-κB signaling pathway. WEAO also inhibited RANKL-induced down-regulation of Id2 and MafB, negative regulators of NFATc1. WEAO does not directly affect bone resorbing activity of mature osteoclasts. In accordance with the in vitro results, WEAO attenuated RANKL-induced bone destruction in mice by inhibiting osteoclast differentiation. This study demonstrates that WEAO exhibits a protective effect against bone loss by inhibiting RANKL-induced osteoclast differentiation. These findings suggest that WEAO might be useful for the prevention and treatment of bone diseases associated with excessive bone resorption.

[Hearing and balance in metabolic bone diseases].

PubMed

Zatoński, Tomasz; Temporale, Hanna; Krecicki, Tomasz

2012-03-01

There are reports that hearing loss is one of the clinical manifestations of metabolic bone diseases. Demineralization can lead to a reduction in ossicular mass. Paget's disease can reveal loss of mineral density of the cochlear bone. Ear bone remodeling in osteoporosis is similar to the changes in otosclerosis. Moreover, osteoporosis, osteogenesis imperfecta and otosclerosis have a similar genetic mechanism. According to some researchers osteopenia and osteoporosis may well be associated with idiopathic benign positional vertigo (BPV). Dysfunction of the organ of hearing and balance in patients with renal insufficiency may be due to disturbances in calcium phosphate balance and renal osteodystrophy in the course of the disease. Proving the presence of hearing loss in patients with metabolic bone diseases may lead to determining the new indications for bone densitometry in some patients with hearing impairment. Furthermore, audiological examination in patients with osteoporosis may be important because of the impact of hearing loss on prognosis for patients with metabolic bone diseases.

Bone formation: roles of genistein and daidzein

USDA-ARS?s Scientific Manuscript database

Bone remodeling consists of a balance between bone formation by osteoblasts and bone resorption by osteoclasts. Osteoporosis is the result of increased bone resorption and decreased bone formation causing a decreased bone mass density, loss of bone microarchitecture, and an increased risk of fractu...

Decreased bone turnover with balanced resorption and formation prevent cortical bone loss during disuse (hibernation) in grizzly bears (Ursus arctos horribilis).

PubMed

McGee, Meghan E; Maki, Aaron J; Johnson, Steven E; Nelson, O Lynne; Robbins, Charles T; Donahue, Seth W

2008-02-01

Disuse uncouples bone formation from resorption, leading to increased porosity, decreased bone geometrical properties, and decreased bone mineral content which compromises bone mechanical properties and increases fracture risk. However, black bear bone properties are not adversely affected by aging despite annual periods of disuse (i.e., hibernation), which suggests that bears either prevent bone loss during disuse or lose bone and subsequently recover it at a faster rate than other animals. Here we show decreased cortical bone turnover during hibernation with balanced formation and resorption in grizzly bear femurs. Hibernating grizzly bear femurs were less porous and more mineralized, and did not demonstrate any changes in cortical bone geometry or whole bone mechanical properties compared to active grizzly bear femurs. The activation frequency of intracortical remodeling was 75% lower during hibernation than during periods of physical activity, but the normalized mineral apposition rate was unchanged. These data indicate that bone turnover decreases during hibernation, but osteons continue to refill at normal rates. There were no changes in regional variation of porosity, geometry, or remodeling indices in femurs from hibernating bears, indicating that hibernation did not preferentially affect one region of the cortex. Thus, grizzly bears prevent bone loss during disuse by decreasing bone turnover and maintaining balanced formation and resorption, which preserves bone structure and strength. These results support the idea that bears possess a biological mechanism to prevent disuse osteoporosis.

PRMT5 is essential for the maintenance of chondrogenic progenitor cells in the limb bud

PubMed Central

Norrie, Jacqueline L.; Li, Qiang; Co, Swanie; Huang, Bau-Lin; Ding, Ding; Uy, Jann C.; Ji, Zhicheng; Mackem, Susan; Bedford, Mark T.; Galli, Antonella; Ji, Hongkai

2016-01-01

During embryonic development, undifferentiated progenitor cells balance the generation of additional progenitor cells with differentiation. Within the developing limb, cartilage cells differentiate from mesodermal progenitors in an ordered process that results in the specification of the correct number of appropriately sized skeletal elements. The internal pathways by which these cells maintain an undifferentiated state while preserving their capacity to differentiate is unknown. Here, we report that the arginine methyltransferase PRMT5 has a crucial role in maintaining progenitor cells. Mouse embryonic buds lacking PRMT5 have severely truncated bones with wispy digits lacking joints. This novel phenotype is caused by widespread cell death that includes mesodermal progenitor cells that have begun to precociously differentiate into cartilage cells. We propose that PRMT5 maintains progenitor cells through its regulation of Bmp4. Intriguingly, adult and embryonic stem cells also require PRMT5 for maintaining pluripotency, suggesting that similar mechanisms might regulate lineage-restricted progenitor cells during organogenesis. PMID:27827819

PRMT5 is essential for the maintenance of chondrogenic progenitor cells in the limb bud.

PubMed

Norrie, Jacqueline L; Li, Qiang; Co, Swanie; Huang, Bau-Lin; Ding, Ding; Uy, Jann C; Ji, Zhicheng; Mackem, Susan; Bedford, Mark T; Galli, Antonella; Ji, Hongkai; Vokes, Steven A

2016-12-15

During embryonic development, undifferentiated progenitor cells balance the generation of additional progenitor cells with differentiation. Within the developing limb, cartilage cells differentiate from mesodermal progenitors in an ordered process that results in the specification of the correct number of appropriately sized skeletal elements. The internal pathways by which these cells maintain an undifferentiated state while preserving their capacity to differentiate is unknown. Here, we report that the arginine methyltransferase PRMT5 has a crucial role in maintaining progenitor cells. Mouse embryonic buds lacking PRMT5 have severely truncated bones with wispy digits lacking joints. This novel phenotype is caused by widespread cell death that includes mesodermal progenitor cells that have begun to precociously differentiate into cartilage cells. We propose that PRMT5 maintains progenitor cells through its regulation of Bmp4 Intriguingly, adult and embryonic stem cells also require PRMT5 for maintaining pluripotency, suggesting that similar mechanisms might regulate lineage-restricted progenitor cells during organogenesis. © 2016. Published by The Company of Biologists Ltd.

Acute ketamine administration corrects abnormal inflammatory bone markers in major depressive disorder.

PubMed

Kadriu, B; Gold, P W; Luckenbaugh, D A; Lener, M S; Ballard, E D; Niciu, M J; Henter, I D; Park, L T; De Sousa, R T; Yuan, P; Machado-Vieira, R; Zarate, C A

2017-05-30

Patients with major depressive disorder (MDD) have clinically relevant, significant decreases in bone mineral density (BMD). We sought to determine if predictive markers of bone inflammation-the osteoprotegerin (OPG)-RANK-RANKL system or osteopontin (OPN)-play a role in the bone abnormalities associated with MDD and, if so, whether ketamine treatment corrected the abnormalities. The OPG-RANK-RANKL system plays the principal role in determining the balance between bone resorption and bone formation. RANKL is the osteoclast differentiating factor and diminishes BMD. OPG is a decoy receptor for RANKL, thereby increasing BMD. OPN is the bone glue that acts as a scaffold between bone tissues matrix composition to bind them together and is an important component of bone strength and fracture resistance. Twenty-eight medication-free inpatients with treatment-resistant MDD and 16 healthy controls (HCs) participated in the study. Peripheral bone marker levels and their responses to IV ketamine infusion in MDD patients and HCs were measured at four time points: at baseline, and post-infusion at 230 min, Day 1, and Day 3. Patients with MDD had significant decreases in baseline OPG/RANKL ratio and in plasma OPN levels. Ketamine significantly increased both the OPG/RANKL ratio and plasma OPN levels, and significantly decreased RANKL levels. Bone marker levels in HCs remained unaltered. We conclude that the OPG-RANK-RANKL system and the OPN system play important roles in the serious bone abnormalities associated with MDD. These data suggest that, in addition to its antidepressant effects, ketamine also has a salutary effect on a major medical complication of depressive illness.Molecular Psychiatry advance online publication, 30 May 2017; doi:10.1038/mp.2017.109.

Effects of Arsenic on Osteoblast Differentiation in Vitro and on Bone Mineral Density and Microstructure in Rats

PubMed Central

Wu, Cheng-Tien; Lu, Tung-Ying; Chan, Ding-Cheng; Tsai, Keh-Sung; Yang, Rong-Sen

2014-01-01

Background: Arsenic is a ubiquitous toxic element and is known to contaminate drinking water in many countries. Several epidemiological studies have shown that arsenic exposure augments the risk of bone disorders. However, the detailed effect and mechanism of inorganic arsenic on osteoblast differentiation of bone marrow stromal cells and bone loss still remain unclear. Objectives: We investigated the effects and mechanism of arsenic on osteoblast differentiation in vitro and evaluated bone mineral density (BMD) and bone microstructure in rats at doses relevant to human exposure from drinking water. Methods: We used a cell model of rat primary bone marrow stromal cells (BMSCs) and a rat model of long-term exposure with arsenic-contaminated drinking water, and determined bone microstructure and BMD in rats by microcomputed tomography (μCT). Results: We observed significant attenuation of osteoblast differentiation after exposure of BMSCs to arsenic trioxide (0.5 or 1 μM). After arsenic treatment during differentiation, expression of runt-related transcription factor-2 (Runx2), bone morphogenetic protein-2 (BMP-2), and osteocalcin in BMSCs was inhibited and phosphorylation of enhanced extracellular signal-regulated kinase (ERK) was increased. These altered differentiation-related molecules could be reversed by the ERK inhibitor PD98059. Exposure of rats to arsenic trioxide (0.05 or 0.5 ppm) in drinking water for 12 weeks altered BMD and microstructure, decreased Runx2 expression, and increased ERK phosphorylation in bones. In BMSCs isolated from arsenic-treated rats, osteoblast differentiation was inhibited. Conclusions: Our results suggest that arsenic is capable of inhibiting osteoblast differentiation of BMSCs via an ERK-dependent signaling pathway and thus increasing bone loss. Citation: Wu CT, Lu TY, Chan DC, Tsai KS, Yang RS, Liu SH. 2014. Effects of arsenic on osteoblast differentiation in vitro and on bone mineral density and microstructure in rats. Environ Health Perspect 122:559–565; http://dx.doi.org/10.1289/ehp.1307832 PMID:24531206

Epigenetic regulation of fetal bone development and placental transfer of nutrients: progress for osteoporosis.

PubMed

Bocheva, Georgeta; Boyadjieva, Nadka

2011-12-01

Osteoporosis is a common age-related disorder and causes acute and long-term disability and economic cost. Many factors influence the accumulation of bone minerals, including heredity, diet, physical activity, gender, endocrine functions, and risk factors such as alcohol, drug abuse, some pharmacological drugs or cigarette smoking. The pathology of bone development during intrauterine life is a factor for osteoporosis. Moreover, the placental transfer of nutrients plays an important role in the building of bones of fetuses. The importance of maternal calcium intake and vitamin D status are highlighted in this review. Various environmental factors including nutrition state or maternal stress may affect the epigenetic state of a number of genes during fetal development of bones. Histone modifications as histone hypomethylation, histone hypermethylation, hypoacetylation, etc. are involved in chromatin remodeling, known to contribute to the epigenetic landscape of chromosomes, and play roles in both fetal bone development and osteoporosis. This review will give an overview of epigenetic modulation of bone development and placental transfer of nutrients. In addition, the data from animal and human studies support the role of epigenetic modulation of calcium and vitamin D in the pathogenesis of osteoporosis. We review the evidence suggesting that various genes are involved in regulation of osteoclast formation and differentiation by osteoblasts and stem cells. Epigenetic changes in growth factors as well as cytokines play a rol in fetal bone development. On balance, the data suggest that there is a link between epigenetic changes in placental transfer of nutrients, including calcium and vitamin D, abnormal intrauterine bone development and pathogenesis of osteoporosis.

Hypercalciuric Bone Disease

NASA Astrophysics Data System (ADS)

Favus, Murray J.

2008-09-01

Hypercalciuria plays an important causal role in many patients with calcium oxalate (CaOx) stones. The source of the hypercalciuria includes increased intestinal Ca absorption and decreased renal tubule Ca reabsorption. In CaOx stone formers with idiopathic hypercalciuria (IH), Ca metabolic balance studies have revealed negative Ca balance and persistent hypercalciuria in the fasting state and during low dietary Ca intake. Bone resorption may also contribute to the high urine Ca excretion and increase the risk of bone loss. Indeed, low bone mass by DEXA scanning has been discovered in many IH patients. Thiazide diuretic agents reduce urine Ca excretion and may increase bone mineral density (BMD), thereby reducing fracture risk. Dietary Ca restriction that has been used unsuccessfully in the treatment of CaOx nephrolithiasis in the past may enhance negative Ca balance and accelerate bone loss. DEXA scans may demonstrate low BMD at the spine, hip, or forearm, with no predictable pattern. The unique pattern of bone histologic changes in IH differs from other causes of low DEXA bone density including postmenopausal osteoporosis, male hypogonadal osteoporosis, and glucocorticoid-induced osteoporosis. Hypercalciuria appears to play an important pathologic role in the development of low bone mass, and therefore correction of urine Ca losses should be a primary target for treatment of the bone disease accompanying IH.

Enhanced accumulation of adipocytes in bone marrow stromal cells in the presence of increased extracellular and intracellular [Ca{sup 2+}

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hashimoto, Ryota, E-mail: hryota@juntendo.ac.jp; Katoh, Youichi, E-mail: katoyo@juntendo-urayasu.jp; Nakamura, Kyoko

2012-07-13

Highlights: Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} enhances adipocyte accumulation in the presence of adipogenic inducers. Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} enhances both proliferation and adipocyte differentiation in BMSCs. Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub o} in BMSCs. Black-Right-Pointing-Pointer An intracellular Ca{sup 2+} chelator suppresses the enhancement in adipocyte accumulation. Black-Right-Pointing-Pointer Controlling [Ca{sup 2+}]{sub o} may govern the balance of adipocyte and osteoblast development. -- Abstract: The bone marrow stroma contains osteoblasts and adipocytes that have a common precursor: the pluripotent mesenchymal stem cell found in bone marrow stromal cells (BMSCs). Local bone marrow Ca{sup 2+}more » levels can reach high concentrations due to bone resorption, which is one of the notable features of the bone marrow stroma. Here, we describe the effects of high [Ca{sup 2+}]{sub o} on the accumulation of adipocytes in the bone marrow stroma. Using primary mouse BMSCs, we evaluated the level of adipocyte accumulation by measuring Oil Red O staining and glycerol-3-phosphate dehydrogenase (GPDH) activity. High [Ca{sup 2+}]{sub o} enhanced the accumulation of adipocytes following treatment with both insulin and dexamethasone together but not in the absence of this treatment. This enhanced accumulation was the result of both the accelerated proliferation of BMSCs and their differentiation into adipocytes. Using the fura-2 method, we also showed that high [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub i}. An intracellular Ca{sup 2+} chelator suppressed the enhancement in adipocyte accumulation due to increased [Ca{sup 2+}]{sub o} in BMSCs. These data suggest a new role for extracellular Ca{sup 2+} in the bone marrow stroma: increased [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub i} levels, which in turn enhances the accumulation of adipocytes under certain conditions.« less

Generation of novel bone forming cells (monoosteophils) from the cathelicidin-derived peptide LL-37 treated monocytes.

PubMed

Zhang, Zhifang; Shively, John E

2010-11-15

Bone generation and maintenance involve osteoblasts, osteoclasts, and osteocytes which originate from unique precursors and rely on key growth factors for differentiation. However, an incomplete understanding of bone forming cells during wound healing has led to an unfilled clinical need such as nonunion of bone fractures. Since circulating monocytes are often recruited to sites of injury and may differentiate into various cell types including osteoclasts, we investigated the possibility that circulating monocytes in the context of tissue injury may also contribute to bone repair. In particular, we hypothesized that LL-37 (produced from hCAP-18, cathelicidin), which recruits circulating monocytes during injury, may play a role in bone repair. Treatment of monocytes from blood with LL-37 for 6 days resulted in their differentiation to large adherent cells. Growth of LL-37-differentiated monocytes on osteologic discs reveals bone-like nodule formation by scanning electron microscopy (SEM). In vivo transplantation studies in NOD/SCID mice show that LL-37-differentiated monocytes form bone-like structures similar to endochondral bone formation. Importantly, LL-37-differentiated monocytes are distinct from conventional monocyte-derived osteoclasts, macrophages, and dendritic cells and do not express markers of the mesenchymal stem cells (MSC) lineage, distinguishing them from the conventional precursors of osteoblasts. Furthermore, LL-37 differentiated monocytes express intracellular proteins of both the osteoblast and osteoclast lineage including osteocalcin (OC), osteonectin (ON), bone sialoprotein II (BSP II), osteopontin (OP), RANK, RANKL, MMP-9, tartrate resistant acid phosphatase (TRAP), and cathepsin K (CK). Blood derived monocytes treated with LL-37 can be differentiated into a novel bone forming cell that functions both in vitro and in vivo. We propose the name monoosteophil to indicate their monocyte derived lineage and their bone forming phenotype. These cells may have wide ranging implications in the clinic including repair of broken bones and treatment of osteoporosis.

Anabolic activity of ursolic acid in bone: Stimulating osteoblast differentiation in vitro and inducing new bone formation in vivo.

PubMed

Lee, Su-Ui; Park, Sang-Joon; Kwak, Han Bok; Oh, Jaemin; Min, Yong Ki; Kim, Seong Hwan

2008-01-01

In the field of osteoporosis, there has been growing interest in anabolic agents that enhance bone mass and improve bone architecture. In this study, we demonstrated that the ubiquitous plant triterpenoid, ursolic acid, enhances differentiation and mineralization of osteoblasts in vitro. We found that ursolic acid induced the expression of osteoblast-specific genes with the activation of mitogen-activated protein kinases, nuclear factor-kappaB, and activator protein-1. Additionally, noggin, an antagonist of bone morphogenetic proteins (BMPs), inhibited ursolic acid-induced osteoblast differentiation. Noggin also inhibited the activation of Smad and the induction of BMP-2 mRNA expression by ursolic acid in the late stage of osteoblast differentiation. Importantly, ursolic acid was shown to have bone-forming activity in vivo in a mouse calvarial bone formation model. A high proportion of positive immunostaining of BMP-2 was found in the nuclear region of woven bone formed by ursolic acid. These results suggested that ursolic acid has the anabolic potential to stimulate osteoblast differentiation and enhance new bone formation.

Evaluation of Bone Marrow Processing Protocol for Therapeutic Applications via Culture and Characterization of Mesenchymal Stem Cells.

PubMed

Verma, Poonam; Bansal, Himanshu; Agrawal, Anupama; Leon, Jerry; Sundell, I Birgitta; Koka, Prasad S

Human mesenchymal stem cells from bone marrow (hMSCs) have broad therapeutic potential. These cells can be are readily isolated from bone marrow by their property to adhere to tissue culture treated culture wares. However, the proliferation rates and other properties of the cells gradually change during expansion. This study aims to validate the protocol of isolation and differentiation of hMSCs from bone marrow for therapeutic applications. Sixty ml of bone marrow was extracted from 5 patients and MSCs were isolated. These were characterized by Flow Cytometry, CFU assay and were differentiated into bone, fat cells and neurocytes. The cells were having healthy morphology. These were positive for the markers CD105, CD90 and CD73 and negative for CD45, CD34 and HLA-DR. The cells could differentiate into fat, bone and neural cells. MSCs from the bone marrow were isolated and differentiated. These cells were morphologically healthy and passed CFU assay. The cells exhibited differentiation potential into bone, fat and neural tissue. These cells can be used in therapeutic applications.

On the role of subtype selective adenosine receptor agonists during proliferation and osteogenic differentiation of human primary bone marrow stromal cells.

PubMed

Costa, M Adelina; Barbosa, A; Neto, E; Sá-e-Sousa, A; Freitas, R; Neves, J M; Magalhães-Cardoso, T; Ferreirinha, F; Correia-de-Sá, P

2011-05-01

Purines are important modulators of bone cell biology. ATP is metabolized into adenosine by human primary osteoblast cells (HPOC); due to very low activity of adenosine deaminase, the nucleoside is the end product of the ecto-nucleotidase cascade. We, therefore, investigated the expression and function of adenosine receptor subtypes (A(1) , A(2A) , A(2B) , and A(3) ) during proliferation and osteogenic differentiation of HPOC. Adenosine A(1) (CPA), A(2A) (CGS21680C), A(2B) (NECA), and A(3) (2-Cl-IB-MECA) receptor agonists concentration-dependently increased HPOC proliferation. Agonist-induced HPOC proliferation was prevented by their selective antagonists, DPCPX, SCH442416, PSB603, and MRS1191. CPA and NECA facilitated osteogenic differentiation measured by increases in alkaline phosphatase (ALP) activity. This contrasts with the effect of CGS21680C which delayed HPOC differentiation; 2-Cl-IB-MECA was devoid of effect. Blockade of the A(2B) receptor with PSB603 prevented osteogenic differentiation by NECA. In the presence of the A(1) antagonist, DPCPX, CPA reduced ALP activity at 21 and 28 days in culture. At the same time points, blockade of A(2A) receptors with SCH442416 transformed the inhibitory effect of CGS21680C into facilitation. Inhibition of adenosine uptake with dipyridamole caused a net increase in osteogenic differentiation. The presence of all subtypes of adenosine receptors on HPOC was confirmed by immunocytochemistry. Data show that adenosine is an important regulator of osteogenic cell differentiation through the activation of subtype-specific receptors. The most abundant A(2B) receptor seems to have a consistent role in cell differentiation, which may be balanced through the relative strengths of A(1) or A(2A) receptors determining whether osteoblasts are driven into proliferation or differentiation. Copyright © 2010 Wiley-Liss, Inc.

The Central Nervous System and Bone Metabolism: An Evolving Story.

PubMed

Dimitri, Paul; Rosen, Cliff

2017-05-01

Our understanding of the control of skeletal metabolism has undergone a dynamic shift in the last two decades, primarily driven by our understanding of energy metabolism. Evidence demonstrating that leptin not only influences bone cells directly, but that it also plays a pivotal role in controlling bone mass centrally, opened up an investigative process that has changed the way in which skeletal metabolism is now perceived. Other central regulators of bone metabolism have since been identified including neuropeptide Y (NPY), serotonin, endocannabinoids, cocaine- and amphetamine-regulated transcript (CART), adiponectin, melatonin and neuromedin U, controlling osteoblast and osteoclast differentiation, proliferation and function. The sympathetic nervous system was originally identified as the predominant efferent pathway mediating central signalling to control skeleton metabolism, in part regulated through circadian genes. More recent evidence points to a role of the parasympathetic nervous system in the control of skeletal metabolism either through muscarinic influence of sympathetic nerves in the brain or directly via nicotinic receptors on osteoclasts, thus providing evidence for broader autonomic skeletal regulation. Sensory innervation of bone has also received focus again widening our understanding of the complex neuronal regulation of bone mass. Whilst scientific advance in this field of bone metabolism has been rapid, progress is still required to understand how these model systems work in relation to the multiple confounders influencing skeletal metabolism, and the relative balance in these neuronal systems required for skeletal growth and development in childhood and maintaining skeletal integrity in adulthood.

The Rho-GEF Kalirin regulates bone mass and the function of osteoblasts and osteoclasts.

PubMed

Huang, Su; Eleniste, Pierre P; Wayakanon, Kornchanok; Mandela, Prashant; Eipper, Betty A; Mains, Richard E; Allen, Matthew R; Bruzzaniti, Angela

2014-03-01

Bone homeostasis is maintained by the balance between bone resorption by osteoclasts and bone formation by osteoblasts. Dysregulation in the activity of the bone cells can lead to osteoporosis, a disease characterized by low bone mass and an increase in bone fragility and risk of fracture. Kalirin is a novel GTP-exchange factor protein that has been shown to play a role in cytoskeletal remodeling and dendritic spine formation in neurons. We examined Kalirin expression in skeletal tissue and found that it was expressed in osteoclasts and osteoblasts. Furthermore, micro-CT analyses of the distal femur of global Kalirin knockout (Kal-KO) mice revealed significantly reduced trabecular and cortical bone parameters in Kal-KO mice, compared to WT mice, with significantly reduced bone mass in 8, 14 and 36week-old female Kal-KO mice. Male mice also exhibited a decrease in bone parameters but not to the level seen in female mice. Histomorphometric analyses also revealed decreased bone formation rate in 14week-old female Kal-KO mice, as well as decreased osteoblast number/bone surface and increased osteoclast surface/bone surface. Consistent with our in vivo findings, the bone resorbing activity and differentiation of Kal-KO osteoclasts was increased in vitro. Although alkaline phosphatase activity by Kal-KO osteoblasts was increased in vitro, Kal-KO osteoblasts showed decreased mineralizing activity, as well as decreased secretion of OPG, which was inversely correlated with ERK activity. Taken together, our findings suggest that deletion of Kalirin directly affects osteoclast and osteoblast activity, leading to decreased OPG secretion by osteoblasts which is likely to alter the RANKL/OPG ratio and promote osteoclastogenesis. Therefore, Kalirin may play a role in paracrine and/or endocrine signaling events that control skeletal bone remodeling and the maintenance of bone mass. Copyright © 2013 Elsevier Inc. All rights reserved.

Osteogenically differentiated mesenchymal stem cells and ceramics for bone tissue engineering.

PubMed

Ohgushi, Hajime

2014-02-01

In the human body, cells having self-renewal and multi-differentiation capabilities reside in many tissues and are called adult stem cells. In bone marrow tissue, two types of stem cells are well known: hematopoietic stem cells and mesenchymal stem cells (MSCs). Though the number of MSCs in bone marrow tissue is very low, it can be increased by in vitro culture of the marrow, and culture-expanded MSCs are available for various tissue regeneration. The culture-expanded MSCs can further differentiate into osteogenic cells such as bone forming osteoblasts by culturing the MSCs in an osteogenic medium. This paper discusses osteogenically differentiated MSCs derived from the bone marrow of patients. Importantly, the differentiation can be achieved on ceramic surfaces which demonstrate mineralized bone matrix formation as well as appearance of osteogenic cells. The cell/matrix/ceramic constructs could show immediate in vivo bone formation and are available for bone reconstruction surgery. Currently, MSCs are clinically available for the regeneration of various tissues due to their high proliferation/differentiation capabilities. However, the capabilities are still limited and thus technologies to improve or recover the inherent capabilities of MSCs are needed.

Utility of unenhanced fat-suppressed T1-weighted MRI in children with sickle cell disease -- can it differentiate bone infarcts from acute osteomyelitis?

PubMed

Delgado, Jorge; Bedoya, Maria A; Green, Abby M; Jaramillo, Diego; Ho-Fung, Victor

2015-12-01

Children with sickle cell disease (SCD) are at risk of bone infarcts and acute osteomyelitis. The clinical differentiation between a bone infarct and acute osteomyelitis is a diagnostic challenge. Unenhanced T1-W fat-saturated MR images have been proposed as a potential tool to differentiate bone infarcts from osteomyelitis. To evaluate the reliability of unenhanced T1-W fat-saturated MRI for differentiation between bone infarcts and acute osteomyelitis in children with SCD. We retrospectively reviewed the records of 31 children (20 boys, 11 girls; mean age 10.6 years, range 1.1-17.9 years) with SCD and acute bone pain who underwent MR imaging including unenhanced T1-W fat-saturated images from 2005 to 2010. Complete clinical charts were reviewed by a pediatric hematologist with training in infectious diseases to determine a clinical standard to define the presence or absence of osteomyelitis. A pediatric radiologist reviewed all MR imaging and was blinded to clinical information. Based on the signal intensity in T1-W fat-saturated images, the children were further classified as positive for osteomyelitis (low bone marrow signal intensity) or positive for bone infarct (high bone marrow signal intensity). Based on the clinical standard, 5 children were classified as positive for osteomyelitis and 26 children as positive for bone infarct (negative for osteomyelitis). The bone marrow signal intensity on T1-W fat-saturated imaging was not significant for the differentiation between bone infarct and osteomyelitis (P = 0.56). None of the additional evaluated imaging parameters on unenhanced MRI proved reliable in differentiating these diagnoses. The bone marrow signal intensity on unenhanced T1-W fat-saturated MR images is not a reliable criterion to differentiate bone infarcts from osteomyelitis in children.

Novel curcumin analogue UBS109 potently stimulates osteoblastogenesis and suppresses osteoclastogenesis: involvement in Smad activation and NF-κB inhibition.

PubMed

Yamaguchi, Masayoshi; Moore, Terry W; Sun, Aiming; Snyder, James P; Shoji, Mamoru

2012-08-01

Bone homeostasis is maintained through a balance between osteoblastic bone formation and osteoclastic bone resorption. Bone loss is induced due to decreased osteoblastic bone formation and increased osteoclastic bone resorption with various pathologic states. Osteoporosis with its accompanying decrease in bone mass is widely recognized as a major public health problem. Pharmacologic and functional food factors may play a role in the prevention of bone loss with aging. This study was undertaken to determine the effect of curcumin analogues (curcumin, EF31, ECMN909, and UBS109), which were newly synthesized, on osteoblastogenesis and osteoclastogenesis in vitro. Among these compounds, UBS109 had a unique stimulatory effect on osteoblastic differentiation and mineralization. UBS109 stimulated both basal and bone morphogenic protein-2 (BMP2)-increased Smad-luciferase activity, the Smad signaling of which is related to osteoblastogenesis. Such an effect was not seen with other compounds. Moreover, UBS109 potently suppressed tumor necrosis factor-α (TNF-α)-increased osteoblastic nuclear factor kappa B (NF-κB)-luciferase activity. In addition, EF31, ECMN909, and UBS109 had a suppressive effect on osteoclastogenesis as compared with that of curcumin. ECMN909 and UBS109 potently inhibited the receptor activator of NF-κB (RANK) ligand (RANKL)-increased preosteoclastic NF-κB-luciferase activity, in which NF-κB signaling plays a pivotal role in osteoclastogenesis. In the present study, curcumin analogue UBS109 was found to have a stimulating effect on osteoblastogenesis and a suppressive effect on osteoclastogenesis in vitro, suggesting an anabolic effect of the compound on bone mass.

Photobiomodulation of mesenchymal stem cells encapsulated in an injectable rhBMP4-loaded hydrogel directs hard tissue bioengineering.

PubMed

Diniz, Ivana M A; Carreira, Ana C O; Sipert, Carla R; Uehara, Cindi M; Moreira, Maria S N; Freire, Laila; Pelissari, Cibele; Kossugue, Patrícia M; de Araújo, Daniele R; Sogayar, Mari C; Marques, Márcia M

2018-06-01

Photobiomodulation (PBM) therapy displays relevant properties for tissue healing and regeneration, which may be of interest for the tissue engineering field. Here, we show that PBM is able to improve cell survival and to interact with recombinant human Bone Morphogenetic Protein 4 (rhBMP4) to direct and accelerate odonto/osteogenic differentiation of dental derived mesenchymal stem cells (MSCs). MSCs were encapsulated in an injectable and thermo-responsive cell carrier (Pluronic ® F-127) loaded with rhBMP4 and then photoactivated. PBM improved MSCs self-renewal and survival upon encapsulation in the Pluronic ® F-127. In the presence of rhBMP4, cell odonto/osteogenic differentiation was premature and markedly improved in the photoactivated MSCs. An in vivo calvarial critical sized defect model demonstrated significant increase in bone formation after PBM treatment. Finally, a balance in the reactive oxygen species levels may be related to the favorable results of PBM and rhBMP4 association. PBM may act in synergism with rhBMP4 and is a promise candidate to direct and accelerate hard tissue bioengineering. © 2017 Wiley Periodicals, Inc.

Mitochondrial Complex I Activity Suppresses Inflammation and Enhances Bone Resorption by Tipping the Balance of Macrophage-Osteoclast Polarization

PubMed Central

Jin, Zixue; Wei, Wei; Yang, Marie; Du, Yang; Wan, Yihong

2014-01-01

SUMMARY Mitochondrial complex I (CI) deficiency is associated with multiple neurological and metabolic disorders. However, its effect on innate immunity and bone remodeling is unclear. Using deletion of the essential CI subunit Ndufs4 as a model for mitochondrial dysfunction, we report that mitochondria suppress macrophage activation and inflammation while promoting osteoclast differentiation and bone resorption via both cell-autonomous and systemic regulation. Global Ndufs4 deletion causes systemic inflammation and osteopetrosis. Hematopoietic Ndufs4 deletion causes an intrinsic lineage shift from osteoclast to macrophage. Liver Ndufs4 deletion causes a metabolic shift from fatty acid oxidation to glycolysis, accumulating fatty acids and lactate (FA/LAC) in circulation. FA/LAC further activates Ndufs4−/− macrophages via ROS induction, and diminishes osteoclast lineage commitment in Ndufs4−/− progenitors; both inflammation and osteopetrosis in Ndufs4−/− mice are attenuated by TLR4/2 deletion. Together, these findings reveal mitochondrial CI as a critical rheostat of innate immunity and skeletal homeostasis. PMID:25130399

Activation of Hedgehog signaling by loss of GNAS causes heterotopic ossification

PubMed Central

Regard, Jean B.; Malhotra, Deepti; Gvozdenovic-Jeremic, Jelena; Josey, Michelle; Chen, Min; Weinstein, Lee S.; Lu, Jianming; Shore, Eileen M.; Kaplan, Frederick S.; Yang, Yingzi

2014-01-01

Bone formation is exquisitely controlled in space and time. Heterotopic ossification (HO), the pathologic formation of extra-skeletal bone, occurs as a common complication of trauma or in genetic disorders and can be disabling and lethal. However, the underlying molecular mechanisms are largely unknown. Here we demonstrate that Gαs restricts bone formation to the skeleton by inhibiting Hedgehog (Hh) signaling in mesenchymal progenitor cells. In progressive osseous heteroplasia (POH), a human disease caused by null mutations in GNAS that encodes Gαs, HH signaling is upregulated in ectopic osteoblasts and progenitor cells. Ectopic Hh signaling is sufficient to induce HO, while Hh signaling inhibition blocks HO in animal models. As our previous work has shown that GNAS gain of function mutations upregulate WNT/β-Catenin signaling in fibrous dysplasia (FD), our findings identify Gαs as a critical regulator of osteoblast differentiation by maintaining a balance between two key signaling pathways: Wnt/β-catenin and Hh. HH signaling inhibitors developed for cancer therapy may be repurposed to treat HO and other diseases caused by GNAS inactivation. PMID:24076664

Dasatinib inhibits both osteoclast activation and prostate cancer PC-3-cell-induced osteoclast formation.

PubMed

Araujo, John C; Poblenz, Ann; Corn, Paul; Parikh, Nila U; Starbuck, Michael W; Thompson, Jerry T; Lee, Francis; Logothetis, Christopher J; Darnay, Bryant G

2009-11-01

Therapies to target prostate cancer bone metastases have only limited effects. New treatments are focused on the interaction between cancer cells, bone marrow cells and the bone matrix. Osteoclasts play an important role in the development of bone tumors caused by prostate cancer. Since Src kinase has been shown to be necessary for osteoclast function, we hypothesized that dasatinib, a Src family kinase inhibitor, would reduce osteoclast activity and prostate cancer (PC-3) cell-induced osteoclast formation. Dasatinib inhibited RANKL-induced osteoclast differentiation of bone marrow-derived monocytes with an EC(50) of 7.5 nM. PC-3 cells, a human prostate cancer cell line, were able to differentiate RAW 264.7 cells, a murine monocytic cell line, into osteoclasts, and dasatinib inhibited this differentiation. In addition, conditioned medium from PC-3 cell cultures was able to differentiate RAW 264.7 cells into osteoclasts and this too, was inhibited by dasatinib. Even the lowest concentration of dasatinib, 1.25 nmol, inhibited osteoclast differentiation by 29%. Moreover, dasatinib inhibited osteoclast activity by 58% as measured by collagen 1 release. We performed in vitro experiments utilizing the Src family kinase inhibitor dasatinib to target osteoclast activation as a means of inhibiting prostate cancer bone metastases. Dasatinib inhibits osteoclast differentiation of mouse primary bone marrow-derived monocytes and PC-3 cell-induced osteoclast differentiation. Dasatinib also inhibits osteoclast degradation activity. Inhibiting osteoclast differentiation and activity may be an effective targeted therapy in patients with prostate cancer bone metastases.

Rapidly Assessing Changes in Bone Mineral Balance Using Natural Stable Calcium Isotopes

NASA Technical Reports Server (NTRS)

Morgan, J. L. L.; Gordon, G. W.; Romaniello, S. J.; Skulan, J. L.; Smith, S. M.; Anbar, A. D.

2011-01-01

We demonstrate that variations in the Ca isotope ratios in urine rapidly and quantitatively reflect changes in bone mineral balance. This variation occurs because bone formation depletes soft tissue of light Ca isotopes, while bone resorption releases that isotopically light Ca back into soft tissue. In a study of 12 individuals confined to bed rest, a condition known to induce bone resorption, we show that Ca isotope ratios shift in a direction consistent with net bone loss after just 7 days, long before detectible changes in bone density occur. Consistent with this interpretation, the Ca isotope variations track changes observed in N-teleopeptide, a bone resorption biomarker, while bone-specific alkaline phosphatase, a bone formation biomarker, is unchanged. Ca isotopes can in principle be used to quantify net changes in bone mass. Ca isotopes indicate an average loss of 0.62 +/- 0.16 % in bone mass over the course of this 30-day study. The Ca isotope technique should accelerate the pace of discovery of new treatments for bone disease and provide novel insights into the dynamics of bone metabolism.

Synergetic effect of topological cue and periodic mechanical tension-stress on osteogenic differentiation of rat bone mesenchymal stem cells.

PubMed

Liu, Yao; Yang, Guang; Ji, Huanzhong; Xiang, Tao; Luo, En; Zhou, Shaobing

2017-06-01

Mesenchymal stem cells (MSCs) are able to self-renew and differentiate into tissues of mesenchymal origin, making them to be significant for cell-based therapies, such as metabolic bone diseases and bone repair. Regulating the differentiation of MSCs is significant for bone regeneration. Electrospun fibers mimicking natural extracellular matrix (ECM), is an effective artificial ECM to regulate the behaviors and fates of MSCs. The aligned electrospun fibers can modulate polar cell pattern of bone mesenchymal stem cells, which leads to more obvious osteogenic differentiation. Apart from the topographic effect of electrospun fibers, mechanical cues can also intervene the cell behaviors. In this study, the osteogenic differentiation of rat bone mesenchymal stem cells was evaluated, which were cultured on aligned/random electrospun fiber mats materials under mechanical tension intervention. Scanning electron microscope and immune-fluorescent staining were used to directly observe the polarity changing of cellular morphology and cytoskeleton. The results proved that aligned electrospun fibers could be more conducive to promote osteogenic differentiation of rat bone mesenchymal stem cells and this promotion of osteogenic differentiation was enhanced by tension intervention. These results were correlated to the quantitative real-time PCR assay. In general, culturing rat bone mesenchymal stem cells on electrospun fibers under the intervention of mechanical tension is an effective way to mimic a more real cellular microenvironment. Copyright © 2017 Elsevier B.V. All rights reserved.

Hepatocyte growth factor improves bone regeneration via the bone morphogenetic protein‑2‑mediated NF‑κB signaling pathway.

PubMed

Zhen, Ruixin; Yang, Jianing; Wang, Yu; Li, Yubo; Chen, Bin; Song, Youxin; Ma, Guiyun; Yang, Bo

2018-04-01

Bone regeneration is an important process associated with the treatment of osteonecrosis, which is caused by various factors. Hepatocyte growth factor (HGF) is an active biological factor that has multifunctional roles in cell biology, life sciences and clinical medicine. It has previously been suggested that bone morphogenetic protein (BMP)‑2 exerts beneficial roles in bone formation, repair and angiogenesis in the femoral head. The present study aimed to investigate the benefits and molecular mechanisms of HGF in bone regeneration. The viability of osteoblasts and osteoclasts were studied in vitro. In addition, the expression levels of tumor necrosis factor (TNF)‑α, monocyte chemotactic protein (MCP)‑1, interleukin (IL)‑1 and IL‑6 were detected in a mouse fracture model following treatment with HGF. The expression and activity of nuclear factor (NF)‑κB were also analyzed in osteocytes post‑treatment with HGF. Histological analysis was used to determine the therapeutic effects of HGF on mice with fractures. The migration and differentiation of osteoblasts and osteoclasts were investigated in HGF‑incubated cells. Furthermore, angiogenesis and BMP‑2 expression were analyzed in the mouse fracture model post‑treatment with HGF. The results indicated that HGF regulates the cell viability of osteoblasts and osteoclasts, and also balanced the ratio between osteoblasts and osteoclasts. In addition, HGF decreased the serum expression levels of TNF‑α, MCP‑1, IL‑1 and IL‑6 in experimental mice. The results of a mechanistic analysis demonstrated that HGF upregulated p65, IκB kinase‑β and IκBα expression in osteoblasts from experimental mice. In addition, the expression levels of vascular endothelial growth factor, BMP‑2 receptor, receptor activator of NF‑κB ligand and macrophage colony‑stimulating factor were upregulated by HGF, which may effectively promote blood vessel regeneration, and contribute to the formation and revascularization of tissue‑engineered bone. Furthermore, HGF promoted BMP‑2 expression and enhanced angiogenesis at the fracture location. These results suggested that HGF treatment may significantly promote bone regeneration in a mouse fracture model. In conclusion, these results indicated that HGF is involved in bone regeneration, angiogenesis and the balance between osteoblasts and osteoclasts, thus suggesting that HGF may be considered a potential agent for the treatment of fractures via the promotion of bone regeneration through regulation of the BMP‑2‑mediated NF‑κB signaling pathway.

Ablation of the Sam68 RNA Binding Protein Protects Mice from Age-Related Bone Loss

PubMed Central

Richard, Stéphane; Torabi, Nazi; Franco, Gladys Valverde; Tremblay, Guy A; Chen, Taiping; Vogel, Gillian; Morel, Mélanie; Cléroux, Patrick; Forget-Richard, Alexandre; Komarova, Svetlana; Tremblay, Michel L; Li, Wei; Li, Ailian; Gao, Yun Jing; Henderson, Janet E

2005-01-01

The Src substrate associated in mitosis of 68 kDa (Sam68) is a KH-type RNA binding protein that has been shown to regulate several aspects of RNA metabolism; however, its physiologic role has remained elusive. Herein we report the generation of Sam68-null mice by homologous recombination. Aged Sam68−/− mice preserved their bone mass, in sharp contrast with 12-month-old wild-type littermates in which bone mass was decreased up to approximately 75%. In fact, the bone volume of the 12-month-old Sam68−/− mice was virtually indistinguishable from that of 4-month-old wild-type or Sam68−/− mice. Sam68−/− bone marrow stromal cells had a differentiation advantage for the osteogenic pathway. Moreover, the knockdown of Sam68 using short hairpin RNA in the embryonic mesenchymal multipotential progenitor C3H10T1/2 cells resulted in more pronounced expression of the mature osteoblast marker osteocalcin when differentiation was induced with bone morphogenetic protein-2. Cultures of mouse embryo fibroblasts generated from Sam68+/+ and Sam68−/− littermates were induced to differentiate into adipocytes with culture medium containing pioglitazone and the Sam68−/− mouse embryo fibroblasts shown to have impaired adipocyte differentiation. Furthermore, in vivo it was shown that sections of bone from 12-month-old Sam68−/− mice had few marrow adipocytes compared with their age-matched wild-type littermate controls, which exhibited fatty bone marrow. Our findings identify endogenous Sam68 as a positive regulator of adipocyte differentiation and a negative regulator of osteoblast differentiation, which is consistent with Sam68 being a modulator of bone marrow mesenchymal cell differentiation, and hence bone metabolism, in aged mice. PMID:16362077

Differentiating human bone from animal bone: a review of histological methods.

PubMed

Hillier, Maria L; Bell, Lynne S

2007-03-01

This review brings together a complex and extensive literature to address the question of whether it is possible to distinguish human from nonhuman bone using the histological appearance of cortical bone. The mammalian species included are rat, hare, badger, racoon dog, cat, dog, pig, cow, goat, sheep, deer, horse, water buffalo, bear, nonhuman primates, and human and are therefore not exhaustive, but cover those mammals that may contribute to a North American or Eurasian forensic assemblage. The review has demonstrated that differentiation of human from certain nonhuman species is possible, including small mammals exhibiting Haversian bone tissue and large mammals exhibiting plexiform bone tissue. Pig, cow, goat, sheep, horse, and water buffalo exhibit both plexiform and Haversian bone tissue and where only Haversian bone tissue exists in bone fragments, differentiation of these species from humans is not possible. Other primate Haversian bone tissue is also not distinguishable from humans. Where differentiation using Haversian bone tissue is undertaken, both the general microstructural appearance and measurements of histological structures should be applied. Haversian system diameter and Haversian canal diameter are the most optimal and diagnostic measurements to use. Haversian system density may be usefully applied to provide an upper and lower limit for humans.

Mechanical stress regulates osteogenic differentiation and RANKL/OPG ratio in periodontal ligament stem cells by the Wnt/β-catenin pathway.

PubMed

Zhang, Liqiang; Liu, Wenjia; Zhao, Jiangdong; Ma, Xiaojie; Shen, Lin; Zhang, Yongjie; Jin, Fang; Jin, Yan

2016-10-01

The balance between osteoblastic and osteoclastic activity is critical in orthodontic tooth movement (OTM). Mesenchymal stem cells (MSCs) play an important role in maintaining bone homeostasis, and periodontal ligament stem cells (PDLSCs) are tissue-specific MSCs in the periodontal ligament. However, whether PDLSCs are required for periodontal tissue remodeling during OTM is not fully understood. Here, we used PDGFRα and Nestin to trace PDLSCs during OTM in rats. We treat human PDLSCs with 100kpa static pressure for 1h or 12h in vitro, and examined the phenotypic changes and expression of RANKL and OPG in these cells. In vivo, we found that positive signals of PDGFRα and Nestin in the PDL gradually increased and then decreased on the pressure side to which pressure was applied. In vitro, the osteogenic differentiation of PDLSCs was significantly increased after force treatment for 1h relative to 12h. In contrast, the expression ratio of RANKL/OPG was reduced at 1h and significantly increased at 12h. Furthermore, we found that the Wnt/β-catenin pathway was dynamically activated in the PDL and in PDLSCs after mechanical stimulation. Importantly, the canonical Wnt pathway inhibitor DKK1 blocked the osteogenesis effect and rescued the ratio of RANKL/OPG in PDLSCs under force treatment for 1h. Our findings reveal that PDLSCs participate in OTM and that the Wnt/β-catenin pathway maintains bone homeostasis during tooth movement by regulating the balance between osteoblastic and osteoclastic activity. We describe a novel potential mechanism related to tooth movement. Copyright © 2016 Elsevier B.V. All rights reserved.

Dasatinib inhibits both osteoclast activation and prostate cancer PC-3 cell-induced osteoclast formation

PubMed Central

Araujo, John C.; Poblenz, Ann; Corn, Paul G.; Parikh, Nila U.; Starbuck, Michael W.; Thompson, Jerry T.; Lee, Francis; Logothetis, Christopher J.; Darnay, Bryant G.

2013-01-01

Purpose Therapies to target prostate cancer bone metastases have only limited effects. New treatments are focused on the interaction between cancer cells, bone marrow cells and the bone matrix. Osteoclasts play an important role in the development of bone tumors caused by prostate cancer. Since Src kinase has been shown to be necessary for osteoclast function, we hypothesized that dasatinib, a Src family kinase inhibitor, would reduce osteoclast activity and prostate cancer (PC-3) cell-induced osteoclast formation. Results Dasatinib inhibited RANKL-induced osteoclast differentiation of bone marrow-derived monocytes with an EC50 of 7.5 nM. PC-3 cells, a human prostate cancer cell line, were able to differentiate RAW 264.7 cells, a murine monocytic cell line, into osteoclasts and dasatinib inhibited this differentiation. In addition, conditioned medium from PC-3 cell cultures was able to differentiate RAW 264.7 cells into osteoclasts and this too, was inhibited by dasatinib. Even the lowest concentration of dasatinib, 1.25 nmol, inhibited osteoclast differentiation by 29%. Moreover, dasatinib inhibited osteoclast activity by 58% as measured by collagen 1 release. Experimental design We performed in vitro experiments utilizing the Src family kinase inhibitor dasatinib to target osteoclast activation as a means of inhibiting prostate cancer bone metastases. Conclusion Dasatinib inhibits osteoclast differentiation of mouse primary bone marrow-derived monocytes and PC-3 cell-induced osteoclast differentiation. Dasatinib also inhibits osteoclast degradation activity. Inhibiting osteoclast differentiation and activity may be an effective targeted therapy in patients with prostate cancer bone metastases. PMID:19855158

Mechanisms in endocrinology: micro-RNAs: targets for enhancing osteoblast differentiation and bone formation.

PubMed

Taipaleenmäki, Hanna; Bjerre Hokland, Lea; Chen, Li; Kauppinen, Sakari; Kassem, Moustapha

2012-03-01

Osteoblast differentiation and bone formation (osteogenesis) are regulated by transcriptional and post-transcriptional mechanisms. Recently, a novel class of regulatory factors termed micro-RNAs (miRNAs) has been identified as playing an important role in the regulation of many aspects of osteoblast biology including proliferation, differentiation, metabolism and apoptosis. Also, preliminary data from animal disease models suggest that targeting miRNAs in bone can be a novel approach to increase bone mass. This review highlights the current knowledge of miRNA biology and their role in bone formation and discusses their potential use in future therapeutic applications for metabolic bone diseases.

Osteoblast and osteoclast responses to A/B type carbonate-substituted hydroxyapatite ceramics for bone regeneration.

PubMed

Germaini, Marie-Michèle; Detsch, Rainer; Grünewald, Alina; Magnaudeix, Amandine; Lalloue, Fabrice; Boccaccini, Aldo R; Champion, Eric

2017-06-06

The influence of carbonate substitution (4.4 wt%, mixed A/B type) in hydroxyapatite ceramics for bone remodeling scaffolds was investigated by separately analyzing the response of pre-osteoblasts and osteoclast-like cells. Carbonated hydroxyapatite (CHA) (Ca 9.5 (PO 4 ) 5.5 (CO 3 ) 0.5 (OH)(CO 3 ) 0.25 -CHA), mimicking the chemical composition of natural bone mineral, and pure hydroxyapatite (HA) (Ca 10 (PO 4 ) 6 (OH) 2 -HA) porous ceramics were processed to obtain a similar microstructure and surface physico-chemical properties (grain size, porosity ratio and pore size, surface roughness and zeta potential). The biological behavior was studied using MC3T3-E1 pre-osteoblastic and RAW 264.7 monocyte/macrophage cell lines. Chemical dissolution in the culture media and resorption lacunae produced by osteoclasts occur with both HA and CHA ceramics, but CHA exhibits much higher dissolution and greater bioresorption ability. CHA ceramics promoted a significantly higher level of pre-osteoblast proliferation. Osteoblastic differentiation, assessed by qRT-PCR of RUNX2 and COLIA2, and pre-osteoclastic proliferation and differentiation were not significantly different on CHA or HA ceramics but cell viability and metabolism were significantly greater on CHA ceramics. Thus, the activity of both osteoclast-like and osteoblastic cells was influenced by the carbonate substitution in the apatite structure. Furthermore, CHA showed a particularly interesting balance between biodegradation, by osteoclasts and chemical dissolution, and osteogenesis through osteoblasts' activity, to stimulate bone regeneration. It is hypothesized that this amount of 4.4 wt% carbonate substitution leads to an adapted concentration of calcium in the fluid surrounding the ceramic to stimulate the activity of cells. These results highlight the superior biological behavior of microporous 4.4 wt% A/B CHA ceramics that could beneficially replace the commonly used HA of biphasic calcium phosphates for future applications in bone tissue engineering.

Natural calcium isotonic composition of urine as a marker of bone mineral balance

USGS Publications Warehouse

Skulan, J.; Bullen, T.; Anbar, A.D.; Puzas, J.E.; Shackelford, L.; LeBlanc, A.; Smith, S.M.

2007-01-01

Background: We investigated whether changes in the natural isotopic composition of calcium in human urine track changes in net bone mineral balance, as predicted by a model of calcium isotopic behavior in vertebrates. If so, isotopic analysis of natural urine or blood calcium could be used to monitor short-term changes in bone mineral balance that cannot be detected with other techniques. Methods: Calcium isotopic compositions are expressed as ??44Ca, or the difference in parts per thousand between the 44Ca/40Ca of a sample and the 44Ca/ 40Ca of a standard reference material. ??44Ca was measured in urine samples from 10 persons who participated in a study of the effectiveness of countermeasures to bone loss in spaceflight, in which 17 weeks of bed rest was used to induce bone loss. Study participants were assigned to 1 of 3 treatment groups: controls received no treatment, one treatment group received alendronate, and another group performed resistive exercise. Measurements were made on urine samples collected before, at 2 or 3 points during, and after bed rest. Results: Urine ??44Ca values during bed rest were lower in controls than in individuals treated with alendronate (P <0.05, ANOVA) or exercise (P <0.05), and lower than the control group baseline (P <0.05, Mest). Results were consistent with the model and with biochemical and bone mineral density data. Conclusion: Results confirm the predicted relationship between bone mineral balance and calcium isotopes, suggesting that calcium isotopic analysis of urine might be refined into a clinical and research tool. ?? 2007 American Association for Clinical Chemistry.

Natural calcium isotopic composition of urine as a marker of bone mineral balance.

PubMed

Skulan, Joseph; Bullen, Thomas; Anbar, Ariel D; Puzas, J Edward; Shackelford, Linda; LeBlanc, Adrian; Smith, Scott M

2007-06-01

We investigated whether changes in the natural isotopic composition of calcium in human urine track changes in net bone mineral balance, as predicted by a model of calcium isotopic behavior in vertebrates. If so, isotopic analysis of natural urine or blood calcium could be used to monitor short-term changes in bone mineral balance that cannot be detected with other techniques. Calcium isotopic compositions are expressed as delta(44)Ca, or the difference in parts per thousand between the (44)Ca/(40)Ca of a sample and the (44)Ca/(40)Ca of a standard reference material. delta(44)Ca was measured in urine samples from 10 persons who participated in a study of the effectiveness of countermeasures to bone loss in spaceflight, in which 17 weeks of bed rest was used to induce bone loss. Study participants were assigned to 1 of 3 treatment groups: controls received no treatment, one treatment group received alendronate, and another group performed resistive exercise. Measurements were made on urine samples collected before, at 2 or 3 points during, and after bed rest. Urine delta(44)Ca values during bed rest were lower in controls than in individuals treated with alendronate (P <0.05, ANOVA) or exercise (P <0.05), and lower than the control group baseline (P <0.05, t-test). Results were consistent with the model and with biochemical and bone mineral density data. Results confirm the predicted relationship between bone mineral balance and calcium isotopes, suggesting that calcium isotopic analysis of urine might be refined into a clinical and research tool.

MRI differentiation of low-grade from high-grade appendicular chondrosarcoma.

PubMed

Douis, Hassan; Singh, Leanne; Saifuddin, Asif

2014-01-01

To identify magnetic resonance imaging (MRI) features which differentiate low-grade chondral lesions (atypical cartilaginous tumours/grade 1 chondrosarcoma) from high-grade chondrosarcomas (grade 2, grade 3 and dedifferentiated chondrosarcoma) of the major long bones. We identified all patients treated for central atypical cartilaginous tumours and central chondrosarcoma of major long bones (humerus, femur, tibia) over a 13-year period. The MRI studies were assessed for the following features: bone marrow oedema, soft tissue oedema, bone expansion, cortical thickening, cortical destruction, active periostitis, soft tissue mass and tumour length. The MRI-features were compared with the histopathological tumour grading using univariate, multivariate logistic regression and receiver operating characteristic curve (ROC) analyses. One hundred and seventy-nine tumours were included in this retrospective study. There were 28 atypical cartilaginous tumours, 79 grade 1 chondrosarcomas, 36 grade 2 chondrosarcomas, 13 grade 3 chondrosarcomas and 23 dedifferentiated chondrosarcomas. Multivariate analysis demonstrated that bone expansion (P = 0.001), active periostitis (P = 0.001), soft tissue mass (P < 0.001) and tumour length (P < 0.001) were statistically significant differentiating factors between low-grade and high-grade chondral lesions with an area under the ROC curve of 0.956. On MRI, bone expansion, active periostitis, soft tissue mass and tumour length can reliably differentiate high-grade chondrosarcomas from low-grade chondral lesions of the major long bones. • Accurate differentiation of low-grade from high-grade chondrosarcomas is essential before surgery • MRI can reliably differentiate high-grade from low-grade chondrosarcomas of long bone • Differentiating features are bone expansion, periostitis, soft tissue mass and tumour length • Presence of these four MRI features demonstrated a diagnostic accuracy (AUC) of 95.6 % • The findings may result in more accurate diagnosis before definitive surgery.

Influence of bone marrow-derived mesenchymal stem cells pre-implantation differentiation approach on periodontal regeneration in vivo.

PubMed

Cai, Xinjie; Yang, Fang; Yan, Xiangzhen; Yang, Wanxun; Yu, Na; Oortgiesen, Daniel A W; Wang, Yining; Jansen, John A; Walboomers, X Frank

2015-04-01

The implantation of bone marrow-derived mesenchymal stem cells (MSCs) has previously been shown successful to achieve periodontal regeneration. However, the preferred pre-implantation differentiation strategy (e.g. maintenance of stemness, osteogenic or chondrogenic induction) to obtain optimal periodontal regeneration is still unknown. This in vivo study explored which differentiation approach is most suitable for periodontal regeneration. Mesenchymal stem cells were obtained from Fischer rats and seeded onto poly(lactic-co-glycolic acid)/poly(ɛ-caprolactone) electrospun scaffolds, and then pre-cultured under different in vitro conditions: (i) retention of multilineage differentiation potential; (ii) osteogenic differentiation approach; and (iii) chondrogenic differentiation approach. Subsequently, the cell-scaffold constructs were implanted into experimental periodontal defects of Fischer rats, with empty scaffolds as controls. After 6 weeks of implantation, histomorphometrical analyses were applied to evaluate the regenerated periodontal tissues. The chondrogenic differentiation approach showed regeneration of alveolar bone and ligament tissues. The retention of multilineage differentiation potential supported only ligament regeneration, while the osteogenic differentiation approach boosted alveolar bone regeneration. Chondrogenic differentiation of MSCs before implantation is a useful strategy for regeneration of alveolar bone and periodontal ligament, in the currently used rat model. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Experimental variation of the level and the ratio of angiogenic and osteogenic signaling affects the spatiotemporal expression of bone-specific markers and organization of bone formation in ectopic sites.

PubMed

Moser, Norman; Goldstein, Jan; Kauffmann, Phillip; Epple, Matthias; Schliephake, Henning

2018-04-01

The aim of the present study was to test the hypothesis that the ratio of angiogenic and osteogenic signaling affects ectopic bone formation when delivered in different amounts. Porous composite PDLLA/CaCO 3 scaffolds were loaded with rhBMP2 and rhVEGF in different dosage combinations and implanted into the gluteal muscles of 120 adult male Wistar rats. Bone formation and expression of alkaline phosphatase and Runx2 were quantified by histomorphometry. Spatial distribution across the scaffolds was assessed by using a grid that discriminated between the periphery and center of the scaffolds. The evaluation showed that the combined delivery of bone morphogenetic protein BMP2 and VEGF in different dosage combinations did not enhance the overall quantity of ectopic bone formation compared to the delivery of BMP2 alone. The addition of VEGF generally upregulated Runx2 after 4 weeks, which may have retarded terminal osteogenic differentiation. However, slow combined delivery of 1.5-2.0 μg BMP2 combined with 50 ng VEGF165 over a period of 5 weeks supported a more even distribution of bone formation across the implanted scaffolds whereas higher amounts of VEGF did not elicit this effect. The findings suggest that structural organization rather than the quantity of ectopic bone formation is affected by the dosage and the ratio of BMP2 and VEGF levels at the observed intervals. The development of carriers for dual growth factor delivery has to take into account the necessity to carefully balance the ratio of growth release.

Isolation and hepatocyte differentiation of mesenchymal stem cells from porcine bone marrow--"surgical waste" as a novel MSC source.

PubMed

Brückner, S; Tautenhahn, H-M; Winkler, S; Stock, P; Jonas, S; Dollinger, M; Christ, B

2013-06-01

Mesenchymal stem cells (MSC) isolated from bone marrow and differentiated into hepatocyte-like cells have increasingly gained attention for clinical cell therapy of liver diseases because of their high regenerative capacity. They are available from bone marrow aspirates of the os coxae after puncture of the crista iliaca or from bone marrow "surgical waste" gained from amputations or knee and hip operations. Thus, the aim of the study was to demonstrate whether these pBM-MSC (porcine bone marrow-derived mesenchymal stem cells) displayed mesenchymal features and hepatocyte differentiation potential. MSC were isolated either from crista iliaca punctures or after sampling and collagenase digestion of bone marrow from the os femoris. Mesenchymal features were assessed by flow cytometry for specific surface antigens and their ability to differentiate into at least 3 lineages. Functional properties, such as urea or glycogen synthesis and cytochrome P450 activity, as well as the cell morphology were examined during hepatocyte differentiation. pBM-MSC from both sources lacked the hematopoietic markers CD14 and CD45 but expressed the typical mesenchymal markers CD44, CD29, CD90, and CD105. Both cell types could differentiate into adipocyte, osteocyte, and hepatocyte lineages. After hepatocyte differentiation, CD105 expression decreased significantly and cells changed morphology from fibroblastoid into polygonal, displaying significantly increased glycogen storage, urea synthesis, and cytochrome activity. pBM-MSC from various sources were identical in respect to their mesenchymal features and their hepatocyte differentiation potential. Hence, long bones might be a particularly useful resource to isolate bone marrow mesenchymal stem cells for transplantation. Copyright © 2013 Elsevier Inc. All rights reserved.

A Comparison between the Effects of Aerobic Dance Training on Mini-Trampoline and Hard Wooden Surface on Bone Resorption, Health-Related Physical Fitness, Balance, and Foot Plantar Pressure in Thai Working Women.

PubMed

Sukkeaw, Wittawat; Kritpet, Thanomwong; Bunyaratavej, Narong

2015-09-01

To compare the effects of aerobic dance training on mini-trampoline and hard wooden surface on bone resorption, health-related physical fitness, balance, and foot plantar pressure in Thai working women. Sixty-three volunteered females aged 35-45 years old participated in the study and were divided into 3 groups: A) aerobic dance on mini-trampoline (21 females), B) aerobic dance on hard wooden surface (21 females), and C) control group (21 females). All subjects in the aerobic dance groups wore heart rate monitors during exercise. Aerobic dance worked out 3 times a week, 40 minutes a day for 12 weeks. The intensity was set at 60-80% of the maximum heart rate. The control group engaged in routine physical activity. The collected data were bone formation (N-terminal propeptine of procollagen type I: P1NP) bone resorption (Telopeptide cross linked: β-CrossLaps) health-related physical fitness, balance, and foot plantar pressure. The obtained data from pre- and post trainings were compared and analyzed by paired samples t-test and one way analysis of covariance. The significant difference was at 0.05 level. After the 12-week training, the biochemical bone markers of both mini-trampoline and hard wooden surface aerobic dance training subjects decreased in bone resorption (β-CrossLaps) but increased in boneformation (P1NP). Health-related physical fitness, balance, and foot plantar pressure were not only better when comparing to the pre-test result but also significantly different when comparing to the control group (p < 0.05). The aerobic dance on mini-trampoline showed that leg muscular strength, balance and foot plantar pressure were significantly better than the aerobic dance on hard wooden surface (p < 0.05). The aerobic dance on mini-trampoline and hard wooden surface had positive effects on biochemical bone markers. However, the aerobic dance on mini-trampoline had more leg muscular strength and balance including less foot plantar pressure. It is considered to be an appropriate exercise programs in working women.

Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold

PubMed Central

Feng, Yuping; Wang, Jiao; Ling, Shixin; Li, Zhuo; Li, Mingsheng; Li, Qiongyi; Ma, Zongren; Yu, Sijiu

2014-01-01

The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells following induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined specific neuronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuronal-specific proteins, including βIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differentiation medium differentiated into a multilayered neural network-like structure with long nerve fibers that was composed of several parallel microfibers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sectioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve. PMID:25598779

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tu Qisheng; Valverde, Paloma; Chen, Jake

Osterix (Osx) is a zinc-finger-containing transcription factor that is expressed in osteoblasts of all endochondral and membranous bones. In Osx null mice osteoblast differentiation is impaired and bone formation is absent. In this study, we hypothesized that overexpression of Osx in murine bone marrow stromal cells (BMSC) would be able to enhance their osteoblastic differentiation and mineralization in vitro. Retroviral transduction of Osx in BMSC cultured in non-differentiating medium did not affect expression of Runx2/Cbfa1, another key transcription factor of osteoblast differentiation, but induced an increase in the expression of other markers associated with the osteoblastic lineage including alkaline phosphatase,more » bone sialoprotein, osteocalcin, and osteopontin. Retroviral transduction of Osx in BMSC also increased their proliferation, alkaline phosphatase activity, and ability to form bone nodules. These events occurred without significant changes in the expression of {alpha}1(II) procollagen or lipoprotein lipase, which are markers of chondrogenic and adipogenic differentiation, respectively.« less

Effects of dietary bread crust Maillard reaction products on calcium and bone metabolism in rats.

PubMed

Roncero-Ramos, Irene; Delgado-Andrade, Cristina; Haro, Ana; Ruiz-Roca, Beatriz; Morales, Francisco J; Navarro, María Pilar

2013-06-01

Maillard reaction products (MRP) consumption has been related with the development of bone degenerative disorders, probably linked to changes in calcium metabolism. We aimed to investigate the effects of MRP intake from bread crust on calcium balance and its distribution, and bone metabolism. During 88 days, rats were fed control diet or diets containing bread crust as source of MRP, or its soluble high molecular weight, soluble low molecular weight or insoluble fractions (bread crust, HMW, LMW and insoluble diets, respectively). In the final week, a calcium balance was performed, then animals were sacrified and some organs removed to analyse calcium levels. A second balance was carried out throughout the experimental period to calculate global calcium retention. Biochemical parameters and bone metabolism markers were measured in serum or urine. Global calcium bioavailability was unmodified by consumption of bread crust or its isolate fractions, corroborating the previously described low affinity of MRP to bind calcium. Despite this, a higher calcium concentration was found in femur due to smaller bones having a lower relative density. The isolate consumption of the fractions altered some bone markers, reflecting a situation of increased bone resorption or higher turnover; this did not take place in the animals fed the bread crust diet. Thus, the bread crust intake does not affect negatively calcium bioavailability and bone metabolism.

Dexamethasone Enhances Osteogenic Differentiation of Bone Marrow- and Muscle-Derived Stromal Cells and Augments Ectopic Bone Formation Induced by Bone Morphogenetic Protein-2

PubMed Central

Yuasa, Masato; Yamada, Tsuyoshi; Taniyama, Takashi; Masaoka, Tomokazu; Xuetao, Wei; Yoshii, Toshitaka; Horie, Masaki; Yasuda, Hiroaki; Uemura, Toshimasa; Okawa, Atsushi; Sotome, Shinichi

2015-01-01

We evaluated whether dexamethasone augments the osteogenic capability of bone marrow-derived stromal cells (BMSCs) and muscle tissue-derived stromal cells (MuSCs), both of which are thought to contribute to ectopic bone formation induced by bone morphogenetic protein-2 (BMP-2), and determined the underlying mechanisms. Rat BMSCs and MuSCs were cultured in growth media with or without 10-7 M dexamethasone and then differentiated under osteogenic conditions with dexamethasone and BMP-2. The effects of dexamethasone on cell proliferation and osteogenic differentiation, and also on ectopic bone formation induced by BMP-2, were analyzed. Dexamethasone affected not only the proliferation rate but also the subpopulation composition of BMSCs and MuSCs, and subsequently augmented their osteogenic capacity during osteogenic differentiation. During osteogenic induction by BMP-2, dexamethasone also markedly affected cell proliferation in both BMSCs and MuSCs. In an in vivo ectopic bone formation model, bone formation in muscle-implanted scaffolds containing dexamethasone and BMP-2 was more than two fold higher than that in scaffolds containing BMP-2 alone. Our results suggest that dexamethasone potently enhances the osteogenic capability of BMP-2 and may thus decrease the quantity of BMP-2 required for clinical application, thereby reducing the complications caused by excessive doses of BMP-2. Highlights: 1. Dexamethasone induced selective proliferation of bone marrow- and muscle-derived cells with higher differentiation potential. 2. Dexamethasone enhanced the osteogenic capability of bone marrow- and muscle-derived cells by altering the subpopulation composition. 3. Dexamethasone augmented ectopic bone formation induced by bone morphogenetic protein-2. PMID:25659106

microRNAs as regulators of adipogenic differentiation of mesenchymal stem cells.

PubMed

Hamam, Dana; Ali, Dalia; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

2015-02-15

microRNAs (miRNAs) constitute complex regulatory network, fine tuning the expression of a myriad of genes involved in different biological and physiological processes, including stem cell differentiation. Mesenchymal stem cells (MSCs) are multipotent stem cells present in the bone marrow stroma, and the stroma of many other tissues, and can give rise to a number of mesoderm-type cells including adipocytes and osteoblasts, which form medullary fat and bone tissues, respectively. The role of bone marrow fat in bone mass homeostasis is an area of intensive investigation with the aim of developing novel approaches for enhancing osteoblastic bone formation through inhibition of bone marrow fat formation. A number of recent studies have reported several miRNAs that enhance or inhibit adipogenic differentiation of MSCs and with potential use in microRNA-based therapy to regulate adipogenesis in the context of treating bone diseases and metabolic disorders. The current review focuses on miRNAs and their role in regulating adipogenic differentiation of MSCs.

Activation of GLP-1 Receptor Promotes Bone Marrow Stromal Cell Osteogenic Differentiation through β-Catenin

PubMed Central

Meng, Jingru; Ma, Xue; Wang, Ning; Jia, Min; Bi, Long; Wang, Yunying; Li, Mingkai; Zhang, Huinan; Xue, Xiaoyan; Hou, Zheng; Zhou, Ying; Yu, Zhibin; He, Gonghao; Luo, Xiaoxing

2016-01-01

Summary Glucagon-like peptide 1 (GLP-1) plays an important role in regulating bone remodeling, and GLP-1 receptor agonist shows a positive relationship with osteoblast activity. However, GLP-1 receptor is not found in osteoblast, and the mechanism of GLP-1 receptor agonist on regulating bone remodeling is unclear. Here, we show that the GLP-1 receptor agonist exendin-4 (Ex-4) promoted bone formation and increased bone mass and quality in a rat unloading-induced bone loss model. These functions were accompanied by an increase in osteoblast number and serum bone formation markers, while the adipocyte number was decreased. Furthermore, GLP-1 receptor was detected in bone marrow stromal cells (BMSCs), but not in osteoblast. Activation of GLP-1 receptor by Ex-4 promoted the osteogenic differentiation and inhibited BMSC adipogenic differentiation through regulating PKA/β-catenin and PKA/PI3K/AKT/GSK3β signaling. These findings reveal that GLP-1 receptor regulates BMSC osteogenic differentiation and provide a molecular basis for therapeutic potential of GLP-1 against osteoporosis. PMID:26947974

Tridax procumbens flavonoids promote osteoblast differentiation and bone formation.

PubMed

Al Mamun, Md Abdullah; Hosen, Mohammad Jakir; Islam, Kamrul; Khatun, Amina; Alam, M Masihul; Al-Bari, Md Abdul Alim

2015-11-18

Tridax procumbens flavonoids (TPFs) are well known for their medicinal properties among local natives. Besides traditionally used for dropsy, anemia, arthritis, gout, asthma, ulcer, piles, and urinary problems, it is also used in treating gastric problems, body pain, and rheumatic pains of joints. TPFs have been reported to increase osteogenic functioning in mesenchymal stem cells. Our previous study showed that TPFs were significantly suppressed the RANKL-induced differentiation of osteoclasts and bone resorption. However, the effects of TPFs to promote osteoblasts differentiation and bone formation remain unclear. TPFs were isolated from Tridax procumbens and investigated for their effects on osteoblasts differentiation and bone formation by using primary mouse calvarial osteoblasts. TPFs promoted osteoblast differentiation in a dose-dependent manner demonstrated by up-regulation of alkaline phosphatase and osteocalcin. TPFs also upregulated osteoblast differentiation related genes, including osteocalcin, osterix, and Runx2 in primary osteoblasts. TPFs treated primary osteoblast cells showed significant upregulation of bone morphogenetic proteins (BMPs) including Bmp-2, Bmp-4, and Bmp-7. Addition of noggin, a BMP specific-antagonist, inhibited TPFs induced upregulation of the osteocalcin, osterix, and Runx2. Our findings point towards the induction of osteoblast differentiation by TPFs and suggested that TPFs could be a potential anabolic agent to treat patients with bone loss-associated diseases such as osteoporosis.

Effect of bone sialoprotein coating of ceramic and synthetic polymer materials on in vitro osteogenic cell differentiation and in vivo bone formation.

PubMed

Schaeren, Stefan; Jaquiéry, Claude; Wolf, Francine; Papadimitropoulos, Adam; Barbero, Andrea; Schultz-Thater, Elke; Heberer, Michael; Martin, Ivan

2010-03-15

In this study, we addressed whether Bone Sialoprotein (BSP) coating of various substrates could enhance the in vitro osteogenic differentiation and in vivo bone formation capacity of human Bone Marrow Stromal Cells (BMSC). Moreover, we tested whether synthetic polymer-based porous scaffolds, despite the absence of a mineral component, could support ectopic bone formation by human BMSC if coated with BSP. Adsorption of recombinant human BSP on tissue culture-treated polystyrene (TCTP), beta-tricalcium phosphate (Osteologic) or synthetic polymer (Polyactive) substrates was dose dependent, but did not consistently accelerate or enhance in vitro BMSC osteogenic differentiation, as assessed by the mRNA expression of osteoblast-related genes. Similarly, BSP coating of porous beta-tricalcium phosphate scaffolds (Skelite) did not improve the efficiency of bone tissue formation following loading with BMSC and ectopic implantation in nude mice. Finally, Polyactive foams seeded with BMSC did not form bone tissue in the same ectopic assay, even if coated with BSP. We conclude that BSP coating of a variety of substrates is not directly associated with an enhancement of osteoprogenitor cell differentiation in vitro or in vivo, and that presentation of BSP on polymeric materials is not sufficient to prime BMSC functional osteoblastic differentiation in vivo. (c) 2009 Wiley Periodicals, Inc.

Gastrodin inhibits osteoclastogenesis via down-regulating the NFATc1 signaling pathway and stimulates osseointegration in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Feng; Shen, Yi; Liu, Bo

Bone is a rigid yet dynamic organ, and this dynamism is mediated by the delicate balance between osteoclastic bone resorption and osteoblastic bone formation. However, excessive activation of osteoclasts is responsible for many bone diseases such as osteoporosis, Paget disease, and tumor bone metastasis. Agents that could inhibit osteoclast formation or function are regarded as promising alternatives to treat osteoclast-related diseases. Recently, traditional Chinese medicine has attracted attention because of its multiple activities in bone metabolism. Among them, gastrodin has been reported as an anti-osteoporosis agent that reduces reactive oxygen species. However, the direct action of gastrodin on osteoclast differentiationmore » and bone resorption, and its underlying molecular mechanism, remain unknown. In this study, we investigated the effects of gastrodin on receptor activator NF-κB ligand (RANKL)-activated osteoclasts formation and bone resorption. Our results showed that gastrodin retarded RANKL-induced osteoclast differentiation efficiently by downregulating transcriptional and translational expression of nuclear factor of activated T cells cl (NFATc1), a major factor in RANKL-mediated osteoclastogenesis. Meanwhile, gastrodin prevented osteoclast maturation and migration by inhibiting the gene expression of dendrocyte expressed seven transmembrane protein (DC-STAMP), an osteoclastic-specific gene that controls cells fusion and movement. And gastrodin prevented RANKL-induced osteoclastic bone erosion in vitro. In addition, gastrodin also stimulated bone mesenchymal stem cell (BMSC) spreading and osseointegration in titanium plate. In summary, gastrodin could prevent osteoclasts formation and bone resorption via blockage of NFATc1 activity, and stimulate osseointegration in vitro. Gastrodin could be developed as a potent phytochemical candidate to treat osteolytic diseases. - Highlights: • Gastrodin suppresses osteoclasts formation and function in vitro. • Gastrodin impairs NFATc1 activation. • Gastrodin stimulates osseointegration in vitro. • Gastrodin may be used for treating osteoclast related diseases.« less

Sonochemical synthesis of fructose 1,6-bisphosphate dicalcium porous microspheres and their application in promotion of osteogenic differentiation.

PubMed

Qi, Chao; Zhou, Ding; Zhu, Ying-Jie; Sun, Tuan-Wei; Chen, Feng; Zhang, Chang-Qing

2017-08-01

Human bone mesenchymal stem cells (hBMSCs) have the ability to differentiate into bone and cartilage for clinical bone regeneration. Biomaterials with an innate ability to stimulate osteogenic differentiation of hBMSCs into bone and cartilage are considered attractive candidates for the applications in bone tissue engineering and regeneration. In this paper, we synthesized fructose 1,6-bisphosphate dicalcium (Ca 2 FBP) porous microspheres by the sonochemical method, and investigated the ability of Ca 2 FBP for the promotion of the osteogenic differentiation of hBMSCs. After the hBMSCs were co-cultured with the sterilized powder of Ca 2 FBP porous microspheres for different times, the cell proliferation assay, alkaline phosphatase activity assay, quantitative real-time polymerase chain reaction and western blotting were performed to investigate the bioactivity and osteogenic differentiation performance of the as-prepared product. Compared with hydroxyapatite nanorods, Ca 2 FBP porous microspheres show a superior bioactivity and osteoinductive potential, and can promote the cell differentiation of hBMSCs in vitro, thus, they are promising for applications in the tissue engineering field such as dental and bone defect repair. Copyright © 2017 Elsevier B.V. All rights reserved.

SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells

PubMed Central

de Kroon, Laurie M. G.; Narcisi, Roberto; van den Akker, Guus G. H.; Vitters, Elly L.; Blaney Davidson, Esmeralda N.; van Osch, Gerjo J. V. M.; van der Kraan, Peter M.

2017-01-01

To improve cartilage formation by bone marrow-derived mesenchymal stem cells (BMSCs), the signaling mechanism governing chondrogenic differentiation requires better understanding. We previously showed that the transforming growth factor-β (TGFβ) receptor ALK5 is crucial for chondrogenesis induced by TGFβ. ALK5 phosphorylates SMAD2 and SMAD3 proteins, which then form complexes with SMAD4 to regulate gene transcription. By modulating the expression of SMAD2, SMAD3 and SMAD4 in human BMSCs, we investigated their role in TGFβ-induced chondrogenesis. Activation of TGFβ signaling, represented by SMAD2 phosphorylation, was decreased by SMAD2 knockdown and highly increased by SMAD2 overexpression. Moreover, TGFβ signaling via the alternative SMAD1/5/9 pathway was strongly decreased by SMAD4 knockdown. TGFβ-induced chondrogenesis of human BMSCs was strongly inhibited by SMAD4 knockdown and only mildly inhibited by SMAD2 knockdown. Remarkably, both knockdown and overexpression of SMAD3 blocked chondrogenic differentiation. Chondrogenesis appears to rely on a delicate balance in the amount of SMAD3 and SMAD4 as it was not enhanced by SMAD4 overexpression and was inhibited by SMAD3 overexpression. Furthermore, this study reveals that TGFβ-activated phosphorylation of SMAD2 and SMAD1/5/9 depends on the abundance of SMAD4. Overall, our findings suggest a more dominant role for SMAD3 and SMAD4 than SMAD2 in TGFβ-induced chondrogenesis of human BMSCs. PMID:28240243

SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells.

PubMed

de Kroon, Laurie M G; Narcisi, Roberto; van den Akker, Guus G H; Vitters, Elly L; Blaney Davidson, Esmeralda N; van Osch, Gerjo J V M; van der Kraan, Peter M

2017-02-27

To improve cartilage formation by bone marrow-derived mesenchymal stem cells (BMSCs), the signaling mechanism governing chondrogenic differentiation requires better understanding. We previously showed that the transforming growth factor-β (TGFβ) receptor ALK5 is crucial for chondrogenesis induced by TGFβ. ALK5 phosphorylates SMAD2 and SMAD3 proteins, which then form complexes with SMAD4 to regulate gene transcription. By modulating the expression of SMAD2, SMAD3 and SMAD4 in human BMSCs, we investigated their role in TGFβ-induced chondrogenesis. Activation of TGFβ signaling, represented by SMAD2 phosphorylation, was decreased by SMAD2 knockdown and highly increased by SMAD2 overexpression. Moreover, TGFβ signaling via the alternative SMAD1/5/9 pathway was strongly decreased by SMAD4 knockdown. TGFβ-induced chondrogenesis of human BMSCs was strongly inhibited by SMAD4 knockdown and only mildly inhibited by SMAD2 knockdown. Remarkably, both knockdown and overexpression of SMAD3 blocked chondrogenic differentiation. Chondrogenesis appears to rely on a delicate balance in the amount of SMAD3 and SMAD4 as it was not enhanced by SMAD4 overexpression and was inhibited by SMAD3 overexpression. Furthermore, this study reveals that TGFβ-activated phosphorylation of SMAD2 and SMAD1/5/9 depends on the abundance of SMAD4. Overall, our findings suggest a more dominant role for SMAD3 and SMAD4 than SMAD2 in TGFβ-induced chondrogenesis of human BMSCs.

Selective resistance of CD44hi T cells to p53-dependent cell death results in persistence of immunologic memory after total body irradiation.

PubMed

Yao, Zhenyu; Jones, Jennifer; Kohrt, Holbrook; Strober, Samuel

2011-10-15

Our previous studies showed that treatment of mice with total body irradiation (TBI) or total lymphoid tissue irradiation markedly changes the balance of residual T cell subsets to favor CD4(+)CD44(hi) NKT cells because of the differential resistance of the latter subset to cell death. The object of the current study was to further elucidate the changed balance and mechanisms of differential radioresistance of T cell subsets after graded doses of TBI. The experimental results showed that CD4(+) T cells were markedly more resistant than CD8(+) T cells, and CD44(hi) T cells, including NKT cells and memory T cells, were markedly more resistant than CD44(lo) (naive) T cells. The memory T cells immunized to alloantigens persisted even after myeloablative (1000 cGy) TBI and were able to prevent engraftment of bone marrow transplants. Although T cell death after 1000 cGy was prevented in p53(-/-) mice, there was progressive T cell death in p53(-/-) mice at higher doses. Although p53-dependent T cell death changed the balance of subsets, p53-independent T cell death did not. In conclusion, resistance of CD44(hi) T cells to p53-dependent cell death results in the persistence of immunological memory after TBI and can explain the immune-mediated rejection of marrow transplants in sensitized recipients.

Diagnosing aneurysmal and unicameral bone cysts with magnetic resonance imaging.

PubMed

Sullivan, R J; Meyer, J S; Dormans, J P; Davidson, R S

1999-09-01

The differential between aneurysmal bone cysts and unicameral bone cysts usually is clear clinically and radiographically. Occasionally there are cases in which the diagnosis is not clear. Because natural history and treatment are different, the ability to distinguish between these two entities before surgery is important. The authors reviewed, in a blinded fashion, the preoperative magnetic resonance images to investigate criteria that could be used to differentiate between the two lesions. All patients had operative or pathologic confirmation of an aneurysmal bone cyst or unicameral bone cyst. The authors analyzed the preoperative magnetic resonance images of 14 patients with diagnostically difficult bone cysts (eight children with unicameral bone cysts and six children with aneurysmal bone cysts) and correlated these findings with diagnosis after biopsy or cyst aspiration and contrast injection. The presence of a double density fluid level within the lesion strongly indicated that the lesion was an aneurysmal bone cyst, rather than a unicameral bone cyst. Other criteria that suggested the lesion was an aneurysmal bone cyst were the presence of septations within the lesion and signal characteristics of low intensity on T1 images and high intensity on T2 images. The authors identified a way of helping to differentiate between aneurysmal bone cysts and unicameral bone cysts on magnetic resonance images. Double density fluid level, septation, and low signal on T1 images and high signal on T2 images strongly suggest the bone cyst in question is an aneurysmal bone cyst, rather than a unicameral bone cyst. This may be helpful before surgery for the child who has a cystic lesion for which radiographic features do not allow a clear differentiation of unicameral bone cyst from aneurysmal bone cyst.

Changes in the population of perivascular cells in the bone tissue remodeling zones under microgravity

NASA Astrophysics Data System (ADS)

Katkova, Olena; Rodionova, Natalia; Shevel, Ivan

2016-07-01

Microgravity and long-term hypokinesia induce reduction both in bone mass and mineral saturation, which can lead to the development of osteoporosis and osteopenia. (Oganov, 2003). Reorganizations and adaptive remodeling processes in the skeleton bones occur in the topographical interconnection with blood capillaries and perivascular cells. Radioautographic studies with 3H- thymidine (Kimmel, Fee, 1980; Rodionova, 1989, 2006) have shown that in osteogenesis zones there is sequential differentiation process of the perivascular cells into osteogenic. Hence the study of populations of perivascular stromal cells in areas of destructive changes is actual. Perivascular cells from metaphysis of the rat femoral bones under conditions of modeling microgravity were studied using electron microscopy and cytochemistry (hindlimb unloading, 28 days duration) and biosatellite «Bion-M1» (duration of flight from April 19 till May 19, 2013 on C57, black mice). It was revealed that both control and test groups populations of the perivascular cells are not homogeneous in remodeling adaptive zones. These populations comprise of adjacent to endothelium poorly differentiated forms and isolated cells with signs of differentiation (specific increased volume of rough endoplasmic reticulum in cytoplasm). Majority of the perivascular cells in the control group (modeling microgravity) reveals reaction to alkaline phosphatase (marker of the osteogenic differentiation). In poorly differentiated cells this reaction is registered in nucleolus, nucleous and cytoplasm. In differentiating cells activity of the alkaline phosphatase is also detected on the outer surface of the cellular membrane. Unlike the control group in the bones of experimental animals reaction to the alkaline phosphatase is registered not in all cells of perivascular population. Part of the differentiating perivascular cells does not contain a product of the reaction. Under microgravity some poorly differentiated perivascular cells reveal signs of destruction. Thus it was found that number of the alkaline phosphatase containing cells (i.e. osteogenic cells) declines in perivascular cells population. It is one of the mechanisms of the osteogenic process decrease of intensity in bones because of lessening support loading on the bone skeleton. In the adaptive remodeling zones of bone tissue (near the vascular canals) in experiments fibroblasts and fibrosis zones were found - areas filled with non-mineralized collagen fibrils on the bones surfaces. Hence it should be considered that decrease (removal) of support loading slows down osteogenic differentiation of the part of perivascular cells and stimulates differentiation of the fibroblast cells. Obtained data is considered as one of the cellular mechanisms of the adaptive reactions development in spongy bone under microgravity which could lead to the bone mass loss.

NF-κB RelB Negatively Regulates Osteoblast Differentiation and Bone Formation

PubMed Central

Yao, Zhenqiang; Li, Yanyun; Yin, Xiaoxiang; Dong, Yufeng; Xing, Lianping; Boyce, Brendan F.

2013-01-01

RelA-mediated NF-κB canonical signaling promotes mesenchymal progenitor cell (MPC) proliferation, but inhibits differentiation of mature osteoblasts (OBs) and thus negatively regulates bone formation. Previous studies suggest that NF-κB RelB may also negatively regulate bone formation through non-canonical signaling, but they involved a complex knockout mouse model and the molecular mechanisms involved were not investigated. Here, we report that RelB−/− mice develop age-related increased trabecular bone mass associated with increased bone formation. RelB−/− bone marrow stromal cells expanded faster in vitro and have enhanced OB differentiation associated with increased expression of the osteoblastogenic transcription factor, Runx2. In addition, RelB directly targeted the Runx2 promoter to inhibit its activation. Importantly, RelB−/− bone-derived MPCs formed bone more rapidly than wild-type cells after they were injected into a murine tibial bone defect model. Our findings indicate that RelB negatively regulates bone mass as mice age and limits bone formation in healing bone defects, suggesting that inhibition of RelB could reduce age-related bone loss and enhance bone repair. PMID:24115294

[Development, physiology, and cell activity of bone].

PubMed

de Baat, P; Heijboer, M P; de Baat, C

2005-07-01

Bones are of crucial importance for the human body, providing skeletal support, serving as a home for the formation of haematopoietic cells, and reservoiring calcium and phosphate. Long bones develop by endochondral ossification. Flat bones develop by intramembranous ossification. Bone tissue contains hydroxyapatite and various extracellular proteins, producing bone matrix. Two biological mechanisms, determining the strength of bone, are modelling and remodelling. Modelling can change bone shape and size through bone formation by osteoblasts at some sites and through bone destruction by osteoclasts at other sites. Remodelling is bone turnover, also performed by osteoclasts and osteoblasts. The processes of modelling and remodelling are induced by mechanical loads, predominantly muscle loads. Osteoblasts develop from mesenchymal stem cells. Many stimulating factors are known to activate the differentiation. Mature osteoblasts synthesize bone matrix and may further differentiate into osteocytes. Osteocytes maintain structural bone integrity and allow bone to adapt to any mechanical and chemical stimulus. Osteoclasts derive from haematopoietic stem cells. A number of transcription and growth factors have been identified essential for osteoclast differentiation and function. Finally, there is a complex interaction between osteoblasts and osteoclasts. Bone destruction starts by attachment of osteoclasts to the bone surface. Following this, osteoclasts undergo specific morphological changes. The process of bone destruction starts by acid dissolution of hydroxyapatite. After that osteoclasts start to destruct the organic matrix.

Calcium Kinetics During Space Flight

NASA Technical Reports Server (NTRS)

Smith, Scott M.; OBrien, K. O.; Abrams, S. A.; Wastney, M. E.

2005-01-01

Bone loss during space flight is one of the most critical challenges to astronaut health on space exploration missions. Defining the time course and mechanism of these changes will aid in developing means to counteract bone loss during space flight, and will have relevance for other clinical situations that impair weight-bearing activity. Bone health is a product of the balance between bone formation and bone resorption. Early space research could not clearly identify which of these was the main process altered in bone loss, but identification of the collagen crosslinks in the 1990s made possible a clear understanding that the impact of space flight was greater on bone resorption, with bone formation being unchanged or only slightly decreased. Calcium kinetics data showed that bone resorption was greater during flight than before flight (668 plus or minus 130 vs. 427 plus or minus 153 mg/d, p less than 0.001), and clearly documented that true intestinal calcium absorption was lower during flight than before flight (233 plus or minus 87 vs. 460 plus or minus 47 mg/d, p less than 0.01). Weightlessness had a detrimental effect on the balance in bone turnover: the difference between daily calcium balance during flight (-234 plus or minus 102 mg/d) and calcium balance before flight (63 plus or minus 75 mg/d) approached 300 mg/d (p less than 0.01). These data demonstrate that the bone loss that occurs during space flight is a consequence of increased bone resorption and decreased intestinal calcium absorption. Examining the changes in bone and calcium homeostasis in the initial days and weeks of space flight, as well as at later times on missions longer than 6 months, is critical to understanding the nature of bone adaptation to weightlessness. To increase knowledge of these changes, we studied bone adaptation to space flight on the 16-day Space Shuttle Columbia (STS-107) mission. When the brave and talented crew of Columbia were lost during reentry on the tragic morning of February 1, 2003, in a much smaller matter, the scientific products of this experiment, successfully obtained on orbit, were lost as well. As we begin to plan for missions back to the Moon, and even off to Mars, many questions remain to be answered. Counteracting bone loss is one of the greatest challenges. Calcium kinetics studies provide a valuable tool for assessing this loss, and evaluating countermeasures.

Gene expression profiling of bone marrow mesenchymal stem cells from Osteogenesis Imperfecta patients during osteoblast differentiation.

PubMed

Kaneto, Carla Martins; Pereira Lima, Patrícia S; Prata, Karen Lima; Dos Santos, Jane Lima; de Pina Neto, João Monteiro; Panepucci, Rodrigo Alexandre; Noushmehr, Houtan; Covas, Dimas Tadeu; de Paula, Francisco José Alburquerque; Silva, Wilson Araújo

2017-06-01

Mesenchymal stem cells (MSCs) are precursors present in adult bone marrow that are able to differentiate into osteoblasts, adipocytes and chondroblasts that have gained great importance as a source for cell therapy. Recently, a number of studies involving the analysis of gene expression of undifferentiated MSCs and of MSCs in the differentiation into multiple lineage processes were observed but there is no information concerning the gene expression of MSCs from Osteogenesis Imperfecta (OI) patients. Osteogenesis Imperfecta is characterized as a genetic disorder in which a generalized osteopenia leads to excessive bone fragility and severe bone deformities. The aim of this study was to analyze gene expression profile during osteogenic differentiation from BMMSCs (Bone Marrow Mesenchymal Stem Cells) obtained from patients with Osteogenesis Imperfecta and from control subjects. Bone marrow samples were collected from three normal subjects and five patients with OI. Mononuclear cells were isolated for obtaining mesenchymal cells that had been expanded until osteogenic differentiation was induced. RNA was harvested at seven time points during the osteogenic differentiation period (D0, D+1, D+2, D+7, D+12, D+17 and D+21). Gene expression analysis was performed by the microarray technique and identified several differentially expressed genes. Some important genes for osteoblast differentiation had lower expression in OI patients, suggesting a smaller commitment of these patient's MSCs with the osteogenic lineage. Other genes also had their differential expression confirmed by RT-qPCR. An increase in the expression of genes related to adipocytes was observed, suggesting an increase of adipogenic differentiation at the expense osteogenic differentiation. Copyright © 2017. Published by Elsevier Masson SAS.

Differentiation potentials of perivascular cells in the bone tissue remodeling zones under microgravity

NASA Astrophysics Data System (ADS)

Rodionova, Natalia; Katkova, Olena

Adaptive remodeling processes in the skeleton bones occur in the close topographical interconnection with blood capillaries followed by perivascular cells. Radioautographic studies with 3H- thymidine (Kimmel D.B., Fee W.S., 1980; Rodionova N.V., 1989, 2006) has shown that in osteogenesis zones there is sequential differentiation process of the perivascular cells into osteogenic ones. Using electron microscopy and cytochemistry we studied perivsacular cells in metaphysis of the rats femoral bones under conditions of modeling microgravity (28 days duration) and in femoral bones metaphyses of rats flown on board of the space laboratory (Spacelab - 2) It was revealed that population of the perivascular cells is not homogeneous in adaptive zones of the remodeling in both control and test groups (lowering support loading). This population comprises adjacent to endothelium little differentiated forms and isolated cells with differentiation features (specific volume of rough endoplasmic reticulum in cytoplasm is increased). Majority of the perivascular cells in the control group reveals reaction to alkaline phosphatase (marker of the osteogenic differentiation). In little differentiated cells this reaction is registered in nucleolus, nucleous and cytoplasm. In differentiating cells activity of the alkaline phosphatase is also detected on the outer surface of the cellular membrane. Unlike the control group in the bones of animals under microgravitaty reaction to the alkaline phosphatase is registered not for all cells of perivascular population. Part of the differentiating perivascular cells does not contain a product of the reaction. There is also visible trend of individual alkaline phosphatase containing perivascular cells amounts decrease (i.e. osteogenic cells-precursors). Under microgravity some little differentiated perivascular cells reveal destruction signs. Found decrease trend of the alkaline phosphatase containing cells (i.e. osteogenic cells) number in perivascular cells population. It is one of the mechanisms of the osteogenic process intensity decrease in bones due to lowering support loading on the bone skeleton. In particular this is confirmed by the fact that in the zones of adaptive remodeling we found fibroblasts and fibrosis zones - areas filled with non mineralized collagen fibrils on the bones surfaces. Hence it should be considered that lowering (removal) support loading slows down (or blocks) osteogenic differentiation of the perivascular cells part and stimulates differentiation of the fibroblast cells. Obtained data considered as one of the cellular mechanisms of the adaptive reactions development in spongy bone under microgravity which could lead to the bone mass loss.

Morphological and proteomic analysis of early stage of osteoblast differentiation in osteoblastic progenitor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hong, Dun; Orthopedic Department, Taizhou Hospital, Wenzhou Medical College, Linhai, Zhejiang 317000; Chen, Hai-Xiao, E-mail: Hxchen-1@163.net

Bone remodeling relies on a dynamic balance between bone formation and resorption, mediated by osteoblasts and osteoclasts, respectively. Under certain stimuli, osteoprogenitor cells may differentiate into premature osteoblasts and further into mature osteoblasts. This process is marked by increased alkaline phosphatase (ALP) activity and mineralized nodule formation. In this study, we induced osteoblast differentiation in mouse osteoprogenitor MC3T3-E1 cells and divided the process into three stages. In the first stage (day 3), the MC3T3-E1 cell under osteoblast differentiation did not express ALP or deposit a mineralized nodule. In the second stage, the MC3T3-E1 cell expressed ALP but did not formmore » a mineralized nodule. In the third stage, the MC3T3-E1 cell had ALP activity and formed mineralized nodules. In the present study, we focused on morphological and proteomic changes of MC3T3-E1 cells in the early stage of osteoblast differentiation - a period when premature osteoblasts transform into mature osteoblasts. We found that mean cell area and mean stress fiber density were increased in this stage due to enhanced cell spreading and decreased cell proliferation. We further analyzed the proteins in the signaling pathway of regulation of the cytoskeleton using a proteomic approach and found upregulation of IQGAP1, gelsolin, moesin, radixin, and Cfl1. After analyzing the focal adhesion signaling pathway, we found the upregulation of FLNA, LAMA1, LAMA5, COL1A1, COL3A1, COL4A6, and COL5A2 as well as the downregulation of COL4A1, COL4A2, and COL4A4. In conclusion, the signaling pathway of regulation of the cytoskeleton and focal adhesion play critical roles in regulating cell spreading and actin skeleton formation in the early stage of osteoblast differentiation.« less

Maintenance of exercise-induced benefits in physical functioning and bone among elderly women.

PubMed

Karinkanta, S; Heinonen, A; Sievänen, H; Uusi-Rasi, K; Fogelholm, M; Kannus, P

2009-04-01

This study showed that about a half of the exercise-induced gain in dynamic balance and bone strength was maintained one year after cessation of the supervised high-intensity training of home-dwelling elderly women. However, to maintain exercise-induced gains in lower limb muscle force and physical functioning, continued training seems necessary. Maintenance of exercise-induced benefits in physical functioning and bone structure was assessed one year after cessation of 12-month randomized controlled exercise intervention. Originally 149 healthy women 70-78 years of age participated in the 12-month exercise RCT and 120 (81%) of them completed the follow-up study. Self-rated physical functioning, dynamic balance, leg extensor force, and bone structure were assessed. During the intervention, exercise increased dynamic balance by 7% in the combination resistance and balance-jumping training group (COMB). At the follow-up, a 4% (95% CI: 1-8%) gain compared with the controls was still seen, while the exercise-induced isometric leg extension force and self-rated physical functioning benefits had disappeared. During the intervention, at least twice a week trained COMB subjects obtained a significant 2% benefit in tibial shaft bone strength index compared to the controls. A half of this benefit seemed to be maintained at the follow-up. Exercise-induced benefits in dynamic balance and rigidity in the tibial shaft may partly be maintained one year after cessation of a supervised 12-month multi-component training in initially healthy elderly women. However, to maintain the achieved gains in muscle force and physical functioning, continued training seems necessary.

A pilot study of the feasibility of long-term human bone balance during perimenopause using a 41Ca tracer

NASA Astrophysics Data System (ADS)

Hui, S. K.; Prior, J.; Gelbart, Z.; Johnson, R. R.; Lentle, B. C.; Paul, M.

2007-06-01

The mechanisms governing calcium fluxes during bone remodeling processes in perimenopausal women are poorly known. Despite higher, albeit erratic, estradiol levels in perimenopause, spine bone loss is greater than during the first five years past the final menstrual flow when estradiol becomes low. Understanding changes during this dynamic transition are important to prevent fragility fractures in midlife and older women. The exploration of long-lived 41Ca (T1/2 = 1.04 × 105 yrs) tracer measurements using accelerator mass spectrometry (AMS) leads to the possibility of monitoring bone remodeling balance. With this new technology, we explored a pilot long-term feasibility study of bone health by measuring the 41Ca trace element in urine for six years from premenopausal to later perimenopausal phases in one midlife woman. We measured bone mineral density in parallel.

The effects and mechanisms of clinorotation on proliferation and differentiation in bone marrow mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yan, Ming; Wang, Yongchun; Yang, Min

Data from human and rodent studies have demonstrated that microgravity induces observed bone loss in real spaceflight or simulated experiments. The decrease of bone formation and block of maturation may play important roles in bone loss induced by microgravity. The aim of this study was to investigate the changes of proliferation and differentiation in bone marrow mesenchymal stem cells (BMSCs) induced by simulated microgravity and the mechanisms underlying it. We report here that clinorotation, a simulated model of microgravity, decreased proliferation and differentiation in BMSCs after exposure to 48 h simulated microgravity. The inhibited proliferation are related with blocking the cellmore » cycle in G2/M and enhancing the apoptosis. While alterations of the osteoblast differentiation due to the decreased SATB2 expression induced by simulated microgravity in BMSCs. - Highlights: • Simulated microgravity inhibited proliferation and differentiation in BMSCs. • The decreased proliferation due to blocked cell cycle and enhanced the apoptosis. • The inhibited differentiation accounts for alteration of SATB2, Hoxa2 and Cbfa1.« less

Osteoclast differentiation inhibitors: a patent review (2008 - 2012).

PubMed

Kim, Seong Hwan; Moon, Seong-Hee

2013-12-01

Mononuclear macrophage/monocyte-lineage hematopoietic precursors differentiate into multinucleated osteoclasts. Abnormally increased numbers and/or overactivation of osteoclasts can lead to bone loss. Therefore, pharmaceutical inhibition of osteoclast differentiation is one therapeutic strategy for mitigating the occurrence of bone loss-associated disorders and related fractures. This review surveys the patents and patent applications from 2008 to 2012 that are related to inventions of therapeutics and/or methods for inhibiting osteoclast differentiation. Over the past 20 years, the identification and validation of signaling molecules involved in osteoclast differentiation has led to a better understanding of the molecular mechanism, and to the development of new therapeutic agents for treating bone loss-associated disorders. Since 2008, 34 WO patents or patent applications have been filed that relate to inventions of therapeutics and/or methods for chemical-based, natural product-based, or biological-based inhibitors of osteoclast differentiation. Here, analysis of these patents and patent applications is presented, and summarize the disclosed osteoclast differentiation-inhibiting target molecules. This report can support further advances in the development of anti-osteoclastogenic therapeutics for bone loss-associated disorders, including osteoporosis, rheumatoid arthritis, Paget's disease, periodontal disease, osteosarcoma, and cancer bone metastasis.

Mangiferin positively regulates osteoblast differentiation and suppresses osteoclast differentiation

PubMed Central

Sekiguchi, Yuusuke; Mano, Hiroshi; Nakatani, Sachie; Shimizu, Jun; Kataoka, Aya; Ogura, Kana; Kimira, Yoshifumi; Ebata, Midori; Wada, Masahiro

2017-01-01

Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3-E1 osteoblastic cells and osteoclast-like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST-1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that mangiferin significantly increased the mRNA level of runt-related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate-resistant acid phosphatase-positive multinuclear cells. RT-PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast-associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3-E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes. PMID:28627701

Mangiferin positively regulates osteoblast differentiation and suppresses osteoclast differentiation.

PubMed

Sekiguchi, Yuusuke; Mano, Hiroshi; Nakatani, Sachie; Shimizu, Jun; Kataoka, Aya; Ogura, Kana; Kimira, Yoshifumi; Ebata, Midori; Wada, Masahiro

2017-08-01

Mangiferin is a polyphenolic compound present in Salacia reticulata. It has been reported to reduce bone destruction and inhibit osteoclastic differentiation. This study aimed to determine whether mangiferin directly affects osteoblast and osteoclast proliferation and differentiation, and gene expression in MC3T3‑E1 osteoblastic cells and osteoclast‑like cells derived from primary mouse bone marrow macrophage cells. Mangiferin induced significantly greater WST‑1 activity, indicating increased cell proliferation. Mangiferin induced significantly increased alkaline phosphatase staining, indicating greater cell differentiation. Reverse transcription‑polymerase chain reaction (RT‑PCR) demonstrated that mangiferin significantly increased the mRNA level of runt‑related transcription factor 2 (RunX2), but did not affect RunX1 mRNA expression. Mangiferin significantly reduced the formation of tartrate‑resistant acid phosphatase‑positive multinuclear cells. RT‑PCR demonstrated that mangiferin significantly increased the mRNA level of estrogen receptor β (ERβ), but did not affect the expression of other osteoclast‑associated genes. Mangiferin may inhibit osteoclastic bone resorption by suppressing differentiation of osteoclasts and promoting expression of ERβ mRNA in mouse bone marrow macrophage cells. It also has potential to promote osteoblastic bone formation by promoting cell proliferation and inducing cell differentiation in preosteoblast MC3T3‑E1 cells via RunX2. Mangiferin may therefore be useful in improving bone disease outcomes.

Engineered Fibrin Gels for Parallel Stimulation of Mesenchymal Stem Cell Proangiogenic and Osteogenic Potential

PubMed Central

Murphy, Kaitlin C.; Hughbanks, Marissa L.; Binder, Bernard Y.K.; Vissers, Caroline B.; Leach, J. Kent

2014-01-01

Mesenchymal stem/stromal cells (MSCs) are under examination for use in cell therapies to repair bone defects resulting from trauma or disease. MSCs secrete proangiogenic cues and can be induced to differentiate into bone-forming osteoblasts, yet there is limited evidence that these events can be achieved in parallel. Manipulation of the cell delivery vehicle properties represents a candidate approach for directing MSC function in bone healing. We hypothesized that the biophysical properties of a fibrin gel could simultaneously regulate the proangiogenic and osteogenic potential of entrapped MSCs. Fibrin gels were formed by supplementation with NaCl (1.2, 2.3, and 3.9% w/v) to modulate gel biophysical properties without altering protein concentrations. MSCs entrapped in 1.2% w/v NaCl gels were the most proangiogenic in vitro, yet cells in 3.9% w/v gels exhibited the greatest osteogenic response. Compared to the other groups, MSCs entrapped in 2.3% w/v gels provided the best balance between proangiogenic potential, osteogenic potential, and gel contractility. The contribution of MSCs to bone repair was then examined when deployed in 2.3% w/v NaCl gels and implanted into an irradiated orthotopic bone defect. Compared to acellular gels after 3 weeks of implantation, defects treated with MSC-loaded fibrin gels exhibited significant increases in vessel density, early osteogenesis, superior morphology, and increased cellularity of repair tissue. Defects treated with MSC-loaded gels exhibited increased bone formation after 12 weeks compared to blank gels. These results confirm that fibrin gel properties can be modulated to simultaneously promote both the proangiogenic and osteogenic potential of MSCs, and fibrin gels modified by supplementation with NaCl are promising carriers for MSCs to stimulate bone repair in vivo. PMID:25527322

Pleiotrophin Commits Human Bone Marrow Mesenchymal Stromal Cells towards Hypertrophy during Chondrogenesis

PubMed Central

Bouderlique, Thibault; Henault, Emilie; Lebouvier, Angelique; Frescaline, Guilhem; Bierling, Phillipe; Rouard, Helene; Courty, José

2014-01-01

Pleiotrophin (PTN) is a growth factor present in the extracellular matrix of the growth plate during bone development and in the callus during bone healing. Bone healing is a complicated process that recapitulates endochondral bone development and involves many cell types. Among those cells, mesenchymal stromal cells (MSC) are able to differentiate toward chondrogenic and osteoblastic lineages. We aimed to determine PTN effects on differentiation properties of human bone marrow stromal cells (hBMSC) under chondrogenic induction using histological analysis and quantitative reverse transcription polymerase chain reaction. PTN dramatically potentiated chondrogenic differentiation as indicated by a strong increase of collagen 2 protein, and cartilage-related gene expression. Moreover, PTN increased transcription of hypertrophic chondrocyte markers such as MMP13, collagen 10 and alkaline phosphatase and enhanced calcification and the content of collagen 10 protein. These effects are dependent on PTN receptors signaling and PI3 K pathway activation. These data suggest a new role of PTN in bone regeneration as an inducer of hypertrophy during chondrogenic differentiation of hBMSC. PMID:24516627

Gradients in pore size enhance the osteogenic differentiation of human mesenchymal stromal cells in three-dimensional scaffolds

NASA Astrophysics Data System (ADS)

di Luca, Andrea; Ostrowska, Barbara; Lorenzo-Moldero, Ivan; Lepedda, Antonio; Swieszkowski, Wojcech; van Blitterswijk, Clemens; Moroni, Lorenzo

2016-03-01

Small fractures in bone tissue can heal by themselves, but in case of larger defects current therapies are not completely successful due to several drawbacks. A possible strategy relies on the combination of additive manufactured polymeric scaffolds and human mesenchymal stromal cells (hMSCs). The architecture of bone tissue is characterized by a structural gradient. Long bones display a structural gradient in the radial direction, while flat bones in the axial direction. Such gradient presents a variation in bone density from the cancellous bone to the cortical bone. Therefore, scaffolds presenting a gradient in porosity could be ideal candidates to improve bone tissue regeneration. In this study, we present a construct with a discrete gradient in pore size and characterize its ability to further support the osteogenic differentiation of hMSCs. Furthermore, we studied the behaviour of hMSCs within the different compartments of the gradient scaffolds, showing a correlation between osteogenic differentiation and ECM mineralization, and pore dimensions. Alkaline phosphatase activity and calcium content increased with increasing pore dimensions. Our results indicate that designing structural porosity gradients may be an appealing strategy to support gradual osteogenic differentiation of adult stem cells.

LPS-treated bone marrow-derived dendritic cells induce immune tolerance through modulating differentiation of CD4+ regulatory T cell subpopulations mediated by 3G11 and CD127.

PubMed

Zhou, Fang; Zhang, Guang-Xian; Rostami, Abdolmohamad

2017-06-01

Intravenous transfer of LPS-treated bone marrow-derived dendritic cells blocks development of autoimmunity induced by CD4 + T cells in vivo. However, cellular mechanisms of dendritic cell-mediated immune tolerance have not yet been fully elucidated. Here, we report that there are two new subpopulations of CD4 + CD25 + FoxP3 + GITR + regulatory T cells (CD127 + 3G11 + and CD127 + 3G11 - cells). LPS-treated dendritic cells facilitate development of CD4 + CD127 + 3G11 - regulatory T cells but inhibit that of CD4 + CD127 + 3G11 + regulatory T cells. LPS-induced tolerogenic dendritic cells may cause immune tolerance through modulating balance of different subsets of CD4 + regulatory T cells mediated by CD127 and 3G11. Our results imply a new potential cellular mechanism of dendritic cell-mediated immune tolerance.

HLA genetic diversity in Hungarians and Hungarian Gypsies: complementary differentiation patterns and demographic signals revealed by HLA-A, -B and -DRB1 in Central Europe.

PubMed

Inotai, D; Szilvasi, A; Benko, S; Boros-Major, A; Illes, Z; Bors, A; Kiss, K P; Rajczy, K; Gelle-Hossó, A; Buhler, S; Nunes, J M; Sanchez-Mazas, A; Tordai, A

2015-08-01

Systematic analyses of human leukocyte antigen (HLA) profiles in different populations may increase the efficiency of bone marrow donor selection and help reconstructing human peopling history. We typed HLA-A, -B, and -DRB1 allele groups in two bone marrow donor cohorts of 2402 Hungarians and 186 Hungarian Gypsies and compared them with several Central-European, Spanish Gypsy, and Indian populations. Our results indicate that different European Gypsy populations share a common origin but diverged genetically as a consequence of founder effect and rapid genetic drift, whereas other European populations are related genetically in relation to geography. This study also suggests that while HLA-A accurately depicts the effects of genetic drift, HLA-B, and -DRB1 conserve more signatures of ancient population relationships, as a result of balancing selection. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Deficits in distal radius bone strength, density and microstructure are associated with forearm fractures in girls: an HR-pQCTstudy

PubMed Central

Määttä, M.; Macdonald, H. M.; Mulpuri, K.

2016-01-01

Summary Forearm fractures are common during growth. We studied bone strength in youth with a recent forearm fracture. In girls, suboptimal bone strength was associated with fractures. In boys, poor balance and physical inactivity may lead to fractures. Prospective studies will confirm these relationships and identify targets for prevention strategies. Introduction The etiology of pediatric forearm fractures is unclear. Thus, we examined distal radius bone strength, microstructure, and density in children and adolescents with a recent low- or moderate-energy forearm fracture and those without forearm fractures. Methods We assessed the non-dominant (controls) and non-fractured (cases) distal radius (7 % site) using high-resolution peripheral quantitative computed tomography (HR-pQCT) (Scanco Medical AG) in 270 participants (girls: cases n=47, controls n=61 and boys: cases n=88, controls n=74) aged 8–16 years. We assessed standard anthropometry, maturity, body composition (dual energy X-ray absorptiometry (DXA), Hologic QDR 4500 W) physical activity, and balance. We fit sex-specific logistic regression models for each bone outcome adjusting for maturity, ethnicity, height, and percent body fat. Results In girls, impaired bone strength (failure load, ultimate stress) and a high load-to-strength ratio were associated with low-energy fractures (odds ratios (OR) 2.8–4.3). Low total bone mineral density (Tt.BMD), bone volume ratio, trabecular thickness, and cortical BMD and thickness were also associated with low-energy fractures (ORs 2.0–7.0). In boys, low Tt.BMD, but not bone strength, was associated with low-energy fractures (OR=1.8). Boys with low-energy fractures had poor balance and higher percent body fat compared with controls (p<0.05). Boys with fractures (both types) were less active than controls (p<0.05). Conclusions Forearm fracture etiology appears to be sex-specific. In girls, deficits in bone strength are associated with fractures. In boys, a combination of poor balance, excess body fat, and low physical activity may lead to fractures. Prospective studies are needed to confirm these relationships and clarify targets for prevention strategies. PMID:25572041

Can bone marrow differentiate into renal cells?

PubMed

Imai, Enyu; Ito, Takahito

2002-10-01

A considerable plasticity of adult stem cells has been confirmed in a wide variety of tissues. In particular, the pluripotency of bone marrow-derived stem cells may influence the regeneration of injured tissues and may provide novel avenues in regenerative medicine. Bone marrow contains at least hematopoietic and mesenchymal stem cells, and both can differentiate into a wide range of differentiated cells. Side population (SP) cells, which are originally defined in bone marrow cells by high efflux of DNA-binding dye, seem to be a new class of multipotent stem cells. Irrespective of the approach used to obtain stem cells, the fates of marrow-derived cells following bone marrow transplantation can be traced by labeling donor cells with green fluorescence protein or by identifying donor Y chromosome in female recipients. So far, bone marrow-derived cells have been reported to differentiate into renal cells, including mesangial cells, endothelial cells, podocytes, and tubular cells in the kidney, although controversy exists. Further studies are required to address this issue. Cell therapy will be promising when we learn to control stem cells such as bone marrow-derived stem cells, embryonic stem cells, and resident stem cells in the kidney. Identification of factors that support stem cells or promote their differentiation should provide a relevant step towards cell therapy.

The role of carboxy-terminal cross-linking telopeptide of type I collagen, dual x-ray absorptiometry bone strain and Romberg test in a new osteoporotic fracture risk evaluation: A proposal from an observational study.

PubMed

Ulivieri, Fabio M; Piodi, Luca P; Grossi, Enzo; Rinaudo, Luca; Messina, Carmelo; Tassi, Anna P; Filopanti, Marcello; Tirelli, Anna; Sardanelli, Francesco

2018-01-01

The consolidated way of diagnosing and treating osteoporosis in order to prevent fragility fractures has recently been questioned by some papers, which complained of overdiagnosis and consequent overtreatment of this pathology with underestimating other causes of the fragility fractures, like falls. A new clinical approach is proposed for identifying the subgroup of patients prone to fragility fractures. This retrospective observational study was conducted from January to June 2015 at the Nuclear Medicine-Bone Metabolic Unit of the of the Fondazione IRCCS Ca' Granda, Milan, Italy. An Italian population of 125 consecutive postmenopausal women was investigated for bone quantity and bone quality. Patients with neurological diseases regarding balance and vestibular dysfunction, sarcopenia, past or current history of diseases and use of drugs known to affect bone metabolism were excluded. Dual X-ray absorptiometry was used to assess bone quantity (bone mineral density) and bone quality (trabecular bone score and bone strain). Biochemical markers of bone turnover (type I collagen carboxy-terminal telopeptide, alkaline phosphatase, vitamin D) have been measured. Morphometric fractures have been searched by spine radiography. Balance was evaluated by the Romberg test. The data were evaluated with the neural network analysis using the Auto Contractive Map algorithm. The resulting semantic map shows the Minimal Spanning Tree and the Maximally Regular Graph of the interrelations between bone status parameters, balance conditions and fractures of the studied population. A low fracture risk seems to be related to a low carboxy-terminal cross-linking telopeptide of type I collagen level, whereas a positive Romberg test, together with compromised bone trabecular microarchitecture DXA parameters, appears to be strictly connected with fragility fractures. A simple assessment of the risk of fragility fracture is proposed in order to identify those frail patients at risk for osteoporotic fractures, who may have the best benefit from a pharmacological and physiotherapeutic approach.

Human decellularized bone scaffolds from aged donors show improved osteoinductive capacity compared to young donor bone.

PubMed

Smith, Christopher A; Board, Tim N; Rooney, Paul; Eagle, Mark J; Richardson, Stephen M; Hoyland, Judith A

2017-01-01

To improve the safe use of allograft bone, decellularization techniques may be utilized to produce acellular scaffolds. Such scaffolds should retain their innate biological and biomechanical capacity and support mesenchymal stem cell (MSC) osteogenic differentiation. However, as allograft bone is derived from a wide age-range, this study aimed to determine whether donor age impacts on the ability an osteoinductive, acellular scaffold produced from human bone to promote the osteogenic differentiation of bone marrow MSCs (BM-MSC). BM-MSCs from young and old donors were seeded on acellular bone cubes from young and old donors undergoing osteoarthritis related hip surgery. All combinations resulted in increased osteogenic gene expression, and alkaline phosphatase (ALP) enzyme activity, however BM-MSCs cultured on old donor bone displayed the largest increases. BM-MSCs cultured in old donor bone conditioned media also displayed higher osteogenic gene expression and ALP activity than those exposed to young donor bone conditioned media. ELISA and Luminex analysis of conditioned media demonstrated similar levels of bioactive factors between age groups; however, IGF binding protein 1 (IGFBP1) concentration was significantly higher in young donor samples. Additionally, structural analysis of old donor bone indicated an increased porosity compared to young donor bone. These results demonstrate the ability of a decellularized scaffold produced from young and old donors to support osteogenic differentiation of cells from young and old donors. Significantly, the older donor bone produced greater osteogenic differentiation which may be related to reduced IGFBP1 bioavailability and increased porosity, potentially explaining the excellent clinical results seen with the use of allograft from aged donors.

Tridax procumbens flavonoids: a prospective bioactive compound increased osteoblast differentiation and trabecular bone formation.

PubMed

Al Mamun, Md Abdullah; Hosen, Mohammad Jakir; Khatun, Amina; Alam, M Masihul; Al-Bari, Md Abdul Alim

2017-09-08

The Tridax procumbens extracts (TPE) are known for their ethno-medicinal properties to increase osteogenic functioning in mesenchymal stem cells. Recently, we found that the T. procumbens flavonoids (TPF) significantly suppressed the RANKL-induced osteoclasts differentiation and bone resorption. The TPF also promoted osteoblasts differentiation and bone formation demonstrated by increasing bone formation markers in cultured mouse primary osteoblasts. However, the effects of the TPF on in vivo bone formation remain unclear. In this study, we investigated the effects of the TPF on in vivo bone formation, injected the TPF (20 mg/kg) twice a day in the low calcium diet mice and killed them after 21 day. Radiographic and histomorphometric analyses were performed on the dissected bones to determine the anabolic effects of the TPF. Bone mineral density and bone mineral content of the TPF-treated mice were significantly increased compared to the control mice. Bone formation-related indices like osteoblast number, osteoblast surface, bone volume, mineralizing surface, mineral apposition rate and bone formation rate were significantly increased in the TPF-treated mice compared to the control mice. Our findings point towards the stimulation of bone formation by TPF, suggested that the TPF could be a potential natural anabolic agent to treat patients with bone loss-associated diseases such as osteoporosis.

Perfluoroalkyl substances in human bone: concentrations in bones and effects on bone cell differentiation.

PubMed

Koskela, A; Koponen, J; Lehenkari, P; Viluksela, M; Korkalainen, M; Tuukkanen, J

2017-07-28

Perfluoroalkyl substances (PFAS), including two most commonly studied compounds perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), are widely distributed environmental pollutants, used extensively earlier. Due to their toxicological effects the use of PFAS is now regulated. Based on earlier studies on PFOA's distribution in bone and bone marrow in mice, we investigated PFAS levels and their possible link to bone microarchitecture of human femoral bone samples (n = 18). Soft tissue and bone biopsies were also taken from a 49-year old female cadaver for PFAS analyses. We also studied how PFOA exposure affects differentiation of human osteoblasts and osteoclasts. PFAS were detectable from all dry bone and bone marrow samples, PFOS and PFOA being the most prominent. In cadaver biopsies, lungs and liver contained the highest concentrations of PFAS, whereas PFAS were absent in bone marrow. Perfluorononanoic acid (PFNA) was present in the bones, PFOA and PFOS were absent. In vitro results showed no disturbance in osteogenic differentiation after PFOA exposure, but in osteoclasts, lower concentrations led to increased resorption, which eventually dropped to zero after increase in PFOA concentration. In conclusion, PFAS are present in bone and have the potential to affect human bone cells partly at environmentally relevant concentrations.

Gene expression of runx2, Osterix, c-fos, DLX-3, DLX-5, and MSX-2 in dental follicle cells during osteogenic differentiation in vitro.

PubMed

Morsczeck, C

2006-02-01

Recently, osteogenic precursor cells were isolated from human dental follicles, which differentiate into cementoblast- or osteoblast- like cells under in vitro conditions. However, mechanisms for osteogenic differentiation are not known in detail. Dental follicle cell long-term cultures supplemented with dexamethasone or with insulin resulted in mineralized nodules, whereas no mineralization or alkaline phosphatase activity was detected in the control culture without an osteogenic stimulus. A real-time reverse-transcriptase polymerase chain reaction (PCR) analysis was developed to investigate gene expression during osteogenic differentiation in vitro. Expression of the alkaline phosphatase (ALP) gene was detected during differentiation in the control culture and was similar to that in cultures with dexamethasone and insulin. DLX-3, DLX-5, runx2, and MSX-2 are differentially expressed during osteogenic differentiation in bone marrow mesenchymal stem cells. In dental follicle cells, gene expression of runx2, DLX-5, and MSX-2 was unaffected during osteogenic differentiation in vitro. Osteogenic differentiation appeared to be independent of MSX-2 expression; the same was true of runx2 and DLX-5, which were protagonists of osteogenic differentiation and osteocalcin promoter activity in bone marrow mesenchymal stem cells. Like in bone marrow-derived stem cells, DLX-3 gene expression was increased in dental follicle cells during osteogenic differentiation but similar to control cultures. However, gene expression of osterix was not detected in dental follicle cells during osteogenic differentiation; this gene is expressed during osteogenic differentiation in bone marrow stem cells. These real-time PCR results display molecular mechanisms in dental follicle precursor cells during osteogenic differentiation that are different from those in bone marrow-derived mesenchymal stem cells.

Effect of in vitro chondrogenic differentiation of autologous mesenchymal stem cells on cartilage and subchondral cancellous bone repair in osteoarthritis of temporomandibular joint.

PubMed

Chen, K; Man, C; Zhang, B; Hu, J; Zhu, S S

2013-02-01

This study investigated the effects of in vitro chondrogenic differentiated mesenchymal stem cells (MSCs) on cartilage and subchondral cancellous bone in temporomandibular joint osteoarthritis (TMJOA). Four weeks after induction of osteoarthritis (OA), the joints received hylartin solution, non-chondrogenic MSCs or in vitro chondrogenic differentiated MSCs. The changes in cartilage and subchondral cancellous bone were evaluated by histology, reverse transcription polymerase chain reaction and micro-computed tomography (CT). Implanted cells were tracked using Adeno-LacZ labelling. The differentiated MSC-treated group had better histology than the MSC-treated group at 4 and 12 weeks, but no difference at 24 weeks. Increased mRNA expression of collegan II, aggeran, Sox9 and decreased matrix metalloproteinase 13 (MMP13) were observed in differentiated MSC-treated groups compared to the undifferentiated MSC-treated group at 4 weeks. The differentiated MSC-treated group had decreased bone volume fraction, trabecular thickness and bone surface density, and increased trabecular spacing in the subchondral cancellous bone than the undifferentiated MSC-treated group. Transplanted cells were observed at cartilage, subchondral bone, and the synovial membrane lining at 4 weeks. Intra-articular injection of MSCs could delay the progression of TMJOA, and in vitro chondrogenic induction of MSCs could enhance the therapeutic effects. This provides new insights into the role of MSCs in cell-based therapies for TMJOA. Copyright © 2012 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

Graphene supports in vitro proliferation and osteogenic differentiation of goat adult mesenchymal stem cells: potential for bone tissue engineering.

PubMed

Elkhenany, Hoda; Amelse, Lisa; Lafont, Andersen; Bourdo, Shawn; Caldwell, Marc; Neilsen, Nancy; Dervishi, Enkeleda; Derek, Oshin; Biris, Alexandru S; Anderson, David; Dhar, Madhu

2015-04-01

Current treatments for bone loss injuries involve autologous and allogenic bone grafts, metal alloys and ceramics. Although these therapies have proved useful, they suffer from inherent challenges, and hence, an adequate bone replacement therapy has not yet been found. We hypothesize that graphene may be a useful nanoscaffold for mesenchymal stem cells and will promote proliferation and differentiation into bone progenitor cells. In this study, we evaluate graphene, a biocompatible inert nanomaterial, for its effect on in vitro growth and differentiation of goat adult mesenchymal stem cells. Cell proliferation and differentiation are compared between polystyrene-coated tissue culture plates and graphene-coated plates. Graphitic materials are cytocompatible and support cell adhesion and proliferation. Importantly, cells seeded on to oxidized graphene films undergo osteogenic differentiation in fetal bovine serum-containing medium without the addition of any glucocorticoid or specific growth factors. These findings support graphene's potential to act as an osteoinducer and a vehicle to deliver mesenchymal stem cells, and suggest that the combination of graphene and goat mesenchymal stem cells provides a promising construct for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

Comparative effect of soy protein, soy isoflavones, and 17beta-estradiol on bone metabolism in adult ovariectomized rats.

PubMed

Cai, David J; Zhao, Yongdong; Glasier, Jennifer; Cullen, Diane; Barnes, Stephen; Turner, Charles H; Wastney, Meryl; Weaver, Connie M

2005-05-01

This study provided a comprehensive investigation on the effect of soy protein and soy isoflavones on both calcium and bone metabolism in virgin adult rats. The measurements included bone histology, calcium kinetic modeling, calcium balance, bone densitometry, and whole body densitometry. Results confirmed the bone-preserving effect of estrogen but did not support a bone-sparing role of soy isoflavones. Several animal and short-term human studies have indicated that soy protein isolate enriched with isoflavones may be used as an alternative therapy to estrogen replacement therapy. However, none of the previous studies have investigated this estrogenic effect on both calcium and bone metabolism in animals or humans, which is essential in ascertaining the mode of action of isoflavones. This study was designed to determine the effects of soy protein versus isoflavones on calcium and bone metabolism in an ovariectomized rat model. Unmated 6-month-old ovariectomized and sham-operated female Sprague-Dawley rats were randomly assigned to nine groups (16 rats/group) and pair-fed soy- or casein-based diets with or without isoflavones for 8 weeks. A reference group was administered estrogen through subcutaneous implants (20-35 pg/liter plasma). Bone densitometry, histomorphometry, and mechanical testing were used to study bone metabolism and quality. Calcium metabolism was studied using calcium tracer balance and kinetics. After ovariectomy, estrogen prevented bone loss in trabecular bone and suppressed formation on both trabecular and cortical bone surfaces. Isoflavones given as enriched soy protein isolate or supplements did not prevent trabecular bone loss. Combining isoflavones with estrogen had no additional benefits over estrogen alone. There were no differences in response to isoflavones caused by protein source. None of the treatments significantly affected either total Ca balance or (45)Ca absorption. However, soy protein showed significant effects on reducing urinary loss of Ca in animals, irrespective of isoflavone level, perhaps because of the lower amount of sulfur-containing amino acids in soy protein. Estrogen, but not isoflavones at the levels tested, suppressed bone remodeling in both trabecular and cortical bone after ovariectomy.

Effects of Mesenchymal Stem Cell Derivatives on Hematopoiesis and Hematopoietic Stem Cells

PubMed Central

Aqmasheh, Sara; Shamsasanjan, karim; Akbarzadehlaleh, Parvin; Pashoutan Sarvar, Davod; Timari, Hamze

2017-01-01

Hematopoiesis is a balance among quiescence, self-renewal, proliferation, and differentiation, which is believed to be firmly adjusted through interactions between hematopoietic stem and progenitor cells (HSPCs) with the microenvironment. This microenvironment is derived from a common progenitor of mesenchymal origin and its signals should be capable of regulating the cellular memory of transcriptional situation and lead to an exchange of stem cell genes expression. Mesenchymal stem cells (MSCs) have self-renewal and differentiation capacity into tissues of mesodermal origin, and these cells can support hematopoiesis through release various molecules that play a crucial role in migration, homing, self-renewal, proliferation, and differentiation of HSPCs. Studies on the effects of MSCs on HSPC differentiation can develop modern solutions in the treatment of patients with hematologic disorders for more effective Bone Marrow (BM) transplantation in the near future. However, considerable challenges remain on realization of how paracrine mechanisms of MSCs act on the target tissues, and how to design a therapeutic regimen with various paracrine factors in order to achieve optimal results for tissue conservation and regeneration. The aim of this review is to characterize and consider the related aspects of the ability of MSCs secretome in protection of hematopoiesis. PMID:28761818

Skeletal (stromal) stem cells: an update on intracellular signaling pathways controlling osteoblast differentiation.

PubMed

Abdallah, Basem M; Jafari, Abbas; Zaher, Walid; Qiu, Weimin; Kassem, Moustapha

2015-01-01

Skeletal (marrow stromal) stem cells (BMSCs) are a group of multipotent cells that reside in the bone marrow stroma and can differentiate into osteoblasts, chondrocytes and adipocytes. Studying signaling pathways that regulate BMSC differentiation into osteoblastic cells is a strategy for identifying druggable targets for enhancing bone formation. This review will discuss the functions and the molecular mechanisms of action on osteoblast differentiation and bone formation; of a number of recently identified regulatory molecules: the non-canonical Notch signaling molecule Delta-like 1/preadipocyte factor 1 (Dlk1/Pref-1), the Wnt co-receptor Lrp5 and intracellular kinases. This article is part of a Special Issue entitled: Stem Cells and Bone. Copyright © 2014 Elsevier Inc. All rights reserved.

Legumain Regulates Differentiation Fate of Human Bone Marrow Stromal Cells and Is Altered in Postmenopausal Osteoporosis.

PubMed

Jafari, Abbas; Qanie, Diyako; Andersen, Thomas L; Zhang, Yuxi; Chen, Li; Postert, Benno; Parsons, Stuart; Ditzel, Nicholas; Khosla, Sundeep; Johansen, Harald Thidemann; Kjærsgaard-Andersen, Per; Delaisse, Jean-Marie; Abdallah, Basem M; Hesselson, Daniel; Solberg, Rigmor; Kassem, Moustapha

2017-02-14

Secreted factors are a key component of stem cell niche and their dysregulation compromises stem cell function. Legumain is a secreted cysteine protease involved in diverse biological processes. Here, we demonstrate that legumain regulates lineage commitment of human bone marrow stromal cells and that its expression level and cellular localization are altered in postmenopausal osteoporotic patients. As shown by genetic and pharmacological manipulation, legumain inhibited osteoblast (OB) differentiation and in vivo bone formation through degradation of the bone matrix protein fibronectin. In addition, genetic ablation or pharmacological inhibition of legumain activity led to precocious OB differentiation and increased vertebral mineralization in zebrafish. Finally, we show that localized increased expression of legumain in bone marrow adipocytes was inversely correlated with adjacent trabecular bone mass in a cohort of patients with postmenopausal osteoporosis. Our data suggest that altered proteolytic activity of legumain in the bone microenvironment contributes to decreased bone mass in postmenopausal osteoporosis. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Associations of components of sarcopenic obesity with bone health and balance in older adults.

PubMed

Scott, David; Shore-Lorenti, Catherine; McMillan, Lachlan; Mesinovic, Jakub; Clark, Ross A; Hayes, Alan; Sanders, Kerrie M; Duque, Gustavo; Ebeling, Peter R

To determine characteristics of sarcopenic obesity that are independently associated with bone health and balance in older adults. Cross-sectional study of 168 community-dwelling older adults (mean age 67.7 ± 8.4 years; 55% women). Appendicular lean mass (ALM), whole-body areal BMD (aBMD) and body fat percentage were assessed by dual-energy X-ray absorptiometry. Peripheral quantitative computed tomography assessed muscle density and cortical volumetric BMD (vBMD), area, thickness, and strength-strain index (SSI) at 66% tibial length. Hand grip strength (dynamometry) and balance path length (computerised posturography) were assessed. Obesity was defined as high body fat percentage. Greater lower-leg muscle density was associated with lower balance path length in men (r = -0.36; P < .01) and women (r = -0.40; P = < .01). Obese participants by body fat percentage did not differ to non-obese on bone indices, although a trend towards lower cortical vBMD was observed in obese compared with non-obese men (1041.4 ± 39.8 vs 1058.8 ± 36.1 mg/cm 3 ; P = .051). In multivariable models, ALM was positively associated with all bone parameters in obese women, and with whole-body aBMD, proximal tibial cortical area and SSI in non-obese women, and both non-obese and obese men (all P < .05). Lower-leg muscle density was also positively associated with cortical vBMD (B = 2.91; 95% CI 0.02, 5.80) and area (2.70; 0.06, 5.33) in obese women. Amongst components of sarcopenic obesity, higher ALM is a consistent independent predictor of better bone health. Low muscle density may also compromise bone health and balance. Interventions which improve muscle mass and composition may lower fracture risk in sarcopenic obesity. Copyright © 2017 Elsevier B.V. All rights reserved.

Ectopic bone regeneration by human bone marrow mononucleated cells, undifferentiated and osteogenically differentiated bone marrow mesenchymal stem cells in beta-tricalcium phosphate scaffolds.

PubMed

Ye, Xinhai; Yin, Xiaofan; Yang, Dawei; Tan, Jian; Liu, Guangpeng

2012-07-01

Tissue engineering approaches using the combination of porous ceramics and bone marrow mesenchymal stem cells (BMSCs) represent a promising bone substitute for repairing large bone defects. Nevertheless, optimal conditions for constructing tissue-engineered bone have yet to be determined. It remains unclear if transplantation of predifferentiated BMSCs is superior to undifferentiated BMSCs or freshly isolated bone marrow mononucleated cells (BMNCs) in terms of new bone formation in vivo. The aim of this study was to investigate the effect of in vitro osteogenic differentiation (β-glycerophosphate, dexamethasone, and l-ascorbic acid) of human BMSCs on the capability to form tissue-engineered bone in unloaded conditions after subcutaneous implantation in nude mice. After isolation from human bone marrow aspirates, BMNCs were divided into three parts: one part was seeded onto porous beta-tricalcium phosphate ceramics immediately and transplanted in a heterotopic nude mice model; two parts were expanded in vitro to passage 2 before cell seeding and in vivo transplantation, either under osteogenic conditions or not. Animals were sacrificed for micro-CT and histological evaluation at 4, 8, 12, 16, and 20 weeks postimplantation. The results showed that BMSCs differentiated into osteo-progenitor cells after induction, as evidenced by the altered cell morphology and elevated alkaline phosphatase activity and calcium deposition, but their clonogenicity, proliferating rate, and seeding efficacy were not significantly affected by osteogenic differentiation, compared with undifferentiated cells. Extensive new bone formed in the pores of all the scaffolds seeded with predifferentiated BMSCs at 4 weeks after implantation, and maintained for 20 weeks. On the contrary, scaffolds containing undifferentiated BMSCs revealed limited bone formation only in 1 out of 6 cases at 8 weeks, and maintained for 4 weeks. For scaffolds with BMNCs, woven bone was observed sporadically only in one case at 8 weeks. Overall, this study suggests that ectopic osteogenesis of cell/scaffold composites is more dependent on the in vitro expansion condition, and osteo-differentiated BMSCs hold the highest potential concerning in vivo bone regeneration.

Osteoblast Differentiation and Bone Matrix Formation In Vivo and In Vitro.

PubMed

Blair, Harry C; Larrouture, Quitterie C; Li, Yanan; Lin, Hang; Beer-Stoltz, Donna; Liu, Li; Tuan, Rocky S; Robinson, Lisa J; Schlesinger, Paul H; Nelson, Deborah J

2017-06-01

We review the characteristics of osteoblast differentiation and bone matrix synthesis. Bone in air breathing vertebrates is a specialized tissue that developmentally replaces simpler solid tissues, usually cartilage. Bone is a living organ bounded by a layer of osteoblasts that, because of transport and compartmentalization requirements, produce bone matrix exclusively as an organized tight epithelium. With matrix growth, osteoblasts are reorganized and incorporated into the matrix as living cells, osteocytes, which communicate with each other and surface epithelium by cell processes within canaliculi in the matrix. The osteoblasts secrete the organic matrix, which are dense collagen layers that alternate parallel and orthogonal to the axis of stress loading. Into this matrix is deposited extremely dense hydroxyapatite-based mineral driven by both active and passive transport and pH control. As the matrix matures, hydroxyapatite microcrystals are organized into a sophisticated composite in the collagen layer by nucleation in the protein lattice. Recent studies on differentiating osteoblast precursors revealed a sophisticated proton export network driving mineralization, a gene expression program organized with the compartmentalization of the osteoblast epithelium that produces the mature bone matrix composite, despite varying serum calcium and phosphate. Key issues not well defined include how new osteoblasts are incorporated in the epithelial layer, replacing those incorporated in the accumulating matrix. Development of bone in vitro is the subject of numerous projects using various matrices and mesenchymal stem cell-derived preparations in bioreactors. These preparations reflect the structure of bone to variable extents, and include cells at many different stages of differentiation. Major challenges are production of bone matrix approaching the in vivo density and support for trabecular bone formation. In vitro differentiation is limited by the organization and density of osteoblasts and by endogenous and exogenous inhibitors.

Osteoblast Differentiation and Bone Matrix Formation In Vivo and In Vitro

PubMed Central

Larrouture, Quitterie C.; Li, Yanan; Lin, Hang; Beer-Stoltz, Donna; Liu, Li; Tuan, Rocky S.; Robinson, Lisa J.; Schlesinger, Paul H.; Nelson, Deborah J.

2017-01-01

We review the characteristics of osteoblast differentiation and bone matrix synthesis. Bone in air breathing vertebrates is a specialized tissue that developmentally replaces simpler solid tissues, usually cartilage. Bone is a living organ bounded by a layer of osteoblasts that, because of transport and compartmentalization requirements, produce bone matrix exclusively as an organized tight epithelium. With matrix growth, osteoblasts are reorganized and incorporated into the matrix as living cells, osteocytes, which communicate with each other and surface epithelium by cell processes within canaliculi in the matrix. The osteoblasts secrete the organic matrix, which are dense collagen layers that alternate parallel and orthogonal to the axis of stress loading. Into this matrix is deposited extremely dense hydroxyapatite-based mineral driven by both active and passive transport and pH control. As the matrix matures, hydroxyapatite microcrystals are organized into a sophisticated composite in the collagen layer by nucleation in the protein lattice. Recent studies on differentiating osteoblast precursors revealed a sophisticated proton export network driving mineralization, a gene expression program organized with the compartmentalization of the osteoblast epithelium that produces the mature bone matrix composite, despite varying serum calcium and phosphate. Key issues not well defined include how new osteoblasts are incorporated in the epithelial layer, replacing those incorporated in the accumulating matrix. Development of bone in vitro is the subject of numerous projects using various matrices and mesenchymal stem cell-derived preparations in bioreactors. These preparations reflect the structure of bone to variable extents, and include cells at many different stages of differentiation. Major challenges are production of bone matrix approaching the in vivo density and support for trabecular bone formation. In vitro differentiation is limited by the organization and density of osteoblasts and by endogenous and exogenous inhibitors. PMID:27846781

Osthole Stimulates Osteoblast Differentiation and Bone Formation by Activation of β-Catenin–BMP Signaling

PubMed Central

Tang, De-Zhi; Hou, Wei; Zhou, Quan; Zhang, Minjie; Holz, Jonathan; Sheu, Tzong-Jen; Li, Tian-Fang; Cheng, Shao-Dan; Shi, Qi; Harris, Stephen E; Chen, Di; Wang, Yong-Jun

2010-01-01

Osteoporosis is defined as reduced bone mineral density with a high risk of fragile fracture. Current available treatment regimens include antiresorptive drugs such as estrogen receptor analogues and bisphosphates and anabolic agents such as parathyroid hormone (PTH). However, neither option is completely satisfactory because of adverse effects. It is thus highly desirable to identify novel anabolic agents to improve future osteoporosis treatment. Osthole, a coumarin-like derivative extracted from Chinese herbs, has been shown to stimulate osteoblast proliferation and differentiation, but its effect on bone formation in vivo and underlying mechanism remain unknown. In this study, we found that local injection of Osthole significantly increased new bone formation on the surface of mouse calvaria. Ovariectomy caused evident bone loss in rats, whereas Osthole largely prevented such loss, as shown by improved bone microarchitecture, histomorphometric parameters, and biomechanical properties. In vitro studies demonstrated that Osthole activated Wnt/β-catenin signaling, increased Bmp2 expression, and stimulated osteoblast differentiation. Targeted deletion of the β-catenin and Bmp2 genes abolished the stimulatory effect of Osthole on osteoblast differentiation. Since deletion of the Bmp2 gene did not affect Osthole-induced β-catenin expression and the deletion of the β-catenin gene inhibited Osthole-regulated Bmp2 expression in osteoblasts, we propose that Osthole acts through β-catenin–BMP signaling to promote osteoblast differentiation. Our findings demonstrate that Osthole could be a potential anabolic agent to stimulate bone formation and prevent estrogen deficiency–induced bone loss. © 2010 American Society for Bone and Mineral Research. PMID:20200936

Enamel Matrix Derivative has No Effect on the Chondrogenic Differentiation of Mesenchymal Stem Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Groeneveldt, Lisanne C.; Knuth, Callie; Witte-Bouma, Janneke

2014-09-02

Background: Treatment of large bone defects due to trauma, tumor resection, or congenital abnormalities is challenging. Bone tissue engineering using mesenchymal stem cells (MSCs) represents a promising treatment option. However, the quantity and quality of engineered bone tissue are not sufficient to fill large bone defects. The aim of this study was to determine if the addition of enamel matrix derivative (EMD) improves in vitro chondrogenic priming of MSCs to ultimately improve in vivo MSC mediated endochondral bone formation. Methods: MSCs were chondrogenically differentiated in 2.0 × 10{sup 5} cell pellets in medium supplemented with TGFβ3 in the absence ormore » presence of 1, 10, or 100 μg/mL EMD. Samples were analyzed for gene expression of RUNX2, Col II, Col X, and Sox9. Protein and glycoaminoglycan (GAG) production were also investigated via DMB assays, histology, and immunohistochemistry. Osteogenic and adipogenic differentiation capacity were also assessed. Results: The addition of EMD did not negatively affect chondrogenic differentiation of adult human MSCs. EMD did not appear to alter GAG production or expression of chondrogenic genes. Osteogenic and adipogenic differentiation were also unaffected though a trend toward decreased adipogenic gene expression was observed. Conclusion: EMD does not affect chondrogenic differentiation of adult human MSCs. As such the use of EMD in combination with chondrogenically primed MSCs for periodontal bone tissue repair is unlikely to have negative effects on MSC differentiation.« less

Subprimal purchasing and merchandising decisions for pork: relationship to retail value.

PubMed

Lorenzen, C L; Walter, J P; Dockerty, T R; Griffin, D B; Johnson, H K; Savell, J W

1996-01-01

To assess retail value and profitability, cutting test data were obtained in a simulated retail cutting room for boxed pork subprimals, bone-in loins (n = 180), boneless loins (n = 94), Boston butts (n = 148), fresh hams (n = 28), and boneless hams (n = 23). Processing times (seconds) and retail weights (kilograms) were used to determine relative value. Cutting style affected (P < .05) value differential (US$/subprimal) for bone-in and boneless loins. When cutting styles within subprimals were pooled, value differential was affected (P < .05) by purchasing specification for bone-in loins, boneless loins, Boston butts, and inside fresh hams. Processing bone-in loins to a boneless end point produced a greater (P < .05) value differential and percentage of gross margin than a bone-in retail end point. Bone-in loins fabricated to a boneless retail end point produced a greater (P < .05) value differential and percentage of gross margin than boneless loins fabricated to the same end point. The increase in retail value can be attributed to the increased number and weight of retail cuts produced from bone-in loins. The thick, boneless loin cutting style produced a greater (P < .05) value differential and percentage of gross margin as a result of a lower (P < .05) cost of fabrication and increased value of retail cuts than the thin, boneless cutting style. In general, boneless pork cutting methods were more profitable than bone-in cutting methods regardless of subprimal.

The orally available Btk inhibitor ibrutinib (PCI-32765) protects against osteoclast-mediated bone loss.

PubMed

Shinohara, Masahiro; Chang, Betty Y; Buggy, Joseph J; Nagai, Yusuke; Kodama, Tatsuhiko; Asahara, Hiroshi; Takayanagi, Hiroshi

2014-03-01

Bone-resorbing osteoclasts play an essential role in normal bone homeostasis, as well as in various bone disorders such as osteoporosis and rheumatoid arthritis. Previously we showed that the Tec family of tyrosine kinases is essential for the differentiation of osteoclasts and the inhibition of Btk is a promising strategy for the prevention of the bone loss in osteoclast-associated bone disorders. Here we demonstrate that an orally available Btk inhibitor, ibrutinib (PCI-32765), suppresses osteoclastic bone resorption by inhibiting both osteoclast differentiation and function. Ibrutinib downregulated the expression of NFATc1, the key transcription factor for osteoclastogenesis, and disrupted the formation of the actin ring in mature osteoclasts. In addition, genome-wide screening revealed that Btk regulates the expression of the genes involved in osteoclast differentiation and function in both an NFATc1-dependent and -independent manner. Finally, we showed that ibrutinib administration ameliorated the bone loss that developed in a RANKL-induced osteoporosis mouse model. Thus, this study suggests ibrutinib to be a promising therapeutic agent for osteoclast-associated bone diseases. Copyright © 2013 Elsevier Inc. All rights reserved.

BMP-2 Derived Peptide and Dexamethasone Incorporated Mesoporous Silica Nanoparticles for Enhanced Osteogenic Differentiation of Bone Mesenchymal Stem Cells.

PubMed

Zhou, Xiaojun; Feng, Wei; Qiu, Kexin; Chen, Liang; Wang, Weizhong; Nie, Wei; Mo, Xiumei; He, Chuanglong

2015-07-29

Bone morphogenetic protein-2 (BMP-2), a growth factor that induces osteoblast differentiation and promotes bone regeneration, has been extensively investigated in bone tissue engineering. The peptides of bioactive domains, corresponding to residues 73-92 of BMP-2 become an alternative to reduce adverse side effects caused by the use of high doses of BMP-2 protein. In this study, BMP-2 peptide functionalized mesoporous silica nanoparticles (MSNs-pep) were synthesized by covalently grafting BMP-2 peptide on the surface of nanoparticles via an aminosilane linker, and dexamethasone (DEX) was then loaded into the channel of MSNs to construct nanoparticulate osteogenic delivery systems (DEX@MSNs-pep). The in vitro cell viability of MSNs-pep was tested with bone mesenchymal stem cells (BMSCs) exposure to different particle concentrations, revealing that the functionalized MSNs had better cytocompatibility than their bare counterparts, and the cellular uptake efficiency of MSNs-pep was remarkably larger than that of bare MSNs. The in vitro results also show that the MSNs-pep promoted osteogenic differentiation of BMSCs in terms of the levels of alkaline phosphatase (ALP) activity, calcium deposition, and expression of bone-related protein. Moreover, the osteogenic differentiation of BMSCs can be further enhanced by incorporating of DEX into MSNs-pep. After intramuscular implantation in rats for 3 weeks, the computed tomography (CT) images and histological examination indicate that this nanoparticulate osteogenic delivery system induces effective osteoblast differentiation and bone regeneration in vivo. Collectively, the BMP-2 peptide and DEX incorporated MSNs can act synergistically to enhance osteogenic differentiation of BMSCs, which have potential applications in bone tissue engineering.

A Cbfa1-dependent genetic pathway controls bone formation beyond embryonic development

PubMed Central

Ducy, Patricia; Starbuck, Michael; Priemel, Matthias; Shen, Jianhe; Pinero, Gerald; Geoffroy, Valerie; Amling, Michael; Karsenty, Gerard

1999-01-01

The molecular mechanisms controlling bone extracellular matrix (ECM) deposition by differentiated osteoblasts in postnatal life, called hereafter bone formation, are unknown. This contrasts with the growing knowledge about the genetic control of osteoblast differentiation during embryonic development. Cbfa1, a transcriptional activator of osteoblast differentiation during embryonic development, is also expressed in differentiated osteoblasts postnatally. The perinatal lethality occurring in Cbfa1-deficient mice has prevented so far the study of its function after birth. To determine if Cbfa1 plays a role during bone formation we generated transgenic mice overexpressing Cbfa1 DNA-binding domain (ΔCbfa1) in differentiated osteoblasts only postnatally. ΔCbfa1 has a higher affinity for DNA than Cbfa1 itself, has no transcriptional activity on its own, and can act in a dominant-negative manner in DNA cotransfection assays. ΔCbfa1-expressing mice have a normal skeleton at birth but develop an osteopenic phenotype thereafter. Dynamic histomorphometric studies show that this phenotype is caused by a major decrease in the bone formation rate in the face of a normal number of osteoblasts thus indicating that once osteoblasts are differentiated Cbfa1 regulates their function. Molecular analyses reveal that the expression of the genes expressed in osteoblasts and encoding bone ECM proteins is nearly abolished in transgenic mice, and ex vivo assays demonstrated that ΔCbfa1-expressing osteoblasts were less active than wild-type osteoblasts. We also show that Cbfa1 regulates positively the activity of its own promoter, which has the highest affinity Cbfa1-binding sites characterized. This study demonstrates that beyond its differentiation function Cbfa1 is the first transcriptional activator of bone formation identified to date and illustrates that developmentally important genes control physiological processes postnatally. PMID:10215629

Organotins Are Potent Activators of PPARγ and Adipocyte Differentiation in Bone Marrow Multipotent Mesenchymal Stromal Cells

PubMed Central

Yanik, Susan C.; Baker, Amelia H.; Mann, Koren K.; Schlezinger, Jennifer J.

2011-01-01

Adipocyte differentiation in bone marrow is potentially deleterious to both bone integrity and lymphopoiesis. Here, we examine the hypothesis that organotins, common environmental contaminants that are dual ligands for peroxisome proliferator–activated receptor (PPAR) γ and its heterodimerization partner retinoid X receptor (RXR), are potent activators of bone marrow adipogenesis. A C57Bl/6-derived bone marrow multipotent mesenchymal stromal cell (MSC) line, BMS2, was treated with rosiglitazone, a PPARγ agonist, bexarotene, an RXR agonist, or a series of organotins. Rosiglitazone and bexarotene potently activated adipocyte differentiation; however, bexarotene had a maximal efficacy of only 20% of that induced by rosiglitazone. Organotins (tributyltin [TBT], triphenyltin, and dibutyltin) also stimulated adipocyte differentiation (EC50 of 10–20nM) but with submaximal, structure-dependent efficacy. In coexposures, both bexarotene and TBT enhanced rosiglitazone-induced adipogenesis. To investigate the contribution of PPARγ to TBT-induced adipogenesis, we examined expression of PPARγ2, as well as its transcriptional target FABP4. TBT-induced PPARγ2 and FABP4 protein expression with an efficacy intermediate between rosiglitazone and bexarotene, similar to lipid accumulation. A PPARγ antagonist and PPARγ-specific small hairpin RNA suppressed TBT-induced differentiation, although to a lesser extent than rosiglitazone-induced differentiation, suggesting that TBT may engage alternate pathways. TBT and bexarotene, but not rosiglitazone, also induced the expression of TGM2 (an RXR target) and ABCA1 (a liver X receptor target). The results show that an environmental contaminant, acting with the same potency as a therapeutic drug, induces PPARγ-dependent adipocyte differentiation in bone marrow MSCs. Activation of multiple nuclear receptor pathways by organotins may have significant implications for bone physiology. PMID:21622945

Arctigenin suppresses receptor activator of nuclear factor κB ligand (RANKL)-mediated osteoclast differentiation in bone marrow-derived macrophages.

PubMed

Kim, A-Ram; Kim, Hyuk Soon; Lee, Jeong Min; Choi, Jung Ho; Kim, Se Na; Kim, Do Kyun; Kim, Ji Hyung; Mun, Se Hwan; Kim, Jie Wan; Jeon, Hyun Soo; Kim, Young Mi; Choi, Wahn Soo

2012-05-05

Osteoclasts, multinucleated bone-resorbing cells, are closely associated with bone diseases such as rheumatoid arthritis and osteoporosis. Osteoclasts are derived from hematopoietic precursor cells, and their differentiation is mediated by two cytokines, including macrophage colony stimulating factor and receptor activator of nuclear factor κB ligand (RANKL). Previous studies have shown that arctigenin exhibits an anti-inflammatory effect. However, the effect of arctigenin on osteoclast differentiation is yet to be elucidated. In this study, we found that arctigenin inhibited RANKL-mediated osteoclast differentiation in bone marrow macrophages in a dose-dependent manner and suppressed RANKL-mediated bone resorption. Additionally, the expression of typical marker proteins, such as NFATc1, c-Fos, TRAF6, c-Src, and cathepsin K, were significantly inhibited. Arctigenin inhibited the phosphorylation of Erk1/2, but not p38 and JNK, in a dose-dependent manner. Arctigenin also dramatically suppressed immunoreceptor tyrosine-based activation motif-mediated costimulatory signaling molecules, including Syk and PLCγ2, and Gab2. Notably, arctigenin inhibited the activation of Syk through RANKL stimulation. Furthermore, arctigenin prevented osteoclast differentiation in the calvarial bone of mice following stimulation with lipopolysaccharide. Our results show that arctigenin inhibits osteoclast differentiation in vitro and in vivo. Therefore, arctigenin may be useful for treating rheumatoid arthritis and osteoporosis. Copyright © 2012 Elsevier B.V. All rights reserved.

Assessment of the Role of Noni (Morinda citrifolia) Juice for Inducing Osteoblast Differentiation in Isolated Rat Bone Marrow Derived Mesenchymal Stem Cells

PubMed Central

Hussain, Sharmila; Tamizhselvi, Ramasamy; George, Leema; Manickam, Venkatraman

2016-01-01

Background and Objectives Morinda citrifolia (Noni), an important traditional medicinal plant still used in patients with bone fractures or dislocation to promote connective tissue repair and to reduce inflammation. However, the effects of Noni on bone metabolism and whether it influences the osteogenic differentiation is yet to be clarified. In this study, we investigated the effect of Morinda citrifolia (Noni) juice on the proliferation rate of rat bone marrow derived mesenchymal stem cells (BMSC) and the osteoblastic differentiation as shown by alkaline phosphatase (ALP), Runt-related transcription factor 2 (Runx2) and osteocalcin (OCN) mRNA expression in vitro. Methods and Results Treatment with 200 μg/ml Noni juice enhanced the proliferation rate of the BMSC and also upregulated the osteogenic differentiation marker genes ALP and OCN, and Runx2 measured by RTPCR. Consistent with these results collagen scaffolds implanted in vivo, which were loaded with BMSC pre-exposed to Noni, showed increased bone density measured by computed tomography and histological analysis revealed neo-angiogenesis for bone formation. Conclusions These results suggest that Noni stimulates osteoblastogenesis and can be used as adjuvant natural medicine for bone diseases such as osteoporosis. PMID:27572713

Assessment of the Role of Noni (Morinda citrifolia) Juice for Inducing Osteoblast Differentiation in Isolated Rat Bone Marrow Derived Mesenchymal Stem Cells.

PubMed

Hussain, Sharmila; Tamizhselvi, Ramasamy; George, Leema; Manickam, Venkatraman

2016-11-30

Morinda citrifolia (Noni), an important traditional medicinal plant still used in patients with bone fractures or dislocation to promote connective tissue repair and to reduce inflammation. However, the effects of Noni on bone metabolism and whether it influences the osteogenic differentiation is yet to be clarified. In this study, we investigated the effect of Morinda citrifolia (Noni) juice on the proliferation rate of rat bone marrow derived mesenchymal stem cells (BMSC) and the osteoblastic differentiation as shown by alkaline phosphatase (ALP), Runt-related transcription factor 2 (Runx2) and osteocalcin (OCN) mRNA expression in vitro . Treatment with 200 μg/ml Noni juice enhanced the proliferation rate of the BMSC and also upregulated the osteogenic differentiation marker genes ALP and OCN, and Runx2 measured by RTPCR. Consistent with these results collagen scaffolds implanted in vivo , which were loaded with BMSC pre-exposed to Noni, showed increased bone density measured by computed tomography and histological analysis revealed neo-angiogenesis for bone formation. These results suggest that Noni stimulates osteoblastogenesis and can be used as adjuvant natural medicine for bone diseases such as osteoporosis.

Human decellularized bone scaffolds from aged donors show improved osteoinductive capacity compared to young donor bone

PubMed Central

Smith, Christopher A.; Board, Tim N.; Rooney, Paul; Eagle, Mark J.; Richardson, Stephen M.

2017-01-01

To improve the safe use of allograft bone, decellularization techniques may be utilized to produce acellular scaffolds. Such scaffolds should retain their innate biological and biomechanical capacity and support mesenchymal stem cell (MSC) osteogenic differentiation. However, as allograft bone is derived from a wide age-range, this study aimed to determine whether donor age impacts on the ability an osteoinductive, acellular scaffold produced from human bone to promote the osteogenic differentiation of bone marrow MSCs (BM-MSC). BM-MSCs from young and old donors were seeded on acellular bone cubes from young and old donors undergoing osteoarthritis related hip surgery. All combinations resulted in increased osteogenic gene expression, and alkaline phosphatase (ALP) enzyme activity, however BM-MSCs cultured on old donor bone displayed the largest increases. BM-MSCs cultured in old donor bone conditioned media also displayed higher osteogenic gene expression and ALP activity than those exposed to young donor bone conditioned media. ELISA and Luminex analysis of conditioned media demonstrated similar levels of bioactive factors between age groups; however, IGF binding protein 1 (IGFBP1) concentration was significantly higher in young donor samples. Additionally, structural analysis of old donor bone indicated an increased porosity compared to young donor bone. These results demonstrate the ability of a decellularized scaffold produced from young and old donors to support osteogenic differentiation of cells from young and old donors. Significantly, the older donor bone produced greater osteogenic differentiation which may be related to reduced IGFBP1 bioavailability and increased porosity, potentially explaining the excellent clinical results seen with the use of allograft from aged donors. PMID:28505164

The Role of the Progressive Ankylosis Protein (ANK) in Adipogenic/Osteogenic Fate Decision of Precursor Cells

PubMed Central

Minashima, Takeshi; Quirno, Martin; Lee, You Jin; Kirsch, Thorsten

2017-01-01

The progressive ankylosis protein (ANK) is a transmembrane protein that transports intracellular pyrophosphate (PPi) to the extracellular milieu. In this study we show increased fatty degeneration of the bone marrow of adult ank/ank mice, which lack a functional ANK protein. In addition, isolated bone marrow stromal cells (BMSCs) isolated from ank/ank mice showed a decreased proliferation rate and osteogenic differentiation potential, and an increased adipogenic differentiation potential compared to BMSCs isolated from wild type (WT) littermates. Wnt signaling pathway PCR array analysis revealed that Wnt ligands, Wnt receptors and Wnt signaling proteins that stimulate osteoblast differentiation were expressed at markedly lower levels in ank/ank BMSCs than in WT BMSCs. Lack of ANK function also resulted in impaired bone fracture healing, as indicated by a smaller callus formed and delayed bone formation in the callus site. Whereas 5 weeks after fracture, the fractured bone in WT mice was further remodeled and restored to original shape, the fractured bone in ank/ank mice was not fully restored and remodeled to original shape. In conclusion, our study provides evidence that ANK plays a critical role in the adipogenic/osteogenic fate decision of adult mesenchymal precursor cells. ANK functions in precursor cells are required for osteogenic differentiation of these cells during adult bone homeostasis and repair, whereas lack of ANK functions favors adipogenic differentiation. PMID:28286238

Chronic High Dose Alcohol Induces Osteopenia via Activation of mTOR Signaling in Bone Marrow Mesenchymal Stem Cells.

PubMed

Liu, Yao; Kou, Xiaoxing; Chen, Chider; Yu, Wenjing; Su, Yingying; Kim, Yong; Shi, Songtao; Liu, Yi

2016-08-01

Chronic consumption of excessive alcohol results in reduced bone mass, impaired bone structure, and increased risk of bone fracture. However, the mechanisms underlying alcohol-induced osteoporosis are not fully understood. Here, we show that high dose chronic alcohol consumption reduces osteogenic differentiation and enhances adipogenic differentiation of bone marrow mesenchymal stem cells (BMMSCs), leading to osteopenia in a mouse model. Mechanistically, impaired osteo/adipogenic lineage differentiation of BMMSCs is due to activation of a phosphatidylinositide 3-kinase/AKT/mammalian target of rapamycin (mTOR) signaling cascade, resulting in downregulation of runt-related transcription factor 2 and upregulation of peroxisome proliferator-activated receptor gamma via activation of p70 ribosomal protein S6 kinase. Blockage of the mTOR pathway by rapamycin treatment ameliorates alcohol-induced osteopenia by rescuing impaired osteo/adipogenic lineage differentiation of BMMSCs. In this study, we identify a previously unknown mechanism by which alcohol impairs BMMSC lineage differentiation and reveal a potential rapamycin-based drug therapy for alcohol-induced osteoporosis. Stem Cells 2016;34:2157-2168. © 2016 AlphaMed Press.

Galectin-3 is essential for proper bone cell differentiation and activity, bone remodeling and biomechanical competence in mice.

PubMed

Iacobini, Carla; Fantauzzi, Claudia Blasetti; Bedini, Rossella; Pecci, Raffaella; Bartolazzi, Armando; Amadio, Bruno; Pesce, Carlo; Pugliese, Giuseppe; Menini, Stefano

2018-02-09

Galectin-3 is constitutively expressed in bone cells and was recently shown to modulate osteogenic transdifferentiation of vascular smooth muscle cells and atherosclerotic calcification. However, the role of galectin-3 in bone physiology is largely undefined. To address this issue, we analyzed (1) the skeletal features of 1-, 3- and 6-month-old galectin-3 null (Lgals3 -/- ) and wild type (WT) mice and (2) the differentiation and function of osteoblasts and osteoclasts derived from these animals. Long bone phenotype, gene expression profile, and remodeling were investigated by micro-computed tomography, real time-PCR, static and dynamic histomorphometry, and assessment of biochemical markers of bone resorption and formation. Bone competence was also evaluated by biomechanical testing at 3 months. In vitro, the effects of galectin-3 deficiency on bone cell differentiation and function were investigated by assessing (a) gene expression of osteoblast markers, alkaline phosphatase activity, mineralization assay, and WNT/β-catenin signaling (of which galectin-3 is a known regulator) in osteoblasts; and (b) tartrate-resistant acid phosphatase activity and bone resorption activity in osteoclasts. Lgals3 -/- mice revealed a wide range of age-dependent alterations including lower bone formation and higher bone resorption, accelerated age-dependent trabecular bone loss (p < 0.01 vs. WT at 3 months) and reduced bone strength (p < 0.01 vs. WT at 3 months). These abnormalities were accompanied by a steady inflammatory state, as revealed by higher bone expression of the pro-inflammatory cytokines interleukin (IL)-1β and IL-6 (p < 0.001 vs. WT at 3 months), increased content of osteal macrophages (p < 0.01 vs. WT at 3 months), and reduced expression of markers of alternative (M2) macrophage activation. Lgals3 -/- osteoblasts and osteoclasts showed impaired terminal differentiation, reduced mineralization capacity (p < 0.01 vs. WT cells) and resorption activity (p < 0.01 vs. WT cells). Mechanistically, impaired differentiation and function of Lgals3 -/- osteoblasts was associated with altered WNT/β-catenin signaling (p < 0.01 vs. WT cells). These data provide evidence for a contribution of galectin-3 to bone cell maturation and function, bone remodeling, and biomechanical competence, thus identifying galectin-3 as a promising therapeutic target for age-related disorders of bone remodeling. Copyright © 2018. Published by Elsevier Inc.

Scaffold preferences of mesenchymal stromal cells and adipose-derived stem cells from green fluorescent protein transgenic mice influence the tissue engineering of bone.

PubMed

Wittenburg, Gretel; Flade, Viktoria; Garbe, Annette I; Lauer, Günter; Labudde, Dirk

2014-05-01

We have analysed the growth and differentiation of mesenchymal stromal cells (MSC) from bone marrow, and of adipose derived stem cells (ASC) from murine abdominal fat tissue, of green fluorescent protein (GFP) transgenic animals grown directly on two types of hydroxyapatite ceramic bone substitutes. BONITmatrix® and NanoBone® have specific mechanical and physiochemical properties such as porosity and an inner surface that influence cellular growth. Both MSC and ASC were separately seeded on 200mg of each biomaterial and cultured for 3 weeks under osteogenic differentiation conditions. The degree of mineralisation was assessed by alizarin red dye and the specific alkaline phosphatase activity of the differentiated cells. The morphology of the cells was examined by scanning electron microscopy and confocal microscopy. The osteoblastic phenotype of the cells was confirmed by analysing the expression of bone-specific genes (Runx2, osteocalcin, osteopontin, and osteonectin) by semiquantitative reverse transcriptase polymerase chain reaction (PCR). Comparison of BONITmatrix® and NanoBone® showed cell type-specific preferences in terms of osteogenic differentiation. MSC-derived osteoblast-like cells spread optimally on the surface of NanoBone® but not BONITmatrix® granules. In contrast BONITmatrix® granules conditioned the growth of osteoblast-like cells derived from ASC. The osteoblastic phenotype of the cultured cells on all matrices was confirmed by specific gene expression. Our results show that the in vitro growth and osteogenic differentiation of murine MSC or ASC of GFP transgenic mice are distinctly influenced by the ceramic substratum. While NanoBone® granules support the proliferation and differentiation of murine MSC isolated from bone marrow, the growth of murine ASC is supported by BONITmatrix® granules. NanoBone® is therefore recommended for use as scaffold in tissue engineering that requires MSC, whereas ASC can be combined with BONITmatrix® for in vitro bone engineering. Copyright © 2014 The British Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

Selective Resistance of CD44hi T Cells to p53 Dependent Cell Death Results in Persistence of Immunologic Memory after Total Body Irradiation1,2,3

PubMed Central

Yao, Zhenyu; Jones, Jennifer; Kohrt, Holbrook; Strober, Samuel

2011-01-01

Our previous studies showed that treatment of mice with total body irradiation (TBI) or total lymphoid tissue irradiation (TLI) markedly changes the balance of residual T cell subsets to favor CD4+CD44hi natural killer T (NKT) cells due to differential resistance of the latter subset to cell death. The object of the current study was to further elucidate the changed balance and mechanisms of differential radioresistance of T cell subsets after graded doses of TBI. The experimental results show that CD4+ T cells were markedly more resistant than CD8+ T cells, and CD44hi T cells including NKT cells and memory T cells were markedly more resistant than CD44lo (naïve) T cells. The memory T cells immunized to alloantigens persisted even after myeloabloative (1,000cGy) TBI, and were able to prevent engraftment of bone marrow transplants. Although T cell death after 1,000cGy was prevented in p53−/− mice, there was progressive T cell death in p53−/− mice at higher doses. Whereas, p53 dependent T cell death changed the balance of subsets, the p53 independent T cell death did not. In conclusion, resistance of CD44hi T cells to p53 dependent cell death results in the persistence of immunological memory after TBI, and can explain the immune mediated rejection of marrow transplants in sensitized recipients. PMID:21930972

Differential effects of calorie restriction and involuntary wheel running on body composition and bone structure in diet-induced obese rats

USDA-ARS?s Scientific Manuscript database

Weight reduction is recommended to reduce obesity-related health disorders. This study investigated the differential effects of weight reduction through caloric restriction and/or physical activity on bone structure and molecular characteristics of bone metabolism in an obese rat model. We tested th...

Monosodium urate monohydrate crystals inhibit osteoblast viability and function: implications for development of bone erosion in gout.

PubMed

Chhana, Ashika; Callon, Karen E; Pool, Bregina; Naot, Dorit; Watson, Maureen; Gamble, Greg D; McQueen, Fiona M; Cornish, Jillian; Dalbeth, Nicola

2011-09-01

Bone erosion is a common manifestation of chronic tophaceous gout. To investigate the effects of monosodium urate monohydrate (MSU) crystals on osteoblast viability and function. The MTT assay and flow cytometry were used to assess osteoblast cell viability in the MC3T3-E1 and ST2 osteoblast-like cell lines, and primary rat and primary human osteoblasts cultured with MSU crystals. Quantitative real-time PCR and von Kossa stained mineralised bone formation assays were used to assess the effects of MSU crystals on osteoblast differentiation using MC3T3-E1 cells. The numbers of osteoblasts and bone lining cells were quantified in bone samples from patients with gout. MSU crystals rapidly reduced viability in all cell types in a dose-dependent manner. The inhibitory effect on cell viability was independent of crystal phagocytosis and was not influenced by differing crystal length or addition of serum. Long-term culture of MC3T3-E1 cells with MSU crystals showed a reduction in mineralisation and decreased mRNA expression of genes related to osteoblast differentiation such as Runx2, Sp7 (osterix), Ibsp (bone sialoprotein), and Bglap (osteocalcin). Fewer osteoblast and lining cells were present on bone directly adjacent to gouty tophus than bone unaffected by tophus in patients with gout. MSU crystals have profound inhibitory effects on osteoblast viability and differentiation. These data suggest that bone erosion in gout occurs at the tophus-bone interface through alteration of physiological bone turnover, with both excessive osteoclast formation, and reduced osteoblast differentiation from mesenchymal stem cells.

Natural Ca Isotope Composition of Urine as a Rapid Measure of Bone Mineral Balance

NASA Astrophysics Data System (ADS)

Skulan, J.; Gordon, G. W.; Morgan, J.; Romaniello, S. J.; Smith, S. M.; Anbar, A. D.

2011-12-01

Naturally occurring stable Ca isotope variations in urine are emerging as a powerful tool to detect changes in bone mineral balance. Bone formation depletes soft tissue of light Ca isotopes while bone resorption releases isotopically light Ca into soft tissue. Previously published work found that variations in Ca isotope composition could be detected at 4 weeks of bed rest in a 90-day bed rest study (data collected at 4, 8 and 12 weeks). A new 30-day bed rest study involved 12 patients on a controlled diet, monitored for 7 days prior to bed rest and 7 days post bed rest. Samples of urine, blood and food were collected throughout the study. Four times daily blood samples and per void urine samples were collected to monitor diurnal or high frequency variations. An improved chemical purification protocol, followed by measurement using multiple collector inductively coupled plasma mass spectrometry (MC-ICP-MS) allowed accurate and precise determinations of mass-dependent Ca isotope variations in these biological samples to better than ±0.2% (δ44/42Ca) on <25 μg of Ca. Results from this new study show that Ca isotope ratios shift in a direction consistent with net bone loss after just 7 days, long before detectible changes in bone density by X-ray measurements occur. Consistent with this interpretation, the Ca isotope variations track changes observed in N-teleopeptide, a bone resorption biomarker. Bone-specific alkaline phosphatase, a bone formation biomarker, is unchanged over this period. Ca isotopes can in principle be used to quantify net changes in bone mass. Using a mass-balance model, our results indicate an average loss of 0.62 ± 0.16 % in bone mass over the course of this 30-day study. This is consistent with the rate of bone loss in longer-term studies as seen by X-ray measurements. This Ca isotope technique should accelerate the pace of discovery of new treatments for bone disease and provide novel insights into the dynamics of bone metabolism.

Using Natural Stable Calcium Isotopes to Rapidly Assess Changes in Bone Mineral Balance Using a Bed Rest Model to Induce Bone Loss

NASA Technical Reports Server (NTRS)

Morgan, J. L. L.; Skulan, J. L.; Gordon, G. E.; Smith, Scott M.; Romaniello, S. J.; Anbar, A. D.

2012-01-01

Metabolic bone diseases like osteoporosis result from the disruption of normal bone mineral balance (BMB) resulting in bone loss. During spaceflight astronauts lose substantial bone. Bed rest provides an analog to simulate some of the effects of spaceflight; including bone and calcium loss and provides the opportunity to evaluate new methods to monitor BMB in healthy individuals undergoing environmentally induced-bone loss. Previous research showed that natural variations in the Ca isotope ratio occur because bone formation depletes soft tissue of light Ca isotopes while bone resorption releases that isotopically light Ca back into soft tissue (Skulan et al, 2007). Using a bed rest model, we demonstrate that the Ca isotope ratio of urine shifts in a direction consistent with bone loss after just 7 days of bed rest, long before detectable changes in bone mineral density (BMD) occur. The Ca isotope variations tracks changes observed in urinary N-teleopeptide, a bone resorption biomarker. Bone specific alkaline phosphatase, a bone formation biomarker, is unchanged. The established relationship between Ca isotopes and BMB can be used to quantitatively translate the changes in the Ca isotope ratio to changes in BMD using a simple mathematical model. This model predicts that subjects lost 0.25 0.07% ( SD) of their bone mass from day 7 to day 30 of bed rest. Given the rapid signal observed using Ca isotope measurements and the potential to quantitatively assess bone loss; this technique is well suited to study the short-term dynamics of bone metabolism.

The effect of feeding different sugar-sweetened beverages to growing female Sprague-Dawley rats on bone mass and strength.

PubMed

Tsanzi, Embedzayi; Light, Heather R; Tou, Janet C

2008-05-01

Consumption of sugar beverages has increased among adolescents. Additionally, the replacement of sucrose with high fructose corn syrup (HFCS) as the predominant sweetener has resulted in higher fructose intake. Few studies have investigated the effect of drinking different sugar-sweetened beverages on bone, despite suggestions that sugar consumption negatively impacts mineral balance. The objective of this study was to determine the effect of drinking different sugar-sweetened beverages on bone mass and strength. Adolescent (age 35d) female Sprague-Dawley rats were randomly assigned (n=8-9/group) to consume deionized distilled water (ddH2O, control) or ddH2O containing 13% w/v glucose, sucrose, fructose or high fructose corn syrup (HFCS-55) for 8weeks. Tibia and femur measurements included bone morphometry, bone turnover markers, determination of bone mineral density (BMD) and bone mineral content (BMC) by dual energy X-ray absorptiometry (DXA) and bone strength by three-point bending test. The effect of sugar-sweetened beverage consumption on mineral balance, urinary and fecal calcium (Ca) and phosphorus (P) was measured by inductively coupled plasma optical emission spectrometry. The results showed no difference in the bone mass or strength of rats drinking the glucose-sweetened beverage despite their having the lowest food intake, but the highest beverage and caloric consumption. Only in comparisons among the rats provided sugar-sweetened beverage were femur and tibia BMD lower in rats drinking the glucose-sweetened beverage. Differences in bone and mineral measurements appeared most pronounced between rats drinking glucose versus fructose-sweetened beverages. Rats provided the glucose-sweetened beverage had reduced femur and tibia total P, reduced P and Ca intake and increased urinary Ca excretion compared to the rats provided the fructose-sweetened beverage. The results suggested that glucose rather than fructose exerted more deleterious effects on mineral balance and bone.

In vitro comparison of the efficacy of TGF-β1 and PDGF-BB in combination with freeze-dried bone allografts for induction of osteogenic differentiation in MG-63 osteoblast-like cells.

PubMed

Vahabi, Surena; Torshabi, Maryam; Esmaeil Nejad, Azadeh

2016-12-01

Predictable regeneration of alveolar bone defects has always been a challenge in implant dentistry. Bone allografts are widely used bone substitutes with controversial osteoinductive activity. This in vitro study aimed to assess the osteogenic potential of some commercially available freeze-dried bone allografts supplemented with human recombinant platelet-derived growth factor-BB and transforming growth factor beta-1. Cell viability, mineralization, and osteogenic gene expression of MG-63 osteoblast-like cells were compared among the allograft alone, allograft/platelet-derived growth factor-BB, allograft/transforming growth factor beta-1, and allograft/platelet-derived growth factor-BB/transforming growth factor beta-1 groups. The methyl thiazol tetrazolium assay, real-time quantitative reverse transcription polymerase chain reaction and alizarin red staining were performed, respectively, for assessment of cell viability, differentiation, and mineralization at 24-72 h post treatment. The allograft with greater cytotoxic effect on MG-63 cells caused the lowest differentiation among the groups. In comparison with allograft alone, allograft/transforming growth factor beta-1, and allograft/transforming growth factor beta-1/platelet-derived growth factor-BB caused significant upregulation of bone sialoprotein and osteocalcin osteogenic mid-late marker genes, and resulted in significantly higher amounts of calcified nodules especially in mineralized non-cytotoxic allograft group. Supplementation of platelet-derived growth factor-BB alone in 5 ng/mL concentration had no significant effect on differentiation or mineralization markers. According to the results, transforming growth factor beta-1 acts synergistically with bone allografts to enhance the osteogenic differentiation potential. Therefore, this combination may be useful for rapid transformation of undifferentiated cells into bone-forming cells for bone regeneration. However, platelet-derived growth factor-BB supplementation did not support this synergistic ability to enhance osteogenic differentiation and thus, further investigations are required.

Assessment of the Genetic Variation in Bone Fracture Healing

DTIC Science & Technology

2004-10-01

decrease in the expression of the major mRNA markers of differentiated osteogenic cells (osteocalcin type I collagen and bone sialoprotein ) (Kon et al., 9...and higher levels of bone sialoprotein which are seen both during cartilage hypertrophy and as a marker of early to mid osteogenic differentiation...Biochem 89(2):401-26. Barnes GL, Della Torre T, Sommer B, Young MF, Gerstenfeld LC. 2002. Transcriptional regulation restricting bone sialoprotein gene

Osteoporosis

MedlinePlus

Osteoporosis is a condition that leads to loss of bone mass. From the outside, osteoporotic bone is ... disease. Prevention is the best measure for treating osteoporosis by eating a recommended balanced diet including foods ...

A Conditional Knockout Mouse Model Reveals a Critical Role of PKD1 in Osteoblast Differentiation and Bone Development

PubMed Central

Li, Shao; Xu, Wanfu; Xing, Zhe; Qian, Jiabi; Chen, Liping; Gu, Ruonan; Guo, Wenjing; Lai, Xiaoju; Zhao, Wanlu; Li, Songyu; Wang, Yaodong; Wang, Q. Jane; Deng, Fan

2017-01-01

The protein kinase D family of serine/threonine kinases, particularly PKD1, has been implicated in the regulation of a complex array of fundamental biological processes. However, its function and mechanism underlying PKD1-mediated the bone development and osteoblast differentiation are not fully understood. Here we demonstrate that loss of PKD1 function led to impaired bone development and osteoblast differentiation through STAT3 and p38 MAPK signaling using in vitro and in vivo bone-specific conditional PKD1-knockout (PKD1-KO) mice models. These mice developed markedly craniofacial dysplasia, scapula dysplasia, long bone length shortage and body weight decrease compared with wild-type littermates. Moreover, deletion of PKD1 in vivo reduced trabecular development and activity of osteoblast development, confirmed by Micro-CT and histological staining as well as expression of osteoblastic marker (OPN, Runx2 and OSX). Mechanistically, loss of PKD1 mediated the downregulation of osteoblast markers and impaired osteoblast differentiation through STAT3 and p38 MAPK signaling pathways. Taken together, these results demonstrated that PKD1 contributes to the osteoblast differentiation and bone development via elevation of osteoblast markers through activation of STAT3 and p38 MAPK signaling pathways. PMID:28084409

Differential growth factor control of bone formation through osteoprogenitor differentiation.

PubMed

Chaudhary, L R; Hofmeister, A M; Hruska, K A

2004-03-01

The osteogenic factors bone morphogenetic protein (BMP-7), platelet-derived growth factor (PDGF)-BB, and fibroblast growth factor (FGF-2) regulate the recruitment of osteoprogenitor cells and their proliferation and differentiation into mature osteoblasts. However, their mechanisms of action on osteoprogenitor cell growth, differentiation, and bone mineralization remain unclear. Here, we tested the hypothesis that these osteogenic agents were capable of regulating osteoblast differentiation and bone formation in vitro. Normal human bone marrow stromal (HBMS) cells were treated with BMP-7 (40 ng ml(-1)), PDGF-BB (20 ng ml(-1)), FGF-2 (20 ng ml(-1)), or FGF-2 plus BMP-7 for 28 days in a serum-containing medium with 10 mM beta-glycerophosphate and 50 microg ml(-1) ascorbic acid. BMP-7 stimulated a morphological change to cuboidal-shaped cells, increased alkaline phosphatase (ALKP) activity, bone sialoprotein (BSP) gene expression, and alizarin red S positive nodule formation. Hydroxyapatite (HA) crystal deposition in the nodules was demonstrated by Fourier transform infrared (FTIR) spectroscopy only in BMP-7- and dexamethasone (DEX)-treated cells. DEX-treated cells appeared elongated and fibroblast-like compared to BMP-7-treated cells. FGF-2 did not stimulate ALKP, and cell morphology was dystrophic. PDGF-BB had little or no effect on ALKP activity and biomineralization. Alizarin Red S staining of cells and calcium assay indicated that BMP-7, DEX, and FGF-2 enhanced calcium mineral deposition, but FTIR spectroscopic analysis demonstrated no formation of HA similar to human bone in control, PDGF-BB-, and FGF-2-treated samples. Thus, FGF-2 stimulated amorphous octacalcium phosphate mineral deposition that failed to mature into HA. Interestingly, FGF-2 abrogated BMP-7-induced ALKP activity and HA formation. Results demonstrate that BMP-7 was competent as a sole factor in the differentiation of human bone marrow stromal cells to bone-forming osteoblasts confirmed by FTIR examination of mineralized matrix. Other growth factors, PDGF, and FGF-2 were incompetent as sole factors, and FGF-2 inhibited BMP-7-stimulated osteoblast differentiation.

Bone biomaterials and interactions with stem cells

PubMed Central

Gao, Chengde; Peng, Shuping; Feng, Pei; Shuai, Cijun

2017-01-01

Bone biomaterials play a vital role in bone repair by providing the necessary substrate for cell adhesion, proliferation, and differentiation and by modulating cell activity and function. In past decades, extensive efforts have been devoted to developing bone biomaterials with a focus on the following issues: (1) developing ideal biomaterials with a combination of suitable biological and mechanical properties; (2) constructing a cell microenvironment with pores ranging in size from nanoscale to submicro- and microscale; and (3) inducing the oriented differentiation of stem cells for artificial-to-biological transformation. Here we present a comprehensive review of the state of the art of bone biomaterials and their interactions with stem cells. Typical bone biomaterials that have been developed, including bioactive ceramics, biodegradable polymers, and biodegradable metals, are reviewed, with an emphasis on their characteristics and applications. The necessary porous structure of bone biomaterials for the cell microenvironment is discussed, along with the corresponding fabrication methods. Additionally, the promising seed stem cells for bone repair are summarized, and their interaction mechanisms with bone biomaterials are discussed in detail. Special attention has been paid to the signaling pathways involved in the focal adhesion and osteogenic differentiation of stem cells on bone biomaterials. Finally, achievements regarding bone biomaterials are summarized, and future research directions are proposed. PMID:29285402

Pharmacological activation of aldehyde dehydrogenase 2 promotes osteoblast differentiation via bone morphogenetic protein-2 and induces bone anabolic effect

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mittal, Monika; Pal, Subhashis; China, Shyamsundar

Aldehyde dehydrogenases (ALDHs) are a family of enzymes involved in detoxifying aldehydes. Previously, we reported that an ALDH inhibitor, disulfiram caused bone loss in rats and among ALDHs, osteoblast expressed only ALDH2. Loss-of-function mutation in ALDH2 gene is reported to cause bone loss in humans which suggested its importance in skeletal homeostasis. We thus studied whether activating ALDH2 by N-(1, 3-benzodioxol-5-ylmethyl)-2, 6-dichlorobenzamide (alda-1) had osteogenic effect. We found that alda-1 increased and acetaldehyde decreased the differentiation of rat primary osteoblasts and expressions of ALDH2 and bone morphogenetic protein-2 (BMP-2). Silencing ALDH2 in osteoblasts abolished the alda-1 effects. Further, alda-1 attenuatedmore » the acetaldehyde-induced lipid-peroxidation and oxidative stress. BMP-2 is essential for bone regeneration and alda-1 increased its expression in osteoblasts. We then showed that alda-1 (40 mg/kg dose) augmented bone regeneration at the fracture site with concomitant increase in BMP-2 protein compared with control. The osteogenic dose (40 mg/kg) of alda-1 attained a bone marrow concentration that was stimulatory for osteoblast differentiation, suggesting that the tissue concentration of alda-1 matched its pharmacologic effect. In addition, alda-1 promoted modeling-directed bone growth and peak bone mass achievement, and increased bone mass in adult rats which reiterated its osteogenic effect. In osteopenic ovariectomized (OVX) rats, alda-1 reversed trabecular osteopenia with attendant increase in serum osteogenic marker (procollagen type I N-terminal peptide) and decrease in oxidative stress. Alda-1 has no effect on liver and kidney function. We conclude that activating ALDH2 by alda-1 had an osteoanabolic effect involving increased osteoblastic BMP-2 production and decreased OVX-induced oxidative stress. - Highlights: • Alda-1 induced osteoblast differentiation that involved upregulation of ALDH2 and BMP-2 • Alda-1 attenuated acetaldehyde-induced inhibition of osteoblast differentiation • Alda-1 enhanced bone regeneration at the fracture site and peak bone mass achievement • Alda-1 reversed trabecular osteopenia in OVX rats via an osteoanabolic mechanism.« less

Constitutive β-catenin activation in osteoblasts impairs terminal osteoblast differentiation and bone quality

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bao, Quanwei; Chen, Sixu; Qin, Hao

Accumulating evidence suggests that Wnt/β-catenin signaling plays a central role in controlling bone mass. We previously reported that constitutive activation of β-catenin (CA-β-catenin) in osteoblasts potentially has side effects on the bone growth and bone remodeling process, although it could increase bone mass. The present study aimed to observe the effects of osteoblastic CA-β-catenin on bone quality and to investigate possible mechanisms of these effects. It was found that CA-β-catenin mice exhibited lower mineralization levels and disorganized collagen in long bones as confirmed by von Kossa staining and sirius red staining, respectively. Also, bone strength decreased significantly in CA-β-catenin mice.more » Then the effect of CA-β-catenin on biological functions of osteoblasts were investigated and it was found that the expression levels of osteocalcin, a marker for the late differentiation of osteoblasts, decreased in CA-β-catenin mice, while the expression levels of osterix and alkaline phosphatase, two markers for the early differentiation of osteoblasts, increased in CA-β-catenin mice. Furthermore, higher proliferation rate were revealed in osteoblasts that were isolated from CA-β-catenin mice. The Real-time PCR and western blot examination found that the expression level of c-myc and cyclin D1, two G1 progression-related molecules, increased in osteoblasts that were isolated from the CA-β-catenin mice, and the expression levels of CDK14 and cyclin Y, two mitotic-related molecules that can accelerate cells entering into S and G2/M phases, increased in osteoblasts that were isolated from the CA-β-catenin mice. In summary, osteoblastic CA-β-catenin kept osteoblasts in high proliferative state and impaired the terminal osteoblast differentiation, and this led to changed bone structure and decreased bone strength. - Highlights: • Wnt/β-catenin signaling plays a central role in controlling bone mass. • CA-β-catenin has side effects on the bone strength and bone qulity. • CA-β-catenin kept osteoblasts in high proliferative state. • Osteoblastic CA-β-catenin impaired the terminal osteoblast differentiation.« less

Magnesium and zinc borate enhance osteoblastic differentiation of stem cells from human exfoliated deciduous teeth in vitro.

PubMed

Liu, Yao-Jen; Su, Wen-Ta; Chen, Po-Hung

2018-01-01

Various biocompatible and biodegradable scaffolds blended with biochemical signal molecules with adequate osteoinductive and osteoconductive properties have attracted significant interest in hard tissue engineering regeneration. We evaluated the distinct effects of magnesium borate, zinc borate, and boric acid blended into chitosan scaffold for osteogenic differentiation of stem cells from exfoliated deciduous teeth. Stem cells from exfoliated deciduous teeth cells are a potential source of functional osteoblasts for applications in bone tissue engineering, but the efficiency of osteoblastic differentiation is low, thereby significantly limiting their clinical applications. Divalent metal borates have potential function in bone remodeling because they can simulate bone formation and decrease bone resorption. These magnesium, zinc, and B ions can gradually be released into the culture medium from the scaffold and induce advanced osteoblastic differentiation from stem cells from exfoliated deciduous teeth. Stem cells from exfoliated deciduous teeth with magnesium borate or zinc borate as inducer demonstrated more osteoblastic differentiation after 21 days of culture. Differentiated cells exhibited activity of alkaline phosphatase, bone-related gene expression of collagen type I, runt-related transcription factor 2, osteopontin, osteocalcin, vascular endothelial growth factor, and angiopoietin-1, as noted via real-time polymerase chain reaction analysis, as well as significant deposits of calcium minerals. Divalent mental magnesium and zinc and nonmetal boron can be an effective inducer of osteogenesis for stem cells from exfoliated deciduous teeth. This experiment might provide useful inducers for osteoblastic differentiation of stem cells from exfoliated deciduous teeth for tissue engineering and bone repair.

A novel rat fibrosarcoma cell line from transformed bone marrow-derived mesenchymal stem cells with maintained in vitro and in vivo stemness properties.

PubMed

Wang, Meng-Yu; Nestvold, Janne; Rekdal, Øystein; Kvalheim, Gunnar; Fodstad, Øystein

2017-03-15

Increasing evidence suggests a possible relationship between mesenchymal stem cells (MSCs) and sarcoma. MSCs are hypothesized to be the cells initiating sarcomagenesis, and cancer stem cells (CSCs) sharing features of MSCs have been identified in sarcomas. Here, we report on the characteristics of a bone marrow-derived rat mesenchymal stem cell line that spontaneously transformed in long-term culture. The rat transformed mesenchymal stem cells (rTMSCs) produced soft-tissue fibrosarcomas in immunocompromised mice and immunocompetent rats. In vitro, the rTMSCs displayed increased proliferation capacity compared to the untransformed cell line. The transformed MSCs maintained the mesenchymal phenotype by expression of the stem cell marker CD 90 and the lack of hematopoietic and endothelial markers. Cytogenetic analysis detected trisomy 6 in the rTMSCs. Side population (SP) isolation and tumorsphere cultivation of the transformed cells confirmed the presence of CSCs among the rTMSCs. Importantly, the rTMSCs retained their differentiation capacity towards osteogenic and adipogenic lineages. This transformed MSC-based cell line may be valuable in examining the balance in a mixed cell population between cancer stem cell properties and the ability to differentiate to specific non-transformed cell populations. Moreover, it may also be a useful tool to evaluate the efficacy of novel targeted immunotherapies in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

Regulation of Bone Metabolism by Serotonin.

PubMed

Lavoie, Brigitte; Lian, Jane B; Mawe, Gary M

2017-01-01

The processes of bone growth and turnover are tightly regulated by the actions of various signaling molecules, including hormones, growth factors, and cytokines. Imbalances in these processes can lead to skeletal disorders such as osteoporosis or high bone mass disease. It is becoming increasingly clear that serotonin can act through a number of mechanisms, and at different locations in the body, to influence the balance between bone formation and resorption. Its actions on bone metabolism can vary, based on its site of synthesis (central or peripheral) as well as the cells and subtypes of receptors that are activated. Within the central nervous system, serotonergic neurons act via the hypothalamus to suppress sympathetic input to the bone. Since sympathetic input inhibits bone formation, brain serotonin has a net positive effect on bone growth. Gut-derived serotonin is thought to inhibit bone growth by attenuating osteoblast proliferation via activation of receptors on pre-osteoblasts. There is also evidence that serotonin can be synthesized within the bone and act to modulate bone metabolism. Osteoblasts, osteoclasts, and osteocytes all have the machinery to synthesize serotonin, and they also express the serotonin-reuptake transporter (SERT). Understanding the roles of serotonin in the tightly balanced system of bone modeling and remodeling is a clinically relevant goal. This knowledge can clarify bone-related side effects of drugs that affect serotonin signaling, including serotonin-specific reuptake inhibitors (SSRIs) and receptor agonists and antagonists, and it can potentially lead to therapeutic approaches for alleviating bone pathologies.

How B cells influence bone biology in health and disease.

PubMed

Horowitz, Mark C; Fretz, Jackie A; Lorenzo, Joseph A

2010-09-01

It is now well established that important regulatory interactions occur between the cells in the hematopoietic, immune and skeletal systems (osteoimmunology). B lymphocytes (B cells) are responsible for the generation and production of antibodies or immunoglobulins in the body. Together with T cells these lymphocytes comprise the adaptive immune system, which allows an individual to develop specific responses to an infection and retain memory of that infection, allowing for a faster and more robust response if that same infection occurs again. In addition to this immune function, B cells have a close and multifaceted relationship with bone cells. B cells differentiate from hematopoietic stem cells (HSCs) in supportive niches found on endosteal bone surfaces. Cells in the osteoblast lineage support HSC and B cell differentiation in these niches. B cell differentiation is regulated, at least in part, by a series of transcription factors that function in a temporal manner. While these transcription factors are required for B cell differentiation, their loss causes profound changes in the bone phenotype. This is due, in part, to the close relationship between macrophage/osteoclast and B cell differentiation. Cross talk between B cells and bone cells is reciprocal with defects in the RANKL-RANK, OPG signaling axis resulting in altered bone phenotypes. While the role of B cells during normal bone remodeling appears minimal, activated B cells play an important role in many inflammatory diseases with associated bony changes. This review examines the relationship between B cells and bone cells and how that relationship affects the skeleton and hematopoiesis during health and disease. Copyright 2010 Elsevier Inc. All rights reserved.

The Use of Patient-Specific Induced Pluripotent Stem Cells (iPSCs) to Identify Osteoclast Defects in Rare Genetic Bone Disorders

PubMed Central

Chen, I-Ping

2014-01-01

More than 500 rare genetic bone disorders have been described, but for many of them only limited treatment options are available. Challenges for studying these bone diseases come from a lack of suitable animal models and unavailability of skeletal tissues for studies. Effectors for skeletal abnormalities of bone disorders may be abnormal bone formation directed by osteoblasts or anomalous bone resorption by osteoclasts, or both. Patient-specific induced pluripotent stem cells (iPSCs) can be generated from somatic cells of various tissue sources and in theory can be differentiated into any desired cell type. However, successful differentiation of hiPSCs into functional bone cells is still a challenge. Our group focuses on the use of human iPSCs (hiPSCs) to identify osteoclast defects in craniometaphyseal dysplasia. In this review, we describe the impact of stem cell technology on research for better treatment of such disorders, the generation of hiPSCs from patients with rare genetic bone disorders and current protocols for differentiating hiPSCs into osteoclasts. PMID:25621177

Clec11a/osteolectin is an osteogenic growth factor that promotes the maintenance of the adult skeleton

PubMed Central

Yue, Rui; Shen, Bo; Morrison, Sean J

2016-01-01

Bone marrow stromal cells maintain the adult skeleton by forming osteoblasts throughout life that regenerate bone and repair fractures. We discovered that subsets of these stromal cells, osteoblasts, osteocytes, and hypertrophic chondrocytes secrete a C-type lectin domain protein, Clec11a, which promotes osteogenesis. Clec11a-deficient mice appeared developmentally normal and had normal hematopoiesis but reduced limb and vertebral bone. Clec11a-deficient mice exhibited accelerated bone loss during aging, reduced bone strength, and delayed fracture healing. Bone marrow stromal cells from Clec11a-deficient mice showed impaired osteogenic differentiation, but normal adipogenic and chondrogenic differentiation. Recombinant Clec11a promoted osteogenesis by stromal cells in culture and increased bone mass in osteoporotic mice in vivo. Recombinant human Clec11a promoted osteogenesis by human bone marrow stromal cells in culture and in vivo. Clec11a thus maintains the adult skeleton by promoting the differentiation of mesenchymal progenitors into mature osteoblasts. In light of this, we propose to call this factor Osteolectin. DOI: http://dx.doi.org/10.7554/eLife.18782.001 PMID:27976999

Physiological Stresses Related to Hypercapnia during Patrols on Submarines

DTIC Science & Technology

1975-12-01

Acid- base balance, CO., storage, and calcium homeostasis | I am trying to show that this delayed renal response in low level chronic hypercapnia is 1...C02 Co, P BONE 4 1 BLOOD Fig. 11. Cycles in acid- base balance, bone buffering, and renal regulation during prolonged exposure to 0.7...patrols on submarines K. E. SCHAEFER Naval Submarine Medical Research Laboratory, Naval Submarine Base . Groton. CT 06340 Schaefer, K. E. 1979

Transgenic expression of BRCA1 disturbs hematopoietic stem and progenitor cells quiescence and function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bai, Lin; Shi, Guiying; Zhang, Xu

The balance between quiescence and proliferation of HSCs is an important regulator of hematopoiesis. Loss of quiescence frequently results in HSCs exhaustion, which underscores the importance of tight regulation of proliferation in these cells. Studies have indicated that cyclin-dependent kinases are involved in the regulation of quiescence in HSCs. BRCA1 plays an important role in the repair of DNA double-stranded breaks, cell cycle, apoptosis and transcription. BRCA1 is expressed in the bone marrow. However, the function of BRCA1 in HSCs is unknown. In our study, we generated BRCA1 transgenic mice to investigate the effects of BRCA1 on the mechanisms ofmore » quiescence and differentiation in HSCs. The results demonstrate that over-expression of BRCA1 in the bone marrow impairs the development of B lymphocytes. Furthermore, BRCA1 induced an increase in the number of LSKs, LT-HSCs, ST-HSCs and MPPs. A competitive transplantation assay found that BRCA1 transgenic mice failed to reconstitute hematopoiesis. Moreover, BRCA1 regulates the expression of p21{sup waf1}/cip1 and p57{sup kip2}, which results in a loss of quiescence in LSKs. Together, over-expression of BRCA1 in bone marrow disrupted the quiescent of LSKs, induced excessive accumulation of LSKs, and disrupted differentiation of the HSCs, which acts through the down-regulated of p21{sup waf1}/cip1 and p57{sup kip2}. - Highlights: • Over-expression of BRCA1 results in impaired B lymphocyte development. • BRCA1 transgenic mice disrupted the quiescent of LSKs, induced excessive accumulation of LSKs. • BRCA1 impairs the function of HSCs through the down-regulated of p21{sup waf1/cip1} and p57{sup kip2}.« less

The effects of vibration loading on adipose stem cell number, viability and differentiation towards bone-forming cells

PubMed Central

Tirkkonen, Laura; Halonen, Heidi; Hyttinen, Jari; Kuokkanen, Hannu; Sievänen, Harri; Koivisto, Anna-Maija; Mannerström, Bettina; Sándor, George K. B.; Suuronen, Riitta; Miettinen, Susanna; Haimi, Suvi

2011-01-01

Mechanical stimulation is an essential factor affecting the metabolism of bone cells and their precursors. We hypothesized that vibration loading would stimulate differentiation of human adipose stem cells (hASCs) towards bone-forming cells and simultaneously inhibit differentiation towards fat tissue. We developed a vibration-loading device that produces 3g peak acceleration at frequencies of 50 and 100 Hz to cells cultured on well plates. hASCs were cultured using either basal medium (BM), osteogenic medium (OM) or adipogenic medium (AM), and subjected to vibration loading for 3 h d–1 for 1, 7 and 14 day. Osteogenesis, i.e. differentiation of hASCs towards bone-forming cells, was analysed using markers such as alkaline phosphatase (ALP) activity, collagen production and mineralization. Both 50 and 100 Hz vibration frequencies induced significantly increased ALP activity and collagen production of hASCs compared with the static control at 14 day in OM. A similar trend was detected for mineralization, but the increase was not statistically significant. Furthermore, vibration loading inhibited adipocyte differentiation of hASCs. Vibration did not affect cell number or viability. These findings suggest that osteogenic culture conditions amplify the stimulatory effect of vibration loading on differentiation of hASCs towards bone-forming cells. PMID:21613288

Cytokines TNF-α, IL-6, IL-17F, and IL-4 Differentially Affect Osteogenic Differentiation of Human Adipose Stem Cells

PubMed Central

Bravenboer, Nathalie

2016-01-01

During the initial stages of bone repair, proinflammatory cytokines are released within the injury site, quickly followed by a shift to anti-inflammatory cytokines. The effect of pro- and anti-inflammatory cytokines on osteogenic differentiation of mesenchymal stem cells is controversial. Here, we investigated the effect of the proinflammatory cytokines TNF-α, IL-6, IL-8, and IL-17F and the anti-inflammatory cytokine IL-4 on proliferation and osteogenic differentiation of human adipose stem cells (hASCs). hASCs were treated with TNF-α, IL-6, IL-8, IL-17F, or IL-4 (10 ng/mL) for 72 h mimicking bone repair. TNF-α reduced collagen type I gene expression but increased hASC proliferation and ALP activity. IL-6 also strongly enhanced ALP activity (18-fold), as well as bone nodule formation by hASCs. IL-8 did not affect proliferation or osteogenic gene expression but reduced bone nodule formation. IL-17F decreased hASC proliferation but enhanced ALP activity. IL-4 enhanced osteocalcin gene expression and ALP activity but reduced RUNX2 gene expression and bone nodule formation. In conclusion, all cytokines studied have both enhancing and reducing effects on osteogenic differentiation of hASCs, even when applied for 72 h only. Some cytokines, specifically IL-6, may be suitable to induce osteogenic differentiation of mesenchymal stem cells as a strategy for enhancing bone repair. PMID:27667999

Murine bone cell lines as models for spaceflight induced effects on differentiation and gene expression

NASA Astrophysics Data System (ADS)

Lau, P.; Hellweg, C. E.; Baumstark-Khan, C.; Reitz, G.

Critical health factors for space crews especially on long-term missions are radiation exposure and the absence of gravity DNA double strand breaks DSB are presumed to be the most deleterious DNA lesions after radiation as they disrupt both DNA strands in close proximity Besides radiation risk the absence of gravity influences the complex skeletal apparatus concerning muscle and especially bone remodelling which results from mechanical forces exerting on the body Bone is a dynamic tissue which is life-long remodelled by cells from the osteoblast and osteoclast lineage Any imbalance of this system leads to pathological conditions such as osteoporosis or osteopetrosis Osteoblastic cells play a crucial role in bone matrix synthesis and differentiate either into bone-lining cells or into osteocytes Premature terminal differentiation has been reported to be induced by a number of DNA damaging or cell stress inducing agents including ionising and ultraviolet radiation as well as treatment with mitomycin C In the present study we compare the effects of sequential differentiation by adding osteoinductive substances ss -glycerophosphate and ascorbic acid Radiation-induced premature differentiation was investigated regarding the biosynthesis of specific osteogenic marker molecules and the differentiation dependent expression of marker genes The bone cell model established in our laboratory consists of the osteocyte cell line MLO-Y4 the osteoblast cell line OCT-1 and the subclones 4 and 24 of the osteoblast cell line MC3T3-E1 expressing several

The use of SHP-2 gene transduced bone marrow mesenchymal stem cells to promote osteogenic differentiation and bone defect repair in rat.

PubMed

Fan, Dapeng; Liu, Shen; Jiang, Shichao; Li, Zhiwei; Mo, Xiumei; Ruan, Hongjiang; Zou, Gang-Ming; Fan, Cunyi

2016-08-01

Bone tissue engineering is a promising approach for bone regeneration, in which growth factors play an important role. The tyrosine phosphatase Src-homology region 2-containing protein tyrosine phosphatase 2 (SHP2), encoded by the PTPN11 gene, is essential for the differentiation, proliferation and metabolism of osteoblasts. However, SHP-2 has never been systematically studied for its effect in osteogenesis. We predicted that overexpression of SHP-2 could promote bone marrow-derived mesenchymal stem cell (BMSC)osteogenic differentiation and SHP-2 transduced BMSCs could enhance new bone formation, determined using the following study groups: (1) BMSCs transduced with SHP-2 and induced with osteoblast-inducing liquid (BMSCs/SHP-2/OL); (2) BMSCs transduced with SHP-2 (BMSCs/-SHP-2); (3) BMSCs induced with osteoblast-inducing liquid (BMSCs/OL) and (4) pure BMSCs. Cells were assessed for osteogenic differentiation by quantitative real-time polymerase chain reaction analysis, western blot analysis, alkaline phosphatase activity and alizarin red S staining. For in vivo assessment, cells were combined with beta-tricalcium phosphate scaffolds and transplanted into rat calvarial defects for 8 weeks. Following euthanasia, skull samples were explanted for osteogenic evaluation, including micro-computed tomography measurement, histology and immunohistochemistry staining. SHP-2 and upregulation of its gene promoted BMSC osteogenic differentiation and therefore represents a potential new therapeutic approach to bone repair. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1871-1881, 2016. © 2016 Wiley Periodicals, Inc.

Low skeletal muscle mass is associated with poor structural parameters of bone and impaired balance in elderly men--the MINOS study.

PubMed

Szulc, Pawel; Beck, Tom J; Marchand, François; Delmas, Pierre D

2005-05-01

In 796 men, 50-85 years of age, decreased relative skeletal muscle mass index was associated with narrower bones, thinner cortices, and a consequent decreased bending strength (lower section modulus), as well as with impaired balance and an increased risk of falls. In men, appendicular skeletal muscle mass (ASM) is correlated positively with BMC and areal BMD (aBMD). In elderly men, low muscle mass and strength (sarcopenia) is associated with difficulties in daily living activities. The aim of this study was to evaluate if ASM is correlated with bone size, mechanical properties of bones, balance, and risk of falls in elderly men. This study used 796 men, 50-85 years of age, belonging to the MINOS cohort. Lifestyle factors were evaluated by standardized questionnaires. Estimates of mechanical bone properties were derived from aBMD measured by DXA. ASM was estimated by DXA. The relative skeletal muscle mass index (RASM) was calculated as ASM/(body height)(2.3). After adjustment for age, body size, tobacco smoking, professional physical activity, and 17beta-estradiol concentration, RASM was correlated positively with BMC, aBMD, external diameter, and cortical thickness (r = 0.17-0.34, p < 0.0001) but not with volumetric BMD. Consequently, RASM was correlated with section modulus (r = 0.29-0.39, p < 0.0001). Men in the lowest quartile of RASM had section modulus of femoral neck and distal radius lower by 12-18% in comparison with men in the highest quartile of RASM. In contrast, bone width was not correlated with fat mass, reflecting the load of body weight (except for L(3)), which suggests that the muscular strain may exert a direct stimulatory effect on periosteal apposition. After adjustment for confounding variables, a decrease in RASM was associated with increased risk of falls and of inability to accomplish clinical tests of muscle strength, static balance, and dynamic balance (odds ratio per 1 SD decrease in RASM, 1.31-2.23; p < 0.05-0.001). In elderly men, decreased RASM is associated with narrower bones and thinner cortices, which results in a lower bending strength. Low RASM is associated with impaired balance and with an increased risk of falls in elderly men. It remains to be studied whether low RASM is associated with decreased periosteal apposition and with increased fracture risk in elderly men, and whether the difference in skeletal muscle mass between men and women contributes to the between-sex difference in fracture incidence.

An age-dependent interaction with leptin unmasks ghrelin's bone-protective effects

USDA-ARS?s Scientific Manuscript database

The mutual interplay between energy homeostasis and bone metabolism is an important emerging concept. Ghrelin and leptin antagonize each other in regulating energy balance, but the role of this interaction in bone metabolism is unknown. Using ghrelin receptor and leptin-deficient mice, we show that ...

Effects of Ving Tsun Chinese martial art training on musculoskeletal health, balance performance, and self-efficacy in community-dwelling older adults.

PubMed

Lip, Ryan W T; Fong, Shirley S M; Ng, Shamay S M; Liu, Karen P Y; Guo, X

2015-03-01

[Purpose] The aim of this study was to investigate the effects of Ving Tsun (VT) Chinese martial art training on radial bone strength, upper- and lower-limb muscular strength, shoulder joint mobility, balance performance, and self-efficacy in elderly participants. [Subjects and Methods] Twelve seniors voluntarily joined the VT training group, and twenty-seven seniors voluntarily joined the control group. The VT group received VT training for three months, while the control group received no training. The bone strength of the distal radius was assessed using an ultrasound bone sonometer. Muscular strength in the limbs was evaluated using a Jamar handgrip dynamometer and the five times sit-to-stand test. Shoulder joint mobility was examined using a goniometer. Balance performance and self-efficacy were evaluated using the Berg Balance Scale and the Chinese version of the Activities-specific Balance Confidence Scale, respectively. [Results] The results revealed a nonsignificant group-by-time interaction effect, group effect, and time effect for all outcome variables. However, general trends of maintenance or improvement in all outcome parameters were observed to a greater extent in the VT group than in the control group. [Conclusion] VT training might be a potential fall-prevention exercise that can be used to maintain general physique, balance, and confidence in the elderly population. A further randomized controlled trial is needed to confirm this postulation.

Effects of Ving Tsun Chinese martial art training on musculoskeletal health, balance performance, and self-efficacy in community-dwelling older adults

PubMed Central

Lip, Ryan W.T.; Fong, Shirley S.M.; Ng, Shamay S.M.; Liu, Karen P.Y.; Guo, X.

2015-01-01

[Purpose] The aim of this study was to investigate the effects of Ving Tsun (VT) Chinese martial art training on radial bone strength, upper- and lower-limb muscular strength, shoulder joint mobility, balance performance, and self-efficacy in elderly participants. [Subjects and Methods] Twelve seniors voluntarily joined the VT training group, and twenty-seven seniors voluntarily joined the control group. The VT group received VT training for three months, while the control group received no training. The bone strength of the distal radius was assessed using an ultrasound bone sonometer. Muscular strength in the limbs was evaluated using a Jamar handgrip dynamometer and the five times sit-to-stand test. Shoulder joint mobility was examined using a goniometer. Balance performance and self-efficacy were evaluated using the Berg Balance Scale and the Chinese version of the Activities-specific Balance Confidence Scale, respectively. [Results] The results revealed a nonsignificant group-by-time interaction effect, group effect, and time effect for all outcome variables. However, general trends of maintenance or improvement in all outcome parameters were observed to a greater extent in the VT group than in the control group. [Conclusion] VT training might be a potential fall-prevention exercise that can be used to maintain general physique, balance, and confidence in the elderly population. A further randomized controlled trial is needed to confirm this postulation. PMID:25931704

Concise Review: Stem Cells in Osteoimmunology.

PubMed

Fierro, Fernando A; Nolta, Jan A; Adamopoulos, Iannis E

2017-06-01

Bone remodeling is a lifelong process in which mature bone tissue is removed from the skeleton by bone resorption and is replenished by new during ossification or bone formation. The remodeling cycle requires both the differentiation and activation of two cell types with opposing functions; the osteoclast, which orchestrates bone resorption, and the osteoblast, which orchestrates bone formation. The differentiation of these cells from their respective precursors is a process which has been overshadowed by enigma, particularly because the precise osteoclast precursor has not been identified and because the identification of skeletal stem cells, which give rise to osteoblasts, is very recent. Latest advances in the area of stem cell biology have enabled us to gain a better understanding of how these differentiation processes occur in physiological and pathological conditions. In this review we postulate that modulation of stem cells during inflammatory conditions is a necessary prerequisite of bone remodeling and therefore an essential new component to the field of osteoimmunology. In this context, we highlight the role of transcription factor nuclear factor of activated T cells cytoplasmic 1 (NFATc1), because it directly links inflammation with differentiation of osteoclasts and osteoblasts. Stem Cells 2017;35:1461-1467. © 2017 The Authors Stem Cells published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Global MicroRNA Profiling in Human Bone Marrow Skeletal-Stromal or Mesenchymal-Stem Cells Identified Candidates for Bone Regeneration.

PubMed

Chang, Chi-Chih; Venø, Morten T; Chen, Li; Ditzel, Nicholas; Le, Dang Q S; Dillschneider, Philipp; Kassem, Moustapha; Kjems, Jørgen

2018-02-07

Bone remodeling and regeneration are highly regulated multistep processes involving posttranscriptional regulation by microRNAs (miRNAs). Here, we performed a global profiling of differentially expressed miRNAs in bone-marrow-derived skeletal cells (BMSCs; also known as stromal or mesenchymal stem cells) during in vitro osteoblast differentiation. We functionally validated the regulatory effects of several miRNAs on osteoblast differentiation and identified 15 miRNAs, most significantly miR-222 and miR-423, as regulators of osteoblastogenesis. In addition, we tested the possible targeting of miRNAs for enhancing bone tissue regeneration. Scaffolds functionalized with miRNA nano-carriers enhanced osteoblastogenesis in 3D culture and retained this ability at least 2 weeks after storage. Additionally, anti-miR-222 enhanced in vivo ectopic bone formation through targeting the cell-cycle inhibitor CDKN1B (cyclin-dependent kinase inhibitor 1B). A number of additional miRNAs exerted additive osteoinductive effects on BMSC differentiation, suggesting that pools of miRNAs delivered locally from an implanted scaffold can provide a promising approach for enhanced bone regeneration. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Mouse fibroblasts homozygous for c-Src oncogene disruption shows dramatic suppression of expression of the gene encoding osteopontin, and adhesive phosphoprotein implicated in bone differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chackalaparampil, I.; Mukherjee, B.B.; Peri, A.

1994-09-01

Osteopetrosis, affecting mice and humans alike, arises from reduced or impaired bone resorption, causing abnormally dense bone formation. Normal bone differentiation requires continuous resorption and remodeling by osteoclasts which are derived from monocyte/macrophage lineage in the bone marrow. It has been reported that targeted homozygous disruption of c-src proto-oncogene in mice results in the development of osteopetrosis due to impaired bone-resorbing function of osteoclast cells. However, the molecular mechanism(s) which leads to osteoclast dysfunction in c-src deficient (src{sup -/-}) mice remains unclear. Here, we report that in embryonic fibroblasts derived from homozygous Src{sup -/-} mice, the expression of the genemore » coding for osteopontin (OP), a phosphorylated glycoprotein involved in bone differentiation, is drastically repressed. OP gene expression is not, however, affected in the heterozygous (Src{sup +/-}) mutant cells of identical origin, or in the c-src expression and OP production. Moreover, OP expression in c-src-deficient cells could be rescued upon treatment with 12-0-tetradecanoyl phorbol-13-myristate-acetate or okadaic acid. These observations indicate that OP expression is regulated via an src-mediated protein kinase C signaling pathway. Since it is known that OP mediates osteoclast adherence to the bone matrix, a key event in bone differentiation, our data is most significant in that they strongly suggest that drastic inhibition of synthesis of OP prevents osteoclasts in Src{sup -/-} mice from anchoring to the bone matrix. Consequently, this disruption of osteoclast adherence impairs their ability to form bone-resorbing ruffled border, causing osteopetrosis.« less

Histone deacetylase inhibitors reduce differentiating osteoblast-mediated protection of acute myeloid leukemia cells from cytarabine

PubMed Central

Sterner, Rosalie M.; Kremer, Kimberly N.; Al-Kali, Aref; Patnaik, Mrinal M.; Gangat, Naseema; Litzow, Mark R.; Kaufmann, Scott H.; Westendorf, Jennifer J.; van Wijnen, Andre J.; Hedin, Karen E.

2017-01-01

The bone marrow microenvironment protects acute myeloid leukemia (AML) cells during chemotherapy and is a major factor in relapse. Here, we examined which type(s) of bone marrow cells are responsible for the relapse of AML following treatment with cytarabine (Ara-C), and we identified a means to inhibit this protection. To determine the protective cell type(s), AML cells were treated with Ara-C, and AML cell survival in the presence or absence of osteoblast lineage cells was assessed. Cultured AML cells and patient bone marrow isolates were each significantly protected from Ara-C-induced apoptosis by co-culture with differentiating osteoblasts. Moreover, pretreating differentiating osteoblasts with the histone deacetylase inhibitors (HDACi) vorinostat and panobinostat abrogated the ability of the differentiating osteoblasts to protect AML cells. Together, our results indicate that differentiating osteoblasts have the potential to promote residual AML in the bone marrow following standard chemotherapy and act via a mechanism requiring HDACi-sensitive gene expression. Using HDACi to target the leukemic microenvironment in combination with Ara-C could potentially improve treatment of AML. Moreover, other strategies for manipulating bone marrow osteoblasts may also help eradicate AML cells and reduce relapse. PMID:29212250

A novel bioactive osteogenesis scaffold delivers ascorbic acid, β-glycerophosphate, and dexamethasone in vivo to promote bone regeneration.

PubMed

Wang, Chao; Cao, Xuecheng; Zhang, Yongxian

2017-05-09

Ascorbic acid, β-glycerophosphate, and dexamethasone have been used in osteogenesis differentiation medium for in vitro cell culture, nothing is known for delivering these three bioactive compounds in vivo. In this study, we synthesized a novel bioactive scaffold by combining these three compounds with a lysine diisocyanate-based polyurethane. These bioactive compounds were released from the scaffold during the degradation process. The cell culture showed that the sponge-like structure in the scaffold was critical in providing a large surface area to support cell growth and all degradation products of the polymer were non-toxic. This bioactive scaffold enhanced the bone regeneration as evidenced by increasing the expression of three bone-related genes including collagen type I, Runx-2 and osteocalcin in rabbit bone marrow stem cells (BMSCs) in vitro and in vivo. The osteogenesis differentiation of BMSCs cultured in this bioactive scaffold was similar to that in osteogenesis differentiation medium and more extensive in this bioactive scaffold compared to the scaffold without these three bioactive compounds. These results indicated that the scaffold containing three bioactive compounds was a good osteogenesis differentiation promoter to enhance the osteogenesis differentiation and new bone formation in vivo.

A novel bioactive osteogenesis scaffold delivers ascorbic acid, β-glycerophosphate, and dexamethasone in vivo to promote bone regeneration

PubMed Central

Wang, Chao; Cao, Xuecheng; Zhang, Yongxian

2017-01-01

Ascorbic acid, β-glycerophosphate, and dexamethasone have been used in osteogenesis differentiation medium for in vitro cell culture, nothing is known for delivering these three bioactive compounds in vivo. In this study, we synthesized a novel bioactive scaffold by combining these three compounds with a lysine diisocyanate-based polyurethane. These bioactive compounds were released from the scaffold during the degradation process. The cell culture showed that the sponge-like structure in the scaffold was critical in providing a large surface area to support cell growth and all degradation products of the polymer were non-toxic. This bioactive scaffold enhanced the bone regeneration as evidenced by increasing the expression of three bone-related genes including collagen type I, Runx-2 and osteocalcin in rabbit bone marrow stem cells (BMSCs) in vitro and in vivo. The osteogenesis differentiation of BMSCs cultured in this bioactive scaffold was similar to that in osteogenesis differentiation medium and more extensive in this bioactive scaffold compared to the scaffold without these three bioactive compounds. These results indicated that the scaffold containing three bioactive compounds was a good osteogenesis differentiation promoter to enhance the osteogenesis differentiation and new bone formation in vivo. PMID:28404942

Transforming growth factor-{beta} inhibits CCAAT/enhancer-binding protein expression and PPAR{gamma} activity in unloaded bone marrow stromal cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ahdjoudj, S.; Kaabeche, K.; Holy, X.

2005-02-01

The molecular mechanisms regulating the adipogenic differentiation of bone marrow stromal cells in vivo remain largely unknown. In this study, we investigated the regulatory effects of transforming growth factor beta-2 (TGF-{beta}2) on transcription factors involved in adipogenic differentiation induced by hind limb suspension in rat bone marrow stromal cells in vivo. Time course real-time quantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis of gene expression showed that skeletal unloading progressively increases the expression of CCAAT/enhancer-binding protein (C/EBP){alpha} and C/EBP{beta} {alpha} at 5 days in bone marrow stromal cells resulting in increased peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}2) transcripts at 7 days. TGF-{beta}2more » administration in unloaded rats corrected the rise in C/EBP{alpha} and C/EBP{beta} transcripts induced by unloading in bone marrow stromal cells. This resulted in inhibition of PPAR{gamma}2 expression that was associated with increased Runx2 expression. Additionally, the inhibition of C/EBP{alpha} and C/EBP{beta} expression by TGF-{beta}2 was associated with increased PPAR{gamma} serine phosphorylation in bone marrow stromal cells, a mechanism that inhibits PPAR{gamma} transactivating activity. The sequential inhibitory effect of TGF-{beta}2 on C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma}2 resulted in reduced LPL expression and abolition of bone marrow stromal cell adipogenic differentiation, which contributed to prevent bone loss induced by skeletal unloading. We conclude that TGF-{beta}2 inhibits the excessive adipogenic differentiation of bone marrow stromal cells induced by skeletal unloading by inhibiting C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma} expression and activity, which provides a sequential mechanism by which TGF-{beta}2 regulates adipogenic differentiation of bone marrow stromal cells in vivo.« less

Monocytes Induce STAT3 Activation in Human Mesenchymal Stem Cells to Promote Osteoblast Formation

PubMed Central

Nicolaidou, Vicky; Wong, Mei Mei; Redpath, Andia N.; Ersek, Adel; Baban, Dilair F.; Williams, Lynn M.; Cope, Andrew P.; Horwood, Nicole J.

2012-01-01

A major therapeutic challenge is how to replace bone once it is lost. Bone loss is a characteristic of chronic inflammatory and degenerative diseases such as rheumatoid arthritis and osteoporosis. Cells and cytokines of the immune system are known to regulate bone turnover by controlling the differentiation and activity of osteoclasts, the bone resorbing cells. However, less is known about the regulation of osteoblasts (OB), the bone forming cells. This study aimed to investigate whether immune cells also regulate OB differentiation. Using in vitro cell cultures of human bone marrow-derived mesenchymal stem cells (MSC), it was shown that monocytes/macrophages potently induced MSC differentiation into OBs. This was evident by increased alkaline phosphatase (ALP) after 7 days and the formation of mineralised bone nodules at 21 days. This monocyte-induced osteogenic effect was mediated by cell contact with MSCs leading to the production of soluble factor(s) by the monocytes. As a consequence of these interactions we observed a rapid activation of STAT3 in the MSCs. Gene profiling of STAT3 constitutively active (STAT3C) infected MSCs using Illumina whole human genome arrays showed that Runx2 and ALP were up-regulated whilst DKK1 was down-regulated in response to STAT3 signalling. STAT3C also led to the up-regulation of the oncostatin M (OSM) and LIF receptors. In the co-cultures, OSM that was produced by monocytes activated STAT3 in MSCs, and neutralising antibodies to OSM reduced ALP by 50%. These data indicate that OSM, in conjunction with other mediators, can drive MSC differentiation into OB. This study establishes a role for monocyte/macrophages as critical regulators of osteogenic differentiation via OSM production and the induction of STAT3 signalling in MSCs. Inducing the local activation of STAT3 in bone cells may be a valuable tool to increase bone formation in osteoporosis and arthritis, and in localised bone remodelling during fracture repair. PMID:22802946

Skeletal unloading inhibits the in vitro proliferation and differentiation of rat osteoprogenitor cells

NASA Technical Reports Server (NTRS)

Kostenuik, P. J.; Halloran, B. P.; Morey-Holton, E. R.; Bikle, D. D.

1997-01-01

Loss of weight bearing in the growing rat decreases bone formation, osteoblast numbers, and bone maturation in unloaded bones. These responses suggest an impairment of osteoblast proliferation and differentiation. To test this assumption, we assessed the effects of skeletal unloading using an in vitro model of osteoprogenitor cell differentiation. Rats were hindlimb elevated for 0 (control), 2, or 5 days, after which their tibial bone marrow stromal cells (BMSCs) were harvested and cultured. Five days of hindlimb elevation led to significant decreases in proliferation, alkaline phosphatase (AP) enzyme activity, and mineralization of BMSC cultures. Differentiation of BMSCs was analyzed by quantitative competitive polymerase chain reaction of cDNA after 10, 15, 20, and 28 days of culture. cDNA pools were analyzed for the expression of c-fos (an index of proliferation), AP (an index of early osteoblast differentiation), and osteocalcin (a marker of late differentiation). BMSCs from 5-day unloaded rats expressed 50% less c-fos, 61% more AP, and 35% less osteocalcin mRNA compared with controls. These data demonstrate that cultured osteoprogenitor cells retain a memory of their in vivo loading history and indicate that skeletal unloading inhibits proliferation and differentiation of osteoprogenitor cells in vitro.

Global deletion of tetraspanin CD82 attenuates bone growth and enhances bone marrow adipogenesis.

PubMed

Bergsma, Alexis; Ganguly, Sourik S; Dick, Daniel; Williams, Bart O; Miranti, Cindy K

2018-05-18

CD82 is a widely expressed member of the tetraspanin family of transmembrane proteins known to control cell signaling, adhesion, and migration. Tetraspanin CD82 is induced over 9-fold during osteoclast differentiation in vitro; however, its role in bone homeostasis is unknown. A globally deleted CD82 mouse model was used to assess the bone phenotype. Based on microCT and 4-point bending tests, CD82-deficient bones are smaller in diameter and weaker, but display no changes in bone density. Histomorphometry shows a decrease in size, erosion perimeter, and number of osteoclasts in situ, with a corresponding increase in trabecular surface area, specifically in male mice. Male-specific alterations are observed in trabecular structure by microCT and in vitro differentiated osteoclasts are morphologically abnormal. Histomorphometry did not reveal a significant reduction in osteoblast number; however, dynamic labeling reveals a significant decrease in bone growth. Consistent with defects in OB function, OB differentiation and mineralization are defective in vitro, whereas adipogenesis is enhanced. There is a corresponding increase in bone marrow adipocytes in situ. Thus, combined defects in both osteoclasts and osteoblasts can account for the observed bone phenotypes, and suggests a role for CD82 in both bone mesenchyme and myeloid cells. Copyright © 2018 Elsevier Inc. All rights reserved.

Leptin regulation of bone resorption by the sympathetic nervous system and CART.

PubMed

Elefteriou, Florent; Ahn, Jong Deok; Takeda, Shu; Starbuck, Michael; Yang, Xiangli; Liu, Xiuyun; Kondo, Hisataka; Richards, William G; Bannon, Tony W; Noda, Masaki; Clement, Karine; Vaisse, Christian; Karsenty, Gerard

2005-03-24

Bone remodelling, the mechanism by which vertebrates regulate bone mass, comprises two phases, namely resorption by osteoclasts and formation by osteoblasts; osteoblasts are multifunctional cells also controlling osteoclast differentiation. Sympathetic signalling via beta2-adrenergic receptors (Adrb2) present on osteoblasts controls bone formation downstream of leptin. Here we show, by analysing Adrb2-deficient mice, that the sympathetic nervous system favours bone resorption by increasing expression in osteoblast progenitor cells of the osteoclast differentiation factor Rankl. This sympathetic function requires phosphorylation (by protein kinase A) of ATF4, a cell-specific CREB-related transcription factor essential for osteoblast differentiation and function. That bone resorption cannot increase in gonadectomized Adrb2-deficient mice highlights the biological importance of this regulation, but also contrasts sharply with the increase in bone resorption characterizing another hypogonadic mouse with low sympathetic tone, the ob/ob mouse. This discrepancy is explained, in part, by the fact that CART ('cocaine amphetamine regulated transcript'), a neuropeptide whose expression is controlled by leptin and nearly abolished in ob/ob mice, inhibits bone resorption by modulating Rankl expression. Our study establishes that leptin-regulated neural pathways control both aspects of bone remodelling, and demonstrates that integrity of sympathetic signalling is necessary for the increase in bone resorption caused by gonadal failure.

Human fetal bone cells in delivery systems for bone engineering.

PubMed

Tenorio, Diene M H; Scaletta, Corinne; Jaccoud, Sandra; Hirt-Burri, Nathalie; Pioletti, Dominique P; Jaques, Bertrand; Applegate, Lee Ann

2011-11-01

The aim of this study was to culture human fetal bone cells (dedicated cell banks of fetal bone derived from 14 week gestation femurs) within both hyaluronic acid gel and collagen foam, to compare the biocompatibility of both matrices as potential delivery systems for bone engineering and particularly for oral application. Fetal bone cell banks were prepared from one organ donation and cells were cultured for up to 4 weeks within hyaluronic acid (Mesolis®) and collagen foams (TissueFleece®). Cell survival and differentiation were assessed by cell proliferation assays and histology of frozen sections stained with Giemsa, von Kossa and ALP at 1, 2 and 4 weeks of culture. Within both materials, fetal bone cells could proliferate in three-dimensional structure at ∼70% capacity compared to monolayer culture. In addition, these cells were positive for ALP and von Kossa staining, indicating cellular differentiation and matrix production. Collagen foam provides a better structure for fetal bone cell delivery if cavity filling is necessary and hydrogels would permit an injectable technique for difficult to treat areas. In all, there was high biocompatibility, cellular differentiation and matrix deposition seen in both matrices by fetal bone cells, allowing for easy cell delivery for bone stimulation in vivo. Copyright © 2011 John Wiley & Sons, Ltd.

MiRNAs in bone diseases.

PubMed

Moore, Benjamin T; Xiao, Peng

2013-01-01

MicroRNAs (miRNAs), which mainly inhibit protein expression by targeting the 3'UTR (untranslated region) of mRNAs, are known to play various roles in the pathogenesis of many different types of diseases. Specifically, in bone diseases, recent emphasis has been placed on the involvement of miRNAs in the differentiation and proliferation of bone and cartilage cells, particularly with regards to how these mechanisms contribute to bone homeostasis. In this review, we summarize miRNAs that are important in the differentiation and proliferation of bone cells, and specific miRNAs associated with bone diseases, such as osteoporosis, osteoarthritis and rheumatoid arthritis. This review also provides the perspective that miRNA studies will identify not only new mechanisms in basic bone research, but also potential novel diagnostic biomarkers and drug targets for bone diseases.

Noggin inactivation affects the number and differentiation potential of muscle progenitor cells in vivo

PubMed Central

Costamagna, Domiziana; Mommaerts, Hendrik; Sampaolesi, Maurilio; Tylzanowski, Przemko

2016-01-01

Inactivation of Noggin, a secreted antagonist of Bone Morphogenetic Proteins (BMPs), in mice leads, among others, to severe malformations of the appendicular skeleton and defective skeletal muscle fibers. To determine the molecular basis of the phenotype, we carried out a histomorphological and molecular analysis of developing muscles Noggin−/− mice. We show that in 18.5 dpc embryos there is a marked reduction in muscle fiber size and a failure of nuclei migration towards the cell membrane. Molecularly, the absence of Noggin results in an increased BMP signaling in muscle tissue as shown by the increase in SMAD1/5/8 phosphorylation, concomitant with the induction of BMP target genes such as Id1, 2, 3 as well as Msx1. Finally, upon removal of Noggin, the number of mesenchymal Pax7+ muscle precursor cells is reduced and they are more prone to differentiate into adipocytes in vitro. Thus, our results highlight the importance of Noggin/BMP balance for myogenic commitment of early fetal progenitor cells. PMID:27573479

Costal chondrosarcoma requiring differential diagnosis from metastatic tumor.

PubMed

Matsuoka, Katsunari; Ueda, Mitsuhiro; Miyamoto, Yoshihiro

2017-02-01

Although chondrosarcoma is a common malignant bone tumor, cases arising in the rib are relatively rare. We experienced a case of chondrosarcoma arising in the right 10th rib during follow-up after lung cancer surgery. Although the finding of an osteolytic mass suggested a metastatic bone tumor, 18F-fluorodeoxyglucose positron-emission tomography demonstrated low fluorodeoxyglucose uptake, and a primary bone tumor was suspected. The bone tumor was resected and diagnosed as chondrosarcoma. Four years after resection, there has been no recurrence or metastasis. Positron-emission tomography was useful for differential diagnosis between a chondrosarcoma and a metastatic bone tumor.

Differences in responses to X-ray exposure between osteoclast and osteoblast cells

PubMed Central

Zhang, Jian; Wang, Ziyang; Wu, Anqing; Nie, Jing; Pei, Hailong; Hu, Wentao; Wang, Bing; Shang, Peng; Li, Bingyan

2017-01-01

Abstract Radiation-induced bone loss is a potential health concern for cancer patients undergoing radiotherapy. Enhanced bone resorption by osteoclasts and decreased bone formation by osteoblasts were thought to be the main reasons. In this study, we showed that both pre-differentiating and differentiating osteoclasts were relatively sensitive to X-rays compared with osteoblasts. X-rays decreased cell viability to a greater degree in RAW264.7 cells and in differentiating cells than than in osteoblastic MC3T3-E1 cells. X-rays at up to 8 Gy had little effects on osteoblast mineralization. In contrast, X-rays at 1 Gy induced enhanced osteoclastogenesis by enhanced cell fusion, but had no effects on bone resorption. A higher dose of X-rays at 8 Gy, however, had an inhibitory effect on bone resorption. In addition, actin ring formation was disrupted by 8 Gy of X-rays and reorganized into clusters. An increased activity of Caspase 3 was found after X-ray exposure. Actin disorganization and increased apoptosis may be the potential effects of X-rays at high doses, by inhibiting osteoclast differentiation. Taken together, our data indicate high radiosensitivity of osteoclasts. X-ray irradiation at relatively low doses can activate osteoclastogenesis, but not osteogenic differentiation. The radiosensitive osteoclasts are the potentially responsive cells for X-ray-induced bone loss. PMID:28541506

Can one generate stable hyaline cartilage from adult mesenchymal stem cells? A developmental approach.

PubMed

Hellingman, Catharine A; Koevoet, Wendy; van Osch, Gerjo J V M

2012-11-01

Chondrogenically differentiating bone marrow-derived mesenchymal stem cells (BMSCs) display signs of chondrocyte hypertrophy, such as production of collagen type X, MMP13 and alkaline phosphatase (ALPL). For cartilage reconstructions this is undesirable, as terminally differentiated cartilage produced by BMSCs mineralizes when implanted in vivo. Terminal differentiation is not restricted to BMSCs but is also encountered in chondrogenic differentiation of adipose-derived mesenchymal stem cells (MSCs) as well as embryonic stem cells, which by definition should be able to generate all types of tissues, including stable cartilage. Therefore, we propose that the currently used culture conditions may drive the cells towards terminal differentiation. In this manuscript we aim to review the literature, supplemented by our own data to answer the question, is it possible to generate stable hyaline cartilage from adult MSCs? We demonstrate that recently published methods for inhibiting terminal differentiation (through PTHrP, MMP13 or blocking phosphorylation of Smad1/5/8) result in cartilage formation with reduction of hypertrophic markers, although this does not reach the low level of stable chondrocytes. A set of hypertrophy markers should be included in future studies to characterize the phenotype more precisely. Finally, we used what is currently known in developmental biology about the differential development of hyaline and terminally differentiated cartilage to provide thought and insights to change current culture models for creating hyaline cartilage. Inhibiting terminal differentiation may not result in stable hyaline cartilage if the right balance of signals has not been created from the start of culture onwards. Copyright © 2011 John Wiley & Sons, Ltd.

Nuclear factor I-C reciprocally regulates adipocyte and osteoblast differentiation via control of canonical Wnt signaling.

PubMed

Zhou, Jie; Wang, Shan; Qi, Qi; Yang, Xiaoyue; Zhu, Endong; Yuan, Hairui; Li, Xuemei; Liu, Ying; Li, Xiaoxia; Wang, Baoli

2017-05-01

Nuclear factor I-C (NFIC) has recently been identified as an important player in osteogenesis and bone homeostasis in vivo However, the molecular mechanisms involved have yet to be defined. In the current study, Nfic expression was altered in primary marrow stromal cells and established progenitor lines after adipogenic and osteogenic treatment. Overexpression of Nfic in stromal cells ST2, mesenchymal cells C3H10T1/2, and primary marrow stromal cells inhibited adipogenic differentiation, whereas it promoted osteogenic differentiation. Conversely, silencing of endogenous Nfic in the cell lines enhanced adipogenic differentiation, whereas it blocked osteogenic differentiation. Mechanism investigations revealed that Nfic overexpression promoted nuclear translocation of β-catenin and increased nuclear protein levels of β-catenin and transcription factor 7-like 2 (TCF7L2). Promoter studies and the chromatin immunoprecipitation (ChIP) assay revealed that NFIC directly binds to the promoter of low-density lipoprotein receptor-related protein 5 (Lrp5) and thereafter transactivates the promoter. Finally, inactivation of canonical Wnt signaling in ST2 attenuated the inhibition of adipogenic differentiation and stimulation of osteogenic differentiation by NFIC. Our study suggests that NFIC balances adipogenic and osteogenic differentiation from progenitor cells through controlling canonical Wnt signaling and highlights the potential of NFIC as a target for new therapies to control metabolic disorders like osteoporosis and obesity.-Zhou, J., Wang, S., Qi, Q., Yang, X., Zhu, E., Yuan, H., Li, X., Liu, Y., Li, X., Wang, B. Nuclear factor I-C reciprocally regulates adipocyte and osteoblast differentiation via control of canonical Wnt signaling. © FASEB.

In vitro effect of mineralized and demineralized bone allografts on proliferation and differentiation of MG-63 osteoblast-like cells.

PubMed

Lafzi, Ardeshir; Vahabi, Surena; Ghods, Shadab; Torshabi, Maryam

2016-03-01

Due to the extensive use of bone allografts in bone reconstruction and periodontal therapy as suitable alternatives to autografts, they are now marketed under different commercial brands. Considering the controversial reports regarding the osteoinductive properties of bone allografts, this study sought to assess the effect of type (mineralized/demineralized), amount and particle size of several allografts on the proliferation and differentiation of MG-63 osteoblast-like cells. MG-63 cells (24-h culture) were exposed to 20 and 40 mg amounts of nine different commercially available freeze-dried bone allografts. After 24 and 72 h of incubation, the effect of water-soluble allograft released materials on cell viability and proliferation was assessed using methyl thiazol tetrazolium (MTT) assay after 24 and 72 h of exposure. Cell differentiation and mineralization was assessed by real-time quantitative reverse transcription PCR and alizarin red staining after 72 h of exposure. The amount and particle size of understudy allografts had significant effects on cell viability after 24 h of exposure (in contrast to 72 h). Higher rate of proliferation was seen in non-differentiated or slow-differentiated groups. The amount and particle size factors had no significant effect on the amount of calcified nodules or the expression of osteogenic marker genes in most groups. Faster and more distinct differentiation and mineralization was noted in mineralized compared to demineralized groups during the 3-day study period. Based on the results, the understudy mineralized (non-demineralized) bone allografts had greater effect on osteogenic differentiation of the MG-63 cells and showed more in vitro osteoinductive activity compared to partially demineralized and fully demineralized types.

MiR-27a is Essential for the Shift from Osteogenic Differentiation to Adipogenic Differentiation of Mesenchymal Stem Cells in Postmenopausal Osteoporosis.

PubMed

You, Li; Pan, Ling; Chen, Lin; Gu, Wensha; Chen, Jinyu

2016-01-01

Osteoporosis is a progressive bone disease characterized by a decrease in bone mass and density, which results in an increased risk of fractures. Mesenchymal stem cells (MSCs) are progenitor cells that can differentiate into osteoblasts, osteocytes and adipocytes in bone and fat formation. A reduction in the differentiation of MSCs into osteoblasts contributes to the impaired bone formation observed in osteoporosis. MicroRNAs (miRNAs) play a regulatory role in osteogenesis and MSC differentiation. MiR-27a has been reported to be down-regulated in the development of osteoporosis and during adipogenic differentiation. In this study, a miRNA microarray analysis was used to investigate expression profiles of miRNA in the serum of osteoporotic patients and healthy controls and this data was validated by quantitative real-time PCR (qRT-PCR). MSCs isolated from human and mice with miR-27a inhibition or overexpression were induced to differentiate into osteoblasts or adipocytes. TargetScan and PicTar were used to predict the target gene of miR-27a. The mRNA or protein levels of several specific proteins in MSCs were detected using qRT-PCR or western blot analysis. Ovariectomized mice were used as in vivo model of human postmenopausal osteoporosis for bone mineral density measurement, micro-CT analysis and histomorphometric analysis. Here, we analyzed the role of miR-27a in bone metabolism. Microarray analysis indicated that miR-27a expression was significantly reduced in osteoporotic patients. Analysis on MSCs derived from patients with osteoporosis indicated that osteoblastogenesis was reduced, whereas adipogenesis was increased. MSCs that had undergone osteoblast induction showed a significant increase in miR-27a expression, whereas cells that had undergone adipocyte induction showed a significant decrease in miR-27a expression, indicating that miR-27a was essential for MSC differentiation. We demonstrated that myocyte enhancer factor 2 c (Mef2c), a transcription factor, was the direct target of miR-27a using a dual luciferase assay. An inverse relationship between miR-27a expression and Mef2c expression in osteoporotic patients was shown. Silencing of miR-27a decreased bone formation, confirming the role of miR-27a in bone formation in vivo. In summary, miR-27a was essential for the shift of MSCs from osteogenic differentiation to adipogenic differentiation in osteoporosis by targeting Mef2c. © 2016 S. Karger AG, Basel.

Determination of osteogenic or adipogenic lineages in muscle-derived stem cells (MDSCs) by a collagen-binding peptide (CBP) derived from bone sialoprotein (BSP)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Choi, Yoon Jung; Lee, Jue Yeon; Lee, Seung Jin

Highlights: Black-Right-Pointing-Pointer CBP sequence is identified from BSP and has collagen binding activity. Black-Right-Pointing-Pointer CBP directly activates the MAPK signaling, especially ERK1/2. Black-Right-Pointing-Pointer CBP increase osteoblastic differentiation by the activation of Runx2. Black-Right-Pointing-Pointer CBP decrease adipogenic differentiation by the inhibition of PPAR{gamma}. -- Abstract: Bone sialoprotein (BSP) is a mineralized, tissue-specific, non-collagenous protein that is normally expressed only in mineralized tissues such as bone, dentin, cementum, and calcified cartilage, and at sites of new mineral formation. The binding of BSP to collagen is thought to be important for initiating bone mineralization and bone cell adhesion to the mineralized matrix. Severalmore » recent studies have isolated stem cells from muscle tissue, but their functional properties are still unclear. In this study, we examined the effects of a synthetic collagen-binding peptide (CBP) on the differentiation efficiency of muscle-derived stem cells (MDSCs). The CBP sequence (NGVFKYRPRYYLYKHAYFYPHLKRFPVQ) corresponds to residues 35-62 of bone sialoprotein (BSP), which are located within the collagen-binding domain in BSP. Interestingly, this synthetic CBP inhibited adipogenic differentiation but increased osteogenic differentiation in MDSCs. The CBP also induced expression of osteoblastic marker proteins, including alkaline phosphatase (ALP), type I collagen, Runt-related transcription factor 2 (Runx2), and osteocalcin; prevented adipogenic differentiation in MDSCs; and down-regulated adipose-specific mRNAs, such as adipocyte protein 2 (aP2) and peroxisome proliferator-activated receptor {gamma}. The CBP increased Extracellular signal-regulated kinases (ERK) 1/2 protein phosphorylation, which is important in lineage determination. These observations suggest that this CBP determines the osteogenic or adipogenic lineage in MDSCs by activating ERK1/2. Taken together, a novel CBP could be a useful candidate for regenerating bone and treating osteoporosis, which result from an imbalance in osteogenesis and adipogenesis differentiation.« less

Bone health nutraceuticals alter microarray mRNA gene expression: A randomized, parallel, open-label clinical study.

PubMed

Lin, Yumei; Kazlova, Valentina; Ramakrishnan, Shyam; Murray, Mary A; Fast, David; Chandra, Amitabh; Gellenbeck, Kevin W

2016-01-15

Dietary intake of fruits and vegetables has been suggested to have a role in promoting bone health. More specifically, the polyphenols they contain have been linked to physiological effects related to bone mineral density and bone metabolism. In this research, we use standard microarray analyses of peripheral whole blood from post-menopausal women treated with two fixed combinations of plant extracts standardized to polyphenol content to identify differentially expressed genes relevant to bone health. In this 28-day open-label study, healthy post-menopausal women were randomized into three groups, each receiving one of three investigational fixed combinations of plant extracts: an anti-resorptive (AR) combination of pomegranate fruit (Punica granatum L.) and grape seed (Vitis vinifera L.) extracts; a bone formation (BF) combination of quercetin (Dimorphandra mollis Benth) and licorice (Glycyrrhiza glabra L.) extracts; and a fixed combination of all four plant extracts (AR plus BF). Standard microarray analysis was performed on peripheral whole blood samples taken before and after each treatment. Annotated genes were analyzed for their association to bone health by comparison to a gene library. The AR combination down-regulated a number of genes involved in reduction of bone resorption including cathepsin G (CTSG) and tachykinin receptor 1 (TACR1). The AR combination also up-regulated genes associated with formation of extracellular matrix including heparan sulfate proteoglycan 2 (HSPG2) and hyaluronoglucosaminidase 1 (HYAL1). In contrast, treatment with the BF combination resulted in up-regulation of bone morphogenetic protein 2 (BMP-2) and COL1A1 (collagen type I α1) genes which are linked to bone and collagen formation while down-regulating genes linked to osteoclastogenesis. Treatment with a combination of all four plant extracts had a distinctly different effect on gene expression than the results of the AR and BF combinations individually. These results could be due to multiple feedback systems balancing activities of osteoblasts and osteoclasts. In summary, this ex-vivo microarray study indicated that the pomegranate, grape seed, quercetin and licorice combinations of plant extracts modulated gene expression for both osteoclastic and osteogenic processes. Copyright © 2015 The Authors. Published by Elsevier GmbH.. All rights reserved.

Expression profile of osteoprotegerin, RANK and RANKL genes in the femoral head of patients with avascular necrosis.

PubMed

Samara, Stavroula; Dailiana, Zoe; Chassanidis, Christos; Koromila, Theodora; Papatheodorou, Loukia; Malizos, Konstantinos N; Kollia, Panagoula

2014-02-01

Femoral head avascular necrosis (AVN) is a recalcitrant disease of the hip that leads to joint destruction. Osteoprotegerin (OPG), Receptor Activator of Nuclear Factor kappa-B (RANK) and RANK ligand (RANKL) regulate the balance between osteoclasts-osteoblasts. The expression of these genes affects the maturation and function of osteoblasts-osteoclasts and bone remodeling. In this study, we investigated the molecular pathways leading to AVN by studying the expression profile of OPG, RANK and RANKL genes. Quantitative Real Time-PCR was performed for evaluation of OPG, RANK and RANKL expression. Analysis was based on parallel evaluation of mRNA and protein levels in normal/necrotic sites of 42 osteonecrotic femoral heads (FHs). OPG and RANKL protein levels were estimated by western blotting. The OPG mRNA levels were higher (insignificantly) in the necrotic than the normal site (p > 0.05). Although the expression of RANK and RANKL was significantly lower than OPG in both sites, RANK and RANKL mRNA levels were higher in the necrotic part than the normal (p < 0.05). Protein levels of OPG and RANKL showed no remarkable divergence. Our results indicate that differential expression mechanisms for OPG, RANK and RANKL that could play an important role in the progress of bone remodeling in the necrotic area, disturbing bone homeostasis. This finding may have an effect on the resulting bone destruction and the subsequent collapse of the hip joint. Copyright © 2013. Published by Elsevier Inc.

Dynamic Cross Talk between S1P and CXCL12 Regulates Hematopoietic Stem Cells Migration, Development and Bone Remodeling

PubMed Central

Golan, Karin; Kollet, Orit; Lapidot, Tsvee

2013-01-01

Hematopoietic stem cells (HSCs) are mostly retained in a quiescent non-motile mode in their bone marrow (BM) niches, shifting to a migratory cycling and differentiating state to replenish the blood with mature leukocytes on demand. The balance between the major chemo-attractants CXCL12, predominantly in the BM, and S1P, mainly in the blood, dynamically regulates HSC recruitment to the circulation versus their retention in the BM. During alarm situations, stress-signals induce a decrease in CXCL12 levels in the BM, while S1P levels are rapidly and transiently increased in the circulation, thus favoring mobilization of stem cells as part of host defense and repair mechanisms. Myeloid cytokines, including G-CSF, up-regulate S1P signaling in the BM via the PI3K pathway. Induced CXCL12 secretion from stromal cells via reactive oxygen species (ROS) generation and increased S1P1 expression and ROS signaling in HSCs, all facilitate mobilization. Bone turnover is also modulated by both CXCL12 and S1P, regulating the dynamic BM stromal microenvironment, osteoclasts and stem cell niches which all functionally express CXCL12 and S1P receptors. Overall, CXCL12 and S1P levels in the BM and circulation are synchronized to mutually control HSC motility, leukocyte production and osteoclast/osteoblast bone turnover during homeostasis and stress situations. PMID:24276423

Keeping Your Balance

MedlinePlus

... Advocacy Be Bone Strong TCS New York City Marathon 2018 Paget’s Disease What is Paget’s Disease? Symptoms, ... Advocacy Be Bone Strong TCS New York City Marathon 2018 Paget’s Disease What is Paget’s Disease? Symptoms, ...

Body/bone-marrow differential-temperature sensor

NASA Technical Reports Server (NTRS)

Anselmo, V. J.; Berdahl, C. M.

1978-01-01

Differential-temperature sensor developed to compare bone-marrow and body temperature in leukemia patients uses single stable amplifier to monitor temperature difference recorded by thermocouples. Errors are reduced by referencing temperatures to each other, not to separate calibration points.

Role of inflammation in the aging bones.

PubMed

Abdelmagid, Samir M; Barbe, Mary F; Safadi, Fayez F

2015-02-15

Chronic inflammation in aging is characterized by increased inflammatory cytokines, bone loss, decreased adaptation, and defective tissue repair in response to injury. Aging leads to inherent changes in mesenchymal stem cell (MSC) differentiation, resulting in impaired osteoblastogenesis. Also, the pro-inflammatory cytokines increase with aging, leading to enhanced myelopoiesis and osteoclastogenesis. Bone marrow macrophages (BMMs) play pivotal roles in osteoblast differentiation, the maintenance of hematopoietic stem cells (HSCs), and subsequent bone repair. However, during aging, little is known about the role of macrophages in the differentiation and function of MSC and HSC. Aged mammals have higher circulating pro-inflammatory cytokines than young adults, supporting the hypothesis of increased inflammation with aging. This review will aid in the understanding of the potential role(s) of pro-inflammatory (M1) and anti-inflammatory (M2) macrophages in differentiation and function of osteoblasts and osteoclasts in relation to aging. Copyright © 2014 Elsevier Inc. All rights reserved.

Tenascin-W inhibits proliferation and differentiation of preosteoblasts during endochondral bone formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kimura, Hiroaki; Akiyama, Haruhiko; Nakamura, Takashi

We identified a cDNA encoding mouse Tenascin-W (TN-W) upregulated by bone morphogenetic protein (Bmp)2 in ATDC5 osteo-chondroprogenitors. In adult mice, TN-W was markedly expressed in bone. In mouse embryos, during endochondral bone formation TN-W was localized in perichondrium/periosteum, but not in trabecular and cortical bones. During bone fracture repair, cells in the newly formed perichondrium/periosteum surrounding the cartilaginous callus expressed TN-W. Furthermore, TN-W was detectable in perichondrium/periosteum of Runx2-null and Osterix-null embryos, indicating that TN-W is expressed in preosteoblasts. In CFU-F and -O cells, TN-W had no effect on initiation of osteogenesis of bone marrow cells, and in MC3T3-E1 osteoblasticmore » cells TN-W inhibited cell proliferation and Col1a1 expression. In addition, TN-W suppressed canonical Wnt signaling which stimulates osteoblastic differentiation. Our results indicate that TN-W is a novel marker of preosteoblasts in early stage of osteogenesis, and that TN-W inhibits cell proliferation and differentiation of preosteoblasts mediated by canonical Wnt signaling.« less

Insulin antagonises pigment epithelium-derived factor (PEDF)-induced modulation of lineage commitment of myocytes and heterotrophic ossification.

PubMed

Carnagarin, Revathy; Elahy, Mina; Dharmarajan, Arun M; Dass, Crispin R

2017-12-16

Extensive bone defects arising as a result of trauma, infection and tumour resection and other bone pathologies necessitates the identification of effective strategies in the form of tissue engineering, gene therapy and osteoinductive agents to enhance the bone repair process. PEDF is a multifunctional glycoprotein which plays an important role in regulating osteoblastic differentiation and bone formation. PEDF treatment of mice and human skeletal myocytes at physiological concentration inhibited myogenic differentiation and activated Erk1/2 MAPK- dependent osteogenic transdifferentiation of myocytes. In mice, insulin, a promoter of bone regeneration, attenuated PEDF-induced expression of osteogenic markers such as osteocalcin, alkaline phosphatase and mineralisation for bone formation in the muscle and surrounding adipose tissue. These results provide new insights into the molecular aspects of the antagonising effect of insulin on PEDF-dependent modulation of the differentiation commitment of musculoskeletal environment into osteogenesis, and suggest that PEDF may be developed as an effective clinical therapy for bone regeneration as its heterotopic ossification can be controlled via co-administration of insulin. Copyright © 2017 Elsevier B.V. All rights reserved.

Inhibition of Osteoclast Differentiation and Bone Resorption by N-Methylpyrrolidone*

PubMed Central

Ghayor, Chafik; Correro, Rita M.; Lange, Katrin; Karfeld-Sulzer, Lindsay S.; Grätz, Klaus W.; Weber, Franz E.

2011-01-01

Regulation of RANKL (receptor activator of nuclear factor κB ligand)-induced osteoclast differentiation is of current interest in the development of antiresorptive agents. Osteoclasts are multinucleated cells that play a crucial role in bone resorption. In this study, we investigated the effects of N-methylpyrrolidone (NMP) on the regulation of RANKL-induced osteoclastogenesis. NMP inhibited RANKL-induced tartrate-resistant acid phosphatase activity and the formation of tartrate-resistant acid phosphatase-positive multinucleated cells. The RANKL-induced expression of NFATc1 (nuclear factor of activated T cells, cytoplasmic 1) and c-Fos, which are key transcription factors for osteoclastogenesis, was also reduced by treatment with NMP. Furthermore, NMP induced disruption of the actin rings and decreased the mRNAs of cathepsin K and MMP-9 (matrix metalloproteinase-9), both involved in bone resorption. Taken together, these results suggest that NMP inhibits osteoclast differentiation and attenuates bone resorption. Therefore, NMP could prove useful for the treatment of osteoporosis or other bone diseases associated with excessive bone resorption. PMID:21613210

Utilization of microgravity bioreactors for differentiation of mammalian skeletal tissue

NASA Technical Reports Server (NTRS)

Klement, B. J.; Spooner, B. S.

1993-01-01

Bioreactor cell and tissue culture vessels can be used to study bone development in a simulated microgravity environment. These vessels will also provide an advantageous, low maintenance culture system on space station Freedom. Although many types of cells and tissues can potentially utilize this system, our particular interest is in developing bone tissue. We have characterized an organ culture system utilizing embryonic mouse pre-metatarsal mesenchyme, documenting morphogenesis and differentiation as cartilage rods are formed, with subsequent terminal chondrocyte differentiation to hypertrophied cells. Further development to form bone tissue is achieved by supplementation of the culture medium. Research using pre-metatarsal tissue, combined with the bioreactor culture hardware, could give insight into the advantages and/or disadvantages of conditions experienced in microgravity. Studies such as these have the potential to enhance understanding of bone development and adult bone physiology, and may help define the processes of bone demineralization experienced in space and in pathological conditions here on earth.

Spontaneous Differentiation of Human Mesenchymal Stem Cells on Poly-Lactic-Co-Glycolic Acid Nano-Fiber Scaffold.

PubMed

Sonomoto, Koshiro; Yamaoka, Kunihiro; Kaneko, Hiroaki; Yamagata, Kaoru; Sakata, Kei; Zhang, Xiangmei; Kondo, Masahiro; Zenke, Yukichi; Sabanai, Ken; Nakayamada, Shingo; Sakai, Akinori; Tanaka, Yoshiya

2016-01-01

Mesenchymal stem cells (MSCs) have immunosuppressive activity and can differentiate into bone and cartilage; and thus seem ideal for treatment of rheumatoid arthritis (RA). Here, we investigated the osteogenesis and chondrogenesis potentials of MSCs seeded onto nano-fiber scaffolds (NFs) in vitro and possible use for the repair of RA-affected joints. MSCs derived from healthy donors and patients with RA or osteoarthritis (OA) were seeded on poly-lactic-glycolic acid (PLGA) electrospun NFs and cultured in vitro. Healthy donor-derived MSCs seeded onto NFs stained positive with von Kossa at Day 14 post-stimulation for osteoblast differentiation. Similarly, MSCs stained positive with Safranin O at Day 14 post-stimulation for chondrocyte differentiation. Surprisingly, even cultured without any stimulation, MSCs expressed RUNX2 and SOX9 (master regulators of bone and cartilage differentiation) at Day 7. Moreover, MSCs stained positive for osteocalcin, a bone marker, and simultaneously also with Safranin O at Day 14. On Day 28, the cell morphology changed from a spindle-like to an osteocyte-like appearance with processes, along with the expression of dentin matrix protein-1 (DMP-1) and matrix extracellular phosphoglycoprotein (MEPE), suggesting possible differentiation of MSCs into osteocytes. Calcification was observed on Day 56. Expression of osteoblast and chondrocyte differentiation markers was also noted in MSCs derived from RA or OA patients seeded on NFs. Lactic acid present in NFs potentially induced MSC differentiation into osteoblasts. Our PLGA scaffold NFs induced MSC differentiation into bone and cartilage. NFs induction process resembled the procedure of endochondral ossification. This finding indicates that the combination of MSCs and NFs is a promising therapeutic technique for the repair of RA or OA joints affected by bone and cartilage destruction.

Angiopoietin-like protein 2 promotes chondrogenic differentiation during bone growth as a cartilage matrix factor.

PubMed

Tanoue, H; Morinaga, J; Yoshizawa, T; Yugami, M; Itoh, H; Nakamura, T; Uehara, Y; Masuda, T; Odagiri, H; Sugizaki, T; Kadomatsu, T; Miyata, K; Endo, M; Terada, K; Ochi, H; Takeda, S; Yamagata, K; Fukuda, T; Mizuta, H; Oike, Y

2018-01-01

Chondrocyte differentiation is crucial for long bone growth. Many cartilage extracellular matrix (ECM) proteins reportedly contribute to chondrocyte differentiation, indicating that mechanisms underlying chondrocyte differentiation are likely more complex than previously appreciated. Angiopoietin-like protein 2 (ANGPTL2) is a secreted factor normally abundantly produced in mesenchymal lineage cells such as adipocytes and fibroblasts, but its loss contributes to the pathogenesis of lifestyle- or aging-related diseases. However, the function of ANGPTL2 in chondrocytes, which are also differentiated from mesenchymal stem cells, remains unclear. Here, we investigate whether ANGPTL2 is expressed in or functions in chondrocytes. First, we evaluated Angptl2 expression during chondrocyte differentiation using chondrogenic ATDC5 cells and wild-type epiphyseal cartilage of newborn mice. We next assessed ANGPTL2 function in chondrogenic differentiation and associated signaling using Angptl2 knockdown ATDC5 cells and Angptl2 knockout mice. ANGPTL2 is expressed in chondrocytes, particularly those located in resting and proliferative zones, and accumulates in ECM surrounding chondrocytes. Interestingly, long bone growth was retarded in Angptl2 knockout mice from neonatal to adult stages via attenuation of chondrocyte differentiation. Both in vivo and in vitro experiments show that changes in ANGPTL2 expression can also alter p38 mitogen-activated protein kinase (MAPK) activity mediated by integrin α5β1. ANGPTL2 contributes to chondrocyte differentiation and subsequent endochondral ossification through α5β1 integrin and p38 MAPK signaling during bone growth. Our findings provide insight into molecular mechanisms governing communication between chondrocytes and surrounding ECM components in bone growth activities. Copyright © 2017. Published by Elsevier Ltd.

CUDC-907 Promotes Bone Marrow Adipocytic Differentiation Through Inhibition of Histone Deacetylase and Regulation of Cell Cycle.

PubMed

Ali, Dalia; Alshammari, Hassan; Vishnubalaji, Radhakrishnan; Chalisserry, Elna Paul; Hamam, Rimi; Alfayez, Musaad; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

2017-03-01

The role of bone marrow adipocytes (BMAs) in overall energy metabolism and their effects on bone mass are currently areas of intensive investigation. BMAs differentiate from bone marrow stromal cells (BMSCs); however, the molecular mechanisms regulating BMA differentiation are not fully understood. In this study, we investigated the effect of CUDC-907, identified by screening an epigenetic small-molecule library, on adipocytic differentiation of human BMSCs (hBMSCs) and determined its molecular mechanism of action. Human bone marrow stromal cells exposed to CUDC-907 (500 nM) exhibited enhanced adipocytic differentiation (∼2.9-fold increase, P < 0.005) compared with that of control cells. Global gene expression and signaling pathway analyses of differentially expressed genes revealed a strong enrichment of genes involved in adipogenesis, cell cycle, and DNA replication. Chromatin immune precipitation combined with quantitative polymerase chain reaction showed significant increase in H3K9ac epigenetic marker in the promoter regions of AdipoQ, FABP4, PPARγ, KLF15, and CEBPA in CUDC-907-treated hBMSCs. Follow-up experiments corroborated that the inhibition of histone deacetylase (HDAC) activity enhanced adipocytic differentiation, while the inhibition of PI3K decreased adipocytic differentiation. In addition, CUDC-907 arrested hBMSCs in the G0-G1 phase of the cell cycle and reduced the number of S-phase cells. Our data reveal that HDAC, PI3K, and cell cycle genes are important regulators of BMA formation and demonstrate that adipocyte differentiation of hBMSCs is associated with complex changes in a number of epigenetic and genetic pathways, which can be targeted to regulate BMA formation.

Myeloid cell recruitment versus local proliferation differentiates susceptibility from resistance to filarial infection

PubMed Central

Campbell, Sharon M; Knipper, Johanna A; Ruckerl, Dominik; Finlay, Conor M; Logan, Nicola; Minutti, Carlos M; Mack, Matthias; Jenkins, Stephen J; Taylor, Matthew D

2018-01-01

Both TH2-dependent helminth killing and suppression of the TH2 effector response have been attributed to macrophages (MΦ) activated by IL-4 (M(IL-4)). To investigate how M(IL-4) contribute to diverse infection outcomes, the MΦ compartment of susceptible BALB/c mice and more resistant C57BL/6 mice was profiled during infection of the pleural cavity with the filarial nematode, Litomosoides sigmodontis. C57BL/6 mice exhibited a profoundly expanded resident MΦ (resMΦ) population, which was gradually replenished from the bone marrow in an age-dependent manner. Infection status did not alter the bone-marrow derived contribution to the resMΦ population, confirming local proliferation as the driver of resMΦ expansion. Significantly less resMΦ expansion was observed in the susceptible BALB/c strain, which instead exhibited an influx of monocytes that assumed an immunosuppressive PD-L2+ phenotype. Inhibition of monocyte recruitment enhanced nematode killing. Thus, the balance of monocytic vs. resident M(IL-4) numbers varies between inbred mouse strains and impacts infection outcome. PMID:29299998

PTH promotes allograft integration in a calvarial bone defect.

PubMed

Sheyn, Dmitriy; Cohn Yakubovich, Doron; Kallai, Ilan; Su, Susan; Da, Xiaoyu; Pelled, Gadi; Tawackoli, Wafa; Cook-Weins, Galen; Schwarz, Edward M; Gazit, Dan; Gazit, Zulma

2013-12-02

Allografts may be useful in craniofacial bone repair, although they often fail to integrate with the host bone. We hypothesized that intermittent administration of parathyroid hormone (PTH) would enhance mesenchymal stem cell recruitment and differentiation, resulting in allograft osseointegration in cranial membranous bones. Calvarial bone defects were created in transgenic mice, in which luciferase is expressed under the control of the osteocalcin promoter. The mice were given implants of allografts with or without daily PTH treatment. Bioluminescence imaging (BLI) was performed to monitor host osteprogenitor differentiation at the implantation site. Bone formation was evaluated with the aid of fluorescence imaging (FLI) and microcomputed tomography (μCT) as well as histological analyses. Reverse transcription polymerase chain reaction (RT-PCR) was performed to evaluate the expression of key osteogenic and angiogenic genes. Osteoprogenitor differentiation, as detected by BLI, in mice treated with an allograft implant and PTH was over 2-fold higher than those in mice treated with an allograft implant without PTH. FLI also demonstrated that the bone mineralization process in PTH-treated allografts was significantly higher than that in untreated allografts. The μCT scans revealed a significant increase in bone formation in allograft + PTH treated mice comparing to allograft + PBS treated mice. The osteogenic genes osteocalcin (Oc/Bglap) and integrin binding sialoprotein (Ibsp) were upregulated in the allograft + PTH treated animals. In summary, PTH treatment enhances osteoprogenitor differentiation and augments bone formation around structural allografts. The precise mechanism is not clear, but we show that infiltration pattern of mast cells, associated with the formation of fibrotic tissue, in the defect site is significantly affected by the PTH treatment.

Growth-differentiation balance: a basis for understanding southern pine beetle - tree interactions

Treesearch

Peter L. Lorio

1986-01-01

Interrelationships between the southern pine beetle (SPB), Dendroctonus frontalis Zimm.) and its host pines are explained in terms of the growth-differentiation balance concept.A general hypothesis is proposed based on growth-differentiation balance in southern pines (radial growth of stems versus synthesis and yield of oleoresin) and seasonal activity of the SPB based...

NHE10, a novel osteoclast-specific member of the Na{sup +}/H{sup +} exchanger family, regulates osteoclast differentiation and survival

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Seoung Hoon; Kim, Taesoo; Park, Eui-Soon

2008-05-02

Bone homeostasis is tightly regulated by the balanced actions of osteoblasts (OBs) and osteoclasts (OCs). We previously analyzed the gene expression profile of OC differentiation using a cDNA microarray, and identified a novel osteoclastogenic gene candidate, clone OCL-1-E7 [J. Rho, C.R. Altmann, N.D. Socci, L. Merkov, N. Kim, H. So, O. Lee, M. Takami, A.H. Brivanlou, Y. Choi, Gene expression profiling of osteoclast differentiation by combined suppression subtractive hybridization (SSH) and cDNA microarray analysis, DNA Cell Biol. 21 (2002) 541-549]. In this study, we have isolated full-length cDNAs corresponding to this clone from mice and humans to determine the functionalmore » roles of this gene in osteoclastogenesis. The full-length cDNA of OCL-1-E7 encodes 12 membrane-spanning domains that are typical of isoforms of the Na{sup +}/H{sup +} exchangers (NHEs), indicating that this clone is a novel member of the NHE family (hereafter referred to as NHE10). Here, we show that NHE10 is highly expressed in OCs in response to receptor activator of nuclear factor-{kappa}B ligand signaling and is required for OC differentiation and survival.« less

Pro416Arg cherubism mutation in Sh3bp2 knock-in mice affects osteoblasts and alters bone mineral and matrix properties

PubMed Central

Wang, Chiachien J.; Chen, I-Ping; Koczon-Jaremko, Boguslawa; Boskey, Adele L.; Ueki, Yasuyoshi; Kuhn, Liisa; Reichenberger, Ernst J.

2010-01-01

Cherubism is an autosomal dominant disorder in children characterized by unwarranted symmetrical bone resorption of the jaws with fibrous tissue deposition. Mutations causing cherubism have been identified in the adaptor protein SH3BP2. Knock-in mice with a Pro416Arg mutation in Sh3bp2 exhibit a generalized osteoporotic bone phenotype. In this study, we examined the effects of this “cherubism” mutation on spectroscopic indices of “bone quality” and on osteoblast differentiation. Fourier-transform infrared imaging (FTIRI) analysis of femurs from wild-type and Sh3bp2 knock-in mice showed decreased mineral content, decreased mineral crystallinity/crystal size, and increased collagen maturity in homozygous mutants. To assess osteoblast maturation in vivo, knock-in mice were crossed with transgenic mice over-expressing GFP driven by 3.6-kb or 2.3-kb Col1a1 promoter fragments. Reduced numbers of mature osteoblasts were observed in homozygous mice. Neonatal calvarial cultures, which were enriched for osteoblasts by depletion of hematopoietic cells (negative selection for Ter119- and CD45-positive cells) were investigated for osteoblast-specific gene expression and differentiation, which demonstrated that differentiation and mineralization in homozygous osteoblast cultures was impaired. Co-cultures with calvarial osteoblasts and bone marrow macrophages showed that mutant osteoblasts appear to increase osteoclastogenesis resulting in increased bone resorption on bone chips. In summary, the Sh3bp2 mutation in cherubism mice alters bone quality, reduces osteoblast function, and may contribute to excessive bone resorption by osteoclasts. Our data, together with previous osteoclast studies, demonstrate a critical role of Sh3bp2 in bone remodeling and osteoblast differentiation. PMID:20117257

Phenotypic Spectrum in Osteogenesis Imperfecta Due to Mutations in TMEM38B: Unraveling a Complex Cellular Defect.

PubMed

Webb, Emma A; Balasubramanian, Meena; Fratzl-Zelman, Nadja; Cabral, Wayne A; Titheradge, Hannah; Alsaedi, Atif; Saraff, Vrinda; Vogt, Julie; Cole, Trevor; Stewart, Susan; Crabtree, Nicola J; Sargent, Brandi M; Gamsjaeger, Sonja; Paschalis, Eleftherios P; Roschger, Paul; Klaushofer, Klaus; Shaw, Nick J; Marini, Joan C; Högler, Wolfgang

2017-06-01

Recessive mutations in TMEM38B cause type XIV osteogenesis imperfecta (OI) by dysregulating intracellular calcium flux. Clinical and bone material phenotype description and osteoblast differentiation studies. Natural history study in pediatric research centers. Eight patients with type XIV OI. Clinical examinations included bone mineral density, radiographs, echocardiography, and muscle biopsy. Bone biopsy samples (n = 3) were analyzed using histomorphometry, quantitative backscattered electron microscopy, and Raman microspectroscopy. Cellular differentiation studies were performed on proband and control osteoblasts and normal murine osteoclasts. Type XIV OI clinical phenotype ranges from asymptomatic to severe. Previously unreported features include vertebral fractures, periosteal cloaking, coxa vara, and extraskeletal features (muscular hypotonia, cardiac abnormalities). Proband lumbar spine bone density z score was reduced [median -3.3 (range -4.77 to +0.1; n = 7)] and increased by +1.7 (1.17 to 3.0; n = 3) following bisphosphonate therapy. TMEM38B mutant bone has reduced trabecular bone volume, osteoblast, and particularly osteoclast numbers, with >80% reduction in bone resorption. Bone matrix mineralization is normal and nanoporosity low. We demonstrate a complex osteoblast differentiation defect with decreased expression of early markers and increased expression of late and mineralization-related markers. Predominance of trimeric intracellular cation channel type B over type A expression in murine osteoclasts supports an intrinsic osteoclast defect underlying low bone turnover. OI type XIV has a bone histology, matrix mineralization, and osteoblast differentiation pattern that is distinct from OI with collagen defects. Probands are responsive to bisphosphonates and some show muscular and cardiovascular features possibly related to intracellular calcium flux abnormalities. Copyright © 2017 Endocrine Society

TOPICAL REVIEW: Stem cell technology using bioceramics: hard tissue regeneration towards clinical application

NASA Astrophysics Data System (ADS)

Ohnishi, Hiroe; Oda, Yasuaki; Ohgushi, Hajime

2010-02-01

Mesenchymal stem cells (MSCs) are adult stem cells which show differentiation capabilities toward various cell lineages. We have already used MSCs for treatments of osteoarthritis, bone necrosis and bone tumor. For this purpose, culture expanded MSCs were combined with various ceramics and then implanted. Because of rejection response to allogeneic MSC implantation, we have utilized patients' own MSCs for the treatment. Bone marrow is a good cell source of MSCs, although the MSCs also exist in adipose tissue. When comparing osteogenic differentiation of these MSCs, bone marrow MSCs show more extensive bone forming capability than adipose MSCs. Thus, the bone marrow MSCs are useful for bone tissue regeneration. However, the MSCs show limited proliferation and differentiation capabilities that hindered clinical applications in some cases. Recent advances reveal that transduction of plural transcription factors into human adult cells results in generation of new type of stem cells called induced pluripotent stem cells (iPS cells). A drawback of the iPS cells for clinical applications is tumor formation after their in vivo implantation; therefore it is difficult to use iPS cells for the treatment. To circumvent the problem, we transduced a single factor of either SOX2 or NANOG into the MSCs and found high proliferation as well as osteogenic differentiation capabilities of the MSCs. The stem cells could be combined with bioceramics for clinical applications. Here, we summarize our recent technologies using adult stem cells in viewpoints of bone tissue regeneration.

Determination of osteogenic or adipogenic lineages in muscle-derived stem cells (MDSCs) by a collagen-binding peptide (CBP) derived from bone sialoprotein (BSP).

PubMed

Choi, Yoon Jung; Lee, Jue Yeon; Lee, Seung Jin; Chung, Chong-Pyoung; Park, Yoon Jeong

2012-03-09

Bone sialoprotein (BSP) is a mineralized, tissue-specific, non-collagenous protein that is normally expressed only in mineralized tissues such as bone, dentin, cementum, and calcified cartilage, and at sites of new mineral formation. The binding of BSP to collagen is thought to be important for initiating bone mineralization and bone cell adhesion to the mineralized matrix. Several recent studies have isolated stem cells from muscle tissue, but their functional properties are still unclear. In this study, we examined the effects of a synthetic collagen-binding peptide (CBP) on the differentiation efficiency of muscle-derived stem cells (MDSCs). The CBP sequence (NGVFKYRPRYYLYKHAYFYPHLKRFPVQ) corresponds to residues 35-62 of bone sialoprotein (BSP), which are located within the collagen-binding domain in BSP. Interestingly, this synthetic CBP inhibited adipogenic differentiation but increased osteogenic differentiation in MDSCs. The CBP also induced expression of osteoblastic marker proteins, including alkaline phosphatase (ALP), type I collagen, Runt-related transcription factor 2 (Runx2), and osteocalcin; prevented adipogenic differentiation in MDSCs; and down-regulated adipose-specific mRNAs, such as adipocyte protein 2 (aP2) and peroxisome proliferator-activated receptor γ. The CBP increased Extracellular signal-regulated kinases (ERK) 1/2 protein phosphorylation, which is important in lineage determination. These observations suggest that this CBP determines the osteogenic or adipogenic lineage in MDSCs by activating ERK1/2. Taken together, a novel CBP could be a useful candidate for regenerating bone and treating osteoporosis, which result from an imbalance in osteogenesis and adipogenesis differentiation. Copyright © 2012 Elsevier Inc. All rights reserved.

Genetic and molecular control of Osterix in skeletal formation

PubMed Central

Sinha, Krishna M.; Zhou, Xin

2013-01-01

Osteoblast differentiation is a multi-step process where mesenchymal cells differentiate into osteoblast lineage cells including osteocytes. Osterix (Osx) is an osteoblast-specific transcription factor which activates a repertoire of genes during differentiation of preosteoblasts into mature osteoblasts and osteocytes. The essential role of Osx in the genetic program of bone formation and in bone homeostasis is well established. Osx mutant embryos do not form bone and fail to express osteoblast-specific marker genes. Inactivation of Osx in mice after birth causes multiple skeletal phenotypes including lack of new bone formation, absence of resorption of mineralized cartilage, and defects in osteocyte maturation and function. Since Osx is a major effector in skeletal formation, studies on Osx gained momentum over the last five-seven years and implicated its important function in tooth formation as well as in healing of bone fractures. This review outlines mouse genetic studies that establish the essential role of Osx in bone and tooth formation as well as in healing of bone fractures. We also discuss the recent advances in regulation of Osx expression which is under control of a transcriptional network, signaling pathways, and epigenetic regulation. Finally we summarize important findings on the positive and negative regulation of Osx’s transcriptional activity through protein-protein interactions in expression of its target genes during osteoblast differentiation. In particular, the identification of the histone demethylase NO66 as an Osx-interacting protein which negatively regulates Osx activity opens further avenues in studying epigenetic control of Osx target genes during differentiation and maturation of osteoblasts. PMID:23225263

Bmp 2 and bmp 7 induce odonto- and osteogenesis of human tooth germ stem cells.

PubMed

Taşlı, P Neslihan; Aydın, Safa; Yalvaç, Mehmet Emir; Sahin, Fikrettin

2014-03-01

Bone morphogenetic proteins (BMPs) initiate, promote, and maintain odontogenesis and osteogenesis. In this study, we studied the effect of bone morphogenic protein 2 (BMP 2) and bone morphogenic protein 7 (BMP 7) as differentiation inducers in tooth and bone regeneration. We compared the effect of BMP 2 and BMP 7 on odontogenic and osteogenic differentiation of human tooth germ stem cells (hTGSCs). Third molar-derived hTGSCs were characterized with mesenchymal stem cell surface markers by flow cytometry. BMP 2 and BMP 7 were transfected into hTGSCs and the cells were seeded onto six-well plates. One day after the transfection, hTGSCs were treated with odontogenic and osteogenic mediums for 14 days. For confirmation of odontogenic and osteogenic differentiation, mRNA levels of BMP2, BMP 7, collagen type 1 (COL1A), osteocalsin (OCN), and dentin sialophosphoprotein (DSPP) genes were measured by quantitative real-time PCR. In addition to this, immunocytochemistry was performed by odontogenic and osteogenic antibodies and mineralization obtained by von Kossa staining. Our results showed that the BMP 2 and BMP 7 both promoted odontogenic and osteogenic differentiation of hTGSCs. Data indicated that BMP 2 treatment and BMP 7 treatment induce odontogenic differentiation without affecting each other, whereas they induce osteogenic differentiation by triggering expression of each other. These findings provide a feasible tool for tooth and bone tissue engineering.

Differentiation of osteoblast and osteoclast precursors on pure and silicon-substituted synthesized hydroxyapatites.

PubMed

Lehmann, Giorgia; Cacciotti, Ilaria; Palmero, Paola; Montanaro, Laura; Bianco, Alessandra; Campagnolo, Luisa; Camaioni, Antonella

2012-10-01

Calcium phosphate-based materials should show excellent bone-bonding and cell-mediated resorption characteristics at the same time, in order to be employed for bone replacement. In this perspective, pure (HAp) and silicon-substituted hydroxyapatite (Si-HAp, 1.4% wt) porous cylinders were prepared starting from synthesized powders and polyethylene spheres used as porogens, and investigated as supports for osteoblast and osteoclast progenitor differentiation. A systematic and detailed biological characterization is reported, in terms of cell adhesion, viability, proliferation, differentiation and bioresorption, aimed at proposing a complete and reliable picture of bone cell in vitro behavior, comprehensive of both the osteogenesis and the bone resorption processes. In order to achieve this purpose, cytocompatibility, differentiation and gene expression by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) were carried out using parietal bone-derived pre-osteoblasts obtained from neonatal mice and the bioresorption capability was assessed by seeding human peripheral blood monocytes, as osteoclast precursors. It resulted that both pure and Si-substituted HAps were able to promote differentiation of precursor cells in mature osteoblasts and osteoclasts. In particular, the Si-HAps enhanced the pre-osteoblast proliferation and showed higher osteoclast-mediated bioresorption capability, as supported by the presence of larger and more numerous resorption lacunae, whereas HAps promoted a more robust cell differentiation in terms of both osteocalcin gene expression by qRT-PCR and cell morphological evaluation by SEM analysis.

A comparative study of the proliferation and osteogenic differentiation of human periodontal ligament cells cultured on β-TCP ceramics and demineralized bone matrix with or without osteogenic inducers in vitro.

PubMed

An, Shaofeng; Gao, Yan; Huang, Xiangya; Ling, Junqi; Liu, Zhaohui; Xiao, Yin

2015-05-01

The repair of bone defects that result from periodontal diseases remains a clinical challenge for periodontal therapy. β-tricalcium phosphate (β-TCP) ceramics are biodegradable inorganic bone substitutes with inorganic components that are similar to those of bone. Demineralized bone matrix (DBM) is an acid-extracted organic matrix derived from bone sources that consists of the collagen and matrix proteins of bone. A few studies have documented the effects of DBM on the proliferation and osteogenic differentiation of human periodontal ligament cells (hPDLCs). The aim of the present study was to investigate the effects of inorganic and organic elements of bone on the proliferation and osteogenic differentiation of hPDLCs using three-dimensional porous β-TCP ceramics and DBM with or without osteogenic inducers. Primary hPDLCs were isolated from human periodontal ligaments. The proliferation of the hPDLCs on the scaffolds in the growth culture medium was examined using a Cell-Counting kit-8 (CCK-8) and scanning electron microscopy (SEM). Alkaline phosphatase (ALP) activity and the osteogenic differentiation of the hPDLCs cultured on the β-TCP ceramics and DBM were examined in both the growth culture medium and osteogenic culture medium. Specific osteogenic differentiation markers were examined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). SEM images revealed that the cells on the β-TCP were spindle-shaped and much more spread out compared with the cells on the DBM surfaces. There were no significant differences observed in cell proliferation between the β-TCP ceramics and the DBM scaffolds. Compared with the cells that were cultured on β-TCP ceramics, the ALP activity, as well as the Runx2 and osteocalcin (OCN) mRNA levels in the hPDLCs cultured on DBM were significantly enhanced both in the growth culture medium and the osteogenic culture medium. The organic elements of bone may exhibit greater osteogenic differentiation effects on hPDLCs than the inorganic elements.

Long-term culture and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized mesenchymal cells.

PubMed

Garba, Abubakar; Acar, Delphine D; Roukaerts, Inge D M; Desmarets, Lowiese M B; Devriendt, Bert; Nauwynck, Hans J

2017-09-01

Mesenchymal cells are multipotent stromal cells with self-renewal, differentiation and immunomodulatory capabilities. We aimed to develop a co-culture model for differentiating hematopoietic cells on top of immortalized mesenchymal cells for studying interactions between hematopoietic and mesenchymal cells, useful for adequately exploring the therapeutic potential of mesenchymal cells. In this study, we investigated the survival, proliferation and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized porcine bone marrow mesenchymal cells for a period of five weeks. Directly after collection, primary porcine bone marrow mesenchymal cells adhered firmly to the bottom of the culture plates and showed a fibroblast-like appearance, one week after isolation. Upon immortalization, porcine bone marrow mesenchymal cells were continuously proliferating. They were positive for simian virus 40 (SV40) large T antigen and the mesenchymal cell markers CD44 and CD55. Isolated red bone marrow cells were added to these immortalized mesenchymal cells. Five weeks post-seeding, 92±6% of the red bone marrow hematopoietic cells were still alive and their number increased 3-fold during five weekly subpassages on top of the immortalized mesenchymal cells. The red bone marrow hematopoietic cells were originally small and round; later, the cells increased in size. Some of them became elongated, while others remained round. Tiny dendrites appeared attaching hematopoietic cells to the underlying immortalized mesenchymal cells. Furthermore, weekly differential-quick staining of the cells indicated the presence of monoblasts, monocytes, macrophages and lymphocytes in the co-cultures. At three weeks of co-culture, flow cytometry analysis showed an increased surface expression of CD172a, CD14, CD163, CD169, CD4 and CD8 up to 37±0.8%, 40±8%, 41±4%, 23±3% and 19±5% of the hematopoietic cells, respectively. In conclusion, continuous mesenchymal cell cultures were successfully established and characterized and they supported the proliferation of red bone marrow hematopoietic cells, which finally differentiated into monocytic cells and CD4 + and CD8 + cells. Copyright © 2017. Published by Elsevier B.V.

Effects of Polymethoxyflavonoids on Bone Loss Induced by Estrogen Deficiency and by LPS-Dependent Inflammation in Mice.

PubMed

Matsumoto, Shigeru; Tominari, Tsukasa; Matsumoto, Chiho; Yoshinouchi, Shosei; Ichimaru, Ryota; Watanabe, Kenta; Hirata, Michiko; Grundler, Florian M W; Miyaura, Chisato; Inada, Masaki

2018-01-20

Polymethoxyflavonoids (PMFs) are a family of the natural compounds that mainly compise nobiletin, tangeretin, heptamethoxyflavone (HMF), and tetramethoxyflavone (TMF) in citrus fruits. PMFs have shown various biological functions, including anti-oxidative effects. We previously showed that nobiletin, tangeretin, and HMF all inhibited interleukin (IL)-1-mediated osteoclast differentiation via the inhibition of prostaglandin E2 synthesis. In this study, we created an original mixture of PMFs (nobiletin, tangeretin, HMF, and TMF) and examined whether or not PMFs exhibit co-operative inhibitory effects on osteoclastogenesis and bone resorption. In a coculture of bone marrow cells and osteoblasts, PMFs dose-dependently inhibited IL-1-induced osteoclast differentiation and bone resorption. The optimum concentration of PMFs was lower than that of nobiletin alone in the suppression of osteoclast differentiation, suggesting that the potency of PMFs was stronger than that of nobiletin in vitro. The oral administration of PMFs recovered the femoral bone loss induced by estrogen deficiency in ovariectomized mice. We further tested the effects of PMFs on lipopolysaccharide-induced bone resorption in mouse alveolar bone. In an ex vivo experimental model for periodontitis, PMFs significantly suppressed the bone-resorbing activity in organ cultures of mouse alveolar bone. These results indicate that a mixture of purified nobiletin, tangeretin, HMF, and TMF exhibits a co-operative inhibitory effect for the protection against bone loss in a mouse model of bone disease, suggesting that PMFs may be potential candidates for the prevention of bone resorption diseases, such as osteoporosis and periodontitis.

Mechanism of Action of Bortezomib and the New Proteasome Inhibitors on Myeloma Cells and the Bone Microenvironment: Impact on Myeloma-Induced Alterations of Bone Remodeling

PubMed Central

Accardi, Fabrizio; Toscani, Denise; Dalla Palma, Benedetta; Aversa, Franco; Giuliani, Nicola

2015-01-01

Multiple myeloma (MM) is characterized by a high capacity to induce alterations in the bone remodeling process. The increase in osteoclastogenesis and the suppression of osteoblast formation are both involved in the pathophysiology of the bone lesions in MM. The proteasome inhibitor (PI) bortezomib is the first drug designed and approved for the treatment of MM patients by targeting the proteasome. However, recently novel PIs have been developed to overcome bortezomib resistance. Interestingly, several preclinical data indicate that the proteasome complex is involved in both osteoclast and osteoblast formation. It is also evident that bortezomib either inhibits osteoclast differentiation induced by the receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL) or stimulates the osteoblast differentiation. Similarly, the new PIs including carfilzomib and ixazomib can inhibit bone resorption and stimulate the osteoblast differentiation. In a clinical setting, PIs restore the abnormal bone remodeling by normalizing the levels of bone turnover markers. In addition, a bone anabolic effect was described in responding MM patients treated with PIs, as demonstrated by the increase in the osteoblast number. This review summarizes the preclinical and clinical evidence on the effects of bortezomib and other new PIs on myeloma bone disease. PMID:26579531

Differential effects of intermittent and continuous administration of parathyroid hormone on bone histomorphometry and gene expression

NASA Technical Reports Server (NTRS)

Lotinun, Sutada; Sibonga, Jean D.; Turner, Russell T.

2002-01-01

A mechanism explaining the differential skeletal effects of intermittent and continuous elevation of serum parathyroid hormone (PTH) remains elusive. Intermittent PTH increases bone formation and bone mass and is being investigated as a therapy for osteoporosis. By contrast, chronic hyperparathyroidism results in the metabolic bone disease osteitis fibrosa characterized by osteomalacia, focal bone resorption, and peritrabecular bone marrow fibrosis. Intermittent and continuous PTH have similar effects on the number of osteoblasts and bone-forming activity. Many of the beneficial as well as detrimental effects of the hormone appear to be mediated by osteoblast-derived growth factors. This hypothesis was tested using cDNA microgene arrays to compare gene expression in tibia of rats treated with continuous and pulsatile administration of PTH. These treatments result in differential expression of many genes, including growth factors. One of the genes whose steady-state mRNA levels was increased by continuous but not pulsatile administration was platelet-derived growth factor-A (PDGF-A). Administration of a PDGF-A antagonist greatly reduced bone resorption, osteomalacia, and bone marrow fibrosis in a rat model for hyperparathyroidism, suggesting that PDGF-A is a causative agent for this disease. These findings suggest that profiling changes in gene expression can help identify the metabolic pathways responsible for the skeletal responses to the hormone.

Reactive oxygen species and oxidative stress in osteoclastogenesis, skeletal aging and bone diseases.

PubMed

Callaway, Danielle A; Jiang, Jean X

2015-07-01

Osteoclasts are cells derived from bone marrow macrophages and are important in regulating bone resorption during bone homeostasis. Understanding what drives osteoclast differentiation and activity is important when studying diseases characterized by heightened bone resorption relative to formation, such as osteoporosis. In the last decade, studies have indicated that reactive oxygen species (ROS), including superoxide and hydrogen peroxide, are crucial components that regulate the differentiation process of osteoclasts. However, there are still many unanswered questions that remain. This review will examine the mechanisms by which ROS can be produced in osteoclasts as well as how it may affect osteoclast differentiation and activity through its actions on osteoclastogenesis signaling pathways. In addition, the contribution of ROS to the aging-associated disease of osteoporosis will be addressed and how targeting ROS may lead to the development of novel therapeutic treatment options.

Fall-related self-efficacy, not balance and mobility performance, is related to accidental falls in chronic stroke survivors with low bone mineral density.

PubMed

Pang, M Y C; Eng, J J

2008-07-01

Chronic stroke survivors with low hip bone density are particularly prone to fractures. This study shows that fear of falling is independently associated with falls in this population. Thus, fear of falling should not be overlooked in the prevention of fragility fractures in these patients. Chronic stroke survivors with low bone mineral density (BMD) are particularly prone to fragility fractures. The purpose of this study was to identify the determinants of balance, mobility and falls in this sub-group of stroke patients. Thirty-nine chronic stroke survivors with low hip BMD (T-score <-1.0) were studied. Each subject was evaluated for the following: balance, mobility, leg muscle strength, spasticity, and fall-related self-efficacy. Any falls in the past 12 months were also recorded. Multiple regression analysis was used to identify the determinants of balance and mobility performance, whereas logistic regression was used to identify the determinants of falls. Multiple regression analysis revealed that after adjusting for basic demographics, fall-related self-efficacy remained independently associated with balance/mobility performance (R2 = 0.494, P < 0.001). Logistic regression showed that fall-related self-efficacy, but not balance and mobility performance, was a significant determinant of falls (odds ratio: 0.18, P = 0.04). Fall-related self-efficacy, but not mobility and balance performance, was the most important determinant of accidental falls. This psychological factor should not be overlooked in the prevention of fragility fractures among chronic stroke survivors with low hip BMD.

TGF-β and BMP Signaling in Osteoblast Differentiation and Bone Formation

PubMed Central

Chen, Guiqian; Deng, Chuxia; Li, Yi-Ping

2012-01-01

Transforming growth factor-beta (TGF-β)/bone morphogenic protein (BMP) signaling is involved in a vast majority of cellular processes and is fundamentally important throughout life. TGF-β/BMPs have widely recognized roles in bone formation during mammalian development and exhibit versatile regulatory functions in the body. Signaling transduction by TGF-β/BMPs is specifically through both canonical Smad-dependent pathways (TGF-β/BMP ligands, receptors and Smads) and non-canonical Smad-independent signaling pathway (e.g. p38 mitogen-activated protein kinase pathway, MAPK). Following TGF-β/BMP induction, both the Smad and p38 MAPK pathways converge at the Runx2 gene to control mesenchymal precursor cell differentiation. The coordinated activity of Runx2 and TGF-β/BMP-activated Smads is critical for formation of the skeleton. Recent advances in molecular and genetic studies using gene targeting in mice enable a better understanding of TGF-β/BMP signaling in bone and in the signaling networks underlying osteoblast differentiation and bone formation. This review summarizes the recent advances in our understanding of TGF-β/BMP signaling in bone from studies of genetic mouse models and human diseases caused by the disruption of TGF-β/BMP signaling. This review also highlights the different modes of cross-talk between TGF-β/BMP signaling and the signaling pathways of MAPK, Wnt, Hedgehog, Notch, and FGF in osteoblast differentiation and bone formation. PMID:22298955

Peptide-laden mesoporous silica nanoparticles with promoted bioactivity and osteo-differentiation ability for bone tissue engineering.

PubMed

Luo, Zuyuan; Deng, Yi; Zhang, Ranran; Wang, Mengke; Bai, Yanjie; Zhao, Qiang; Lyu, Yalin; Wei, Jie; Wei, Shicheng

2015-07-01

Combination of mesoporous silica materials and bioactive factors is a promising niche-mimetic solution as a hybrid bone substitution for bone tissue engineering. In this work, we have synthesized biocompatible silica-based nanoparticles with abundant mesoporous structure, and incorporated bone-forming peptide (BFP) derived from bone morphogenetic protein-7 (BMP-7) into the mesoporous silica nanoparticles (MSNs) to obtain a slow-release system for osteogenic factor delivery. The chemical characterization demonstrates that the small osteogenic peptide is encapsulated in the mesoporous successfully, and the nitrogen adsorption-desorption isotherms suggest that the peptide encapsulation has no influence on mesoporous structure of MSNs. In the cell experiment, the peptide-laden MSNs (p-MSNs) show higher MG-63 cell proliferation, spreading and alkaline phosphatase (ALP) activity than the bare MSNs, indicating good in vitro cytocompatibility. Simultaneously, the osteogenesis-related proteins expression and calcium mineral deposition disclose enhanced osteo-differentiation of human mesenchymal stem cells (hMSCs) under the stimulation of the p-MSNs, confirming that BFP released from MSNs could significantly promote the osteogenic differentiation of hMSCs, especially at 500μg/mL of p-MSNs concentration. The peptide-modified MSNs with better bioactivity and osteogenic differentiation make it a potential candidate as bioactive material for bone repairing, bone regeneration, and bio-implant coating applications. Copyright © 2015 Elsevier B.V. All rights reserved.

Novel antiosteoclastogenic activity of phloretin antagonizing RANKL-induced osteoclast differentiation of murine macrophages.

PubMed

Kim, Jung-Lye; Kang, Min-Kyung; Gong, Ju-Hyun; Park, Sin-Hye; Han, Seon-Young; Kang, Young-Hee

2012-08-01

Bone-remodeling imbalance resulting in more bone resorption than bone formation is known to cause skeletal diseases such as osteoporosis. Phloretin, a natural dihydrochalcone compound largely present in apple peels, possesses antiphotoaging, and antiinflammatory activity. Phloretin inhibited receptor activator of NF-κB ligand (RANKL)-induced formation of multinucleated osteoclasts and diminished bone resorption area produced during the osteoclast differentiation process. It was also found that ≥ 10 μM phloretin reduced RANKL-enhanced tartrate-resistance acid phosphatase activity and matrix metalloproteinase-9 secretion in a dose-dependent manner. The phloretin treatment retarded RANKL-induced expression of carbonic anhydrase II, vacuolar-type H(+) -ATPase D2 and β3 integrin, all involved in the bone resorption. Furthermore, submicromolar phloretin diminished the expression and secretion of cathepsin K elevated by RANKL, being concurrent with inhibition of TRAF6 induction and NF-κB activation. RANKL-induced activation of nuclear factor of activated T cells c1 (NFATc1) and microphthalmia-associated transcription factor was also suppressed by phloretin. These results demonstrate that the inhibition of osteoclast differentiation and bone resorption by phloretin entail a disturbance of TRAF6-NFATc1-NF-κB pathway triggered by RANKL. Therefore, phloretin may be a potential therapeutic agent targeting osteoclast differentiation and bone resorption in skeletal diseases such as osteoporosis. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Prostaglandin E2 acts via bone marrow macrophages to block PTH-stimulated osteoblast differentiation in vitro

PubMed Central

Choudhary, Shilpa; Blackwell, Katherine; Voznesensky, Olga; Roy, Abhijit Deb; Pilbeam, Carol

2014-01-01

Intermittent PTH is the major anabolic therapy for osteoporosis while continuous PTH causes bone loss. PTH acts on the osteoblast (OB) lineage to regulate bone resorption and formation. PTH also induces cyclooxygenase-2 (COX-2), producing prostaglandin E2 (PGE2) that can act on both OBs and osteoclasts (OCs). Because intermittent PTH is more anabolic in Cox-2 knockout (KO) than wild type (WT) mice, we hypothesized COX-2 might contribute to the effects of continuous PTH by suppressing PTH-stimulated differentiation of mesenchymal stem cells into OBs. We compared effects of continuous PTH on bone marrow stromal cells (BMSCs) and primary OBs (POBs) from Cox-2 KO mice, mice with deletion of PGE2 receptors (Ptger4 and Ptger2 KO mice), and WT controls. PTH increased OB differentiation in BMSCs only in the absence of COX-2 expression or activity. In the absence of COX-2, PTH stimulated differentiation if added during the first week of culture. In Cox-2 KO BMSCs, PTH-stimulated differentiation was prevented by adding PGE2 to cultures. Co-culture of POBs with M-CSF-expanded bone marrow macrophages (BMMs) showed that the inhibition of PTH-stimulated OB differentiation required not only COX-2 or PGE2 but also BMMs. Sufficient PGE2 to mediate the inhibitory effect was made by either WT POBs or WT BMMs. The inhibitory effect mediated by COX-2/PGE2 was transferred by conditioned media from RANKL-treated BMMs and could be blocked by osteoprotegerin, which interferes with RANKL binding to its receptor on OC lineage cells. Deletion of Ptger4, but not Ptger2, in BMMs prevented the inhibition of PTH-stimulated OB differentiation. As expected, PGE2 also stimulated OB differentiation, but when given in combination with PTH, the stimulatory effects of both were abrogated. These data suggest that PGE2, acting via EP4R on BMMs committed to the OC lineage, stimulated secretion of a factor or factors that acted to suppress PTH-stimulated OB differentiation. This suppression of OB differentiation could contribute to the bone loss seen with continuous PTH in vivo. PMID:23639875

Myelopotentiating effect of curcumin in tumor-bearing host: Role of bone marrow resident macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vishvakarma, Naveen Kumar; Kumar, Anjani; Kumar, Ajay

2012-08-15

The present investigation was undertaken to study if curcumin, which is recognized for its potential as an antineoplastic and immunopotentiating agent, can also influence the process of myelopoiesis in a tumor-bearing host. Administration of curcumin to tumor-bearing host augmented count of bone marrow cell (BMC) accompanied by an up-regulated BMC survival and a declined induction of apoptosis. Curcumin administration modulated expression of cell survival regulatory molecules: Bcl2, p53, caspase-activated DNase (CAD) and p53-upregulated modulator of apoptosis (PUMA) along with enhanced expression of genes of receptors for M-CSF and GM-CSF in BMC. The BMC harvested from curcumin-administered hosts showed an up-regulatedmore » colony forming ability with predominant differentiation into bone marrow-derived macrophages (BMDM), responsive for activation to tumoricidal state. The number of F4/80 positive bone marrow resident macrophages (BMM), showing an augmented expression of M-CSF, was also augmented in the bone marrow of curcumin-administered host. In vitro reconstitution experiments indicated that only BMM of curcumin-administered hosts, but not in vitro curcumin-exposed BMM, augmented BMC survival. It suggests that curcumin-dependent modulation of BMM is of indirect nature. Such prosurvival action of curcumin is associated with altered T{sub H1}/T{sub H2} cytokine balance in serum. Augmented level of serum-borne IFN-γ was found to mediate modulation of BMM to produce enhanced amount of monokines (IL-1, IL-6, TNF-α), which are suggested to augment the BMC survival. Taken together the present investigation indicates that curcumin can potentiate myelopoiesis in a tumor-bearing host, which may have implications in its therapeutic utility. Highlights: ► Curcumin augments myelopoiesis in tumor-bearing host. ► Bone marrow resident macrophages mediate curcumin-dependent augmented myelopoiesis. ► Serum borne cytokine are implicated in modulation of bone marrow resident macrophages.« less

Future human bone research in space

NASA Technical Reports Server (NTRS)

LeBlanc, A.; Shackelford, L.; Schneider, V.

1998-01-01

Skylab crewmembers demonstrated negative calcium (Ca) balance reaching about -300 mg/day by flight day 84. Limited bone density (BMD) measurements documented that bone was not lost equally from all parts of the skeleton. Subsequent BMD studies during long duration Russian flights documented the regional extent of bone loss. These studies demonstrated mean losses in the spine, femur neck, trochanter, and pelvis of about 1%-1.6% with large differences between individuals as well as between bone sites in a given individual. Limited available data indicate postflight bone recovery occurred in some individuals, but may require several years for complete restoration. Long duration bedrest studies showed a similar pattern of bone loss and calcium balance (-180 mg/day) as spaceflight. During long duration bedrest, resorption markers were elevated, formation markers were unchanged, 1,25 vitamin D (VitD) and calcium absorption were decreased, and serum ionized Ca was increased. Although this information is a good beginning, additional spaceflight research is needed to assess architectural and subregional bone changes, elucidate mechanisms, and develop efficient as well as effective countermeasures. Space research poses a number of unique problems not encountered in ground-based laboratory research. Therefore, researchers contemplating human spaceflight research need to consider a number of unique problems related to spaceflight in their experimental design.

Bone formation transcripts dominate the differential gene expression profile in an equine osteoporotic condition associated with pulmonary silicosis.

PubMed

Zavodovskaya, Regina; Stover, Susan M; Murphy, Brian G; Katzman, Scott; Durbin-Johnson, Blythe; Britton, Monica; Finno, Carrie J

2018-01-01

Osteoporosis has been associated with pulmonary silicosis in California horses exposed to soils rich in cytotoxic silica dioxide crystals, a syndrome termed silicate associated osteoporosis (SAO). The causal mechanism for the development of osteoporosis is unknown. Osteoporotic lesions are primarily located in bone marrow-rich sites such as ribs, scapula and pelvis. Gene transcription patterns within bone marrow and pulmonary lymph nodes of affected horses may offer clues to disease pathobiology. Bone marrow core and tracheobronchial lymph node tissue samples harvested postmortem from affected and unaffected horses were examined histologically and subjected to RNA sequencing (RNA-seq). Sequenced data were analyzed for differential gene expression and gene ontology. Metatranscriptomic and metagenomic assays evaluated samples for infectious agents. Thirteen of 17 differentially expressed transcripts in bone marrow were linked to bone and cartilage formation such as integrin binding bone sialoprotein (log2FC = 3.39, PFDR = 0.013) and chondroadherin (log2FC = 4.48, PFDR = 0.031). Equus caballus solute carrier family 9, subfamily A2 (log2FC = 3.77, PFDR = 0.0034) was one of the four differentially expressed transcripts linked to osteoclast activity. Osteoblasts were hyperplastic and hypertrophic in bone marrow from affected horses. Biological pathways associated with skeletal morphogenesis were significantly enriched in affected horses. The 30 differentially expressed genes in affected lymph nodes were associated with inflammatory responses. Evidence of infectious agents was not found. The SAO affected bone marrow molecular signature demonstrated increased transcription and heightened activation of osteoblasts. Increased osteoblastic activity could be part of the pathological mechanism for osteoporosis or a compensatory response to the accelerated osteolysis. Transcriptome data offer gene targets for inquiries into the role of osteocytes and osteoblasts in SAO pathogenesis. Viral or bacterial infectious etiology in SAO is less likely based on metatranscriptomic and metagenomic data but cannot be completely ruled out.

Osteoinductive ceramics as a synthetic alternative to autologous bone grafting

PubMed Central

Yuan, Huipin; Fernandes, Hugo; Habibovic, Pamela; de Boer, Jan; Barradas, Ana M. C.; de Ruiter, Ad; Walsh, William R.; van Blitterswijk, Clemens A.; de Bruijn, Joost D.

2010-01-01

Biomaterials can be endowed with biologically instructive properties by changing basic parameters such as elasticity and surface texture. However, translation from in vitro proof of concept to clinical application is largely missing. Porous calcium phosphate ceramics are used to treat small bone defects but in general do not induce stem cell differentiation, which is essential for regenerating large bone defects. Here, we prepared calcium phosphate ceramics with varying physicochemical and structural characteristics. Microporosity correlated to their propensity to stimulate osteogenic differentiation of stem cells in vitro and bone induction in vivo. Implantation in a large bone defect in sheep unequivocally demonstrated that osteoinductive ceramics are equally efficient in bone repair as autologous bone grafts. Our results provide proof of concept for the clinical application of “smart” biomaterials. PMID:20643969

Rapidly assessing changes in bone mineral balance using natural stable calcium isotopes

PubMed Central

Morgan, Jennifer L. L.; Skulan, Joseph L.; Gordon, Gwyneth W.; Romaniello, Stephen J.; Smith, Scott M.; Anbar, Ariel D.

2012-01-01

The ability to rapidly detect changes in bone mineral balance (BMB) would be of great value in the early diagnosis and evaluation of therapies for metabolic bone diseases such as osteoporosis and some cancers. However, measurements of BMB are hampered by difficulties with using biochemical markers to quantify the relative rates of bone resorption and formation and the need to wait months to years for altered BMB to produce changes in bone mineral density large enough to resolve by X-ray densitometry. We show here that, in humans, the natural abundances of Ca isotopes in urine change rapidly in response to changes in BMB. In a bed rest experiment, use of high-precision isotope ratio MS allowed the onset of bone loss to be detected in Ca isotope data after about 1 wk, long before bone mineral density has changed enough to be detectable with densitometry. The physiological basis of the relationship between Ca isotopes and BMB is sufficiently understood to allow quantitative translation of changes in Ca isotope abundances to changes in bone mineral density using a simple model. The rate of change of bone mineral density inferred from Ca isotopes is consistent with the rate observed by densitometry in long-term bed rest studies. Ca isotopic analysis provides a powerful way to monitor bone loss, potentially making it possible to diagnose metabolic bone disease and track the impact of treatments more effectively than is currently possible. PMID:22652567

Osteoinduction and survival of osteoblasts and bone-marrow stromal cells in 3D biphasic calcium phosphate scaffolds under static and dynamic culture conditions.

PubMed

Rath, Subha N; Strobel, Leonie A; Arkudas, Andreas; Beier, Justus P; Maier, Anne-Kathrin; Greil, Peter; Horch, Raymund E; Kneser, Ulrich

2012-10-01

In many tissue engineering approaches, the basic difference between in vitro and in vivo conditions for cells within three-dimensional (3D) constructs is the nutrition flow dynamics. To achieve comparable results in vitro, bioreactors are advised for improved cell survival, as they are able to provide a controlled flow through the scaffold. We hypothesize that a bioreactor would enhance long-term differentiation conditions of osteogenic cells in 3D scaffolds. To achieve this either primary rat osteoblasts or bone marrow stromal cells (BMSC) were implanted on uniform-sized biphasic calcium phosphate (BCP) scaffolds produced by a 3D printing method. Three types of culture conditions were applied: static culture without osteoinduction (Group A); static culture with osteoinduction (Group B); dynamic culture with osteoinduction (Group C). After 3 and 6 weeks, the scaffolds were analysed by alkaline phosphatase (ALP), dsDNA amount, SEM, fluorescent labelled live-dead assay, and real-time RT-PCR in addition to weekly alamarBlue assays. With osteoinduction, increased ALP values and calcium deposition are observed; however, under static conditions, a significant decrease in the cell number on the biomaterial is observed. Interestingly, the bioreactor system not only reversed the decreased cell numbers but also increased their differentiation potential. We conclude from this study that a continuous flow bioreactor not only preserves the number of osteogenic cells but also keeps their differentiation ability in balance providing a suitable cell-seeded scaffold product for applications in regenerative medicine. © 2012 The Authors Journal of Cellular and Molecular Medicine © 2012 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

Dual Role of Cyanidin-3-glucoside on the Differentiation of Bone Cells.

PubMed

Park, K H; Gu, D R; So, H S; Kim, K J; Lee, S H

2015-12-01

Cyanidin-3-glucoside (C3G) is one of the major components of anthocyanin, a water-soluble phytochemical. Recent studies demonstrated the chemopreventive and chemotherapeutic activities of C3G in various conditions, including cancer, although the precise effects of C3G on osteoclast and osteoblast differentiation remain unclear. Here, we investigated the role of C3G in the differentiation of bone-associated cells and its underlying mechanism. C3G inhibited receptor activator of nuclear factor kappa-B ligand (RANKL)-mediated osteoclast differentiation and formation in a dose-dependent manner and downregulated the expression of osteoclast differentiation marker genes. Pretreatment with C3G considerably reduced the induction of extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated kinases activation by RANKL in osteoclast precursor cells. Furthermore, C3G dramatically inhibited the expression of c-Fos and nuclear factor of activated T-cells, cytoplasmic 1, which are important transcription factors for osteoclast differentiation and activation. The formation of osteoclasts in coculture of bone marrow cells and calvaria-derived osteoblasts was also inhibited by C3G treatment, although the expression of macrophage colony-stimulating factor and RANKL (master factors for osteoclast differentiation and formation) and osteoprotegerin (a decoy receptor for RANKL) on osteoblasts was unaffected. The inhibitory effect of C3G on osteoclastogenesis is therefore targeted specifically to osteoclasts but not osteoblasts. Moreover, analysis of the expression levels of osteoblast differentiation marker genes and alizarin red staining showed that osteoblast differentiation and matrix formation increased after C3G treatment. Taken together, these results strongly suggest that C3G has a dual role in bone metabolism, as an effective inhibitor of osteoclast differentiation but also as an activator of osteoblast differentiation. Therefore, C3G may be used as a potent preventive or therapeutic agent for bone-related diseases, such as osteoporosis, rheumatoid arthritis, and periodontitis. © International & American Associations for Dental Research 2015.

Role of apparent diffusion coefficients with diffusion-weighted magnetic resonance imaging in differentiating between benign and malignant bone tumors.

PubMed

Wang, Tingting; Wu, Xiangru; Cui, Yanfen; Chu, Caiting; Ren, Gang; Li, Wenhua

2014-11-29

Benign and malignant bone tumors can present similar imaging features. This study aims to evaluate the significance of apparent diffusion coefficients (ADC) in differentiating between benign and malignant bone tumors. A total of 187 patients with 198 bone masses underwent diffusion-weighted (DW) magnetic resonance (MR) imaging. The ADC values in the solid components of the bone masses were assessed. Statistical differences between the mean ADC values in the different tumor types were determined by Student's t-test. Histological analysis showed that 84/198 (42.4%) of the bone masses were benign and 114/198 (57.6%) were malignant. There was a significant difference between the mean ADC values in the benign and malignant bone lesions (P<0.05). However, no significant difference was found in the mean ADC value between non-ossifying fibromas, osteofibrous dysplasia, and malignant bone tumors. When an ADC cutoff value≥1.10×10(-3) mm2/s was applied, malignant bone lesions were excluded with a sensitivity of 89.7%, a specificity of 84.5%, a positive predictive value of 82.6%, and a negative predictive value of 95.3%. The combination of DW imaging with ADC quantification and T2-weighted signal characteristics of the solid components in lesions can facilitate differentiation between benign and malignant bone tumors.

Receptor tyrosine kinase inhibition causes simultaneous bone loss and excess bone formation within growing bone in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nurmio, Mirja, E-mail: Mirja.Nurmio@utu.fi; Department of Pediatrics, University of Turku; Joki, Henna, E-mail: Henna.Joki@utu.fi

During postnatal skeletal growth, adaptation to mechanical loading leads to cellular activities at the growth plate. It has recently become evident that bone forming and bone resorbing cells are affected by the receptor tyrosine kinase (RTK) inhibitor imatinib mesylate (STI571, Gleevec (registered)) . Imatinib targets PDGF, ABL-related gene, c-Abl, c-Kit and c-Fms receptors, many of which have multiple functions in the bone microenvironment. We therefore studied the effects of imatinib in growing bone. Young rats were exposed to imatinib (150 mg/kg on postnatal days 5-7, or 100 mg/kg on postnatal days 5-13), and the effects of RTK inhibition on bonemore » physiology were studied after 8 and 70 days (3-day treatment), or after 14 days (9-day treatment). X-ray imaging, computer tomography, histomorphometry, RNA analysis and immunohistochemistry were used to evaluate bone modeling and remodeling in vivo. Imatinib treatment eliminated osteoclasts from the metaphyseal osteochondral junction at 8 and 14 days. This led to a resorption arrest at the growth plate, but also increased bone apposition by osteoblasts, thus resulting in local osteopetrosis at the osteochondral junction. The impaired bone remodelation observed on day 8 remained significant until adulthood. Within the same bone, increased osteoclast activity, leading to bone loss, was observed at distal bone trabeculae on days 8 and 14. Peripheral quantitative computer tomography (pQCT) and micro-CT analysis confirmed that, at the osteochondral junction, imatinib shifted the balance from bone resorption towards bone formation, thereby altering bone modeling. At distal trabecular bone, in turn, the balance was turned towards bone resorption, leading to bone loss. - Research Highlights: > 3-Day imatinib treatment. > Causes growth plate anomalies in young rats. > Causes biomechanical changes and significant bone loss at distal trabecular bone. > Results in loss of osteoclasts at osteochondral junction.« less

Bone Marrow Stromal Cells Contribute to Bone Formation Following Infusion into Femoral Cavities of a Mouse Model of Osteogenesis Imperfecta

PubMed Central

Li, Feng; Wang, Xujun; Niyibizi, Christopher

2010-01-01

Currently, there are conflicting data in literature regarding contribution of bone marrow stromal cells (BMSCs) to bone formation when the cells are systemically delivered in recipient animals. To understand if BMSCs contribute to bone cell phenotype and bone formation in osteogenesis imperfecta bones (OI), MSCs marked with GFP were directly infused into the femurs of a mouse model of OI (oim). The contribution of the cells to the cell phenotype and bone formation was assessed by histology, immunohistochemistry and biomechanical loading of recipient bones. Two weeks following infusion of BMSCs, histological examination of the recipient femurs demonstrated presence of new bone when compared to femurs injected with saline which showed little or no bone formation. The new bone contained few donor cells as demonstrated by GFP fluorescence. At six weeks following cell injection, new bone was still detectable in the recipient femurs but was enhanced by injection of the cells suspended in pepsin solublized type I collagen. Immunofluorescence and immunohistochemical staining showed that donor GFP positive cells in the new bone were localized with osteocalcin expressing cells suggesting that the cells differentiated into osteoblasts in vivo. Biomechanical loading to failure in thee point bending, revealed that, femurs infused with BMSCs in PBS or in soluble type I collagen were biomechanically stronger than those injected with PBS or type I collagen alone. Taken together, the results indicate that transplanted cells differentiated into osteoblasts in vivo and contributed to bone formation in vivo; we also speculate that donor cells induced differentiation or recruitment of endogenous cells to initiate reparative process at early stages following transplantation. PMID:20570757

Divalent Metal Ions Induced Osteogenic Differentiation of MC3T3E1

NASA Astrophysics Data System (ADS)

Wang, Guoshou; Su, Wenta; Chen, Pohung; Huang, Teyang

2017-12-01

Biomaterial scaffolds blended with biochemical signal molecules with adequate osteoinductive and osteoconductive properties have attracted significant interest in bone tissue engineering regeneration. The divalent metal ions can gradually release from the scaffold into the culture medium and then induced osteoblastic differentiation of MC3T3E1. These MC3T3E1 cells expressed high activity of alkaline phosphatase, bone-related gene expression of collagen type I, Runx2, osteopontin, osteocalcin, and significantly enhanced deposited minerals on scaffold after 21 days of culture. This experiment provided a useful inducer for osteogenic differentiation in bone repair.

A fat option for the pig: Hepatocytic differentiated mesenchymal stem cells for translational research

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brückner, Sandra, E-mail: sandra.brueckner@medizin.uni-leipzig.de; Tautenhahn, Hans-Michael, E-mail: hans-michael.tautenhahn@medizin.uni-leipzig.de; TRM, Translational Centre for Regenerative Medicine, Philipp-Rosenthal-Str. 55, Leipzig D-04103

Study background: Extended liver resection is the only curative treatment option of liver cancer. Yet, the residual liver may not accomplish the high metabolic and regenerative capacity needed, which frequently leads to acute liver failure. Because of their anti-inflammatory and -apoptotic as well as pro-proliferative features, mesenchymal stem cells differentiated into hepatocyte-like cells might provide functional and regenerative compensation. Clinical translation of basic research requires pre-clinical approval in large animals. Therefore, we characterized porcine mesenchymal stem cells (MSC) from adipose tissue and bone marrow and their hepatocyte differentiation potential for future assessment of functional liver support after surgical intervention inmore » the pig model. Methods: Mesenchymal surface antigens and multi-lineage differentiation potential of porcine MSC isolated by collagenase digestion either from bone marrow or adipose tissue (subcutaneous/visceral) were assessed by flow cytometry. Morphology and functional properties (urea-, glycogen synthesis and cytochrome P450 activity) were determined during culture under differentiation conditions and compared with primary porcine hepatocytes. Results: MSC from porcine adipose tissue and from bone marrow express the typical mesenchymal markers CD44, CD29, CD90 and CD105 but not haematopoietic markers. MSC from both sources displayed differentiation into the osteogenic as well as adipogenic lineage. After hepatocyte differentiation, expression of CD105 decreased significantly and cells adopted the typical polygonal morphology of hepatocytes. Glycogen storage was comparable in adipose tissue- and bone marrow-derived cells. Urea synthesis was about 35% lower in visceral than in subcutaneous adipose tissue-derived MSC. Cytochrome P450 activity increased significantly during differentiation and was twice as high in hepatocyte-like cells generated from bone marrow as from adipose tissue. Conclusion: The hepatocyte differentiation of porcine adipose tissue-derived MSC was shown for the first time yielding hepatocyte-like cells with specific functions similar in bone marrow and subcutaneous adipose tissue-derived MSC. That makes them good pre-clinical candidates for supportive approaches after liver resection in the pig. - Highlights: • First time to show hepatocytic differentiation of porcine adipose tissue-derived MSC. • Hepatocytic-differentiated MSC display metabolic qualities of primary hepatocytes. • Metabolic potency varies between differentiated MSC from different tissues. • MSC are good candidates for pre-clinical evaluation of stem cell-based therapies.« less

Intestinal absorption and renal reabsorption of calcium throughout postnatal development

PubMed Central

Beggs, Megan R

2017-01-01

Calcium is vital for many physiological functions including bone mineralization. Postnatal deposition of calcium into bone is greatest in infancy and continues through childhood and adolescence until peek mineral density is reached in early adulthood. Thereafter, bone mineral density remains static until it eventually declines in later life. A positive calcium balance, i.e. more calcium absorbed than excreted, is crucial to bone deposition during growth and thus to peek bone mineral density. Dietary calcium is absorbed from the intestine into the blood. It is then filtered by the renal glomerulus and either reabsorbed by the tubule or excreted in the urine. Calcium can be (re)absorbed across intestinal and renal epithelia via both transcellular and paracellular pathways. Current evidence suggests that significant intestinal and renal calcium transport changes occur throughout development. However, the molecular details of these alterations are incompletely delineated. Here we first briefly review the current model of calcium transport in the intestine and renal tubule in the adult. Then, we describe what is known with regard to calcium handling through postnatal development, and how alterations may aid in mediating a positive calcium balance. The role of transcellular and paracellular calcium transport pathways and the contribution of specific intestinal and tubular segments vary with age. However, the current literature highlights knowledge gaps in how specifically intestinal and renal calcium (re)absorption occurs early in postnatal development. Future research should clarify the specific changes in calcium transport throughout early postnatal development including mediators of these alterations enabling appropriate bone mineralization. Impact statement This mini review outlines the current state of knowledge pertaining to the molecules and mechanisms maintaining a positive calcium balance throughout postnatal development. This process is essential to achieving optimal bone mineral density in early adulthood, thereby lowering the lifetime risk of osteoporosis. PMID:28346014

Long Noncoding RNAs: New Players in the Osteogenic Differentiation of Bone Marrow- and Adipose-Derived Mesenchymal Stem Cells.

PubMed

Yang, Qiaolin; Jia, Lingfei; Li, Xiaobei; Guo, Runzhi; Huang, Yiping; Zheng, Yunfei; Li, Weiran

2018-06-01

Mesenchymal stem cells (MSCs) are an important population of multipotent stem cells that differentiate into multiple lineages and display great potential in bone regeneration and repair. Although the role of protein-coding genes in the osteogenic differentiation of MSCs has been extensively studied, the functions of noncoding RNAs in the osteogenic differentiation of MSCs are unclear. The recent application of next-generation sequencing to MSC transcriptomes has revealed that long noncoding RNAs (lncRNAs) are associated with the osteogenic differentiation of MSCs. LncRNAs are a class of non-coding transcripts of more than 200 nucleotides in length. Noncoding RNAs are thought to play a key role in osteoblast differentiation through various regulatory mechanisms including chromatin modification, transcription factor binding, competent endogenous mechanism, and other post-transcriptional mechanisms. Here, we review the roles of lncRNAs in the osteogenic differentiation of bone marrow- and adipose-derived stem cells and provide a theoretical foundation for future research.

Adrenaline inhibits osteogenesis via repressing miR-21 expression.

PubMed

Chen, Danying; Wang, Zuolin

2017-01-01

Sympathetic signaling is involved in bone homeostasis; however, the cellular and molecular mechanisms remain unknown. In this study, we found that the psychological stress mediator adrenaline inhibited osteogenic differentiation of human bone marrow-derived stem cells (hMSC) by reducing microRNA-21 (miR-21) expression. Briefly, adrenaline significantly inhibited the osteogenic differentiation of hMSCs, as observed with both Alizarin red staining and maker gene expression (RUNX2, OSX, OCN, and OPN). During this process, miR-21 was suppressed by adrenaline via inhibition of histone acetylation, as verified by H3K9Ac chromatin immunoprecipitation (ChIP) assay. MiR-21 was confirmed to promote hMSC osteogenic differentiation, and overexpression of miR-21 reversed the impeditive effect of adrenaline on hMSC osteogenic differentiation. Our results demonstrate that down-regulation of miR-21 is responsible for the adrenaline-mediated inhibition of hMSC osteogenic differentiation. These findings indicate a regulation of bone metabolism by psychological stress and also provide a molecular basis for psychological stress-associated bone diseases. © 2016 International Federation for Cell Biology.

A novel pyrazole derivative protects from ovariectomy-induced osteoporosis through the inhibition of NADPH oxidase

PubMed Central

Joo, Jung Hee; Huh, Jeong-Eun; Lee, Jee Hyun; Park, Doo Ri; Lee, Yoonji; Lee, Seul Gee; Choi, Sun; Lee, Hwa Jeong; Song, Seong-Won; Jeong, Yongmi; Goo, Ja-Il; Choi, Yongseok; Baek, Hye Kyung; Yi, Sun Shin; Park, Soo Jin; Lee, Ji Eun; Ku, Sae Kwang; Lee, Won Jae; Lee, Kee-In; Lee, Soo Young; Bae, Yun Soo

2016-01-01

Osteoclast cells (OCs) are differentiated from bone marrow-derived macrophages (BMMs) by activation of receptor activator of nuclear factor κB (NF-κB) ligand (RANKL). Activation of NADPH oxidase (Nox) isozymes is involved in RANKL-dependent OC differentiation, implicating Nox isozymes as therapeutic targets for treatment of osteoporosis. Here, we show that a novel pyrazole derivative, Ewha-18278 has high inhibitory potency on Nox isozymes. Blocking the activity of Nox with Ewha-18278 inhibited the responses of BMMs to RANKL, including reactive oxygen species (ROS) generation, activation of mitogen-activated protein (MAP) kinases and NF-κB, and OC differentiation. To evaluate the anti-osteoporotic function of Ewha-18278, the derivative was applied to estrogen-deficient ovariectomized (OVX) ddY mice. Oral administration of Ewha-18278 (10 mg/kg/daily, 4 weeks) into the mice recovered bone mineral density, trabecular bone volume, trabecular bone length, number and thickness, compared to control OVX ddY mice. Moreover, treatment of OVX ddY mice with Ewha-18278 increased bone strength by increasing cortical bone thickness. We provide that Ewha-18278 displayed Nox inhibition and blocked the RANKL-dependent cell signaling cascade leading to reduced differentiation of OCs. Our results implicate Ewha-18278 as a novel therapeutic agent for the treatment of osteoporosis. PMID:26975635

The effect of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow‑derived mesenchymal stem cells.

PubMed

Javanmard, F; Azadbakht, M; Pourmoradi, M

2016-01-01

In this study, the role of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow mesenchymal stem cells were investigated. The cells were cultured in treatment medium containing 100 nM of staurosporine for 4 hours; then the cells were affected by hydrostatic pressure (0, 25,50, 100 mmHg). The percentage of cell viability by trypan blue staining and the percentage of cell death by Hoechst/PI differential staining were assessed. We obtained the total neurite length. Expression of β-tubulin III and GFAP (Glial fibrillary acidic protein) proteins were also analyzed by immunocytochemistry. The percentage of cell viability in treatments decreased relative to the increase in hydrostatic pressure and time (p Keywords: bone marrow mesenchymal stem cell, hydrostatic pressure, immunocytochemistry, neural differentiation, neurite length, cell differentiation.

Inhibition of bone resorption by Tanshinone VI isolated from Salvia miltiorrhiza Bunge.

PubMed

Nicolin, V; Dal Piaz, F; Nori, S L; Narducci, P; De Tommasi, N

2010-05-10

During the last decade, a more detailed knowledge of molecular mechanisms involved in osteoclastogenesis has driven research efforts in the development and screening of compound libraries of several small molecules that specifically inhibit the pathway involved in the commitment of the osteoclast precursor cells. Natural compounds that suppress osteoclast differentiation may have therapeutic value in treating osteoporosis and other bone erosive diseases such as rheumatoid arthritis or metastasis associated with bone loss. In ongoing investigation into anti-osteoporotic compounds from natural products we have analyzed the effect of Tanshinone VI on osteoclasts differentiation, using a physiologic three-dimensional osteoblast/bone marrow model of cell co-culture. Tanshinone VI is an abietane diterpene extracted from the root of Salvia miltiorrhiza Bunge (Labiatae), a Chinese traditional crude drug, "Tan-Shen". Tashinone has been widely used in clinical practice for the prevention of cardiac diseases, arthritis and other inflammation-related disorders based on its pharmacological actions in multiple tissues. Although Tanshinone VI A has been used as a medicinal agent in the treatment of many diseases, its role in osteoclast-related bone diseases remains unknown. We showed previously that Tanshinone VI greatly inhibits osteoclast differentiation and suppresses bone resorption through disruption of the actin ring; subsequently, we intended to examine the precise inhibitory mechanism of Tanshinone VI on osteoclast differentiating factor. This study shows, for the first time, that Tanshinone VI prevents osteoclast differentiation by inhibiting RANKL expression and NFkB induction.

Long-Duration Spaceflight During the Bion-M1 Spaceflight Experiment Resulted in Significant Bone Loss in the Femoral Head and Alterations in Stem Cell Differentiation Potential in Male Mice

NASA Astrophysics Data System (ADS)

Blaber, Elizabeth; Almeida, Eduardo; Grigoryan, Eleonora; Globus, Ruth

Scientific understanding of the effects of microgravity on mammalian physiology has been limited to short duration spaceflight experiments (10-15 days). As long duration and inter-planetary missions are being initiated, there is a great need to understand the long-term effects of spaceflight on various physiological processes, including stem cell-based tissue regeneration. Bion-M1, for the first time, enabled the possibility of studying the effects of 30-days of microgravity exposure on a mouse model with sufficient sample size to enable statistical analysis. In this experiment, we hypothesized that microgravity negatively impacts stem cell based tissue regeneration, such as bone remodeling and regeneration from hematopoietic and mesenchymal precursors, thereby resulting in tissue degeneration in mice exposed to spaceflight. To test this hypothesis we collected the pelvis and proximal femur from space-flown mice and asynchronous ground controls and analyzed bone and bone marrow using techniques including Microcomputed Tomography (MicroCT), and in-vitro differentiation and differentiating cell motility assays. To determine the effects of 30-days spaceflight on bone tissue mass, we used MicroCT to analyze the trabecular bone of the femoral head and the cortical bone of the femoral neck and mid-shaft. We found that spaceflight caused a 45% decrease in bone volume ratio, a 17% decrease in trabecular thickness, a 25% decrease in trabecular number, and a 17% increase in trabecular spacing of trabecular bone. Furthermore, structural model index and trabecular pattern factor were increased by 32% and 82% respectively indicating that 30-days spaceflight resulted not only in a large loss of trabecular bone but also in a decrease of bone strength indicators. Analysis of the femoral neck cortical bone showed an increase in marrow area and cortical porosity indicating an overall widening of the femoral neck. Interestingly, no significant alterations were found in the cortical bone of the femoral mid-shaft. To determine the regenerative potential of osteoblasts derived from mesenchymal stem cells flown in microgravity we conducted post-flight in-vitro osteoblastogenesis and mineralized nodule formation assays. We found an increase in post-flight differentiation and mineralization of microgravity-flown osteogenic cells, suggesting an accumulation of precursor cells that fail to fully differentiate in space, and then resume vigorous osteogenesis upon reloading at 1g. Overall, these preliminary results indicate that exposure to 30-days spaceflight causes significant trabecular bone loss in the femoral head, a decrease in trabecular bone strength indicators, and compensatory widening of the femoral neck. These results, coupled with diminished regenerative potential of bone marrow stem cells during mechanical unloading in microgravity, have potentially serious implications for bone health and fracture risk during long-duration spaceflight.

Treatment of giant cell tumor of bone: Current concepts.

PubMed

Puri, Ajay; Agarwal, Manish

2007-04-01

Giant cell tumor (GCT) of bone though one of the commonest bone tumors encountered by an orthopedic surgeon continues to intrigue treating surgeons. Usually benign, they are locally aggressive and may occasionally undergo malignant transformation. The surgeon needs to strike a balance during treatment between reducing the incidence of local recurrence while preserving maximal function.Differing opinions pertaining to the use of adjuvants for extension of curettage, the relative role of bone graft or cement to pack the defect and the management of recurrent lesions are some of the issues that offer topics for eternal debate.Current literature suggests that intralesional curettage strikes the best balance between controlling disease and preserving optimum function in the majority of the cases though there may be occasions where the extent of the disease mandates resection to ensure adequate disease clearance.An accompanying treatment algorithm helps outline the management strategy in GCT.

Non-invasive photo acoustic approach for human bone diagnosis.

PubMed

Thella, Ashok Kumar; Rizkalla, James; Helmy, Ahdy; Suryadevara, Vinay Kumar; Salama, Paul; Rizkalla, Maher

2016-12-01

The existing modalities of bone diagnosis including X-ray and ultrasound may cite drawback in some cases related to health issues and penetration depth, while the ultrasound modality may lack image quality. Photo acoustic approach however, provides light energy to the acoustic wave, enabling it to activate and respond according to the propagating media (which is type of bones in this case). At the same time, a differential temperature change may result in the bio heat response, resulting from the heat absorbed across the multiple materials under study. In this work, we have demonstrated the features of using photo acoustic modality in order to non-invasively diagnose the type of human bones based on their electrical, thermal, and acoustic properties that differentiate the output response of each type. COMSOL software was utilized to combine both acoustic equations and bio heat equations, in order to study both the thermal and acoustic responses through which the differential diagnosis can be obtained. In this study, we solved both the acoustic equation and bio heat equations for four types of bones, bone (cancellous), bone (cortical), bone marrow (red), and bone marrow (yellow). 1 MHz acoustic source frequency was chosen and 10(5) W/m(2) power source was used in the simulation. The simulation tested the dynamic response of the wave over a distance of 5 cm from each side for the source. Near 2.4 cm was detected from simulation from each side of the source with a temperature change of within 0.5 K for various types of bones, citing a promising technique for a practical model to detect the type of bones via the differential temperature as well as the acoustic was response via the multiple materials associated with the human bones (skin and blood). The simulation results suggest that the PA technique may be applied to non-invasive diagnosis for the different types of bones, including cancerous bones. A practical model for detecting both the temperature change via IR sensors, and acoustic wave signals may be detected via sensitive pressure transducer, which is reserved for future realization.

The transcription factor E74-like factor controls quiescence of endothelial cells and their resistance to myeloablative treatments in bone marrow

USDA-ARS?s Scientific Manuscript database

The regeneration of the hematopoietic system in bone marrow after chemotherapy depends on a balance between the quiescence and proliferation of lineage-specific progenitor cells. Even though the vascular network in bone is damaged by cytoablation, the transcriptional control of quiescence in endothe...

Osteoporosis and gait and balance disturbances in older sarcopenic obese New Zealanders.

PubMed

Waters, D L; Hale, L; Grant, A M; Herbison, P; Goulding, A

2010-02-01

Bone, muscle, and fat may affect gait and balance in older adults. Osteoporosis was prevalent in low muscle mass participants and related to gait and balance deficits. Low muscle combined with high fat mass had more functional deficits and poorer bone health, which has implications for falls risk and fractures. Decreasing bone density and muscle mass and increasing fat mass may act synergistically to affect gait and balance in older adults. One hundred eighty-three older adults (age 72.7 +/- 6 years, range 56-93; body mass index 28.2 +/- 4.9, range 16.6-46.0) were recruited from a New Zealand falls prevention intervention trial. Total and appendicular skeletal muscle mass (ASM), percent fat, and bone mineralization were assessed by dual energy X-ray absorptiometry and used to characterize normal lean (NL, n = 51), sarcopenic (SS, n = 18), sarcopenic obese (SO, n = 29), and obese (OO, n = 85) phenotypes. Functional performance was assessed using timed up and go, chair stand, single leg stand, and step test. Regression models were adjusted for age, sex, medications, and physical activity. Femoral neck osteoporosis was present in 22% SS, 17% SO, 12% NL, and 7% OO. Femoral neck osteoporosis with low ASM predicted poor chair stand performance (beta -3.3, standard error 1.6, p = 0.04). SO scored lowest on the chair stand (p = 0.03) and step test (p = 0.03). Higher ASM predicted faster timed up and go performance (p = 0.001). Osteoporosis was prevalent in low ASM groups (SS and SO) and related to gait and balance deficits, particularly in the SO. This has implications for falls risk, fractures, and interventions.

[Homeostasis and Disorder of Musculoskeletal System.Molecular mechanism of bone metabolism and future therapeutic strategies.

PubMed

Nakashima, Tomoki

Recent studies of mouse genetics and human gene mutations has greatly contributed to clarifying the molecular mechanism of bone metabolism. Bone is constantly renewed by the balanced action of osteoblastic bone formation and osteoclastic bone resorption both of which mainly occur at the bone surface. This restructuring process called "bone remodeling" is important not only for normal bone mass and strength, but also for mineral homeostasis. Bone remodeling is stringently regulated by communication among bone component cells such as osteoclasts, osteoblasts, osteocytes and endothelial cells. An imbalance of this process is often linked to various bone diseases. Thus, the elucidation of the molecular mechanisms involved in bone remodeling is critical for a deeper understanding of the maintenance of healthy skeleton and bone disease.

Canonical Wnt signaling differently modulates osteogenic differentiation of mesenchymal stem cells derived from bone marrow and from periodontal ligament under inflammatory conditions.

PubMed

Liu, Wenjia; Konermann, Anna; Guo, Tao; Jäger, Andreas; Zhang, Liqiang; Jin, Yan

2014-03-01

Cellular plasticity and complex functional requirements of the periodontal ligament (PDL) assume a local stem cell (SC) niche to maintain tissue homeostasis and repair. Here, pathological alterations caused by inflammatory insults might impact the regenerative capacities of these cells. As bone homeostasis is fundamentally controlled by Wnt-mediated signals, it was the aim of this study to characterize the SC-like capacities of cells derived from PDL and to investigate their involvement in bone pathophysiology especially regarding the canonical Wnt pathway. PDLSCs were investigated for their SC characteristics via analysis of cell surface marker expression, colony forming unit efficiency, proliferation, osteogenic differentiation and adipogenic differentiation, and compared to bone marrow derived mesenchymal SCs (BMMSCs). To determine the impact of both inflammation and the canonical Wnt pathway on osteogenic differentiation, cells were challenged with TNF-α, maintained with or without Wnt3a or DKK-1 under osteogenic induction conditions and investigated for p-IκBα, p-NF-κB, p-Akt, β-catenin, p-GSK-3β, ALP and Runx2. PDLSCs exhibit weaker adipogenic and osteogenic differentiation capacities compared to BMMSCs. TNF-α inhibited osteogenic differentiation of PDLSCs more than BMMSCs mainly through regulating canonical Wnt pathway. Blocking the canonical Wnt pathway by DKK-1 reconstituted osteogenic differentiation of PDLSCs under inflammatory conditions, whereas activation by Wnt3a increased osteogenic differentiation of BMMSCs. Our results suggest a diverse regulation of the inhibitory effect of TNF-α in BMMSCs and PDLSCs via canonical Wnt pathway modulation. These findings provide novel insights on PDLSC SC-like capacities and their involvement in bone pathophysiology under the impact of the canonical Wnt pathway. Copyright © 2013 Elsevier B.V. All rights reserved.

Retinoic acid receptor signalling directly regulates osteoblast and adipocyte differentiation from mesenchymal progenitor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Green, A.C.; Department of Medicine at St. Vincent's Hospital, The University of Melbourne, Victoria 3065; Kocovski, P.

Low and high serum retinol levels are associated with increased fracture risk and poor bone health. We recently showed retinoic acid receptors (RARs) are negative regulators of osteoclastogenesis. Here we show RARs are also negative regulators of osteoblast and adipocyte differentiation. The pan-RAR agonist, all-trans retinoic acid (ATRA), directly inhibited differentiation and mineralisation of early osteoprogenitors and impaired the differentiation of more mature osteoblast populations. In contrast, the pan-RAR antagonist, IRX4310, accelerated differentiation of early osteoprogenitors. These effects predominantly occurred via RARγ and were further enhanced by an RARα agonist or antagonist, respectively. RAR agonists similarly impaired adipogenesis in osteogenicmore » cultures. RAR agonist treatment resulted in significant upregulation of the Wnt antagonist, Sfrp4. This accompanied reduced nuclear and cytosolic β-catenin protein and reduced expression of the Wnt target gene Axin2, suggesting impaired Wnt/β-catenin signalling. To determine the effect of RAR inhibition in post-natal mice, IRX4310 was administered to male mice for 10 days and bones were assessed by µCT. No change to trabecular bone volume was observed, however, radial bone growth was impaired. These studies show RARs directly influence osteoblast and adipocyte formation from mesenchymal cells, and inhibition of RAR signalling in vivo impairs radial bone growth in post-natal mice. - Graphical abstract: Schematic shows RAR ligand regulation of osteoblast differentiation in vitro. RARγ antagonists±RARα antagonists promote osteoblast differentiation. RARγ and RARα agonists alone or in combination block osteoblast differentiation, which correlates with upregulation of Sfrp4, and downregulation of nuclear and cytosolic β-catenin and reduced expression of the Wnt target gene Axin2. Red arrows indicate effects of RAR agonists on mediators of Wnt signalling.« less

Bone and fat connection in aging bone.

PubMed

Duque, Gustavo

2008-07-01

The fat and bone connection plays an important role in the pathophysiology of age-related bone loss. This review will focus on the age-induced mechanisms regulating the predominant differentiation of mesenchymal stem cells into adipocytes. Additionally, bone marrow fat will be considered as a diagnostic and therapeutic approach to osteoporosis. There are two types of bone and fat connection. The 'systemic connection', usually seen in obese patients, is hormonally regulated and associated with high bone mass and strength. The 'local connection' happens inside the bone marrow. Increasing amounts of bone marrow fat affect bone turnover through the inhibition of osteoblast function and survival and the promotion of osteoclast differentiation and activation. This interaction is regulated by paracrine secretion of fatty acids and adipokines. Additionally, bone marrow fat could be quantified using noninvasive methods and could be used as a therapeutic approach due to its capacity to transdifferentiate into bone without affecting other types of fat in the body. The bone and fat connection within the bone marrow constitutes a typical example of lipotoxicity. Additionally, bone marrow fat could be used as a new diagnostic and therapeutic approach for osteoporosis in older persons.

Balancing the Rates of New Bone Formation and Polymer Degradation Enhances Healing of Weight-Bearing Allograft/Polyurethane Composites in Rabbit Femoral Defects

PubMed Central

Dumas, Jerald E.; Prieto, Edna M.; Zienkiewicz, Katarzyna J.; Guda, Teja; Wenke, Joseph C.; Bible, Jesse; Holt, Ginger E.

2014-01-01

There is a compelling clinical need for bone grafts with initial bone-like mechanical properties that actively remodel for repair of weight-bearing bone defects, such as fractures of the tibial plateau and vertebrae. However, there is a paucity of studies investigating remodeling of weight-bearing bone grafts in preclinical models, and consequently there is limited understanding of the mechanisms by which these grafts remodel in vivo. In this study, we investigated the effects of the rates of new bone formation, matrix resorption, and polymer degradation on healing of settable weight-bearing polyurethane/allograft composites in a rabbit femoral condyle defect model. The grafts induced progressive healing in vivo, as evidenced by an increase in new bone formation, as well as a decrease in residual allograft and polymer from 6 to 12 weeks. However, the mismatch between the rates of autocatalytic polymer degradation and zero-order (independent of time) new bone formation resulted in incomplete healing in the interior of the composite. Augmentation of the grafts with recombinant human bone morphogenetic protein-2 not only increased the rate of new bone formation, but also altered the degradation mechanism of the polymer to approximate a zero-order process. The consequent matching of the rates of new bone formation and polymer degradation resulted in more extensive healing at later time points in all regions of the graft. These observations underscore the importance of balancing the rates of new bone formation and degradation to promote healing of settable weight-bearing bone grafts that maintain bone-like strength, while actively remodeling. PMID:23941405

PTH promotes allograft integration in a calvarial bone defect

PubMed Central

Sheyn, Dmitriy; Yakubovich, Doron Cohn; Kallai, Ilan; Su, Susan; Da, Xiaoyu; Pelled, Gadi; Tawackoli, Wafa; Cook-Weins, Galen; Schwarz, Edward M.; Gazit, Dan; Gazit, Zulma

2013-01-01

Allografts may be useful in craniofacial bone repair, although they often fail to integrate with the host bone. We hypothesized that intermittent administration of parathyroid hormone (PTH) would enhance mesenchymal stem cell recruitment and differentiation, resulting in allograft osseointegration in cranial membranous bones. Calvarial bone defects were created in transgenic mice, in which luciferase is expressed under the control of the osteocalcin promoter. The mice were given implants of allografts with or without daily PTH treatment. Bioluminescence imaging (BLI) was performed to monitor host osteprogenitor differentiation at the implantation site. Bone formation was evaluated with the aid of fluorescence imaging (FLI) and micro–computed tomography (μCT) as well as histological analyses. Reverse transcription polymerase chain reaction (RT-PCR) was performed to evaluate the expression of key osteogenic and angiogenic genes. Osteoprogenitor differentiation, as detected by BLI, in mice treated with an allograft implant and PTH was over 2-fold higher than those in mice treated with an allograft implant without PTH. FLI also demonstrated that the bone mineralization process in PTH-treated allografts was significantly higher than that in untreated allografts. The μCT scans revealed a significant increase in bone formation in Allograft + PTH–treated mice comparing to Allograft + PBS treated mice. The osteogenic genes osteocalcin (Oc/Bglap) and integrin binding sialoprotein (Ibsp) were upregulated in the Allograft + PTH–treated animals. In summary, PTH treatment enhances osteoprogenitor differentiation and augments bone formation around structural allografts. The precise mechanism is not clear, but we show that infiltration pattern of mast cells, associated with the formation of fibrotic tissue, in the defect site is significantly affected by the PTH treatment. PMID:24131143

Thiazide-sensitive Na+ -Cl- cotransporter (NCC) gene inactivation results in increased duodenal Ca2+ absorption, enhanced osteoblast differentiation and elevated bone mineral density.

PubMed

Hsu, Yu-Juei; Yang, Sung-Sen; Cheng, Chih-Jen; Liu, Shu-Ting; Huang, Shih-Ming; Chau, Tom; Chu, Pauling; Salter, Donald M; Lee, Herng-Sheng; Lin, Shih-Hua

2015-01-01

Inactivation of the thiazide-sensitive sodium chloride cotransporter (NCC) due to genetic mutations in Gitelman's syndrome (GS) or pharmacological inhibition with thiazide diuretics causes hypocalciuria and increased bone mineral density (BMD) with unclear extrarenal calcium (Ca(2+) ) regulation. We investigated intestinal Ca(2+) absorption and bone Ca(2+) metabolism in nonsense Ncc Ser707X (S707X) homozygous knockin mice (Ncc(S707X/S707X) mice). Compared to wild-type and heterozygous knockin littermates, Ncc(S707X/S707X) mice had increased intestinal absorption of (45) Ca(2+) and expression of the active Ca(2+) transport machinery (transient receptor potential vanilloid 6, calbindin-D9K , and plasma membrane Ca(2+) ATPase isoform 1b). Ncc(S707X/S707X) mice had also significantly increased Ca(2+) content accompanied by greater mineral apposition rate (MAR) in their femurs and higher trabecular bone volume, cortical bone thickness, and BMD determined by μCT. Their osteoblast differentiation markers, such as bone alkaline phosphatase, procollagen I, osteocalcin, and osterix, were also significantly increased while osteoclast activity was unaffected. Analysis of marrow-derived bone cells, either treated with thiazide or directly cultured from Ncc S707X knockin mice, showed that the differentiation of osteoblasts was associated with increased phosphorylation of mechanical stress-induced focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK). In conclusion, NCC inhibition stimulates duodenal Ca(2+) absorption as well as osteoblast differentiation and bone Ca(2+) storage, possibly through a FAK/ERK dependent mechanism. © 2014 American Society for Bone and Mineral Research.

Novel oxysterols have pro-osteogenic and anti-adipogenic effects in vitro and induce spinal fusion in vivo.

PubMed

Johnson, Jared S; Meliton, Vicente; Kim, Woo Kyun; Lee, Kwang-Bok; Wang, Jeffrey C; Nguyen, Khanhlinh; Yoo, Dongwon; Jung, Michael E; Atti, Elisa; Tetradis, Sotirios; Pereira, Renata C; Magyar, Clara; Nargizyan, Taya; Hahn, Theodore J; Farouz, Francine; Thies, Scott; Parhami, Farhad

2011-06-01

Stimulation of bone formation by osteoinductive materials is of great clinical importance in spinal fusion surgery, repair of bone fractures, and in the treatment of osteoporosis. We previously reported that specific naturally occurring oxysterols including 20(S)-hydroxycholesterol (20S) induce the osteogenic differentiation of pluripotent mesenchymal cells, while inhibiting their adipogenic differentiation. Here we report the characterization of two structural analogues of 20S, Oxy34 and Oxy49, which induce the osteogenic and inhibit the adipogenic differentiation of bone marrow stromal cells (MSC) through activation of Hedgehog (Hh) signaling. Treatment of M2-10B4 MSC with Oxy34 or Oxy49 induced the expression of osteogenic differentiation markers Runx2, Osterix (Osx), alkaline phosphatase (ALP), bone sialoprotein (BSP), and osteocalcin (OCN), as well as ALP enzymatic activity and robust mineralization. Treatment with oxysterols together with PPARγ activator, troglitazone (Tro), inhibited mRNA expression for adipogenic genes PPARγ, LPL, and aP2, and inhibited the formation of adipocytes. Efficacy of Oxy34 and Oxy49 in stimulating bone formation in vivo was assessed using the posterolateral intertransverse process rat spinal fusion model. Rats receiving collagen implants with Oxy 34 or Oxy49 showed comparable osteogenic efficacy to BMP2/collagen implants as measured by radiography, MicroCT, and manual inspection. Histological analysis showed trabecular and cortical bone formation by oxysterols and rhBMP2 within the fusion mass, with robust adipogenesis in BMP2-induced bone and significantly less adipocytes in oxysterol-induced bone. These data suggest that Oxy34 and Oxy49 are effective novel osteoinductive molecules and may be suitable candidates for further development and use in orthopedic indications requiring local bone formation. Copyright © 2011 Wiley-Liss, Inc.

Transgenic Expression of Osteoactivin/gpnmb Enhances Bone Formation In Vivo and Osteoprogenitor Differentiation Ex Vivo.

PubMed

Frara, Nagat; Abdelmagid, Samir M; Sondag, Gregory R; Moussa, Fouad M; Yingling, Vanessa R; Owen, Thomas A; Popoff, Steven N; Barbe, Mary F; Safadi, Fayez F

2016-01-01

Initial identification of osteoactivin (OA)/glycoprotein non-melanoma clone B (gpnmb) was demonstrated in an osteopetrotic rat model, where OA expression was increased threefold in mutant bones, compared to normal. OA mRNA and protein expression increase during active bone regeneration post-fracture, and primary rat osteoblasts show increased OA expression during differentiation in vitro. To further examine OA/gpnmb as an osteoinductive agent, we characterized the skeletal phenotype of transgenic mouse overexpressing OA/gpnmb under the CMV-promoter (OA-Tg). Western blot analysis showed increased OA/gpnmb in OA-Tg osteoblasts, compared to wild-type (WT). In OA-Tg mouse femurs versus WT littermates, micro-CT analysis showed increased trabecular bone volume and thickness, and cortical bone thickness; histomorphometry showed increased osteoblast numbers, bone formation and mineral apposition rates in OA-Tg mice; and biomechanical testing showed higher peak moment and stiffness. Given that OA/gpnmb is also over-expressed in osteoclasts in OA-Tg mice, we evaluated bone resorption by ELISA and histomorphometry, and observed decreased serum CTX-1 and RANK-L, and decreased osteoclast numbers in OA-Tg, compared to WT mice, indicating decreased bone remodeling in OA-Tg mice. The proliferation rate of OA-Tg osteoblasts in vitro was higher, compared to WT, as was alkaline phosphatase staining and activity, the latter indicating enhanced differentiation of OA-Tg osteoprogenitors. Quantitative RT-PCR analysis showed increased TGF-β1 and TGF-β receptors I and II expression in OA-Tg osteoblasts, compared to WT. Together, these data suggest that OA overexpression has an osteoinductive effect on bone mass in vivo and stimulates osteoprogenitor differentiation ex vivo. © 2015 Wiley Periodicals, Inc.

Zoledronic acid has differential antitumor activity in the pre- and postmenopausal bone microenvironment in vivo.

PubMed

Ottewell, Penelope D; Wang, Ning; Brown, Hannah K; Reeves, Kimberly J; Fowles, C Anne; Croucher, Peter I; Eaton, Colby L; Holen, Ingunn

2014-06-01

Clinical trials in early breast cancer have suggested that benefits of adjuvant bone-targeted treatments are restricted to women with established menopause. We developed models that mimic pre- and postmenopausal status to investigate effects of altered bone turnover on growth of disseminated breast tumor cells. Here, we report a differential antitumor effect of zoledronic acid (ZOL) in these two settings. Twleve-week-old female Balb/c-nude mice with disseminated MDA-MB-231 breast tumor cells in bone underwent sham operation or ovariectomy (OVX), mimicking the pre- and postmenopausal bone microenvironment, respectively. To determine the effects of bone-targeted therapy, sham/OVX animals received saline or 100 μg/kg ZOL weekly. Tumor growth was assessed by in vivo imaging and effects on bone by real-time PCR, micro-CT, histomorphometry, and measurements of bone markers. Disseminated tumor cells were detected by two-photon microscopy. OVX increased bone resorption and induced growth of disseminated tumor cells in bone. Tumors were detected in 83% of animals following OVX (postmenopausal model) compared with 17% following sham operation (premenopausal model). OVX had no effect on tumors outside of bone. OVX-induced tumor growth was completely prevented by ZOL, despite the presence of disseminated tumor cells. ZOL did not affect tumor growth in bone in the sham-operated animals. ZOL increased bone volume in both groups. This is the first demonstration that tumor growth is driven by osteoclast-mediated mechanisms in models that mimic post- but not premenopausal bone, providing a biologic rationale for the differential antitumor effects of ZOL reported in these settings. Clin Cancer Res; 20(11); 2922-32. ©2014 AACR. ©2014 American Association for Cancer Research.

Anti-osteoporotic activity of harpagide by regulation of bone formation in osteoblast cell culture and ovariectomy-induced bone loss mouse models.

PubMed

Chung, Hwa-Jin; Kyung Kim, Won; Joo Park, Hyen; Cho, Lan; Kim, Me-Riong; Kim, Min Jeong; Shin, Joon-Shik; Ho Lee, Jin; Ha, In-Hyuk; Kook Lee, Sang

2016-02-17

Harpagide, an iridoid glucoside, is a constituent of the root of Harpagophytum procumbens var. sublobatum (Engl.) Stapf, Devil's claw which has been used in patients with osteoarthritis (OA). In the present study, we investigated the anti-osteoporotic potential of harpagide and its underlying mechanism of action in in vitro cell culture and in vivo bone loss animal models. Harpagide was obtained from the alkalic hydrolysis of harpagoside, a major constituent of H. procumbens var. sublobatum Analysis of biomarkers for bone formation in osteoblastic MC3T3-E1 cells and bone resorption in osteoclast cells derived from mouse bone marrow cells was performed to evaluate the mechanism of action. The protective activity of harpagide against bone loss was also evaluated in ovariectomized (OVX) mouse model. Harpagide improved bone properties by stimulating the process of differentiation and maturation of osteoblast cells and suppressing the process of RANKL-induced differentiation of osteoclast cells. In OVX-induced bone loss mouse model, oral administration of harpagide significantly improved recovery of bone mineral density, trabecular bone volume, and trabecular number in the femur. Harpagide also prevented increase of trabecular separation and structure model index induced by OVX. Harpagide effectively inhibited the serum levels of biochemical markers of bone loss, including alkaline phosphatase, osteocalcin, C-terminal telopeptide, and tartrate-resistant acid phosphatase. Taken together, the present study demonstrates that harpagide has a potential for prevention of bone loss in OVX mice by regulating the stimulation of osteoblast differentiation and the suppression of osteoclast formation. Therefore, these findings suggest that harpagide might serve as a bioactive compound derived from H. procumbens var. sublobatum for improvement of age-dependent bone destruction disease. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Strong and rapid induction of osteoblast differentiation by Cbfa1/Til-1 overexpression for bone regeneration.

PubMed

Kojima, Hiroko; Uemura, Toshimasa

2005-01-28

Core binding factor alpha-1 (Cbfa1), known as an essential transcription factor for osteogenic lineage, has two major N-terminal isoforms: Pebp2alphaA and Til-1. To study the roles of these isoforms in bone regeneration, we applied an adenoviral vector carrying their genes to transduce primary osteoprogenitor cells in vitro and in vivo. Overexpression of the two isoforms induced rapid and marked osteoblast differentiation, with Til-1 being more effective in vitro, by examination of the alkaline phosphatase activity, calcium content, and Alizarin red staining. Til-1 overexpressing cells/porous ceramic composites were transplanted into subcutaneous and bone defect sites in Fischer rats (cultured bone transplantation model) and markedly affected in vivo bone formation and osteoblast markers. The results demonstrated that the reconstitution of bone tissues, such as cortical bone and trabecular bone was accelerated by implantation of Til-1 overexpressing cells/porous ceramic composites. Moreover, the new bone formation by Til-1 overexpression appeared to reflect replacement of new bone within the implant boundaries. To ascertain whether implanted Cbfa1 overexpressing cells could differentiate into osteogenic cells to create bone or whether it stimulated the surrounding recipient tissue to regenerate bone, implanted male donor cells were visualized by fluorescent in situ hybridization analysis. The proportion of implanted cells in the presumptive bone forming region was over 80% and did not change throughout from 3 days to 8 weeks after implantation. These findings suggested that the newly formed bone in the porous area of the scaffold is mostly produced by the implanted donor cells or their derived cells, effectively by Til-1 overexpression.

Enhanced osteoporotic bone regeneration by strontium-substituted calcium silicate bioactive ceramics.

PubMed

Lin, Kaili; Xia, Lunguo; Li, Haiyan; Jiang, Xinquan; Pan, Haobo; Xu, Yuanjin; Lu, William W; Zhang, Zhiyuan; Chang, Jiang

2013-12-01

The regeneration capacity of the osteoporotic bones is generally lower than that of the normal bones. Current methods of bone defect treatment for osteoporosis are not always satisfactory. Recent studies have shown that the silicate based biomaterials can stimulate osteogenesis and angiogenesis due to the silicon (Si) ions released from the materials, and enhance bone regeneration in vivo. Other studies showed that strontium (Sr) plays a distinct role on inhibiting bone resorption. Based on the hypothesis that the combination of Si and Sr may have synergetic effects on osteoporotic bone regeneration, the porous Sr-substituted calcium silicate (SrCS) ceramic scaffolds combining the functions of Sr and Si elements were developed with the goals to promote osteoporotic bone defect repair. The effects of the ionic extract from SrCS on osteogenic differentiation of bone marrow mesenchymal stem cells derived from ovariectomized rats (rBMSCs-OVX), angiogenic differentiation of human umbilical vein endothelial cells (HUVECs) were investigated. The in vitro results showed that Sr and Si ions released from SrCS enhanced cell viability, alkaline phosphatase (ALP) activity, and mRNA expression levels of osteoblast-related genes of rBMSCs-OVX and expression of vascular endothelial growth factor (VEGF) without addition of extra osteogenic and angiogenic reagents. The activation in extracellular signal-related kinases (ERK) and p38 signaling pathways were observed in rBMSCs-OVX cultured in the extract of SrCS, and these effects could be blocked by ERK inhibitor PD98059, and P38 inhibitor SB203580, respectively. Furthermore, the ionic extract of SrCS stimulated HUVECs proliferation, differentiation and angiogenesis process. The in vivo experiments revealed that SrCS dramatically stimulated bone regeneration and angiogenesis in a critical sized OVX calvarial defect model, and the enhanced bone regeneration might be attributed to the modulation of osteogenic differentiation of endogenous mesenchymal stem cells (MSCs) and the inhibition of osteoclastogenesis, accompanying with the promotion of the angiogenic activity of endothelial cells (ECs). Copyright © 2013 Elsevier Ltd. All rights reserved.

Alendronate promotes osteoblast differentiation and bone formation in ovariectomy-induced osteoporosis through interferon-β/signal transducer and activator of transcription 1 pathway

PubMed Central

Ma, Xiaoqing; Xu, Zhongyang; Ding, Shaofeng; Yi, Guangkun; Wang, Qian

2018-01-01

Alendronate is commonly used for the treatment of postmenopausal osteoporosis; however, the underlying pathological molecular mechanisms of its action remain unclear. In the present study, the alendronate-treated signaling pathway in bone metabolism in rats with ovariectomy induced by osteoporosis was investigated. Rats with osteoporosis were orally administered alendronate or phosphate-buffered saline (control). In addition, the interferon-β (IFN-β)/signal transducer and activator of transcription 1 (STAT1) signaling pathway was investigated in osteoblasts following treatment with alendronate in vitro and in vivo. During the differentiation period, IFN-β (100 ng/ml) was used to treat the osteoblast cells, and the activity, viability and bone metabolism-associated gene expression levels (STAT1, p-STAT1, Fra1, TRAF6 and SOCS1) were analyzed in osteoblast cells. Histopathological changes were used to evaluate osteoblasts, osteoclasts, inflammatory phase of bone healing and osteonecrotic areas. The results demonstrated that alendronate significantly inhibited the activity of osteoporotic osteoclasts by stimulating expression of IFN-β, as well as markedly improved the viability and activity of osteoblasts compared with the control group. In addition, alendronate increased the expression and phosphorylation levels of STAT1 in osteoclasts, enhanced osteoblast differentiation, upregulated the expression levels of alkaline phosphatase and osteocalcin, and increased the expression of osteoblast differentiation-associated genes (osteocalcin, osterix and Runx2). Inhibition of IFN-β expression canceled the benefits of alendronate-mediated osteoblast differentiation. Notably, alendronate enhanced bone formation in rats with osteoporosis induced by ovariectomy. In conclusion, these findings suggest that alendronate can regulate osteoblast differentiation and bone formation in rats with osteoporosis induced by ovariectomy through upregulation of IFN-β/STAT1 signaling pathway. PMID:29375681

Fucoidan-induced osteogenic differentiation promotes angiogenesis by inducing vascular endothelial growth factor secretion and accelerates bone repair.

PubMed

Kim, Beom-Su; Yang, Sun-Sik; You, Hyung-Keun; Shin, Hong-In; Lee, Jun

2018-03-01

Osteogenesis and angiogenesis, including cell-cell communication between blood vessel cells and bone cells, are essential for bone repair. Fucoidan is a chemical compound that has a variety of biological activities. It stimulates osteoblast differentiation in human mesenchymal stem cells (MSCs), which in turn induces angiogenesis. However, the mechanism by which this communication between osteoblasts and endothelial cells is mediated remains unclear. Thus, the aim of this study was to clarify the relationship between fucoidan-induced osteoblastic differentiation in MSCs and angiogenesis in endothelial cells. First, the effect was confirmed of fucoidan on osteoblast differentiation in MSCs and obtained conditioned media from these cells (Fucoidan-MSC-CM). Next, the angiogenic activity of Fucoidan-MSC-CM was investigated and it was found that it stimulated angiogenesis, demonstrated by proliferation, tube formation, migration and sprout capillary formation in human umbilical vein endothelial cells. Messenger ribonucleic acid expression and protein secretion of vascular endothelial growth factor (VEGF) were dramatically increased during fucoidan-induced osteoblast differentiation and that its angiogenic activities were reduced by a VEGF/VEGF receptor-specific binding inhibitor. Furthermore, Fucoidan-MSC-CM increased the phosphorylation of mitogen-activated protein kinase and PI3K/AKT/eNOS signalling pathway, and that its angiogenic effects were markedly suppressed by SB203580 and AKT 1/2 inhibitor. Finally, an in vivo study was conducted and it was found that fucoidan accelerated new blood vessel formation and partially promoted bone formation in a rabbit model of a calvarial bone defect. This is the first study to investigate the angiogenic effect of fucoidan-induced osteoblastic differentiation through VEGF secretion, suggesting the therapeutic potential of fucoidan for enhancing bone repair. Copyright © 2017 John Wiley & Sons, Ltd.

Validation of a simple and fast method to quantify in vitro mineralization with fluorescent probes used in molecular imaging of bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Moester, Martiene J.C.; Schoeman, Monique A.E.; Oudshoorn, Ineke B.

2014-01-03

Highlights: •We validate a simple and fast method of quantification of in vitro mineralization. •Fluorescently labeled agents can detect calcium deposits in the mineralized matrix of cell cultures. •Fluorescent signals of the probes correlated with Alizarin Red S staining. -- Abstract: Alizarin Red S staining is the standard method to indicate and quantify matrix mineralization during differentiation of osteoblast cultures. KS483 cells are multipotent mouse mesenchymal progenitor cells that can differentiate into chondrocytes, adipocytes and osteoblasts and are a well-characterized model for the study of bone formation. Matrix mineralization is the last step of differentiation of bone cells and ismore » therefore a very important outcome measure in bone research. Fluorescently labelled calcium chelating agents, e.g. BoneTag and OsteoSense, are currently used for in vivo imaging of bone. The aim of the present study was to validate these probes for fast and simple detection and quantification of in vitro matrix mineralization by KS483 cells and thus enabling high-throughput screening experiments. KS483 cells were cultured under osteogenic conditions in the presence of compounds that either stimulate or inhibit osteoblast differentiation and thereby matrix mineralization. After 21 days of differentiation, fluorescence of stained cultures was quantified with a near-infrared imager and compared to Alizarin Red S quantification. Fluorescence of both probes closely correlated to Alizarin Red S staining in both inhibiting and stimulating conditions. In addition, both compounds displayed specificity for mineralized nodules. We therefore conclude that this method of quantification of bone mineralization using fluorescent compounds is a good alternative for the Alizarin Red S staining.« less

Myostatin inhibits osteoblastic differentiation by suppressing osteocyte-derived exosomal microRNA-218: A novel mechanism in muscle-bone communication.

PubMed

Qin, Yiwen; Peng, Yuanzhen; Zhao, Wei; Pan, Jianping; Ksiezak-Reding, Hanna; Cardozo, Christopher; Wu, Yingjie; Divieti Pajevic, Paola; Bonewald, Lynda F; Bauman, William A; Qin, Weiping

2017-06-30

Muscle and bone are closely associated in both anatomy and function, but the mechanisms that coordinate their synergistic action remain poorly defined. Myostatin, a myokine secreted by muscles, has been shown to inhibit muscle growth, and the disruption of the myostatin gene has been reported to cause muscle hypertrophy and increase bone mass. Extracellular vesicle-exosomes that carry microRNA (miRNA), mRNA, and proteins are known to perform an important role in cell-cell communication. We hypothesized that myostatin may play a crucial role in muscle-bone interactions and may promote direct effects on osteocytes and on osteocyte-derived exosomal miRNAs, thereby indirectly influencing the function of other bone cells. We report herein that myostatin promotes expression of several bone regulators such as sclerostin (SOST), DKK1, and RANKL in cultured osteocytic (Ocy454) cells, concomitant with the suppression of miR-218 in both parent Ocy454 cells and derived exosomes. Exosomes produced by Ocy454 cells that had been pretreated with myostatin could be taken up by osteoblastic MC3T3 cells, resulting in a marked reduction of Runx2, a key regulator of osteoblastic differentiation, and in decreased osteoblastic differentiation via the down-regulation of the Wnt signaling pathway. Importantly, the inhibitory effect of myostatin-modified osteocytic exosomes on osteoblast differentiation is completely reversed by expression of exogenous miR-218, through a mechanism involving miR-218-mediated inhibition of SOST. Together, our findings indicate that myostatin directly influences osteocyte function and thereby inhibits osteoblastic differentiation, at least in part, through the suppression of osteocyte-derived exosomal miR-218, suggesting a novel mechanism in muscle-bone communication. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Synergistic effect of scaffold composition and dynamic culturing environment in multilayered systems for bone tissue engineering.

PubMed

Rodrigues, Márcia T; Martins, Albino; Dias, Isabel R; Viegas, Carlos A; Neves, Nuno M; Gomes, Manuela E; Reis, Rui L

2012-11-01

Bone extracellular matrix (ECM) is composed of mineralized collagen fibrils which support biological apatite nucleation that participates in bone outstanding properties. Understanding and mimicking bone morphological and physiological parameters at a biological scale is a major challenge in tissue engineering scaffolding. Using emergent (nano)technologies scaffold designing may be critically improved, enabling highly functional tissue substitutes for bone applications. This study aims to develop novel biodegradable composite scaffolds of tricalcium phosphate (TCPs) and electrospun nanofibers of poly(ϵ-caprolactone) (PCL), combining TCPs osteoconductivity with PCL biocompatibility and elasticity, mimicking bone structure and composition. We hypothesized that scaffolds with such structure/composition would stimulate the proliferation and differentiation of bone marrow stromal cells (BMSCs) towards the osteogenic phenotype. Composite scaffolds, developed by electrospining using consecutive stacked layers of PCL and TCPs, were characterized by FTIR spectroscopy, X-Ray diffraction and scanning electronic microscopy. Cellular behavior was assessed in goat BMSCs seeded onto composite scaffolds and cultured in static or dynamic conditions, using basal or osteogenic media during 7, 14 or 21 days. Cellular proliferation was quantified and osteogenic differentiation confirmed by alkaline phosphatase activity, alizarin red staining and immunocytochemistry for osteocalcin and collagen I. Results suggest that PCL-TCP scaffolds provide a 3D support for gBMSCs proliferation and osteogenic differentiation with production of ECM. TCPs positively stimulate the osteogenic process, especially under dynamic conditions, where PCL-TCP scaffolds are sufficient to promote osteogenic differentiation even in basal medium conditions. The enhancement of the osteogenic potential in dynamic conditions evidences the synergistic effect of scaffold composition and dynamic stimulation in gBMSCs osteogenic differentiation. Copyright © 2012 John Wiley & Sons, Ltd.

Silk fibroin/chitosan thin film promotes osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

PubMed

Li, Da-Wei; He, Jin; He, Feng-Li; Liu, Ya-Li; Liu, Yang-Yang; Ye, Ya-Jing; Deng, Xudong; Yin, Da-Chuan

2018-04-01

As a biodegradable polymer thin film, silk fibroin/chitosan composite film overcomes the defects of pure silk fibroin and chitosan films, respectively, and shows remarkable biocompatibility, appropriate hydrophilicity and mechanical properties. Silk fibroin/chitosan thin film can be used not only as metal implant coating for bone injury repair, but also as tissue engineering scaffold for skin, cornea, adipose, and other soft tissue injury repair. However, the biocompatibility of silk fibroin/chitosan thin film for mesenchymal stem cells, a kind of important seed cell of tissue engineering and regenerative medicine, is rarely reported. In this study, silk fibroin/chitosan film was prepared by solvent casting method, and the rat bone marrow-derived mesenchymal stem cells were cultured on the silk fibroin/chitosan thin film. Osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells were induced, respectively. The proliferation ability, osteogenic and adipogenic differentiation abilities of rat bone marrow-derived mesenchymal stem cells were systematically compared between silk fibroin/chitosan thin film and polystyrene tissue culture plates. The results showed that silk fibroin/chitosan thin film not only provided a comparable environment for the growth and proliferation of rat bone marrow-derived mesenchymal stem cells but also promoted their osteogenic and adipogenic differentiation. This work provided information of rat bone marrow-derived mesenchymal stem cells behavior on silk fibroin/chitosan thin film and extended the application of silk fibroin/chitosan thin film. Based on the results, we suggested that the silk fibroin/chitosan thin film could be a promising material for tissue engineering of bone, cartilage, adipose, and skin.

A novel strontium(II)-modified calcium phosphate bone cement stimulates human-bone-marrow-derived mesenchymal stem cell proliferation and osteogenic differentiation in vitro.

PubMed

Schumacher, M; Lode, A; Helth, A; Gelinsky, M

2013-12-01

In the present study, the in vitro effects of novel strontium-modified calcium phosphate bone cements (SrCPCs), prepared using two different approaches on human-bone-marrow-derived mesenchymal stem cells (hMSCs), were evaluated. Strontium ions, known to stimulate bone formation and therefore already used in systemic osteoporosis therapy, were incorporated into a hydroxyapatite-forming calcium phosphate bone cement via two simple approaches: incorporation of strontium carbonate crystals and substitution of Ca(2+) by Sr(2+) ions during cement setting. All modified cements released 0.03-0.07 mM Sr(2+) under in vitro conditions, concentrations that were shown not to impair the proliferation or osteogenic differentiation of hMSCs. Furthermore, strontium modification led to a reduced medium acidification and Ca(2+) depletion in comparison to the standard calcium phosphate cement. In indirect and direct cell culture experiments with the novel SrCPCs significantly enhanced cell proliferation and differentiation were observed. In conclusion, the SrCPCs described here could be beneficial for the local treatment of defects, especially in the osteoporotic bone. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Physalis angulata induces in vitro differentiation of murine bone marrow cells into macrophages.

PubMed

da Silva, Bruno José Martins; Rodrigues, Ana Paula D; Farias, Luis Henrique S; Hage, Amanda Anastácia P; Do Nascimento, Jose Luiz M; Silva, Edilene O

2014-10-03

The bone marrow is a hematopoietic tissue that, in the presence of cytokines and growth factors, generates all of the circulating blood cells. These cells are important for protecting the organism against pathogens and for establishing an effective immune response. Previous studies have shown immunomodulatory effects of different products isolated from plant extracts. This study aimed to evaluate the immunomodulatory properties of aqueous Physalis angulata (AEPa) extract on the differentiation of bone marrow cells. Increased cellular area, higher spreading ability and several cytoplasmatic projections were observed in the treated cells, using optical microscopy, suggesting cell differentiation. Furthermore, AEPa did not promote the proliferation of lymphocytes and polymorphonuclear leukocytes, however promotes increased the number of macrophages in the culture. The ultrastructural analysis by Transmission Electron Microscopy of treated cells showed spreading ability, high number of cytoplasmatic projections and increase of autophagic vacuoles. Moreover, a high level of LC3b expression by treated cells was detected by flow cytometry, suggesting an autophagic process. Cell surface expression of F4/80 and CD11b also indicated that AEPa may stimulate differentiation of bone marrow cells mainly into macrophages. In addition, AEPa did not differentiate cells into dendritic cells, as assessed by CD11c analysis. Furthermore, no cytotoxic effects were observed in the cells treated with AEPa. Results demonstrate that AEPa promotes the differentiation of bone marrow cells, particularly into macrophages and may hold promise as an immunomodulating agent.

Physalis angulata induces in vitro differentiation of murine bone marrow cells into macrophages

PubMed Central

2014-01-01

Background The bone marrow is a hematopoietic tissue that, in the presence of cytokines and growth factors, generates all of the circulating blood cells. These cells are important for protecting the organism against pathogens and for establishing an effective immune response. Previous studies have shown immunomodulatory effects of different products isolated from plant extracts. This study aimed to evaluate the immunomodulatory properties of aqueous Physalis angulata (AEPa) extract on the differentiation of bone marrow cells. Results Increased cellular area, higher spreading ability and several cytoplasmatic projections were observed in the treated cells, using optical microscopy, suggesting cell differentiation. Furthermore, AEPa did not promote the proliferation of lymphocytes and polymorphonuclear leukocytes, however promotes increased the number of macrophages in the culture. The ultrastructural analysis by Transmission Electron Microscopy of treated cells showed spreading ability, high number of cytoplasmatic projections and increase of autophagic vacuoles. Moreover, a high level of LC3b expression by treated cells was detected by flow cytometry, suggesting an autophagic process. Cell surface expression of F4/80 and CD11b also indicated that AEPa may stimulate differentiation of bone marrow cells mainly into macrophages. In addition, AEPa did not differentiate cells into dendritic cells, as assessed by CD11c analysis. Furthermore, no cytotoxic effects were observed in the cells treated with AEPa. Conclusion Results demonstrate that AEPa promotes the differentiation of bone marrow cells, particularly into macrophages and may hold promise as an immunomodulating agent. PMID:25281406

Cellular Mechanisms of Multiple Myeloma Bone Disease

PubMed Central

Oranger, Angela; Carbone, Claudia; Izzo, Maddalena; Grano, Maria

2013-01-01

Multiple myeloma (MM) is a hematologic malignancy of differentiated plasma cells that accumulates and proliferates in the bone marrow. MM patients often develop bone disease that results in severe bone pain, osteolytic lesions, and pathologic fractures. These skeletal complications have not only a negative impact on quality of life but also a possible effect in overall survival. MM osteolytic bone lesions arise from the altered bone remodeling due to both increased osteoclast activation and decreased osteoblast differentiation. A dysregulated production of numerous cytokines that can contribute to the uncoupling of bone cell activity is well documented in the bone marrow microenvironment of MM patients. These molecules are produced not only by malignant plasma cells, that directly contribute to MM bone disease, but also by bone, immune, and stromal cells interacting with each other in the bone microenvironment. This review focuses on the current knowledge of MM bone disease biology, with particular regard on the role of bone and immune cells in producing cytokines critical for malignant plasma cell proliferation as well as in osteolysis development. Therefore, the understanding of MM pathogenesis could be useful to the discovery of novel agents that will be able to both restore bone remodelling and reduce tumor burden. PMID:23818912

Understanding leptin-dependent regulation of skeletal homeostasis

PubMed Central

Motyl, Katherine J.; Rosen, Clifford J.

2012-01-01

Despite growing evidence for adipose tissue regulation of bone mass, the role of the adipokine leptin in bone remodeling remains controversial. The majority of in vitro studies suggest leptin enhances osteoblastic proliferation and differentiation while inhibiting adipogenic differentiation from marrow stromal cells. Alternatively, some evidence demonstrates either no effect or a pro-apoptotic action of leptin on stromal cells. Similarly, in vivo work has demonstrated both positive and negative effects of leptin on bone mass. Most of the literature supports the idea that leptin suppresses bone mass by acting in the brainstem to reduce serotonin-dependent sympathetic signaling from the ventromedial hypothalamus to bone. However, other studies have found partly or entirely contrasting actions of leptin. Recently one study found a significant effect of surgery alone with intracerebroventricular administration of leptin, a technique crucial for understanding centrally-mediated leptin regulation of bone. Thus, two mainstream hypotheses for the role of leptin on bone emerge: 1) direct regulation through increased osteoblast proliferation and differentiation and 2) indirect suppression of bone formation through a hypothalamic relay. At the present time, it remains unclear whether these effects are relevant in only extreme circumstances (i.e. models with complete deficiency) or play an important homeostatic role in the regulation of peak bone acquisition and skeletal remodeling. Ultimately, determining the actions of leptin on the skeleton will be critical for understanding how the obesity epidemic may be impacting the prevalence of osteoporosis. PMID:22534195

Bone Morphogenetic Protein-2 Promotes Human Mesenchymal Stem Cell Survival and Resultant Bone Formation When Entrapped in Photocrosslinked Alginate Hydrogels.

PubMed

Ho, Steve S; Vollmer, Nina L; Refaat, Motasem I; Jeon, Oju; Alsberg, Eben; Lee, Mark A; Leach, J Kent

2016-10-01

There is a substantial need to prolong cell persistence and enhance functionality in situ to enhance cell-based tissue repair. Bone morphogenetic protein-2 (BMP-2) is often used at high concentrations for osteogenic differentiation of mesenchymal stem cells (MSCs) but can induce apoptosis. Biomaterials facilitate the delivery of lower doses of BMP-2, reducing side effects and localizing materials at target sites. Photocrosslinked alginate hydrogels (PAHs) can deliver osteogenic materials to irregular-sized bone defects, providing improved control over material degradation compared to ionically cross-linked hydrogels. It is hypothesized that the delivery of MSCs and BMP-2 from a PAH increases cell persistence by reducing apoptosis, while promoting osteogenic differentiation and enhancing bone formation compared to MSCs in PAHs without BMP-2. BMP-2 significantly decreases apoptosis and enhances survival of photoencapsulated MSCs, while simultaneously promoting osteogenic differentiation in vitro. Bioluminescence imaging reveals increased MSC survival when implanted in BMP-2 PAHs. Bone defects treated with MSCs in BMP-2 PAHs demonstrate 100% union as early as 8 weeks and significantly higher bone volumes at 12 weeks, while defects with MSC-entrapped PAHs alone do not fully bridge. This study demonstrates that transplantation of MSCs with BMP-2 in PAHs achieves robust bone healing, providing a promising platform for bone repair. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

ATF4, A Novel Mediator of the Anabolic Actions of PTH on Bone

DTIC Science & Technology

2010-07-01

increased the expression of genes known to be associated with osteoblast differentiation including osteocalcin (Ocn), bone sialoprotein (Bsp...differentiation including osteocalcin (Ocn) (2.2-fold), bone sialoprotein (Bsp) (4.2-fold), alkaline phosphatase (Alp) (3.2-fold), a1(I) collagen (Col1... sialoprotein (BSP) gene transcription is mediated through a pituitary-specific transcription factor-1 (Pit-1) motif in the rat BSP gene promoter. Matrix Biol

ATF4, A Novel Mediator of the Anabolic Actions of PTH on Bone

DTIC Science & Technology

2011-07-01

Accumulating evidence estab- lishes that ubiquitin-proteasome pathways control osteoblast differentiation and bone forma - tion. For example, the proteasome...tide ligase, accelerated Runx2 ubiquitin-proteasomal degrada- tion and inhibited osteoblast differentiation and bone forma - FIGURE 5. TFIIA activates...36. Zhao, C., Irie , N., Takada, Y., Shimoda, K., Miyamoto, T., Nishiwaki, T., Suda, T., and Matsuo, K. (2006) Cell Metab. 4, 111–121 37. Geoffroy, V

ATF4, A Novel Mediator of the Anabolic Actions of PTH on Bone

DTIC Science & Technology

2008-07-01

lishes that ubiquitin-proteasome pathways control osteoblast differentiation and bone forma - tion. For example, the proteasome inhibitors epoxomicin...Runx2 ubiquitin-proteasomal degrada- tion and inhibited osteoblast differentiation and bone forma - FIGURE 5. TFIIA activates endogenous Ocn gene...2565 35. Duvall, J. F., Kashanchi, F., Cvekl, A., Radonovich, M. F., Piras, G., and Brady, J. N. (1995) J. Virol. 69, 5077–5086 36. Zhao, C., Irie , N

Soft matrix supports osteogenic differentiation of human dental follicle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Viale-Bouroncle, Sandra; Voellner, Florian; Moehl, Christoph

Highlights: {yields} Rigid stiffness supports osteogenic differentiation in mesenchymal stem cells (MSCs). {yields} Our study examined stiffness and differentiation of dental follicle cells (DFCs). {yields} Soft ECMs have a superior capacity to support the osteogenic differentiation of DFCs. {yields} DFCs and MSCs react contrarily to soft and rigid surface stiffness. -- Abstract: The differentiation of stem cells can be directed by the grade of stiffness of the developed tissue cells. For example a rigid extracellular matrix supports the osteogenic differentiation in bone marrow derived mesenchymal stem cells (MSCs). However, less is known about the relation of extracellular matrix stiffness andmore » cell differentiation of ectomesenchymal dental precursor cells. Our study examined for the first time the influence of the surface stiffness on the proliferation and osteogenic differentiation of human dental follicle cells (DFCs). Cell proliferation of DFCs was only slightly decreased on cell culture surfaces with a bone-like stiffness. The osteogenic differentiation in DFCs could only be initiated with a dexamethasone based differentiation medium after using varying stiffness. Here, the softest surface improved the induction of osteogenic differentiation in comparison to that with the highest stiffness. In conclusion, different to bone marrow derived MSCs, soft ECMs have a superior capacity to support the osteogenic differentiation of DFCs.« less

Impact of ionizing radiation exposure on in vitro differentiation of preosteoblastic cell lines

NASA Astrophysics Data System (ADS)

Hu, Yueyuan; Lau, Patrick; Hellweg, Christine; Baumstark-Khan, Christa; Reitz, Guenther

Bone demineralization of astronauts during residence in microgravity is a well known phe-nomenon during space travel. Besides altered gravity conditions, radiation risk is considered to be one of the major health hazards for astronauts in both orbital and interplanetary space. Un-til know, little is known about the effects of space radiation on the skeletal system especially on the bone forming osteoblasts. Accelerator facilities are used to simulate parts of the radiation environment in space. We examined the effects of heavy ion exposure on osteoblastic differ-entiation of murine preosteoblastic cell lines to gain insight into potential cellular mechanisms involved in bone cellular response after exposure to heavy ions. Therefore, we examined gene expression modulation of bone specific transcription factors, osteoblast specific marker genes as well as genes function as coupling factors that link bone resorption to bone formation. mRNA levels were determined using quantitative real time reverse transcriptase PCR (qRT-PCR). Expression of a target gene was standardized to unregulated reference genes. We investigated the transcriptional regulation of Osteocalcin (OCN) as well as TGF-β1, p21(CDKN1A) and the bone specific transcription factor Runx2 (cbfa1). We investigated gene expression modula-tions after exposure to energetic carbon ions (35 MeV/u, 73 keV/µm), iron ions (1000 MeV/u, 150 keV/µm) and lead ions (29 MeV/u, 9600 keV/µm) versus low LET X-rays. X-irradiation dose-dependently increased the mRNA levels of p21(CDKN1A) and Runx2 (cbfa1) whereas expression of OCN and TGF-β1 were elevated at later time points. Exposure to heavy ions provoked a more pronounced effect on osteoblastic specific gene expression within the dif-ferentiation process. Collectively, our results indicate that heavy ions facilitate osteoblastic differentiation more effectively than X-ray. Using the proposed in vitro model we confirmed that exposure to ionizing radiation significantly modulates gene expression levels of marker genes involved in the differentiation of osteoblasts. The data presented allow us to suggest that exposure to ionizing radiation interferes with bone formation at the level of cell differentiation.

Differential osteogenic activity of osteoprogenitor cells on HA and TCP/HA scaffold of tissue engineered bone.

PubMed

Ng, Angela M H; Tan, K K; Phang, M Y; Aziyati, O; Tan, G H; Isa, M R; Aminuddin, B S; Naseem, M; Fauziah, O; Ruszymah, B H I

2008-05-01

Biomaterial, an essential component of tissue engineering, serves as a scaffold for cell attachment, proliferation, and differentiation; provides the three dimensional (3D) structure and, in some applications, the mechanical strength required for the engineered tissue. Both synthetic and naturally occurring calcium phosphate based biomaterial have been used as bone fillers or bone extenders in orthopedic and reconstructive surgeries. This study aims to evaluate two popular calcium phosphate based biomaterial i.e., hydroxyapatite (HA) and tricalcium phosphate/hydroxyapatite (TCP/HA) granules as scaffold materials in bone tissue engineering. In our strategy for constructing tissue engineered bone, human osteoprogenitor cells derived from periosteum were incorporated with human plasma-derived fibrin and seeded onto HA or TCP/HA forming 3D tissue constructs and further maintained in osteogenic medium for 4 weeks to induce osteogenic differentiation. Constructs were subsequently implanted intramuscularly in nude mice for 8 weeks after which mice were euthanized and constructs harvested for evaluation. The differential cell response to the biomaterial (HA or TCP/HA) adopted as scaffold was illustrated by the histology of undecalcified constructs and evaluation using SEM and TEM. Both HA and TCP/HA constructs showed evidence of cell proliferation, calcium deposition, and collagen bundle formation albeit lesser in the former. Our findings demonstrated that TCP/HA is superior between the two in early bone formation and hence is the scaffold material of choice in bone tissue engineering. Copyright 2007 Wiley Periodicals, Inc.

Impairment of osteoclastic bone resorption in rapidly growing female p47phox knockout mice

USDA-ARS?s Scientific Manuscript database

Bone formation is dependent on the activity and differentiation of osteoblasts; whereas resorption of preexisting mineralized bone matrix by osteoclasts is necessary not only for bone development but also for regeneration and remodeling. Bone remodeling is a process in which osteoblasts and osteocla...

Bone Formation Rate in Experimental Disuse Osteoporosis in Monkeys

NASA Technical Reports Server (NTRS)

Cann, Christopher; Young, Donald R.

1976-01-01

Specific mechanisms underlying weightless and hypodynamic bone loss are obscure. A principal relationship which must be affected is the balance between bone formation and bone resorption rates. In order to better define the influence of those parameters on bone loss, and also to develop measurements in other species as a useful adjunct to human research, studies were undertaken with experimental monkeys. Tests were conducted with a total of 6 adult male monkeys, weighing 10-13 kg, and approximately 10-12 yrs. of age to evaluate specifically bone formation rate during the development of disuse osteoporosis and osteopenia. Three animals were restrained in a semi-recumbent position for six months; three animals served as normal caged controls. Food intake (Purina) was held relatively constant at 200g/day for each animal. Using a Norland Bone Mineral Analyzer, bone mineral losses of 3.5 to 6% were seen in the mid-shaft of the tibia and in the distal radius. Bone loss was confirmed radiographically, with observation of thinning of the proximal tibial cortex and trabeculae in the calcaneus. Bone formation rate was determined using standard Ca-47 kinetics under metabolic balance conditions. After six months of restraint, accretion was 7.2-13.2 mg Ca/kg/day, compared to 3.2-4.1 mg Ca/kg/day in caged controls and 3-8 mg Ca/kg/day in normal adult humans. Fecal and urine calcium was 25-40% higher in restrained animals than in controls. Dietary calcium absorption decreases during restraint, and calcium turnover increases, implying a rise in bone resorption rate concommitant with the observed rise in bone accretion rate. Further studies dealing specifically with bone resorption are underway to define this more fully.

Screening of Osteogenic-Enhancing Short Peptides from BMPs for Biomimetic Material Applications

PubMed Central

Kanie, Kei; Kurimoto, Rio; Tian, Jing; Ebisawa, Katsumi; Narita, Yuji; Honda, Hiroyuki; Kato, Ryuji

2016-01-01

Bone regeneration is an important issue in many situations, such as bone fracture and surgery. Umbilical cord mesenchymal stem cells (UC-MSCs) are promising cell sources for bone regeneration. Bone morphogenetic proteins and their bioactive peptides are biomolecules known to enhance the osteogenic differentiation of MSCs. However, fibrosis can arise during the development of implantable biomaterials. Therefore, it is important to control cell organization by enhancing osteogenic proliferation and differentiation and inhibiting fibroblast proliferation. Thus, we focused on the screening of such osteogenic-enhancing peptides. In the present study, we developed new peptide array screening platforms to evaluate cell proliferation and alkaline phosphatase activity in osteoblasts, UC-MSCs and fibroblasts. The conditions for the screening platform were first defined using UC-MSCs and an osteogenic differentiation peptide known as W9. Next, in silico screening to define the candidate peptides was carried out to evaluate the homology of 19 bone morphogenetic proteins. Twenty-five candidate 9-mer peptides were selected for screening. Finally, the screening of osteogenic-enhancing (osteogenic cell-selective proliferation and osteogenic differentiation) short peptide was carried out using the peptide array method, and three osteogenic-enhancing peptides were identified, confirming the validity of this screening. PMID:28773850

Bradykinin regulates osteoblast differentiation by Akt/ERK/NFκB signaling axis.

PubMed

Srivastava, Swati; Sharma, Kirti; Kumar, Narender; Roy, Partha

2014-12-01

Bradykinin (BK), a well known mediator of pain and inflammation, is also known to be involved in the process of bone resorption. The present study therefore evaluated the role of BK in osteoblast lineage commitment. Our data showed that BK inhibits the migration of bone marrow mesenchymal stem cells, but does not affect their viability. Moreover, BK also inhibits osteoblastic differentiation by significantly downregulating the levels of mRNAs for osteopontin, runX2, col24, osterix, osteocalcin genes and bone mineralization (P < 0.05). Further, BK was found to elicit the BK receptors (BDKR1 and BDKR2) mediated activation of ERK1/2 and Akt pathways, which finally led to the activation of NFκB. BK also promoted the osteoclast differentiation of bone marrow derived preosteoclast cells by upregulating the expression of c-fos, NFATC1, TRAP, clcn7, cathK, and OSCAR genes and increasing TRAP activity through NFκB pathway. In conclusion, our data suggest that BK decreases the differentiation of osteoblasts with concomitant increase in osteoclast formation and thus provides new insight into the mechanism of action of BK in modulating bone resorption. © 2014 Wiley Periodicals, Inc.

Embroidered and surface modified polycaprolactone-co-lactide scaffolds as bone substitute: in vitro characterization.

PubMed

Rentsch, Barbe; Hofmann, Andre; Breier, Annette; Rentsch, Claudia; Scharnweber, Dieter

2009-10-01

The aim of this study was to evaluate an embroidered polycaprolactone-co-lactide (trade name PCL) scaffold for the application in bone tissue engineering. The surface of the PCL scaffolds was hydrolyzed with NaOH and coated with collagen I (coll I) and chondroitin sulfate (CS). It was investigated if a change of the surface properties and the application of coll I and CS could promote cell adhesion, proliferation, and osteogenic differentiation of human mesenchymal stem cells (hMSC). The porosity (80%) and pore size (0.2-1 mm) of the scaffold could be controlled by embroidery technique and should be suitable for bone ingrowth. The treatment with NaOH made the polymer surface more hydrophilic (water contact angle dropped to 25%), enhanced the coll I adsorption (up to 15%) and the cell attachment (two times). The coll I coated scaffold improved cell attachment and proliferation (three times). CS, as part of the artificial matrix, could induce the osteogenic differentiation of hMSC without other differentiation additives. The investigated scaffolds could act not just as temporary matrix for cell migration, proliferation, and differentiation in bone tissue engineering but also have a great potential as bioartificial bone substitute.

Clinical efficacy of stem cell mediated osteogenesis and bioceramics for bone tissue engineering.

PubMed

Neman, Josh; Hambrecht, Amanda; Cadry, Cherie; Goodarzi, Amir; Youssefzadeh, Jonathan; Chen, Mike Y; Jandial, Rahul

2012-01-01

Lower back pain is a common disorder that often requires bony spinal fusion for long-term relief. Current arthrodesis procedures use bone grafts from autogenous bone, allogenic backed bone or synthetic materials. Autogenous bone grafts can result in donor site morbidity and pain at the donor site, while allogenic backed bone and synthetic materials have variable effectiveness. Given these limitations, researchers have focused on new treatments that will allow for safe and successful bone repair and regeneration. Mesenchymal stem cells (MSCs) have received attention for their ability to differentiate into osteoblasts, cells that synthesize the extracellular matrix and regulate matrix mineralization. Successful bone regeneration requires three elements: MSCs that serve as osteoblastic progenitors, osteoinductive growth factors and their pathways that promote development and differentiation of the cells as well as an osteoconductive scaffold that allows for the formation of a vascular network. Future treatments should strive to combine mesenchymal stem cells, cell-seeded scaffolds and gene therapy to optimize the efficiency and safety of tissue repair and bone regeneration.

Inactivation of the Progesterone Receptor in Mx1+ Cells Potentiates Osteogenesis in Calvaria but Not in Long Bone.

PubMed

Zhong, Zhendong A; Sun, Weihua; Chen, Haiyan; Zhang, Hongliang; Lane, Nancy E; Yao, Wei

2015-01-01

The effect of progesterone on bone remains elusive. We previously reported that global progesterone receptor (PR) knockout mice displayed high bone mass phenotype, suggesting that PR influences bone growth and modeling. Recently, Mx1+ cells were characterized to be mesenchymal stem cell-like pluripotent Cells. The aim of this study was to evaluate whether the PR in Mx1+ cells regulates osteogenesis. Using the Mx1-Cre;mT/mG reporter mouse model, we found that the calvarial cells exhibited minimal background Mx1-Cre activity prior to Cre activation by IFNα treatment as compared to the bone marrow stromal cells. IFNα treatment significantly activated Mx1-Cre in the calvarial cells. When the PR gene was deleted in the Mx1-Cre;PR-flox calvarial cells in vitro, significantly higher levels of expression of osteoblast maturation marker genes (RUNX2, Osteocalcin, and Dmp1) and osteogenic potential were detected. The PR-deficient calvariae exhibited greater bone volume, especially in the males. Although Mx1-Cre activity could be induced on the bone surface in vivo, the Mx1+ cells did not differentiate into osteocytes in long bones. Bone volumes at the distal femurs and the bone turnover marker serum Osteocalcin were similar between the Mx1-Cre;PR-flox mutant mice and the corresponding wild types in both sexes. In conclusion, our data demonstrates that blocking progesterone signaling via PRs in calvarial Mx1+ cells promoted osteoblast differentiation in the calvaria. Mx1+ was expressed by heterogeneous cells in bone marrow and did not differentiate into osteocyte during long bone development in vivo. Selectively inactivating the PR gene in Mx1+ cells affected the membrane bone formation but did not affect peripheral skeletal homeostasis.

Quantitation of calcium metabolism in postmenopausal osteoporosis and in scoliosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bronner, F.; Richelle, L. J.; Saville, P. D.

1963-06-01

By a combination of balance and isotope techniques, the following parameters of Ca metabolism were measured: pool size, rate of loss from pool, urinary excretion, fecal excretion, intake, endogenous fecal Ca, absorption, balance, bone formation, and bone resorption. The subjects were two normal women and five women with postmenopausal osteoporosis, aged 41 to 74 years, and four patients with scoliosis, aged 12 to 22 years. The latter were studied before, shortly after, and many months after immobilization in plaster casts. On the basis of observed relationships, it appeared that the negative Ca balance observed in the older women was duemore » to the low intensity of the various vectors of Ca metabolism, without clearcut distinction between the subjects with and without osteoporosis. Conversely, in the young patients with scoliosis, the negative balance incident to treatment by immobilization was associated with vectors of relatively high intensity whose relationships were altered temporarily.« less

Transforming Growth Factor β1/Smad4 Signaling Affects Osteoclast Differentiation via Regulation of miR-155 Expression.

PubMed

Zhao, Hongying; Zhang, Jun; Shao, Haiyu; Liu, Jianwen; Jin, Mengran; Chen, Jinping; Huang, Yazeng

2017-03-01

Transforming growth factor β1 (TGFβ1)/Smad4 signaling plays a pivotal role in maintenance of the dynamic balance between bone formation and resorption. The microRNA miR-155 has been reported to exert a significant role in the differentiation of macrophage and dendritic cells. The goal of this study was to determine whether miR-155 regulates osteoclast differentiation through TGFβ1/Smad4 signaling. Here, we present that TGFβ1 elevated miR-155 levels during osteoclast differentiation through the stimulation of M-CSF and RANKL. Additionally, we found that silencing Smad4 attenuated the upregulation of miR-155 induced by TGFβ1. The results of luciferase reporter experiments and ChIP assays demonstrated that TGFβ1 promoted the binding of Smad4 to the miR-155 promoter at a site located in 454 bp from the transcription start site in vivo , further verifying that miR-155 is a transcriptional target of the TGFβ1/Smad4 pathway. Subsequently, TRAP staining and qRT-PCR analysis revealed that silencing Smad4 impaired the TGFβ1-mediated inhibition on osteoclast differentiation. Finally, we found that miR-155 may target SOCS1 and MITF to suppress osteoclast differentiation. Taken together, we provide the first evidence that TGFβ1/Smad4 signaling affects osteoclast differentiation by regulation of miR-155 expression and the use of miR-155 as a potential therapeutic target for osteoclast-related diseases shows great promise.

Transforming Growth Factor β1/Smad4 Signaling Affects Osteoclast Differentiation via Regulation of miR-155 Expression

PubMed Central

Zhao, Hongying; Zhang, Jun; Shao, Haiyu; Liu, Jianwen; Jin, Mengran; Chen, Jinping; Huang, Yazeng

2017-01-01

Transforming growth factor β1 (TGFβ1)/Smad4 signaling plays a pivotal role in maintenance of the dynamic balance between bone formation and resorption. The microRNA miR-155 has been reported to exert a significant role in the differentiation of macrophage and dendritic cells. The goal of this study was to determine whether miR-155 regulates osteoclast differentiation through TGFβ1/Smad4 signaling. Here, we present that TGFβ1 elevated miR-155 levels during osteoclast differentiation through the stimulation of M-CSF and RANKL. Additionally, we found that silencing Smad4 attenuated the upregulation of miR-155 induced by TGFβ1. The results of luciferase reporter experiments and ChIP assays demonstrated that TGFβ1 promoted the binding of Smad4 to the miR-155 promoter at a site located in 454 bp from the transcription start site in vivo, further verifying that miR-155 is a transcriptional target of the TGFβ1/Smad4 pathway. Subsequently, TRAP staining and qRT-PCR analysis revealed that silencing Smad4 impaired the TGFβ1-mediated inhibition on osteoclast differentiation. Finally, we found that miR-155 may target SOCS1 and MITF to suppress osteoclast differentiation. Taken together, we provide the first evidence that TGFβ1/Smad4 signaling affects osteoclast differentiation by regulation of miR-155 expression and the use of miR-155 as a potential therapeutic target for osteoclast-related diseases shows great promise. PMID:28359146

Bone marrow-derived mesenchymal stem cells assembled with low-dose BMP-2 in a three-dimensional hybrid construct enhances posterolateral spinal fusion in syngeneic rats.

PubMed

Hu, Tao; Abbah, Sunny Akogwu; Toh, Soo Yein; Wang, Ming; Lam, Raymond Wing Moon; Naidu, Mathanapriya; Bhakta, Gajadhar; Cool, Simon M; Bhakoo, Kishore; Li, Jun; Goh, James Cho-Hong; Wong, Hee-Kit

2015-12-01

The combination of potent osteoinductive growth factor, functional osteoblastic cells, and osteoconductive materials to induce bone formation is a well-established concept in bone tissue engineering. However, supraphysiological dose of growth factor, such as recombinant human bone morphogenetic protein 2 (rhBMP-2), which is necessary in contemporary clinical application, have been reported to result in severe side effects. We hypothesize that the synergistic osteoinductive capacity of low-dose bone morphogenetic protein 2 (BMP-2) combined with undifferentiated bone marrow-derived stromal cells (BMSCs) is comparable to that of osteogenically differentiated BMSCs when used in a rodent model of posterolateral spinal fusion. A prospective study using a rodent model of posterolateral spinal fusion was carried out. Thirty-six syngeneic Fischer rats comprised the patient sample. Six groups of implants were evaluated as follows (n=6): (1) 10 µg BMP-2 with undifferentiated BMSCs; (2) 10 µg BMP-2 with osteogenic-differentiated BMSCs; (3) 2.5 µg BMP-2 with undifferentiated BMSCs; (4) 2.5 µg BMP-2 with osteogenic-differentiated BMSCs; (5) 0.5 µg BMP-2 with undifferentiated BMSCs; and (6) 0.5 µg BMP-2 with osteogenic-differentiated BMSCs. Optimal in vitro osteogenic differentiation of BMSCs was determined by quantitative real-time polymerase chain reaction (qRT-PCR) gene analysis whereas in vivo bone formation capacity was evaluated by manual palpation, micro-computed tomography, and histology. Rat BMSCs cultured in fibrin matrix that was loaded into the pores of medical-grade poly epsilon caprolactone tricalcium phosphate scaffolds differentiated toward osteogenic lineage by expressing osterix, runt-related transcription factor 2, and osteocalcium mRNA when supplemented with dexamethasone, ascorbic acid, and β-glycerophosphate. Whereas qRT-PCR revealed optimal increase in osteogenic genes expression after 7 days of in vitro culture, in vivo transplantation study showed that pre-differentiation of BMSCs before transplantation failed to promote posterolateral spinal fusion when co-delivered with low-dose BMP-2 (1/6 or 17% fusion rate). In contrast, combined delivery of undifferentiated BMSCs with low-dose BMP-2 (2.5 µg) demonstrated significantly higher fusion rate (4/6 or 67%) as well as significantly increased volume of new bone formation (p<.05). In summary, this study supports the combination of undifferentiated BMSCs and low-dose rhBMP-2 for bone tissue engineering construct. Copyright © 2015 Elsevier Inc. All rights reserved.

Effect of space relevant radiation exposure on differentiation and mineralization of murine osteoprogenitor cells

NASA Astrophysics Data System (ADS)

Lau, Patrick; Hu, Yueyuan; Hellweg, Christine; Baumstark-Khan, Christa; Reitz, Guenther

Extended exposure to altered gravity conditions like during long-term space flight results in significant bone loss. Exposure to ionizing radiation for cancer therapy causes bone damage and may increase the risk of fractures. Similarly, besides altered gravity conditions, astronauts on exploratory missions beyond low-Earth orbit will be exposed to high-energy heavy ions in addition to proton and photon radiation, although for prolonged periods and at lower doses and dose rates compared with therapy. Space conditions may place astronauts at a greater risk for mission-critical fractures. Until now, little is known about the effects of space radiation on the skeletal system especially on osteoprogenitor cells. Accelerator facilities are used to simulate parts of the radiation environment in space. Heavy ion accelerators therefore could be used to assess radiation risks for astronauts who will be exposed to higher radiation doses e.g. on a Mars mission. The aim of the present study was to determine the biological effects of spaceflight-relevant radiation exposure on the cellular level using murine osteoprogenitor cell lines compared to nonirradiated controls. To gain a deeper understanding of bone cell differenti-ation and mineralization after exposure to heavy ions, we examined gene expression modulation of bone specific transcription factors, osteoblast specific marker genes as well as genes function as coupling factors that link bone resorption to bone formation. We investigated the transcrip-tional modulation of type I collagen (Col I), osteocalcin (Ocn), Transforming growth factor-β1 (TGF-β1), interleukin-6 (IL-6) and the bone specific transcription factor Runx2 (Cbfa1). To gain deeper insight into potential cellular mechanisms involved in cellular response after ex-posure to heavy ions, we investigated gene expression modulations after exposure to energetic carbon ions (35 MeV/u, 73.2 keV/µm), iron ions (1000 MeV/u, 150 keV/µm) and lead ions (29 MeV/u, 9600 keV/µm) versus low LET X-rays. Exposure to X-irradiation dose-dependently increased the mRNA levels of Runx2 (cbfa1) whereas expression values of OCN and TGF-β1 were elevated at later time points. Exposure to heavy ions provoked a more marked effect on bone specific gene expression within the differentiation process. Collectively, our results indi-cate that heavy ions facilitate differentiation more effectively than X-rays as a major response in the progeny of irradiated osteoprogenitor cells. The data presented allow us to suggest that exposure to ionizing radiation interferes with bone formation at the level of cellular differenti-ation. In this regard, further experiments are needed to investigate gene expression patterns in mammalian cells that respond to differentiation after exposure to ionizing radiation.

Effect of oxygen plasma etching on pore size-controlled 3D polycaprolactone scaffolds for enhancing the early new bone formation in rabbit calvaria.

PubMed

Kook, Min-Suk; Roh, Hee-Sang; Kim, Byung-Hoon

2018-05-02

This study was to investigate the effects of O 2 plasma-etching of the 3D polycaprolactone (PCL) scaffold surface on preosteoblast cell proliferation and differentiation, and early new bone formation. The PCL scaffolds were fabricated by 3D printing technique. After O 2 plasma treatment, surface characterizations were examined by scanning electron microscopy, atomic force microscopy, and contact angle. MTT assay was used to determine cell proliferation. To investigate the early new bone formation, rabbits were sacrificed at 2 weeks for histological analyses. As the O 2 plasma etching time is increased, roughness and hydrophilicity of the PCL scaffold surface increased. The cell proliferation and differentiation on plasma-etched samples was significantly increased than on untreated samples. At 2 weeks, early new bone formation in O 2 plasma-etched PCL scaffolds was the higher than that of untreated scaffolds. The O 2 plasma-etched PCL scaffolds showed increased preosteoblast differentiation as well as increased new bone formation.

Direct effects of mitochondrial dysfunction on poor bone health in Leigh syndrome.

PubMed

Kato, Hiroki; Han, Xu; Yamaza, Haruyoshi; Masuda, Keiji; Hirofuji, Yuta; Sato, Hiroshi; Pham, Thanh Thi Mai; Taguchi, Tomoaki; Nonaka, Kazuaki

2017-11-04

Mitochondrial diseases are the result of aberrant mitochondrial function caused by mutations in either nuclear or mitochondrial DNA. Poor bone health has recently been suggested as a symptom of mitochondrial diseases; however, a direct link between decreased mitochondrial function and poor bone health in mitochondrial disease has not been demonstrated. In this study, stem cells from human exfoliated deciduous teeth (SHED) were isolated from a child with Leigh syndrome (LS), a mitochondrial disease, and the effects of decreased mitochondrial function on poor bone health were analyzed. Compared with control SHED, LS SHED displayed decreased osteoblastic differentiation and calcium mineralization. The intracellular and mitochondrial calcium levels were lower in LS SHED than in control SHED. Furthermore, the mitochondrial activity of LS SHED was decreased compared with control SHED both with and without osteoblastic differentiation. Our results indicate that decreased osteoblast differentiation potential and osteoblast function contribute to poor bone health in mitochondrial diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Platelet-Poor and Platelet-Rich Plasma Stimulate Bone Lineage Differentiation in Periodontal Ligament Stem Cells.

PubMed

Martínez, Constanza E; González, Sergio A; Palma, Verónica; Smith, Patricio C

2016-02-01

Plasma-derived fractions have been used as an autologous source of growth factors; however, limited knowledge concerning their biologic effects has hampered their clinical application. In this study, the authors analyze the content and specific effect of both platelet-rich plasma (PRP) and platelet-poor plasma (PPP) on osteoblastic differentiation using primary cultures of human periodontal ligament stem cells (HPLSCs). The authors evaluated the growth factor content of PRP and PPP using a proteome profiler array and enzyme-linked immunosorbent assay. HPLSCs were characterized by flow cytometry and differentiation assays. The effect of PRP and PPP on HPLSC bone differentiation was analyzed by quantifying calcium deposition after 14 and 21 days of treatment. Albeit at different concentrations, the two fractions had similar profiles of growth factors, the most representative being platelet-derived growth factor (PDGF) isoforms (PDGF-AA, -BB, and -AB), insulin-like growth factor binding protein (IGFBP)-2, and IGFBP-6. Both formulations exerted a comparable stimulus on osteoblastic differentiation even at low doses (2.5%), increasing calcium deposits in HPLSCs. PRP and PPP showed a similar protein profile and exerted comparable effects on bone differentiation. Further studies are needed to characterize and compare the effects of PPP and PRP on bone healing in vivo.

Dedifferentiated Liposarcoma of the Retroperitoneum With Heterologous Osteosarcomatous Differentiation and a Striking Aneurysmal Bone Cyst-Like Morphology.

PubMed

Van Haverbeke, Carole; Van Dorpe, Jo; Lecoutere, Evelyne; Flucke, Uta; Ferdinande, Liesbeth; Creytens, David

2017-06-01

A 69-year-old woman with a 10-year medical history of recurrent retroperitoneal dedifferentiated liposarcoma presented with a 3-cm large hemorrhagic and multicystic left-sided retroperitoneal mass. Histopathological examination of the resected specimen showed a heterogeneous, high-grade mesenchymal nonlipogenic tumor with areas of osteoblastic/osteosarcomatous differentiation and aneurysmal bone cyst-like features. Based on the clinical presentation, the morphology, and the supportive immunohistochemical and molecular findings (MDM2 overexpression and amplification of the MDM2 gene, respectively), a diagnosis of a dedifferentiated liposarcoma with heterologous osteosarcomatous differentiation and an aneurysmal bone cyst-like morphology was made. To the best of our knowledge, this is the first description of aneurysmal bone cyst-like morphology in dedifferentiated liposarcoma, further expanding the broad morphological spectrum of dedifferentiated liposarcoma.

Interaction between bone marrow stromal cells and neuroblastoma cells leads to a VEGFA-mediated osteoblastogenesis

PubMed Central

HaDuong, Josephine H.; Blavier, Laurence; Baniwal, Sanjeev K.; Frenkel, Baruch; Malvar, Jemily; Punj, Vasu; Sposto, Richard; DeClerck, Yves A.

2017-01-01

The potential role of osteoblasts in bone and bone marrow (BM) metastases in neuroblastoma (NBL) remains unclear. In this study, we examined the effect of NBL cells on the osteoblastic differentiation of bone marrow-derived mesenchymal stromal cells (BMMSC). We show that the presence of NBL cells enhanced the osteoblastic differentiation of BMMSC driven by bone morphogenetic protein (BMP)-4, in the absence of any effect on NBL cell proliferation. Expression profiles of BMMSC driven towards osteoblastic differentiation revealed an increase in vascular endothelial growth factor A (Vegfa) expression in the presence of NBL cells. We demonstrated that NBL cells increased BMMSC-derived VEGFA mRNA and protein and that this was enhanced by BMP-4. However, in similar conditions, neither the addition of an mVEGFA blocking antibody nor exogenous recombinant (r) mVEGFA affected osteoblastic differentiation. In contrast, siRNA-mediated knock-down of VEGFA in BMMSC prevented osteoblastic differentiation in BMP-4-treated co-cultures, an effect that was not reversed in the presence of rmVEGFA. An analysis of murine bones injected with hNBL cells revealed an increase of mVEGFA producing cells near tumor cells concomitantly with an increase in Vegfa and Runx2 mRNA. This coincided with an increase in osteoclasts, in Rankl/Opg mRNA ratio and with the formation of osteolytic lesions. Thus NBL cells promote osteoblastogenesis in the BM by increasing VEGFA expression in BMMSC. Our study provides a new insight into the role of VEGFA in NBL metastases by pointing to the role of stroma-derived intracrine VEGFA in osteoblastogenesis. PMID:25648303

Osteogenic differentiation capacity of human skeletal muscle-derived progenitor cells.

PubMed

Oishi, Teruyo; Uezumi, Akiyoshi; Kanaji, Arihiko; Yamamoto, Naoki; Yamaguchi, Asami; Yamada, Harumoto; Tsuchida, Kunihiro

2013-01-01

Heterotopic ossification (HO) is defined as the formation of ectopic bone in soft tissue outside the skeletal tissue. HO is thought to result from aberrant differentiation of osteogenic progenitors within skeletal muscle. However, the precise origin of HO is still unclear. Skeletal muscle contains two kinds of progenitor cells, myogenic progenitors and mesenchymal progenitors. Myogenic and mesenchymal progenitors in human skeletal muscle can be identified as CD56(+) and PDGFRα(+) cells, respectively. The purpose of this study was to investigate the osteogenic differentiation potential of human skeletal muscle-derived progenitors. Both CD56(+) cells and PDGFRα(+) cells showed comparable osteogenic differentiation potential in vitro. However, in an in vivo ectopic bone formation model, PDGFRα(+) cells formed bone-like tissue and showed successful engraftment, while CD56(+) cells did not form bone-like tissue and did not adapt to an osteogenic environment. Immunohistological analysis of human HO sample revealed that many PDGFRα(+) cells were localized in proximity to ectopic bone formed in skeletal muscle. MicroRNAs (miRNAs) are known to regulate many biological processes including osteogenic differentiation. We investigated the participation of miRNAs in the osteogenic differentiation of PDGFRα(+) cells by using microarray. We identified miRNAs that had not been known to be involved in osteogenesis but showed dramatic changes during osteogenic differentiation of PDGFRα(+) cells. Upregulation of miR-146b-5p and -424 and downregulation of miR-7 during osteogenic differentiation of PDGFRα(+) cells were confirmed by quantitative real-time RT-PCR. Inhibition of upregulated miRNAs, miR-146b-5p and -424, resulted in the suppression of osteocyte maturation, suggesting that these two miRNAs have the positive role in the osteogenesis of PDGFRα(+) cells. Our results suggest that PDGFRα(+) cells may be the major source of HO and that the newly identified miRNAs may regulate osteogenic differentiation process of PDGFRα(+) cells.

Endochondral bone formation in embryonic mouse pre-metatarsals

NASA Technical Reports Server (NTRS)

Klement, B. J.; Spooner, B. S.

1992-01-01

Long term exposure to a reduced gravitational environment has a deleterious effect on bone. The developmental events which occur prior to initial bone deposition will provide insight into the regulation of mature bone physiology. We have characterized a system in which the events preceding bone formation take place in an isolated in vitro organ culture environment. We show that cultured pre-metatarsal tissue parallels development of pre-metatarsal tissue in the embryo. Both undergo mesenchyme differentiation and morphogenesis to form a cartilage rod, which resembles the future bone, followed by terminal chondrocyte differentiation in a definite morphogenetic pattern. These sequential steps occur prior to osteoblast maturation and bone matrix deposition in the developing organism. Alkaline phosphatase (ALP) activity is a distinctive enzymatic marker for mineralizing tissues. We have measured this activity throughout pre-metatarsal development and show (a) where in the tissue it is predominantly found, and (b) that this is indeed the mineralizing isoform of the enzyme.

Osteoblast dysfunctions in bone diseases: from cellular and molecular mechanisms to therapeutic strategies.

PubMed

Marie, Pierre J

2015-04-01

Several metabolic, genetic and oncogenic bone diseases are characterized by defective or excessive bone formation. These abnormalities are caused by dysfunctions in the commitment, differentiation or survival of cells of the osteoblast lineage. During the recent years, significant advances have been made in our understanding of the cellular and molecular mechanisms underlying the osteoblast dysfunctions in osteoporosis, skeletal dysplasias and primary bone tumors. This led to suggest novel therapeutic approaches to correct these abnormalities such as the modulation of WNT signaling, the pharmacological modulation of proteasome-mediated protein degradation, the induction of osteoprogenitor cell differentiation, the repression of cancer cell proliferation and the manipulation of epigenetic mechanisms. This article reviews our current understanding of the major cellular and molecular mechanisms inducing osteoblastic cell abnormalities in age-related bone loss, genetic skeletal dysplasias and primary bone tumors, and discusses emerging therapeutic strategies to counteract the osteoblast abnormalities in these disorders of bone formation.

Critical-Size Calvarial Bone Defects Healing in a Mouse Model with Silk Scaffolds and SATB2- Modified iPSCs

PubMed Central

Ye, Jin-Hai; Xu, Yuan-Jin; Gao, Jun; Yan, Shi-Guo; Zhao, Jun; Tu, Qisheng; Zhang, Jin; Duan, Xue-Jing; Sommer, Cesar A.; Mostoslavsky, Gustavo; Kaplan, David; Wu, Yu-Nong; Zhang, Chen-Ping; Wang, Lin; Chen, Jake

2011-01-01

Induced pluripotent stem cells (iPSCs) can differentiate into mineralizing cells and thus have a great potential in application in engineered bone substitutes with bioactive scaffolds in regeneration medicine. In the current study we characterized and demonstrated the pluripotency and osteogenic differentiation of mouse iPSCs. To enhance the osteogenic differentiation of iPSCs, we then transduced the iPSCs with the potent transcription factor, nuclear matrix protein SATB2. We observed that in SATB2-overexpressing iPSCs there were increased mineral nodule formation and elevated mRNA levels of key osteogenic genes, osterix (OSX), Runx2, bone sialoprotein (BSP) and osteocalcin (OCN). Moreover, the mRNA levels of HoxA2 was reduced after SATB2 overexpression in iPSCs. The SATB2-overexpressing iPSCs were then combined with silk scaffolds and transplanted into critical-size calvarial bone defects created in nude mice. Five weeks post-surgery, radiological and micro-CT analysis revealed enhanced new bone formation in calvarial defects in SATB2 group. Histological analysis also showed increased new bone formation and mineralization in the SATB2 group. In conclusion, the results demonstrate that SATB2 facilitates the differentiation of iPSCs towards osteoblast-lineage cells by repressing HoxA2 and augmenting the functions of the osteoblast determinants Runx2, BSP and OCN. PMID:21492931

NF-κB inhibits osteogenic differentiation of mesenchymal stem cells by promoting β-catenin degradation

PubMed Central

Chang, Jia; Liu, Fei; Lee, Min; Wu, Benjamin; Ting, Kang; Zara, Janette N.; Soo, Chia; Al Hezaimi, Khalid; Zou, Weiping; Chen, Xiaohong; Mooney, David J.; Wang, Cun-Yu

2013-01-01

Mesenchymal stem cell (MSC)-based transplantation is a promising therapeutic approach for bone regeneration and repair. In the realm of therapeutic bone regeneration, the defect or injured tissues are frequently inflamed with an abnormal expression of inflammatory mediators. Growing evidence suggests that proinflammatory cytokines inhibit osteogenic differentiation and bone formation. Thus, for successful MSC-mediated repair, it is important to overcome the inflammation-mediated inhibition of tissue regeneration. In this study, using genetic and chemical approaches, we found that proinflammatory cytokines TNF and IL-17 stimulated IκB kinase (IKK)–NF-κB and impaired osteogenic differentiation of MSCs. In contrast, the inhibition of IKK–NF-κB significantly enhanced MSC-mediated bone formation. Mechanistically, we found that IKK–NF-κB activation promoted β-catenin ubiquitination and degradation through induction of Smurf1 and Smurf2. To translate our basic findings to potential clinic applications, we showed that the IKK small molecule inhibitor, IKKVI, enhanced osteogenic differentiation of MSCs. More importantly, the delivery of IKKVI promoted MSC-mediated craniofacial bone regeneration and repair in vivo. Considering the well established role of NF-κB in inflammation and infection, our results suggest that targeting IKK–NF-κB may have dual benefits in enhancing bone regeneration and repair and inhibiting inflammation, and this concept may also have applicability in many other tissue regeneration situations. PMID:23690607

Effects of intermittent versus continuous parathyroid hormone administration on condylar chondrocyte proliferation and differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Qi; Wan, Qilong; Yang, Rongtao

Highlights: Black-Right-Pointing-Pointer Different PTH administration exerts different effects on condylar chondrocyte. Black-Right-Pointing-Pointer Intermittent PTH administration suppresses condylar chondrocyte proliferation. Black-Right-Pointing-Pointer Continuous PTH administration maintains condylar chondrocyte proliferating. Black-Right-Pointing-Pointer Intermittent PTH administration enhances condylar chondrocyte differentiation. -- Abstract: Endochondral ossification is a complex process involving chondrogenesis and osteogenesis regulated by many hormones and growth factors. Parathyroid hormone (PTH), one of the key hormones regulating bone metabolism, promotes osteoblast differentiation and osteogenesis by intermittent administration, whereas continuous PTH administration inhibits bone formation. However, the effects of PTH on chondrocyte proliferation and differentiation are still unclear. In this study, intermittent PTH administration presentedmore » enhanced effects on condylar chondrocyte differentiation and bone formation, as demonstrated by increased mineral nodule formation and alkaline phosphatase (ALP) activity, up-regulated runt-related transcription factor 2 (RUNX2), ALP, collagen type X (COL10a1), collagen type I (COL1a1), osteocalcin (OCN), bone sialoprotein (BSP), bone morphogenetic protein 2 (BMP2) and osterix (OSX) mRNA and/or protein expression. On the contrary, continuous PTH administration promoted condylar chondrocyte proliferation and suppressed its differentiation, as demonstrated by up-regulated collagen type II (COL2a1) mRNA expression, reduced mineral nodule formation and down-regulated expression of the mRNAs and/or proteins mentioned above. Our data suggest that PTH can regulate condylar chondrocyte proliferation and differentiation, depending on the type of PTH administration. These results provide new insight into the effects of PTH on condylar chondrocytes and new evidence for using local PTH administration to cure mandibular asymmetry.« less

Stromal derived factor-1 regulates bone morphogenetic protein 2-induced osteogenic differentiation of primary mesenchymal stem cells

PubMed Central

Hosogane, Naobumi; Huang, Zhiping; Rawlins, Bernard A.; Liu, Xia; Boachie-Adjei, Oheneba; Boskey, Adele L.; Zhu, Wei

2010-01-01

Stromal derived factor-1 (SDF-1) is a chemokine signaling molecule that binds to its transmembrane receptor CXC chemokine receptor-4 (CXCR4). While we previously detected that SDF-1 was co-required with bone morphogenetic protein 2 (BMP2) for differentiating mesenchymal C2C12 cells into osteoblastic cells, it is unknown whether SDF-1 is similarly involved in the osteogenic differentiation of mesenchymal stem cells (MSCs). Therefore, here we examined the role of SDF-1 signaling during BMP2-induced osteogenic differentiation of primary MSCs that were derived from human and mouse bone marrow. Our data showed that blocking of the SDF-1/CXCR4 signal axis or adding SDF-1 protein to MSCs significantly affected BMP2-induced alkaline phosphatase (ALP) activity and osteocalcin (OCN) synthesis, markers of preosteoblasts and mature osteoblasts, respectively. Moreover, disrupting the SDF-1 signaling impaired bone nodule mineralization during terminal differentiation of MSCs. Furthermore, we detected that blocking of the SDF-1 signaling inhibited the BMP2-induced early expression of Runt-related factor-2 (Runx2) and osterix (Osx), two “master” regulators of osteogenesis, and the SDF-1 effect was mediated via intracellular Smad and Erk activation. In conclusion, our results demonstrated a regulatory role of SDF-1 in BMP2-induced osteogenic differentiation of MSCs, as perturbing the SDF-1 signaling affected the differentiation of MSCs towards osteoblastic cells in response to BMP2 stimulation. These data provide novel insights into molecular mechanisms underlying MSC osteogenesis, and will contribute to the development of MSC therapies for enhancing bone formation and regeneration in broad orthopaedic situations. PMID:20362069

Bropirimine inhibits osteoclast differentiation through production of interferon-β

DOE Office of Scientific and Technical Information (OSTI.GOV)

Suzuki, Hiroaki; Mochizuki, Ayako; Yoshimura, Kentaro

Bropirimine is a synthetic agonist for toll-like receptor 7 (TLR7). In this study, we investigated the effects of bropirimine on differentiation and bone-resorbing activity of osteoclasts in vitro. Bropirimine inhibited osteoclast differentiation of mouse bone marrow-derived macrophages (BMMs) induced by receptor activator of nuclear factor κB ligand (RANKL) in a concentration-dependent manner. Furthermore, it suppressed the mRNA expression of nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 1 (NFATc1), a master transcription factor for osteoclast differentiation, without affecting BMM viability. Bropirimine also inhibited osteoclast differentiation induced in co-cultures of mouse bone marrow cells (BMCs) and mouse osteoblastic UAMS-32 cells in the presencemore » of activated vitamin D{sub 3}. Bropirimine partially suppressed the expression of RANKL mRNA in UAMS-32 cells induced by activated vitamin D{sub 3}. Finally, the anti-interferon-β (IFN-β) antibody restored RANKL-dependent differentiation of BMMs into osteoclasts suppressed by bropirimine. These results suggest that bropirimine inhibits differentiation of osteoclast precursor cells into osteoclasts via TLR7-mediated production of IFN-β.« less

Isometric Scaling in Developing Long Bones Is Achieved by an Optimal Epiphyseal Growth Balance

PubMed Central

Stern, Tomer; Aviram, Rona; Rot, Chagai; Galili, Tal; Sharir, Amnon; Kalish Achrai, Noga; Keller, Yosi; Shahar, Ron; Zelzer, Elazar

2015-01-01

One of the major challenges that developing organs face is scaling, that is, the adjustment of physical proportions during the massive increase in size. Although organ scaling is fundamental for development and function, little is known about the mechanisms that regulate it. Bone superstructures are projections that typically serve for tendon and ligament insertion or articulation and, therefore, their position along the bone is crucial for musculoskeletal functionality. As bones are rigid structures that elongate only from their ends, it is unclear how superstructure positions are regulated during growth to end up in the right locations. Here, we document the process of longitudinal scaling in developing mouse long bones and uncover the mechanism that regulates it. To that end, we performed a computational analysis of hundreds of three-dimensional micro-CT images, using a newly developed method for recovering the morphogenetic sequence of developing bones. Strikingly, analysis revealed that the relative position of all superstructures along the bone is highly preserved during more than a 5-fold increase in length, indicating isometric scaling. It has been suggested that during development, bone superstructures are continuously reconstructed and relocated along the shaft, a process known as drift. Surprisingly, our results showed that most superstructures did not drift at all. Instead, we identified a novel mechanism for bone scaling, whereby each bone exhibits a specific and unique balance between proximal and distal growth rates, which accurately maintains the relative position of its superstructures. Moreover, we show mathematically that this mechanism minimizes the cumulative drift of all superstructures, thereby optimizing the scaling process. Our study reveals a general mechanism for the scaling of developing bones. More broadly, these findings suggest an evolutionary mechanism that facilitates variability in bone morphology by controlling the activity of individual epiphyseal plates. PMID:26241802

Postradiation atrophy of mature bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Erguen, H.; Howland, W.J.

1980-01-01

The growing number of oncological patients subjected to radiotherapy require the diagnostic radiologist to be aware of expected bone changes following irradiation and the differentiation of this entity from metastasis. The primary event of radiation damage to bone is atrophy and true necrosis of bone is uncommon. The postradiation atrophic changes of bone are the result of combined cellular and vascular damage, the former being more important. The damage to the osteoblast resulting in decreased matrix production is apparently the primary histopathologic event. Radiation damaged bone is susceptible to superimposed complications of fracture, infection, necrosis, and sarcoma. The primary radiographicmore » evidence of atrophy, localized osteopenia, is late in appearing, mainly because of the relative insensitivity of radiographs in detecting demineralization. Contrary to former views, the mature bone is quite radiosensitive and reacts quickly to even small doses of radiation. In vivo midrodensitometric analysis and radionuclide bone and bone marrow scans can reveal early changes following irradiation. The differentiation of postirradiation atrophy and metastasis may be difficult. Biopsy should be the last resort because of the possibility of causing true necrosis in atrophic bone by trauma and infection.« less

Postradiation atrophy of mature bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ergun, H.; Howland, W.J.

1980-01-01

The growing number of oncological patients subjected to radiotherapy require the diagnostic radiologist to be aware of expected bone changes following irradiation and the differentiation of this entity from metastasis. The primary event of radiation damage to bone is atrophy and true necrosis of bone is uncommon. The postradiation atrophic changes of bone are the result of combined cellular and vascular damage, the former being more important. The damage to the osteoblast resulting in decreased matrix production is apparently the primary histopathologic event. Radiation damaged bone is susceptible to superimposed complications of fracture, infection, necrosis, and sarcoma. The primary radiographicmore » evidence of atrophy, localized osteopenia, is late in appearing, mainly because of the relative insensitivity of radiographs in detecing demineralization. Contrary to former views, the mature bone is quite radiosensitive and reacts quickly to even small doses of radiation. In vivo midrodensitometric analysis and radionuclide bone and bone marrow scans can reveal early changes following irradiation. The differentiation of postirradiation atrophy and metastasis may be difficult. Biopsy should be the last resort because of the possibility of causing true necrosis in atrophic bone by trauma and infection.« less

Bone conditioned media (BCM) improves osteoblast adhesion and differentiation on collagen barrier membranes.

PubMed

Fujioka-Kobayashi, Masako; Caballé-Serrano, Jordi; Bosshardt, Dieter D; Gruber, Reinhard; Buser, Daniel; Miron, Richard J

2016-07-04

The use of autogenous bone chips during guided bone regeneration procedures has remained the gold standard for bone grafting due to its excellent combination of osteoconduction, osteoinduction and osteogenesis. Recent protocols established by our group have characterized specific growth factors and cytokines released from autogenous bone that have the potential to be harvested and isolated into bone conditioned media (BCM). Due to the advantageous osteo-promotive properties of BCM, the aims of the present study was to pre-coat collagen barrier membranes with BCM and investigate its effect on osteoblast adhesion, proliferation and differentiation for possible future clinical use. Scanning electron microscopy (SEM) was first used to qualitative assess BCM protein accumulation on the surface of collagen membranes. Thereafter, undifferentiated mouse ST2 stromal bone marrow cells were seeded onto BioGide porcine derived collagen barrier membranes (control) or barrier membranes pre-coated with BCM (test group). Control and BCM samples were compared for cell adhesion at 8 h, cell proliferation at 1, 3 and 5 days and real-time PCR at 5 days for osteoblast differentiation markers including Runx2, alkaline phosphatase (ALP), osteocalcin (OCN) and bone sialoprotein (BSP). Mineralization was further assessed with alizarin red staining at 14 days post seeding. SEM images demonstrated evidence of accumulated proteins found on the surface of collagen membranes following coating with BCM. Analysis of total cell numbers revealed that the additional pre-coating with BCM markedly increased cell attachment over 4 fold when compared to cells seeded on barrier membranes alone. No significant difference could be observed for cell proliferation at all time points. BCM significantly increased mRNA levels of osteoblast differentiation markers including ALP, OCN and BSP at 5 days post seeding. Furthermore, barrier membranes pre-coated with BCM demonstrated a 5-fold increase in alizarin red staining at 14 days. The results from the present study suggest that the osteoconductive properties of porcine-derived barrier membranes could be further improved by BCM by significantly increasing cell attachment, differentiation and mineralization of osteoblasts in vitro. Future animal testing is required to fully characterize the additional benefits of BCM for guided bone regeneration.

Sphere-shaped nano-hydroxyapatite/chitosan/gelatin 3D porous scaffolds increase proliferation and osteogenic differentiation of human induced pluripotent stem cells from gingival fibroblasts.

PubMed

Ji, Jun; Tong, Xin; Huang, Xiaofeng; Wang, Tiancong; Lin, Zitong; Cao, Yazhou; Zhang, Junfeng; Dong, Lei; Qin, Haiyan; Hu, Qingang

2015-07-08

Hydroxyapatite (HA) is an important component of human bone and bone tissue engineering scaffolds. A plethora of bone tissue engineering scaffolds have been synthesized so far, including nano-HA/chitosan/gelatin (nHA/CG) scaffolds; and for seeding cells, stem cells, especially induced pluripotent stem cells (iPSCs), have been a promising cell source for bone tissue engineering recently. However, the influence of different HA nano-particle morphologies on the osteogenic differentiation of human iPSCs (hiPSCs) from human gingival fibroblasts (hGFs) is unknown. The purpose of this study was to investigate the osteogenic differentiation of hiPSCs from hGFs seeded on nHA/CG scaffolds with 2 shapes (rod and sphere) of nHA particles. Firstly, hGFs isolated from discarded normal gingival tissues were reprogrammed into hiPSCs. Secondly, hiPSCs were seeded on rod-like nHA/CG (rod-nHA/CG) and sphere-shaped nHA/CG (sphere-nHA/CG) scaffolds respectively and then cell/scaffold complexes were cultured in vitro. Scanning electron microscope, hematoxyline and eosin (HE) staining, Masson's staining, and quantitative real-time polymerase chain reaction techniques were used to examine hiPSC morphology, proliferation, and differentiation on rod-nHA/CG and sphere-nHA/CG scaffolds. Finally, hiPSCs composited with 2 kinds of nHA/CG were transplanted in vivo in a subcutaneous implantation model for 12 weeks; pure scaffolds were also transplanted as a blank control. HE, Masson's, and immunohistochemistry staining were applied to detect new bone regeneration ability. The results showed that sphere-nHA/CG significantly increased hiPSCs from hGF proliferation and osteogenic differentiation in vitro. hiPSCs and sphere-nHA/CG composities generated large bone, whereas hiPSCs and rod-nHA/CG composities produced tiny bone in vivo. Moreover, pure scaffolds without cells almost produced no bone. In conclusion, our work provided a potential innovative bone tissue engineering approach using clinically discarded gingival tissues and sphere-nHA/CG scaffolds.

Tributyltin induces distinct effects on cortical and trabecular bone in female C57Bl/6J mice.

PubMed

Watt, James; Baker, Amelia H; Meeks, Brett; Pajevic, Paola D; Morgan, Elise F; Gerstenfeld, Louis C; Schlezinger, Jennifer J

2018-09-01

The retinoid X receptors (RXR), peroxisome proliferator activated receptor gamma (PPARγ), and liver X receptors (LXR) all have been shown to regulate bone homeostasis. Tributyltin (TBT) is an environmental contaminant that is a dual RXRα/β and PPARγ agonist. TBT induces RXR, PPARγ, and LXR-mediated gene transcription and suppresses osteoblast differentiation in vitro. Bone marrow multipotent mesenchymal stromal cells derived from female C57BL/6J mice were more sensitive to suppression of osteogenesis by TBT than those derived from male mice. In vivo, oral gavage of 12 week old female, C57Bl/6J mice with 10 mg/kg TBT for 10 weeks resulted in femurs with a smaller cross-sectional area and thinner cortex. Surprisingly, TBT induced significant increases in trabecular thickness, number, and bone volume fraction. TBT treatment did not change the Rankl:Opg RNA ratio in whole bone, and histological analyses showed that osteoclasts in the trabecular space were minimally reduced. In contrast, expression of cardiotrophin-1, an osteoblastogenic cytokine secreted by osteoclasts, increased. In primary bone marrow macrophage cultures, TBT marginally inhibited the number of osteoclasts that differentiated, in spite of significantly suppressing expression of osteoclast markers Nfatc1, Acp5, and Ctsk and resorptive activity. TBT induced expression of RXR- and LXR-dependent genes in whole bone and in vitro osteoclast cultures. However, only an RXR antagonist, but not an LXR antagonist, significantly inhibited TBTs ability to suppress osteoclast differentiation. These results suggest that TBT has distinct effects on cortical versus trabecular bone, likely resulting from independent effects on osteoblast and osteoclast differentiation that are mediated through RXR. © 2018 Wiley Periodicals, Inc.

The effect of dexamethasone and triiodothyronine on terminal differentiation of primary bovine chondrocytes and chondrogenically differentiated mesenchymal stem cells.

PubMed

Randau, Thomas M; Schildberg, Frank A; Alini, Mauro; Wimmer, Matthias D; Haddouti, El-Mustapha; Gravius, Sascha; Ito, Keita; Stoddart, Martin J

2013-01-01

The newly evolved field of regenerative medicine is offering solutions in the treatment of bone or cartilage loss and deficiency. Mesenchymal stem cells, as well as articular chondrocytes, are potential cells for the generation of bone or cartilage. The natural mechanism of bone formation is that of endochondral ossification, regulated, among other factors, through the hormones dexamethasone and triiodothyronine. We investigated the effects of these hormones on articular chondrocytes and chondrogenically differentiated mesenchymal stem cells, hypothesizing that these hormones would induce terminal differentiation, with chondrocytes and differentiated stem cells being similar in their response. Using a 3D-alginate cell culture model, bovine chondrocytes and chondrogenically differentiated stem cells were cultured in presence of triiodothyronine or dexamethasone, and cell proliferation and extracellular matrix production were investigated. Collagen mRNA expression was measured by real-time PCR. Col X mRNA and alkaline phosphatase were monitored as markers of terminal differentiation, a prerequisite of endochondral ossification. The alginate culture system worked well, both for the culture of chondrocytes and for the chondrogenic differentiation of mesenchymal stem cells. Dexamethasone led to an increase in glycosaminoglycan production. Triiodothyronine increased the total collagen production only in chondrocytes, where it also induced signs of terminal differentiation, increasing both collagen X mRNA and alkaline phosphatase activity. Dexamethasone induced terminal differentiation in the differentiated stem cells. The immature articular chondrocytes used in this study seem to be able to undergo terminal differentiation, pointing to their possible role in the onset of degenerative osteoarthritis, as well as their potential for a cell source in bone tissue engineering. When chondrocyte-like cells, after their differentiation, can indeed be moved on towards terminal differentiation, they can be used to generate a model of endochondral ossification, but this limitation must be kept in mind when using them in cartilage tissue engineering application.

Influence of pore size of porous titanium fabricated by vacuum diffusion bonding of titanium meshes on cell penetration and bone ingrowth.

PubMed

Chang, Bei; Song, Wen; Han, Tianxiao; Yan, Jun; Li, Fuping; Zhao, Lingzhou; Kou, Hongchao; Zhang, Yumei

2016-03-01

The present work assesses the potential of three-dimensional (3D) porous titanium (pore size of 188-390 μm and porosity of 70%) fabricated by vacuum diffusion bonding of titanium meshes for applications in bone engineering. Rat bone marrow mesenchymal stem cells were used to investigate the proliferation and differentiation of cells on titanium scaffolds with different pore sizes at day 7, day 14 and day 21 based on DNA contents, alkaline phosphatase (ALP) activity, collagen (COL) secretion and osteogenic gene expressions including ALP, COL-1, bone morphogenetic protein-2 (BMP-2), osteopontin (OPN), runt-related transcription factor 2 (RUNX2), using smooth solid titanium plate as reference material. The rabbit models with distal femoral condyles defect were used to investigate the bone ingrowth into the porous titanium. All samples were subjected to Micro-CT and histological analysis after 4 and 12 weeks of healing. A one-way ANOVA followed by Tukey post hoc tests was used to analyze the data. It was found that the differentiation stage of cells on the porous titanium delayed compared with the smooth solid titanium plate and Ti 188 was more inclined to promote cell differentiation at the initial stage (day 14) while cell proliferation (day 1, 4, 7, 10, 14 and 21) and bone ingrowth (4 and 12 weeks) were biased to Ti 313 and Ti 390. The study indicates that the hybrid porous implant design which combines the advantages of different pore sizes may be meaningful and promising for bone defect restoration. One of the significant challenges in bone defect restoration is the integration of biomaterials and surrounding bone tissue. Porous titanium may be a promising choice for bone ingrowth and mineralization with appropriate mechanical and biological properties. In this study, based on porous titanium fabricated by vacuum diffusion bonding of titanium meshes, we have evaluated the influence of various pore sizes on rat bone marrow mesenchymal stem cells (rBMMSCs) penetration in vitro and bone ingrowth in vivo. It was interesting that we found the proliferation and differentiation abilities of rBMMSCs, as well as bone ingrowth were related to different pore sizes of such porous scaffolds. The results may provide guidance for porous titanium design for bone defect restoration. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Amniotic fluid-derived mesenchymal stem cells lead to bone differentiation when cocultured with dental pulp stem cells.

PubMed

De Rosa, Alfredo; Tirino, Virginia; Paino, Francesca; Tartaglione, Antonella; Mitsiadis, Thimios; Feki, Anis; d'Aquino, Riccardo; Laino, Luigi; Colacurci, Nicola; Papaccio, Gianpaolo

2011-03-01

Mesenchymal stem cells are present in many tissues of the human body, including amniotic fluid (AF) and dental pulp (DP). Stem cells of both AF and DP give rise to a variety of differentiated cells. In our experience, DP stem cells (DPSCs) display a high capacity to produce bone. Therefore, our aim was to investigate if AF-derived stem cells (AFSCs) were able to undergo bone differentiation in the presence of DPSCs. AFSCs were seeded under three different conditions: (i) cocultured with DPSCs previously differentiated into osteoblasts; (ii) cultured in the conditioned medium of osteoblast-differentiated DPSCs; (iii) cultured in the osteogenic medium supplemented with vascular endothelial growth factor and bone morphogenetic protein-2 (BMP-2). Results showed that AFSCs were positive for mesenchymal markers, and expressed high levels of Tra1-60, Tra1-80, BMPR1, BMPR2, and BMP-2. In contrast, AFSCs were negative for epithelial and hematopoietic/endothelial markers. When AFSCs were cocultured with DPSCs-derived osteoblasts, they differentiated into osteoblasts. A similar effect was observed when AFSCs were cultured in the presence of a conditioned medium originated from DPSCs. We found that osteoblasts derived from DPSCs released large amounts of BMP-2 and vascular endothelial growth factor into the culture medium and that those morphogens significantly upregulate RUNX-2 gene, stimulating osteogenesis. This study highlights the mechanisms of osteogenesis and strongly suggests that the combination of AFSCs with DPSCs may provide a rich source of soluble proteins useful for bone engineering purposes.

Complex effect of hydroxyapatite nanoparticles on the differentiation and functional activity of human pre-osteoclastic cells.

PubMed

Costa-Rodrigues, João; Silva, Ana; Santos, Catarina; Almeida, Maria Margarida; Costa, Maria Elisabete; Fernandes, Maria Helena

2014-12-01

Nanosized hydroxyapatite (HA) is a promising material in clinical applications targeting the bone tissue. NanoHA is able to modulate bone cellular events, which accounts for its potential utility, but also raises safety concerns regarding the maintenance of the bone homeostasis. This work analyses the effects of HA nanoparticles (HAnp) on osteoclastic differentiation and activity, an issue that has been barely addressed. Rod-like HAnp, produced by a hydrothermal precipitation method, were tested on peripheral blood mononuclear cells (PBMC), which contains the CD14+ osteoclastic precursors, in unstimulated or osteoclastogenic-induced conditions. HAnp were added at three time-points during the osteoclastic differentiation pathway, and cell response was evaluated for osteoclastic related parameters. Results showed that HAnp modulated the differentiation and function of osteoclastic cells in a dose- and time-dependent manner. In addition, the effects were dependent on the stage of osteoclastic differentiation. In unstimulated PBMC, HAnp significantly increased osteoclastogenesis, leading to the formation of mature osteoclasts, as evident by the significant increase of TRAP activity, number of TRAP-positive multinucleated cells, osteoclastic gene expression and resorbing ability. However, in a population of mature osteoclasts (formed in osteoclastogenic-induced PBMC cultures), HAnp caused a dose-dependent decrease on the osteoclastic-related parameters. These results highlight the complex effects of HAnp in osteoclastic differentiation and activity, and suggest the possibility of HAnp to modulate/disrupt osteoclastic behavior, with eventual imbalances in the bone metabolism. This should be carefully considered in bone-related and other established and prospective biomedical applications of HAnp.

Resveratrol Increases Osteoblast Differentiation In Vitro Independently of Inflammation.

PubMed

Ornstrup, Marie Juul; Harsløf, Torben; Sørensen, Lotte; Stenkjær, Liselotte; Langdahl, Bente Lomholt; Pedersen, Steen Bønløkke

2016-08-01

Low-grade inflammation negatively affects bone. Resveratrol is a natural compound proven to possess both anti-inflammatory and bone protective properties. However, it is uncertain if the bone effects are mediated though anti-inflammatory effects. Firstly, we investigated if resveratrol affects proliferation and differentiation of human bone marrow-derived mesenchymal stem cells. Secondly, we investigated if inflammation negatively affects proliferation and differentiation, and if resveratrol counteracts this through anti-inflammatory effects. Mesenchymal stem cells were obtained from bone marrow aspiration in 13 healthy individuals and cultured towards the osteoblast cell lineage. The cells were stimulated with resveratrol, lipopolysaccharide (LPS), LPS + resveratrol, or vehicle (control) for 21 days. Compared to control, resveratrol decreased cell number by 35 % (p < 0.05) and induced differentiation (a 3-fold increase in alkaline phosphatase (p < 0.002), while P1NP and OPG showed similar trends). LPS induced inflammation with a 44-fold increase in interleukin-6 (p < 0.05) and an extremely prominent increase in interleukin-8 production (p < 0.05) relative to control. In addition, LPS increased cell count (p < 0.05) and decreased differentiation (a reduction in P1NP production (p < 0.02)). Co-stimulation with LPS + resveratrol did not reduce interleukin-6 or interleukin-8, but nonetheless, cell count was reduced (p < 0.05) and alkaline phosphatase, P1NP, and OPG increased (p < 0.05 for all). Thus, resveratrol stimulates osteoblast differentiation independently of inflammation.

Osterix/Sp7 limits cranial bone initiation sites and is required for formation of sutures

PubMed Central

Kague, Erika; Roy, Paula; Asselin, Garrett; Hu, Gui; Stanley, Alexandra; Albertson, Craig; Simonet, Jacqueline; Fisher, Shannon

2017-01-01

During growth, individual skull bones overlap at sutures, where osteoblast differentiation and bone deposition occur. Mutations causing skull malformations have revealed some required genes, but many aspects of suture regulation remain poorly understood. We describe a zebrafish mutation in osterix/sp7, which causes a generalized delay in osteoblast maturation. While most of the skeleton is patterned normally, mutants have specific defects in the anterior skull and upper jaw, and the top of the skull comprises a random mosaic of bones derived from individual initiation sites. Osteoblasts at the edges of the bones are highly proliferative and fail to differentiate, consistent with global changes in gene expression. We propose that signals from the bone itself are required for orderly recruitment of precursor cells and growth along the edges. The delay in bone maturation caused by loss of Sp7 leads to unregulated bone formation, revealing a new mechanism for patterning the skull and sutures. PMID:26992365

Automated morphological analysis of bone marrow cells in microscopic images for diagnosis of leukemia: nucleus-plasma separation and cell classification using a hierarchical tree model of hematopoesis

NASA Astrophysics Data System (ADS)

Krappe, Sebastian; Wittenberg, Thomas; Haferlach, Torsten; Münzenmayer, Christian

2016-03-01

The morphological differentiation of bone marrow is fundamental for the diagnosis of leukemia. Currently, the counting and classification of the different types of bone marrow cells is done manually under the use of bright field microscopy. This is a time-consuming, subjective, tedious and error-prone process. Furthermore, repeated examinations of a slide may yield intra- and inter-observer variances. For that reason a computer assisted diagnosis system for bone marrow differentiation is pursued. In this work we focus (a) on a new method for the separation of nucleus and plasma parts and (b) on a knowledge-based hierarchical tree classifier for the differentiation of bone marrow cells in 16 different classes. Classification trees are easily interpretable and understandable and provide a classification together with an explanation. Using classification trees, expert knowledge (i.e. knowledge about similar classes and cell lines in the tree model of hematopoiesis) is integrated in the structure of the tree. The proposed segmentation method is evaluated with more than 10,000 manually segmented cells. For the evaluation of the proposed hierarchical classifier more than 140,000 automatically segmented bone marrow cells are used. Future automated solutions for the morphological analysis of bone marrow smears could potentially apply such an approach for the pre-classification of bone marrow cells and thereby shortening the examination time.

Skeletal stem cell isolation: A review on the state-of-the-art microfluidic label-free sorting techniques.

PubMed

Xavier, Miguel; Oreffo, Richard O C; Morgan, Hywel

2016-01-01

Skeletal stem cells (SSC) are a sub-population of bone marrow stromal cells that reside in postnatal bone marrow with osteogenic, chondrogenic and adipogenic differentiation potential. SSCs reside only in the bone marrow and have organisational and regulatory functions in the bone marrow microenvironment and give rise to the haematopoiesis-supportive stroma. Their differentiation capacity is restricted to skeletal lineages and therefore the term SSC should be clearly distinguished from mesenchymal stem cells which are reported to exist in extra-skeletal tissues and, critically, do not contribute to skeletal development. SSCs are responsible for the unique regeneration capacity of bone and offer unlimited potential for application in bone regenerative therapies. A current unmet challenge is the isolation of homogeneous populations of SSCs, in vitro, with homogeneous regeneration and differentiation capacities. Challenges that limit SSC isolation include a) the scarcity of SSCs in bone marrow aspirates, estimated at between 1 in 10-100,000 mononuclear cells; b) the absence of specific markers and thus the phenotypic ambiguity of the SSC and c) the complexity of bone marrow tissue. Microfluidics provides innovative approaches for cell separation based on bio-physical features of single cells. Here we review the physical principles underlying label-free microfluidic sorting techniques and review their capacity for stem cell selection/sorting from complex (heterogeneous) samples. Copyright © 2016 Elsevier Inc. All rights reserved.

Balancing the Rates of New Bone Formation and Polymer Degradation Enhances Healing of Weight-Bearing Allograft/Polyurethane Composites in Rabbit Femoral Defects

DTIC Science & Technology

2014-10-03

frac tures? J Orthop Res 29, 33, 2011. Epub 2010/07/08. 2. Russell, T.A., and Leighton , R.K. Comparison of autogenous bone graft and endothermic...Boyd, S.K., Christiansen , B.A., Guldberg, R.E., Jepsen, K.J., and Muller, R. Guidelines for assessment of bone microstructure in rodents using micro

PDGFBB promotes PDGFR{alpha}-positive cell migration into artificial bone in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yoshida, Shigeyuki; Center for Human Metabolomic Systems Biology, Keio University School of Medicine, 35 Shinano-machi, Shinjuku-ku, Tokyo 160-8582; Iwasaki, Ryotaro

2012-05-18

Highlights: Black-Right-Pointing-Pointer We examined effects of PDGFBB in PDGFR{alpha} positive cell migration in artificial bones. Black-Right-Pointing-Pointer PDGFBB was not expressed in osteoblastic cells but was expressed in peripheral blood cells. Black-Right-Pointing-Pointer PDGFBB promoted PDGFR{alpha} positive cell migration into artificial bones but not osteoblast proliferation. Black-Right-Pointing-Pointer PDGFBB did not inhibit osteoblastogenesis. -- Abstract: Bone defects caused by traumatic bone loss or tumor dissection are now treated with auto- or allo-bone graft, and also occasionally by artificial bone transplantation, particularly in the case of large bone defects. However, artificial bones often exhibit poor affinity to host bones followed by bony union failure.more » Thus therapies combining artificial bones with growth factors have been sought. Here we report that platelet derived growth factor bb (PDGFBB) promotes a significant increase in migration of PDGF receptor {alpha} (PDGFR{alpha})-positive mesenchymal stem cells/pre-osteoblastic cells into artificial bone in vivo. Growth factors such as transforming growth factor beta (TGF{beta}) and hepatocyte growth factor (HGF) reportedly inhibit osteoblast differentiation; however, PDGFBB did not exhibit such inhibitory effects and in fact stimulated osteoblast differentiation in vitro, suggesting that combining artificial bones with PDGFBB treatment could promote host cell migration into artificial bones without inhibiting osteoblastogenesis.« less

Three-Dimensional Mechanical Loading Modulates the Osteogenic Response of Mesenchymal Stem Cells to Tumor-Derived Soluble Signals.

PubMed

Lynch, Maureen E; Chiou, Aaron E; Lee, Min Joon; Marcott, Stephen C; Polamraju, Praveen V; Lee, Yeonkyung; Fischbach, Claudia

2016-08-01

Dynamic mechanical loading is a strong anabolic signal in the skeleton, increasing osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BM-MSCs) and increasing the bone-forming activity of osteoblasts, but its role in bone metastatic cancer is relatively unknown. In this study, we integrated a hydroxyapatite-containing three-dimensional (3D) scaffold platform with controlled mechanical stimulation to investigate the effects of cyclic compression on the interplay between breast cancer cells and BM-MSCs as it pertains to bone metastasis. BM-MSCs cultured within mineral-containing 3D poly(lactide-co-glycolide) (PLG) scaffolds differentiated into mature osteoblasts, and exposure to tumor-derived soluble factors promoted this process. When BM-MSCs undergoing osteogenic differentiation were exposed to conditioned media collected from mechanically loaded breast cancer cells, their gene expression of osteopontin was increased. This was further enhanced when mechanical compression was simultaneously applied to BM-MSCs, leading to more uniformly deposited osteopontin within scaffold pores. These results suggest that mechanical loading of 3D scaffold-based culture models may be utilized to evaluate the role of physiologically relevant physical cues on bone metastatic breast cancer. Furthermore, our data imply that cyclic mechanical stimuli within the bone microenvironment modulate interactions between tumor cells and BM-MSCs that are relevant to bone metastasis.

Selective differentiation and proliferation of hematopoietic cells induced by recombinant human interleukins.

PubMed Central

Saito, H; Hatake, K; Dvorak, A M; Leiferman, K M; Donnenberg, A D; Arai, N; Ishizaka, K; Ishizaka, T

1988-01-01

Effects of recombinant human interleukins on hematopoiesis were explored by using suspension cultures of mononuclear cells of human umbilical-cord blood and bone marrow. The results showed that interleukin 5 induced the selective differentiation and proliferation of eosinophils. After 3 weeks in culture with interleukin 5, essentially all nonadherent cells in both bone marrow and cord blood cell cultures became eosinophilic myelocytes. Culture of the same cells with interleukin 4 resulted in the selective growth of OKT3+ lymphocytes. However, OKT3+ cells did not develop if the bone marrow cells were depleted of OKT3+/OKT11+ cells prior to the culture, indicating that interleukin 4 induced the proliferation of a subpopulation of resting T cells present in cord blood and bone marrow cell preparations. In suspension cultures of bone marrow cells and cord blood cells grown in the presence of interleukin 3, basophilic, eosinophilic, and neutrophilic myelocytes and macrophages developed within 2 weeks. By 3 weeks, however, the majority of nonadherent cells became eosinophilic myelocytes. In contrast to mouse bone marrow cell cultures, neither interleukin 3 nor a combination of interleukins 3 and 4 induced the differentiation of mast cells in human bone marrow or cord blood cell cultures. Images PMID:3258425

MicroRNA Functions in Osteogenesis and Dysfunctions in Osteoporosis

PubMed Central

van Wijnen, Andre J.; van de Peppel, Jeroen; van Leeuwen, Johannes P.; Lian, Jane B.; Stein, Gary S.; Westendorf, Jennifer J.; Oursler, Merry-Jo; Sampen, Hee-Jeong Im; Taipaleenmaki, Hanna; Hesse, Eric; Riester, Scott; Kakar, Sanjeev

2013-01-01

MicroRNAs (miRNAs) are critical post-transcriptional regulators of gene expression that control osteoblast mediated bone formation and osteoclast-related bone remodelling. Deregulation of miRNA mediated mechanisms is emerging as an important pathological factor in bone degeneration (e.g., osteoporosis) and other bone-related diseases. MiRNAs are intriguing regulatory molecules that are networked with cell signaling pathways and intricate transcriptional programs through ingenuous circuits with remarkably simple logic. This overview examines key principles by which miRNAs control differentiation of osteoblasts as they evolve from mesenchymal stromal cells during osteogenesis, or of osteoclasts as they originate from monocytic precursors in the hematopoietic lineage during osteoclastogenesis. Of particular note are miRNAs that are temporally up-regulated during osteoblastogenesis (e.g., miR-218) or osteoclastogenesis (e.g., miR-148a). Each miRNA stimulates differentiation by suppressing inhibitory signalling pathways (‘double-negative’ regulation). The excitement surrounding miRNAs in bone biology stems from the prominent effects that individual miRNAs can have on biological transitions during differentiation of skeletal cells and correlations of miRNA dysfunction with bone diseases. MiRNAs have significant clinical potential which is reflected by their versatility as disease-specific biomarkers and their promise as therapeutic agents to ameliorate or reverse bone tissue degeneration. PMID:23605904

[Current status of bone/cartilage tissue engineering towards clinical applications].

PubMed

Ohgushi, Hajime

2014-10-01

Osteo/chondrogenic differentiation capabilities are seen after in vivo implantation of mesenchymal stem cells (MSCs), which are currently used for the patients having bone/cartilage defects. Importantly, the differentiation capabilities are induced by culturing technology, resulting in in vitro bone/cartilage formation. Especially, the in vitro bone tissue is useful for bone tissue regeneration. For cartilage regeneration, culture expanded chondrocytes derived from patient's normal cartilage are also used for the patients having cartilage damages. Recently, the cultured chondrocytes embedded in atelocollagen gel are obtainable as tissue engineered products distributed by Japan Tissue Engineering Co. Ltd. The products are available in the well-regulated hospitals by qualified orthopedic surgeons. The criteria for these hospitals/surgeons have been established. This review paper focuses on current status of bone/cartilage tissue engineering towards clinical applications in Japan.

Cigarette Smoke Inhibits Recruitment of Bone-Marrow-Derived Stem cells to The Uterus

PubMed Central

Zhou, Yuping; Gan, Ye; Taylor, Hugh S.

2011-01-01

Cigarette smoking leads to female infertility and a decreased incidence of endometriosis. Bone marrow derived stem cells are recruited to uterine endometrium and endometriosis. The effect of cigarette smoking on stem cell recruitment to any organ is uncharacterized. We hypothesized that bone marrow-derived mesenchymal stem cell recruitment to the uterus and differentiation would be diminished by cigarette smoke. We used human mesenchymal stem cells (hMSC) in vitro and a mouse model of cigarette smoke exposure. After myeloablation female C57BL/6J received bone marrow cells from males. Mice were exposed to room air or smoke from unfiltered cigarettes. Immunofluorescence and Y-FISH was performed on uterine sections. In vitro hMSCs were treated with 8-Br-cAMP to induce endometrial cell differentiation with or without cigarette smoke extract (CSE) and decidualization assessed morphologically and by prolactin expression. After 4 weeks the total number of Y-chromosome cells in the uterus was reduced by 68% in the smoke exposed mice. Both leukocytes and bone marrow derived endometrial cells were reduced by 60% and 73%, respectively. Differentiation of bone marrow derived cell to endometrial epithelial cells was reduced by 84%. hMSC treated with CSE failed to show cytological characteristics of decidualization. mRNA levels of the decidualization marker prolactin were decreased by 90% in CSE treated cells. Smoking inhibits both recruitment of bone marrow derived stem cells to uterus and stem cell differentiation. Inhibition of stem cells recruitment may be a general mechanism by which smoking leads to long term organ damage through inability to repair or regenerate multiple tissues. PMID:20955787

Differentiating osteomyelitis from bone infarction in sickle cell disease.

PubMed

Wong, A L; Sakamoto, K M; Johnson, E E

2001-02-01

This brief review discusses one possible approach to evaluating the sickle cell patient with bone pain. The major differential diagnoses include osteomyelitis and bone infarction. Based on previous studies, we provide an approach to assessing and treating patients with the possible diagnosis of osteomyelitis. An algorithm has been provided, which emphasizes the importance of the initial history and physical examination. Specific radiographic studies are recommended to aid in making the initial assessment and to determine whether the patient has an infarct or osteomyelitis. Differentiating osteomyelitis from infarction in sickle cell patients remains a challenge for the pediatrician. This algorithm can be used as a guide for physicians who evaluate such patients in the acute care setting.

Study protocol for prevention of falls: A randomized controlled trial of effects of vitamin D and exercise on falls prevention

PubMed Central

2012-01-01

Background Falls are the leading cause of unintentional injury and injury-related death among older people. In addition to physical activity, vitamin D also may affect balance and neuromuscular function. Low serum 25-hydroksivitamin D level increases the risk of bone loss, falls and fractures. Thus, an appropriate exercise program and sufficient vitamin D intake may significantly improve not only functional balance, but also balance confidence. Balance represents a complex motor skill determined by reaction time, muscle strength, and speed and coordination of movement. Methods/Design A 2-year randomized double-blind placebo-controlled vitamin D and open exercise trial of 409 home-dwelling women 70 to 80 years of age comprising four study arms: 1) exercise + vitamin D (800 IU/d), 2) exercise + placebo, 3) no exercise + vitamin D (800 IU/d), 4) no exercise + placebo. In addition to monthly fall diaries, general health status, life style, bone health, physical functioning, and vitamin D metabolism will be assessed. The primary outcomes are the rate of falls and fall-related injuries. Secondary outcomes include changes in neuromuscular functioning (e.g. body balance, muscle strength), ADL- and mobility functions, bone density and structure, cardiovascular risk factors, quality of life and fear of falling. Discussion The successful completion of this trial will provide evidence on the effectiveness of exercise and vitamin D for falls reduction. Trial Registration ClinicalTrial.gov -register (NCT00986466). PMID:22448872

Secondary osteoporosis.

PubMed

Gennari, C; Martini, G; Nuti, R

1998-06-01

Generalized osteoporosis currently represents a heterogeneous group of conditions with many different causes and pathogenetic mechanisms, that often are variably associated. The term "secondary" is applied to all patients with osteoporosis in whom the identifiable causal factors are other than menopause and aging. In this heterogeneous group of conditions, produced by many different pathogenetic mechanisms, a negative bone balance may be variably associated with low, normal or increased bone remodeling states. A consistent group of secondary osteoporosis is related to endocrinological or iatrogenic causes. Exogenous hypercortisolism may be considered an important risk factor for secondary osteoporosis in the community, and probably glucocorticoid-induced osteoporosis is the most common type of secondary osteoporosis. Supraphysiological doses of corticosteroids cause two abnormalities in bone metabolism: a relative increase in bone resorption, and a relative reduction in bone formation. Bone loss, mostly of trabecular bone, with its resultant fractures is the most incapacitating consequence of osteoporosis. The estimated incidence of fractures in patients prescribed corticosteroid is 30% to 50%. Osteoporosis is considered one of the potentially serious side effects of heparin therapy. The occurrence of heparin-induced osteoporosis appeared to be strictly related to the length of treatment (over 4-5 months), and the dosage (15,000 U or more daily), but the pathogenesis is poorly understood. It has been suggested that heparin could cause an increase in bone resorption by increasing the number of differentiated osteoclasts, and by enhancing the activity of individual osteoclasts. Hyperthyroidism is frequently associated with loss of trabecular and cortical bone; the enhanced bone turnover that develops in thyrotoxicosis is characterized by an increase in the number of osteoclasts and resorption sites, and an increase in the ratio of resorptive to formative bone surfaces, with the net result of bone loss. Despite these findings, the occurrence of pathological fractures in patients with hyperthyroidism is relatively low, and probably due to the fact that deficiencies in bone mass may be reversed by treatment of the thyroid disease. Most, but not all, studies on insulin-dependent diabetes mellitus (IDDM) report an association with osteopenia. In IDDM, the extent of bone loss is usually slight, which helps explain the discrepancy between the frequency of decreased bone mineral density, and the frequency of osteoporotic fractures in long-standing diabetes. Contradictory results have been obtained in non-insulin-dependent diabetes mellitus (NIDDM) patients. Increased rates of bone loss at the radius and lumbar spine were demonstrated either in patients with two-thirds gastric resection and Billroth II reconstruction, or in those with one-third resection and Billroth I anastomosis, and the metabolic bone disease following gastrectomy may consist also of osteomalacia or mixed pattern of osteoporosis-osteomalacia, with secondary hyperparathyroidism. Miscellaneous causes of secondary osteoporosis are also immobilization, pregnancy and lactation, and alcohol abuse.

Safflower bud inhibits RANKL-induced osteoclast differentiation and prevents bone loss in ovariectomized mice.

PubMed

Choi, Joo-Hee; Lim, Seul-Ki; Kim, Dong-Il; Park, Min-Jung; Kim, Young-Kuk; Lee, An-Chul; Kim, Young-Min; Yang, Soo-Jin; Park, Jong-Hwan

2017-10-15

The powder and extract of safflower seeds are known to be effective in the prevention of bone loss in ovariectomized animals. However, the inhibitory effect and molecular mechanisms of safflower bud (SB), the germinated safflower, on bone destruction is unclear. The present study was designed to investigate the inhibitory effect and molecular mechanism of SB on osteoclastic differentiation and on bone loss in ovarietomized (OVX) mice. Osteoclastogenesis was determined by TRAP staining, F-actin ring formation, and bone resorption assay. NF-κB and MAPKs activation was analyzed by transfection assay and Western blot, respectively. Real-time PCR was performed to examine the expression of osteoclastogenesis-related genes. Histological changes, increases in TRAP-positive cells, and cathepsin K expression were examined in the metaphysis of OVX mice. Density of bone marrow was evaluated by µCT. SB inhibited the RANKL-induced differentiation of BMDMs into osteoclasts in a dose-dependent manner. F-actin ring formation and bone resorption were also reduced by SB in RANKL-treated BMDMs. In addition, SB decreased the activation of NF-κB and MAPKs and the expression of osteoclastogenesis-related genes in BMDMs treated with RANKL. Feeding of SB-included diet prevented bone loss in OVX mice. The number of TRAP-positive cells and level of protein expression of cathepsin K was reduced and bone mineral density was increased in the metaphysis of mice fed SB compared with OVX mice. These findings suggest that SB can be a preventive and therapeutic candidate for destructive bone diseases. Copyright © 2017. Published by Elsevier GmbH.

N-AC-l-Leu-PEI-mediated miR-34a delivery improves osteogenic differentiation under orthodontic force.

PubMed

Yu, Wenwen; Zheng, Yi; Yang, Zhujun; Fei, Hongbo; Wang, Yang; Hou, Xu; Sun, Xinhua; Shen, Yuqin

2017-12-15

Rare therapeutic genes or agents are reported to control orthodontic bone remodeling. MicroRNAs have recently been associated with bone metabolism. Here, we report the in vitro and in vivo effects of miR-34a on osteogenic differentiation under orthodontic force using an N -acetyl-L-leucine-modified polyethylenimine ( N -Ac-l-Leu-PEI) carrier. N -Ac-l-Leu-PEI exhibited low cytotoxicity and high miR-34a transfection efficiency in rat bone mineral stem cells and local alveolar bone tissue. After transfection, miR-34a enhanced the osteogenic differentiation of Runx2 and ColI , Runx2 and ColI protein levels, and early osteogenesis function under orthodontic strain in vitro . MiR-34a also enhanced alveolar bone remodeling under orthodontic force in vivo , as evidenced by elevated gene and protein expression, upregulated indices of alveolar bone anabolism, and diminished tooth movement. We determined that the mechanism miR-34a in osteogenesis under orthodontic force may be associated with GSK-3β. These results suggested that miR-34a delivered by N -Ac-l-Leu-PEI could be a potential therapeutic target for orthodontic treatment.

The Nell-1 Growth Factor Stimulates Bone Formation by Purified Human Perivascular Cells

PubMed Central

Zhang, Xinli; Péault, Bruno; Chen, Weiwei; Li, Weiming; Corselli, Mirko; James, Aaron W.; Lee, Min; Siu, Ronald K.; Shen, Pang; Zheng, Zhong; Shen, Jia; Kwak, Jinny; Zara, Janette N.; Chen, Feng; Zhang, Hong; Yin, Zack; Wu, Ben; Ting, Kang

2011-01-01

The search for novel sources of stem cells other than bone marrow mesenchymal stem cells (MSCs) for bone regeneration and repair has been a critical endeavor. We previously established an effective protocol to homogeneously purify human pericytes from multiple fetal and adult tissues, including adipose, bone marrow, skeletal muscle, and pancreas, and identified pericytes as a primitive origin of human MSCs. In the present study, we further characterized the osteogenic potential of purified human pericytes combined with a novel osteoinductive growth factor, Nell-1. Purified pericytes grown on either standard culture ware or human cancellous bone chip (hCBC) scaffolds exhibited robust osteogenic differentiation in vitro. Using a nude mouse muscle pouch model, pericytes formed significant new bone in vivo as compared to scaffold alone (hCBC). Moreover, Nell-1 significantly increased pericyte osteogenic differentiation, both in vitro and in vivo. Interestingly, Nell-1 significantly induced pericyte proliferation and was observed to have pro-angiogenic effects, both in vitro and in vivo. These studies suggest that pericytes are a potential new cell source for future efforts in skeletal regenerative medicine, and that Nell-1 is a candidate growth factor able to induce pericyte osteogenic differentiation. PMID:21615216

Cell-Autonomous Control of IL-7 Response Revealed In a Novel Stage of Precursor B Cells

PubMed Central

Sandoval, Gabriel J.; Graham, Daniel B.; Bhattacharya, Deepta; Sleckman, Barry P.; Xavier, Ramnik J.; Swat, Wojciech

2013-01-01

During early stages of B-lineage differentiation in bone marrow, signals emanating from IL-7 receptor and pre-B cell receptor (pre-BCR) are thought to synergistically induce proliferative expansion of progenitor cells. Paradoxically, loss of pre-BCR signaling components is associated with leukemia in both mice and humans. Exactly how progenitor B cells perform the task of balancing proliferative burst dependent on IL-7 with the termination of IL-7 signals and the initiation of LC gene rearrangement remains to be elucidated. In this report, we provide genetic and functional evidence that the cessation of IL-7 response of pre-B cells is controlled via a cell-autonomous mechanism that operates at a discreet developmental transition inside Fraction C’ (Large Pre-BII) marked by transient expression of c-Myc. Our data indicates that pre-BCR cooperates with IL-7R in expanding pre-B cell pool, but it is also critical to control differentiation program shutting off c-Myc gene in large pre-B cells. PMID:23420891

Feeding soy protein isolate and oils rich in omega-3 polyunsaturated fatty acids affected mineral balance, but not bone in a rat model of autosomal recessive polycystic kidney disease.

PubMed

Maditz, Kaitlin H; Smith, Brenda J; Miller, Matthew; Oldaker, Chris; Tou, Janet C

2015-02-10

Polycystic kidney disease (PKD), a genetic disorder characterized by multiple cysts and renal failure at an early age. In children, kidney disease is often accompanied by disordered mineral metabolism, failure to achieve peak bone mass, and reduced adult height. Optimizing bone health during the growth stage may preserve against bone loss associated with early renal dysfunction in PKD. Dietary soy protein and omega-3 polyunsaturated fatty acid (n-3 PUFA) have been reported to ameliorate PKD and to promote bone health. The study objective was to determine the bone effects of feeding soy protein and/or n-3 PUFAs in a rat model of PKD. Weanling female PCK rats (n = 12/group) were randomly assigned to casein + corn oil (Casein + CO), casein + soybean oil (Casein + SO), soy protein isolate + soybean oil (SPI + SO) or soy protein isolate + 1:1 soybean oil:salmon oil blend (SPI + SB) for 12 weeks. Rats fed SPI + SO diet had shorter (P = 0.001) femur length than casein-fed rats. Rats fed SPI + SO and SPI + SB diet had higher (P = 0.04) calcium (Ca) and phosphorus (P) retention. However, there were no significant differences in femur and tibial Ca, P or bone mass between diet groups. There were also no significant difference in bone microarchitecture measured by micro-computed tomography or bone strength determined by three-point bending test between diet groups. Early diet management of PKD using SPI and/or n-3 PUFAs influenced bone longitudinal growth and mineral balance, but neither worsened nor enhanced bone mineralization, microarchitecture or strength.

Enhanced adipogenic differentiation of bovine bone marrow-derived mesenchymal stem cells

USDA-ARS?s Scientific Manuscript database

Until now, the isolation and characterization of bovine bone marrow-derived mesenchymal stem cells (bBM-MSCs) have not been established, which prompted us to optimize the differentiation protocol for bBM-MSCs. In this study, bBM-MSCs were freshly isolated from three 6-month-old cattle and used for p...

Infection of chicken bone marrow mononuclear cells with subgroup J avian leukosis virus inhibits dendritic cell differentiation and alters cytokine expression.

PubMed

Liu, Di; Qiu, Qianqian; Zhang, Xu; Dai, Manman; Qin, Jianru; Hao, Jianjong; Liao, Ming; Cao, Weisheng

2016-10-01

Subgroup J avian leukosis virus (ALV-J) is an oncogenic retrovirus known to induce tumor formation and immunosuppression in infected chickens. One of the organs susceptible to ALV-J is the bone marrow, from which specialized antigen-presenting cells named dendritic cells (BM-DCs) are derived. Notably, these cells possess the unique ability to induce primary immune responses. In the present study, a method of cultivating and purifying DCs from chicken bone marrow in vitro was established to investigate the effects of ALV-J infection on BM-DC differentiation or generation. The results indicated that ALV-J not only infects the chicken bone marrow mononuclear cells but also appears to inhibit the differentiation and maturation of BM-DCs and to trigger apoptosis. Moreover, substantial reductions in the mRNA expression of TLR1, TLR2, TLR3, MHCI, and MHCII and in cytokine production were detected in the surviving BM-DCs following ALV-J infection. These findings indicate that ALV-J infection disrupts the process of bone marrow mononuclear cell differentiation into BM-DCs likely via altered antigen presentation, resulting in a downstream immune response in affected chickens. Copyright © 2016 Elsevier B.V. All rights reserved.

Regulation of bone formation by osteoclasts involves Wnt/BMP signaling and the chemokine sphingosine-1-phosphate

PubMed Central

Pederson, Larry; Ruan, Ming; Westendorf, Jennifer J.; Khosla, Sundeep; Oursler, Merry Jo

2008-01-01

Under most conditions, resorbed bone is nearly precisely replaced in location and amount by new bone. Thus, it has long been recognized that bone loss through osteoclast-mediated bone resorption and bone replacement through osteoblast-mediated bone formation are tightly coupled processes. Abundant data conclusively demonstrate that osteoblasts direct osteoclast differentiation. Key questions remain, however, as to how osteoblasts are recruited to the resorption site and how the amount of bone produced is so precisely controlled. We hypothesized that osteoclasts play a crucial role in the promotion of bone formation. We found that osteoclast conditioned medium stimulates human mesenchymal stem (hMS) cell migration and differentiation toward the osteoblast lineage as measured by mineralized nodule formation in vitro. We identified candidate osteoclast-derived coupling factors using the Affymetrix microarray. We observed significant induction of sphingosine kinase 1 (SPHK1), which catalyzes the phosphorylation of sphingosine to form sphingosine 1-phosphate (S1P), in mature multinucleated osteoclasts as compared with preosteoclasts. S1P induces osteoblast precursor recruitment and promotes mature cell survival. Wnt10b and BMP6 also were significantly increased in mature osteoclasts, whereas sclerostin levels decreased during differentiation. Stimulation of hMS cell nodule formation by osteoclast conditioned media was attenuated by the Wnt antagonist Dkk1, a BMP6-neutralizing antibody, and by a S1P antagonist. BMP6 antibodies and the S1P antagonist, but not Dkk1, reduced osteoclast conditioned media-induced hMS chemokinesis. In summary, our findings indicate that osteoclasts may recruit osteoprogenitors to the site of bone remodeling through SIP and BMP6 and stimulate bone formation through increased activation of Wnt/BMP pathways. PMID:19075223

High levels of β-catenin signaling reduce osteogenic differentiation of stem cells in inflammatory microenvironments through inhibition of the noncanonical Wnt pathway.

PubMed

Liu, Na; Shi, Songtao; Deng, Manjing; Tang, Liang; Zhang, Guangjing; Liu, Ning; Ding, Bofu; Liu, Wenjia; Liu, Yali; Shi, Haigang; Liu, Luchuan; Jin, Yan

2011-09-01

Periodontal ligament stem cells (PDLSCs), a new population of mesenchymal stem cells (MSCs), have been isolated from the periodontal ligament (PDL). The capacity of multipotency and self-renewal makes them an excellent cell source for bone regeneration and repair. However, their bone-regeneration ability could be awakened in inflammatory microenvironments, which may be the result of changes in their differentiation potential. Recently, genetic evidences has shown that the Wnt pathway plays an important role in bone homeostasis. In this study we have determined the specific role of β-catenin in osteogenic differentiation of PDLSCs obtained from inflammatory microenvironments (P-PDLSCs). The inflammatory microenvironment, while inhibiting osteogenic differentiation potential, promotes proliferation of MSCs. A higher the level of β-catenin in P-PDLSCs than in H-PDLSCs (PDLSCs obtained from a healthy microenvironment) resulted in the same disparity in canonical Wnt signaling pathway activation between each cell type. Here we show that activation of β-catenin suppresses the noncanonical Wnt/Ca(2+) pathway, leading to increased proliferation but reduced osteogenic differentiation of P-PDLSCs. Downregulation of the levels of β-catenin by treatment with dickkopf-1 (DKK-1) leads to activation of the noncanonical Wnt/Ca(2+) pathway, which, in turn, results in the promotion of osteogenic differentiation in P-PDLSCs. Interestingly, β-catenin can affect both the canonical Wnt/β-catenin pathway and the noncanonical Wnt/Ca(2+) pathway. Our data indicate that β-catenin plays a central role in regulating osteogenic differentiation of MSCs in inflammatory microenvironments. Given the important role of Wnt signaling in osteogenic differentiation, it is possible that agents that can modify this pathway may be of value in bone regeneration by MSCs in chronic inflammatory microenvironments. Copyright © 2011 American Society for Bone and Mineral Research.

Comparative Analysis of Mouse-Induced Pluripotent Stem Cells and Mesenchymal Stem Cells During Osteogenic Differentiation In Vitro

PubMed Central

Kayashima, Hiroki; Miura, Jiro; Uraguchi, Shinya; Wang, Fangfang; Okawa, Hiroko; Sasaki, Jun-Ichi; Saeki, Makio; Matsumoto, Takuya; Yatani, Hirofumi

2014-01-01

Induced pluripotent stem cells (iPSCs) can differentiate into mineralizing cells and are, therefore, expected to be useful for bone regenerative medicine; however, the characteristics of iPSC-derived osteogenic cells remain unclear. Here, we provide a direct in vitro comparison of the osteogenic differentiation process in mesenchymal stem cells (MSCs) and iPSCs from adult C57BL/6J mice. After 30 days of culture in osteogenic medium, both MSCs and iPSCs produced robustly mineralized bone nodules that contained abundant calcium phosphate with hydroxyapatite crystal formation. Mineral deposition was significantly higher in iPSC cultures than in MSC cultures. Scanning electron microscopy revealed budding matrix vesicles in early osteogenic iPSCs; subsequently, the vesicles propagated to exhibit robust mineralization without rich fibrous structures. Early osteogenic MSCs showed deposition of many matrix vesicles in abundant collagen fibrils that became solid mineralized structures. Both cell types demonstrated increased expression of osteogenic marker genes, such as runx2, osterix, dlx5, bone sialoprotein (BSP), and osteocalcin, during osteogenesis; however, real-time reverse transcription–polymerase chain reaction array analysis revealed that osteogenesis-related genes encoding mineralization-associated molecules, bone morphogenetic proteins, and extracellular matrix collagens were differentially expressed between iPSCs and MSCs. These data suggest that iPSCs are capable of differentiation into mature osteoblasts whose associated hydroxyapatite has a crystal structure similar to that of MSC-associated hydroxyapatite; however, the transcriptional differences between iPSCs and MSCs could result in differences in the mineral and matrix environments of the bone nodules. Determining the biological mechanisms underlying cell-specific differences in mineralization during in vitro iPSC osteogenesis may facilitate the development of clinically effective engineered bone. PMID:24625139

Validation of osteogenic properties of Cytochalasin D by high-resolution RNA-sequencing in mesenchymal stem cells derived from bone marrow and adipose tissues.

PubMed

Samsonraj, Rebekah; Paradise, Christopher R; Dudakovic, Amel; Sen, Buer; Nair, Asha A; Dietz, Allan B; Deyle, David R; Cool, Simon M; Rubin, Janet; van Wijnen, Andre

2018-06-08

Differentiation of mesenchymal stromal/stem cells (MSCs) involves a series of molecular signals and gene transcription events required for attaining cell lineage commitment. Modulation of the actin cytoskeleton using cytochalasin D (CytoD) drives osteogenesis at early time points in bone marrow-derived MSCs, and also initiates a robust osteogenic differentiation program in adipose-derived MSCs. To understand the molecular basis for these pronounced effects on osteogenic differentiation, we investigated global changes in gene expression in CytoD-treated murine and human MSCs by high-resolution RNA-sequencing (RNA-seq) analysis. A three-way bioinformatic comparison between human adipose-derived, human bone marrow-derived and mouse bone marrow-derived MSCs revealed significant upregulation of genes linked to extracellular matrix organization, cell adhesion and bone metabolism. As anticipated, the activation of these differentiation related genes is accompanied by a downregulation of nuclear and cell cycle-related genes presumably reflecting cytostatic effects of CytoD. We also identified eight novel CytoD activated genes - VGLL4, ARHGAP24, KLHL24, RCBTB2, BDH2, SCARF2, ACAD10, HEPH - which are commonly upregulated across the two species and tissue sources of our MSC samples. We selected the Hippo-pathway related VGLL4 gene, which encodes the transcriptional co-factor Vestigial-like 4, for further study because this pathway is linked to osteogenesis. VGLL4 siRNA depletion reduces mineralization of adipose-derived MSCs during CytoD-induced osteogenic differentiation. Together, our RNA-seq analyses suggest that while the stimulatory effects of CytoD on osteogenesis are pleiotropic and depend on the biological state of the cell type, a small group of genes including VGLL4 may contribute to MSC commitment towards the bone lineage.

Reduced graphene oxide-coated hydroxyapatite composites stimulate spontaneous osteogenic differentiation of human mesenchymal stem cells

NASA Astrophysics Data System (ADS)

Lee, Jong Ho; Shin, Yong Cheol; Jin, Oh Seong; Kang, Seok Hee; Hwang, Yu-Shik; Park, Jong-Chul; Hong, Suck Won; Han, Dong-Wook

2015-07-01

Human mesenchymal stem cells (hMSCs) have great potential as cell sources for bone tissue engineering and regeneration, but the control and induction of their specific differentiation into bone cells remain challenging. Graphene-based nanomaterials are considered attractive candidates for biomedical applications such as scaffolds in tissue engineering, substrates for SC differentiation and components of implantable devices, due to their biocompatible and bioactive properties. Despite the potential biomedical applications of graphene and its derivatives, only limited information is available regarding their osteogenic activity. This study concentrates upon the effects of reduced graphene oxide (rGO)-coated hydroxyapatite (HAp) composites on osteogenic differentiation of hMSCs. The average particle sizes of HAp and rGO were 1270 +/- 476 nm and 438 +/- 180 nm, respectively. When coated on HAp particulates, rGO synergistically enhanced spontaneous osteogenic differentiation of hMSCs, without hampering their proliferation. This result was confirmed by determining alkaline phosphatase activity and mineralization of calcium and phosphate as early and late stage markers of osteogenic differentiation. It is suggested that rGO-coated HAp composites can be effectively utilized as dental and orthopedic bone fillers since these graphene-based particulate materials have potent effects on stimulating the spontaneous differentiation of MSCs and show superior bioactivity and osteoinductive potential.Human mesenchymal stem cells (hMSCs) have great potential as cell sources for bone tissue engineering and regeneration, but the control and induction of their specific differentiation into bone cells remain challenging. Graphene-based nanomaterials are considered attractive candidates for biomedical applications such as scaffolds in tissue engineering, substrates for SC differentiation and components of implantable devices, due to their biocompatible and bioactive properties. Despite the potential biomedical applications of graphene and its derivatives, only limited information is available regarding their osteogenic activity. This study concentrates upon the effects of reduced graphene oxide (rGO)-coated hydroxyapatite (HAp) composites on osteogenic differentiation of hMSCs. The average particle sizes of HAp and rGO were 1270 +/- 476 nm and 438 +/- 180 nm, respectively. When coated on HAp particulates, rGO synergistically enhanced spontaneous osteogenic differentiation of hMSCs, without hampering their proliferation. This result was confirmed by determining alkaline phosphatase activity and mineralization of calcium and phosphate as early and late stage markers of osteogenic differentiation. It is suggested that rGO-coated HAp composites can be effectively utilized as dental and orthopedic bone fillers since these graphene-based particulate materials have potent effects on stimulating the spontaneous differentiation of MSCs and show superior bioactivity and osteoinductive potential. Electronic supplementary information (ESI) available: Additional figures. See DOI: 10.1039/c5nr01580d

Development of a rapid culture method to induce adipocyte differentiation of human bone marrow-derived mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ninomiya, Yuichi; Sugahara-Yamashita, Yzumi; Nakachi, Yutaka

2010-04-02

Human mesenchymal stem cells (hMSCs) derived from bone marrow are multipotent stem cells that can regenerate mesenchymal tissues such as adipose, bone or muscle. It is thought that hMSCs can be utilized as a cell resource for tissue engineering and as human models to study cell differentiation mechanisms, such as adipogenesis, osteoblastogenesis and so on. Since it takes 2-3 weeks for hMSCs to differentiate into adipocytes using conventional culture methods, the development of methods to induce faster differentiation into adipocytes is required. In this study we optimized the culture conditions for adipocyte induction to achieve a shorter cultivation time formore » the induction of adipocyte differentiation in bone marrow-derived hMSCs. Briefly, we used a cocktail of dexamethasone, insulin, methylisobutylxanthine (DIM) plus a peroxisome proliferator-activated receptor {gamma} agonist, rosiglitazone (DIMRo) as a new adipogenic differentiation medium. We successfully shortened the period of cultivation to 7-8 days from 2-3 weeks. We also found that rosiglitazone alone was unable to induce adipocyte differentiation from hMSCs in vitro. However, rosiglitazone appears to enhance hMSC adipogenesis in the presence of other hormones and/or compounds, such as DIM. Furthermore, the inhibitory activity of TGF-{beta}1 on adipogenesis could be investigated using DIMRo-treated hMSCs. We conclude that our rapid new culture method is very useful in measuring the effect of molecules that affect adipogenesis in hMSCs.« less

Optimisation of the differing conditions required for bone formation in vitro by primary osteoblasts from mice and rats

PubMed Central

ORRISS, ISABEL R.; HAJJAWI, MARK O.R.; HUESA, CARMEN; MACRAE, VICKY E.; ARNETT, TIMOTHY R.

2014-01-01

The in vitro culture of calvarial osteoblasts from neonatal rodents remains an important method for studying the regulation of bone formation. The widespread use of transgenic mice has created a particular need for a reliable, simple method that allows the differentiation and bone-forming activity of murine osteoblasts to be studied. In the present study, we established such a method and identified key differences in optimal culture conditions between mouse and rat osteoblasts. Cells isolated from neonatal rodent calvariae by collagenase digestion were cultured for 14–28 days before staining for tissue non-specific alkaline phosphatase (TNAP) and bone mineralisation (alizarin red). The reliable differentiation of mouse osteoblasts, resulting in abundant TNAP expression and the formation of mineralised ‘trabecular-shaped’ bone nodules, occurred only following culture in α minimum essential medium (αMEM) and took 21–28 days. Dexamethasone (10 nM) inhibited bone mineralisation in the mouse osteoblasts. By contrast, TNAP expression and bone formation by rat osteoblasts were observed following culture in both αMEM and Dulbecco’s modified Eagle’s medium (DMEM) after approximately 14 days (although ~3-fold more effectively in αMEM) and was strongly dependent on dexamethasone. Both the mouse and rat osteoblasts required ascorbate (50 μg/ml) for osteogenic differentiation and β-glycerophosphate (2 mM) for mineralisation. The rat and mouse osteoblasts showed similar sensitivity to the well-established inhibitors of mineralisation, inorganic pyrophosphate (PPi) and adenosine triphosphate (ATP; 1–100 μM). The high efficiency of osteogenic differentiation observed following culture in αMEM, compared with culture in DMEM possibly reflects the richer formulation of the former. These findings offer a reliable technique for inducing mouse osteoblasts to form bone in vitro and a more effective method for culturing bone-forming rat osteoblasts. PMID:25200658

High Salt Diets, Bone Strength and Mineral Content of Mature Femur After Skeletal Unloading

NASA Technical Reports Server (NTRS)

Liang, Michael T. C.

1998-01-01

It is known that high salt diets increase urinary calcium (Ca) loss, but it is not known whether this effect weakens bone during space flight. The Bone Hormone Lab has studied the effect of high salt diets on Ca balance and whole body Ca in a space flight model (2,8). Neither the strength nor mineral content of the femurs from these studies has been evaluated. The purpose of this study was to determine the effect of high salt diets (HiNa) and skeletal unloading on femoral bone strength and bone mineral content (BMC) in mature rats.

Osteogenic differentiation of human mesenchymal stem cells in mineralized alginate matrices.

PubMed

Westhrin, Marita; Xie, Minli; Olderøy, Magnus Ø; Sikorski, Pawel; Strand, Berit L; Standal, Therese

2015-01-01

Mineralized biomaterials are promising for use in bone tissue engineering. Culturing osteogenic cells in such materials will potentially generate biological bone grafts that may even further augment bone healing. Here, we studied osteogenic differentiation of human mesenchymal stem cells (MSC) in an alginate hydrogel system where the cells were co-immobilized with alkaline phosphatase (ALP) for gradual mineralization of the microenvironment. MSC were embedded in unmodified alginate beads and alginate beads mineralized with ALP to generate a polymer/hydroxyapatite scaffold mimicking the composition of bone. The initial scaffold mineralization induced further mineralization of the beads with nanosized particles, and scanning electron micrographs demonstrated presence of collagen in the mineralized and unmineralized alginate beads cultured in osteogenic medium. Cells in both types of beads sustained high viability and metabolic activity for the duration of the study (21 days) as evaluated by live/dead staining and alamar blue assay. MSC in beads induced to differentiate in osteogenic direction expressed higher mRNA levels of osteoblast-specific genes (RUNX2, COL1AI, SP7, BGLAP) than MSC in traditional cell cultures. Furthermore, cells differentiated in beads expressed both sclerostin (SOST) and dental matrix protein-1 (DMP1), markers for late osteoblasts/osteocytes. In conclusion, Both ALP-modified and unmodified alginate beads provide an environment that enhance osteogenic differentiation compared with traditional 2D culture. Also, the ALP-modified alginate beads showed profound mineralization and thus have the potential to serve as a bone substitute in tissue engineering.

Osteogenic Differentiation of Human Mesenchymal Stem Cells in Mineralized Alginate Matrices

PubMed Central

Westhrin, Marita; Xie, Minli; Olderøy, Magnus Ø.; Sikorski, Pawel

2015-01-01

Mineralized biomaterials are promising for use in bone tissue engineering. Culturing osteogenic cells in such materials will potentially generate biological bone grafts that may even further augment bone healing. Here, we studied osteogenic differentiation of human mesenchymal stem cells (MSC) in an alginate hydrogel system where the cells were co-immobilized with alkaline phosphatase (ALP) for gradual mineralization of the microenvironment. MSC were embedded in unmodified alginate beads and alginate beads mineralized with ALP to generate a polymer/hydroxyapatite scaffold mimicking the composition of bone. The initial scaffold mineralization induced further mineralization of the beads with nanosized particles, and scanning electron micrographs demonstrated presence of collagen in the mineralized and unmineralized alginate beads cultured in osteogenic medium. Cells in both types of beads sustained high viability and metabolic activity for the duration of the study (21 days) as evaluated by live/dead staining and alamar blue assay. MSC in beads induced to differentiate in osteogenic direction expressed higher mRNA levels of osteoblast-specific genes (RUNX2, COL1AI, SP7, BGLAP) than MSC in traditional cell cultures. Furthermore, cells differentiated in beads expressed both sclerostin (SOST) and dental matrix protein-1 (DMP1), markers for late osteoblasts/osteocytes. In conclusion, Both ALP-modified and unmodified alginate beads provide an environment that enhance osteogenic differentiation compared with traditional 2D culture. Also, the ALP-modified alginate beads showed profound mineralization and thus have the potential to serve as a bone substitute in tissue engineering. PMID:25769043

Fall-related self-efficacy, not balance and mobility performance, is related to accidental falls in chronic stroke survivors with low bone mineral density

PubMed Central

Pang, Marco Y.C.; Eng, Janice J.

2011-01-01

Introduction Chronic stroke survivors with low bone mineral density (BMD) are particularly prone to fragility fractures. The purpose of this study was to identify the determinants of balance, mobility and falls in this sub-group of stroke patients. Methods Thirty nine chronic stroke survivors with low hip BMD (T-score <-1.0) were studied. Each subject was evaluated for: balance, mobility, leg muscle strength, spasticity, and falls-related self-efficacy. Any falls in the past 12 months were also recorded. Multiple regression analysis was used to identify the determinants of balance and mobility performance whereas logistic regression was used to identify the determinants of falls. Results Multiple regression analysis revealed that after adjusting for basic demographics, falls-related self-efficacy remained independently associated with balance/mobility performance (R2=0.494, P<0.001). Logistic regression showed that falls-related self-efficacy, but not balance and mobility performance, was a significant determinant of falls (odds ratio: 0.18, P=0.04). Conclusions Falls-related self-efficacy, but not mobility and balance performance, was the most important determinant of accidental falls. This psychological factor should not be overlooked in the prevention of fragility fractures among chronic stroke survivors with low hip BMD. PMID:18097709

Investigation of gene expressions in differentiated cell derived bone marrow stem cells during bone morphogenetic protein-4 treatments with Fourier transform infrared spectroscopy

NASA Astrophysics Data System (ADS)

Zafari, Jaber; Jouni, Fatemeh Javani; Ahmadvand, Ali; Abdolmaleki, Parviz; Soodi, Malihe; Zendehdel, Rezvan

2017-02-01

A model was set up to predict the differentiation patterns based on the data extracted from FTIR spectroscopy. For this reason, bone marrow stem cells (BMSCs) were differentiated to primordial germ cells (PGCs). Changes in cellular macromolecules in the time of 0, 24, 48, 72, and 96 h of differentiation, as different steps of the differentiation procedure were investigated by using FTIR spectroscopy. Also, the expression of pluripotency (Oct-4, Nanog and c-Myc) and specific genes (Mvh, Stella and Fragilis) were investigated by real-time PCR. However, the expression of genes in five steps of differentiation was predicted by FTIR spectroscopy. FTIR spectra showed changes in the template of band intensities at different differentiation steps. There are increasing changes in the stepwise differentiation procedure for the ratio area of CH2, which is symmetric to CH2 asymmetric stretching. An ensemble of expert methods, including regression tree (RT), boosting algorithm (BA), and generalized regression neural network (GRNN), was the best method to predict the gene expression by FTIR spectroscopy. In conclusion, the model was able to distinguish the pattern of different steps from cell differentiation by using some useful features extracted from FTIR spectra.

Effects of water extract of Cajanus cajan leaves on the osteogenic and adipogenic differentiation of mouse primary bone marrow stromal cells and the adipocytic trans-differentiation of mouse primary osteoblasts.

PubMed

Zhang, Jinchao; Liu, Cuilian; Sun, Jing; Liu, Dandan; Wang, Peng

2010-01-01

The effects of water extract of Cajanus cajan (Linn.) Millsp. (Leguminosae) leaves (WECML) on the osteogenic and adipogenic differentiation of mouse primary bone marrow stromal cells (BMSCs) and the adipocytic trans-differentiation of mouse primary osteoblasts (OBs) were studied. The results indicated that WECML promoted the proliferation of BMSCs and OBs at most concentrations. WECML promoted the osteogenic differentiation and formation of mineralized matrix nodules of BMSCs at concentrations of 0.1, 1, and 10 microg/mL, but inhibited the osteogenic differentiation and formation of mineralized matrix nodules of BMSCs at concentration of 0.01 microg/mL. WECML inhibited the adipogenic differentiation of BMSCs and adipocytic trans-differentiation of OBs at concentrations of 0.001, 0.1, 1, 10, and 100 microg/mL, but had no effects at concentration of 0.01 microg/mL. The results suggest that WECML has protective effects on bone and these protective effects may be mediated by decreasing adipocytic cell formation from BMSCs, which may promote the proliferation, differentiation, and mineralization function of OBs. The defined active ingredients in the WECML and the active mechanism need to be further studied.

Screening for Key Pathways Associated with the Development of Osteoporosis by Bioinformatics Analysis

PubMed Central

Liu, Yanqing; Wang, Yueqiu; Zhang, Yanxia; Liu, Zhiyong; Xiang, Hongfei; Peng, Xianbo

2017-01-01

Objectives. We aimed to find the key pathways associated with the development of osteoporosis. Methods. We downloaded expression profile data of GSE35959 and analyzed the differentially expressed genes (DEGs) in 3 comparison groups (old_op versus middle, old_op versus old, and old_op versus senescent). KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analyses were carried out. Besides, Venn diagram analysis and gene functional interaction (FI) network analysis were performed. Results. Totally 520 DEGs, 966 DEGs, and 709 DEGs were obtained in old_op versus middle, old_op versus old, and old_op versus senescent groups, respectively. Lysosome pathway was the significantly enriched pathways enriched by intersection genes. The pathways enriched by subnetwork modules suggested that mitotic metaphase and anaphase and signaling by Rho GTPases in module 1 had more proteins from module. Conclusions. Lysosome pathway, mitotic metaphase and anaphase, and signaling by Rho GTPases may be involved in the development of osteoporosis. Furthermore, Rho GTPases may regulate the balance of bone resorption and bone formation via controlling osteoclast and osteoblast. These 3 pathways may be regarded as the treatment targets for osteoporosis. PMID:28466021

The Role of Calcium in Osteoporosis

NASA Technical Reports Server (NTRS)

Arnaud, C. D.; Sanchez, S. D.

1991-01-01

Calcium requirements may vary throughout the lifespan. During the growth years and up to age 25 to 30, it is important to maximize dietary intake of calcium to maintain positive calcium balance and achieve peak bone mass, thereby possibly decreasing the risk of fracture when bone is subsequently lost. Calcium intake need not be greater than 800 mg/day during the relatively short period of time between the end of bone building and the onset of bone loss (30 to 40 years). Starting at age 40 to 50, both men and women lose bone slowly, but women lose bone more rapidly around the menopause and for about 10 years after. Intestinal calcium absorption and the ability to adapt to low calcium diets are impaired in many postmenopausal women and elderly persons owing to a suspected functional or absolute decrease in the ability of the kidney to produce 1,25(OH)2D2. The bones then become more and more a source of calcium to maintain critical extracellular fluid calcium levels. Excessive dietary intake of protein and fiber may induce significant negative calcium balance and thus increase dietary calcium requirements. Generally, the strongest risk factors for osteoporosis are uncontrollable (e.g., sex, age, and race) or less controllable (e.g., disease and medications). However, several factors such as diet, physical activity, cigarette smoking, and alcohol use are lifestyle related and can be modified to help reduce the risk of osteoporosis.

Induction of DNA-strand breaks after X-irradiation in murine bone cells of various differentiation capacities

NASA Astrophysics Data System (ADS)

Lau, Patrick; Hellweg, Christine E.; Kirchner, Simone; Baumstark-Khan, Christa

During longterm space missions, astronauts suffer from the loss of minerals especially from weightbearing bones due to prolonged sojourn under microgravity. In addition to weightlessness, exposure to cosmic ionization radiation is another space related factor endangering health and productivity of astronauts. In order to elucidate changes in bone cell metabolism induced by ionizing radiation, ground-based bone cell models have been developed. The differentiation level of the bone cells may influence their radiation sensitivity. Therefore, our cell model comprises a collection of immortalized murine pre-osteoblast, osteoblast and osteocyte cell lines representing discrete stages of differentiation: the subclones 4 and 24 of the osteoblast cell line MC3T3-E1, the osteoblast cell line OCT-1 and the osteocyte cell line MLO-Y4 display varying potential to produce mineralized bone matrix upon incubation with ascorbic acid and β-glycerophosphate (osteogenic medium). The MLO-Y4 cells showed the highest and subclone 24 the lowest proliferation rate. The most intense von Kossa reaction after culture in osteogenic medium was observed in subclone 4, indicating mineralized bone matrix. The bone cell markers alkaline phosphatase and osteocalcin were determined to further characterize the differentiation stage. All cell lines expressed osteocalcin, as determined by reverse transcriptase polymerase chain reaction. The activity of alkaline phosphatase was highest in the cell line OCT-1 and very low in MLO-Y4 and S4. The peculiarity of the markers suggests a characterization of OCT-1 and S24 as preosteoblast, S4 as (mature) osteoblast, and MLO-Y4 as osteocyte. Survival after exposure to X-rays was determined using the colony forming ability test. The resulting dose-effect relationships revealed normal radiation sensitivity (compared to human fibroblasts). Cell clone specific variations (subclones 4 and 24) in the radiation sensitivity may be due to the differentiation level. The survival curve of MLO-Y4 shows a broad shoulder, suggesting a high repair capacity or a high DNA damage or misrepair tolerance. The quantitative acquisition of DNA-strand breaks was performed by fluorescent analysis of DNA unwinding and revealed a high level of DNA damage immediately after X-irradiation, which increases dose dependently. In conclusion, the cell line with the highest differentiation level (MLO-Y4) displays lower radiation sensitivity, regarding the shoulder width of the dose-effect curve, compared to the less differentiated osteoblast cell lines.

ATF4, A Novel Mediator of the Anabolic Actions of PTH on Bone

DTIC Science & Technology

2009-07-01

formation rate and bone mineral density (severe osteoporosis) that persists throughout life. The expression of both osteocalcin (Ocn) and bone sialoprotein ...established that ATF4 is critical for osteoblast differentiation as demonstrated by dramatically reduced expression of osteocalcin and bone sialoprotein mRNA

N-acetylcysteine supplementation decreases osteoclast differentiation and increases bone mass in mice fed a high-fat diet

USDA-ARS?s Scientific Manuscript database

Studies have demonstrated that obesity induced by high-fat diets increases bone resorption, decreases trabecular bone mass, and reduces bone strength in various animal models. This study investigated whether N-acetylcysteine (NAC), an antioxidant and a glutathione precursor, alters glutathione statu...

Bioenergetics during calvarial osteoblast differentiation reflect strain differences in bone mass.

PubMed

Guntur, Anyonya R; Le, Phuong T; Farber, Charles R; Rosen, Clifford J

2014-05-01

Osteoblastogenesis is the process by which mesenchymal stem cells differentiate into osteoblasts that synthesize collagen and mineralize matrix. The pace and magnitude of this process are determined by multiple genetic and environmental factors. Two inbred strains of mice, C3H/HeJ and C57BL/6J, exhibit differences in peak bone mass and bone formation. Although all the heritable factors that differ between these strains have not been elucidated, a recent F1 hybrid expression panel (C3H × B6) revealed major genotypic differences in osteoblastic genes related to cellular respiration and oxidative phosphorylation. Thus, we hypothesized that the metabolic rate of energy utilization by osteoblasts differed by strain and would ultimately contribute to differences in bone formation. In order to study the bioenergetic profile of osteoblasts, we measured oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) first in a preosteoblastic cell line MC3T3-E1C4 and subsequently in primary calvarial osteoblasts from C3H and B6 mice at days 7, 14, and 21 of differentiation. During osteoblast differentiation in media containing ascorbic acid and β-glycerophosphate, all 3 cell types increased their oxygen consumption and extracellular acidification rates compared with the same cells grown in regular media. These increases are sustained throughout differentiation. Importantly, C3H calvarial osteoblasts had greater oxygen consumption rates than B6 consistent with their in vivo phenotype of higher bone formation. Interestingly, osteoblasts utilized both oxidative phosphorylation and glycolysis during the differentiation process although mature osteoblasts were more dependent on glycolysis at the 21-day time point than oxidative phosphorylation. Thus, determinants of oxygen consumption reflect strain differences in bone mass and provide the first evidence that during collagen synthesis osteoblasts use both glycolysis and oxidative phosphorylation to synthesize and mineralize matrix.

Bioenergetics During Calvarial Osteoblast Differentiation Reflect Strain Differences in Bone Mass

PubMed Central

Le, Phuong T.; Farber, Charles R.; Rosen, Clifford J.

2014-01-01

Osteoblastogenesis is the process by which mesenchymal stem cells differentiate into osteoblasts that synthesize collagen and mineralize matrix. The pace and magnitude of this process are determined by multiple genetic and environmental factors. Two inbred strains of mice, C3H/HeJ and C57BL/6J, exhibit differences in peak bone mass and bone formation. Although all the heritable factors that differ between these strains have not been elucidated, a recent F1 hybrid expression panel (C3H × B6) revealed major genotypic differences in osteoblastic genes related to cellular respiration and oxidative phosphorylation. Thus, we hypothesized that the metabolic rate of energy utilization by osteoblasts differed by strain and would ultimately contribute to differences in bone formation. In order to study the bioenergetic profile of osteoblasts, we measured oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) first in a preosteoblastic cell line MC3T3-E1C4 and subsequently in primary calvarial osteoblasts from C3H and B6 mice at days 7, 14, and 21 of differentiation. During osteoblast differentiation in media containing ascorbic acid and β-glycerophosphate, all 3 cell types increased their oxygen consumption and extracellular acidification rates compared with the same cells grown in regular media. These increases are sustained throughout differentiation. Importantly, C3H calvarial osteoblasts had greater oxygen consumption rates than B6 consistent with their in vivo phenotype of higher bone formation. Interestingly, osteoblasts utilized both oxidative phosphorylation and glycolysis during the differentiation process although mature osteoblasts were more dependent on glycolysis at the 21-day time point than oxidative phosphorylation. Thus, determinants of oxygen consumption reflect strain differences in bone mass and provide the first evidence that during collagen synthesis osteoblasts use both glycolysis and oxidative phosphorylation to synthesize and mineralize matrix. PMID:24437492

White blood cell differential count of maturation stages in bone marrow smear using dual-stage convolutional neural networks.

PubMed

Choi, Jin Woo; Ku, Yunseo; Yoo, Byeong Wook; Kim, Jung-Ah; Lee, Dong Soon; Chai, Young Jun; Kong, Hyoun-Joong; Kim, Hee Chan

2017-01-01

The white blood cell differential count of the bone marrow provides information concerning the distribution of immature and mature cells within maturation stages. The results of such examinations are important for the diagnosis of various diseases and for follow-up care after chemotherapy. However, manual, labor-intensive methods to determine the differential count lead to inter- and intra-variations among the results obtained by hematologists. Therefore, an automated system to conduct the white blood cell differential count is highly desirable, but several difficulties hinder progress. There are variations in the white blood cells of each maturation stage, small inter-class differences within each stage, and variations in images because of the different acquisition and staining processes. Moreover, a large number of classes need to be classified for bone marrow smear analysis, and the high density of touching cells in bone marrow smears renders difficult the segmentation of single cells, which is crucial to traditional image processing and machine learning. Few studies have attempted to discriminate bone marrow cells, and even these have either discriminated only a few classes or yielded insufficient performance. In this study, we propose an automated white blood cell differential counting system from bone marrow smear images using a dual-stage convolutional neural network (CNN). A total of 2,174 patch images were collected for training and testing. The dual-stage CNN classified images into 10 classes of the myeloid and erythroid maturation series, and achieved an accuracy of 97.06%, a precision of 97.13%, a recall of 97.06%, and an F-1 score of 97.1%. The proposed method not only showed high classification performance, but also successfully classified raw images without single cell segmentation and manual feature extraction by implementing CNN. Moreover, it demonstrated rotation and location invariance. These results highlight the promise of the proposed method as an automated white blood cell differential count system.

White blood cell differential count of maturation stages in bone marrow smear using dual-stage convolutional neural networks

PubMed Central

Choi, Jin Woo; Ku, Yunseo; Yoo, Byeong Wook; Kim, Jung-Ah; Lee, Dong Soon; Chai, Young Jun; Kong, Hyoun-Joong

2017-01-01

The white blood cell differential count of the bone marrow provides information concerning the distribution of immature and mature cells within maturation stages. The results of such examinations are important for the diagnosis of various diseases and for follow-up care after chemotherapy. However, manual, labor-intensive methods to determine the differential count lead to inter- and intra-variations among the results obtained by hematologists. Therefore, an automated system to conduct the white blood cell differential count is highly desirable, but several difficulties hinder progress. There are variations in the white blood cells of each maturation stage, small inter-class differences within each stage, and variations in images because of the different acquisition and staining processes. Moreover, a large number of classes need to be classified for bone marrow smear analysis, and the high density of touching cells in bone marrow smears renders difficult the segmentation of single cells, which is crucial to traditional image processing and machine learning. Few studies have attempted to discriminate bone marrow cells, and even these have either discriminated only a few classes or yielded insufficient performance. In this study, we propose an automated white blood cell differential counting system from bone marrow smear images using a dual-stage convolutional neural network (CNN). A total of 2,174 patch images were collected for training and testing. The dual-stage CNN classified images into 10 classes of the myeloid and erythroid maturation series, and achieved an accuracy of 97.06%, a precision of 97.13%, a recall of 97.06%, and an F-1 score of 97.1%. The proposed method not only showed high classification performance, but also successfully classified raw images without single cell segmentation and manual feature extraction by implementing CNN. Moreover, it demonstrated rotation and location invariance. These results highlight the promise of the proposed method as an automated white blood cell differential count system. PMID:29228051

The role of bone marrow-derived cells during the bone healing process in the GFP mouse bone marrow transplantation model.

PubMed

Tsujigiwa, Hidetsugu; Hirata, Yasuhisa; Katase, Naoki; Buery, Rosario Rivera; Tamamura, Ryo; Ito, Satoshi; Takagi, Shin; Iida, Seiji; Nagatsuka, Hitoshi

2013-03-01

Bone healing is a complex and multistep process in which the origin of the cells participating in bone repair is still unknown. The involvement of bone marrow-derived cells in tissue repair has been the subject of recent studies. In the present study, bone marrow-derived cells in bone healing were traced using the GFP bone marrow transplantation model. Bone marrow cells from C57BL/6-Tg (CAG-EGFP) were transplanted into C57BL/6 J wild mice. After transplantation, bone injury was created using a 1.0-mm drill. Bone healing was histologically assessed at 3, 7, 14, and 28 postoperative days. Immunohistochemistry for GFP; double-fluorescent immunohistochemistry for GFP-F4/80, GFP-CD34, and GFP-osteocalcin; and double-staining for GFP and tartrate-resistant acid phosphatase were performed. Bone marrow transplantation successfully replaced the hematopoietic cells into GFP-positive donor cells. Immunohistochemical analyses revealed that osteoblasts or osteocytes in the repair stage were GFP-negative, whereas osteoclasts in the repair and remodeling stages and hematopoietic cells were GFP-positive. The results indicated that bone marrow-derived cells might not differentiate into osteoblasts. The role of bone marrow-derived cells might be limited to adjustment of the microenvironment by differentiating into inflammatory cells, osteoclasts, or endothelial cells in immature blood vessels.

Hypercalcaemia and hypocalcaemia: finding the balance.

PubMed

Body, Jean-Jacques; Niepel, Daniela; Tonini, Giuseppe

2017-05-01

The balance between bone formation and resorption may be disrupted in patients with cancer, leading either to increased bone resorption, calcium release, and possibly hypercalcaemia, or to increased bone formation, sequestration of calcium, and possibly hypocalcaemia. In adults, hypercalcaemia of malignancy is most common in patients with tumours that produce factors that induce osteoclast activation and enhance bone resorption. Impaired renal function and increased renal tubular calcium resorption may further affect calcium levels. Inhibitors of bone resorption, first the bisphosphonates and, later, denosumab, have been shown to be effective in hypercalcaemia treatment. Bisphosphonates (which are administered intravenously) are approved for hypercalcaemia of malignancy and are the current mainstay of treatment, whereas denosumab (which is administered subcutaneously) may offer an option for patients who do not respond to bisphosphonates or suffer from renal insufficiency. TREATMENT AND PREVENTION: Hypocalcaemia is most common in patients with prostate cancer and osteoblastic bone metastases, but can occur in patients with a variety of tumour types who are receiving inhibitors of bone resorption. While patients often respond to calcium and vitamin D supplementation, prevention should be the aim; at-risk patients should be identified before starting treatment with inhibitors of bone resorption, be closely monitored during at least the first few months of treatment, and receive concomitant calcium and vitamin D supplementation unless hypercalcaemia is present. Both hypercalcaemia and hypocalcaemia can be serious if left untreated. It is therefore important that patients with cancer are closely monitored and receive adequate prevention and treatment measures to maintain normal blood calcium levels.

TULA-2, a novel histidine phosphatase regulates bone remodeling by modulating osteoclast function

PubMed Central

Back, Steven H.; Adapala, Naga Suresh; Barbe, Mary F.; Carpino, Nick C.; Tsygankov, Alexander Y.; Sanjay, Archana

2013-01-01

Bone is a dynamic tissue that depends on the intricate relationship between protein tyrosine kinases (PTK) and protein tyrosine phosphatases (PTP) for maintaining homeostasis. PTKs and PTPs act like molecular on and off switches and help modulate differentiation and the attachment of osteoclasts to bone matrix regulating bone resorption. The novel protein T-cell Ubiquitin Ligand-2 (TULA-2), which is abundantly expressed in osteoclasts, is a novel histidine phosphatase. Our results show that of the two family members only TULA-2 is expressed in osteoclasts and that its expression is sustained throughout the course of osteoclast differentiation suggesting that TULA-2 may play a role during early as well late stages of osteoclast differentiation. Skeletal analysis of mice that do not express TULA or TULA-2 proteins (DKO Mice) revealed that there was a decrease in bone volume due to increased osteoclast numbers and function. Furthermore, in vitro experiments indicated that bone marrow precursor cells from DKO mice have an increased potential to form osteoclasts. At the molecular level, the absence of TULA-2 in osteoclasts results in increased Syk phosphorylation at the Y352 and Y525/526 residues and activation of phospholipase C gamma 2 (PLCγ2) upon engagement of Immune-receptor-Tyrosine-based-Activation-Motif (ITAM)–mediated signaling. Furthermore, expression of a phosphatase-dead TULA-2 leads to increased osteoclast function. Taken together, these results suggest that TULA-2 negatively regulates osteoclast differentiation and function. PMID:23149425

TULA-2, a novel histidine phosphatase, regulates bone remodeling by modulating osteoclast function.

PubMed

Back, Steven H; Adapala, Naga Suresh; Barbe, Mary F; Carpino, Nick C; Tsygankov, Alexander Y; Sanjay, Archana

2013-04-01

Bone is a dynamic tissue that depends on the intricate relationship between protein tyrosine kinases (PTK) and protein tyrosine phosphatases (PTP) for maintaining homeostasis. PTKs and PTPs act like molecular on and off switches and help modulate differentiation and the attachment of osteoclasts to bone matrix regulating bone resorption. The protein T cell ubiquitin ligand-2 (TULA-2), which is abundantly expressed in osteoclasts, is a novel histidine phosphatase. Our results show that of the two family members, only TULA-2 is expressed in osteoclasts and that its expression is sustained throughout the course of osteoclast differentiation, suggesting that TULA-2 may play a role during early as well late stages of osteoclast differentiation. Skeletal analysis of mice that do not express TULA or TULA-2 proteins (DKO mice) revealed that there was a decrease in bone volume due to increased osteoclast numbers and function. Furthermore, in vitro experiments indicated that bone marrow precursor cells from DKO mice have an increased potential to form osteoclasts. At the molecular level, the absence of TULA-2 in osteoclasts results in increased Syk phosphorylation at the Y352 and Y525/526 residues and activation of phospholipase C gamma 2 (PLCγ2) upon engagement of immune-receptor-tyrosine-based-activation-motif (ITAM)-mediated signaling. Furthermore, expression of a phosphatase-dead TULA-2 leads to increased osteoclast function. Taken together, these results suggest that TULA-2 negatively regulates osteoclast differentiation and function.

Conjugated linoleic acid prevents age-induced bone loss in mice by regulating both osteoblastogenesis and adipogenesis.

PubMed

Lin, Guanlin; Wang, Huan; Dai, Jun; Li, Xiao; Guan, Ming; Gao, Shutao; Ding, Qing; Wang, Huaixi; Fang, Huang

2017-08-26

Osteoporosis (OP) can increase the risk of bone fracture and other complications, which is a major clinical problem. Previous researches have revealed that conjugated linoleic acid (CLA) can promote the bone formation. But the mechanisms are not clear. Thus, we tested the hypothesis that CLA acts on bone formation might be via mTOR Complex1 (mTORC 1) pathway by in vitro and vivo assays. We studied the effect of CLA mix on MC3T3-E1 pre-osteoblasts differentiation into osteoblasts, and bone formation under osteoporotic conditions. At the same time, 3T3-L1 pre-adipocyte with the same CLA mix concentration gradient for 8 days with adipogenic differentiation medium. We found that Alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2) and osteocalcin (OCN) expressions of pre-osteoblasts were up-regulated. Moreover in presence of CLA, peroxisome proliferators-activated receptor γ(PPARγ) and CCAAT/enhancer-binding protein (C/EBPα) were down-regulated. Osteoporosis mice bone parameters in the distal femoral meraphysis were significantly increased compared with placebo mice. Furthermore, the phosphor-S6 (P-S6) was suppressed and phosphor-AKT (P-AKT) was up-regulated. Consistently, CLA can stimulate differentiation of osteoblasts and inhibited pre-adipocytes differentiated into adipocytes via AKT/mTORC1 signal pathway. Overall CLA thus be a suitable candidate for the treatment of patients with postmenopausal osteoporosis and obesity. Copyright © 2017 Elsevier Inc. All rights reserved.

MiR-214 regulates the function of osteoblast under simulated microgravity by targeting ATF4

NASA Astrophysics Data System (ADS)

Li, Yingxian; Wang, Xiaogang; Li, Qi; Lv, Ke; Wan, Yumin; Li, Yinghui; Bai, Yanqiang

Background: MicroRNAs (miRNAs) are small fragments of single-stranded RNA containing 18-24 nucleotides, and are generated from endogenous transcripts. MicroRNAs function in post-transcriptional gene silencing by targeting the 3'-untranslated region (UTR) of mRNAs, resulting in translational repression. Growing evidence shows that microRNAs (miRNAs) regu-late various developmental and homeostatic events in vertebrates and invertebrates. Osteoblast differentiation is a key step in proper skeletal development and acquisition of bone mass; How-ever, the physiological role of non-coding small RNAs, especially miRNAs, in osteoblast dif-ferentiation remains elusive. Methods: To study the potential involvement of miRNAs in osteoblast differentiation under stimulated microgravity, we analyzed the expression of 20 bone relative miRNAs using real time PCR platform to find particularly miRNAs whose expression is altered during osteoblast differentiation. TargetScan, miRBase and Miranda were used to predict the target gene of candidate miRNA. To investigate whether ATF4 can be directly targeted by miR-214, we engineered luciferase reporters that have either the wild-type 3'UTRs of these genes, or the mutant UTRs with a 6 base pair (bp) deletion in the target sites. Lastly, to address the in vivo role of miR-214 in bone formation, tail suspension mice model was used to simulate the change of osteoblast function and bone loss. Results: Recent studies have sug-gested that miRNAs might play a role in osteoblast differentiation and bone formation. Here, we identify miR-214 in MC3T3-E1 cells, which is a primary mouse osteoblasts cell line, to promote osteoblast differentiation by repressing Activating Transcription Factor4 (ATF4) ex-pression at the posttranscriptional level. What is more, miR-214 was found to be transcribed in C2C12 cells during bone morphogenetic protein 2-induced (BMP2-induced) osteogenesis, and overexpression of miR-214 attenuated BMP2-induced osteoblastogenesis, whereas inhibition of miR-214 expression enhanced this progress. The levels of miR-214 increased dramatically in tail suspension mice. Conclusions: Thus, our studies show that miR-214 plays an important role in osteoblast differentiation by targeting ATF4 under stimulated microgravity induced bone loss and contributes to osteoporosis via its effect on osteoblasts.

Selection of an appropriate animal model for study of bone loss in weightlessness

NASA Technical Reports Server (NTRS)

Wolinsky, I.

1986-01-01

Prolonged weightlessness in space flight results in a slow progressive demineralization of bone accompanied by an increased calcium output in the urine resulting in negative calcium balances. This possibly irreversible bone loss may constitute a serious limiting factor to long duration manned space flight. A number of preventative measures have been suggested, i.e., exercise during flight, dietary calcium supplements, use of specific prophylactic drugs. In order to facilitate research in these areas it is necessary to develop appropriate ground-based animal models that simulate the human condition of osteoporsis. An appropriate animal model would permit bone density studies, calcium balance studies, biochemical analyses, ground-based simulation models of weightlessness (bed rest, restraint, immobilization) and the planning of inflight experiments. Several animal models have been proposed in the biomedical research literature, but have inherent deficiencies. The purpose of this project was to evaluate models in the literature and determine which of these most closely simulates the phenomenon of bone loss in humans with regard to growth, bone remodeling, structural, chemical and mineralization similarities to human. This was accomplished by a comprehensive computer assisted literature search and report. Three animal models were examined closely for their relative suitability: the albino rat, monkey, and Beagle.

The influence of exercise on bone atrophy

NASA Technical Reports Server (NTRS)

1978-01-01

The relationship between the skeletal system, the muscular system, and exercise in bed rest studies are described. The regime of exercises performed, the mineral balance data derived, and the bone densitometric data obtained are discussed. A brief review of some of the histological results are also given.

Periprosthetic bone remodelling of short-stem total hip arthroplasty: a systematic review.

PubMed

Yan, Shuang G; Weber, Patrick; Steinbrück, Arnd; Hua, Xingyi; Jansson, Volkmar; Schmidutz, Florian

2017-11-27

Short-stem hip arthroplasty (SHA) was designed to preserve bone stock and provide an improved load transfer. To gain more evidence regarding the load transfer, this review analysed the periprosthetic bone remodelling of SHA in comparison to standard hip arthroplasty (THA). PubMed and ScienceDirect were screened to extract dual-energy X-ray absorptiometry (DXA) studies evaluating the periprosthetic bone remodelling of SHA and two proven THA designs. From the studies included, the postoperative change in periprosthetic bone mineral density (BMD) after one year and the trend over two years was determined. Fifteen studies with four SHAs (CFP, Metha, Nanos, Fitmore) and two THAs (CLS and Bicontact) designs were included. All SHA and THA stems revealed an initial decrease at the calcar and major trochanter (Gruen 1 and 7) with the Metha, Nanos and Fitmore showing a smaller and more balanced remodelling compared to THA. The pattern after one year and the trend over two years argue for a methaphyseal anchorage of the Metha and Nanos, whereas the Fitmore and CFP seem to anchor metha-diaphyseal. Clearly different pattern of bone remodelling were observed between all four SHAs. Periprosthetic bone remodelling is also present in SHA, with the main bone reduction observed proximally. However, certain SHA stems show a more balanced remodelling compared to THA, arguing for a favourable load transfer. Also, the femoral length where bone remodelling occurs is clearly shorter in SHA. As distinctively different pattern between the SHA designs were observed, they should not be judged as a single implant group.

Selection, proliferation and differentiation of bone marrow-derived liver stem cells with a culture system containing cholestatic serum in vitro.

PubMed

Cai, Yun-Feng; Zhen, Zuo-Jun; Min, Jun; Fang, Tian-Ling; Chu, Zhong-Hua; Chen, Ji-Sheng

2004-11-15

To explore the feasibility of direct separation, selective proliferation and differentiation of the bone marrow-derived liver stem cells (BDLSC) from bone marrow cells with a culture system containing cholestatic serum in vitro. Whole bone marrow cells of rats cultured in routine medium were replaced with conditioning selection media containing 20 mL/L, 50 mL/L, 70 mL/L, and 100 mL/L cholestatic sera, respectively, after they attached to the plates. The optimal concentration of cholestatic serum was determined according to the outcome of the selected cultures. Then the selected BDLSC were induced to proliferate and differentiate with the addition of hepatocyte growth factor (HGF). The morphology and phenotypic markers of BDLSC were characterized using immunohistochemistry, RT-PCR and electron microscopy. The metabolic functions of differentiated cells were also determined by glycogen staining and urea assay. Bone marrow cells formed fibroblast-like but not hepatocyte-like colonies in the presence of 20 mL/L cholestatic serum. In 70 mL/L cholestatic serum, BDLSC colonies could be selected but could not maintain good growth status. In 100 mL/L cholestatic serum, all of the bone marrow cells were unable to survive. A 50 mL/L cholestatic serum was the optimal concentration for the selection of BDLSC at which BDLSC could survive while the other populations of the bone marrow cells could not. The selected BDLSC proliferated and differentiated after HGF was added. Hepatocyte-like colony-forming units (H-CFU) then were formed. H-CFU expressed markers of embryonic hepatocytes (AFP, albumin and cytokeratin 8/18), biliary cells (cytokeratin 19), hepatocyte functional proteins (transthyretin and cytochrome P450-2b1), and hepatocyte nuclear factors (HNF-1alpha and HNF-3beta). They also had glycogen storage and urea synthesis functions, two of the critical features of hepatocytes. The selected medium containing cholestatic serum can select BDLSC from whole bone marrow cells. It will be a new way to provide a readily available alternate source of cells for clinical hepatocyte therapy.

Changes of vessel-cells complex in zones of adaptive remodeling of the bone tissue under microgravity conditions

NASA Astrophysics Data System (ADS)

Rodionova, N.; Oganov, V.; Nosova, L.

The development and differentiation of osteogenic cells in organism happen in closely topographical and functional connection with blood capillaries. We formerly proofed, that small-differentiated cells, which are in the population of perivascular cells are osteogenic cells -precursors . At the present time it is actually to clear up, how these biostructures react on conditions of less of biomechanical load on skeleton bones. We researched peculiarities of blood-bed structure and perivascular cells in metaphises of thighbones and tibial bones in rats, which were onboard the American space station SLS-2 and in experiments of modeling hypokinesia. There were used methods of cytochemistry, histology and electron microscopy. We established, that under the support and functional load decreasing in zones of bones adaptive remodeling, comparatively to control, on histosections the own volume of sinusoid capillaries reduces. The small vessels prevail here. The spaces of sinusoid capillaries are limited by 1 2 cells of the endothelia. Endotheliocytes in- general have the typical ultrastructure. Basal membranes are expressed not-distinctly. Perivascular cells don't create the unbroken layer. The population of these cells is not-homogeneous. It includes enclosed to endothelia small-differentiated forms and separating cells with sings of fibroblastic differentiation (the own volume of rough endoplasmic reticulum in cytoplasm induces). The part of these cells reacts on the alkaline phosphatase (the marker of the osteogenic differentiation). Under the conditions of support load decreasing (especially under the microgravity) there is a tendency to reducing of separating osteogenic cells number. We noted the priority of differentiating fibroblasts. It leads to further development in zones of bone remodeling of hearths of fibrous tissue, that doesn't mineralize. The obtained data are seen as one of mechanisms of osteoporosis and osteopenia development under the deficite of support load.

Nanostructured Biomaterials for Tissue Engineered Bone Tissue Reconstruction

PubMed Central

Chiara, Gardin; Letizia, Ferroni; Lorenzo, Favero; Edoardo, Stellini; Diego, Stomaci; Stefano, Sivolella; Eriberto, Bressan; Barbara, Zavan

2012-01-01

Bone tissue engineering strategies are emerging as attractive alternatives to autografts and allografts in bone tissue reconstruction, in particular thanks to their association with nanotechnologies. Nanostructured biomaterials, indeed, mimic the extracellular matrix (ECM) of the natural bone, creating an artificial microenvironment that promotes cell adhesion, proliferation and differentiation. At the same time, the possibility to easily isolate mesenchymal stem cells (MSCs) from different adult tissues together with their multi-lineage differentiation potential makes them an interesting tool in the field of bone tissue engineering. This review gives an overview of the most promising nanostructured biomaterials, used alone or in combination with MSCs, which could in future be employed as bone substitutes. Recent works indicate that composite scaffolds made of ceramics/metals or ceramics/polymers are undoubtedly more effective than the single counterparts in terms of osteoconductivity, osteogenicity and osteoinductivity. A better understanding of the interactions between MSCs and nanostructured biomaterials will surely contribute to the progress of bone tissue engineering. PMID:22312283

Overexpression of HSPA1A enhances the osteogenic differentiation of bone marrow mesenchymal stem cells via activation of the Wnt/β-catenin signaling pathway

PubMed Central

Zhang, Wei; Xue, Deting; Yin, Houfa; Wang, Shengdong; Li, Chao; Chen, Erman; Hu, Dongcai; Tao, Yiqing; Yu, Jiawei; Zheng, Qiang; Gao, Xiang; Pan, Zhijun

2016-01-01

HSPA1A, which encodes cognate heat shock protein 70, plays important roles in various cellular metabolic pathways. To investigate its effects on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), its expression level was compared between undifferentiated and differentiated BMSCs. Rat HSPA1A overexpression in BMSCs increased osteoblast-specific gene expression, alkaline phosphatase activity, and mineral deposition in vitro. Moreover, it upregulated β-catenin and downregulated DKK1 and SOST. The enhanced osteogenesis due to HSPA1A overexpression was partly rescued by a Wnt/β-catenin inhibitor. Additionally, using a rat tibial fracture model, a sheet of HSPA1A-overexpressing BMSCs improved bone fracture healing, as determined by imaging and histological analysis. Taken together, these findings suggest that HSPA1A overexpression enhances osteogenic differentiation of BMSCs, partly through Wnt/β-catenin. PMID:27279016

Biophysical regulation of stem cell differentiation.

PubMed

Govey, Peter M; Loiselle, Alayna E; Donahue, Henry J

2013-06-01

Bone adaptation to its mechanical environment, from embryonic through adult life, is thought to be the product of increased osteoblastic differentiation from mesenchymal stem cells. In parallel with tissue-scale loading, these heterogeneous populations of multipotent stem cells are subject to a variety of biophysical cues within their native microenvironments. Bone marrow-derived mesenchymal stem cells-the most broadly studied source of osteoblastic progenitors-undergo osteoblastic differentiation in vitro in response to biophysical signals, including hydrostatic pressure, fluid flow and accompanying shear stress, substrate strain and stiffness, substrate topography, and electromagnetic fields. Furthermore, stem cells may be subject to indirect regulation by mechano-sensing osteocytes positioned to more readily detect these same loading-induced signals within the bone matrix. Such paracrine and juxtacrine regulation of differentiation by osteocytes occurs in vitro. Further studies are needed to confirm both direct and indirect mechanisms of biophysical regulation within the in vivo stem cell niche.

Effects of the 1, 4-dihydropyridine L-type calcium channel blocker benidipine on bone marrow stromal cells.

PubMed

Ma, Zhong-ping; Liao, Jia-cheng; Zhao, Chang; Cai, Dao-zhang

2015-08-01

Osteoporosis (OP) often increases the risk of bone fracture and other complications and is a major clinical problem. Previous studies have found that high blood pressure is associated with bone formation abnormalities, resulting in increased calcium loss. We have investigated the effect of the antihypertensive drug benidipine on bone marrow stromal cell (BMSC) differentiation into osteoblasts and bone formation under osteoporotic conditions. We used a combination of in vitro and in vivo approaches to test the hypothesis that benidipine promotes murine BMSC differentiation into osteoblasts. Alkaline phosphatase (ALP), osteocalcin (OCN), runt-related transcription factor 2 (RUNX2), β-catenin, and low-density lipoprotein receptor-related protein 5 (LRP5) protein expression was evaluated in primary femoral BMSCs from C57/BL6 mice cultured under osteogenic conditions for 2 weeks to examine the effects of benidipine. An ovariectomized (OVX) mouse model was used to investigate the effect of benidipine treatment for 3 months in vivo. We found that ALP, OCN, and RUNX2 expression was up-regulated and WNT/β-catenin signaling was enhanced in vitro and in vivo. In OVX mice that were intragastrically administered benidipine, bone parameters (trabecular thickness, bone mineral density, and trabecular number) in the distal femoral metaphysis were significantly increased compared with control OVX mice. Consistently, benidipine promoted BMSC differentiation into osteoblasts and protected against bone loss in OVX mice. Therefore, benidipine might be a suitable candidate for the treatment of patients with postmenopausal osteoporosis and hypertension.

Contribution of methylglyoxal to delayed healing of bone injury in diabetes.

PubMed

Aikawa, Takao; Matsubara, Hidenori; Ugaji, Shuhei; Shirakawa, Junichi; Nagai, Ryoji; Munesue, Seiichi; Harashima, Ai; Yamamoto, Yasuhiko; Tsuchiya, Hiroyuki

2017-07-01

Patients with diabetes are vulnerable to delayed bone fracture healing or pseudoarthrosis. Chronic sustained hyperglycemia, reactive intermediate derivatives of glucose metabolism, such as methylglyoxal (MGO), and advanced glycation end‑products (AGEs) are implicated in diabetic complications. In the present study, it was examined whether MGO is able to cause disturbed bone healing in diabetes. Diabetes was induced in male mice by injection of streptozotocin (50 mg/kg) for 5 days. A bone defect (1.0‑mm diameter) was created in the left distal femur, and bone repair was assessed from an examination of computed tomography scans. ST2 cells were exposed to MGO (0‑400 µM) to investigate osteoblastic differentiation, cell viability, and damage. Consequently, blood glucose and hemoglobin A1c levels in diabetic mice were determined to be 493±14.1 mg/dl and 8.0±0.05%, respectively. Compared with non‑diabetic control mice, diabetic mice exhibited markedly delayed bone healing, with increased levels of the MGO‑derived AGEs, Nε‑(carboxymethyl)‑lysine and Nδ‑(5‑hydro‑5‑methyl‑4‑imidazolone‑2‑yl)‑ornithine, in the sera and femurs. MGO inhibited the osteoblastic differentiation of ST2 cells in a dose‑dependent manner, and markedly decreased cell proliferation through cytotoxicity. In conclusion, MGO has been demonstrated to cause impaired osteoblastic differentiation and delayed bone repair in diabetes. Therefore, detoxification of MGO may be a potentially useful strategy against bone problems in patients with diabetes.

Locally Produced BDNF Promotes Sclerotic Change in Alveolar Bone after Nerve Injury

PubMed Central

Ida-Yonemochi, Hiroko; Yamada, Yurie; Yoshikawa, Hiroyuki

2017-01-01

Brain-derived neurotrophic factor (BDNF), which is released due to nerve injury, is known to promote the natural healing of injured nerves. It is often observed that damage of mandibular canal induces local sclerotic changes in alveolar bone. We reported that peripheral nerve injury promotes the local production of BDNF; therefore, it was possible to hypothesize that peripheral nerve injury affects sclerotic changes in the alveolar bone. This study aimed to evaluate the effect of BDNF on osteogenesis using in vitro osteoblast-lineage cell culture and an in vivo rat osteotomy model. MC3T3-E1 cells were cultured with BDNF and were examined for cell proliferative activity, chemotaxis and mRNA expression levels of osteoblast differentiation markers. For in vivo study, inferior alveolar nerve (IAN) injury experiments and mandibular cortical osteotomy were performed using a rat model. In the osteotomy model, exogenous BDNF was applied to bone surfaces after corticotomy of the mandible, and we morphologically analyzed the new bone formation. As a result, mRNA expression of osteoblast differentiation marker, osteocalcin, was significantly increased by BDNF, although cell proliferation and migration were not affected. In the in vivo study, osteopontin-positive new bone formation was significantly accelerated in the BDNF-grafted groups, and active bone remodeling, involving trkB-positive osteoblasts and osteocytes, continued after 28 days. In conclusion, BDNF stimulated the differentiation of MC3T3-E1 cells and it promoted new bone formation and maturation. These results suggested that local BDNF produced by peripheral nerve injury contributes to accelerating sclerotic changes in the alveolar bone. PMID:28072837

TRAF family member-associated NF-κB activator (TANK) is a negative regulator of osteoclastogenesis and bone formation.

PubMed

Maruyama, Kenta; Kawagoe, Tatsukata; Kondo, Takeshi; Akira, Shizuo; Takeuchi, Osamu

2012-08-17

The differentiation of bone-resorbing osteoclasts is induced by RANKL signaling, and leads to the activation of NF-κB via TRAF6 activation. TRAF family member-associated NF-κB activator (TANK) acts as a negative regulator of Toll-like receptors (TLRs) and B-cell receptor (BCR) signaling by inhibiting TRAF6 activation. Tank(-/-) mice spontaneously develop autoimmune glomerular nephritis in an IL-6-dependent manner. Despite its importance in the TCRs and BCR-activated TRAF6 inhibition, the involvement of TANK in RANKL signaling is poorly understood. Here, we report that TANK is a negative regulator of osteoclast differentiation. The expression levels of TANK mRNA and protein were up-regulated during RANKL-induced osteoclastogenesis, and overexpression of TANK in vitro led to a decrease in osteoclast formation. The in vitro osteoclastogenesis of Tank(-/-) cells was significantly increased, accompanied by increased ubiquitination of TRAF6 and enhanced canonical NF-κB activation in response to RANKL stimulation. Tank(-/-) mice showed severe trabecular bone loss, but increased cortical bone mineral density, because of enhanced bone erosion and formation. TANK mRNA expression was induced during osteoblast differentiation and Tank(-/-) osteoblasts exhibited enhaced NF-κB activation, IL-11 expression, and bone nodule formation than wild-type control cells. Finally, wild-type mice transplanted with bone marrow cells from Tank(-/-) mice showed trabecular bone loss analogous to that in Tank(-/-) mice. These findings demonstrate that TANK is critical for osteoclastogenesis by regulating NF-κB, and is also important for proper bone remodeling.

TRAF Family Member-associated NF-κB Activator (TANK) Is a Negative Regulator of Osteoclastogenesis and Bone Formation*

PubMed Central

Maruyama, Kenta; Kawagoe, Tatsukata; Kondo, Takeshi; Akira, Shizuo; Takeuchi, Osamu

2012-01-01

The differentiation of bone-resorbing osteoclasts is induced by RANKL signaling, and leads to the activation of NF-κB via TRAF6 activation. TRAF family member-associated NF-κB activator (TANK) acts as a negative regulator of Toll-like receptors (TLRs) and B-cell receptor (BCR) signaling by inhibiting TRAF6 activation. Tank−/− mice spontaneously develop autoimmune glomerular nephritis in an IL-6-dependent manner. Despite its importance in the TCRs and BCR-activated TRAF6 inhibition, the involvement of TANK in RANKL signaling is poorly understood. Here, we report that TANK is a negative regulator of osteoclast differentiation. The expression levels of TANK mRNA and protein were up-regulated during RANKL-induced osteoclastogenesis, and overexpression of TANK in vitro led to a decrease in osteoclast formation. The in vitro osteoclastogenesis of Tank−/− cells was significantly increased, accompanied by increased ubiquitination of TRAF6 and enhanced canonical NF-κB activation in response to RANKL stimulation. Tank−/− mice showed severe trabecular bone loss, but increased cortical bone mineral density, because of enhanced bone erosion and formation. TANK mRNA expression was induced during osteoblast differentiation and Tank−/− osteoblasts exhibited enhaced NF-κB activation, IL-11 expression, and bone nodule formation than wild-type control cells. Finally, wild-type mice transplanted with bone marrow cells from Tank−/− mice showed trabecular bone loss analogous to that in Tank−/− mice. These findings demonstrate that TANK is critical for osteoclastogenesis by regulating NF-κB, and is also important for proper bone remodeling. PMID:22773835

The control of human mesenchymal cell differentiation using nanoscale symmetry and disorder

NASA Astrophysics Data System (ADS)

Dalby, Matthew J.; Gadegaard, Nikolaj; Tare, Rahul; Andar, Abhay; Riehle, Mathis O.; Herzyk, Pawel; Wilkinson, Chris D. W.; Oreffo, Richard O. C.

2007-12-01

A key tenet of bone tissue engineering is the development of scaffold materials that can stimulate stem cell differentiation in the absence of chemical treatment to become osteoblasts without compromising material properties. At present, conventional implant materials fail owing to encapsulation by soft tissue, rather than direct bone bonding. Here, we demonstrate the use of nanoscale disorder to stimulate human mesenchymal stem cells (MSCs) to produce bone mineral in vitro, in the absence of osteogenic supplements. This approach has similar efficiency to that of cells cultured with osteogenic media. In addition, the current studies show that topographically treated MSCs have a distinct differentiation profile compared with those treated with osteogenic media, which has implications for cell therapies.

Triiodothyronine increases calcium loss in a bed rest antigravity model for space flight.

PubMed

Smith, Steven R; Lovejoy, Jennifer C; Bray, George A; Rood, Jennifer; Most, Marlene M; Ryan, Donna H

2008-12-01

Bed rest has been used as a model to simulate the effects of space flight on bone metabolism. Thyroid hormones accelerate bone metabolism. Thus, supraphysiologic doses of this hormone might be used as a model to accelerate bone metabolism during bed rest and potentially simulate space flight. The objective of the study was to quantitate the changes in bone turnover after low doses of triiodothyronine (T(3)) added to short-term bed rest. Nine men and 5 women were restricted to bed rest for 28 days with their heads positioned 6 degrees below their feet. Subjects were randomly assigned to receive either placebo or oral T(3) at doses of 50 to 75 microg/d in a single-blind fashion. Calcium balance was measured over 5-day periods; and T(3), thyroxine, thyroid-stimulating hormone, immunoreactive parathyroid hormone, osteocalcin, bone alkaline phosphatase, and urinary deoxypyridinoline were measured weekly. Triiodothyronine increased 2-fold in the men and 5-fold in the women during treatment, suppressing both thyroxine and thyroid-stimulating hormone. Calcium balance was negative by 300 to 400 mg/d in the T(3)-treated volunteers, primarily because of the increased fecal loss that was not present in the placebo group. Urinary deoxypyridinoline to creatinine ratio, a marker of bone resorption, increased 60% in the placebo group during bed rest, but more than doubled in the T(3)-treated subjects (P < .01), suggesting that bone resorption was enhanced by treatment with T(3). Changes in serum osteocalcin and bone-specific alkaline phosphatase, markers of bone formation, were similar in T(3)- and placebo-treated subjects. Triiodothyronine increases bone resorption and fecal calcium loss in subjects at bed rest.

Hepatic differentiation capability of rat bone marrow-derived mesenchymal stem cells and hematopoietic stem cells.

PubMed

Shu, Sai-Nan; Wei, Lai; Wang, Jiang-Hua; Zhan, Yu-Tao; Chen, Hong-Song; Wang, Yu

2004-10-01

To investigate the different effects of mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) on hepatic differentiation. MSCs from rat bone marrow were isolated and cultured by standard methods. HSCs from rat bone marrow were isolated and purified by magnetic activated cell sorting. Both cell subsets were induced. Morphology, RT-PCR and immunocytochemistry were used to identify the hepatic differentiation grade. MSCs exhibited round in shape after differentiation, instead of fibroblast-like morphology before differentiation. Albumin mRNA and protein were expressed positively in MSCs, without detection of alpha-fetoprotein (AFP). HSCs were polygonal in shape after differentiation. The expression of albumin signal decreased and AFP signal increased. The expression of CK18 was continuous in MSCs and HSCs both before and after induction. Both MSCs and HSCs have hepatic differentiation capabilities. However, their capabilities are not the same. MSCs can differentiate into mature hepatocyte-like cells, never expressing early hepatic specific genes, while Thy-1.1(+) cells are inclined to differentiate into hepatic stem cell-like cells, with an increasing AFP expression and a decreasing albumin signal. CK18 mRNA is positive in Thy-1.1(+) cells and MSCs, negative in Thy-1.1(-) cells. It seems that CK18 has some relationship with Thy-1.1 antigen, and CK18 may be a predictive marker of hepatic differentiation capability.

Long noncoding RNA H19 mediates LCoR to impact the osteogenic and adipogenic differentiation of mBMSCs in mice through sponging miR-188.

PubMed

Wang, Yijun; Liu, Wentao; Liu, Yadong; Cui, Jianli; Zhao, Zhiwei; Cao, Hui; Fu, Zhuo; Liu, Bin

2018-04-16

The research aimed to examine the expression of lncRNA H19, miR-188, and LCoR in mouse bone marrow stromal stem cells (mBMSCs), and to investigate the regulatory mechanism of lncRNA H19/miR-188/LCoR in osteogenic and adipogenic differentiation of mBMSCs. The expression of miR-188 in mBMSCs and osteogenesis induced mBMSCs was detected by stem-loop RT-PCR, while the expression of H19 and LCoR in mBMSCs and adipogenesis induced mBMSCs was examined by qRT-PCR. Luciferase reporter assay verified the targeted relationship between miR-188 and H19 or LCoR. Cell proliferation ability was determined by MTT assay, while cell surface markers of mBMSCs were analyzed via flow cytometry. Alkaline phosphatase staining and Alizarin red staining was utilized to detect the osteogenic differentiation capability of mBMSCs, whereas Oil red O staining was applied to examine the ability of adipogenic differentiation of mBMSCs. The expression of miR-188 was lower in osteogenesis induced mBMSCs compared with normal mBMSCs, while H19 and LCoR were downregulated in adipogenic induced mBMSCs. Si-H19 could significantly increase the mRNA level of miR-188. Meanwhile, miR-188 directly regulated LCoR in mBMSCs. Overexpression of miR-188 and knockdown of LCoR suppressed osteogenic differentiation and induced adipogenic differentiation in mBMSCs. Long noncoding RNA H19 mediates LCoR to regulate the balance between osteogenic and adipogenic differentiation of mBMSCs in mice through sponging miR-188. © 2018 Wiley Periodicals, Inc.

Low Intensity Pulsed Ultrasound (LIPUS) Influences the Multilineage Differentiation of Mesenchymal Stem and Progenitor Cell Lines through ROCK-Cot/Tpl2-MEK-ERK Signaling Pathway*

PubMed Central

Kusuyama, Joji; Bandow, Kenjiro; Shamoto, Mitsuo; Kakimoto, Kyoko; Ohnishi, Tomokazu; Matsuguchi, Tetsuya

2014-01-01

Mesenchymal stem cells (MSCs) are pluripotent cells that can differentiate into multilineage cell types, including adipocytes and osteoblasts. Mechanical stimulus is one of the crucial factors in regulating MSC differentiation. However, it remains unknown how mechanical stimulus affects the balance between adipogenesis and osteogenesis. Low intensity pulsed ultrasound (LIPUS) therapy is a clinical application of mechanical stimulus and facilitates bone fracture healing. Here, we applied LIPUS to adipogenic progenitor cell and MSC lines to analyze how multilineage cell differentiation was affected. We found that LIPUS suppressed adipogenic differentiation of both cell types, represented by impaired lipid droplet appearance and decreased gene expression of peroxisome proliferator-activated receptor γ2 (Pparg2) and fatty acid-binding protein 4 (Fabp4). LIPUS also down-regulated the phosphorylation level of peroxisome proliferator-activated receptor γ2 protein, inhibiting its transcriptional activity. In contrast, LIPUS promoted osteogenic differentiation of the MSC line, characterized by increased cell calcification as well as inductions of runt-related transcription factor 2 (Runx2) and Osteocalcin mRNAs. LIPUS induced phosphorylation of cancer Osaka thyroid oncogene/tumor progression locus 2 (Cot/Tpl2) kinase, which was essential for the phosphorylation of mitogen-activated kinase kinase 1 (MEK1) and p44/p42 extracellular signal-regulated kinases (ERKs). Notably, effects of LIPUS on both adipogenesis and osteogenesis were prevented by a Cot/Tpl2-specific inhibitor. Furthermore, effects of LIPUS on MSC differentiation as well as Cot/Tpl2 phosphorylation were attenuated by the inhibition of Rho-associated kinase. Taken together, these results indicate that mechanical stimulus with LIPUS suppresses adipogenesis and promotes osteogenesis of MSCs through Rho-associated kinase-Cot/Tpl2-MEK-ERK signaling pathway. PMID:24550383

Low intensity pulsed ultrasound (LIPUS) influences the multilineage differentiation of mesenchymal stem and progenitor cell lines through ROCK-Cot/Tpl2-MEK-ERK signaling pathway.

PubMed

Kusuyama, Joji; Bandow, Kenjiro; Shamoto, Mitsuo; Kakimoto, Kyoko; Ohnishi, Tomokazu; Matsuguchi, Tetsuya

2014-04-11

Mesenchymal stem cells (MSCs) are pluripotent cells that can differentiate into multilineage cell types, including adipocytes and osteoblasts. Mechanical stimulus is one of the crucial factors in regulating MSC differentiation. However, it remains unknown how mechanical stimulus affects the balance between adipogenesis and osteogenesis. Low intensity pulsed ultrasound (LIPUS) therapy is a clinical application of mechanical stimulus and facilitates bone fracture healing. Here, we applied LIPUS to adipogenic progenitor cell and MSC lines to analyze how multilineage cell differentiation was affected. We found that LIPUS suppressed adipogenic differentiation of both cell types, represented by impaired lipid droplet appearance and decreased gene expression of peroxisome proliferator-activated receptor γ2 (Pparg2) and fatty acid-binding protein 4 (Fabp4). LIPUS also down-regulated the phosphorylation level of peroxisome proliferator-activated receptor γ2 protein, inhibiting its transcriptional activity. In contrast, LIPUS promoted osteogenic differentiation of the MSC line, characterized by increased cell calcification as well as inductions of runt-related transcription factor 2 (Runx2) and Osteocalcin mRNAs. LIPUS induced phosphorylation of cancer Osaka thyroid oncogene/tumor progression locus 2 (Cot/Tpl2) kinase, which was essential for the phosphorylation of mitogen-activated kinase kinase 1 (MEK1) and p44/p42 extracellular signal-regulated kinases (ERKs). Notably, effects of LIPUS on both adipogenesis and osteogenesis were prevented by a Cot/Tpl2-specific inhibitor. Furthermore, effects of LIPUS on MSC differentiation as well as Cot/Tpl2 phosphorylation were attenuated by the inhibition of Rho-associated kinase. Taken together, these results indicate that mechanical stimulus with LIPUS suppresses adipogenesis and promotes osteogenesis of MSCs through Rho-associated kinase-Cot/Tpl2-MEK-ERK signaling pathway.

Systematic Identification, Characterization and Target Gene Analysis of microRNAs Involved in Osteoarthritis Subchondral Bone Pathogenesis.

PubMed

Prasadam, Indira; Batra, Jyotsna; Perry, Samuel; Gu, Wenyi; Crawford, Ross; Xiao, Yin

2016-07-01

This study aimed to identify the microRNAs associated with sclerotic status of subchondral bone in the pathogenesis of osteoarthritis (OA). Total RNA was extracted from non-sclerotic and sclerotic OA subchondral bone from patients undergoing knee replacement surgeries. miRCURY™ LNA miRNA chip and qRT-PCR were used to profile and validate differential microRNA expression. In addition, we further confirmed profiles of altered miRNAs in an OA rat meniscectomy animal model and their putative targets of the miRNAs were predicted using ingenuity (IPA) software. Finally, five short-listed miRNAs were reactivated by transient in vitro overexpression (miRNA mimics) in subchondral bone osteoblasts and their phenotypes were assessed. Functional screening identified 30 differentiated miRNAs in sclerotic subchondral bone compared to non-sclerotic bone of OA patients. Data integration resulted in confirmation of the eight miRNAs, with aberrant expression in independent human OA bone sample set. In silico analysis (IPA) identified 732 mRNA transcripts as putative targets of the eight altered miRNAs, of which twenty genes were validated to be differentially expressed in sclerotic compared to non-sclerotic bone samples. Out of eight dysregulated miRNA's, five of them showed consistent time-dependent downregulation in a rat OA model. Furthermore, synthetic miR-199a-3p, miR-199a-5p, miR-590-5p, and miR-211-5p mimics rescued the abnormal osteoarthritic subchondral bone osteoblast gene expression and mineralization. We have identified four novel miRNAs that play important roles in subchondral bone pathogenesis in OA. Additional studies are required to develop these miRNAs into therapeutic modalities for OA.

Influence of different modifications of a calcium phosphate bone cement on adhesion, proliferation, and osteogenic differentiation of human bone marrow stromal cells.

PubMed

Vater, Corina; Lode, Anja; Bernhardt, Anne; Reinstorf, Antje; Heinemann, Christiane; Gelinsky, Michael

2010-03-15

Collagen and noncollagenous proteins of the extracellular bone matrix are able to stimulate bone cell activities and bone healing. The modification of calcium phosphate bone cements used as temporary bone replacement materials with these proteins seems to be a promising approach to accelerate new bone formation. In this study, we investigated adhesion, proliferation, and osteogenic differentiation of human bone marrow stromal cells (hBMSC) on Biocement D/collagen composites which have been modified with osteocalcin and O-phospho-L-serine. Modification with osteocalcin was carried out by its addition to the cement precursor before setting as well as by functionalization of the cement samples after setting and sterilization. hBMSC were cultured on these samples for 28 days with and without osteogenic supplements. We found a positive impact especially of the phosphoserine-modifications but also of both osteocalcin-modifications on differentiation of hBMSC indicated by higher expression of the osteoblastic markers matrix metalloproteinase-13 and bone sialo protein II. For hBMSC cultured on phosphoserine-containing composites, an increased proliferation has been observed. However, in case of the osteocalcin-modified samples, only osteocalcin adsorbed after setting and sterilization of the cement samples was able to promote initial adhesion and proliferation of hBMSC. The addition of osteocalcin before setting results in a finer microstructure but the biological activity of osteocalcin might be impaired due to the sterilization process. Thus, our data indicate that the initial adhesion and proliferation of hBMSC is enhanced rather by the biological activity of osteocalcin than by the finer microstructure. (c) 2009 Wiley Periodicals, Inc.

An essential role for the association of CD47 to SHPS-1 in skeletal remodeling.

PubMed

Maile, Laura A; DeMambro, Victoria E; Wai, Christine; Lotinun, Sutada; Aday, Ariel W; Capps, Byron E; Beamer, Wesley G; Rosen, Clifford J; Clemmons, David R

2011-09-01

Integrin-associated protein (IAP/CD47) has been implicated in macrophage-macrophage fusion. To understand the actions of CD47 on skeletal remodeling, we compared Cd47(-/-) mice with Cd47(+/+) controls. Cd47(-/-) mice weighed less and had decreased areal bone mineral density compared with controls. Cd47(-/-) femurs were shorter in length with thinner cortices and exhibited lower trabecular bone volume owing to decreased trabecular number and thickness. Histomorphometry revealed reduced bone-formation and mineral apposition rates, accompanied by decreased osteoblast numbers. No differences in osteoclast number were observed despite a nonsignificant but 40% decrease in eroded surface/bone surface in Cd47(-/-) mice. In vitro, the number of functional osteoclasts formed by differentiating Cd47(-/-) bone marrow cells was significantly decreased compared with wild-type cultures and was associated with a decrease in bone-resorption capacity. Furthermore, by disrupting the CD47-SHPS-1 association, we found that osteoclastogenesis was markedly impaired. Assays for markers of osteoclast maturation suggested that the defect was at the point of fusion and not differentiation and was associated with a lack of SHPS-1 phosphorylation, SHP-1 phosphatase recruitment, and subsequent dephosphorylation of non-muscle cell myosin IIA. We also demonstrated a significant decrease in osteoblastogenesis in bone marrow stromal cells derived from Cd47(-/-) mice. Our finding of cell-autonomous defects in Cd47(-/-) osteoblast and osteoclast differentiation coupled with the pronounced skeletal phenotype of Cd47(-/-) mice support the conclusion that CD47 plays an important role in regulating skeletal acquisition and maintenance through its actions on both bone formation and bone resorption. Copyright © 2011 American Society for Bone and Mineral Research.

A novel rat fibrosarcoma cell line from transformed bone marrow-derived mesenchymal stem cells with maintained in vitro and in vivo stemness properties

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Meng-Yu; Nestvold, Janne, E-mail: j.m.nestvold@medisin.uio.no; Rekdal, Øystein

Increasing evidence suggests a possible relationship between mesenchymal stem cells (MSCs) and sarcoma. MSCs are hypothesized to be the cells initiating sarcomagenesis, and cancer stem cells (CSCs) sharing features of MSCs have been identified in sarcomas. Here, we report on the characteristics of a bone marrow-derived rat mesenchymal stem cell line that spontaneously transformed in long-term culture. The rat transformed mesenchymal stem cells (rTMSCs) produced soft-tissue fibrosarcomas in immunocompromised mice and immunocompetent rats. In vitro, the rTMSCs displayed increased proliferation capacity compared to the untransformed cell line. The transformed MSCs maintained the mesenchymal phenotype by expression of the stem cellmore » marker CD 90 and the lack of hematopoietic and endothelial markers. Cytogenetic analysis detected trisomy 6 in the rTMSCs. Side population (SP) isolation and tumorsphere cultivation of the transformed cells confirmed the presence of CSCs among the rTMSCs. Importantly, the rTMSCs retained their differentiation capacity towards osteogenic and adipogenic lineages. This transformed MSC-based cell line may be valuable in examining the balance in a mixed cell population between cancer stem cell properties and the ability to differentiate to specific non-transformed cell populations. Moreover, it may also be a useful tool to evaluate the efficacy of novel targeted immunotherapies in vivo. - Highlights: • Spontaneously transformed rat MSCs (rTMSCs) share characteristics with normal MSCs. • rTMSCs possess a side population, enriched with tumorigenic cells. • rTMSCs model fibrosarcoma in vivo.« less

Calcium and vitamin D for bone health in adults

USDA-ARS?s Scientific Manuscript database

The calcium intake requirement is challenging to determine, and the IOM recommendations are based largely on calcium balance studies. The IOM recommends a calcium intake of 1000-1200 mg per day for older adults to support the preservation of bone mass. Food sources of calcium are preferred because h...

MACF1 Overexpression by Transfecting the 21 kbp Large Plasmid PEGFP-C1A-ACF7 Promotes Osteoblast Differentiation and Bone Formation.

PubMed

Zhang, Yan; Yin, Chong; Hu, Lifang; Chen, Zhihao; Zhao, Fan; Li, Dijie; Ma, Jianhua; Ma, Xiaoli; Su, Peihong; Qiu, Wuxia; Yang, Chaofei; Wang, Pai; Li, Siyu; Zhang, Ge; Wang, Liping; Qian, Airong; Xian, Cory J

2018-02-01

Microtubule actin crosslinking factor 1 (MACF1) is a large spectraplakin protein known to have crucial roles in regulating cytoskeletal dynamics, cell migration, growth, and differentiation. However, its role and action mechanism in bone remain unclear. The present study investigated optimal conditions for effective transfection of the large plasmid PEGFP-C1A-ACF7 (∼21 kbp) containing full-length human MACF1 cDNA, as well as the potential role of MACF1 in bone formation. To enhance MACF1 expression, the plasmid was transfected into osteogenic cells by electroporation in vitro and into mouse calvaria with nanoparticles. Then, transfection efficiency, osteogenic marker expression, calvarial thickness, and bone formation were analyzed. Notably, MACF1 overexpression triggered a drastic increase in osteogenic gene expression, alkaline phosphatase activity, and matrix mineralization in vitro. Mouse calvarial thickness, mineral apposition rate, and osteogenic marker protein expression were significantly enhanced by local transfection. In addition, MACF1 overexpression promoted β-catenin expression and signaling. In conclusion, MACF1 overexpression by transfecting the large plasmid containing full-length MACF1 cDNA promotes osteoblast differentiation and bone formation via β-catenin signaling. Current data will provide useful experimental parameters for the transfection of large plasmids and a novel strategy based on promoting bone formation for prevention and therapy of bone disorders.

Epigenetically Modified Bone Marrow Stromal Cells in Silk Scaffolds Promote Craniofacial Bone Repair and Wound Healing.

PubMed

Han, Qianqian; Yang, Pishan; Wu, Yuwei; Meng, Shu; Sui, Lei; Zhang, Lan; Yu, Liming; Tang, Yin; Jiang, Hua; Xuan, Dongying; Kaplan, David L; Kim, Sung Hoon; Tu, Qisheng; Chen, Jake

2015-08-01

Epigenetic regulation of gene expression is a central mechanism that governs cell stemness, determination, commitment, and differentiation. It has been recently found that PHF8, a major H4K20/H3K9 demethylase, plays a critical role in craniofacial and bone development. In this study, we hypothesize that PHF8 promotes osteoblastogenesis by epigenetically regulating the expression of a nuclear matrix protein, special AT-rich sequence-binding protein 2 (SATB2) that plays pivotal roles in skeletal patterning and osteoblast differentiation. Our results showed that expression levels of PHF8 and SATB2 in preosteoblasts and bone marrow stromal cells (BMSCs) increased simultaneously during osteogenic induction. Overexpressing PHF8 in these cells upregulated the expression of SATB2, Runx2, osterix, and bone matrix proteins. Conversely, knockdown of PHF8 reduced the expression of these genes. Furthermore, ChIP assays confirmed that PHF8 specifically bound to the transcription start site (TSS) of the SATB2 promoter, and the expression of H3K9me1 at the TSS region of SATB2 decreased in PHF8 overexpressed group. Implantation of the BMSCs overexpressing PHF8 with silk protein scaffolds promoted bone regeneration in critical-sized defects in mouse calvaria. Taken together, our results demonstrated that PHF8 epigenetically modulates SATB2 activity, triggering BMSCs osteogenic differentiation and facilitating bone formation and regeneration in biodegradable silk scaffolds.

Three-Dimensional Mechanical Loading Modulates the Osteogenic Response of Mesenchymal Stem Cells to Tumor-Derived Soluble Signals

PubMed Central

Lynch, Maureen E.; Chiou, Aaron E.; Lee, Min Joon; Marcott, Stephen C.; Polamraju, Praveen V.; Lee, Yeonkyung

2016-01-01

Dynamic mechanical loading is a strong anabolic signal in the skeleton, increasing osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BM-MSCs) and increasing the bone-forming activity of osteoblasts, but its role in bone metastatic cancer is relatively unknown. In this study, we integrated a hydroxyapatite-containing three-dimensional (3D) scaffold platform with controlled mechanical stimulation to investigate the effects of cyclic compression on the interplay between breast cancer cells and BM-MSCs as it pertains to bone metastasis. BM-MSCs cultured within mineral-containing 3D poly(lactide-co-glycolide) (PLG) scaffolds differentiated into mature osteoblasts, and exposure to tumor-derived soluble factors promoted this process. When BM-MSCs undergoing osteogenic differentiation were exposed to conditioned media collected from mechanically loaded breast cancer cells, their gene expression of osteopontin was increased. This was further enhanced when mechanical compression was simultaneously applied to BM-MSCs, leading to more uniformly deposited osteopontin within scaffold pores. These results suggest that mechanical loading of 3D scaffold-based culture models may be utilized to evaluate the role of physiologically relevant physical cues on bone metastatic breast cancer. Furthermore, our data imply that cyclic mechanical stimuli within the bone microenvironment modulate interactions between tumor cells and BM-MSCs that are relevant to bone metastasis. PMID:27401765

Zebrafish sp7 mutants show tooth cycling independent of attachment, eruption and poor differentiation of teeth.

PubMed

Kague, E; Witten, P E; Soenens, M; Campos, C L; Lubiana, T; Fisher, S; Hammond, C; Brown, K Robson; Passos-Bueno, M R; Huysseune, A

2018-03-15

The capacity to fully replace teeth continuously makes zebrafish an attractive model to explore regeneration and tooth development. The requirement of attachment bone for the appearance of replacement teeth has been hypothesized but not yet investigated. The transcription factor sp7 (osterix) is known in mammals to play an important role during odontoblast differentiation and root formation. Here we study tooth replacement in the absence of attachment bone using sp7 zebrafish mutants. We analysed the pattern of tooth replacement at different stages of development and demonstrated that in zebrafish lacking sp7, attachment bone is never present, independent of the stage of tooth development or fish age, yet replacement is not interrupted. Without bone of attachment we observed abnormal orientation of teeth, and abnormal connection of pulp cavities of predecessor and replacement teeth. Mutants lacking sp7 show arrested dentinogenesis, with non-polarization of odontoblasts and only a thin layer of dentin deposited. Osteoclast activity was observed in sp7 mutants; due to the lack of bone of attachment, remodelling was diminished but nevertheless present along the pharyngeal bone. We conclude that tooth replacement is ongoing in the sp7 mutant despite poor differentiation and defective attachment. Without bone of attachment tooth orientation and pulp organization are compromised. Copyright © 2018 Elsevier Inc. All rights reserved.

Spinal bone marrow necrosis with vertebral compression fracture: differentiation of BMN from AVN.

PubMed

Nix, J S; Fitzgerald, R T; Samant, R S; Harrison, M; Angtuaco, E J

2014-09-01

Bone marrow necrosis (BMN) is a rare malignancy-associated hematologic disorder characterized by necrosis of myeloid and stromal marrow elements with preservation of cortical bone. Overlap between the imaging appearances of BMN and avascular necrosis (AVN) raises the potential for diagnostic confusion. We report a case of BMN presenting with a traumatic multi-level vertebral body collapse, and finding that may potentially confound distinction between the two entities. We discuss important pathophysiologic, clinical, and radiologic differences between BMN and AVN with emphasis on features important in the differential diagnosis.

A single nucleotide polymorphism in osteonectin 3’ untranslated region regulates bone volume and is targeted by miR-433

PubMed Central

Dole, Neha S.; Kapinas, Kristina; Kessler, Catherine B.; Yee, Siu-Pok; Adams, Douglas J.; Pereira, Renata C.; Delany, Anne M.

2014-01-01

Osteonectin/SPARC is one of the most abundant non-collagenous extracellular matrix proteins in bone, regulating collagen fiber assembly and promoting osteoblast differentiation. Osteonectin-null and –haploinsufficient mice have low turnover osteopenia, indicating that osteonectin contributes to normal bone formation. In male idiopathic osteoporosis patients, osteonectin 3’ UTR single nucleotide polymorphism (SNP) haplotypes that differed only at SNP1599 (rs1054204) were previously associated with bone mass. Haplotype A (containing SNP1599G) was more frequent in severely affected patients, whereas haplotype B (containing SNP1599C) was more frequent in less affected patients and healthy controls. We hypothesized that SNP1599 contributes to variability in bone mass by modulating osteonectin levels. Osteonectin 3’UTR reporter constructs demonstrated that haplotype A has a repressive effect on gene expression compared to B. We found that SNP1599G contributed to a miR-433 binding site and miR-433 inhibitor relieved repression of the haplotype A, but not B, 3’ UTR reporter construct. We tested our hypothesis in vivo, using a knock-in approach to replace the mouse osteonectin 3’ UTR with human haplotype A or B 3’ UTR. Compared to haplotype A mice, bone osteonectin levels were higher in haplotype B mice. B mice displayed higher bone formation rate and gained more trabecular bone with age. When parathyroid hormone was administered intermittently, haplotype B mice gained more cortical bone area than A mice. Cultured marrow stromal cells from B mice deposited more mineralized matrix and had higher osteocalcin mRNA compared with A mice, demonstrating a cell-autonomous effect on differentiation. Altogether, SNP1599 differentially regulates osteonectin expression and contributes to variability in bone mass, by a mechanism that may involve differential targeting by miR-433. This work validates the findings of the previous candidate gene study, and it assigns a physiological function to a common osteonectin allele, providing support for its role in the complex trait of skeletal phenotype. PMID:25262637

Effect of high sodium intake during 14 days of bed-rest on acid-base balance

NASA Astrophysics Data System (ADS)

Frings, P.; Baecker, N.; Heer, M.

Lowering mechanical load like in microgravity is the dominant stimulus leading to bone loss However high dietary sodium intake is also considered as a risk factor for osteoporosis and thereby might exacerbate the microgravity induced bone loss In a metabolic balance non bed-rest study we have recently shown that a very high sodium intake leads to an increased bone resorption most likely because of a mild metabolic acidosis Frings et al FASEB J 19 5 A1345 2005 To test if mild metabolic acidosis also occurs during immobilization we examined the effect of increased dietary sodium on bone metabolism and acid-base balance in eight healthy male test subjects mean age 26 25 pm 3 49 years body weight 77 98 pm 4 34 kg in our metabolic ward during a 14-day head-down tilt HDT bed-rest study The study was designed as a randomized crossover study with two study periods Each period was divided into three parts 4 ambulatory days with 200 mmol sodium intake 14 days of bed-rest with either 550 mmol or 50 mmol sodium intake and 3 recovery days with 200 mmol sodium intake The sodium intake was altered by variations in dietary sodium chloride content Blood pH P CO2 and P O2 were analyzed in fasting morning fingertip blood samples several times during the entire study Bicarbonate HCO 3 - and base excess BE were calculated according to the Henderson-Hasselbach equation Preliminary results in the acid-base balance from the first study period 4 subjects with 550 mmol and 4 subjects with 50 mmol sodium intake strongly

Bone metabolism in anorexia nervosa and hypothalamic amenorrhea.

PubMed

Chou, Sharon H; Mantzoros, Christos

2018-03-01

Anorexia nervosa (AN) and hypothalamic amenorrhea (HA) are states of chronic energy deprivation associated with severely compromised bone health. Poor bone accrual during adolescence followed by increased bone loss results in lifelong low bone density, degraded bone architecture, and higher risk of fractures, despite recovery from AN/HA. Amenorrhea is only one of several compensatory responses to the negative energy balance. Other hypothalamic-pituitary hormones are affected and contribute to bone deficits, including activation of hypothalamic-pituitary-adrenal axis and growth hormone resistance. Adipokines, particularly leptin, provide information on fat/energy stores, and gut hormones play a role in the regulation of appetite and food intake. Alterations in all these hormones influence bone metabolism. Restricted in scope, current pharmacologic approaches to improve bone health have had overall limited success. Copyright © 2017 Elsevier Inc. All rights reserved.

Space Flight Calcium: Implications for Astronaut Health, Spacecraft Operations, and Earth

PubMed Central

Smith, Scott M.; McCoy, Torin; Gazda, Daniel; Morgan, Jennifer L. L.; Heer, Martina; Zwart, Sara R.

2012-01-01

The space flight environment is known to induce bone loss and, subsequently, calcium loss. The longer the mission, generally the more bone and calcium are lost. This review provides a history of bone and calcium studies related to space flight and highlights issues related to calcium excretion that the space program must consider so that urine can be recycled. It also discusses a novel technique using natural stable isotopes of calcium that will be helpful in the future to determine calcium and bone balance during space flight. PMID:23250146

Space flight calcium: implications for astronaut health, spacecraft operations, and Earth.

PubMed

Smith, Scott M; McCoy, Torin; Gazda, Daniel; Morgan, Jennifer L L; Heer, Martina; Zwart, Sara R

2012-12-18

The space flight environment is known to induce bone loss and, subsequently, calcium loss. The longer the mission, generally the more bone and calcium are lost. This review provides a history of bone and calcium studies related to space flight and highlights issues related to calcium excretion that the space program must consider so that urine can be recycled. It also discusses a novel technique using natural stable isotopes of calcium that will be helpful in the future to determine calcium and bone balance during space flight.

The interrelationship between bone and fat: from cellular see-saw to endocrine reciprocity.

PubMed

Sadie-Van Gijsen, H; Crowther, N J; Hough, F S; Ferris, W F

2013-07-01

The number of mature osteoblasts and marrow adipocytes in bone is influenced by the differentiation of the common mesenchymal progenitor cell towards one phenotype and away from the other. Consequently, factors which promote adipogenesis not only lead to fatty marrow but also inhibit osteoblastogenesis, resulting in decreased osteoblast numbers, diminished bone formation and, potentially, inadequate bone mass and osteoporosis. In addition to osteoblast and bone adipocyte numbers being influenced by this skewing of progenitor cell differentiation towards one phenotype, mature osteoblasts and adipocytes secrete factors which may evoke changes in the cell fate and function of each other. This review examines the endogenous factors, such as PPAR-γ2, Wnt, IGF-1, GH, FGF-2, oestrogen, the GP130 signalling cytokines, vitamin D and glucocorticoids, which regulate the selection between osteoblastogenesis and adipogenesis and the interrelationship between fat and bone. The role of adipokines on bone, such as adiponectin and leptin, as well as adipose-derived oestrogen, is reviewed and the role of bone as an energy regulating endocrine organ is discussed.

Bone microenvironment-mediated resistance of cancer cells to bisphosphonates and impact on bone osteocytes/stem cells.

PubMed

Alasmari, Abeer; Lin, Shih-Chun; Dibart, Serge; Salih, Erdjan

2016-08-01

Anti-resorptive bisphosphonates (BPs) have been clinically used to prevent cancer-bone metastasis and cancer-induced bone pathologies despite the fact that the phenotypic response of the cancer-bone interactions to BP exposure is "uncharted territory". This study offers unique insights into the interplay between cancer stem cells and osteocytes/osteoblasts and mesenchymal stem cells using a three-dimensional (3D) live cancer-bone interactive model. We provide extraordinary cryptic details of the biological events that occur as a result of alendronate (ALN) treatment using 3D live cancer-bone model systems under specific bone remodeling stages. While cancer cells are susceptible to BP treatment in the absence of bone, they are totally unaffected in the presence of bone. Cancer cells colonize live bone irrespective of whether the bone is committed to bone resorption or formation and hence, cancer-bone metastasis/interactions are though to be "independent of bone remodeling stages". In our 3D live bone model systems, ALN inhibited bone resorption at the osteoclast differentiation level through effects of mineral-bound ALN on osteocytes and osteoblasts. The mineral-bound ALN rendered bone incapable of osteoblast differentiation, while cancer cells colonize the bone with striking morphological adaptations which led to a conclusion that a direct anti-cancer effect of BPs in a "live or in vivo" bone microenvironment is implausible. The above studies were complemented with mass spectrometric analysis of the media from cancer-bone organ cultures in the absence and presence of ALN. The mineral-bound ALN impacts the bone organs by limiting transformation of mesenchymal stem cells to osteoblasts and leads to diminished endosteal cell population and degenerated osteocytes within the mineralized bone matrix.

Strontium attenuates rhBMP-2-induced osteogenic differentiation via formation of Sr-rhBMP-2 complex and suppression of Smad-dependent signaling pathway.

PubMed

Zhang, Wenjing; Tian, Yu; He, Hongyan; Chen, Rui; Ma, Yifan; Guo, Han; Yuan, Yuan; Liu, Changsheng

2016-03-01

Strontium (Sr(2+)) has pronounced effects on stimulating bone formation and inhibiting bone resorption in bone regeneration. In this current study, the effect and the underlying mechanism involved of Sr(2+) on the biological activity of bone morphogenetic protein-2 (BMP-2) were studied in detail with pluripotent skeletal muscle myogenic progenitor C2C12 model cell line. The results indicated that Sr(2+) could bind recombinant human BMP-2 (rhBMP-2) rapidly, even in the presence of Ca(2+) and Mg(2+), and inhibited rhBMP-2-induced osteogenic differentiation in vitro and osteogenetic efficiency in vivo. Further studies demonstrated that Sr(2+) treatment undermined the binding capacity of rhBMP-2 with its receptor BMPRIA and thus attenuated Smad 1/5/8 phosphorylation without affecting their dephosphorylation in C2C12 cells. Furthermore, circular dichroism spectroscopy, fluorescence spectroscopy and X-ray photoelectron spectroscopy all revealed that the inhibitory effect of Sr(2+) on the rhBMP-2 osteogenic activity was associated with the formation of Sr-rhBMP-2 complex and ensuing enhancement of β-sheet structure. Our work suggests the activity of rhBMP-2 to induce osteogenic differentiation was decreased by directly interaction with free Sr ions in solution, which should provide guide and assist for development of BMP-2-based materials for bone regeneration. Due to easy denaturation and ensuing the reduced activity of rhBMP-2, preserving/enhancing the capacity of rhBMP-2 to induce osteogenic differentiation is of critical importance in developing the protein-based therapy. Cations as effective elements influence the conformation and thereby the bioactivity of protein. Strontium (Sr(2+)), stimulating bone formation and inhibiting bone resorption, has been incorporated into biomaterials/scaffold to improve the bioactivity for bone-regeneration applications. However, Sr(2+)-induced changes in the conformation and bioactivity of BMP-2 have never been investigated. In this study, the formation of Sr-rhBMP-2 complex inhibited the osteogenic differentiation in vitro and osteogenetic efficiency in vivo through the inhibition of BMP/Smad signaling pathway, providing guidance for development of Sr-containing BMP-2-based bone scaffold/matrice and other Sr-dopped protein therapy. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Hematopoietic Stem Cells in Neural-crest Derived Bone Marrow.

PubMed

Jiang, Nan; Chen, Mo; Yang, Guodong; Xiang, Lusai; He, Ling; Hei, Thomas K; Chotkowski, Gregory; Tarnow, Dennis P; Finkel, Myron; Ding, Lei; Zhou, Yanheng; Mao, Jeremy J

2016-12-21

Hematopoietic stem cells (HSCs) in the endosteum of mesoderm-derived appendicular bones have been extensively studied. Neural crest-derived bones differ from appendicular bones in developmental origin, mode of bone formation and pathological bone resorption. Whether neural crest-derived bones harbor HSCs is elusive. Here, we discovered HSC-like cells in postnatal murine mandible, and benchmarked them with donor-matched, mesoderm-derived femur/tibia HSCs, including clonogenic assay and long-term culture. Mandibular CD34 negative, LSK cells proliferated similarly to appendicular HSCs, and differentiated into all hematopoietic lineages. Mandibular HSCs showed a consistent deficiency in lymphoid differentiation, including significantly fewer CD229 + fractions, PreProB, ProB, PreB and B220 + slgM cells. Remarkably, mandibular HSCs reconstituted irradiated hematopoietic bone marrow in vivo, just as appendicular HSCs. Genomic profiling of osteoblasts from mandibular and femur/tibia bone marrow revealed deficiencies in several HSC niche regulators among mandibular osteoblasts including Cxcl12. Neural crest derived bone harbors HSCs that function similarly to appendicular HSCs but are deficient in the lymphoid lineage. Thus, lymphoid deficiency of mandibular HSCs may be accounted by putative niche regulating genes. HSCs in craniofacial bones have functional implications in homeostasis, osteoclastogenesis, immune functions, tumor metastasis and infections such as osteonecrosis of the jaw.

Stem cells applications in bone and tooth repair and regeneration: New insights, tools, and hopes.

PubMed

Abdel Meguid, Eiman; Ke, Yuehai; Ji, Junfeng; El-Hashash, Ahmed H K

2018-03-01

The exploration of stem and progenitor cells holds promise for advancing our understanding of the biology of tissue repair and regeneration mechanisms after injury. This will also help in the future use of stem cell therapy for the development of regenerative medicine approaches for the treatment of different tissue-species defects or disorders such as bone, cartilages, and tooth defects or disorders. Bone is a specialized connective tissue, with mineralized extracellular components that provide bones with both strength and rigidity, and thus enable bones to function in body mechanical supports and necessary locomotion process. New insights have been added to the use of different types of stem cells in bone and tooth defects over the last few years. In this concise review, we briefly describe bone structure as well as summarize recent research progress and accumulated information regarding the osteogenic differentiation of stem cells, as well as stem cell contributions to bone repair/regeneration, bone defects or disorders, and both restoration and regeneration of bones and cartilages. We also discuss advances in the osteogenic differentiation and bone regeneration of dental and periodontal stem cells as well as in stem cell contributions to dentine regeneration and tooth engineering. © 2017 Wiley Periodicals, Inc.

Mathematical modeling of postmenopausal osteoporosis and its treatment by the anti-catabolic drug denosumab

PubMed Central

Scheiner, S; Pivonka, P; Smith, D W; Dunstan, C R; Hellmich, C

2014-01-01

Denosumab, a fully human monoclonal antibody, has been approved for the treatment of postmenopausal osteoporosis. The therapeutic effect of denosumab rests on its ability to inhibit osteoclast differentiation. Here, we present a computational approach on the basis of coupling a pharmacokinetics model of denosumab with a pharmacodynamics model for quantifying the effect of denosumab on bone remodeling. The pharmacodynamics model comprises an integrated systems biology-continuum micromechanics approach, including a bone cell population model, considering the governing biochemical factors of bone remodeling (including the action of denosumab), and a multiscale micromechanics-based bone mechanics model, for implementing the mechanobiology of bone remodeling in our model. Numerical studies of postmenopausal osteoporosis show that denosumab suppresses osteoclast differentiation, thus strongly curtailing bone resorption. Simulation results also suggest that denosumab may trigger a short-term bone volume gain, which is, however, followed by constant or decreasing bone volume. This evolution is accompanied by a dramatic decrease of the bone turnover rate by more than one order of magnitude. The latter proposes dominant occurrence of secondary mineralization (which is not anymore impeded through cellular activity), leading to higher mineral concentration per bone volume. This explains the overall higher bone mineral density observed in denosumab-related clinical studies. Copyright © 2013 John Wiley & Sons, Ltd. PMID:24039120

Stimulation of Bone Formation in Cortical Bone of Mice Treated with a Receptor Activator of Nuclear Factor-κB Ligand (RANKL)-binding Peptide That Possesses Osteoclastogenesis Inhibitory Activity

PubMed Central

Furuya, Yuriko; Inagaki, Atsushi; Khan, Masud; Mori, Kaoru; Penninger, Josef M.; Nakamura, Midori; Udagawa, Nobuyuki; Aoki, Kazuhiro; Ohya, Keiichi; Uchida, Kohji; Yasuda, Hisataka

2013-01-01

To date, parathyroid hormone is the only clinically available bone anabolic drug. The major difficulty in the development of such drugs is the lack of clarification of the mechanisms regulating osteoblast differentiation and bone formation. Here, we report a peptide (W9) known to abrogate osteoclast differentiation in vivo via blocking receptor activator of nuclear factor-κB ligand (RANKL)-RANK signaling that we surprisingly found exhibits a bone anabolic effect in vivo. Subcutaneous administration of W9 three times/day for 5 days significantly augmented bone mineral density in mouse cortical bone. Histomorphometric analysis showed a decrease in osteoclastogenesis in the distal femoral metaphysis and a significant increase in bone formation in the femoral diaphysis. Our findings suggest that W9 exerts bone anabolic activity. To clarify the mechanisms involved in this activity, we investigated the effects of W9 on osteoblast differentiation/mineralization in MC3T3-E1 (E1) cells. W9 markedly increased alkaline phosphatase (a marker enzyme of osteoblasts) activity and mineralization as shown by alizarin red staining. Gene expression of several osteogenesis-related factors was increased in W9-treated E1 cells. Addition of W9 activated p38 MAPK and Smad1/5/8 in E1 cells, and W9 showed osteogenesis stimulatory activity synergistically with BMP-2 in vitro and ectopic bone formation. Knockdown of RANKL expression in E1 cells reduced the effect of W9. Furthermore, W9 showed a weak effect on RANKL-deficient osteoblasts in alkaline phosphatase assay. Taken together, our findings suggest that this peptide may be useful for the treatment of bone diseases, and W9 achieves its bone anabolic activity through RANKL on osteoblasts accompanied by production of several autocrine factors. PMID:23319583

Transglutaminases factor XIII-A and TG2 regulate resorption, adipogenesis and plasma fibronectin homeostasis in bone and bone marrow

PubMed Central

Mousa, Aisha; Cui, Cui; Song, Aimei; Myneni, Vamsee D; Sun, Huifang; Li, Jin Jin; Murshed, Monzur; Melino, Gerry; Kaartinen, Mari T

2017-01-01

Appropriate bone mass is maintained by bone-forming osteoblast and bone-resorbing osteoclasts. Mesenchymal stem cell (MSC) lineage cells control osteoclastogenesis via expression of RANKL and OPG (receptor activator of nuclear factor κB ligand and osteoprotegerin), which promote and inhibit bone resorption, respectively. Protein crosslinking enzymes transglutaminase 2 (TG2) and Factor XIII-A (FXIII-A) have been linked to activity of myeloid and MSC lineage cells; however, in vivo evidence has been lacking to support their function. In this study, we show in mice that TG2 and FXIII-A control monocyte-macrophage cell differentiation into osteoclasts as well as RANKL production in MSCs and in adipocytes. Long bones of mice lacking TG2 and FXIII-A transglutaminases, show compromised biomechanical properties and trabecular bone loss in axial and appendicular skeleton. This was caused by increased osteoclastogenesis, a cellular phenotype that persists in vitro. The increased potential of TG2 and FXIII-A deficient monocytes to form osteoclasts was reversed by chemical inhibition of TG activity, which revealed the presence of TG1 in osteoclasts and assigned different roles for the TGs as regulators of osteoclastogenesis. TG2- and FXIII-A-deficient mice had normal osteoblast activity, but increased bone marrow adipogenesis, MSCs lacking TG2 and FXIII-A showed high adipogenic potential and significantly increased RANKL expression as well as upregulated TG1 expression. Chemical inhibition of TG activity in the null cells further increased adipogenic potential and RANKL production. Altered differentiation of TG2 and FXIII-A null MSCs was associated with plasma fibronectin (FN) assembly defect in cultures and FN retention in serum and marrow in vivo instead of assembly into bone. Our findings provide new functions for TG2, FXIII-A and TG1 in bone cells and identify them as novel regulators of bone mass, plasma FN homeostasis, RANKL production and myeloid and MSC cell differentiation. PMID:28387755

Effects of Cinnamoyloxy-mammeisin from Geopropolis on Osteoclast Differentiation and Porphyromonas gingivalis-Induced Periodontitis.

PubMed

da Cunha, Marcos Guilherme; Ramos-Junior, Erivan Schnaider; Franchin, Marcelo; Taira, Thaise Mayumi; Beutler, John A; Franco, Gilson Cesar Nobre; Ikegaki, Masaharu; de Alencar, Severino Matias; Fukada, Sandra Yasuyo; Rosalen, Pedro Luiz

2017-06-23

Bone-loss-related diseases such as rheumatoid arthritis, osteomyelitis, osteoporosis, and periodontitis are associated with high rates of morbidity worldwide. These disorders are characterized by an imbalance between the formation and activity of osteoblasts and osteoclasts, leading to bone loss. In this context, we evaluated the effect of cinnamoyloxy-mammeisin (CNM), an anti-inflammatory coumarin found in Melipona scutellaris geopropolis, on key targets related to bone remodeling. In the present study we investigated the in vitro effects of CNM on osteoclast differentiation and M-CSF+RANKL-induced osteoclastogenic marker expression. Additionally, the interference of CNM treatment on osteoclast activity was evaluated by zymography and resorption area. Finally, we assessed the capacity of the compound to mitigate alveolar bone loss in vivo in experimental murine periodontitis induced by Porphyromonas gingivalis. We observed that treatment with CNM impaired osteoclast differentiation, as evidenced by a reduced number of tartrate-resistant acid-phosphatase-positive multinucleated cells (TRAP+) as well as the expression of osteoclastogenic markers upon M-CSF+RANKL-induced stimulation. Similarly, we observed reduced gelatinolytic and resorption capacity in M-CSF+RANKL-induced cells in vitro. Lastly, CNM attenuated alveolar bone loss in an experimental murine periodontitis model. These findings indicate that CNM may be considered a promising treatment for bone loss diseases.

Autogenous bone chips: influence of a new piezoelectric device (Piezosurgery) on chip morphology, cell viability and differentiation.

PubMed

Chiriac, G; Herten, M; Schwarz, F; Rothamel, D; Becker, J

2005-09-01

The aim of the present study was to investigate the influence of a new piezoelectric device, designed for harvesting autogenous bone chips from intra-oral sites, on chip morphology, cell viability and differentiation. A total of 69 samples of cortical bone chips were randomly gained by either (1) a piezoelectric device (PS), or (2) conventional rotating drills (RD). Shape and size of the bone chips were compared by means of morphometrical analysis. Outgrowing osteoblasts were identified by means of alkaline phosphatase activity (AP), immunhistochemical staining for osteocalcin (OC) synthesis and reverse transcriptase-polymerase chain reaction phenotyping. In 88.9% of the RD and 87.9% of the PS specimens, an outgrowth of adherent cells nearby the bone chips was observed after 6-19 days. Confluence of cells was reached after 4 weeks. Positive staining for AP and OC identified the cells as osteoblasts. The morphometrical analysis revealed a statistically significant more voluminous size of the particles collected with PS than RD. Within the limits of the present study, it may be concluded that both the harvesting methods are not different from each other concerning their detrimental effect on viability and differentiation of cells growing out of autogenous bone chips derived from intra-oral cortical sites.

Cordyceps sinensis health supplement enhances recovery from taxol-induced leukopenia.

PubMed

Liu, Wei-Chung; Chuang, Wei-Ling; Tsai, Min-Lung; Hong, Ji-Hong; McBride, William H; Chiang, Chi-Shiun

2008-04-01

This study aimed to evaluate the ability of the health food supplement Cordyceps sinensis (CS) to ameliorate suppressive effects of chemotherapy on bone marrow function as a model for cancer treatment. Mice were treated with Taxol (17 mg/kg body wt) one day before oral administration of a hot-water extract of CS (50 mg/kg daily) that was given daily for 3 weeks. White blood cell counts in peripheral blood of mice receiving Taxol were at 50% of normal levels on day 28 but had recovered completely in mice treated with CS. In vitro assays showed that CS enhanced the colony-forming ability of both granulocyte macrophage colony forming unit (GM-CFU) and osteogenic cells from bone marrow preparations and promoted the differentiation of bone marrow mesenchymal stromal cells into adipocytes, alkaline phosphatase-positive osteoblasts, and bone tissue. This result could be attributed to enhanced expression of Cbfa1 (core binding factor a) and BMP-2 (bone morphogenetic protein) with concurrent suppression of ODF (osteoclast differentiation factor/RANK [receptor activator of NF-kappaB]) ligand. In summary, CS enhances recovery of mice from leukopenia caused by Taxol treatment. It appears to do so by protecting both hematopoietic progenitor cells directly and the bone marrow stem cell niche through its effects on osteoblast differentiation.

Platelet-Rich Fibrin Promotes Periodontal Regeneration and Enhances Alveolar Bone Augmentation

PubMed Central

Li, Qi; Pan, Shuang; Dangaria, Smit J.; Gopinathan, Gokul; Kolokythas, Antonia; Chu, Shunli; Geng, Yajun; Zhou, Yanmin; Luan, Xianghong

2013-01-01

In the present study we have determined the suitability of platelet-rich fibrin (PRF) as a complex scaffold for periodontal tissue regeneration. Replacing PRF with its major component fibrin increased mineralization in alveolar bone progenitors when compared to periodontal progenitors, suggesting that fibrin played a substantial role in PRF-induced osteogenic lineage differentiation. Moreover, there was a 3.6-fold increase in the early osteoblast transcription factor RUNX2 and a 3.1-fold reduction of the mineralization inhibitor MGP as a result of PRF application in alveolar bone progenitors, a trend not observed in periodontal progenitors. Subcutaneous implantation studies revealed that PRF readily integrated with surrounding tissues and was partially replaced with collagen fibers 2 weeks after implantation. Finally, clinical pilot studies in human patients documented an approximately 5 mm elevation of alveolar bone height in tandem with oral mucosal wound healing. Together, these studies suggest that PRF enhances osteogenic lineage differentiation of alveolar bone progenitors more than of periodontal progenitors by augmenting osteoblast differentiation, RUNX2 expression, and mineralized nodule formation via its principal component fibrin. They also document that PRF functions as a complex regenerative scaffold promoting both tissue-specific alveolar bone augmentation and surrounding periodontal soft tissue regeneration via progenitor-specific mechanisms. PMID:23586051

Platelet-rich fibrin promotes periodontal regeneration and enhances alveolar bone augmentation.

PubMed

Li, Qi; Pan, Shuang; Dangaria, Smit J; Gopinathan, Gokul; Kolokythas, Antonia; Chu, Shunli; Geng, Yajun; Zhou, Yanmin; Luan, Xianghong

2013-01-01

In the present study we have determined the suitability of platelet-rich fibrin (PRF) as a complex scaffold for periodontal tissue regeneration. Replacing PRF with its major component fibrin increased mineralization in alveolar bone progenitors when compared to periodontal progenitors, suggesting that fibrin played a substantial role in PRF-induced osteogenic lineage differentiation. Moreover, there was a 3.6-fold increase in the early osteoblast transcription factor RUNX2 and a 3.1-fold reduction of the mineralization inhibitor MGP as a result of PRF application in alveolar bone progenitors, a trend not observed in periodontal progenitors. Subcutaneous implantation studies revealed that PRF readily integrated with surrounding tissues and was partially replaced with collagen fibers 2 weeks after implantation. Finally, clinical pilot studies in human patients documented an approximately 5 mm elevation of alveolar bone height in tandem with oral mucosal wound healing. Together, these studies suggest that PRF enhances osteogenic lineage differentiation of alveolar bone progenitors more than of periodontal progenitors by augmenting osteoblast differentiation, RUNX2 expression, and mineralized nodule formation via its principal component fibrin. They also document that PRF functions as a complex regenerative scaffold promoting both tissue-specific alveolar bone augmentation and surrounding periodontal soft tissue regeneration via progenitor-specific mechanisms.

Euphorbia factor L1 inhibits osteoclastogenesis by regulating cellular redox status and induces Fas-mediated apoptosis in osteoclast.

PubMed

Hong, Seong-Eun; Lee, Jiae; Seo, Dong-Hyun; In Lee, Hye; Ri Park, Doo; Lee, Gong-Rak; Jo, You-Jin; Kim, Narae; Kwon, Minjung; Shon, Hansem; Kyoung Seo, Eun; Kim, Han-Sung; Young Lee, Soo; Jeong, Woojin

2017-11-01

Excessive bone resorption caused by increased osteoclast number or activity leads to a variety of bone diseases including osteoporosis, rheumatoid arthritis and periodontitis. Thus, the therapeutic strategy for these diseases has been focused primarily on the inhibition of osteoclast formation and function. This study shows that euphorbia factor L1 (EFL1), a diterpenoid isolated from Euphorbia lathyris, inhibited osteoclastogenesis and induced osteoclast apoptosis. EFL1 suppressed osteoclast formation and bone resorption at both initial and terminal differentiation stages. EFL1 inhibited receptor activator of NF-κB ligand (RANKL)-induced NFATc1 induction with attenuated NF-κB activation and c-Fos expression. EFL1 decreased the level of reactive oxygen species by scavenging them or activating Nrf2, and inhibited PGC-1β that regulates mitochondria biogenesis. In addition, EFL1 induced apoptosis in differentiated osteoclasts by increasing Fas ligand expression followed by caspase activation. Moreover, EFL1 inhibited inflammation-induced bone erosion and ovariectomy-induced bone loss in mice. These findings suggest that EFL1 inhibits osteoclast differentiation by regulating cellular redox status and induces Fas-mediated apoptosis in osteoclast, and may provide therapeutic potential for preventing or treating bone-related diseases caused by excessive osteoclast. Copyright © 2017 Elsevier Inc. All rights reserved.

Effects of strontium on proliferation and differentiation of rat bone marrow mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Yunfeng; Li, Jihua; Zhu, Songsong

Highlights: Black-Right-Pointing-Pointer Strontium ranelate (SrR) inhibits proliferation of BMMSCs. Black-Right-Pointing-Pointer SrR increases osteoblastic but decreases adipocytic differentiation of BMMSCs. Black-Right-Pointing-Pointer SrR increases expression of Runx2, BSP and OCN by BMMSCs in osteogenic medium. Black-Right-Pointing-Pointer SrR decreases expression of PPAR{gamma}, aP2/ALBP and LPL by BMMSCs in adipogenic medium. -- Abstract: Strontium ranelate (SrR) was an effective anti-osteoporotic drug to increase bone formation and decrease bone resorption. However, reports about the effect of SR on osteoblastic and adipocytic differentiation from bone marrow mesenchymal stem cells (BMMSCs) are limited. The purpose of this study is to evaluate whether SrR affects the ability ofmore » BMMSCs to differentiate into osteoblasts or adipocytes. Rat BMMSCs were identified by flow cytometry and exposed to SR (0.1 and 1.0 mM Sr{sup 2+}) under osteogenic or adipogenic medium for 1 and 2 weeks. The proliferation and differentiation of BMMSCs were analyzed by MTT, alkaline phosphatase (ALP), Oil red O staining, quantitative real-time RT-PCR and Western blot assays. SrR significantly inhibited the proliferation, increased osteoblastic but decreased adipocytic differentiation of rat BMMSCs dose-dependently. In osteogenic medium, SrR increased the expression of ALP, the mRNA levels of Cbfa1/Runx2, bone sialoprotein, and osteocalcin by RT-PCR, and the protein levels of Cbfa1/Runx2 by Western blot. In adipogenic medium, SrR decreased the mRNA levels of PPAR{gamma}2, adipocyte lipid-binding protein 2 (aP2/ALBP), and lipoprotein lipase (LPL) by RT-PCR, and the protein expression of PPAR{gamma} in Western blot analysis. These results indicated that the effects of SrR to promote osteoblastic but inhibit adipocytic differentiation of BMMSCs might contribute to its effect on osteoporosis treatment.« less

Differential analysis of genome-wide methylation and gene expression in mesenchymal stem cells of patients with fractures and osteoarthritis

PubMed Central

del Real, Alvaro; Pérez-Campo, Flor M.; Fernández, Agustín F.; Sañudo, Carolina; Ibarbia, Carmen G.; Pérez-Núñez, María I.; Criekinge, Wim Van; Braspenning, Maarten; Alonso, María A.; Fraga, Mario F.

2017-01-01

ABSTRACT Insufficient activity of the bone-forming osteoblasts leads to low bone mass and predisposes to fragility fractures. The functional capacity of human mesenchymal stem cells (hMSCs), the precursors of osteoblasts, may be compromised in elderly individuals, in relation with the epigenetic changes associated with aging. However, the role of hMSCs in the pathogenesis of osteoporosis is still unclear. Therefore, we aimed to characterize the genome-wide methylation and gene expression signatures and the differentiation capacity of hMSCs from patients with hip fractures. We obtained hMSCs from the femoral heads of women undergoing hip replacement due to hip fractures and controls with hip osteoarthritis. DNA methylation was explored with the Infinium 450K bead array. Transcriptome analysis was done by RNA sequencing. The genomic analyses revealed that most differentially methylated loci were situated in genomic regions with enhancer activity, distant from gene bodies and promoters. These regions were associated with differentially expressed genes enriched in pathways related to hMSC growth and osteoblast differentiation. hMSCs from patients with fractures showed enhanced proliferation and upregulation of the osteogenic drivers RUNX2/OSX. Also, they showed some signs of accelerated methylation aging. When cultured in osteogenic medium, hMSCs from patients with fractures showed an impaired differentiation capacity, with reduced alkaline phosphatase activity and poor accumulation of a mineralized matrix. Our results point to 2 areas of potential interest for discovering new therapeutic targets for low bone mass disorders and bone regeneration: the mechanisms stimulating MSCs proliferation after fracture and those impairing their terminal differentiation. PMID:27982725

Proton density fat fraction (PDFF) MRI for differentiation of benign and malignant vertebral lesions.

PubMed

Schmeel, Frederic Carsten; Luetkens, Julian Alexander; Wagenhäuser, Peter Johannes; Meier-Schroers, Michael; Kuetting, Daniel Lloyd; Feißt, Andreas; Gieseke, Jürgen; Schmeel, Leonard Christopher; Träber, Frank; Schild, Hans Heinz; Kukuk, Guido Matthias

2018-06-01

To investigate whether proton density fat fraction (PDFF) measurements using a six-echo modified Dixon sequence can help to differentiate between benign and malignant vertebral bone marrow lesions. Sixty-six patients were prospectively enrolled in our study. In addition to conventional MRI at 3.0-Tesla including at least sagittal T2-weighted/spectral attenuated inversion recovery and T1-weighted sequences, all patients underwent a sagittal six-echo modified Dixon sequence of the spine. The mean PDFF was calculated using regions of interest and compared between vertebral lesions. A cut-off value of 6.40% in PDFF was determined by receiver operating characteristic curves and used to differentiate between malignant (< 6.40%) and benign (≥ 6.40%) vertebral lesions. There were 77 benign and 44 malignant lesions. The PDFF of malignant lesions was statistically significant lower in comparison with benign lesions (p < 0.001) and normal vertebral bone marrow (p < 0.001). The areas under the curves (AUC) were 0.97 for differentiating benign from malignant lesions (p < 0.001) and 0.95 for differentiating acute vertebral fractures from malignant lesions (p < 0.001). This yielded a diagnostic accuracy of 96% in the differentiation of both benign lesions and acute vertebral fractures from malignancy. PDFF derived from six-echo modified Dixon allows for differentiation between benign and malignant vertebral lesions with a high diagnostic accuracy. • Establishing a diagnosis of indeterminate vertebral lesions is a common clinical problem • Benign bone marrow processes may mimic the signal alterations observed in malignancy • PDFF differentiates between benign and malignant lesions with a high diagnostic accuracy • PDFF of non-neoplastic vertebral lesions is significantly higher than that of malignancy • PDFF from six-echo modified Dixon may help avoid potentially harmful bone biopsy.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodríguez-Fuentes, Nayeli; Rodríguez-Hernández, Ana G.; Enríquez-Jiménez, Juana

Highlights: •Nukbone showed to be a good scaffold for adhesion, proliferation and differentiation of stem cells. •Nukbone induced osteoblastic differentiation of human mesenchymal stem cells. •Results showed that Nukbone offer an excellent option for bone tissue regeneration due to properties. -- Abstract: Bovine bone matrix Nukbone® (NKB) is an osseous tissue-engineering biomaterial that retains its mineral and organic phases and its natural bone topography and has been used as a xenoimplant for bone regeneration in clinics. There are not studies regarding its influence of the NKB in the behavior of cells during the repairing processes. The aim of this researchmore » is to demonstrate that NKB has an osteoinductive effect in human mesenchymal stem cells from amniotic membrane (AM-hMSCs). Results indicated that NKB favors the AM-hMSCs adhesion and proliferation up to 7 days in culture as shown by the scanning electron microscopy and proliferation measures using an alamarBlue assay. Furthermore, as demonstrated by reverse transcriptase polymerase chain reaction, it was detected that two gene expression markers of osteoblastic differentiation: the core binding factor and osteocalcin were higher for AM-hMSCs co-cultured with NKB in comparison with cultivated cells in absence of the biomaterial. As the results indicate, NKB possess the capability for inducing successfully the osteoblastic differentiation of AM-hMSC, so that, NKB is an excellent xenoimplant option for repairing bone tissue defects.« less

Synergistic effect of defined artificial extracellular matrices and pulsed electric fields on osteogenic differentiation of human MSCs.

PubMed

Hess, Ricarda; Jaeschke, Anna; Neubert, Holger; Hintze, Vera; Moeller, Stephanie; Schnabelrauch, Matthias; Wiesmann, Hans-Peter; Hart, David A; Scharnweber, Dieter

2012-12-01

In vivo, bone formation is a complex, tightly regulated process, influenced by multiple biochemical and physical factors. To develop a vital bone tissue engineering construct, all of these individual components have to be considered and integrated to gain an in vivo-like stimulation of target cells. The purpose of the present studies was to investigate the synergistic role of defined biochemical and physical microenvironments with respect to osteogenic differentiation of human mesenchymal stem cells (MSCs). Biochemical microenvironments have been designed using artificial extracellular matrices (aECMs), containing collagen I (coll) and glycosaminoglycans (GAGs) like chondroitin sulfate (CS), or a high-sulfated hyaluronan derivative (sHya), formulated as coatings on three-dimensional poly(caprolactone-co-lactide) (PCL) scaffolds. As part of the physical microenvironment, cells were exposed to pulsed electric fields via transformer-like coupling (TC). Results showed that aECM containing sHya enhanced osteogenic differentiation represented by increases in ALP activity and gene-expression (RT-qPCR) of several bone-related proteins (RUNX-2, ALP, OPN). Electric field stimulation alone did not influence cell proliferation, but osteogenic differentiation was enhanced if osteogenic supplements were provided, showing synergistic effects by the combination of sHya and electric fields. These results will improve the understanding of bone regeneration processes and support the development of effective tissue engineered bone constructs. Copyright © 2012 Elsevier Ltd. All rights reserved.

Cell studies of hybridized carbon nanofibers containing bioactive glass nanoparticles using bone mesenchymal stromal cells

NASA Astrophysics Data System (ADS)

Zhang, Xiu-Rui; Hu, Xiao-Qing; Jia, Xiao-Long; Yang, Li-Ka; Meng, Qing-Yang; Shi, Yuan-Yuan; Zhang, Zheng-Zheng; Cai, Qing; Ao, Yin-Fang; Yang, Xiao-Ping

2016-12-01

Bone regeneration required suitable scaffolding materials to support the proliferation and osteogenic differentiation of bone-related cells. In this study, a kind of hybridized nanofibrous scaffold material (CNF/BG) was prepared by incorporating bioactive glass (BG) nanoparticles into carbon nanofibers (CNF) via the combination of BG sol-gel and polyacrylonitrile (PAN) electrospinning, followed by carbonization. Three types (49 s, 68 s and 86 s) of BG nanoparticles were incorporated. To understand the mechanism of CNF/BG hybrids exerting osteogenic effects, bone marrow mesenchymal stromal cells (BMSCs) were cultured directly on these hybrids (contact culture) or cultured in transwell chambers in the presence of these materials (non-contact culture). The contributions of ion release and contact effect on cell proliferation and osteogenic differentiation were able to be correlated. It was found that the ionic dissolution products had limited effect on cell proliferation, while they were able to enhance osteogenic differentiation of BMSCs in comparison with pure CNF. Differently, the proliferation and osteogenic differentiation were both significantly promoted in the contact culture. In both cases, CNF/BG(68 s) showed the strongest ability in influencing cell behaviors due to its fastest release rate of soluble silicium-relating ions. The synergistic effect of CNF and BG would make CNF/BG hybrids promising substrates for bone repairing.

Aggregatibacter actinomycetemcomitans, a potent immunoregulator of the periodontal host defense system and alveolar bone homeostasis

PubMed Central

Herbert, Bethany A.; Novince, Chad M.; Kirkwood, Keith L.

2015-01-01

Summary Aggregatibacter actinomycetemcomitans is a perio-pathogenic bacteria that has long been associated with localized aggressive periodontitis. The mechanisms of its pathogenicity have been studied in humans and pre-clinical experimental models. Although different serotypes of A. actinomycetemcomitans have differential virulence factor expression, A. actinomycetemcomitans cytolethal distending toxin (CDT), leukotoxin, and lipopolysaccharide (LPS) have been most extensively studied in the context of modulating the host immune response. Following colonization and attachment in the oral cavity, A. actinomycetemcomitans employs CDT, leukotoxin, and LPS to evade host innate defense mechanisms and drive a pathophysiologic inflammatory response. This supra-physiologic immune response state perturbs normal periodontal tissue remodeling/turnover and ultimately has catabolic effects on periodontal tissue homeostasis. In this review, we have divided the host response into two systems: non-hematopoietic and hematopoietic. Non-hematopoietic barriers include epithelium and fibroblasts that initiate the innate immune host response. The hematopoietic system contains lymphoid and myeloid-derived cell lineages that are responsible for expanding the immune response and driving the pathophysiologic inflammatory state in the local periodontal microenvironment. Effector systems and signaling transduction pathways activated and utilized in response to A. actinomycetemcomitans will be discussed to further delineate immune cell mechanisms during A. actinomycetemcomitans infection. Finally, we will discuss the osteo-immunomodulatory effects induced by A. actinomycetemcomitans and dissect the catabolic disruption of balanced osteoclast-osteoblast mediated bone remodeling, which subsequently leads to net alveolar bone loss. PMID:26197893

Interaction between bone marrow stromal cells and neuroblastoma cells leads to a VEGFA-mediated osteoblastogenesis.

PubMed

HaDuong, Josephine H; Blavier, Laurence; Baniwal, Sanjeev K; Frenkel, Baruch; Malvar, Jemily; Punj, Vasu; Sposto, Richard; DeClerck, Yves A

2015-08-15

The potential role of osteoblasts in bone and bone marrow (BM) metastases in neuroblastoma (NBL) remains unclear. In this study, we examined the effect of NBL cells on the osteoblastic differentiation of BM-derived mesenchymal stromal cells (BMMSC). We show that the presence of NBL cells enhanced the osteoblastic differentiation of BMMSC driven by bone morphogenetic protein (BMP)-4, in the absence of any effect on NBL cell proliferation. Expression profiles of BMMSC driven toward osteoblastic differentiation revealed an increase in vascular endothelial growth factor A (Vegfa) expression in the presence of NBL cells. We demonstrated that NBL cells increased BMMSC-derived VEGFA mRNA and protein and that this was enhanced by BMP-4. However, in similar conditions, neither the addition of an mVEGFA blocking antibody nor exogenous recombinant (r) mVEGFA affected osteoblastic differentiation. In contrast, siRNA- mediated knock-down of VEGFA in BMMSC prevented osteoblastic differentiation in BMP-4-treated cocultures, an effect that was not reversed in the presence of rmVEGFA. An analysis of murine bones injected with hNBL cells revealed an increase of mVEGFA producing cells near tumor cells concomitantly with an increase in Vegfa and Runx2 mRNA. This coincided with an increase in osteoclasts, in Rankl/Opg mRNA ratio and with the formation of osteolytic lesions. Thus NBL cells promote osteoblastogenesis in the BM by increasing VEGFA expression in BMMSC. Our study provides a new insight into the role of VEGFA in NBL metastases by pointing to the role of stroma-derived intracrine VEGFA in osteoblastogenesis. © 2015 UICC.

[Scintigraphic findings in a patient with sickle-cell thalassemia and recurrent pain attacks].

PubMed

Mikosch, Peter; Jauk, Barbara; Kaulfersch, Wilhelm; Gallowitsch, Hans-Jürgen; Lind, Peter

2003-01-01

The case of an eight years old African boy who suffers from sickle cell-thalassemia is presented. In the course of the disease frequent pain attacks occurred within the abdomen and extremities, recently also within the trunk. Local pain, at some occasions in combination with local swelling and always positive laboratory parameters for inflammation, hindered a solely clinical differentiation between bone infarcts and osteomyelitis. Bone scintigraphy, eventually in combination with bone marrow scintigraphy, can assist the clinician in the differentiation of aseptic bone infarcts versus secondary osteomyelitis. Based on the presented case scintigraphic results for bone infarcts, osteomyelitis and special scintigraphic pattern seen in sickle cell disease are presented. Furthermore, problems regarding the interpretation of the scintigraphies in relation to the delayed time after the beginning of pain attacks are discussed.

Identification of DNA methylation changes associated with disease progression in subchondral bone with site-matched cartilage in knee osteoarthritis.

PubMed

Zhang, Yanfei; Fukui, Naoshi; Yahata, Mitsunori; Katsuragawa, Yozo; Tashiro, Toshiyuki; Ikegawa, Shiro; Lee, Ming Ta Michael

2016-09-30

Subchondral bone plays a key role in the development of osteoarthritis, however, epigenetics of subchondral bone has not been extensively studied. In this study, we examined the genome-wide DNA methylation profiles of subchondral bone from three regions on tibial plateau representing disease progression using HumanMethylation450 BeadChip to identify progression associated DNA methylation alterations. Significant differential methylated probes (DMPs) and differential methylated genes (DMGs) were identified in the intermediate and late stages and during the transition from intermediate to late stage of OA in the subchondral bone. Over half of the DMPs were hyper-methylated. Genes associated with OA and bone remodeling were identified. DMGs were enriched in morphogenesis and development of skeletal system, and HOX transcription factors. Comparison of DMGs identified in subchondral bone and site-matched cartilage indicated that DNA methylation changes occurred earlier in subchondral bone and identified different methylation patterns at the late stage of OA. However, shared DMPs, DMGs and common pathways that implicated the tissue reparation were also identified. Methylation is one key mechanism to regulate the crosstalk between cartilage and subchondral bone.

Identification of DNA methylation changes associated with disease progression in subchondral bone with site-matched cartilage in knee osteoarthritis

PubMed Central

Zhang, Yanfei; Fukui, Naoshi; Yahata, Mitsunori; Katsuragawa, Yozo; Tashiro, Toshiyuki; Ikegawa, Shiro; Lee, Ming Ta Michael

2016-01-01

Subchondral bone plays a key role in the development of osteoarthritis, however, epigenetics of subchondral bone has not been extensively studied. In this study, we examined the genome-wide DNA methylation profiles of subchondral bone from three regions on tibial plateau representing disease progression using HumanMethylation450 BeadChip to identify progression associated DNA methylation alterations. Significant differential methylated probes (DMPs) and differential methylated genes (DMGs) were identified in the intermediate and late stages and during the transition from intermediate to late stage of OA in the subchondral bone. Over half of the DMPs were hyper-methylated. Genes associated with OA and bone remodeling were identified. DMGs were enriched in morphogenesis and development of skeletal system, and HOX transcription factors. Comparison of DMGs identified in subchondral bone and site-matched cartilage indicated that DNA methylation changes occurred earlier in subchondral bone and identified different methylation patterns at the late stage of OA. However, shared DMPs, DMGs and common pathways that implicated the tissue reparation were also identified. Methylation is one key mechanism to regulate the crosstalk between cartilage and subchondral bone. PMID:27686527

Development of laminated fiber-reinforced nanocomposites for bone regeneration

NASA Astrophysics Data System (ADS)

Xu, Weijie

There have been numerous efforts to develop synthetic and/or natural tissue engineering scaffolds that are suitable for bone regeneration applications to replace autograft and allograft bones. Current biomaterials as a scaffold for bone regeneration are limited by the extent of degradation concurrent with bone formation, mechanical strength, and the extent of osteogenic differentiation of marrow stromal cells migrating from the surrounding tissues. In this project, a novel laminated nanocomposite scaffold is fabricated, consisting of poly (L-lactide ethylene oxide fumarate) (PLEOF) hydrogel reinforced with poly (L-lactic acid) (PLLA) electrospun nanofibers and hydroxyapatite (HA) nanoparticles. PLEOF is a novel in situ crosslinkable macromer synthesized from biocompatible building units which can be functionalized with bioactive peptides like the cell-adhesive Arg--Gly--Asp (RGD) amino acid sequence. The hydrophilicity and degradation rate of the macromer can be tailored to a particular application by controlling the ratio of PEG to PLA blocks in the macromer and the unsaturated fumarate units can be used for in-situ crosslinking. The PLLA nanofibers were electrospun from high molecular weight PLLA. The laminated nanocomposites were fabricated by dry-hand lay up technique followed by compression molding and thermal crosslinking. The laminated nanocomposites were evaluated with respect to degradation, water uptake, mechanical strength, and the extent of osteogenic differentiation of bone marrow stromal (BMS) cells. Laminates with or without HA nanoparticles showed modulus values much higher than that of trabecular bone (50-100 MPa). The effect of laminated nanocomposites on osteogenic differentiation of BMS cells was determined in terms of cell number, ALPase activity and calcium content. Our results demonstrate that grafting RGD peptide and HA nanoparticles to a PLEOF hydrogel reinforced with PLLA nanofibers synergistically enhance osteogenic differentiation of BMS cells. In conclusion, the laminated nanocomposite with controllable degradation characteristics and robust mechanical properties is attractive as a synthetic bone-mimetic matrix for skeletal tissue regeneration.

miR-383 negatively regulates osteoblastic differentiation of bone marrow mesenchymal stem cells in rats by targeting Satb2.

PubMed

Tang, Jianfei; Zhang, Zeng; Jin, Xiangyun; Shi, Huipeng

2018-06-14

Emerging evidence indicates that microRNAs (miRNAs, miRs) play diverse roles in the regulation of biological processes, including osteoblastic differentiation. In this study, we found that miR-383 is a critical regulator of osteoblastic differentiation. We showed that miR-383 was downregulated during osteoblastic differentiation of rat bone marrow mesenchymal stem cells (BMSCs). Overexpression of miR-383 suppressed osteoblastic differentiation of BMSCs by downregulating alkaline phosphatase (ALP), matrix mineralization, and mRNA and protein levels of RUNX2 and OCN, whereas a knockdown of miR-383 promoted osteoblastic differentiation in vitro. The results of in vivo analysis indicated that inhibition of miR-383 expression enhanced the efficacy of new bone formation in a rat calvarial defect model. Mechanistic experiments revealed that special AT-rich-sequence-binding protein 2 (Satb2) was a direct and functional target of miR-383. Knockdown of Satb2 had inhibitory effects resembling those of miR-383 on the osteoblast differentiation of rat BMSCs. In addition, the positive effect of miR-383 suppression on osteoblastic differentiation was apparently abrogated by Satb2 silencing. Collectively, these results indicate that miR-383 plays an inhibitory role in osteogenic differentiation of rat BMSCs and may act by targeting Satb2. Copyright © 2017. Published by Elsevier Inc.

Secreted Clusterin protein inhibits osteoblast differentiation of bone marrow mesenchymal stem cells by suppressing ERK1/2 signaling pathway.

PubMed

Abdallah, Basem M; Alzahrani, Abdullah M; Kassem, Moustapha

2018-05-01

Secreted Clusterin (sCLU, also known as Apolipoprotein J) is an anti-apoptotic glycoprotein involved in the regulation of cell proliferation, lipid transport, extracellular tissue remodeling and apoptosis. sCLU is expressed and secreted by mouse bone marrow-derived skeletal (stromal or mesenchymal) stem cells (mBMSCs), but its functional role in MSC biology is not known. In this study, we demonstrated that Clusterin mRNA expression and protein secretion in conditioned medium increased during adipocyte differentiation and decreased during osteoblast differentiation of mBMSCs. Treatment of mBMSC cultures with recombinant sCLU protein increased cell proliferation and exerted an inhibitory effect on the osteoblast differentiation while stimulated adipocyte differentiation in a dose-dependent manner. siRNA-mediated silencing of Clu expression in mBMSCs reduced adipocyte differentiation and stimulated osteoblast differentiation of mBMSCs. Furthermore, the inhibitory effect of sCLU on the osteoblast differentiation of mBMSCs was mediated by the suppression of extracellular signal-regulated kinase (ERK1/2) phosphorylation. In conclusion, we identified sCLU as a regulator of mBMSCs lineage commitment to osteoblasts versus adipocytes through a mechanism mediated by ERK1/2 signaling. Inhibiting sCLU is a possible therapeutic approach for enhancing osteoblast differentiation and consequently bone formation. Copyright © 2018 Elsevier Inc. All rights reserved.

The osteogenic differentiation of SSEA-4 sub-population of human adipose derived stem cells using silicate nanoplatelets.

PubMed

Mihaila, Silvia M; Gaharwar, Akhilesh K; Reis, Rui L; Khademhosseini, Ali; Marques, Alexandra P; Gomes, Manuela E

2014-11-01

How to surpass in vitro stem cell differentiation, reducing cell manipulation, and lead the in situ regeneration process after transplantation, remains to be unraveled in bone tissue engineering (bTE). Recently, we showed that the combination of human bone marrow stromal cells with bioactive silicate nanoplatelets (sNPs) promotes the osteogenic differentiation without the use of standard osteogenic inductors. Even more, using SSEA-4(+) cell-subpopulations (SSEA-4(+)hASCs) residing within the adipose tissue, as a single-cellular source to obtain relevant cell types for bone regeneration, was also proposed. Herein, sNPs were used to promote the osteogenic differentiation of SSEA-4(+)hASCs. The interactions between SSEA-4(+)hASCs and sNPs, namely the internalization pathway and effect on cells osteogenic differentiation, were evaluated. SNPs below 100 μg/mL showed high cytocompatibility and fast internalization via clathrin-mediated pathway. SNPs triggered an overexpression of osteogenic-related markers (RUNX2, osteopontin, osteocalcin) accompanied by increased alkaline phosphatase activity and deposition of a predominantly collagen-type I matrix. Consequently, a robust matrix mineralization was achieved, covering >90% of the culturing surface area. Overall, we demonstrated the high osteogenic differentiation potential of SSEA-4(+)hASCs, further enhanced by the addition of sNPs in a dose dependent manner. This strategy endorses the combination of an adipose-derived cell-subpopulation with inorganic compounds to achieve bone matrix-analogs with clinical relevance. Copyright © 2014 Elsevier Ltd. All rights reserved.

ATP6V1H regulates the growth and differentiation of bone marrow stromal cells.

PubMed

Li, Lin; Yang, Shaoqing; Zhang, Yanli; Ji, Dongrui; Jin, Zuolin; Duan, Xiaohong

2018-05-18

ATP6V1H encodes subunit H of vacuolar ATPase (V-ATPase) and may regulate osteoclastic function. The deficiency of ATP6V1H caused bone loss in human, mouse and zebrafish. In this report, we identified the mechanisms by which ATP6V1H regulates proliferation and differentiation of bone marrow stromal cells (BMSCs). We found that ATP6V1H was expressed in BMSCs, andAtp6v1h +/- BMSCs exhibited the lower proliferation rate, cell cycle arrest and reduced osteogenic differentiation capacity, as well as the increased adipogenic potentials. Histologic analysis confirmed less bone formation and more fatty degeneration in Atp6v1h +/- mice in the different age groups. Q-PCR analysis revealed that loss of ATP6V1H function downregulated the mRNA level of TGF-β1 receptor, and its binding molecule, subunit β of adaptor protein complex 2 (AP-2), suggesting ATP6V1H regulates the proliferation and differentiation of BMSCs by interacting with TGF-β receptor I and AP-2 complex. Copyright © 2018. Published by Elsevier Inc.

Direct evidence of macrophage differentiation from bone marrow cells in the liver: a possible origin of Kupffer cells.

PubMed

Takezawa, R; Watanabe, Y; Akaike, T

1995-12-01

Controversy has surrounded origin and differentiation of tissue macrophages. We directly demonstrate the differentiation of bone marrow cells into macrophages in the liver in vivo using a cell-labeling fluorescence dye, PKH-26. Bone marrow cells labeled with PKH26 were intravenously injected into syngenic mice, and these cells were tracked by flow cytometric analysis. The majority of the labeled cells were detected only in the liver after 4 days. Interestingly, antigens specific for macrophage lineage cells (F4/80, Fc gamma RII, and CD14) were detected on the liver-accumulated cells only 4 h after the injection. The pattern of the antigen expression changed to that of Kupffer cells (F4/80+, Fc gamma RII+, Mac-1-) after 4 days and remained so thereafter. These labeled cells in the liver were esterase staining-positive and showed phagocytic activity at day 7. The number of labeled cells among the Kupffer cells in the liver increased with days after injection. This indicates that bone marrow cells accumulate in the liver and differentiate into liver macrophages on site. Roles of factors secreted from hepatocytes are also discussed.

Differential magnesium implant corrosion coat formation and contribution to bone bonding.

PubMed

Rahim, Muhammad Imran; Weizbauer, Andreas; Evertz, Florian; Hoffmann, Andrea; Rohde, Manfred; Glasmacher, Birgit; Windhagen, Henning; Gross, Gerhard; Seitz, Jan-Marten; Mueller, Peter P

2017-03-01

Magnesium alloys are presently under investigation as promising biodegradable implant materials with osteoconductive properties. To study the molecular mechanisms involved, the potential contribution of soluble magnesium corrosion products to the stimulation of osteoblastic cell differentiation was examined. However, no evidence for the stimulation of osteoblast differentiation could be obtained when cultured mesenchymal precursor cells were differentiated in the presence of metallic magnesium or in cell culture medium containing elevated magnesium ion levels. Similarly, in soft tissue no bone induction by metallic magnesium or by the corrosion product magnesium hydroxide could be observed in a mouse model. Motivated by the comparatively rapid accumulation solid corrosion products physicochemical processes were examined as an alternative mechanism to explain the stimulation of bone growth by magnesium-based implants. During exposure to physiological solutions a structured corrosion coat formed on magnesium whereby the elements calcium and phosphate were enriched in the outermost layer which could play a role in the established biocompatible behavior of magnesium implants. When magnesium pins were inserted into avital bones, corrosion lead to increases in the pull out force, suggesting that the expanding corrosion layer was interlocking with the surrounding bone. Since mechanical stress is a well-established inducer of bone growth, volume increases caused by the rapid accumulation of corrosion products and the resulting force development could be a key mechanism and provide an explanation for the observed stimulatory effects of magnesium-based implants in hard tissue. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 697-709, 2017. © 2016 Wiley Periodicals, Inc.

Effect of autogenous and fresh-frozen bone grafts on osteoblast differentiation.

PubMed

Ferraz, E P; Xavier, S P; Azevedo, F G; de Oliveira, F S; Beloti, M M; Rosa, A L

2015-01-01

Fresh-frozen bone allograft (FFBA) is an alternative to autogenous bone (AB) for reconstructing maxillary bone. Despite the promising clinical results, cell responses to FFBA and AB were not evaluated. Thus, our aim was to compare cells harvested from maxillary reconstructed sites with either AB or FFBA in terms of osteoblast differentiation and to evaluate the effect of culturing cells in contact with FFBA. Cells harvested from three patients submitted to bilateral maxillary reconstruction with AB and FFBA were cultured to evaluate: proliferation, alkaline phosphatase activity, extracellular matrix mineralization and gene expression of osteoblastic markers. The effect of FFBA on osteoblast differentiation was studied by culturing cells harvested from AB in contact with FFBA and evaluating the same parameters. Data were compared using either two-way ANOVA followed by Tukey-b test or Student's t test (p≤0.05). Cell proliferation was higher in cultures from AB grafted sites and extracellular matrix mineralization was higher in cultures derived from FFBA grafted sites. The gene expression of alkaline phosphatase, RUNX2, bone sialoprotein and osteocalcin was higher in cells derived from FFBA compared with cells from AB grafted sites. However, the exposure of cells derived from AB to FFBA particles did not have any remarkable effect on osteoblast differentiation. These results indicate the higher osteogenic activity of cells derived from FFBA compared with AB reconstructed sites, offering an explanation at cellular level of why FFBA could be a suitable alternative to AB for reconstructing maxillary bone defects. Copyright © 2014 Elsevier Ltd. All rights reserved.

Clonal precursor of bone, cartilage, and hematopoietic niche stromal cells

PubMed Central

Chan, Charles K. F.; Lindau, Paul; Jiang, Wen; Chen, James Y.; Zhang, Lillian F.; Chen, Ching-Cheng; Seita, Jun; Sahoo, Debashis; Kim, Jae-Beom; Lee, Andrew; Park, Sujin; Nag, Divya; Gong, Yongquan; Kulkarni, Subhash; Luppen, Cynthia A.; Theologis, Alexander A.; Wan, Derrick C.; DeBoer, Anthony; Seo, Eun Young; Vincent-Tompkins, Justin D.; Loh, Kyle; Walmsley, Graham G.; Kraft, Daniel L.; Wu, Joseph C.; Longaker, Michael T.; Weissman, Irving L.

2013-01-01

Organs are composites of tissue types with diverse developmental origins, and they rely on distinct stem and progenitor cells to meet physiological demands for cellular production and homeostasis. How diverse stem cell activity is coordinated within organs is not well understood. Here we describe a lineage-restricted, self-renewing common skeletal progenitor (bone, cartilage, stromal progenitor; BCSP) isolated from limb bones and bone marrow tissue of fetal, neonatal, and adult mice. The BCSP clonally produces chondrocytes (cartilage-forming) and osteogenic (bone-forming) cells and at least three subsets of stromal cells that exhibit differential expression of cell surface markers, including CD105 (or endoglin), Thy1 [or CD90 (cluster of differentiation 90)], and 6C3 [ENPEP glutamyl aminopeptidase (aminopeptidase A)]. These three stromal subsets exhibit differential capacities to support hematopoietic (blood-forming) stem and progenitor cells. Although the 6C3-expressing subset demonstrates functional stem cell niche activity by maintaining primitive hematopoietic stem cell (HSC) renewal in vitro, the other stromal populations promote HSC differentiation to more committed lines of hematopoiesis, such as the B-cell lineage. Gene expression analysis and microscopic studies further reveal a microenvironment in which CD105-, Thy1-, and 6C3-expressing marrow stroma collaborate to provide cytokine signaling to HSCs and more committed hematopoietic progenitors. As a result, within the context of bone as a blood-forming organ, the BCSP plays a critical role in supporting hematopoiesis through its generation of diverse osteogenic and hematopoietic-promoting stroma, including HSC supportive 6C3(+) niche cells. PMID:23858471

Combined VEGF and LMP-1 delivery enhances osteoprogenitor cell differentiation and ectopic bone formation.

PubMed

Wang, Xiuli; Cui, Fuai; Madhu, Vedavathi; Dighe, Abhijit S; Balian, Gary; Cui, Quanjun

2011-02-01

A novel strategy to enhance bone repair is to combine angiogenic factors and osteogenic factors. We combined vascular endothelial growth factor (VEGF) and LIM mineralization protein-1 (LMP-1) by using an internal ribosome entry site to link the genes within a single plasmid. We then evaluated the effects on osteoblastic differentiation in vitro and ectopic bone formation in vivo with a subcutaneously placed PLAGA scaffold loaded with a cloned mouse osteoprogenitor cell line, D1, transfected with plasmids containing VEGF and LMP-1 genes. The cells expressing both genes elevated mRNA expression of RunX2 and β-catenin and alkaline phosphatase activity compared to cells from other groups. In vivo, X-ray and micro-CT analysis of the retrieved implants revealed more ectopic bone formation at 2 and 3 weeks but not at 4 weeks compared to other groups. The results indicate that the combination of the therapeutic growth factors potentiates cell differentiation and may promote osteogenesis.

Mediator MED23 cooperates with RUNX2 to drive osteoblast differentiation and bone development.

PubMed

Liu, Zhen; Yao, Xiao; Yan, Guang; Xu, YiChi; Yan, Jun; Zou, Weiguo; Wang, Gang

2016-04-01

How lineage specifiers are regulated during development is an outstanding question, and the molecular regulation of osteogenic factor RUNX2 remains to be fully understood. Here we report that the Mediator subunit MED23 cooperates with RUNX2 to regulate osteoblast differentiation and bone development. Med23 deletion in mesenchymal stem cells or osteoblast precursors results in multiple bone defects similar to those observed in Runx2(+/-) mice. In vitro, Med23-deficient progenitor cells are refractory to osteoblast differentiation, and Med23 deficiency reduces Runx2-target gene activity without changing Runx2 expression. Mechanistically, MED23 binds to RUNX2 and modulates its transcriptional activity. Moreover, Med23 deficiency in osteoprogenitor cells exacerbates the skeletal abnormalities observed in Runx2(+/-) mice. Collectively, our results establish a genetic and physical interaction between RUNX2 and MED23, suggesting that MED23 constitutes a molecular node in the regulatory network of anabolic bone formation and related diseases.

MicroRNA-188 regulates age-related switch between osteoblast and adipocyte differentiation.

PubMed

Li, Chang-Jun; Cheng, Peng; Liang, Meng-Ke; Chen, Yu-Si; Lu, Qiong; Wang, Jin-Yu; Xia, Zhu-Ying; Zhou, Hou-De; Cao, Xu; Xie, Hui; Liao, Er-Yuan; Luo, Xiang-Hang

2015-04-01

Bone marrow mesenchymal stem cells (BMSCs) exhibit an age-dependent reduction in osteogenesis that is accompanied by an increased propensity toward adipocyte differentiation. This switch increases adipocyte numbers and decreases the number of osteoblasts, contributing to age-related bone loss. Here, we found that the level of microRNA-188 (miR-188) is markedly higher in BMSCs from aged compared with young mice and humans. Compared with control mice, animals lacking miR-188 showed a substantial reduction of age-associated bone loss and fat accumulation in bone marrow. Conversely, mice with transgenic overexpression of miR-188 in osterix+ osteoprogenitors had greater age-associated bone loss and fat accumulation in bone marrow relative to WT mice. Moreover, using an aptamer delivery system, we found that BMSC-specific overexpression of miR-188 in mice reduced bone formation and increased bone marrow fat accumulation. We identified histone deacetylase 9 (HDAC9) and RPTOR-independent companion of MTOR complex 2 (RICTOR) as the direct targets of miR-188. Notably, BMSC-specific inhibition of miR-188 by intra-bone marrow injection of aptamer-antagomiR-188 increased bone formation and decreased bone marrow fat accumulation in aged mice. Together, our results indicate that miR-188 is a key regulator of the age-related switch between osteogenesis and adipogenesis of BMSCs and may represent a potential therapeutic target for age-related bone loss.

Cell Mechanisms of Bone Tissue Loss Under Space Flight Conditions

NASA Astrophysics Data System (ADS)

Rodionova, Natalia

Investigations on the space biosatellites has shown that the bone skeleton is one of the most im-portant targets of the effect space flight factors on the organism. Bone tissue cells were studied by electron microscopy in biosamples of rats' long bones flown on the board american station "SLS-2" and in experiments with modelling of microgravity ("tail suspension" method) with using autoradiography. The analysis of data permits to suppose that the processes of remod-eling in bone tissue at microgravity include the following succession of cell-to-cell interactions. Osteocytes as mechanosensory cells are first who respond to a changing "mechanical field". The next stage is intensification of osteolytic processes in osteocytes, leading to a volume en-largement of the osteocytic lacunae and removal of the "excess bone". Then mechanical signals have been transmitted through a system of canals and processes of the osteocytic syncitium to certain superficial bone zones and are perceived by osteoblasts and bone-lining cells (superficial osteocytes), as well as by the bone-marrow stromal cells. The sensitivity of stromal cells, pre-osteoblasts and osteoblasts, under microgravity was shown in a number of works. As a response to microgravity, the system of stromal cells -preosteoblasts -osteoblasts displays retardation of proliferation, differentiation and specific functions of osteogenetic cells. This is supported by the 3H-thymidine studies of the dynamics of differentiation of osteogenetic cells in remodeling zones. But unloading is not adequate and in part of the osteocytes are apoptotic changes as shown by our electron microscopic investigations. An osteocytic apoptosis can play the role in attraction the osteoclasts and in regulation of bone remodeling. The apoptotic bodies with a liquid flow through a system of canals are transferred to the bone surface, where they fulfil the role of haemoattractants for monocytes come here and form osteoclasts. The osteoclasts destroy bone tissue. The macrophages are incorporated into resorption lacunaes and utilize the organic matrix and cellular detritus. The products are secreted to remodeling zones and act as haemoattractants for recruiting and subsequent differentiation here of the osteogenic precursor cells. However, as shown by our results with 3H-glycine, in absence of mechanical stimulus the activization of osteoblastogenesis either doesn't occur, or takes place on a smaller scale. According to our electron-microscopic data a load deficit leads to an adaptive differentiation of fibroblasts and adipocytes in this remodeling zones. This sequence of events is considered as a mechanism of bone tissue loss which underlies the development of osteopenia and osteoporosis under space flight condition.

Project Healthy Bones: An Osteoporosis Prevention Program for Older Adults.

ERIC Educational Resources Information Center

Klotzbach-Shimomura, Kathleen

2001-01-01

Project Healthy Bones is a 24-week exercise and education program for older women and men at risk for or who have osteoporosis. The exercise component is designed to improve strength, balance, and flexibility. The education curriculum stresses the importance of exercise, nutrition, safety, drug therapy, and lifestyle factors. (SK)

[Therapeutic agents for disorders of bone and calcium metabolism--Parathyroid hormone in weekly subcutaneous injection].

PubMed

Uzawa, Toyonobu

2007-01-01

The parathyroid hormone (PTH) that is marketed outside Japan is for daily administration. It has been proven to increase bone mass and prevent fractures, and the effects are very strong. However, data suggest that daily administration of PTH increases bone resorption. By contrast, weekly administration of PTH, which is being developed in Japan, actually decreases bone resorption, and data suggest that this regimen maintains a good balance between bone formation (predominant) and bone resorption. Furthermore, it has been reported that weekly administration of PTH increases bone mass as much as every day administration of PTH, and as such, weekly administration of PTH has the potential to be a useful regimen with characteristics that are different from those of daily administration of PTH.

A reciprocal regulatory interaction between megakaryocytes, bone cells, and hematopoietic stem cells.

PubMed

Kacena, Melissa A; Gundberg, Caren M; Horowitz, Mark C

2006-11-01

A growing body of evidence suggests that megakaryocytes (MK) or their growth factors play a role in skeletal homeostasis. MK have been shown to express and/or secrete several bone-related proteins including osteocalcin, osteonectin, bone sialoprotein, osteopontin, bone morphogenetic proteins, and osteoprotegerin. In addition, at least 3 mouse models have been described in which MK number was significantly elevated with an accompanying marked increase in bone mineral density. Mice overexpressing thrombopoietin, the major MK growth factor, have an osteosclerotic bone phenotype. Mice deficient in transcription factors GATA-1 and NF-E2, which are required for the differentiation of MK, exhibited a strikingly increased bone mass. Importantly, recent studies have demonstrated that MK can stimulate osteoblast (OB) proliferation and differentiation in vitro and that they can also inhibit osteoclast (OC) formation in vitro. These findings suggest that MK play a dual role in skeletal homeostasis by stimulating formation while simultaneously inhibiting resorption. Conversely, cells of the osteoblast lineage support hematopoiesis, including megakaryopoiesis. Postnatal hematopoiesis occurs almost solely in the bone marrow (BM), close to or on endosteal surfaces. This finding, in conjunction with the observed contact of OB with hematopoietic cells, has lead investigators to explore the molecular and cellular interactions between hematopoietic cells and cells of the OB lineage. Importantly, it has been shown that many of the cytokines that are critical for normal hematopoiesis and megakaryopoiesis are produced by OB. Indeed, culturing osteoblasts with CD34+ BM cells significantly enhances hematopoietic cell number by both enhancing the proliferation of long-term culture initiating cells and the proliferation and differentiation of MK. These data are consistent with cells in the OB lineage playing a critical role in the hematopoietic niche. Overall, these observations demonstrate the importance of MK-bone cell interactions in both skeletal homeostasis and hematopoiesis.

The effect of dipeptidyl peptidase-IV inhibition on bone in a mouse model of type 2 diabetes

PubMed Central

Gallagher, Emily Jane; Sun, Hui; Kornhauser, Caroline; Tobin-Hess, Aviva; Epstein, Sol; Yakar, Shoshana; LeRoith, Derek

2017-01-01

Background Individuals with type 2 diabetes (T2D) are at greater risk of bone fractures than those without diabetes. Certain oral diabetic medications may further increase the risk of fracture. Dipeptidyl peptidase-IV (DPP-IV) inhibitors are incretin-based therapies that are being increasingly used for the management of T2D. It has been hypothesized that these agents may reduce fracture risk in those with T2D. In this study, we used a mouse model of T2D to examine the effects of the DPP-IV inhibitor, MK-0626, on bone. Methods Male wild type (WT) and diabetic muscle-lysine-arginine (MKR) mice were treated with MK-0626, pioglitazone, alendronate or vehicle. The effects of treatment with MK-0626 on bone microarchitecture and turnover were compared with treatment with pioglitazone, alendronate and vehicle. Osteoblast differentiation was determined by alkaline phosphatase staining of bone marrow cells from WT and MKR mice after treatment with pioglitazone, MK-0626 or phosphate buffered saline. Results We found that MK-0626 had neutral effects on cortical and trabecular bone in diabetic mice. Pioglitazone had detrimental effects on the trabecular bone of WT but not of diabetic mice. Alendronate caused improvements in cortical and trabecular bone architecture in diabetic and WT mice. MK-0626 did not alter osteoblast differentiation, but pioglitazone impaired osteoblast differentiation in vitro. Conclusions Overall, the DPP-IV inhibitor, MK-0626, had no adverse effects on bone in an animal model of T2D or directly on osteoblasts in culture. These findings are reassuring as DPP-IV inhibitors are being widely used to treat patients with T2D who are already at an increased risk of fractures. PMID:24023014

Use of platelet lysate for bone regeneration - are we ready for clinical translation?

PubMed Central

Altaie, Ala; Owston, Heather; Jones, Elena

2016-01-01

Current techniques to improve bone regeneration following trauma or tumour resection involve the use of autograft bone or its substitutes supplemented with osteoinductive growth factors and/or osteogenic cells such as mesenchymal stem cells (MSCs). Although MSCs are most commonly grown in media containing fetal calf serum, human platelet lysate (PL) offers an effective alternative. Bone marrow - derived MSCs grown in PL-containing media display faster proliferation whilst maintaining good osteogenic differentiation capacity. Limited pre-clinical investigations using PL-expanded MSCs seeded onto osteoconductive scaffolds indicate good potential of such constructs to repair bone in vivo. In an alternative approach, nude PL-coated scaffolds without seeded MSCs have been proposed as novel regenerative medicine devices. Even though methods to coat scaffolds with PL vary, in vitro studies suggest that PL allows for MSC adhesion, migration and differentiation inside these scaffolds. Increased new bone formation and vascularisation in comparison to uncoated scaffolds have also been observed in vivo. This review outlines the state-of-the-art research in the field of PL for ex vivo MSC expansion and in vivo bone regeneration. To minimise inconsistency between the studies, further work is required towards standardisation of PL preparation in terms of the starting material, platelet concentration, leukocyte depletion, and the method of platelet lysis. PL quality control procedures and its “potency” assessment are urgently needed, which could include measurements of key growth and attachment factors important for MSC maintenance and differentiation. Furthermore, different PL formulations could be tailor-made for specific bone repair indications. Such measures would undoubtedly speed up clinical translation of PL-based treatments for bone regeneration. PMID:26981170

Use of platelet lysate for bone regeneration - are we ready for clinical translation?

PubMed

Altaie, Ala; Owston, Heather; Jones, Elena

2016-02-26

Current techniques to improve bone regeneration following trauma or tumour resection involve the use of autograft bone or its substitutes supplemented with osteoinductive growth factors and/or osteogenic cells such as mesenchymal stem cells (MSCs). Although MSCs are most commonly grown in media containing fetal calf serum, human platelet lysate (PL) offers an effective alternative. Bone marrow - derived MSCs grown in PL-containing media display faster proliferation whilst maintaining good osteogenic differentiation capacity. Limited pre-clinical investigations using PL-expanded MSCs seeded onto osteoconductive scaffolds indicate good potential of such constructs to repair bone in vivo. In an alternative approach, nude PL-coated scaffolds without seeded MSCs have been proposed as novel regenerative medicine devices. Even though methods to coat scaffolds with PL vary, in vitro studies suggest that PL allows for MSC adhesion, migration and differentiation inside these scaffolds. Increased new bone formation and vascularisation in comparison to uncoated scaffolds have also been observed in vivo. This review outlines the state-of-the-art research in the field of PL for ex vivo MSC expansion and in vivo bone regeneration. To minimise inconsistency between the studies, further work is required towards standardisation of PL preparation in terms of the starting material, platelet concentration, leukocyte depletion, and the method of platelet lysis. PL quality control procedures and its "potency" assessment are urgently needed, which could include measurements of key growth and attachment factors important for MSC maintenance and differentiation. Furthermore, different PL formulations could be tailor-made for specific bone repair indications. Such measures would undoubtedly speed up clinical translation of PL-based treatments for bone regeneration.

iNOS-Derived Nitric Oxide Stimulates Osteoclast Activity and Alveolar Bone Loss in Ligature-Induced Periodontitis in Rats

PubMed Central

Herrera, Bruno S.; Martins-Porto, Rodrigo; Maia-Dantas, Aline; Campi, Paula; Spolidorio, Luis C.; Costa, Soraia K.P.; Van Dyke, Thomas E.; Gyurko, Robert; Muscara, Marcelo N.

2012-01-01

Background Inflammatory stimuli activate inducible nitric oxide synthase (iNOS) in a variety of cell types, including osteoclasts (OC) and osteoblasts, resulting in sustained NO production. In this study, we evaluate the alveolar bone loss in rats with periodontitis under long-term iNOS inhibition, and the differentiation and activity of OC from iNOS-knockout (KO) mice in vitro. Methods Oral aminoguanidine (an iNOS inhibitor) or water treatment was started 2 weeks before induction of periodontitis. Rats were sacrificed 3, 7, or 14 days after ligature placement, and alveolar bone loss was evaluated. In vitro OC culture experiments were also performed to study the differentiation of freshly isolated bone marrow cells from both iNOS KO and wild-type C57BL/6 mice. OC were counted 6 days later after tartrate-resistant acid phosphatase staining (a marker of osteoclast identity), and bone resorption activity was assessed by counting the number of resorption pits on dentin disks. Results Rats with ligature showed progressive and significant alveolar bone loss compared to sham animals, and aminoguanidine treatment significantly inhibited ligature-induced bone loss at 7 and 14 days after the induction. In comparison to bone marrow cells from wild-type mice, cells from iNOS KO mice showed decreased OC growth and the resulting OC covered a smaller culture dish area and generated fewer resorption pit counts. Conclusion Our results demonstrate that iNOS inhibition prevents alveolar bone loss in a rat model of ligature-induced periodontitis, thus confirming that iNOS-derived NO plays a crucial role in the pathogenesis of periodontitis, probably by stimulating OC differentiation and activity. PMID:21417589

Lactation induces increases in the RANK/RANKL/OPG system in maxillary bone.

PubMed

Macari, Soraia; Sharma, Lavanya A; Wyatt, Amanda; da Silva, Janine Maíra; Dias, George J; Silva, Tarcília A; Szawka, Raphael E; Grattan, David R

2018-05-01

The underlying causes of maxillary bone loss during lactation remain poorly understood. We evaluated the impact of lactation on physiological and mechanically-induced alveolar bone remodeling. Nulliparous non-lactating (N-LAC) and 21-day lactating (LAC) mice underwent mechanically-induced bone remodeling by orthodontic tooth movement (OTM). Micro-computed tomography (microCT) was performed in the maxilla, femur and vertebra. Tartrate-resistant-acid phosphatase (TRAP) and Masson's trichrome labelling was performed in the maxillary bone and gene expression was determined in the periodontal ligament. The effect of prolactin on osteoclast (OCL) and osteoblast (OBL) differentiation was also investigated in N-LAC and LAC mice. Lactation increased alveolar bone loss in the maxilla, femur and vertebra, while OTM was enhanced. The number of OCL and OBL was higher in the maxilla of LAC mice. OTM increased OCL in both groups; while OBL was increased only in N-LAC but not in LAC mice, in which cell numbers were already elevated. The alveolar bone loss during lactation was associated with increased expression of receptor activator of nuclear factor-KappaB (RANK), RANK ligand (RANKL), and osteoprotegerin (OPG) in the maxilla. OTM induced the same responses in N-LAC mice, whereas it had no further effect in LAC mice. Lactation enhanced differentiation of OCL and OBL from bone marrow cells, and prolactin recapitulated OCL differentiation in N-LAC mice. Thus, lactation increases physiological maxillary bone remodeling and OTM, and both require activation of RANK/RANKL/OPG system. These findings expand our knowledge of lactation-induced osteopenia and have possible impact on clinical practice regarding orthodontic treatments and dental implants in lactating women. Copyright © 2018 Elsevier Inc. All rights reserved.

Icaritin, a novel plant-derived osteoinductive agent, enhances the osteogenic differentiation of human bone marrow- and human adipose tissue-derived mesenchymal stem cells.

PubMed

Wu, Tao; Shu, Tao; Kang, Le; Wu, Jinhui; Xing, Jianzhou; Lu, Zhiqin; Chen, Shuxiang; Lv, Jun

2017-04-01

For the treatment of diseases affecting bones using bone regenerative medicine, there is an urgent need to develop safe, inexpensive drugs that can strongly induce bone formation. In the present study, we systematically investigated the effects of icaritin, a metabolic product of icariin, on the osteogenic differentiation of human bone marrow‑derived mesenchymal stem cells (hBMSCs) and human adipose tissue‑derived stem cells (hADSCs) in vitro. After treatment with icaritin at concentrations of 10‑8-10‑5 M, hBMSCs and hADSCs were examined for alkaline phosphatase activity, osteocalcin (OC) secretion, matrix mineralization and expression levels of bone‑related mRNA and proteins. Data showed that icaritin at concentrations 10‑7-10‑5 M significantly increased alkaline phosphatase activity, OC secretion at different time points, and calcium deposition at day 21. In addition, icaritin upregulated the mRNA expression of genes for bone morphogenetic proteins (BMP‑2, ‑4 and ‑7), bone transcription factors (Runx2 and Dlx5) and bone matrix proteins (ALP, OC and Col‑1). Moreover, icaritin increased the protein levels of BMPs, Runx2 and OC, as detected by western blot analysis. These findings suggest that icaritin enhances the osteogenic differentiation of hBMSCS and hADSCs. Icaritin exerts its potent osteogenic effect possibly by directly stimulating the production of BMPs. Although the osteogenic activity of icaritin in vitro was inferior to that of rhBMP‑2, icaritin displayed better results than icariin. Moreover, the low cost, simple extraction procedure, and an abundance of icaritin make it appealing as a bone regenerative medicine.

IL-20 bone diseases involvement and therapeutic target potential.

PubMed

Wang, Hsiao-Hsuan; Hsu, Yu-Hsiang; Chang, Ming-Shi

2018-04-24

Millions of people around the world suffer from bone disorders, likes osteoporosis, rheumatoid arthritis (RA), and cancer-induced osteolysis. In general, the bone remodeling balance is determined by osteoclasts and osteoblasts, respectively responsible for bone resorption and bone formation. Excessive inflammation disturbs the activities of these two kinds of cells, typically resulting in the bone loss. IL-20 is emerging as a potent angiogenic, chemotactic, and proinflammatory cytokine related to several chronic inflammatory disorders likes psoriasis, atherosclerosis, cancer, liver fibrosis, and RA. IL-20 has an important role in the regulation of osteoclastogenesis and osteoblastogenesis and is upregulated in several bone-related diseases. The anti-IL-20 monoclonal antibody treatment has a therapeutic potential in several experimental disease models including ovariectomy-induced osteoporosis, cancer-induced osteolysis, and bone fracture. This review article provides an overview describing the IL-20's biological functions in the common bone disorders and thus providing a novel therapeutic strategy in the future.

Transcriptional regulation of bone sialoprotein gene expression by Osx.

PubMed

Yang, Ya; Huang, Yehong; Zhang, Li; Zhang, Chi

2016-08-05

Osteoporosis is the most common metabolic bone disease characterized by decreased bone mass, decreased bone strength, and increased risk of fracture. It is due to unbalance between bone formation and bone resorption. Bone formation is a complex process which involves the differentiation of mesenchymal stem cells to osteoblasts. Osteoblasts produce a characteristic extracellular collagenous matrix that subsequently becomes mineralized. Osterix (Osx) is an osteoblast-specific transcription factor required for osteoblast differentiation. Bone sialoprotein (Bsp) is a member of the SIBLING gene family. Expression of Bsp correlates with the differentiation of osteoblasts and the onset of mineralization. Our preliminary data showed that Bsp was abolished in Osx-null mice; however, the detailed mechanism of Osx regulation on Bsp is not fully understood. In this study, regulation of Bsp expression by Osx was further characterized. It was shown that overexpression of Osx led to Bsp upregulation. Inhibition of Osx by small interfering RNA resulted in Bsp downregulation in osteoblast. Transfection assay demonstrated that Osx was able to activate Bsp promoter reporter in a dose-dependent manner. To define minimal region of Bsp promoter activated by Osx, a series of deletion mutants of Bsp promoter were generated, and the minimal region was narrowed down to the proximal 100 bp. Point-mutagenesis studies showed that one GC-rich site was required for Bsp promoter activation by Osx. ChIP assays demonstrated that endogenous Osx associated with native Bsp promoter in primary osteoblasts. Our observations provide evidence that Osx targets Bsp expression directly. Copyright © 2016 Elsevier Inc. All rights reserved.

Polymicrobial periodontal pathogens transcriptomes in calvarial bone and soft tissue

PubMed Central

Bakthavatchalu, Vasudevan; Meka, Archana; Mans, Jeffrey J.; Sathishkumar, Sabapathi; Lopez, M. Cecilia; Bhattacharyya, Indraneel; Boyce, Brendan F.; Baker, Henry V.; Lamont, Richard J.; Ebersole, Jeffrey L.; Kesavalu, L.

2011-01-01

Summary Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia are consistently associated with adult periodontitis. This study sought to document the host transcriptome to a P. gingivalis, T. denticola, and T. forsythia challenge as a polymicrobial infection using a murine calvarial model of acute inflammation and bone resorption. Mice were infected with P. gingivalis, T. denticola, and T. forsythia over the calvaria, after which the soft tissues and calvarial bones were excised. A Murine GeneChip® array analysis of transcript profiles showed that 6997 genes were differentially expressed in calvarial bones (P < 0.05) and 1544 genes were differentially transcribed in the inflamed tissues after the polymicrobial infection. Of these genes, 4476 and 1035 genes in the infected bone and tissues were differentially expressed by upregulation. Biological pathways significantly impacted by the polymicrobial infection in calvarial bone included leukocyte transendothelial migration (LTM), cell adhesion molecules, adherens junction, major histocompatibility complex antigen, extracellular matrix-receptor interaction (ECM), and antigen processing and presentation resulting in inflammatory/cytokine/chemokine transcripts stimulation in bone and soft tissue. Intense inflammation and increased activated osteoclasts was observed in calvarias compared to sham-infected controls. Quantitative real-time RT-PCR analysis confirmed mRNA level of selected genes corresponded with the microarray expression. The polymicrobial infection regulated several LTM and extracellular membrane (ECM) pathway genes in a manner distinct from monoinfection with P. gingivalis, T. denticola, or T. forsythia. To our knowledge, this is the first definition of the polymicrobial induced transcriptome in calvarial bone and soft tissue in response to periodontal pathogens. PMID:21896157

Missing Cells: Pathophysiology, Diagnosis, and Management of (Pan)Cytopenia in Childhood

PubMed Central

Erlacher, Miriam; Strahm, Brigitte

2015-01-01

Peripheral blood cytopenia in children can be due to a variety of acquired or inherited diseases. Genetic disorders affecting a single hematopoietic lineage are frequently characterized by typical bone marrow findings, such as lack of progenitors or maturation arrest in congenital neutropenia or a lack of megakaryocytes in congenital amegakaryocytic thrombocytopenia, whereas antibody-mediated diseases such as autoimmune neutropenia are associated with a rather unremarkable bone marrow morphology. By contrast, pancytopenia is frequently associated with a hypocellular bone marrow, and the differential diagnosis includes acquired aplastic anemia, myelodysplastic syndrome, inherited bone marrow failure syndromes such as Fanconi anemia and dyskeratosis congenita, and a variety of immunological disorders including hemophagocytic lymphohistiocytosis. Thorough bone marrow analysis is of special importance for the diagnostic work-up of most patients. Cellularity, cellular composition, and dysplastic signs are the cornerstones of the differential diagnosis. Pancytopenia in the presence of a normo- or hypercellular marrow with dysplastic changes may indicate myelodysplastic syndrome. More challenging for the hematologist is the evaluation of the hypocellular bone marrow. Although aplastic anemia and hypocellular refractory cytopenia of childhood (RCC) can reliably be differentiated on a morphological level, the overlapping pathophysiology remains a significant challenge for the choice of the therapeutic strategy. Furthermore, inherited bone marrow failure syndromes are usually associated with the morphological picture of RCC, and the recognition of these entities is essential as they often present a multisystem disease requiring different diagnostic and therapeutic approaches. This paper gives an overview over the different disease entities presenting with (pan)cytopenia, their pathophysiology, characteristic bone marrow findings, and therapeutic approaches. PMID:26217651

Osteoblast Menin Regulates Bone Mass in Vivo*

PubMed Central

Kanazawa, Ippei; Canaff, Lucie; Abi Rafeh, Jad; Angrula, Aarti; Li, Jingjing; Riddle, Ryan C.; Boraschi-Diaz, Iris; Komarova, Svetlana V.; Clemens, Thomas L.; Murshed, Monzur; Hendy, Geoffrey N.

2015-01-01

Menin, the product of the multiple endocrine neoplasia type 1 (Men1) tumor suppressor gene, mediates the cell proliferation and differentiation actions of transforming growth factor-β (TGF-β) ligand family members. In vitro, menin modulates osteoblastogenesis and osteoblast differentiation promoted and sustained by bone morphogenetic protein-2 (BMP-2) and TGF-β, respectively. To examine the in vivo function of menin in bone, we conditionally inactivated Men1 in mature osteoblasts by crossing osteocalcin (OC)-Cre mice with floxed Men1 (Men1f/f) mice to generate mice lacking menin in differentiating osteoblasts (OC-Cre;Men1f/f mice). These mice displayed significant reduction in bone mineral density, trabecular bone volume, and cortical bone thickness compared with control littermates. Osteoblast and osteoclast number as well as mineral apposition rate were significantly reduced, whereas osteocyte number was increased. Primary calvarial osteoblasts proliferated more quickly but had deficient mineral apposition and alkaline phosphatase activity. Although the mRNA expression of osteoblast marker and cyclin-dependent kinase inhibitor genes were all reduced, that of cyclin-dependent kinase, osteocyte marker, and pro-apoptotic genes were increased in isolated Men1 knock-out osteoblasts compared with controls. In contrast to the knock-out mice, transgenic mice overexpressing a human menin cDNA in osteoblasts driven by the 2.3-kb Col1a1 promoter, showed a gain of bone mass relative to control littermates. Osteoblast number and mineral apposition rate were significantly increased in the Col1a1-Menin-Tg mice. Therefore, osteoblast menin plays a key role in bone development, remodeling, and maintenance. PMID:25538250

Analysis of Mouse Growth Plate Development

PubMed Central

Mangiavini, Laura; Merceron, Christophe; Schipani, Ernestina

2016-01-01

To investigate skeletal development, pathophysiological mechanisms of cartilage and bone disease, and eventually assess innovative treatments, the mouse is a very important resource. During embryonic development, mesenchymal condensations are formed, and cells within these mesenchymal condensations either directly differentiate into osteoblasts and give origin to intramembranous bone, or differentiate into chondrocytes and form a cartilaginous anlage. The cartilaginous anlage or fetal growth plate is then replaced with bone. This process is also called endochondral bone development, and it is responsible for the generation of most of our skeleton. In this Review, we will discuss in detail the most common in vivo and in vitro techniques our laboratory is currently using for the analysis of the mouse fetal growth plate during development. PMID:26928664

Zinc, copper, and nitrogen balances during bed rest and fluoride supplementation in healthy adult males

NASA Technical Reports Server (NTRS)

Krebs, J. M.; Schneider, V. S.; LeBlanc, A. D.

1988-01-01

The effects of bed rest and fluoride supplementation on zinc, copper, and nitrogen balances and Zn and Cu serum levels were measured in 15 healthy males. Subjects aged 19-54 y remained on a metabolic research ward for 10 wk. During weeks 1-5, subjects were ambulatory. During wks 6-10 they remained in continuous bed rest. During weeks 3-10 nine subjects received 10 or 20 mg F/d as sodium fluoride. Daily urine and weekly fecal composites were made and biweekly fasting blood samples were taken. Dietary intakes were 1.40 +/- 0.17 mg Cu/d (22.0 +/- 2.7 mumol Cu/d), 10.82 +/- 0.49 mg Zn/d (165.6 +/- 7.6 mumol Zn/d), and 14.27 +/- 0.23 g N/d (1019 +/- 16 mmol N/d). Bed rest increased urinary Zn and N excretions and fecal Zn excretions and decreased Zn balance (p less than 0.05) whereas Cu balance was unchanged. During bed rest, F supplementation increased Zn and N balances compared with untreated control subjects (p less than 0.05). These results are compatible with bone and muscle atrophy during bed rest and increased bone formation with F supplementation.

Periodontal Biology: Stem Cells, Bmp2 Gene, Transcriptional Enhancers, and Use of Sclerostin Antibody and Pth for Treatment of Periodontal Disease and Bone Loss

PubMed Central

Harris, Stephen E; Rediske, Michael; Neitzke, Rebecca; Rakian, Audrey

2017-01-01

The periodontium is a complex tissue with epithelial components and a complex set of mesodermal derived alveolar bone, cellular and a cellular cementum, and tendon like ligaments (PDL). The current evidence demonstrates that the major pool of periodontal stem cells is derived from a population of micro vascular associated aSMA-positive stem/progenitor (PSC) cells that by lineage tracing form all three major mesodermal derived components of the periodontium. With in vitro aSMA+ stem cells, transcriptome and chip- seq experiments, the gene network and enhancer maps were determined at several differentiation states of the PSC. Current work on the role of the Bmp2 gene in the periodontal stem cell differentiation demonstrated that this Wnt regulated gene, Bmp2, is necessary for differentiation to all three major mesodermal derived component of the periodontium. The mechanism and use of Sclerostin antibody as an activator of Wnt signaling and Bmp2 gene as a potential route to treat craniofacial bone loss is discussed. As well, the mechanism and use of Pth in the treatment of periodontal bone loss or other craniofacial bone loss is presented in this review. PMID:29457146

Dual reporter transgene driven by 2.3Col1a1 promoter is active in differentiated osteoblasts

NASA Technical Reports Server (NTRS)

Marijanovic, Inga; Jiang, Xi; Kronenberg, Mark S.; Stover, Mary Louise; Erceg, Ivana; Lichtler, Alexander C.; Rowe, David W.

2003-01-01

AIM: As quantitative and spatial analyses of promoter reporter constructs are not easily performed in intact bone, we designed a reporter gene specific to bone, which could be analyzed both visually and quantitatively by using chloramphenicol acetyltransferase (CAT) and a cyan version of green fluorescent protein (GFPcyan), driven by a 2.3-kb fragment of the rat collagen promoter (Col2.3). METHODS: The construct Col2.3CATiresGFPcyan was used for generating transgenic mice. Quantitative measurement of promoter activity was performed by CAT analysis of different tissues derived from transgenic animals; localization was performed by visualized GFP in frozen bone sections. To assess transgene expression during in vitro differentiation, marrow stromal cell and neonatal calvarial osteoblast cultures were analyzed for CAT and GFP activity. RESULTS: In mice, CAT activity was detected in the calvaria, long bone, teeth, and tendon, whereas histology showed that GFP expression was limited to osteoblasts and osteocytes. In cell culture, increased activity of CAT correlated with increased differentiation, and GFP activity was restricted to mineralized nodules. CONCLUSION: The concept of a dual reporter allows a simultaneous visual and quantitative analysis of transgene activity in bone.

The inhibitory effect of vitamin K on RANKL-induced osteoclast differentiation and bone resorption.

PubMed

Wu, Wei-Jie; Kim, Min Seuk; Ahn, Byung-Yong

2015-10-01

To further understand the correlation between vitamin K and bone metabolism, the effects of vitamins K1, menaquinone-4 (MK-4), and menaquinone-7 (MK-7) on RANKL-induced osteoclast differentiation and bone resorption were comparatively investigated. Vitamin K2 groups (MK-4 and MK-7) were found to significantly inhibit RANKL-medicated osteoclast cell formation of bone marrow macrophages (BMMs) in a dose-dependent manner, without any evidence of cytotoxicity. The mRNA expression of specific osteoclast differentiation markers, such as c-Fos, NFATc1, OSCAR, and TRAP, as well as NFATc1 protein expression and TRAP activity in RANKL-treated BMMs were inhibited by vitamin K2, although MK-4 exhibited a significantly greater efficiency compared to MK-7. In contrast, the same dose of vitamin K1 had no inhibitory effect on RANKL-induced osteoclast cell formation, but increased the expression of major osteoclastogenic genes. Interestingly, vitamins K1, MK-4 and MK-7 all strongly inhibited osteoclastic bone resorption (p < 0.01) in a dose dependent manner. These results suggest that vitamins K1, MK-4 and MK-7 have anti-osteoporotic properties, while their regulation effects on osteoclastogenesis are somewhat different.

Osteogenic differentiation of human mesenchymal bone marrow cells in silk scaffolds is regulated by nitric oxide.

PubMed

Damoulis, Petros D; Drakos, Dimitrios E; Gagari, Eleni; Kaplan, David L

2007-11-01

Bone marrow-derived mesenchymal stem cells (BMSC) are a powerful tool for tissue engineering and can be used in the regeneration of bone and other tissues. Nitric oxide (NO) produced by the endothelial NO synthase (eNOS) plays an important role in bone development and healing. We hypothesized that NO plays a role in osteogenic differentiation of BMSC cultured in three-dimensional silk scaffolds. eNOS protein was measured by Western Analysis and its activity was assessed by measuring nitrite in culture supernatants. Mineralization was evaluated through calcium deposition and the expression of genes associated with osteogenic differentiation (collagen I, RUNX2, and osteocalcin) was quantified using real-time RT-PCR. eNOS was consistently expressed with minor fluctuations, but NO production significantly increased at later time points (weeks 4 and 5). Addition of a competitive NOS inhibitor (L-NAME) resulted in a modest decrease in calcium deposition, which became statistically significant in week 5. This was preceded by a dramatic decrease in RUNX2 and osteocalcin expression in week 4. These results support our hypothesis and implicate NO as an important player in bone tissue engineering.

Activation of the mitogen-activated protein kinase pathway by bone sialoprotein regulates osteoblast differentiation.

PubMed

Gordon, Jonathan A R; Hunter, Graeme K; Goldberg, Harvey A

2009-01-01

Bone sialoprotein (BSP) is an abundant protein in the extracellular matrix of bone that has been suggested to have several different physiological functions, including the nucleation of hydroxyapatite (HA), promotion of cell attachment and binding of collagen. Studies in our lab have demonstrated that increased expression of BSP in osteoblast cells can increase expression of the osteoblast-related genes Runx2 and Osx as well as alkaline phosphatase and osteocalcin and increase matrix mineralization. To determine the molecular mechanisms responsible for the BSP-mediated increase in osteoblastic differentiation, several functional domain mutants of BSP were expressed in primary rat bone osteoblastic cells, including the contiguous glutamic acid sequences (polyGlu) and the arginine-glycine-aspartic acid (RGD) motif. Markers of osteoblast differentiation, including matrix mineralization and alkaline phosphatase staining, were increased in cells expressing BSP mutants of the polyGlu sequences but not in cells expressing RGD-mutated BSP. We also determined the dependence on integrin-associated pathways in promoting BSP-mediated differentiation responses in osteoblasts by demonstrating the activation of focal adhesion kinase, MAP kinase-associated proteins ERK1/2, ribosomal s6 kinase 2 and the AP-1 protein cFos. Thus, the mechanism regulating osteoblast differentiation by BSP was determined to be dependent on integrin-mediated intracellular signaling pathways. Copyright 2008 S. Karger AG, Basel.

Additively manufactured 3D porous Ti-6Al-4V constructs mimic trabecular bone structure and regulate osteoblast proliferation, differentiation and local factor production in a porosity and surface roughness dependent manner.

PubMed

Cheng, Alice; Humayun, Aiza; Cohen, David J; Boyan, Barbara D; Schwartz, Zvi

2014-10-07

Additive manufacturing by laser sintering is able to produce high resolution metal constructs for orthopedic and dental implants. In this study, we used a human trabecular bone template to design and manufacture Ti-6Al-4V constructs with varying porosity via laser sintering. Characterization of constructs revealed interconnected porosities ranging from 15-70% with compressive moduli of 2579-3693 MPa. These constructs with macro porosity were further surface-treated to create a desirable multi-scale micro-/nano-roughness, which has been shown to enhance the osseointegration process. Osteoblasts (MG63 cells) exhibited high viability when grown on the constructs. Proliferation (DNA) and alkaline phosphatase specific activity, an early differentiation marker, decreased as porosity increased, while osteocalcin, a late differentiation marker, as well as osteoprotegerin, vascular endothelial growth factor and bone morphogenetic proteins 2 and 4 increased with increasing porosity. Three-dimensional (3D) constructs with the highest porosity and surface modification supported the greatest osteoblast differentiation and local factor production. These results indicate that additively manufactured 3D porous constructs mimicking human trabecular bone and produced with additional surface treatment can be customized for increased osteoblast response. Increased factors for osteoblast maturation and differentiation on high porosity constructs suggest the enhanced performance of these surfaces for increasing osseointegration in vivo.

Promotion of osteoblast differentiation in 3D biomaterial micro-chip arrays comprising fibronectin-coated poly(methyl methacrylate) polycarbonate.

PubMed

Altmann, Brigitte; Steinberg, Thorsten; Giselbrecht, Stefan; Gottwald, Eric; Tomakidi, Pascal; Bächle-Haas, Maria; Kohal, Ralf-Joachim

2011-12-01

Due to the architecture of solid body tissues including bone, three-dimensional (3D) in vitro microenvironments appear favorable, since herein cell growth proceeds under more physiological conditions compared to conventional 2D systems. In the present study we show that a 3D microenvironment comprising a fibronectin-coated PMMA/PC-based micro-chip promotes differentiation of primary human osteoblasts as reflected by the densely-packed 3D bone cell aggregates and expression of biomarkers indicating osteoblast differentiation. Morphogenesis and fluorescence dye-based live/dead staining revealed homogenous cell coverage of the microcavities of the chip array, whereat cells showed high viability up to 14 days. Moreover, Azur II staining proved formation of uniform sized multilayered aggregates, exhibiting progressive intracellular deposition of extracellular bone matrix constituents comprising fibronectin, osteocalcin and osteonectin from day 7 on. Compared to 2D monolayers, osteoblasts grown in the 3D chip environment displayed differential mostly higher gene expression for osteocalcin, osteonectin, and alkaline phosphatase, while collagen type I remained fairly constant in both culture environments. Our results indicate that the 3D microenvironment, based on the PMMA biomaterial chip array promotes osteoblast differentiation, and hereby renders a promising tool for tissue-specific in vitro preconditioning of osteoblasts designated for clinically-oriented bone augmentation or regeneration. Copyright © 2011 Elsevier Ltd. All rights reserved.

Differentiating zones at periodontal ligament-bone and periodontal ligament-cementum entheses.

PubMed

Lee, J-H; Pryce, B A; Schweitzer, R; Ryder, M I; Ho, S P

2015-12-01

The structural and functional integrity of bone-periodontal ligament (PDL)-cementum complex stems from the load-bearing attachment sites (entheses) between soft (PDL) and hard (bone, cementum) tissues. These attachment sites are responsible for the maintenance of a bone-PDL-cementum complex biomechanical function. The objective was to investigate changes in spatiotemporal expression of key biomolecules in developing and functionally active entheses. Multilabeling technique was performed on hemimandibles of 3 wk and 3 mo-old scleraxis-GFP transgenic mice for CD146, CD31, NG2, osterix and bone sialoprotein. Regions of dominant stretch within the PDL were evaluated by identifying directionality of collagen fibrils, PDL fibroblasts and PDL cell cytoskeleton. CD146+ cells adjacent to CD31+ vasculature were identified at PDL-bone enthesis. NG2+ cells were located at coronal bone-PDL and apical cementum-PDL entheses in the 3-wk-old group, but at 3 mo, NG2 was positive at the entheses of the apical region and alveolar crest. NG2 and osterix were colocalized at the osteoid and cementoid regions of the PDL-bone and PDL-cementum entheses. Bone sialoprotein was prominent at the apical region of 3-wk-old mice. The directionality of collagen fibers, fibroblasts and their cytoskeleton overlapped, except in the apical region of 3 wk. Colocalization of biomolecules at zones of the PDL adjacent to attachment sites may be essential for the formation of precementum and osteoid interfaces at a load-bearing bone-PDL-tooth fibrous joint. Biophysical cues resulting from development and function can regulate recruitment and differentiation of stem cells potentially from a vascular origin toward osteo- and cemento-blastic lineages at the PDL-bone and PDL-cementum entheses. Investigating the coupled effect of biophysical and biochemical stimuli leading to cell differentiation at the functional attachment sites is critical for developing regeneration strategies to enable functional reconstruction of the periodontal complex. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

DNA Demethylation Rescues the Impaired Osteogenic Differentiation Ability of Human Periodontal Ligament Stem Cells in High Glucose

PubMed Central

Liu, Zhi; Chen, Tian; Sun, Wenhua; Yuan, Zongyi; Yu, Mei; Chen, Guoqing; Guo, Weihua; Xiao, Jingang; Tian, Weidong

2016-01-01

Diabetes mellitus, characterized by abnormally high blood glucose levels, gives rise to impaired bone remodeling. In response to high glucose (HG), the attenuated osteogenic differentiation capacity of human periodontal ligament stem cells (hPDLSCs) is associated with the loss of alveolar bone. Recently, DNA methylation was reported to affect osteogenic differentiation of stem cells in pathological states. However, the intrinsic mechanism linking DNA methylation to osteogenic differentiation ability in the presence of HG is still unclear. In this study, we found that diabetic rats with increased DNA methylation levels in periodontal ligaments exhibited reduced bone mass and density. In vitro application of 5-aza-2′-deoxycytidine (5-aza-dC), a DNA methyltransferase inhibitor, to decrease DNA methylation levels in hPDLSCs, rescued the osteogenic differentiation capacity of hPDLSCs under HG conditions. Moreover, we demonstrated that the canonical Wnt signaling pathway was activated during this process and, under HG circumstances, the 5-aza-dC-rescued osteogenic differentiation capacity was blocked by Dickkopf-1, an effective antagonist of the canonical Wnt signaling pathway. Taken together, these results demonstrate for the first time that suppression of DNA methylation is able to facilitate the osteogenic differentiation capacity of hPDLSCs exposed to HG, through activation of the canonical Wnt signaling pathway. PMID:27273319