Surface Mechanoengineering of a Zr-based Bulk Metallic Glass via Ar-Nanobubble Doping to Probe Cell Sensitivity to Rigid Materials

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

Huang, Lu; Tian, Mengkun; Wu, Dong

In this paper, a new materials platform, utilizing the amorphous microstructure of bulk metallic glasses (BMGs) and the versatility of ion implantation, was developed for the fundamental investigation of cell responses to substrate-rigidity variations in the gigapascal modulus range, which was previously unattainable with polymeric materials. The surface rigidity of a Zr-Al- Ni-Cu-Y BMG was modulated with low-energy Ar-ion implantation owing to the impartment of Ar nanobubbles into the amorphous matrix. Surface softening was achieved due to the formation of nanobubble-doped transitional zones in the Zrbased BMG substrate. Bone-forming cell studies on this newly designed platform demonstrated that mechanical cues,more » accompanied with the potential effects of other surface properties (i.e. roughness, morphology, and chemistry), contributed to modulating cell behaviors. Cell adhesion and actin filaments were found to be less established on less stiff surfaces, especially on the surface with an elastic modulus of 51 GPa. Cell growth appeared to be affected by surface mechanical properties. A lower stiffness was generally related to a higher growth rate. Findings in this study broadened our fundamental understanding concerning the mechanosensing of bone cells on stiff substrates. It also suggests that surface mechano-engineering of metallic materials could be a potential strategy to promote osseointegration of such materials for bone-implant applications. Further investigations are proposed to fine tune the ion implantation variables in order to further distinguish the surface-mechanical effect on bone-forming cell activities from the contributions of other surface properties.« less

Acceleration of osteogenesis by using barium titanate piezoelectric ceramic as an implant material

NASA Astrophysics Data System (ADS)

Furuya, K.; Morita, Y.; Tanaka, K.; Katayama, T.; Nakamachi, E.

2011-04-01

As bone has piezoelectric properties, it is expected that activity of bone cells and bone formation can be accelerated by applying piezoelectric ceramics to implants. Since lead ions, included in ordinary piezoelectric ceramics, are harmful, a barium titanate (BTO) ceramic, which is a lead-free piezoelectric ceramic, was used in this study. The purpose of this study was to investigate piezoelectric effects of surface charge of BTO on cell differentiation under dynamic loading in vitro. Rat bone marrow cells seeded on surfaces of BTO ceramics were cultured in culture medium supplemented with dexamethasone, β-glycerophosphate and ascorbic acid while a dynamic load was applied to the BTO ceramics. After 10 days of cultivation, the cell layer and synthesized matrix on the BTO surfaces were scraped off, and then DNA content, alkaline phosphtase (ALP) activity and calcium content were measured, to evaluate osteogenic differentiation. ALP activity on the charged BTO surface was slightly higher than that on the non-charged BTO surface. The amount of calcium on the charged BTO surface was also higher than that on the non-charged BTO surface. These results showed that the electric charged BTO surface accelerated osteogenesis.

Biological and Mechanical Effects of Micro-Nanostructured Titanium Surface on an Osteoblastic Cell Line In vitro and Osteointegration In vivo.

PubMed

Hao, Jingzu; Li, Ying; Li, Baoe; Wang, Xiaolin; Li, Haipeng; Liu, Shimin; Liang, Chunyong; Wang, Hongshui

2017-09-01

Hybrid micro-nanostructure implant surface was produced on titanium (Ti) surface by acid etching and anodic oxidation to improve the biological and mechanical properties. The biological properties of the micro-nanostructure were investigated by simulated body fluid (SBF) soaking test and MC3T3-E1 cell co-culture experiment. The cell proliferation, spreading, and bone sialoprotein (BSP) gene expression were examined by MTT, SEM, and reverse transcription-polymerase chain reaction (RT-PCR), respectively. In addition, the mechanical properties were evaluated by instrumented nanoindentation test and friction-wear test. Furthermore, the effect of the micro-nanostructure surface on implant osteointegration was examined by in vivo experiment. The results showed that the formation of bone-like apatite was accelerated on the micro-nanostructured Ti surface after immersion in simulated body fluid, and the proliferation, spreading, and BSP gene expression of the MC3T3-E1 cells were also upregulated on the modified surface. The micro-nanostructured Ti surface displayed decreased friction coefficient, stiffness value, and Young's modulus which were much closer to those of the cortical bone, compared to the polished Ti surface. This suggested much better mechanical match to the surrounding bone tissue of the micro-nanostructured Ti surface. Furthermore, the in vivo animal experiment showed that after implantation in the rat femora, the micro-nanostructure surface displayed higher bonding strength between bone tissues and implant; hematoxylin and eosin (H&E) staining suggested that much compact osteoid tissue was observed at the interface of Micro-nano-Ti-bone than polished Ti-bone interface after implantation. Based on these results mentioned above, it was concluded that the improved biological and mechanical properties of the micro-nanostructure endowed Ti surface with good biocompatibility and better osteointegration, implying the enlarged application of the micro-nanostructure surface Ti implants in future.

Alendronate promotes bone formation by inhibiting protein prenylation in osteoblasts in rat tooth replantation model.

PubMed

Komatsu, Koichiro; Shimada, Akemi; Shibata, Tatsuya; Wada, Satoshi; Ideno, Hisashi; Nakashima, Kazuhisa; Amizuka, Norio; Noda, Masaki; Nifuji, Akira

2013-11-01

Bisphosphonates (BPs) are a major class of antiresorptive drug, and their molecular mechanisms of antiresorptive action have been extensively studied. Recent studies have suggested that BPs target bone-forming cells as well as bone-resorbing cells. We previously demonstrated that local application of a nitrogen-containing BP (N-BP), alendronate (ALN), for a short period of time increased bone tissue in a rat tooth replantation model. Here, we investigated cellular mechanisms of bone formation by ALN. Bone histomorphometry confirmed that bone formation was increased by local application of ALN. ALN increased proliferation of bone-forming cells residing on the bone surface, whereas it suppressed the number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts in vivo. Moreover, ALN treatment induced more alkaline phosphatase-positive and osteocalcin-positive cells on the bone surface than PBS treatment. In vitro studies revealed that pulse treatment with ALN promoted osteocalcin expression. To track the target cells of N-BPs, we applied fluorescence-labeled ALN (F-ALN) in vivo and in vitro. F-ALN was taken into bone-forming cells both in vivo and in vitro. This intracellular uptake was inhibited by endocytosis inhibitors. Furthermore, the endocytosis inhibitor dansylcadaverine (DC) suppressed ALN-stimulated osteoblastic differentiation in vitro and it suppressed the increase in alkaline phosphatase-positive bone-forming cells and subsequent bone formation in vivo. DC also blocked the inhibition of Rap1A prenylation by ALN in the osteoblastic cells. These data suggest that local application of ALN promotes bone formation by stimulating proliferation and differentiation of bone-forming cells as well as inhibiting osteoclast function. These effects may occur through endocytic incorporation of ALN and subsequent inhibition of protein prenylation.

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.

Effect of bone sialoprotein and collagen coating on cell attachment to TICER and pure titanium implant surfaces.

PubMed

Graf, H-L; Stoeva, S; Armbruster, F P; Neuhaus, J; Hilbig, H

2008-07-01

To improve integration between implants and biological tissues, this study compared bone sialoprotein (BSP) as a surface-coating material against the major organic and inorganic components of bone, collagen type I and hydroxyapatite (TICER). The expression of osteocalcin, osteonectin and transforming growth factor ss was evaluated using immunohistochemical staining procedures. The distribution patterns of osteoblasts on the surface of pure titanium with a smooth machined surface and a rough surface (TICER) were determined by image processing using confocal laser scanning microscopy. The results compared to uncoated control materials showed that, at all times investigated, the number of cells on the surface of the TICER and pure titanium samples differed significantly (P<0.1), demonstrating the superiority of TICER over pure titanium in this respect. For pure titanium implants, collagen-precoated surfaces were not beneficial for the attachment of bone-derived cells with the exception of day 3 in vitro (P<0.01). BSP-precoated implant surfaces displayed non-significantly higher numbers of settled cells. BSP-precoated implant surfaces were beneficial for osteoinduction as revealed by osteocalcin and osteonectin expression. BSP precoating of the rough TICER implant surface enhanced the osteoinductive effect much more than did collagen precoating. These results contribute to the consideration of at least two distinct pathways of osseointegration.

Experiment K-6-23. Effect of spaceflight on levels and function of immune cells

NASA Technical Reports Server (NTRS)

Mandel, A. D.; Sonnenfeld, G.; Berry, W.; Taylor, G.; Wellhausen, S. R.; Konstantinova, I.; Lesnyak, A.; Fuchs, B.

1990-01-01

Two different immunology experiments were performed on samples received from rats flown on Cosmos 1887. In the first experiment, rat bone marrow cells were examined in Moscow for their response to colony stimulating factor-M. In the second experiment, rat spleen and bone marrow cells were stained in Moscow with a variety of antibodies directed against cell surface antigenic markers. These cells were preserved and shipped to the United States where they were subjected to analysis on a flow cytometer. The results of the studies indicate that bone marrow cells from flown rats showed a decreased response to colony stimulating factor than did bone marrow cells from control rats. There was a higher percentage of spleen cells from flown rats staining positively for pan-T-cell, suppressor-T-cell and innate interleukin-2 receptor antigens than from control animals. In addition, a higher percentage of cells that appeared to be part of the myelogenous population of bone marrow cells from flown rats stained positively for surface immunoglobulin than did equivalent cells from control rats.

An unscaled parameter to measure the order of surfaces: a new surface elaboration to increase cells adhesion.

PubMed

Bigerelle, M; Anselme, K; Dufresne, E; Hardouin, P; Iost, A

2002-08-01

We present a new parameter to quantify the order of a surface. This parameter is scale-independent and can be used to compare the organization of a surface at different scales of range and amplitude. To test the accuracy of this roughness parameter versus a hundred existing ones, we created an original statistical bootstrap method. In order to assess the physical relevance of this new parameter, we elaborated a great number of surfaces with various roughness amplitudes on titanium and titanium-based alloys using different physical processes. Then we studied the influence of the roughness amplitude on in vitro adhesion and proliferation of human osteoblasts. It was then shown that our new parameter best discriminates among the cell adhesion phenomena than others' parameters (Average roughness (Ra em leader )): cells adhere better on isotropic surfaces with a low order, provided this order is quantified on a scale that is more important than that of the cells. Additionally, on these low ordered metallic surfaces, the shape of the cells presents the same morphological aspect as that we can see on the human bone trabeculae. The method used to prepare these isotropic surfaces (electroerosion) could be undoubtedly and easily applied to prepare most biomaterials with complex geometries and to improve bone implant integration. Moreover, the new order parameter we developed may be particularly useful for the fundamental understanding of the mechanism of bone cell installation on a relief and of the formation of bone cell-material interface.

Research on dental implant and its industrialization stage

NASA Astrophysics Data System (ADS)

Dongjoon, Yang; Sukyoung, Kim

2017-02-01

Bone cell attachment to Ti implant surfaces is the most concerned issue in the clinical implant dentistry. Many attempts to achieve the fast and strong integration between bone and implant have been tried in many ways, such as selection of materials (for example, Ti, ZrO2), shape design of implant (for example, soft tissue level, bone level, taped or conical, etc), and surface modification of implants (for example, roughed. coated, hybrid), etc. Among them, a major consideration is the surface design of dental implants. The surface with proper structural characteristics promotes or induces the desirable responses of cells and tissues. To obtain such surface which has desirable cell and tissue response, a variety of surface modification techniques has been developed and employed for many years. In this review, the method and trend of surface modification will be introduced and explained in terms of the surface topography and chemistry of dental implants.

Osteoclasts but not osteoblasts are affected by a calcified surface treated with zoledronic acid in vitro

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

Schindeler, Aaron; Little, David G.; Discipline of Paediatrics and Child Health, Faculty of Medicine, University of Sydney, Sydney

2005-12-16

Bisphosphonates are potent inhibitors of osteoclast-mediated bone resorption. Recent interest has centered on the effects of bisphosphonates on osteoblasts. Chronic dosing of osteoblasts with solubilized bisphosphonates has been reported to enhance osteogenesis and mineralization in vitro. However, this methodology poorly reflects the in vivo situation, where free bisphosphonate becomes rapidly bound to mineralized bone surfaces. To establish a more clinically relevant cell culture model, we cultured bone cells on calcium phosphate coated quartz discs pre-treated with the potent nitrogen-containing bisphosphonate, zoledronic acid (ZA). Binding studies utilizing [{sup 14}C]-labeled ZA confirmed that the bisphosphonate bound in a concentration-dependent manner over themore » 1-50 {mu}M dose range. When grown on ZA-treated discs, the viability of bone-marrow derived osteoclasts was greatly reduced, while the viability and mineralization of the osteoblastic MC3T3-E1 cell line were largely unaffected. This suggests that only bone resorbing cells are affected by bound bisphosphonate. However, this system does not account for transient exposure to unbound bisphosphonate in the hours following a clinical dosing. To model this event, we transiently treated osteoblasts with ZA in the absence of a calcified surface. Osteoblasts proved highly resistant to all transitory treatment regimes, even when utilizing ZA concentrations that prevented mineralization and/or induced cell death when dosed chronically. This study represents a pharmacologically more relevant approach to modeling bisphosphonate treatment on cultured bone cells and implies that bisphosphonate therapies may not directly affect osteoblasts at bone surfaces.« less

Effects of a hybrid micro/nanorod topography-modified titanium implant on adhesion and osteogenic differentiation in rat bone marrow mesenchymal stem cells.

PubMed

Zhang, Wenjie; Li, Zihui; Huang, Qingfeng; Xu, Ling; Li, Jinhua; Jin, Yuqin; Wang, Guifang; Liu, Xuanyong; Jiang, Xinquan

2013-01-01

Various methods have been used to modify titanium implant surfaces with the aim of achieving better osseointegration. In this study, we fabricated a clustered nanorod structure on an acid-etched, microstructured titanium plate surface using hydrogen peroxide. We also evaluated biofunctionalization of the hybrid micro/nanorod topography on rat bone marrow mesenchymal stem cells. Scanning electron microscopy and x-ray diffraction were used to investigate the surface topography and phase composition of the modified titanium plate. Rat bone marrow mesenchymal stem cells were cultured and seeded on the plate. The adhesion ability of the cells was then assayed by cell counting at one, 4, and 24 hours after cell seeding, and expression of adhesion-related protein integrin β1 was detected by immunofluorescence. In addition, a polymerase chain reaction assay, alkaline phosphatase and Alizarin Red S staining assays, and osteopontin and osteocalcin immunofluorescence analyses were used to evaluate the osteogenic differentiation behavior of the cells. The hybrid micro/nanoscale texture formed on the titanium surface enhanced the initial adhesion activity of the rat bone marrow mesenchymal stem cells. Importantly, the hierarchical structure promoted osteogenic differentiation of these cells. This study suggests that a hybrid micro/nanorod topography on a titanium surface fabricated by treatment with hydrogen peroxide followed by acid etching might facilitate osseointegration of a titanium implant in vivo.

Differential proteomic analysis of a human breast tumor and its matched bone metastasis identifies cell membrane and extracellular proteins associated with bone metastasis.

PubMed

Dumont, Bruno; Castronovo, Vincent; Peulen, Olivier; Blétard, Noëlla; Clézardin, Philippe; Delvenne, Philippe; De Pauw, Edwin A; Turtoi, Andrei; Bellahcène, Akeila

2012-04-06

The classical fate of metastasizing breast cancer cells is to seed and form secondary colonies in bones. The molecules closely associated with these processes are predominantly present at the cell surface and in the extracellular space, establishing the first contacts with the target tissue. In this study, we had the rare opportunity to analyze a bone metastatic lesion and its corresponding breast primary tumor obtained simultaneously from the same patient. Using mass spectrometry, we undertook a proteomic study on cell surface and extracellular protein-enriched material. We provide a repertoire of significantly modulated proteins, some with yet unknown roles in the bone metastatic process as well as proteins notably involved in cancer cell invasiveness and in bone metabolism. The comparison of these clinical data with those previously obtained using a human osteotropic breast cancer cell line highlighted an overlapping group of proteins. Certain differentially expressed proteins are validated in the present study using immunohistochemistry on a retrospective collection of breast tumors and matched bone metastases. Our exclusive set of selected proteins supports the setup of further investigations on both clinical samples and experimental bone metastasis models that will help to reveal the finely coordinated expression of proteins that favor the development of metastases in the bone microenvironment.

Emulating Native Periosteum Cell Population and Subsequent Paracrine Factor Production To Promote Tissue Engineered Periosteum-Mediated Allograft Healing

PubMed Central

Hoffman, Michael D.

2015-01-01

Emulating autograft healing within the context of decellularized bone allografts has immediate clinical applications in the treatment of critical-sized bone defects. The periosteum, a thin, osteogenic tissue that surrounds bone, houses a heterogeneous population of stem cells and osteoprogenitors. There is evidence that periosteum-cell derived paracrine factors, specifically vascular endothelial growth factor (VEGF) and bone morphogenetic protein 2 (BMP2), orchestrate autograft healing through host cell recruitment and subsequent tissue elaboration. In previous work, we demonstrated that the use of poly(ethylene glycol) (PEG) hydrogels as a tissue engineered (T.E.) periosteum to localize mesenchymal stem cells (MSCs) to the surface of decellularized bone enhances allograft healing and integration. Herein, we utilize a mixed population of 50:50 MSCs and osteoprogenitor cells to better mimic native periosteum cell population and paracrine factor production to further promote allograft healing. This mixed cell population was localized to the surface of decellularized allografts within degradable hydrogels and shown to expedite allograft healing. Specifically, bone callus formation and biomechanical graft-host integration are increased as compared to unmodified allografts. These results demonstrate the dual importance of periosteum-mediated paracrine factors orchestrating host cell recruitment as well as new bone formation while developing clinically translatable strategies for allograft healing and integration. PMID:25818449

Biomaterials and bone mechanotransduction

NASA Technical Reports Server (NTRS)

Sikavitsas, V. I.; Temenoff, J. S.; Mikos, A. G.; McIntire, L. V. (Principal Investigator)

2001-01-01

Bone is an extremely complex tissue that provides many essential functions in the body. Bone tissue engineering holds great promise in providing strategies that will result in complete regeneration of bone and restoration of its function. Currently, such strategies include the transplantation of highly porous scaffolds seeded with cells. Prior to transplantation the seeded cells are cultured in vitro in order for the cells to proliferate, differentiate and generate extracellular matrix. Factors that can affect cellular function include the cell-biomaterial interaction, as well as the biochemical and the mechanical environment. To optimize culture conditions, good understanding of these parameters is necessary. The new developments in bone biology, bone cell mechanotransduction, and cell-surface interactions are reviewed here to demonstrate that bone mechanotransduction is strongly influenced by the biomaterial properties.

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

TRAP-Positive Multinucleated Giant Cells Are Foreign Body Giant Cells Rather Than Osteoclasts: Results From a Split-Mouth Study in Humans.

PubMed

Lorenz, Jonas; Kubesch, Alica; Korzinskas, Tadas; Barbeck, Mike; Landes, Constantin; Sader, Robert A; Kirkpatrick, Charles J; Ghanaati, Shahram

2015-12-01

This study compared the material-specific tissue response to the synthetic, hydroxyapatite-based bone substitute material NanoBone (NB) with that of the xenogeneic, bovine-based bone substitute material Bio-Oss (BO). The sinus cavities of 14 human patients were augmented with NB and BO in a split-mouth design. Six months after augmentation, bone biopsies were extracted for histological and histomorphometric investigation prior to dental implant insertion. The following were evaluated: the cellular inflammatory pattern, the induction of multinucleated giant cells, vascularization, the relative amounts of newly formed bone, connective tissue, and the remaining bone substitute material. NB granules were well integrated in the peri-implant tissue and were surrounded by newly formed bone tissue. Multinucleated giant cells were visible on the surfaces of the remaining granules. BO granules were integrated into the newly formed bone tissue, which originated from active osteoblasts on their surface. Histomorphometric analysis showed a significantly higher number of multinucleated giant cells and blood vessels in the NB group compared to the BO group. No statistical differences were observed in regard to connective tissue, remaining bone substitute, and newly formed bone. The results of this study highlight the different cellular reactions to synthetic and xenogeneic bone substitute materials. The significantly higher number of multinucleated giant cells within the NB implantation bed seems to have no effect on its biodegradation. Accordingly, the multinucleated giant cells observed within the NB implantation bed have characteristics more similar to those of foreign body giant cells than to those of osteoclasts.

Enhanced human bone marrow mesenchymal stem cell functions on cathodic arc plasma-treated titanium.

PubMed

Zhu, Wei; Teel, George; O'Brien, Christopher M; Zhuang, Taisen; Keidar, Michael; Zhang, Lijie Grace

2015-01-01

Surface modification of titanium for use in orthopedics has been explored for years; however, an ideal method of integrating titanium with native bone is still required to this day. Since human bone cells directly interact with nanostructured extracellular matrices, one of the most promising methods of improving titanium's osseointegration involves inducing bio-mimetic nanotopography to enhance cell-implant interaction. In this regard, we explored an approach to functionalize the surface of titanium by depositing a thin film of textured titanium nanoparticles via a cathodic arc discharge plasma. The aim is to improve human bone marrow mesenchymal stem cell (MSC) attachment and differentiation and to reduce deleterious effects of more complex surface modification methods. Surface functionalization was analyzed by scanning electron microscopy, atomic force microscopy, contact angle testing, and specific protein adsorption. Scanning electron microscopy and atomic force microscopy examination demonstrate the deposition of titanium nanoparticles and the surface roughness change after coating. The specific fibronectin adsorption was enhanced on the modified titanium surface that associates with the improved hydrophilicity. MSC adhesion and proliferation were significantly promoted on the nanocoated surface. More importantly, compared to bare titanium, greater production of total protein, deposition of calcium mineral, and synthesis of alkaline phosphatase were observed from MSCs on nanocoated titanium after 21 days. The method described herein presents a promising alternative method for inducing more cell favorable nanosurface for improved orthopedic applications.

Bone Marrow Transplantation Results in Human Donor Blood Cells Acquiring and Displaying Mouse Recipient Class I MHC and CD45 Antigens on Their Surface

PubMed Central

Yamanaka, Nobuko; Wong, Christine J.; Gertsenstein, Marina; Casper, Robert F.; Nagy, Andras; Rogers, Ian M.

2009-01-01

Background Mouse models of human disease are invaluable for determining the differentiation ability and functional capacity of stem cells. The best example is bone marrow transplants for studies of hematopoietic stem cells. For organ studies, the interpretation of the data can be difficult as transdifferentiation, cell fusion or surface antigen transfer (trogocytosis) can be misinterpreted as differentiation. These events have not been investigated in hematopoietic stem cell transplant models. Methodology/Principal Findings In this study we investigated fusion and trogocytosis involving blood cells during bone marrow transplantation using a xenograft model. We report that using a standard SCID repopulating assay almost 100% of the human donor cells appear as hybrid blood cells containing both mouse and human surface antigens. Conclusion/Significance Hybrid cells are not the result of cell-cell fusion events but appear to be due to efficient surface antigen transfer, a process referred to as trogocytosis. Antigen transfer appears to be non-random and includes all donor cells regardless of sub-type. We also demonstrate that irradiation preconditioning enhances the frequency of hybrid cells and that trogocytosis is evident in non-blood cells in chimera mice. PMID:20046883

Giant cells around bone biomaterials: Osteoclasts or multi-nucleated giant cells?

PubMed

Miron, Richard J; Zohdi, Hamoon; Fujioka-Kobayashi, Masako; Bosshardt, Dieter D

2016-12-01

Recently accumulating evidence has put into question the role of large multinucleated giant cells (MNGCs) around bone biomaterials. While cells derived from the monocyte/macrophage lineage are one of the first cell types in contact with implanted biomaterials, it was originally thought that specifically in bone tissues, all giant cells were bone-resorbing osteoclasts whereas foreign body giant cells (FBGCs) were found associated with a connective tissue foreign body reaction resulting in fibrous encapsulation and/or material rejection. Despite the great majority of bone grafting materials routinely found with large osteoclasts, a special subclass of bone biomaterials has more recently been found surrounded by large giant cells virtually incapable of resorbing bone grafts even years after their implantation. While original hypotheses believed that a 'foreign body reaction' may be taking place, histological data retrieved from human samples years after their implantation have put these original hypotheses into question by demonstrating better and more stable long-term bone volume around certain bone grafts. Exactly how or why this 'special' subclass of giant cells is capable of maintaining long-term bone volume, or methods to scientifically distinguish them from osteoclasts remains extremely poorly studied. The aim of this review article was to gather the current available literature on giant cell markers and differences in expression patterns between osteoclasts and MNGCs utilizing 19 specific markers including an array of CD-cell surface markers. Furthermore, the concept of now distinguishing between pro-inflammatory M1-MNGCs (previously referred to as FBGCs) as well as wound-healing M2-MNGCs is introduced and discussed. This review article presents 19 specific cell-surface markers to distinguish between osteoclasts and MNGCs including an array of CD-cell surface markers. Furthermore, the concept of now distinguishing between pro-inflammatory M1-MNGCs (often previously referred to as FBGCs) as well as wound-healing M2-MNGCs is introduced and discussed. The proposed concepts and guidelines aims to guide the next wave of research facilitating the differentiation between osteoclast/MNGCs formation, as well as provides the basis for increasing our understanding of the exact function of MNGCs in bone tissue/biomaterial homeostasis. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Cellular Responses Evoked by Different Surface Characteristics of Intraosseous Titanium Implants

PubMed Central

Feller, Liviu; Jadwat, Yusuf; Khammissa, Razia A. G.; Meyerov, Robin; Lemmer, Johan

2015-01-01

The properties of biomaterials, including their surface microstructural topography and their surface chemistry or surface energy/wettability, affect cellular responses such as cell adhesion, proliferation, and migration. The nanotopography of moderately rough implant surfaces enhances the production of biological mediators in the peri-implant microenvironment with consequent recruitment of differentiating osteogenic cells to the implant surface and stimulates osteogenic maturation. Implant surfaces with moderately rough topography and with high surface energy promote osteogenesis, increase the ratio of bone-to-implant contact, and increase the bonding strength of the bone to the implant at the interface. Certain features of implant surface chemistry are also important in enhancing peri-implant bone wound healing. It is the purpose of this paper to review some of the more important features of titanium implant surfaces which have an impact on osseointegration. PMID:25767803

WAIF1 Is a Cell-Surface CTHRC1 Binding Protein Coupling Bone Resorption and Formation.

PubMed

Matsuoka, Kazuhiko; Kohara, Yukihiro; Naoe, Yoshinori; Watanabe, Atsushi; Ito, Masako; Ikeda, Kyoji; Takeshita, Sunao

2018-04-06

The osteoclast-derived collagen triple helix repeat containing 1 (CTHRC1) protein stimulates osteoblast differentiation, but the underlying mechanism remains unclear. Here, we identified Wnt-activated inhibitory factor 1 (WAIF1)/5T4 as a cell-surface protein binding CTHRC1. The WAIF1-encoding Trophoblast glycoprotein (Tpbg) gene, which is abundantly expressed in the brain and bone but not in other tissues, showed the same expression pattern as Cthrc1. Tpbg downregulation in marrow stromal cells reduced CTHRC1 binding and CTHRC1-stimulated alkaline phosphatase activity through PKCδ activation of MEK/ERK, suggesting a novel WAIF1/PKCδ/ERK pathway triggered by CTHRC1. Unexpectedly, osteoblast lineage-specific deletion of Tpbg downregulated Rankl expression in mouse bones and reduced both bone formation and resorption; importantly, it impaired bone mass recovery following RANKL-induced resorption, reproducing the phenotype of osteoclast-specific Cthrc1 deficiency. Thus, the binding of osteoclast-derived CTHRC1 to WAIF1 in stromal cells activates PKCδ-ERK osteoblastogenic signaling and serves as a key molecular link between bone resorption and formation during bone remodeling. © 2018 American Society for Bone and Mineral Research. © 2018 American Society for Bone and Mineral Research.

Effects of Titanium Surface Microtopography and Simvastatin on Growth and Osteogenic Differentiation of Human Mesenchymal Stem Cells in Estrogen-Deprived Cell Culture.

PubMed

Arpornmaeklong, Premjit; Pripatnanont, Prisana; Chookiatsiri, Chonticha; Tangtrakulwanich, Boonsin

This study aimed to investigate the effects of titanium surface topography and simvastatin on growth and osteogenic differentiation of human bone marrow stromal cells (hBMSCs) in estrogen-deprived (ED) cell culture. Human BMSCs were seeded on cell culture plates, smooth-surface titanium (Ti) disks, and sandblasted with large grits and acid etched (SLA)-surface Ti disks; and subsequently cultured in regular (fetal bovine serum [FBS]), ED, and ED-with 100 nM simvastatin (ED-SIM) culture media for 14 to 21 days. Live/dead cell staining, scanning electron microscope examination, and cell viability assay were performed to determine cell attachment, morphology, and growth. Expression levels of osteoblast-associated genes, Runx2 and bone sialoprotein and levels of alkaline phosphatase (ALP) activity, calcium content, and osteocalcin in culture media were measured to determine osteoblastic differentiation. Expression levels of bone morphogenetic protein-2 (BMP-2) were investigated to examine stimulating effects of simvastatin (n = 4 to 5, mean ± SD). In vitro mineralization was verified by calcein staining. Human BMSCs exhibited different attachment and shapes on smooth and SLA titanium surfaces. Estrogen-deprived cell culture decreased cell attachment and growth, particularly on the SLA titanium surface, but cells were able to grow to reach confluence on day 21 in the ED-osteogenic (OS) culture medium. Promoting effects of the SLA titanium surface in ED-OS were significantly decreased. Simvastatin significantly increased osteogenic differentiation of human BMSCs on the SLA titanium surface in the ED-OS medium, and the promoting effects of simvastatin corresponded with the increasing of BMP-2 gene expression on the SLA titanium surface in ED-OS-SIM culture medium. The ED cell culture model provided a well-defined platform for investigating the effects of hormones and growth factors on cells and titanium surface interaction. Titanium, the SLA surface, and simvastatin synergistically promoted osteoblastic differentiation of hBMSCs in ED condition and might be useful to promote osteointegration in osteoporotic bone.

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.

Pattern of uptake of americium-241 by the rat skeleton and its subsequent redistribution and retention: implications for human dosimetry and toxicology.

PubMed

Priest, N D; Howells, G; Green, D; Haines, J W

1983-01-01

The distribution and retention of intravenously injected 241Am in the skeleton of the female rat has been investigated using autoradiographic and radiochemical techniques. The studies were designed to assess the dosimetric and toxicologic implications of an 241Am intake by man. They showed that in the rat approximately one third of the intravenously injected 241Am was deposited in the skeleton where it appeared to be retained with a long biological half-time. The studies also showed: 1 241Am is initially deposited onto all types of bone surface including endosteal surfaces, periosteal surfaces and those of the vascular canals within cortical bone, but seems to be preferentially deposited onto those that are resorbing, 2 Bone accretion results in the burial of surface deposits of 241Am, 3 Bone resorption causes the removal of 241Am from surfaces, 4 Resorbed 241Am is retained by phagocytic cells (probably macrophages) in the bone marrow, 5 The transfer of 241Am from the phagocytic cells in the marrow to adjacent bone surfaces seems to occur, (local recycling). 6 The possibility that some of the 241Am removed from the bone surfaces enters the blood and is redeposited in bone, (systemic recycling) cannot be dismissed. These results show that 241Am deposition and redistribution in bone shares many characteristics with other 'bone surface-seeking radionuclides' typified by 239Pu. Consequently, it is suggested that a similar model to that used to calculate annual limits of intake for 239Pu in man would be suitable for the calculation of corresponding values for the 241Am isotopes.

Culture-expanded mesenchymal stem cell sheets enhance extraction-site alveolar bone growth: An animal study.

PubMed

Mu, S; Tee, B C; Emam, H; Zhou, Y; Sun, Z

2018-04-06

Impaired bone formation of the buccal alveolar plate after tooth extraction during adolescence increases the difficulty of future implant restoration. This study was undertaken to assess the feasibility and efficacy of transplanting autogenous scaffold-free culture-expanded mesenchymal stem cell (MSC) sheets to the buccal alveolar bone surface to stimulate local bone growth. Mandibular bone marrow was aspirated from 3-month-old pigs (n = 5), from which MSCs were isolated and culture expanded. Triple-layer MSC sheets were then fabricated using temperature-responsive tissue culture plates. One month after bone marrow aspirations, the same pigs underwent bilateral extraction of mandibular primary molars, immediately followed by transplantation of 3 autogenous triple-layer MSC sheets on to the subperiosteal buccal alveolar surface of 1 randomly chosen side. The contralateral side (control) underwent the same periosteal reflection surgery without receiving MSC sheet transplantation. Six weeks later, the animals were killed and specimens from both sides were immediately harvested for radiographic and histological analysis. Buccal alveolar bone thickness, tissue mineral density (TMD), mineral apposition and bone volume fraction (BV/TV) were quantified and compared between the MSC sheet and control sides using paired t-tests. Triple-layer MSC sheets were reliably fabricated and the majority of cells remained vital before transplantation. The thickness of buccal bone tended to increase with MSC sheet transplantation (P = .18), with 4 of 5 animals showing an average of 1.82 ± 0.73 mm thicker bone on the MSC sheet side than the control side. After being normalized by the TMD of intracortical bone, the TMD of surface cortical bone was 0.5-fold higher on the MSC sheet side than the control side (P < .05). Likewise, the BV/TV measurements of the buccal surface region were also 0.4-fold higher on the MSC sheet side than the control side (P < .05) after being normalized by measurements from the intracortical region. Mineral apposition measurements were not different between the 2 sides. Mandibular marrow-derived MSCs can be fabricated into cell sheets and autogenous transplantation of MSC sheets onto the subperiosteal buccal alveolar bone surface at the tooth-extraction site may increase local bone density. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

In vivo outcomes of tissue-engineered osteochondral grafts.

PubMed

Bal, B Sonny; Rahaman, Mohamed N; Jayabalan, Prakash; Kuroki, Keiichi; Cockrell, Mary K; Yao, Jian Q; Cook, James L

2010-04-01

Tissue-engineered osteochondral grafts have been synthesized from a variety of materials, with some success at repairing chondral defects in animal models. We hypothesized that in tissue-engineered osteochondral grafts synthesized by bonding mesenchymal stem cell-loaded hydrogels to a porous material, the choice of the porous scaffold would affect graft healing to host bone, and the quality of cell restoration at the hyaline cartilage surface. Bone marrow-derived allogeneic mesenchymal stem cells were suspended in hydrogels that were attached to cylinders of porous tantalum metal, allograft bone, or a bioactive glass. The tissue-engineered osteochondral grafts, thus created were implanted into experimental defects in rabbit knees. Subchondral bone restoration, defect fill, bone ingrowth-implant integration, and articular tissue quality were compared between the three subchondral materials at 6 and 12 weeks. Bioactive glass and porous tantalum were superior to bone allograft in integrating to adjacent host bone, regenerating hyaline-like tissue at the graft surface, and expressing type II collagen in the articular cartilage.

Cisplatin loaded PMMA: mechanical properties, surface analysis and effects on Saos-2 cell culture.

PubMed

Özben, Hakan; Eralp, Levent; Baysal, Gökhan; Cort, Ayşegül; Sarkalkan, Nazli; Özben, Tomris

2013-01-01

Despite wide resection and systemic chemotherapy, bone tumors may present with local recurrences, metastases and pathological fractures. Application of bone cement containing antineoplastic drug to fill the defect after resection of metastatic lesions and to support implants has been suggested to prevent local tumor growth and implant failures. In this study, we aimed to demonstrate the effects of the addition of cisplatin which is a widely used antineoplastic drug for osteosarcoma, on the mechanical properties of bone cement, and to evaluate the cytotoxic effects of eluted cisplatin on Saos-2 cell culture. Two cement samples were prepared by mixing 100 mg and 300 mg of cisplatin powder with 40 g cement powder. The bone cement of the control group did not contain cisplatin. Mechanical analyses included 4-point bending, compression and shear testing. For cytotoxicity analysis, samples were incubated in Dulbecco's Modified Eagle's medium for 15 days. Mediums were applied to Saos-2 cell culture and cell viability was measured. Surface analyses were performed by scanning electron microscope (SEM). The addition of cisplatin did not alter the mechanical properties of bone cement. It was observed that the eluted cisplatin had cytotoxic effects on Saos-2 cells. SEM analyses demonstrated cisplatin granules on the surface of cement samples. Cisplatin maintains its cytotoxic property when released from bone cement without compromising the mechanical stability. Application of cisplatin loaded bone cement may help local control of tumor growth. We believe that our study will shed light on to these new practices for the treatment of bone cancers and will encourage future studies.

Ectopic bone formation by marrow stromal osteoblast transplantation using poly(DL-lactic-co-glycolic acid) foams implanted into the rat mesentery

NASA Technical Reports Server (NTRS)

Ishaug-Riley, S. L.; Crane, G. M.; Gurlek, A.; Miller, M. J.; Yasko, A. W.; Yaszemski, M. J.; Mikos, A. G.; McIntire, L. V. (Principal Investigator)

1997-01-01

Porous biodegradable poly(DL-lactic-co-glycolic acid) foams were seeded with rat marrow stromal cells and implanted into the rat mesentery to investigate in vivo bone formation at an ectopic site. Cells were seeded at a density of 6.83 x 10(5) cells/cm2 onto polymer foams having pore sizes ranging from either 150 to 300 to 710 microns and cultured for 7 days in vitro prior to implantation. The polymer/cell constructs were harvested after 1, 7, 28, or 49 days in vivo and processed for histology and gel permeation chromatography. Visual observation of hematoxylin and eosin-stained sections and von Kossa-stained sections revealed the formation of mineralized bonelike tissue in the constructs within 7 days postimplantation. Ingrowth of vascular tissue was also found adjacent to the islands of bone, supplying the necessary metabolic requirements to the newly formed tissue. Mineralization and bone tissue formation were investigated by histomorphometry. The average penetration depth of mineralized tissue in the construct ranged from 190 +/- 50 microns for foams with 500-710-microns pores to 370 +/- 160 microns for foams with 150-300-microns pores after 49 days in vivo. The mineralized bone volume per surface area and total bone volume per surface area had maximal values of 0.28 +/- 0.21 mm (500-710-microns pore size, day 28) and 0.038 +/- 0.024 mm (150-300-microns, day 28), respectively. As much as 11% of the foam volume penetrated by bone tissue was filled with mineralized tissue. No significant trends over time were observed for any of the measured values (penetration depth, bone volume/surface area, or percent mineralized bone volume). These results suggest the feasibility of bone formation by osteoblast transplantation in an orthotopic site where not only bone formation from transplanted cells but also ingrowth from adjacent bone may occur.

Modulating macrophage polarization with divalent cations in nanostructured titanium implant surfaces

NASA Astrophysics Data System (ADS)

Lee, Chung-Ho; Kim, Youn-Jeong; Jang, Je-Hee; Park, Jin-Woo

2016-02-01

Nanoscale topographical modification and surface chemistry alteration using bioactive ions are centrally important processes in the current design of the surface of titanium (Ti) bone implants with enhanced bone healing capacity. Macrophages play a central role in the early tissue healing stage and their activity in response to the implant surface is known to affect the subsequent healing outcome. Thus, the positive modulation of macrophage phenotype polarization (i.e. towards the regenerative M2 rather than the inflammatory M1 phenotype) with a modified surface is essential for the osteogenesis funtion of Ti bone implants. However, relatively few advances have been made in terms of modulating the macrophage-centered early healing capacity in the surface design of Ti bone implants for the two important surface properties of nanotopography and and bioactive ion chemistry. We investigated whether surface bioactive ion modification exerts a definite beneficial effect on inducing regenerative M2 macrophage polarization when combined with the surface nanotopography of Ti. Our results indicate that nanoscale topographical modification and surface bioactive ion chemistry can positively modulate the macrophage phenotype in a Ti implant surface. To the best of our knowledge, this is the first demonstration that chemical surface modification using divalent cations (Ca and Sr) dramatically induces the regenerative M2 macrophage phenotype of J774.A1 cells in nanostructured Ti surfaces. In this study, divalent cation chemistry regulated the cell shape of adherent macrophages and markedly up-regulated M2 macrophage phenotype expression when combined with the nanostructured Ti surface. These results provide insight into the surface engineering of future Ti bone implants that are harmonized between the macrophage-governed early wound healing process and subsequent mesenchymal stem cell-centered osteogenesis function.

Simulation of bone resorption-repair coupling in vitro.

PubMed

Jones, S J; Gray, C; Boyde, A

1994-10-01

In the normal adult human skeleton, new bone formation by osteoblasts restores the contours of bone surfaces following osteoclastic bone resorption, but the evidence for resorption-repair coupling remains circumstantial. To investigate whether sites of prior resorption, more than the surrounding unresorbed surface, attract osteoblasts or stimulate them to proliferate or make new matrix, we developed a simple in vitro system in which resorption-repair coupling occurs. Resorption pits were produced in mammalian dentine or bone slabs by culturing chick bone-derived cells on them for 2-3 days. The chick cells were swept off and the substrata reseeded with rat calvarial osteoblastic cells, which make bone nodules in vitro, for periods of up to 8 weeks. Cell positions and new bone formation were investigated by ordinary light microscopy, fluorescence and reflection confocal laser microscopy, and SEM, in stained and unstained samples. There was no evidence that the osteoblasts were especially attracted to, or influenced by, the sites of resorption in dentine or bone before cell confluence was reached. Bone formation was identified by light microscopy by the accumulation of matrix, staining with alizarin and calcein and by von Kossa's method, and confirmed by scanning electron microscopy (SEM) by using backscattered electron (BSE) and transmitted electron imaging of unembedded samples and BSE imaging of micro-milled embedded material. These new bone patches were located initially in the resorption pits. The model in vitro system may throw new light on the factors that control resorption-repair coupling in the mineralised tissues in vivo.

Immunolocalization of matrix metalloproteinase-13 on bone surface under osteoclasts in rat tibia.

PubMed

Nakamura, Hiroaki; Sato, Ginga; Hirata, Azumi; Yamamoto, Toshio

2004-01-01

Matrix metalloproteinase (MMP)-13 (an interstitial collagenase also called collagenase 3) is involved in degradation of extracellular matrix in various tissues. Using immunohistochemistry and Western blotting, we investigated localization of MMP-13 in rat tibia, to clarify the role of MMP-13 in bone resorption. MMP-13 reactivity was mainly seen on bone surfaces under osteoclasts, and in some osteocytes and their lacunae near osteoclasts. However, immunoreactivity was not seen in chondrocytes or osteoclasts. MMP-13 was also localized on cement lines in the epiphysis. In the growth plate erosion zone, perivascular cells showed MMP-13 reactivity. Immunoelectron microscopy revealed that MMP-13 was localized on the bone surfaces, under the ruffled borders and some clear zones of osteoclasts. Gold-labeled MMP-13 was closely associated with collagen fibrils. Gold labeling was also detected in Golgi apparatus of osteocytes adjacent to osteoclasts and bone lining cells. Western blotting showed that MMP-13 was mainly associated with mineralized bone matrix. These findings suggest that MMP-13 synthesized and secreted by osteoblast-lineage cells is localized under the ruffled borders of osteoclasts. MMP-13 may play an important role in degradation of type I collagen in bone matrix, acting in concert with cathepsin K and MMP-9 produced by osteoclasts. MMP-13 in perivascular cells may be involved in removal of cartilage matrix proteins such as type II collagen and aggrecan.

Enhanced human bone marrow mesenchymal stem cell functions on cathodic arc plasma-treated titanium

PubMed Central

Zhu, Wei; Teel, George; O’Brien, Christopher M; Zhuang, Taisen; Keidar, Michael; Zhang, Lijie Grace

2015-01-01

Surface modification of titanium for use in orthopedics has been explored for years; however, an ideal method of integrating titanium with native bone is still required to this day. Since human bone cells directly interact with nanostructured extracellular matrices, one of the most promising methods of improving titanium’s osseointegration involves inducing bio-mimetic nanotopography to enhance cell–implant interaction. In this regard, we explored an approach to functionalize the surface of titanium by depositing a thin film of textured titanium nanoparticles via a cathodic arc discharge plasma. The aim is to improve human bone marrow mesenchymal stem cell (MSC) attachment and differentiation and to reduce deleterious effects of more complex surface modification methods. Surface functionalization was analyzed by scanning electron microscopy, atomic force microscopy, contact angle testing, and specific protein adsorption. Scanning electron microscopy and atomic force microscopy examination demonstrate the deposition of titanium nanoparticles and the surface roughness change after coating. The specific fibronectin adsorption was enhanced on the modified titanium surface that associates with the improved hydrophilicity. MSC adhesion and proliferation were significantly promoted on the nanocoated surface. More importantly, compared to bare titanium, greater production of total protein, deposition of calcium mineral, and synthesis of alkaline phosphatase were observed from MSCs on nanocoated titanium after 21 days. The method described herein presents a promising alternative method for inducing more cell favorable nanosurface for improved orthopedic applications. PMID:26677327

UV-killed Staphylococcus aureus enhances adhesion and differentiation of osteoblasts on bone-associated biomaterials.

PubMed

Somayaji, Shankari N; Huet, Yvette M; Gruber, Helen E; Hudson, Michael C

2010-11-01

Titanium alloys (Ti) are the preferred material for orthopedic applications. However, very often, these metallic implants loosen over a long period and mandate revision surgery. For implant success, osteoblasts must adhere to the implant surface and deposit a mineralized extracellular matrix (ECM). Here, we utilized UV-killed Staphylococcus aureus as a novel osteoconductive coating for Ti surfaces. S. aureus expresses surface adhesins capable of binding to bone and biomaterials directly. Furthermore, interaction of S. aureus with osteoblasts activates growth factor-related pathways that potentiate osteogenesis. Although UV-killed S. aureus cells retain their bone-adhesive ability, they do not stimulate significant immune modulator expression. All of the abovementioned properties were utilized for a novel implant coating so as to promote osteoblast recruitment and subsequent cell functions on the bone-implant interface. In this study, osteoblast adhesion, proliferation, and mineralized ECM synthesis were measured on Ti surfaces coated with fibronectin with and without UV-killed bacteria. Osteoblast adhesion was enhanced on Ti alloy surfaces coated with bacteria compared to uncoated surfaces, while cell proliferation was sustained comparably on both surfaces. Osteoblast markers such as collagen, osteocalcin, alkaline phosphatase activity, and mineralized nodule formation were increased on Ti alloy coated with bacteria compared to uncoated surfaces.

Mesenchymal Progenitors Residing Close to the Bone Surface Are Functionally Distinct from Those in the Central Bone Marrow

PubMed Central

Siclari, Valerie A.; Zhu, Ji; Akiyama, Kentaro; Liu, Fei; Zhang, Xianrong; Chandra, Abhishek; Nah-Cederquist, Hyun-Duck; Shi, Songtao; Qin, Ling

2013-01-01

Long bone is an anatomically complicated tissue with trabecular-rich metaphyses at two ends and cortical-rich diaphysis at the center. The traditional flushing method only isolates mesenchymal progenitor cells from the central region of long bones and these cells are distant from the bone surface. We propose that mesenchymal progenitors residing in endosteal bone marrow that is close to the sites of bone formation, such as trabecular bone and endosteum, behave differently from those in the central bone marrow. In this report, we separately isolated endosteal bone marrow using a unique enzymatic digestion approach and demonstrated that it contained a much higher frequency of mesenchymal progenitors than the central bone marrow. Endosteal mesenchymal progenitors express traditional mesenchymal stem cell markers and are capable of multi-lineage differentiation. However, we found that mesenchymal progenitors isolated from different anatomical regions of the marrow did exhibit important functional differences. Compared to their central marrow counterparts, endosteal mesenchymal progenitors have superior proliferative ability with reduced expression of cell cycle inhibitors. They showed greater immunosuppressive activity in culture and in a mouse model of inflammatory bowel disease. Aging is a major contributing factor for trabecular bone loss. We found that old mice have a dramatically decreased number of endosteal mesenchymal progenitors compared to young mice. Parathyroid hormone (PTH) treatment potently stimulates bone formation. A single PTH injection greatly increased the number of endosteal mesenchymal progenitors, particularly those located at the metaphyseal bone, but had no effect on their central counterparts. In summary, endosteal mesenchymal progenitors are more metabolically active and relevant to physiological bone formation than central mesenchymal progenitors. Hence, they represent a biologically important target for future mesenchymal stem cell studies. PMID:23274348

Mechanic stress generated by a time-varying electromagnetic field on bone surface.

PubMed

Ye, Hui

2018-03-19

Bone cells sense mechanical load, which is essential for bone growth and remodeling. In a fracture, this mechanism is compromised. Electromagnetic stimulation has been widely used to assist in bone healing, but the underlying mechanisms are largely unknown. A recent hypothesis suggests that electromagnetic stimulation could influence tissue biomechanics; however, a detailed quantitative understanding of EM-induced biomechanical changes in the bone is unavailable. This paper used a muscle/bone model to study the biomechanics of the bone under EM exposure. Due to the dielectric properties of the muscle/bone interface, a time-varying magnetic field can generate both compressing and shear stresses on the bone surface, where many mechanical sensing cells are available for cellular mechanotransduction. I calculated these stresses and found that the shear stress is significantly greater than the compressing stress. Detailed parametric analysis suggests that both the compressing and shear stresses are dependent on the geometrical and electrical properties of the muscle and the bone. These stresses are also functions of the orientation of the coil and the frequency of the magnetic field. It is speculated that the EM field could apply biomechanical influence to fractured bone, through the fine-tuning of the controllable field parameters. Graphical abstract Mechanic stress on bone surface in a time-varying magnetic field.

Bone Cell–materials Interactions and Ni Ion Release of Anodized Equiatomic NiTi Alloy

PubMed Central

Bernard, Sheldon A.; Balla, Vamsi Krishna; Davies, Neal M.; Bose, Susmita; Bandyopadhyay, Amit

2011-01-01

Laser processed NiTi alloy was anodized for different durations in H2SO4 electrolyte with varying pH to create biocompatible surfaces with low Ni ion release as well as bioactive surfaces to enhance biocompatibility and bone cell-materials interactions. The anodized surfaces were assessed for their in vitro cell-materials interactions using human fetal osteoblast (hFOB) cells for 3, 7 and 11 days, and Ni ion release up to 8 weeks in simulated body fluids. The results were correlated with surface morphologies of anodized surfaces characterized using field-emission scanning electron microscopy (FESEM). The results show that the anodization creates a surface with nano/micro roughness depending on anodization conditions. The hydrophilicity of NiTi surface was found to improve after anodization due to lower contact angles in cell media, which dropped from 32° to < 5°. The improved wettability of anodized surfaces is further corroborated by their high surface energy comparable to that of cp Ti. Relatively high surface energy, especially polar component, and nano/micro surface features of anodized surfaces significantly increased the number of living cells and their adherence and growth on these surfaces. Finally, a significant drop in Ni ion release from 268 ± 11 to 136 ± 15 ppb was observed for NiTi surfaces after anodization. This work indicates that anodization of NiTi alloy has a positive influence on the surface energy and surface morphology, which in turn improve bone cell-materials interactions and reduce Ni ion release in vitro. PMID:21232641

Reduction and shaping of graphene-oxide by laser-printing for controlled bone tissue regeneration and bacterial killing

NASA Astrophysics Data System (ADS)

Palmieri, Valentina; Barba, Marta; Di Pietro, Lorena; Gentilini, Silvia; Chiara Braidotti, Maria; Ciancico, Carlotta; Bugli, Francesca; Ciasca, Gabriele; Larciprete, Rosanna; Lattanzi, Wanda; Sanguinetti, Maurizio; De Spirito, Marco; Conti, Claudio; Papi, Massimiliano

2018-01-01

Graphene and graphene oxide (GO) are capable of inducing stem cells differentiation into bone tissue with variable efficacy depending on reductive state of the material. Thus, modulation of osteogenic process and of bone mineral density distribution is theoretically possible by controlling the GO oxidative state. In this study, we laser-printed GO surfaces in order to obtain both a local photo-thermal GO reduction and the formation of nano-wrinkles along precise geometric pattern. Initially, after cells adhered on the surface, stem cells migrated and accumulated on the reduced and wrinkled surface. When the local density of the stem cells on the reduced stripes was high, cells started to proliferate and occupy the oxidized/flat area. The designed surfaces morphology guided stem cell orientation and the reduction accelerated differentiation. Furthermore the reduced sharp nano-wrinkles were able to enhance the GO antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), a common cause of prosthetic joints infections. This strategy can offer a revolution in present and future trends of scaffolds design for regenerative medicine.

In vitro study on bone formation and surface topography from the standpoint of biomechanics.

PubMed

Kawahara, H; Soeda, Y; Niwa, K; Takahashi, M; Kawahara, D; Araki, N

2004-12-01

Effect of surface topography upon cell-adhesion, -orientation and -differentiation was investigated by in vitro study on cellular responses to titanium substratum with different surface roughness. Cell-shape, -function and -differentiation depending upon the surface topography were clarified by use of bone formative group cells (BFGCs) derived from bone marrow of beagle's femur. BFGCs consisted of hematopoietic stem cells (HSC) and osteogenetic stem cells (OSC). Cell differentiation of BFGCs was expressed and promoted by structural changes of cytoskeleton, and cell-organella, which was caused by mechanical stress with cytoplasmic stretching of cell adhesions to the substratum. Phagocytic monocytes of HSC differentiated to osteomediator cells (OMC) by cytoplasmic stretching with cell adhesion to the substratum. The OMC mediated and promoted cell differentiation from OSC to osteoblast through osteoblastic phenotype cell (OBC) by cell-aggregation of nodules with "pile up" phenomenon of OBC onto OMC. The osteogenesis might be performed by coupling work of both cells, OMC originated from monocyte of HSC and OBC originated from OSC, which were explained by SEM, TEM and fluorescent probe investigation on BFGCs on the test plate of cp titanium plates with different topographies. This osteogenetic process was proved by investigating cell proliferation, DNA contents, cell-adhesion, alkaline phosphatase activity and osteocalcine productivity for cells on the titanium plates with different topographies. The study showed increased osteogenic effects for cells cultured on Ti with increased surface roughness. Possible mechanisms were discussed from a biomechanical perspective.

Mechanical signals promote osteogenic fate through a primary cilia-mediated mechanism

PubMed Central

Chen, Julia C.; Hoey, David A.; Chua, Mardonn; Bellon, Raymond; Jacobs, Christopher R.

2016-01-01

It has long been suspected, but never directly shown, that bone formed to accommodate an increase in mechanical loading is related to the creation of osteoblasts from skeletal stem cells. Indeed, biophysical stimuli potently regulate osteogenic lineage commitment in vitro. In this study, we transplanted bone marrow cells expressing green fluorescent protein, to enable lineage tracing, and subjected mice to a biophysical stimulus, to elicit a bone-forming response. We detected cells derived from transplanted progenitors embedded within the bone matrix near active bone-forming surfaces in response to loading, demonstrating for the first time, that mechanical signals enhance the homing and attachment of bone marrow cells to bone surfaces and the commitment to an osteogenic lineage of these cells in vivo. Furthermore, we used an inducible Cre/Lox recombination system to delete kinesin family member 3A (Kif3a), a gene that is essential for primary cilia formation, at will in transplanted cells and their progeny, regardless of which tissue may have incorporated them. Disruption of the mechanosensing organelle, the primary cilium in a progenitor population, significantly decreased the amount of bone formed in response to mechanical stimulation. The collective results of our study directly demonstrate that, in a novel experimental stem cell mechanobiology model, mechanical signals enhance osteogenic lineage commitment in vivo and that the primary cilium contributes to this process.—Chen, J. C., Hoey, D. A., Chua, M., Bellon, R., Jacobs, C. R. Mechanical signals promote osteogenic fate through a primary cilia-mediated mechanism. PMID:26675708

Bone cell-materials interactions and Ni ion release of anodized equiatomic NiTi alloy.

PubMed

Bernard, Sheldon A; Balla, Vamsi Krishna; Davies, Neal M; Bose, Susmita; Bandyopadhyay, Amit

2011-04-01

A laser processed NiTi alloy was anodized for different times in H(2)SO(4) electrolyte with varying pH to create biocompatible surfaces with low Ni ion release as well as bioactive surfaces to enhance biocompatibility and bone cell-material interactions. The anodized surfaces were assessed for their in vitro cell-material interactions using human fetal osteoblast (hFOB) cells for 3, 7 and 11 days, and Ni ion release up to 8 weeks in simulated body fluids. The results were correlated with the surface morphologies of anodized surfaces characterized using field-emission scanning electron microscopy (FESEM). The results show that anodization creates a surface with nano/micro-roughness depending on the anodization conditions. The hydrophilicity of the NiTi surface was found to improve after anodization, as shown by the lower contact angles in cell medium, which dropped from 32° to <5°. The improved wettability of anodized surfaces is further corroborated by their high surface energy, comparable with that of commercially pure Ti. Relatively high surface energies, especially the polar component, and nano/micro surface features of anodized surfaces significantly increased the number of living cells and their adherence and growth on these surfaces. Finally, a significant drop in Ni ion release from 268±11 to 136±15 ppb was observed for NiTi surfaces after anodization. This work indicates that anodization of a NiTi alloy has a positive influence on the surface energy and surface morphology, which in turn improves bone cell-material interactions and reduces Ni ion release in vitro. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Formation of bone-like mineralized matrix by periodontal ligament cells in vivo: a morphological study in rats.

PubMed

Hiraga, Toru; Ninomiya, Tadashi; Hosoya, Akihiro; Takahashi, Masafumi; Nakamura, Hiroaki

2009-01-01

Periodontal ligament (PDL) is a unique connective tissue that not only connects cementum and alveolar bone to support teeth, but also plays an important role in reconstructing periodontal tissues. Previous studies have suggested that PDL cells have osteogenic potential; however, they lack precise histological examinations. Here, we studied bone-like matrix formation by PDL cells in rats using morphological techniques. Rat and human PDL cells exhibited substantial alkaline phosphatase activity and induced mineralization in vitro. RT-PCR analyses showed that PDL cells expressed the osteoblast markers, Runx2, osterix, and osteocalcin. These results suggest that PDL cells share similar phenotypes with osteoblasts. To examine the bone-like matrix formation in vivo, PDL cells isolated from green fluorescent protein (GFP)-transgenic rats were inoculated with hydroxyapatite (HA) disks into wild-type rats. Five weeks after the implantation, the pores in HA disks were occupied by GFP-positive cells. Mineralized matrix formation was also found on the surface of HA pores. At 12 weeks, some of the pores were filled with bone-like mineralized matrices (BLMM), which were positive for the bone matrix proteins, osteopontin, bone sialoprotein, and osteocalcin. Immunohistochemical examination revealed that most of the osteoblast- and osteocyte-like cells on or in the BLMM were GFP-positive, suggesting that the BLMM were directly formed by the inoculated PDL cells. On the pore surfaces, Sharpey's fiber-like structures embedded in cementum-like mineralized layers were also observed. These results collectively suggest that PDL cells have the ability to form periodontal tissues and could be a useful source for regenerative therapies of periodontal diseases.

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

Nanoceramics on osteoblast proliferation and differentiation in bone tissue engineering.

PubMed

Sethu, Sai Nievethitha; Namashivayam, Subhapradha; Devendran, Saravanan; Nagarajan, Selvamurugan; Tsai, Wei-Bor; Narashiman, Srinivasan; Ramachandran, Murugesan; Ambigapathi, Moorthi

2017-05-01

Bone, a highly dynamic connective tissue, consist of a bioorganic phase comprising osteogenic cells and proteins which lies over an inorganic phase predominantly made of CaPO 4 (biological apatite). Injury to bone can be due to mechanical, metabolic or inflammatory agents also owing pathological conditions like fractures, osteomyelitis, osteolysis or cysts may arise in enameloid, chondroid, cementum, or chondroid bone which forms the intermediate tissues of the body. Bone tissue engineering (BTE) applies bioactive scaffolds, host cells and osteogenic signals for restoring damaged or diseased tissues. Various bioceramics used in BTE can be bioactive (like glass ceramics and hydroxyapatite bioactive glass), bioresorbable (like tricalcium phosphates) or bioinert (like zirconia and alumina). Limiting the size of these materials to nano-scale has resulted in a higher surface area to volume ratio thereby improving multi-functionality, solubility, surface catalytic activity, high heat and electrical conductivity. Nanoceramics have been found to induce osteoconduction, osteointegration, osteogenesis and osteoinduction. The present review aims at summarizing the interactions of nanoceramics and osteoblast/stem cells for promoting the proliferation and differentiation of the osteoblast cells by nanoceramics as superior bone substitutes in bone tissue engineering applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Surface modification of implants in long bone.

PubMed

Förster, Yvonne; Rentsch, Claudia; Schneiders, Wolfgang; Bernhardt, Ricardo; Simon, Jan C; Worch, Hartmut; Rammelt, Stefan

2012-01-01

Coatings of orthopedic implants are investigated to improve the osteoinductive and osteoconductive properties of the implant surfaces and thus to enhance periimplant bone formation. By applying coatings that mimic the extracellular matrix a favorable environment for osteoblasts, osteoclasts and their progenitor cells is provided to promote early and strong fixation of implants. It is known that the early bone ongrowth increases primary implant fixation and reduces the risk of implant failure. This review presents an overview of coating titanium and hydroxyapatite implants with components of the extracellular matrix like collagen type I, chondroitin sulfate and RGD peptide in different small and large animal models. The influence of these components on cells, the inflammation process, new bone formation and bone/implant contact is summarized.

Surface modification of implants in long bone

PubMed Central

Förster, Yvonne; Rentsch, Claudia; Schneiders, Wolfgang; Bernhardt, Ricardo; Simon, Jan C.; Worch, Hartmut; Rammelt, Stefan

2012-01-01

Coatings of orthopedic implants are investigated to improve the osteoinductive and osteoconductive properties of the implant surfaces and thus to enhance periimplant bone formation. By applying coatings that mimic the extracellular matrix a favorable environment for osteoblasts, osteoclasts and their progenitor cells is provided to promote early and strong fixation of implants. It is known that the early bone ongrowth increases primary implant fixation and reduces the risk of implant failure. This review presents an overview of coating titanium and hydroxyapatite implants with components of the extracellular matrix like collagen type I, chondroitin sulfate and RGD peptide in different small and large animal models. The influence of these components on cells, the inflammation process, new bone formation and bone/implant contact is summarized. PMID:23507866

Histomorphometric and immunohistochemical analysis of human maxillary sinus-floor augmentation using porous β-tricalcium phosphate for dental implant treatment.

PubMed

Miyamoto, Shinji; Shinmyouzu, Kouhei; Miyamoto, Ikuya; Takeshita, Kenji; Terada, Toshihisa; Takahashi, Tetsu

2013-08-01

This study utilized the constitution and expression of Runx2/Cbfa1 to conduct 6-month-post-operation histomorphometrical and histochemical analysis of osteocalcin in bone regeneration following sinus-floor augmentation procedures using β-tricalcium phosphate (β-TCP) and autogenous cortical bone. Thirteen sinuses of nine patients were treated with sinus-floor augmentation using 50% β-TCP and 50% autogenous cancellous bone harvested from the ramus of the mandible. Biopsies of augmented sinuses were taken at 6 months for histomorphometric and immunohistochemical measurements. Runx2/Cbfa1- and osteocalcin-positive cells were found around TCP particles and on the bone surface. Approximately 60% of cells found around TCP particles stained positive for Runx2/Cbfa1. Fewer cells stained positive for osteocalcin. These positive cells decreased apically with increasing vertical distance from the maxillary bone surface. Histomorphometric analysis showed that the augmented site close to residual bone and periosteum contained approximately 42% bony tissue and 42% soft connective tissue, and the remaining 16% consisted of TCP particles. On the other hand, the augmented bone far from residual bone and periosteum contained 35% bony tissue and 50% soft connective tissue. Our data suggest that TCP particles attract osteoprogenitor cells that migrate into the interconnecting micropores of the bone-substitute material by 6 months. The augmented site close to residual bone contained a higher proportion of bony tissue and a lower proportion of soft connective tissue than did the augmented site far from residual bone. © 2012 John Wiley & Sons A/S.

Mining for osteogenic surface topographies: In silico design to in vivo osseo-integration.

PubMed

Hulshof, Frits F B; Papenburg, Bernke; Vasilevich, Aliaksei; Hulsman, Marc; Zhao, Yiping; Levers, Marloes; Fekete, Natalie; de Boer, Meint; Yuan, Huipin; Singh, Shantanu; Beijer, Nick; Bray, Mark-Anthony; Logan, David J; Reinders, Marcel; Carpenter, Anne E; van Blitterswijk, Clemens; Stamatialis, Dimitrios; de Boer, Jan

2017-08-01

Stem cells respond to the physicochemical parameters of the substrate on which they grow. Quantitative material activity relationships - the relationships between substrate parameters and the phenotypes they induce - have so far poorly predicted the success of bioactive implant surfaces. In this report, we screened a library of randomly selected designed surface topographies for those inducing osteogenic differentiation of bone marrow-derived mesenchymal stem cells. Cell shape features, surface design parameters, and osteogenic marker expression were strongly correlated in vitro. Furthermore, the surfaces with the highest osteogenic potential in vitro also demonstrated their osteogenic effect in vivo: these indeed strongly enhanced bone bonding in a rabbit femur model. Our work shows that by giving stem cells specific physicochemical parameters through designed surface topographies, differentiation of these cells can be dictated. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mag-seeding of rat bone marrow stromal cells into porous hydroxyapatite scaffolds for bone tissue engineering.

PubMed

Shimizu, Kazunori; Ito, Akira; Honda, Hiroyuki

2007-09-01

Bone tissue engineering has been investigated as an alternative strategy for autograft transplantation. In the process of tissue engineering, cell seeding into three-dimensional (3-D) scaffolds is the first step for constructing 3-D tissues. We have proposed a methodology of cell seeding into 3-D porous scaffolds using magnetic force and magnetite nanoparticles, which we term Mag-seeding. In this study, we applied this Mag-seeding technique to bone tissue engineering using bone marrow stromal cells (BMSCs) and 3-D hydroxyapatite (HA) scaffolds. BMSCs were magnetically labeled with our original magnetite cationic liposomes (MCLs) having a positive surface charge to improve adsorption to cell surface. Magnetically labeled BMSCs were seeded onto a scaffold, and a 1-T magnet was placed under the scaffold. By using Mag-seeding, the cells were successfully seeded into the internal space of scaffolds with a high cell density. The cell seeding efficiency into HA scaffolds by Mag-seeding was approximately threefold larger than that by static-seeding (conventional method, without a magnet). After a 14-d cultivation period using the osteogenic induction medium by Mag-seeding, the level of two representative osteogenic markers (alkaline phosphatase and osteocalcin) were significantly higher than those by static-seeding. These results indicated that Mag-seeding of BMSCs into HA scaffolds is an effective approach to bone tissue engineering.

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

Pecheva, E.; Fingarova, D.; Pramatarova, L.

Polycrystalline silicon (poly-Si) layers were functionalized through the growth of biomimetic hydroxyapatite (HA) on their surface. HA is the mineral component of bones and teeth and thus possesses excellent bioactivity and biocompatibility. MG-63 osteoblast-like cells were cultured on both HA-coated and un-coated poly-Si surfaces for 1, 3, 5 and 7 days and toxicity, proliferation and cell morphology were investigated. The results revealed that the poly-Si layers were bioactive and compatible with the osteoblast-like cells. Nevertheless, the HA coating improved the cell interactions with the poly-Si surfaces based on the cell affinity to the specific chemical composition of the bone-like HAmore » and/or to the higher HA roughness.« less

Polycrystalline Silicon: a Biocompatibility Assay

NASA Astrophysics Data System (ADS)

Pecheva, E.; Laquerriere, P.; Bouthors, Sylvie; Fingarova, D.; Pramatarova, L.; Hikov, T.; Dimova-Malinovska, D.; Montgomery, P.

2010-01-01

Polycrystalline silicon (poly-Si) layers were functionalized through the growth of biomimetic hydroxyapatite (HA) on their surface. HA is the mineral component of bones and teeth and thus possesses excellent bioactivity and biocompatibility. MG-63 osteoblast-like cells were cultured on both HA-coated and un-coated poly-Si surfaces for 1, 3, 5 and 7 days and toxicity, proliferation and cell morphology were investigated. The results revealed that the poly-Si layers were bioactive and compatible with the osteoblast-like cells. Nevertheless, the HA coating improved the cell interactions with the poly-Si surfaces based on the cell affinity to the specific chemical composition of the bone-like HA and/or to the higher HA roughness.

Detachment strength of human osteoblasts cultured on hydroxyapatite with various surface roughness. Contribution of integrin subunits.

PubMed

Kokkinos, Petros A; Koutsoukos, Petros G; Deligianni, Despina D

2012-06-01

Hydroxyapatite (HA) has been widely used as a bone substitute in dental, maxillofacial and orthopaedic surgery and as osteoconductive bone substitute or precoating of pedicle screws and cages in spine surgery. The aim of the present study was to investigate the osteoblastic adhesion strength on HA substrata with different surface topography and biochemistry (pre-adsorption of fibronectin) after blocking of specific integrin subunits with monoclonal antibodies. Stoichiometric HA was prepared by precipitation followed by ageing and characterized by SEM, EDX, powder XRD, Raman spectroscopy, TGA, and specific surface area analysis. Human bone marrow derived osteoblasts were cultured on HA disc-shaped substrata which were sintered and polished resulting in two surface roughness grades. For attachment evaluation, cells were incubated with monoclonal antibodies and seeded for 2 h on the substrata. Cell detachment strength was determined using a rotating disc device. Cell detachment strength was surface roughness, fibronectin preadsorption and intergin subunit sensitive.

Immunophenotypic characterization of bone marrow mast cells in mastocytosis and other mast cell disorders.

PubMed

Sánchez-Muñoz, Laura; Teodósio, Cristina; Morgado, José M; Escribano, Luis

2011-01-01

Mastocytosis is a term used to designate a heterogeneous group of disorders characterized by an abnormal proliferation and accumulation of mast cells (MCs) in one or multiple tissues including skin, bone marrow (BM), liver, spleen, and lymph nodes, among others. Recent advances in our understanding of mast cell biology and disease resulted in the identification of important differences in the expression of mast cell surface antigens between normal and neoplastic mast cells. Most notably, detection of aberrant expression of CD25 and CD2 on the surface of neoplastic mast cells but not on their normal counterparts lead to the inclusion of this immunophenotypic abnormality in the World Health Organization diagnostic criteria for systemic mastocytosis. Aberrant mast cell surface marker expression can be detected in the bone marrow aspirate by flow cytometry, even in patients lacking histopathologically detectable aggregates of mast cells in bone marrow biopsy sections. These aberrant immunophenotypic features are of great relevance for the assessment of tissue involvement in mastocytosis with consequences in the diagnosis, classification, and follow-up of the disease and in its differential diagnosis with other entities. In this chapter, we provide the reader with information for the objective and reproducible identification of pathologic MCs by using quantitative multiparametric flow cytometry, for their phenotypic characterization, and the criteria currently used for correct interpretation of the immunophenotypic results obtained. Copyright © 2011 Elsevier Inc. All rights reserved.

Autologous implantation of BMP2-expressing dermal fibroblasts to improve bone mineral density and architecture in rabbit long bones.

PubMed

Ishihara, Akikazu; Weisbrode, Steve E; Bertone, Alicia L

2015-10-01

Cell-mediated gene therapy may treat bone fragility disorders. Dermal fibroblasts (DFb) may be an alternative cell source to stem cells for orthopedic gene therapy because of their rapid cell yield and excellent plasticity with bone morphogenetic protein-2 (BMP2) gene transduction. Autologous DFb or BMP2-expressing autologous DFb were administered in twelve rabbits by two delivery routes; a transcortical intra-medullar infusion into tibiae and delayed intra-osseous injection into femoral drill defects. Both delivery methods of DFb-BMP2 resulted in a successful cell engraftment, increased bone volume, bone mineral density, improved trabecular bone microarchitecture, greater bone defect filling, external callus formation, and trabecular surface area, compared to non-transduced DFb or no cells. Cell engraftment within trabecular bone and bone marrow tissue was most efficiently achieved by intra-osseous injection of DFb-BMP2. Our results suggested that BMP2-expressing autologous DFb have enhanced efficiency of engraftment in target bones resulting in a measurable biologic response by the bone of improved bone mineral density and bone microarchitecture. These results support that autologous implantation of DFb-BMP2 warrants further study on animal models of bone fragility disorders, such as osteogenesis imperfecta and osteoporosis to potentially enhance bone quality, particularly along with other gene modification of these diseases. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Stem cell niches and other factors that influence the sensitivity of bone marrow to radiation-induced bone cancer and leukaemia in children and adults

PubMed Central

Richardson, Richard B

2011-01-01

Purpose: This paper reviews and reassesses the internationally accepted niches or ‘targets’ in bone marrow that are sensitive to the induction of leukaemia and primary bone cancer by radiation. Conclusions: The hypoxic conditions of the 10 μm thick endosteal/osteoblastic niche where preleukemic stem cells and hematopoietic stem cells (HSC) reside provides a radioprotective microenvironment that is 2-to 3-fold less radiosensitive than vascular niches. This supports partitioning the whole marrow target between the low haematological cancer risk of irradiating HSC in the endosteum and the vascular niches within central marrow. There is a greater risk of induced bone cancer when irradiating a 50 μm thick peripheral marrow adjacent to the remodelling/reforming portion of the trabecular bone surface, rather than marrow next to the quiescent bone surface. This choice of partitioned bone cancer target is substantiated by the greater radiosensitivity of: (i) Bone with high remodelling rates, (ii) the young, (iii) individuals with hypermetabolic benign diseases of bone, and (iv) the epidemiology of alpha-emitting exposures. Evidence is given to show that the absence of excess bone-cancer in atomic-bomb survivors may be partially related to the extremely low prevalence among Japanese of Paget's disease of bone. Radiation-induced fibrosis and the wound healing response may be implicated in not only radiogenic bone cancers but also leukaemia. A novel biological mechanism for adaptive response, and possibility of dynamic targets, is advocated whereby stem cells migrate from vascular niches to stress-mitigated, hypoxic niches. PMID:21204614

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.

Scanning electron microscopy of bone.

PubMed

Boyde, Alan

2012-01-01

This chapter described methods for Scanning Electron Microscopical imaging of bone and bone cells. Backscattered electron (BSE) imaging is by far the most useful in the bone field, followed by secondary electrons (SE) and the energy dispersive X-ray (EDX) analytical modes. This chapter considers preparing and imaging samples of unembedded bone having 3D detail in a 3D surface, topography-free, polished or micromilled, resin-embedded block surfaces, and resin casts of space in bone matrix. The chapter considers methods for fixation, drying, looking at undersides of bone cells, and coating. Maceration with alkaline bacterial pronase, hypochlorite, hydrogen peroxide, and sodium or potassium hydroxide to remove cells and unmineralised matrix is described in detail. Attention is given especially to methods for 3D BSE SEM imaging of bone samples and recommendations for the types of resin embedding of bone for BSE imaging are given. Correlated confocal and SEM imaging of PMMA-embedded bone requires the use of glycerol to coverslip. Cathodoluminescence (CL) mode SEM imaging is an alternative for visualising fluorescent mineralising front labels such as calcein and tetracyclines. Making spatial casts from PMMA or other resin embedded samples is an important use of this material. Correlation with other imaging means, including microradiography and microtomography is important. Shipping wet bone samples between labs is best done in glycerol. Environmental SEM (ESEM, controlled vacuum mode) is valuable in eliminating -"charging" problems which are common with complex, cancellous bone samples.

Altered bone turnover during spaceflight

NASA Technical Reports Server (NTRS)

Turner, R. T.; Morey, E. R.; Liu, C.; Baylink, D. J.

1982-01-01

Modifications in calcium metabolism during spaceflight were studied, using parameters that reflect bone turnover. Bone formation rate, medullary area, bone length, bone density, pore size distribution, and differential bone cell number were evaluated in growing rate both immediately after and 25 days after orbital spaceflights aboard the Soviet biological satellites Cosmos 782 and 936. The primary effect of space flight on bone turnover was a reversible inhibition of bone formation at the periosteal surface. A simultaneous increase in the length of the periosteal arrest line suggests that bone formation ceased along corresponding portions of that surface. Possible reasons include increased secretion of glucocorticoids and mechanical unloading of the skeleton due to near-weightlessness, while starvation and immobilization are excluded as causes.

Bone pulsating metastasis due to renal cell carcinoma.

PubMed

Cınar, Murat; Derincek, Alihan; Karan, Belgin; Akpınar, Sercan; Tuncay, Cengiz

2010-11-01

Pulsation on the bone cortex surface is a rare condition. Pulsative palpation of the superficial-located bone tumors can be misperceived as an aneurysm. Fifty-eight-year-old man is presented with pulsating bone mass in his proximal tibia. During angiographic examination, hypervascular masses were diagnosed both at right kidney and at right proximal tibia. Renal cell carcinoma was diagnosed after abdominal CT scan. Proximal tibia biopsy was complicated with projectile bleeding.

Effect of modified pectin molecules on the growth of bone cells.

PubMed

Kokkonen, Hanna E; Ilvesaro, Joanna M; Morra, Marco; Schols, Henk A; Tuukkanen, Juha

2007-02-01

The aim of this study was to investigate molecular candidates for bone implant nanocoatings, which could improve biocompatibility of implant materials. Primary rat bone cells and murine preosteoblastic MC3T3-E1 cells were cultured on enzymatically modified hairy regions (MHR-A and MHR-B) of apple pectins. MHRs were covalently attached to tissue culture polystyrene (TCPS) or glass. Uncoated substrata or bone slices were used as controls. Cell attachment, proliferation, and differentiation were investigated with fluorescence and confocal microscopy. Bone cells seem to prefer MHR-B coating to MHR-A coating. On MHR-A samples, the overall numbers as well as proportions of active osteoclasts were diminished compared to those on MHR-B, TCPS, or bone. Focal adhesions indicating attachment of the osteoblastic cells were detected on MHR-B and uncoated controls but not on MHR-A. These results demonstrate the possibility to modify surfaces with pectin nanocoatings.

Development of donor-derived thymic lymphomas after allogeneic bone marrow transplantation in AKR/J mice

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

Yasumizu, R.; Hiai, H.; Sugiura, K.

1988-09-15

The transplantation of bone marrow cells from BALB/c (but not C57BL/6 and C3H/HeN) mice was observed to lead to the development of thymic lymphomas (leukemias) in AKR/J mice. Two leukemic cell lines, CAK1.3 and CAK4.4, were established from the primary culture of two thymic lymphoma, and surface phenotypes of these cell lines found to be H-2d and Thy-1.2+, indicating that these lymphoma cells are derived from BALB/c donor bone marrow cells. Further analyses of surface markers revealed that CAK1.3 is L3T4+ Lyt2+ IL2R-, whereas CAK4.4 is L3T4- Lyt2- IL2R+. Both CAK1.3 and CAK4.4 were transplantable into BALB/c but not AKR/Jmore » mice, further indicating that these cells are of BALB/c bone marrow donor origin. The cells were found to produce XC+-ecotropic viruses, but xenotropic and mink cell focus-forming viruses were undetectable. Inasmuch as thymic lymphomas are derived from bone marrow cells of leukemia-resistant BALB/c strain of mice under the allogeneic environment of leukemia-prone AKR/J mice, this animal model may serve as a useful tool not only for the analysis of leukemic relapse after bone marrow transplantation but also for elucidation of the mechanism of leukemogenesis.« less

Targeted delivery of mesenchymal stem cells to the bone.

PubMed

Yao, Wei; Lane, Nancy E

2015-01-01

Osteoporosis is a disease of excess skeletal fragility that results from estrogen loss and aging. Age related bone loss has been attributed to both elevated bone resorption and insufficient bone formation. We developed a hybrid compound, LLP2A-Ale in which LLP2A has high affinity for the α4β1 integrin on mesenchymal stem cells (MSCs) and alendronate has high affinity for bone. When LLP2A-Ale was injected into mice, the compound directed MSCs to both trabecular and cortical bone surfaces and increased bone mass and bone strength. Additional studies are underway to further characterize this hybrid compound, LLP2A-Ale, and how it can be utilized for the treatment of bone loss resulting from hormone deficiency, aging, and inflammation and to augment bone fracture healing. This article is part of a Special Issue entitled "Stem Cells and Bone". Copyright © 2014 Elsevier Inc. All rights reserved.

Biofunctionalization of a titanium surface with a nano-sawtooth structure regulates the behavior of rat bone marrow mesenchymal stem cells

PubMed Central

Zhang, Wenjie; Li, Zihui; Liu, Yan; Ye, Dongxia; Li, Jinhua; Xu, Lianyi; Wei, Bin; Zhang, Xiuli; Liu, Xuanyong; Jiang, Xinquan

2012-01-01

Background: The topography of an implant surface can serve as a powerful signaling cue for attached cells and can enhance the quality of osseointegration. A series of improved implant surfaces functionalized with nanoscale structures have been fabricated using various methods. Methods: In this study, using an H2O2 process, we fabricated two size-controllable sawtooth-like nanostructures with different dimensions on a titanium surface. The effects of the two nano-sawtooth structures on rat bone marrow mesenchymal stem cells (BMMSCs) were evaluated without the addition of osteoinductive chemical factors. Results: These new surface modifications did not adversely affect cell viability, and rat BMMSCs demonstrated a greater increase in proliferation ability on the surfaces of the nano-sawtooth structures than on a control plate. Furthermore, upregulated expression of osteogenic-related genes and proteins indicated that the nano-sawtooth structures promote osteoblastic differentiation of rat BMMSCs. Importantly, the large nano-sawtooth structure resulted in the greatest cell responses, including increased adhesion, proliferation, and differentiation. Conclusion: The enhanced adhesion, proliferation, and osteogenic differentiation abilities of rat BMMSCs on the nano-sawtooth structures suggest the potential to induce improvements in bone-titanium integration in vivo. Our study reveals the key role played by the nano-sawtooth structures on a titanium surface for the fate of rat BMMSCs and provides insights into the study of stem cell-nanostructure relationships and the related design of improved biomedical implant surfaces. PMID:22927760

Cell morphology, viability, osteocalcin activity, and alkaline phosphatase activity in milled versus unmilled surface of the femoral head.

PubMed

Rhyu, Kee Hyung; Cho, Chang Hoon; Yoon, Kyung Sik; Chun, Young Soo

2016-12-01

To evaluate cellular activity in milled versus unmilled surface of the femoral head in 21 patients who underwent robot-assisted total hip arthroplasty(THA). The femoral head of 21 consecutive patients who underwent robot-assisted THA for osteonecrosis was used. 10 cc of trabecular bone from the entire milled surface was obtained using a curette. The same amount of trabecular bone was obtained at least 1 cm away from the milled surface and served as a matched control. Cell morphology, viability, osteocalcin activity, and alkaline phosphatase activity in milled versus unmilled surface were assessed. Cell morphology of the milled or unmilled surface was comparable; cells were smaller in the milled surface. Cell viability was a mean of 40% higher in the milled surface (107.4% vs. 67.2%, p<0.001); cell viability at 5 time points was comparable in each group. Osteocalcin activity of cells was slightly higher in the milled surface (1.43 vs. 1.24 ng/ml, p=0.69). Alkaline phosphatase activity of cells was slightly higher in the unmilled surface (150 105 vs. 141 789 U/L, p=0.078). The milled and unmilled surfaces of the femoral head were comparable in terms of cell morphology, viability, osteocalcin activity, and alkaline phosphatase activity.

Biomimetic materials for controlling bone cell responses.

PubMed

Drevelle, Olivier; Faucheux, Nathalie

2013-01-01

Bone defects that cannot "heal spontaneously during life" will become an ever greater health problem as populations age. Harvesting autografts has several drawbacks, such as pain and morbidity at both donor and acceptor sites, the limited quantity of material available, and frequently its inappropriate shape. Researchers have therefore developed alternative strategies that involve biomaterials to fill bone defects. These biomaterials must be biocompatible and interact with the surrounding bone tissue to allow their colonization by bone cells and blood vessels. The latest generation biomaterials are not inert; they control cell responses like adhesion, proliferation and differentiation. These biomaterials are called biomimetic materials. This review focuses on the development of third generation materials. We first briefly describe the bone tissue with its cells and matrix, and then how bone cells interact with the extracellular matrix. The next section covers the materials currently used to repair bone defects. Finally, we describe the strategies employed to modify the surface of materials, such as coating with hydroxyapatite and grafting biomolecules.

Inflammatory cytokine response to titanium chemical composition and nanoscale calcium phosphate surface modification.

PubMed

Hamlet, Stephen; Ivanovski, Saso

2011-05-01

Nanoscale surface modification of titanium dental implants with calcium phosphate (CaP) has been shown to achieve superior bone wound healing and osseointegration compared with smooth or microrough titanium surfaces alone. As bone healing has been shown to be influenced by the action of cytokines, this study examined whether changes in cytokine gene expression from RAW 264.7 cells cultured on commercially pure and titanium alloy (Ti-6Al-4V) microrough or nanoscale crystalline CaP-modified surfaces, may influence downstream events in bone wound healing and osseointegration. Whilst no significant difference in the attachment or proliferation of RAW 264.7 cells was observed, the nanoscale CaP-modified surface elicited a gene expression profile with marked down-regulation of a number of pro-inflammatory cytokines and chemokines. Inflammatory cytokine gene expression was further influenced by chemical composition, with lower levels of pro-inflammatory markers noted following exposure of the macrophage-like cells to titanium alloy (Ti-6Al-4V) compared with the commercially pure titanium surface. Down-regulation of pro-inflammatory cytokine gene expression (confirmed at the protein level for TNFα and CCL5), may thus facilitate the enhanced bone wound healing and osseointegration observed clinically with nanoscale calcium phosphate-modified implant surfaces. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. 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.

Effects of titanium nanotubes on the osseointegration, cell differentiation, mineralisation and antibacterial properties of orthopaedic implant surfaces.

PubMed

Su, E P; Justin, D F; Pratt, C R; Sarin, V K; Nguyen, V S; Oh, S; Jin, S

2018-01-01

The development and pre-clinical evaluation of nano-texturised, biomimetic, surfaces of titanium (Ti) implants treated with titanium dioxide (TiO 2 ) nanotube arrays is reviewed. In vitro and in vivo evaluations show that TiO 2 nanotubes on Ti surfaces positively affect the osseointegration, cell differentiation, mineralisation, and anti-microbial properties. This surface treatment can be superimposed onto existing macro and micro porous Ti implants creating a surface texture that also interacts with cells at the nano level. Histology and mechanical pull-out testing of specimens in rabbits indicate that TiO 2 nanotubes improves bone bonding nine-fold (p = 0.008). The rate of mineralisation associated with TiO 2 nanotube surfaces is about three times that of non-treated Ti surfaces. In addition to improved osseointegration properties, TiO 2 nanotubes reduce the initial adhesion and colonisation of Staphylococcus epidermidis Collectively, the properties of Ti implant surfaces enhanced with TiO 2 nanotubes show great promise. Cite this article: Bone Joint J 2018;100-B(1 Supple A):9-16. ©2018 The British Editorial Society of Bone & Joint Surgery.

Concave Pit-Containing Scaffold Surfaces Improve Stem Cell-Derived Osteoblast Performance and Lead to Significant Bone Tissue Formation

PubMed Central

Cusella-De Angelis, Maria Gabriella; Laino, Gregorio; Piattelli, Adriano; Pacifici, Maurizio; De Rosa, Alfredo; Papaccio, Gianpaolo

2007-01-01

Background Scaffold surface features are thought to be important regulators of stem cell performance and endurance in tissue engineering applications, but details about these fundamental aspects of stem cell biology remain largely unclear. Methodology and Findings In the present study, smooth clinical-grade lactide-coglyolic acid 85:15 (PLGA) scaffolds were carved as membranes and treated with NMP (N-metil-pyrrolidone) to create controlled subtractive pits or microcavities. Scanning electron and confocal microscopy revealed that the NMP-treated membranes contained: (i) large microcavities of 80–120 µm in diameter and 40–100 µm in depth, which we termed primary; and (ii) smaller microcavities of 10–20 µm in diameter and 3–10 µm in depth located within the primary cavities, which we termed secondary. We asked whether a microcavity-rich scaffold had distinct bone-forming capabilities compared to a smooth one. To do so, mesenchymal stem cells derived from human dental pulp were seeded onto the two types of scaffold and monitored over time for cytoarchitectural characteristics, differentiation status and production of important factors, including bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF). We found that the microcavity-rich scaffold enhanced cell adhesion: the cells created intimate contact with secondary microcavities and were polarized. These cytological responses were not seen with the smooth-surface scaffold. Moreover, cells on the microcavity-rich scaffold released larger amounts of BMP-2 and VEGF into the culture medium and expressed higher alkaline phosphatase activity. When this type of scaffold was transplanted into rats, superior bone formation was elicited compared to cells seeded on the smooth scaffold. Conclusion In conclusion, surface microcavities appear to support a more vigorous osteogenic response of stem cells and should be used in the design of therapeutic substrates to improve bone repair and bioengineering applications in the future. PMID:17551577

Characterization and Classification of Mesenchymal Stem Cells in Several Species Using Surface Markers for Cell Therapy Purposes.

PubMed

Ghaneialvar, Hori; Soltani, Leila; Rahmani, Hamid Reza; Lotfi, Abbas Sahebghadam; Soleimani, Masoud

2018-01-01

Mesenchymal stem cells are multipotent cells capable of replicating as undifferentiated cells, and have the potential of differentiating into mesenchymal tissue lineages such as osteocytes, adipocytes and chondrocytes. Such lineages can then be used in cell therapy. The aim of present study was to characterize bone marrow derived mesenchymal stem cells in four different species, including: sheep, goat, human and mouse. Human bone-marrow mesenchymal stem cells were purchased, those of sheep and goat were isolated from fetal bone marrow, and those of mouse were collected by washing bone cavity of femur and tibia with DMEM/F12. Using flow-cytometry, they were characterized by CD surface antigens. Furthermore, cells of third passage were examined for their osteogenic and adipogenic differentiation potential by oil red and alizarin red staining respectively. According to the results, CD markers studied in the four groups of mesenchymal stem cells showed a different expression. Goat and sheep expressed CD44 and CD166, and weakly expressed CD34, CD45, CD105 and CD90. Similarly, human and mouse mesenchymal cells expressed CD44, CD166, CD105 and CD90 whereas the expression of CD34 and CD45 was negative. In conclusion, although all mesenchymal stem cells display plastic adherence and tri-lineage differentiation, not all express the same panel of surface antigens described for human mesenchymal stem cells. Additional panel of CD markers are necessary to characterize regenerative potential and possible application of these stem cells in regenerative medicine and implantology.

[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.

Is Graphene a Promising Nano-Material for Promoting Surface Modification of Implants or Scaffold Materials in Bone Tissue Engineering?

PubMed Central

Gu, Ming; Liu, Yunsong; Chen, Tong; Du, Feng; Zhao, Xianghui; Xiong, Chunyang

2014-01-01

Bone tissue engineering promises to restore bone defects that are caused by severe trauma, congenital malformations, tumors, and nonunion fractures. How to effectively promote the proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) or seed cells has become a hot topic in this field. Many researchers are studying the ways of conferring a pro-osteodifferentiation or osteoinductive capability on implants or scaffold materials, where osteogenesis of seed cells is promoted. Graphene (G) provides a new kind of coating material that may confer the pro-osteodifferentiation capability on implants and scaffold materials by surface modification. Here, we review recent studies on the effects of graphene on surface modifications of implants or scaffold materials. The ability of graphene to improve the mechanical and biological properties of implants or scaffold materials, such as nitinol and carbon nanotubes, and its ability to promote the adhesion, proliferation, and osteogenic differentiation of MSCs or osteoblasts have been demonstrated in several studies. Most previous studies were performed in vitro, but further studies will explore the mechanisms of graphene's effects on bone regeneration, its in vivo biocompatibility, its ability to promote osteodifferentiation, and its potential applications in bone tissue engineering. PMID:24447041

Is graphene a promising nano-material for promoting surface modification of implants or scaffold materials in bone tissue engineering?

PubMed

Gu, Ming; Liu, Yunsong; Chen, Tong; Du, Feng; Zhao, Xianghui; Xiong, Chunyang; Zhou, Yongsheng

2014-10-01

Bone tissue engineering promises to restore bone defects that are caused by severe trauma, congenital malformations, tumors, and nonunion fractures. How to effectively promote the proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) or seed cells has become a hot topic in this field. Many researchers are studying the ways of conferring a pro-osteodifferentiation or osteoinductive capability on implants or scaffold materials, where osteogenesis of seed cells is promoted. Graphene (G) provides a new kind of coating material that may confer the pro-osteodifferentiation capability on implants and scaffold materials by surface modification. Here, we review recent studies on the effects of graphene on surface modifications of implants or scaffold materials. The ability of graphene to improve the mechanical and biological properties of implants or scaffold materials, such as nitinol and carbon nanotubes, and its ability to promote the adhesion, proliferation, and osteogenic differentiation of MSCs or osteoblasts have been demonstrated in several studies. Most previous studies were performed in vitro, but further studies will explore the mechanisms of graphene's effects on bone regeneration, its in vivo biocompatibility, its ability to promote osteodifferentiation, and its potential applications in bone tissue engineering.

Chronic administration of Glucagon-like peptide-1 receptor agonists improves trabecular bone mass and architecture in ovariectomised mice.

PubMed

Pereira, M; Jeyabalan, J; Jørgensen, C S; Hopkinson, M; Al-Jazzar, A; Roux, J P; Chavassieux, P; Orriss, I R; Cleasby, M E; Chenu, C

2015-12-01

Some anti-diabetic therapies can have adverse effects on bone health and increase fracture risk. In this study, we tested the skeletal effects of chronic administration of two Glucagon-like peptide-1 receptor agonists (GLP-1RA), increasingly used for type 2 diabetes treatment, in a model of osteoporosis associated bone loss and examined the expression and activation of GLP-1R in bone cells. Mice were ovariectomised (OVX) to induce bone loss and four weeks later they were treated with Liraglutide (LIR) 0.3mg/kg/day, Exenatide (Ex-4) 10 μg/kg/day or saline for four weeks. Mice were injected with calcein and alizarin red prior to euthanasia, to label bone-mineralising surfaces. Tibial micro-architecture was determined by micro-CT and bone formation and resorption parameters measured by histomorphometric analysis. Serum was collected to measure calcitonin and sclerostin levels, inhibitors of bone resorption and formation, respectively. GLP-1R mRNA and protein expression were evaluated in the bone, bone marrow and bone cells using RT-PCR and immunohistochemistry. Primary osteoclasts and osteoblasts were cultured to evaluate the effect of GLP-1RA on bone resorption and formation in vitro. GLP-1RA significantly increased trabecular bone mass, connectivity and structure parameters but had no effect on cortical bone. There was no effect of GLP-1RA on bone formation in vivo but an increase in osteoclast number and osteoclast surfaces was observed with Ex-4. GLP-1R was expressed in bone marrow cells, primary osteoclasts and osteoblasts and in late osteocytic cell line. Both Ex-4 and LIR stimulated osteoclastic differentiation in vitro but slightly reduced the area resorbed per osteoclast. They had no effect on bone nodule formation in vitro. Serum calcitonin levels were increased and sclerostin levels decreased by Ex-4 but not by LIR. Thus, GLP-1RA can have beneficial effects on bone and the expression of GLP-1R in bone cells may imply that these effects are exerted directly on the tissue. Copyright © 2015 Elsevier Inc. All rights reserved.

Fluid shear stress stimulates prostaglandin and nitric oxide release in bone marrow-derived preosteoclast-like cells

NASA Technical Reports Server (NTRS)

McAllister, T. N.; Du, T.; Frangos, J. A.

2000-01-01

Bone is a porous tissue that is continuously perfused by interstitial fluid. Fluid flow, driven by both vascular pressure and mechanical loading, may generate significant shear stresses through the canaliculi as well as along the bone lining at the endosteal surface. Both osteoblasts and osteocytes produce signaling factors such as prostaglandins and nitric in response to fluid shear stress (FSS); however, these humoral agents appear to have more profound affects on osteoclast activity at the endosteal surface. We hypothesized that osteoclasts and preosteoclasts may also be mechanosensitive and that osteoclast-mediated autocrine signaling may be important in bone remodeling. In this study, we investigated the effect of FSS on nitric oxide (NO), prostaglandin E(2) (PGE(2)), and prostacyclin (PGI(2)) release by neonatal rat bone marrow-derived preosteoclast-like cells. These cells were tartrate-resistant acid phosphatase (TRAP) positive, weakly nonspecific esterase (NSE) positive, and capable of fusing into calcitonin-responsive, bone-resorbing, multinucleated cells. Bone marrow-derived preosteoclast-like cells exposed for 6 h to a well-defined FSS of 16 dynes/cm(2) produced NO at a rate of 7.5 nmol/mg protein/h, which was 10-fold that of static controls. This response was completely abolished by 100 microM N(G)-amino-L-arginine (L-NAA). Flow also stimulated PGE(2) production (3.9 microg/mg protein/h) and PGI(2) production (220 pg/mg protein/h). L-NAA attenuated flow-induced PGE(2) production by 30%, suggesting that NO may partially modulate PGE(2) production. This is the first report demonstrating that marrow derived cells are sensitive to FSS and that autocrine signaling in these cells may play an important role in load-induced remodeling and signal transduction in bone. Copyright 2000 Academic Press.

Fabrication of mineralized electrospun PLGA and PLGA/gelatin nanofibers and their potential in bone tissue engineering.

PubMed

Meng, Z X; Li, H F; Sun, Z Z; Zheng, W; Zheng, Y F

2013-03-01

Surface mineralization is an effective method to produce calcium phosphate apatite coating on the surface of bone tissue scaffold which could create an osteophilic environment similar to the natural extracellular matrix for bone cells. In this study, we prepared mineralized poly(D,L-lactide-co-glycolide) (PLGA) and PLGA/gelatin electrospun nanofibers via depositing calcium phosphate apatite coating on the surface of these nanofibers to fabricate bone tissue engineering scaffolds by concentrated simulated body fluid method, supersaturated calcification solution method and alternate soaking method. The apatite products were characterized by the scanning electron microscopy (SEM), Fourier transform-infrared spectroscopy (FT-IR), and X-ray diffractometry (XRD) methods. A large amount of calcium phosphate apatite composed of dicalcium phosphate dihydrate (DCPD), hydroxyapatite (HA) and octacalcium phosphate (OCP) was deposited on the surface of resulting nanofibers in short times via three mineralizing methods. A larger amount of calcium phosphate was deposited on the surface of PLGA/gelatin nanofibers rather than PLGA nanofibers because gelatin acted as nucleation center for the formation of calcium phosphate. The cell culture experiments revealed that the difference of morphology and components of calcium phosphate apatite did not show much influence on the cell adhesion, proliferation and activity. Copyright © 2012 Elsevier B.V. All rights reserved.

A small proportion of mesenchymal stem cells strongly expresses functionally active CXCR4 receptor capable of promoting migration to bone marrow.

PubMed

Wynn, Robert F; Hart, Claire A; Corradi-Perini, Carla; O'Neill, Liam; Evans, Caroline A; Wraith, J Ed; Fairbairn, Leslie J; Bellantuono, Ilaria

2004-11-01

Homing of bone marrow stromal cells (MSCs) to bone and bone marrow after transplantation, important for the correction of conditions such as metabolic storage disorders, can occur but with poor efficiency. Substantial improvements in engraftment will be required in order to derive a clinical benefit from MSC transplantation. Chemokines are the most important factors controlling cellular migration. Stromal-derived factor-1 (SDF-1) has been shown to be critical in promoting the migration of cells to the bone marrow, via its specific receptor CXCR4. The aim of our study was to investigate CXCR4 expression on MSCs and its role in mediating migration to bone marrow. We show that CXCR4, although present at the surface of a small subset of MSCs, is important for mediating specific migration of these cells to bone marrow.

Effect of microporosity on scaffolds for bone tissue engineering

PubMed Central

Zhang, Ke; Fan, Yubo; Dunne, Nicholas; Li, Xiaoming

2018-01-01

Abstract Microporosity has a critical role in improving the osteogenesis of scaffolds for bone tissue engineering. Although the exact mechanism, by which it promotes new bone formation, is not well recognized yet, the related hypothesis can be found in many previous studies. This review presents those possible mechanisms about how the microporosity enhances the osteogenic-related functions of cells in vitro and the osteogenic activity of scaffolds in vivo. In summary, the increased specific surface areas by microporosity can offer more protein adsorption sites and accelerate the release of degradation products, which facilitate the interactions between scaffolds and cells. Meanwhile, the unique surface properties of microporous scaffolds have a considerable effect on the protein adsorption. Moreover, capillary force generated by the microporosity can improve the attachment of bone-related cells on the scaffolds surface, and even make the cells achieve penetration into the micropores smaller than them. This review also pays attention to the relationship between the biological and mechanical properties of microporous scaffolds. Although lots of achievements have been obtained, there is still a lot of work to do, some of which has been proposed in the conclusions and perspectives part. PMID:29644093

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.

Surface Functionalization of Orthopedic Titanium Implants with Bone Sialoprotein

PubMed Central

Ritz, Ulrike; Ackermann, Angelika; Anthonissen, Joris; Kaufmann, Kerstin B.; Brendel, Christian; Götz, Hermann; Rommens, Pol M.; Hofmann, Alexander

2016-01-01

Orthopedic implant failure due to aseptic loosening and mechanical instability remains a major problem in total joint replacement. Improving osseointegration at the bone-implant interface may reduce micromotion and loosening. Bone sialoprotein (BSP) has been shown to enhance bone formation when coated onto titanium femoral implants and in rat calvarial defect models. However, the most appropriate method of BSP coating, the necessary level of BSP coating, and the effect of BSP coating on cell behavior remain largely unknown. In this study, BSP was covalently coupled to titanium surfaces via an aminosilane linker (APTES), and its properties were compared to BSP applied to titanium via physisorption and untreated titanium. Cell functions were examined using primary human osteoblasts (hOBs) and L929 mouse fibroblasts. Gene expression of specific bone turnover markers at the RNA level was detected at different intervals. Cell adhesion to titanium surfaces treated with BSP via physisorption was not significantly different from that of untreated titanium at any time point, whereas BSP application via covalent coupling caused reduced cell adhesion during the first few hours in culture. Cell migration was increased on titanium disks that were treated with higher concentrations of BSP solution, independent of the coating method. During the early phases of hOB proliferation, a suppressive effect of BSP was observed independent of its concentration, particularly when BSP was applied to the titanium surface via physisorption. Although alkaline phosphatase activity was reduced in the BSP-coated titanium groups after 4 days in culture, increased calcium deposition was observed after 21 days. In particular, the gene expression level of RUNX2 was upregulated by BSP. The increase in calcium deposition and the stimulation of cell differentiation induced by BSP highlight its potential as a surface modifier that could enhance the osseointegration of orthopedic implants. Both physisorption and covalent coupling of BSP are similarly effective, feasible methods, although a higher BSP concentration is recommended. PMID:27111551

Different culture media affect growth characteristics, surface marker distribution and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells.

PubMed

Hagmann, Sebastien; Moradi, Babak; Frank, Sebastian; Dreher, Thomas; Kämmerer, Peer Wolfgang; Richter, Wiltrud; Gotterbarm, Tobias

2013-07-30

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) play an important role in modern tissue engineering, while distinct variations of culture media compositions and supplements have been reported. Because MSCs are heterogeneous regarding their regenerative potential and their surface markers, these parameters were compared in four widely used culture media compositions. MSCs were isolated from bone marrow and expanded in four established cell culture media. MSC yield/1000 MNCs, passage time and growth index were observed. In P4, typical MSC surface markers were analysed by fluorescence cytometry. Additionally, chondrogenic, adipogenic and osteogenic differentiation potential were evaluated. Growth index and P0 cell yield varied importantly between the media. The different expansion media had a significant influence on the expression of CD10, CD90, CD105, CD140b CD146 and STRO-1. While no significant differences were observed regarding osteogenic and adipogenic differentiation, chondrogenic differentiation was superior in medium A as reflected by GAG/DNA content. The choice of expansion medium can have a significant influence on growth, differentiation potential and surface marker expression of mesenchymal stromal cells, which is of fundamental importance for tissue engineering procedures.

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.

3D printing of biomaterials with mussel-inspired nanostructures for tumor therapy and tissue regeneration.

PubMed

Ma, Hongshi; Luo, Jian; Sun, Zhe; Xia, Lunguo; Shi, Mengchao; Liu, Mingyao; Chang, Jiang; Wu, Chengtie

2016-12-01

Primary bone cancer brings patients great sufferings. To deal with the bone defects resulted from cancer surgery, biomaterials with good bone-forming ability are necessary to repair bone defects. Meanwhile, in order to prevent possible tumor recurrence, it is essential that the remaining tumor cells around bone defects are completely killed. However, there are few biomaterials with the ability of both cancer therapy and bone regeneration until now. Here, we fabricated a 3D-printed bioceramic scaffold with a uniformly self-assembled Ca-P/polydopamine nanolayer surface. Taking advantage of biocompatibility, biodegradability and the excellent photothermal effect of polydopamine, the bifunctional scaffolds with mussel-inspired nanostructures could be used as a satisfactory and controllable photothermal agent, which effectively induced tumor cell death in vitro, and significantly inhibited tumor growth in mice. In addition, owing to the nanostructured surface, the prepared polydopamine-modified bioceramic scaffolds could support the attachment and proliferation of rabbit bone mesenchymal stem cells (rBMSCs), and significantly promoted the formation of new bone tissues in rabbit bone defects even under photothermal treatment. Therefore, the mussel-inspired nanostructures in 3D-printed bioceramic exhibited a remarkable capability for both cancer therapy and bone regeneration, offering a promising strategy to construct bifunctional biomaterials which could be widely used for therapy of tumor-induced tissue defects. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effect of electrical polarization of hydroxyapatite ceramics on new bone formation.

PubMed

Itoh, S; Nakamura, S; Kobayashi, T; Shinomiya, K; Yamashita, K; Itoh, S

2006-03-01

Large surface charges can be induced on hydroxyapatite (HAp) ceramics by proton transport polarization, but this does not affect beta-tricalcium phosphate (TCP) because of its low polarizability. We wished to examine differences in osteogenic cell activity and new bone growth between positively or negatively surface-charged HAp and HAp/TCP plates using a calvarial bone defect model. In the first group of rats, test pieces were placed with their positively charged surfaces face down on the dura mater. In the second group, test pieces were placed with their negatively charged surfaces face down on the dura mater. A third group received noncharged test pieces. Histological examination, including enzymatic staining for osteoblasts and osteoclasts, was carried out. While no bone formation was observed at the pericranium, direct bone formation on the cranial bone debris and new bone growth expanded from the margins of the sites of injury to bridge across both the positively and negatively charged surfaces of HAp and HAp/TCP plates occurred. Electrical polarization of implanted plates, including positive charge, led to enhanced osteoblast activity, though decreased osteoclast activity was seen on the positively charged plate surface. Thus, polarization of HAp ceramics may modulate new bone formation and resorption.

[Biocompatibility research of true bone ceramics].

PubMed

Qiao, Wei; Ren, Xiaoqi; Shi, Hao; Li, Jing; Yang, Ting; Ma, Shaoying; Zhao, Yaping; Su, Chengzhong; Li, Baoxing

2017-10-01

To investigate the biocompatibility of true bone ceramic (TBC) and provide experimental basis for clinic application. TBC was prepared from healthy adult bovine cancellous bone by deproteinization and high temperature calcinations. Mouse fibroblast cell line (L929 cells) were cultured with the leaching liquor of TBC in vitro , and the cytotoxicity was evaluated at 2nd, 4th, and 7th days. L929 cells were inoculated into the TBC and cultured for 4 days. The cell adhesion and proliferation on the surface of the TBC were observed by scanning electron microscopy, and evaluated the cell compatibility of TBC. Ten New Zealand white rabbits were divided into 2 groups, and drilled holes at the tibia of both hind limbs. TBC and hydroxyapatite (HA) were implanted into the left side (experimental group) and the right side (control group), respectively. And the biocompatibility of TBC was evaluated by general observation and histological observation at 4 and 26 weeks after implantation. Cytotoxicity test showed that the cytotoxicity level of leaching liquor of TBC was grade 0-1. Cell compatibility experiments showed that the L929 cells adhered well on the surface of TBC and migrated into the pores. The implantation test in vivo showed that experimental group and control group both had mild or moderate inflammatory response at 4 weeks, and new bone formation occurred. At 26 weeks, there was no inflammatory reaction observed in both groups, and new bone formation was observed in varying degrees. TBC have good biocompatibility and can be used to repair bone defect in clinic.

Interactions between bone cells and biomaterials: An update.

PubMed

Beauvais, Sabrina; Drevelle, Olivier; Jann, Jessica; Lauzon, Marc-Antoine; Foruzanmehr, Mohammadreza; Grenier, Guillaume; Roux, Sophie; Faucheux, Nathalie

2016-06-01

As the populations of the Western world become older, they will suffer more and more from bone defects related to osteoporosis (non-union fractures, vertebral damages), cancers (malignant osteolysis) and infections (osteomyelitis). Autografts are usually used to fill these defects, but they have several drawbacks such as morbidity at the donor site and the amount and quality of bone that can be harvested. Recent scientific milestones made in biomaterials development were shown to be promising to overcome these limitations. Cell interactions with biomaterials can be improved by adding at their surface functional groups such as adhesive peptides and/or growth factors. The development of such biomimetic materials able to control bone cell responses can only proceed if it is based on a sound understanding of bone cell behavior and regulation. This review focuses on bone physiology and the regulation of bone cell differentiation and function, and how the latest advances in biomimetic materials can be translated within promising clinical outcomes.

Ultrasonic and electromagnetic enhancement of a culture of human SAOS-2 osteoblasts seeded onto a titanium plasma-spray surface.

PubMed

Fassina, Lorenzo; Saino, Enrica; Sbarra, Maria Sonia; Visai, Livia; Cusella De Angelis, Maria Gabriella; Mazzini, Giuliano; Benazzo, Francesco; Magenes, Giovanni

2009-06-01

Several studies suggest that the surface coating of titanium could play an important role in bone tissue engineering. In the present study, we have followed a particular biomimetic strategy where ultrasonically or electromagnetically stimulated SAOS-2 human osteoblasts proliferated and built their extracellular matrix on a titanium plasma-spray surface. In comparison with control conditions, the ultrasonic stimulation (average power, 149 mW; frequency, 1.5 MHz) and the electromagnetic stimulation (magnetic field intensity, 2 mT; frequency, 75 Hz) caused higher cell proliferation, and increased surface coating with decorin, osteocalcin, osteopontin, and type I collagen together with higher incorporation of calcium and phosphorus inside the extracellular matrix. The immunofluorescence related to the preceding bone matrix proteins showed their colocalization in the cell-rich areas. The use of the two physical stimulations aimed at obtaining the coating of the rough titanium plasma-spray surface in terms of cell colonization and deposition of extracellular matrix. The superficially cultured biomaterial could be theoretically used, in clinical applications, as an implant for bone repair.

Human Vitronectin-Derived Peptide Covalently Grafted onto Titanium Surface Improves Osteogenic Activity: A Pilot In Vivo Study on Rabbits.

PubMed

Cacchioli, Antonio; Ravanetti, Francesca; Bagno, Andrea; Dettin, Monica; Gabbi, Carlo

2009-10-01

Peptide and protein exploitation for the biochemical functionalization of biomaterial surfaces allowed fabricating biomimetic devices able to evoke and promote specific and advantageous cell functions in vitro and in vivo. In particular, cell adhesion improvement to support the osseointegration of implantable devices has been thoroughly investigated. This study was aimed at checking the biological activity of the (351-359) human vitronectin precursor (HVP) sequence, mapped on the human vitronectin protein; the peptide was covalently linked to the surface of titanium cylinders, surgically inserted in the femurs of New Zealand white rabbits and analyzed at short experimental time points (4, 9, and 16 days after surgery). To assess the osteogenic activity of the peptide, three vital fluorochromic bone markers were used (calcein green, xylenol orange, and calcein blue) to stain the areas of newly grown bone. Static and dynamic histomorphometric parameters were measured at the bone-implant interface and at different distances from the surface. The biological role of the (351-359)HVP sequence was checked by comparing peptide-grafted samples and controls, analyzing how and how much its effects change with time across the bone regions surrounding the implant surface. The results obtained reveal a major activity of the investigated peptide 4 days after surgery, within the bone region closest to the implant surface, and larger bone to implant contact 9 and 16 days after surgery. Thus, improved primary fixation of endosseous devices can be foreseen, resulting in an increased osteointegration.

Preosteocytes/Osteocytes Have the Potential to Dedifferentiate Becoming a Source of Osteoblasts

PubMed Central

Torreggiani, Elena; Matthews, Brya G.; Pejda, Slavica; Matic, Igor; Horowitz, Mark C.; Grcevic, Danka; Kalajzic, Ivo

2013-01-01

Presently there is no clear evidence for the ability of mature osteogenic lineage cells to dedifferentiate. In order to identify and trace mature osteogenic lineage cells, we have utilized transgenic mouse models in which the dentin matrix protein 1 (Dmp1) promoter drives expression of GFP (active marker) or Cre recombinase (historic label) in preosteocytes/osteocytes. In long bone chip outgrowth cultures, in which cells on the bone surface were enzymatically removed, cells with previous activity of the Dmp1 promoter migrated onto plastic and down-regulated Dmp1-GFP expression. Dmp1Cre-labeled cells from these cultures had the potential to re-differentiate into the osteogenic lineage, while the negative population showed evidence of adipogenesis. We observed numerous Dmp1Cre-labeled osteoblasts on the surface of bone chips following their in vivo transplantation. Our data indicate that cells embedded in bone matrix are motile, and once given access to the extra bony milieu will migrate out of their lacunae. This population of cells is phenotypically and functionally heterogeneous in vitro. Once the preosteocytes/osteocytes leave lacunae, they can dedifferentiate, potentially providing an additional source of functional osteoblasts. PMID:24040401

Enhanced osteoblast adhesion on nanostructured selenium compacts for anti-cancer orthopedic applications.

PubMed

Tran, Phong; Webster, Thomas J

2008-01-01

Metallic bone implants possess numerous problems limiting their long-term efficacy, such as poor prolonged osseointegration, stress shielding, and corrosion under in vivo environments. Such problems are compounded for bone cancer patients since numerous patients receive orthopedic implants after cancerous bone resection. Unfortunately, current orthopedic materials were not originally developed to simultaneously increase healthy bone growth (as in traditional orthopedic implant applications) while inhibiting cancerous bone growth. The long-term objective of the present research is to investigate the use of nano-rough selenium to prevent bone cancer from re-occurring while promoting healthy bone growth for this select group of cancer patients. Selenium is a well known anti-cancer chemical. However, what is not known is how healthy bone cells interact with selenium. To determine this, selenium, spherical or semispherical shots, were pressed into cylindrical compacts and these compacts were then etched using 1N NaOH to obtain various surface structures ranging from the micron, submicron to nano scales. Changes in surface chemistry were also analyzed. Through these etching techniques, results of this study showed that biologically inspired surface roughness values were created on selenium compacts to match that of natural bone roughness. Moreover, results showed that healthy bone cell adhesion increased with greater nanometer selenium roughness (more closely matching that of titanium). In this manner, this study suggests that nano-rough selenium should be further tested for orthopedic applications involving bone cancer treatment.

Enhanced osteoblast adhesion on nanostructured selenium compacts for anti-cancer orthopedic applications

PubMed Central

Tran, Phong; Webster, Thomas J

2008-01-01

Metallic bone implants possess numerous problems limiting their long-term efficacy, such as poor prolonged osseointegration, stress shielding, and corrosion under in vivo environments. Such problems are compounded for bone cancer patients since numerous patients receive orthopedic implants after cancerous bone resection. Unfortunately, current orthopedic materials were not originally developed to simultaneously increase healthy bone growth (as in traditional orthopedic implant applications) while inhibiting cancerous bone growth. The long-term objective of the present research is to investigate the use of nano-rough selenium to prevent bone cancer from re-occurring while promoting healthy bone growth for this select group of cancer patients. Selenium is a well known anti-cancer chemical. However, what is not known is how healthy bone cells interact with selenium. To determine this, selenium, spherical or semispherical shots, were pressed into cylindrical compacts and these compacts were then etched using 1N NaOH to obtain various surface structures ranging from the micron, submicron to nano scales. Changes in surface chemistry were also analyzed. Through these etching techniques, results of this study showed that biologically inspired surface roughness values were created on selenium compacts to match that of natural bone roughness. Moreover, results showed that healthy bone cell adhesion increased with greater nanometer selenium roughness (more closely matching that of titanium). In this manner, this study suggests that nano-rough selenium should be further tested for orthopedic applications involving bone cancer treatment. PMID:18990948

Enhanced adhesion of osteoblastic cells on polystyrene films by independent control of surface topography and wettability.

PubMed

Yang, Seung Yun; Kim, Eung-Sam; Jeon, Gumhye; Choi, Kwan Yong; Kim, Jin Kon

2013-04-01

We independently controlled surface topography and wettability of polystyrene (PS) films by CF4 and oxygen plasma treatments, respectively, to evaluate the adhesion and proliferation of human fetal osteoblastic (hFOB) cells on the films. Among the CF4 plasma-treated PS films with the average surface roughness ranging from 0.9 to 70 nm, the highest adhesion of hFOB cells was observed on a PS film with roughness of ~11 nm. When this film was additionally treated by oxygen plasma to provide a hydrophilic surface with a contact angle less than 10°, the proliferation of bone-forming cell was further enhanced. Thus, the plasma-based independent modification of PS film into an optimum nanotexture for human osteoblast cells could be appplied to materials used in bone tissue engineering. Copyright © 2012 Elsevier B.V. All rights reserved.

HA and double-layer HA-P2O5/CaO glass coatings: influence of chemical composition on human bone marrow cells osteoblastic behavior.

PubMed

Ferraz, M P; Fernandes, M H; Santos, J D; Monteiro, F J

2001-07-01

Human osteoblastic bone marrow derived cells were cultured for 28 days onto the surface of a glass reinforced hydroxyapatite (HA) composite and a commercial type HA plasma sprayed coatings, both in the "as-received" condition and after an immersion treatment with culture medium during 21 days. Cell proliferation and differentiation were analyzed as a function of the chemical composition of the coatings and the immersion treatment. Cell attachment, growth and differentiation of osteoblastic bone marrow cells seeded onto "as-received" plasma sprayed coatings were strongly affected by the time-dependent variation of the surface structure occurring during the first hours of culture. Initial interactions leading to higher amounts of adsorbed protein and zeta potential shifts towards negative charges appeared to result in surface structures with better biological performance. Cultures grown onto the pretreated coatings showed higher rate of cell proliferation and increased functional activity, as compared to those grown onto the corresponding "as-received" materials. However, the cell behavior was similar in the glass composite and HA coatings. The results showed that the glass composites present better characteristics for bone cell growth and function than HA. In addition, this work also provide evidence that the biological performance of the glass composites can be modulated and improved by manipulations in the chemical composition, namely in the content of glass added to HA. Copyright 2001 Kluwer Academic Publishers

Enhanced MC3T3-E1 preosteoblast response and bone formation on the addition of nano-needle and nano-porous features to microtopographical titanium surfaces.

PubMed

Zhuang, X-M; Zhou, B; Ouyang, J-L; Sun, H-P; Wu, Y-L; Liu, Q; Deng, F-L

2014-08-01

Micro/nanotopographical modifications on titanium surfaces constitute a new process to increase osteoblast response to enhance bone formation. In this study, we utilized alkali heat treatment at high (SB-AH1) and low temperatures (SB-AH2) to nano-modify sandblasted titanium with microtopographical surfaces. Then, we evaluated the surface properties, biocompatibility and osteogenic capability of SB-AH1 and SB-AH2 in vitro and in vivo, and compared these with conventional sandblast-acid etching (SLA) and Ti control surfaces. SB-AH1 and SB-AH2 surfaces exhibited micro/nanotopographical modifications of nano-needle structures and nano-porous network layers, respectively, compared with the sole microtopographical surface of macro and micro pits on the SLA surface and the relatively smooth surface on the Ti control. SB-AH1 and SB-AH2 showed different roughness and elemental components, but similar wettability. MC3T3-E1 preosteoblasts anchored closely on the nanostructures of SB-AH1 and SB-AH2 surfaces, and these two surfaces more significantly enhanced cell proliferation and alkaline phosphatase (ALP) activity than others, while the SB-AH2 surface exhibited better cell proliferation and higher ALP activity than SB-AH1. All four groups of titanium domes with self-tapping screws were implanted in rabbit calvarial bone models, and these indicated that SB-AH1 and SB-AH2 surfaces achieved better peri-implant bone formation and implant stability, while the SB-AH2 surface achieved the best percentage of bone-implant contact (BIC%). Our study demonstrated that the micro/nanotopographical surface generated by sandblasting and alkali heat treatment significantly enhanced preosteoblast proliferation, ALP activity and bone formation in vitro and in vivo, and nano-porous network topography may further induce better preosteoblast proliferation, ALP activity and BIC%.

Hit rates and radiation doses to nuclei of bone lining cells from alpha-particle-emitting radionuclides

NASA Technical Reports Server (NTRS)

Polig, E.; Jee, W. S.; Kruglikov, I. L.

1992-01-01

Factors relating the local concentration of a bone-seeking alpha-particle emitter to the mean hit rate have been determined for nuclei of bone lining cells using a Monte Carlo procedure. Cell nuclei were approximated by oblate spheroids with dimensions and location taken from a previous histomorphometric study. The Monte Carlo simulation is applicable for planar and diffuse labels at plane or cylindrical bone surfaces. Additionally, the mean nuclear dose per hit, the dose mean per hit, the mean track segment length and its second moment, the percentage of stoppers, and the frequency distribution of the dose have been determined. Some basic features of the hit statistics for bone lining cells have been outlined, and the consequences of existing standards of radiation protection with regard to the hit frequency to cell nuclei are discussed.

[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.

The Foreign Body Giant Cell Cannot Resorb Bone, But Dissolves Hydroxyapatite Like Osteoclasts.

PubMed

ten Harkel, Bas; Schoenmaker, Ton; Picavet, Daisy I; Davison, Noel L; de Vries, Teun J; Everts, Vincent

2015-01-01

Foreign body multinucleated giant cells (FBGCs) and osteoclasts share several characteristics, like a common myeloid precursor cell, multinuclearity, expression of tartrate-resistant acid phosphatase (TRAcP) and dendritic cell-specific transmembrane protein (DC-STAMP). However, there is an important difference: osteoclasts form and reside in the vicinity of bone, while FBGCs form only under pathological conditions or at the surface of foreign materials, like medical implants. Despite similarities, an important distinction between these cell types is that osteoclasts can resorb bone, but it is unknown whether FBGCs are capable of such an activity. To investigate this, we differentiated FBGCs and osteoclasts in vitro from their common CD14+ monocyte precursor cells, using different sets of cytokines. Both cell types were cultured on bovine bone slices and analyzed for typical osteoclast features, such as bone resorption, presence of actin rings, formation of a ruffled border, and characteristic gene expression over time. Additionally, both cell types were cultured on a biomimetic hydroxyapatite coating to discriminate between bone resorption and mineral dissolution independent of organic matrix proteolysis. Both cell types differentiated into multinucleated cells on bone, but FBGCs were larger and had a higher number of nuclei compared to osteoclasts. FBGCs were not able to resorb bone, yet they were able to dissolve the mineral fraction of bone at the surface. Remarkably, FBGCs also expressed actin rings, podosome belts and sealing zones--cytoskeletal organization that is considered to be osteoclast-specific. However, they did not form a ruffled border. At the gene expression level, FBGCs and osteoclasts expressed similar levels of mRNAs that are associated with the dissolution of mineral (e.g., anion exchange protein 2 (AE2), carbonic anhydrase 2 (CAII), chloride channel 7 (CIC7), and vacuolar-type H+-ATPase (v-ATPase)), in contrast the matrix degrading enzyme cathepsin K, which was hardly expressed by FBGCs. Functionally, the latter cells were able to dissolve a biomimetic hydroxyapatite coating in vitro, which was blocked by inhibiting v-ATPase enzyme activity. These results show that FBGCs have the capacity to dissolve the mineral phase of bone, similar to osteoclasts. However, they are not able to digest the matrix fraction of bone, likely due to the lack of a ruffled border and cathepsin K.

The Foreign Body Giant Cell Cannot Resorb Bone, But Dissolves Hydroxyapatite Like Osteoclasts

PubMed Central

ten Harkel, Bas; Schoenmaker, Ton; Picavet, Daisy I.; Davison, Noel L.; de Vries, Teun J.; Everts, Vincent

2015-01-01

Foreign body multinucleated giant cells (FBGCs) and osteoclasts share several characteristics, like a common myeloid precursor cell, multinuclearity, expression of tartrate-resistant acid phosphatase (TRAcP) and dendritic cell-specific transmembrane protein (DC-STAMP). However, there is an important difference: osteoclasts form and reside in the vicinity of bone, while FBGCs form only under pathological conditions or at the surface of foreign materials, like medical implants. Despite similarities, an important distinction between these cell types is that osteoclasts can resorb bone, but it is unknown whether FBGCs are capable of such an activity. To investigate this, we differentiated FBGCs and osteoclasts in vitro from their common CD14+ monocyte precursor cells, using different sets of cytokines. Both cell types were cultured on bovine bone slices and analyzed for typical osteoclast features, such as bone resorption, presence of actin rings, formation of a ruffled border, and characteristic gene expression over time. Additionally, both cell types were cultured on a biomimetic hydroxyapatite coating to discriminate between bone resorption and mineral dissolution independent of organic matrix proteolysis. Both cell types differentiated into multinucleated cells on bone, but FBGCs were larger and had a higher number of nuclei compared to osteoclasts. FBGCs were not able to resorb bone, yet they were able to dissolve the mineral fraction of bone at the surface. Remarkably, FBGCs also expressed actin rings, podosome belts and sealing zones—cytoskeletal organization that is considered to be osteoclast-specific. However, they did not form a ruffled border. At the gene expression level, FBGCs and osteoclasts expressed similar levels of mRNAs that are associated with the dissolution of mineral (e.g., anion exchange protein 2 (AE2), carbonic anhydrase 2 (CAII), chloride channel 7 (CIC7), and vacuolar-type H+-ATPase (v-ATPase)), in contrast the matrix degrading enzyme cathepsin K, which was hardly expressed by FBGCs. Functionally, the latter cells were able to dissolve a biomimetic hydroxyapatite coating in vitro, which was blocked by inhibiting v-ATPase enzyme activity. These results show that FBGCs have the capacity to dissolve the mineral phase of bone, similar to osteoclasts. However, they are not able to digest the matrix fraction of bone, likely due to the lack of a ruffled border and cathepsin K. PMID:26426806

The in vitro viability and growth of fibroblasts cultured in the presence of different bone grafting materials (NanoBone and Straumann Bone Ceramic).

PubMed

Kauschke, E; Rumpel, E; Fanghänel, J; Bayerlein, T; Gedrange, T; Proff, P

2006-02-01

Different clinical applications, including dentistry, are making increasing demands on bone grafting material. In the present study we have analysed the viability, proliferation and growth characteristics of fibroblasts cultured in vitro together with two different bone grafting materials, NanoBone and Straumann Bone Ceramic, over a period of 24 and 28 days respectively. Viability was measured at least every 72 hours by using the alamarBlue assay, a test that measures quantitatively cell proliferation and viability but does not require cell fixation or extraction. After one week of culture fibroblast viability was as high as in controls for both grafting materials and remained high (> 90%) for the duration of the experiment. Cell growth was evaluated microscopically. Scanning electron microscopy revealed a dense fibroblast growth at the surface of both bone grafting materials after three weeks of in vitro culture. Generally, our in vitro analyses contribute to further insights into cell - scaffold interactions.

Fortifying the Bone-Implant Interface Part 1: An In Vitro Evaluation of 3D-Printed and TPS Porous Surfaces.

PubMed

MacBarb, Regina F; Lindsey, Derek P; Bahney, Chelsea S; Woods, Shane A; Wolfe, Mark L; Yerby, Scott A

2017-01-01

An aging society and concomitant rise in the incidence of impaired bone health have led to the need for advanced osteoconductive spinal implant surfaces that promote greater biological fixation ( e.g. for interbody fusion cages, sacroiliac joint fusion implants, and artificial disc replacements). Additive manufacturing, i.e. 3D-printing, may improve bone integration by generating biomimetic spinal implant surfaces that mimic bone morphology. Such surfaces may foster an enhanced cellular response compared to traditional implant surfacing processes. This study investigated the response of human osteoblasts to additive manufactured (AM) trabecular-like titanium implant surfaces compared to traditionally machined base material with titanium plasma spray (TPS) coated surfaces, with and without a nanocrystalline hydroxyapatite (HA) coating. For TPS-coated discs, wrought Ti6Al4V ELI was machined and TPS-coating was applied. For AM discs, Ti6Al4V ELI powder was 3D-printed to form a solid base and trabecular-like porous surface. The HA-coating was applied via a precipitation dip-spin method. Surface porosity, pore size, thickness, and hydrophilicity were characterized. Initial cell attachment, proliferation, alkaline phosphatase (ALP) activity, and calcium production of hFOB cells ( n =5 per group) were measured. Cells on AM discs exhibited expedited proliferative activity. While there were no differences in mean ALP expression and calcium production between TPS and AM discs, calcium production on the AM discs trended 48% higher than on TPS discs ( p =0.07). Overall, HA-coating did not further enhance results compared to uncoated TPS and AM discs. Results demonstrate that additive manufacturing allows for controlled trabecular-like surfaces that promote earlier cell proliferation and trends toward higher calcium production than TPS coating. Results further showed that nanocrystalline HA may not provide an advantage on porous titanium surfaces. Additive manufactured porous titanium surfaces may induce a more osteogenic environment compared to traditional TPS, and thus present as an attractive alternative to TPS-coating for orthopedic spinal implants.

Three-dimensional polycaprolactone-hydroxyapatite scaffolds combined with bone marrow cells for cartilage tissue engineering.

PubMed

Wei, Bo; Yao, Qingqiang; Guo, Yang; Mao, Fengyong; Liu, Shuai; Xu, Yan; Wang, Liming

2015-08-01

The goal of this study was to investigate the chondrogenic potential of three-dimensional polycaprolactone-hydroxyapatite (PCL-HA) scaffolds loaded with bone marrow cells in vitro and the effect of PCL-HA scaffolds on osteochondral repair in vivo. Here, bone marrow was added to the prepared PCL-HA scaffolds and cultured in chondrogenic medium for 10 weeks. Osteochondral defects were created in the trochlear groove of 29 knees in 17 New Zealand white rabbits, which were then divided into four groups that underwent: implantation of PCL-HA scaffolds (left knee, n = 17; Group 1), microfracture (right knee, n = 6; Group 2), autologous osteochondral transplantation (right knee, n = 6; Group 3), and no treatment (right knee, n = 5; Control). Extracellular matrix produced by bone marrow cells covered the surface and filled the pores of PCL-HA scaffolds after 10 weeks in culture. Moreover, many cell-laden cartilage lacunae were observed, and cartilage matrix was concentrated in the PCL-HA scaffolds. After a 12-week repair period, Group 1 showed excellent vertical and lateral integration with host bone, but incomplete cartilage regeneration and matrix accumulation. An uneven surface of regenerated cartilage and reduced distribution of cartilage matrix were observed in Group 2. In addition, abnormal bone growth and unstable integration between repaired and host tissues were detected. For Group 3, the integration between transplanted and host cartilage was interrupted. Our findings indicate that the PCL-HA scaffolds loaded with bone marrow cells improved chondrogenesis in vitro and implantation of PCL-HA scaffolds for osteochondral repairenhanced integration with host bone. However, cartilage regeneration remained unsatisfactory. The addition of trophic factors or the use of precultured cell-PCL-HA constructs for accelerated osteochondral repair requires further investigation. © The Author(s) 2015.

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.

A new implant with solid core and porous surface: the biocompatability with bone.

PubMed

Yang, Xu; Wang, Dihua; Liang, Youde; Yin, Huayi; Zhang, Shuang; Jiang, Tao; Wang, Yining; Zhou, Yi

2014-07-01

This research investigated osteogenic potencies of Farthing-Fray-Chen Titanium (FFcTi) implant with transitional porous-solid structure. The material characteristics, biomechanical property, osteogenic performances were assessed. FFcTi showed similar roughness as sand-blasted and acid etched titanium (SA), but was more hydrophilic than SA and machined commercial pure titanium (MA). Young's modulus of FFcTi implant in compressive tests was 15.8 ± 6.3 GPa, which was close to bone. In vitro observations manifested excellent spreading abilities of MC3T3-E1 cell on FFcTi and SA. Adhesion rates of MC3T3-E1 cells at 4 h gradually decreased on MA, SA, and FFcTi surfaces (MA > SA, p < 0.01; SA > FFcTi, p < 0.05), while cell proliferation ability on FFcTi was weaker than MA during 1-6 days (p < 0.01) and similar to MA and SA in day 11. ALP activity of cells on FFcTi at 14 day was higher than MA and lower than SA (p < 0.01). In a bone defect model of rabbits, BIC and bone volum ratio within 50 μm were significantly higher for FFcTi than MA (BIC, p < 0.01; BT0.05, p < 0.05) while bone volume ratio within 100 and 500 μm were of no differences. Micro CT analysis also showed similar results to the histomorphometric data. Thus, we conclude that FFcTi with melting sphere based multiporous structure has a hydrophilic, rough surface, and close modulus to bone. In vitro, its low proliferation and ALP activity promotion were similar to other micro scale roughed surface. In vivo test showed better osteogenesis ability when compared with MA at least in 2 weeks. Thus, this Farthing-Fray-Chen Titanium implant seems to hold considerable potential for bone implant applications. © 2013 Wiley Periodicals, Inc.

Cell-printing and transfer technology applications for bone defects in mice.

PubMed

Tsugawa, Junichi; Komaki, Motohiro; Yoshida, Tomoko; Nakahama, Ken-ichi; Amagasa, Teruo; Morita, Ikuo

2011-10-01

Bone regeneration therapy based on the delivery of osteogenic factors and/or cells has received a lot of attention in recent years since the discovery of pluripotent stem cells. We reported previously that the implantation of capillary networks engineered ex vivo by the use of cell-printing technology could improve blood perfusion. Here, we developed a new substrate prepared by coating glass with polyethylene glycol (PEG) to create a non-adhesive surface and subsequent photo-lithography to finely tune the adhesive property for efficient cell transfer. We examined the cell-transfer efficiency onto amniotic membrane and bone regenerative efficiency in murine calvarial bone defect. Cell transfer of KUSA-A1 cells (murine osteoblasts) to amniotic membrane was performed for 1 h using the substrates. Cell transfer using the substrate facilitated cell engraftment onto the amniotic membrane compared to that by direct cell inoculation. KUSA-A1 cells transferred onto the amniotic membrane were applied to critical-sized calvarial bone defects in mice. Micro-computed tomography (micro-CT) analysis showed rapid and effective bone formation by the cell-equipped amniotic membrane. These results indicate that the cell-printing and transfer technology used to create the cell-equipped amniotic membrane was beneficial for the cell delivery system. Our findings support the development of a biologically stable and effective bone regeneration therapy. Copyright © 2011 John Wiley & Sons, Ltd.

Long-term three-dimensional perfusion culture of human adult bone marrow mononuclear cells in bioreactors.

PubMed

Schmelzer, Eva; Finoli, Anthony; Nettleship, Ian; Gerlach, Jörg C

2015-04-01

The construction and long-term maintenance of three-dimensional in vitro bone marrow models is of great interest but still quite challenging. Here we describe the use of a multi-compartment hollow-fiber membrane based three-dimensional perfusion bioreactor for long-term culture of whole human bone marrow mononuclear cells. We also investigated bioreactors with incorporated open-porous foamed hydroxyapatite scaffolds, mimicking the in vivo bone matrix. Cells in bioreactors with and without scaffolds were cultured to 6 weeks and compared to Petri dish controls. Cells were analyzed for gene expression, surface markers by flow cytometry, metabolic activity, hematopoietic potential, viability, and attachment by immunocytochemistry. Cells in bioreactors were metabolic active during long-term culture. The percentages of hematopoietic stem cell and mature endothelial cell fractions were maintained in bioreactors. The expression of most of the analyzed genes stabilized and increased after long-term culture of 6 weeks. Compared to Petri dish culture controls, bioreactor perfusion culture improved in both the short and long-term, the colony formation unit capacity of hematopoietic progenitors. Cells attached to the ample surface area provided by hydroxyapatite scaffolds. The implementation of a hydroxyapatite scaffold did not influence colony formation capacity, percentages of cell type specific fractions, gene expression, cell viability or metabolic turnover when compared to control cells cultured in bioreactors without scaffolds. In conclusion, three-dimensional perfusion bioreactor culture enables long-term maintenance of primary human bone marrow cells, with hydroxyapatite scaffolds providing an in vivo-like scaffold for three-dimensional culture. © 2015 Wiley Periodicals, Inc.

Nanocrystallinity effects on osteoblast and osteoclast response to silicon substituted hydroxyapatite.

PubMed

Casarrubios, Laura; Matesanz, María Concepción; Sánchez-Salcedo, Sandra; Arcos, Daniel; Vallet-Regí, María; Portolés, María Teresa

2016-11-15

Silicon substituted hydroxyapatites (SiHA) are highly crystalline bioceramics treated at high temperatures (about 1200°C) which have been approved for clinical use with spinal, orthopedic, periodontal, oral and craniomaxillofacial applications. The preparation of SiHA with lower temperature methods (about 700°C) provides nanocrystalline SiHA (nano-SiHA) with enhanced bioreactivity due to higher surface area and smaller crystal size. The aim of this study has been to know the nanocrystallinity effects on the response of both osteoblasts and osteoclasts (the two main cell types involved in bone remodelling) to silicon substituted hydroxyapatite. Saos-2 osteoblasts and osteoclast-like cells (differentiated from RAW-264.7 macrophages) have been cultured on the surface of nano-SiHA and SiHA disks and different cell parameters have been evaluated: cell adhesion, proliferation, viability, intracellular content of reactive oxygen species, cell cycle phases, apoptosis, cell morphology, osteoclast-like cell differentiation and resorptive activity. This comparative in vitro study evidences that nanocrystallinity of SiHA affects the cell/biomaterial interface inducing bone cell apoptosis by loss of cell anchorage (anoikis), delaying osteoclast-like cell differentiation and decreasing the resorptive activity of this cell type. These results suggest the potential use of nano-SiHA biomaterial for preventing bone resorption in treatment of osteoporotic bone. Copyright © 2016 Elsevier Inc. All rights reserved.

Characterization of the bone marrow adipocyte niche with three-dimensional electron microscopy.

PubMed

Robles, Hero; Park, SungJae; Joens, Matthew S; Fitzpatrick, James A J; Craft, Clarissa S; Scheller, Erica L

2018-01-27

Unlike white and brown adipose tissues, the bone marrow adipocyte (BMA) exists in a microenvironment containing unique populations of hematopoietic and skeletal cells. To study this microenvironment at the sub-cellular level, we performed a three-dimensional analysis of the ultrastructure of the BMA niche with focused ion beam scanning electron microscopy (FIB-SEM). This revealed that BMAs display hallmarks of metabolically active cells including polarized lipid deposits, a dense mitochondrial network, and areas of endoplasmic reticulum. The distinct orientations of the triacylglycerol droplets suggest that fatty acids are taken up and/or released in three key areas - at the endothelial interface, into the hematopoietic milieu, and at the bone surface. Near the sinusoidal vasculature, endothelial cells send finger-like projections into the surface of the BMA which terminate near regions of lipid within the BMA cytoplasm. In some regions, perivascular cells encase the BMA with their flattened cellular projections, limiting contacts with other cells in the niche. In the hematopoietic milieu, BMAT adipocytes of the proximal tibia interact extensively with maturing cells of the myeloid/granulocyte lineage. Associations with erythroblast islands are also prominent. At the bone surface, the BMA extends organelle and lipid-rich cytoplasmic regions toward areas of active osteoblasts. This suggests that the BMA may serve to partition nutrient utilization between diverse cellular compartments, serving as an energy-rich hub of the stromal-reticular network. Lastly, though immuno-EM, we've identified a subset of bone marrow adipocytes that are innervated by the sympathetic nervous system, providing an additional mechanism for regulation of the BMA. In summary, this work reveals that the bone marrow adipocyte is a dynamic cell with substantial capacity for interactions with the diverse components of its surrounding microenvironment. These local interactions likely contribute to its unique regulation relative to peripheral adipose tissues. Copyright © 2018 Elsevier Inc. All rights reserved.

The effect of layer-by-layer chitosan-hyaluronic acid coating on graft-to-bone healing of a poly(ethylene terephthalate) artificial ligament.

PubMed

Li, Hong; Ge, Yunsheng; Zhang, Pengyun; Wu, Lingxiang; Chen, Shiyi

2012-01-01

Surface coating with an organic layer-by-layer self-assembled template of chitosan and hyaluronic acid on a poly(ethylene terephthalate) (PET) artificial ligament was designed for the promotion and enhancement of graft-to-bone healing after artificial ligament implantation in a bone tunnel. The results of in vitro culturing of MC3T3-E1 mouse osteoblastic cells supported the hypothesis that the layer-by-layer coating of chitosan and hyaluronic acid could promote the cell compatibility of grafts and could promote osteoblast proliferation. A rabbit extra-articular tendon-to-bone healing model was used to evaluate the effect of this kind of surface-modified stainless artificial ligament in vivo. The final results proved that this organic compound coating could significantly promote and enhance new bone formation at the graft-bone interface histologically and, correspondingly, the experimental group with coating had significantly higher biomechanical properties compared with controls at 8 weeks (P < 0.05).

Osteoinductive activity of insulin-functionalized cell culture surfaces obtained using diazonium chemistry

NASA Astrophysics Data System (ADS)

Mikulska, Anna; Filipowska, Joanna; Osyczka, Anna; Nowakowska, Maria; Szczubiałka, Krzysztof

2014-12-01

Polymeric surfaces suitable for cell culture (DR/Pec) were constructed from diazoresin (DR) and pectin (Pec) in a form of ultrathin films using the layer-by-layer (LbL) technique. The surfaces were functionalized with insulin using diazonium chemistry. Such functionalized surfaces were used to culture human mesenchymal stem cells (hMSCs) to assess their suitability for bone tissue engineering and regeneration. The activity of insulin immobilized on the surfaces (DR/Pec/Ins) was compared to that of insulin dissolved in the culture medium. Human MSC grown on insulin-immobilized DR/Pec surfaces displayed increased proliferation and higher osteogenic activity. The latter was determined by means of alkaline phosphatase (ALP) activity, which increases at early stages of osteoblasts differentiation. Insulin dissolved in the culture medium did not stimulate cell proliferation and its osteogenic activity was significantly lower. Addition of recombinant human bone morphogenetic protein 2 (rhBMP-2) to the culture medium further increased ALP activity in hMSCs indicating additive osteogenic action of immobilized insulin and rhBMP-2

Conception on the cell mechanisms of bone tissue loss under spase flight conditions

NASA Astrophysics Data System (ADS)

Rodionova, Natalia; Oganov, Victor; Kabitskaya, Olga

Basing on the analysis of available literature and the results of our own electron microscopic and radioautographic researches the data are presented about the morpho-functional peculiarities and succession of cellular interactions in adaptive remodeling of bone structures under normal conditions and after exposure of animals (rats, monkeys, mice) to microgravity (SLS-2, Bion-11, BionM-1). The probable cellular mechanisms of the development of osteopenia and osteoporosis are considered. Our conception on remodeling proposes the following sequence in the development of cellular interactions after decrease of the mechanical loading: a primary response of osteocytes (mechanosensory cells) to the mechanical stimulus; osteocytic remodeling (osteolysis); transmission of the mechanical signals through a system of canals and processes to functionally active osteoblasts and surface osteocytes as well as to the bone-marrow stromal cells and to those lying on bone surfaces. As a response to the mechanical stimulus (microgravity) the system of stromal cell-preosteoblast-osteoblast shows a delay in proliferation, differentiation and specific functioning of the osteogenetic cells, some of the osteoblasts undergo apoptosis. Then the osteoclastic reaction occurs (attraction of monocytes and formation of osteoclasts and bone matrix resorption in the loci of apoptosis of osteoblasts and osteocytes). The macrophagal reaction is followed by osteoblastogenesis, which appears to be a rehabilitating process. However, during prolonged absence of mechanical stimuli (microgravity, long-time immobilization) the adaptive activization of osteoblastogenesis doesn’t occur (as it is the case during the physiological remodeling of bone tissue) or it occurs to a smaller degree. The loading deficit leads to an adaptive differentiation of stromal cells to fibroblastic cells and adipocytes in these remodeling loci. These cell reactions are considered as adaptive-compensatory, but they don’t result in rehabilitation of the resorbed bone tissue. This sequence of events is considered as a mechanism of bone tissue loss which underlies the development of osteopenia and osteoporosis under the mechanical loading deficit.

Development of a biodegradable scaffold with interconnected pores by heat fusion and its application to bone tissue engineering.

PubMed

Shin, Michael; Abukawa, Harutsugi; Troulis, Maria J; Vacanti, Joseph P

2008-03-01

Tissue engineering has been proposed as an approach to alleviate the shortage of donor tissue and organs by combining cells and a biodegradable scaffold as a temporary extracellular matrix. While numerous scaffold fabrication methods have been proposed, tissue formation is typically limited to the surface of the scaffolds in bone tissue engineering applications due to early calcification on the surface. To improve tissue formation, a novel scaffold with a hierarchical interconnected pore structure on two distinct length scales has been developed. Here we present the fabrication process and the application of the scaffold to bone tissue engineering. Porous poly(lactide-co-glycolide) (PLGA) scaffolds were made by combining solvent casting/particulate leaching with heat fusion. Porcine bone marrow-derived mesenchymal stem cells (MSCs) were differentiated into osteoblasts and cultured on these scaffolds in vitro for 2, 4, and 6 weeks. Subsequently, the constructs were assessed using histology and scanning electron microscopy. The bone marrow-derived osteoblasts attached well on these scaffolds. Cells were observed throughout the scaffolds. These initial results show promise for this scaffold to aid in the regeneration of bone. (c) 2007 Wiley Periodicals, Inc.

Osteogenic potential of in situ TiO2 nanowire surfaces formed by thermal oxidation of titanium alloy substrate

NASA Astrophysics Data System (ADS)

Tan, A. W.; Ismail, R.; Chua, K. H.; Ahmad, R.; Akbar, S. A.; Pingguan-Murphy, B.

2014-11-01

Titanium dioxide (TiO2) nanowire surface structures were fabricated in situ by a thermal oxidation process, and their ability to enhance the osteogenic potential of primary osteoblasts was investigated. Human osteoblasts were isolated from nasal bone and cultured on a TiO2 nanowires coated substrate to assess its in vitro cellular interaction. Bare featureless Ti-6Al-4V substrate was used as a control surface. Initial cell adhesion, cell proliferation, cell differentiation, cell mineralization, and osteogenic related gene expression were examined on the TiO2 nanowire surfaces as compared to the control surfaces after 2 weeks of culturing. Cell adhesion and cell proliferation were assayed by field emission scanning electron microscope (FESEM) and Alamar Blue reduction assay, respectively. The nanowire surfaces promoted better cell adhesion and spreading than the control surface, as well as leading to higher cell proliferation. Our results showed that osteoblasts grown onto the TiO2 nanowire surfaces displayed significantly higher production levels of alkaline phosphatase (ALP), extracellular (ECM) mineralization and genes expression of runt-related transcription factor (Runx2), bone sialoprotein (BSP), ostoepontin (OPN) and osteocalcin (OCN) compared to the control surfaces. This suggests the potential use of such surface modification on Ti-6Al-4V substrates as a promising means to improve the osteointegration of titanium based implants.

Tetraspanin CD9 and ectonucleotidase CD73 identify an osteochondroprogenitor population with elevated osteogenic properties.

PubMed

Singh, Anju; Lester, Chantel; Drapp, Rebecca; Hu, Dorothy Z; Glimcher, Laurie H; Jones, Dallas

2015-02-01

Cell-based bone regeneration strategies offer promise for traumatic bone injuries, congenital defects, non-union fractures and other skeletal pathologies. Postnatal bone remodeling and fracture healing provide evidence that an osteochondroprogenitor cell is present in adult life that can differentiate to remodel or repair the fractured bone. However, cell-based skeletal repair in the clinic is still in its infancy, mostly due to poor characterization of progenitor cells and lack of knowledge about their in vivo behavior. Here, we took a combined approach of high-throughput screening, flow-based cell sorting and in vivo transplantation to isolate markers that identify osteochondroprogenitor cells. We show that the presence of tetraspanin CD9 enriches for osteochondroprogenitors within CD105(+) mesenchymal cells and that these cells readily form bone upon transplantation. In addition, we have used Thy1.2 and the ectonucleotidase CD73 to identify subsets within the CD9(+) population that lead to endochondral or intramembranous-like bone formation. Utilization of this unique cell surface phenotype to enrich for osteochondroprogenitor cells will allow for further characterization of the molecular mechanisms that regulate their osteogenic properties. © 2015. Published by The Company of Biologists Ltd.

CD56-positive small round cell tumor: osseous plasmacytoma manifested in osteolytic tumors of the iliac bone and femora.

PubMed

Kouno, Tsutomu; Watanabe, Takashi; Umeda, Toru; Beppu, Yasuo; Kojima, Rie; Sungwon, Kim; Kobayashi, Yukio; Tobinai, Kensei; Hasegawa, Tadashi; Matsuno, Yoshihiro

2005-02-01

Monoclonal gammopathy of undetermined significance does not overexpress cluster of differentiation (CD) 56, but plasma cell myeloma frequently overexpressed it. However, plasma cell leukemia and extramedullary plasmacytoma usually down-regulate CD56 expression. Plasmacytoma, especially 'solitary plasmacytoma of bone', is difficult to diagnose as plasma cell neoplasm, because it occasionally appears similar to other bone tumors, both clinically and pathologically, and is rarely accompanied by monoclonal protein in the serum or urine. The present case was a patient with an osteolytic 'small round cell tumor' of the iliac bone, which also invaded the femora. An immunohistopathological finding of CD56 expression played a key role in making a diagnosis. The definitive diagnosis of plasmacytoma was made based on the electron microscopic findings. The plasma cells which infiltrated her sternum showed the same restriction to kappa light chain expression in their cytoplasms as that of the iliac bone tumor cells, but did not express CD56. Locally infiltrating osteolytic bone tumors should be examined for surface immunoglobulin light chains as well as CD56 expression when plasmacytoma is suspected.

The effects of hierarchical micro/nanosurfaces decorated with TiO2 nanotubes on the bioactivity of titanium implants in vitro and in vivo

PubMed Central

Ding, Xianglong; Zhou, Lei; Wang, Jingxu; Zhao, Qingxia; Lin, Xi; Gao, Yan; Li, Shaobing; Wu, Jingyi; Rong, Mingdeng; Guo, Zehong; Lai, Chunhua; Lu, Haibin; Jia, Fang

2015-01-01

In the present work, a hierarchical hybrid micro/nanostructured titanium surface was obtained by sandblasting with large grit and acid etching (SLA), and nanotubes of different diameters (30 nm, 50 nm, and 80 nm) were superimposed by anodization. The effect of each SLA-treated surface decorated with nanotubes (SLA + 30 nm, SLA + 50 nm, and SLA + 80 nm) on osteogenesis was studied in vitro and in vivo. The human MG63 osteosarcoma cell line was used for cytocompatibility evaluation, which showed that cell adhesion and proliferation were dramatically enhanced on SLA + 30 nm. In comparison with cells grown on the other tested surfaces, those grown on SLA + 80 nm showed an enhanced expression of osteogenesis-related genes. Cell spread was also enhanced on SLA + 80 nm. A canine model was used for in vivo evaluation of bone bonding. Histological examination demonstrated that new bone was formed more rapidly on SLA-treated surfaces with nanotubes (especially SLA + 80 nm) than on those without nanotubes. All of these results indicate that SLA + 80 nm is favorable for promoting the activity of osteoblasts and early bone bonding. PMID:26635472

Induction of experimental bone metastasis in mice by transfection of integrin alpha 4 beta 1 into tumor cells.

PubMed Central

Matsuura, N.; Puzon-McLaughlin, W.; Irie, A.; Morikawa, Y.; Kakudo, K.; Takada, Y.

1996-01-01

Cell adhesion receptors (eg, integrins and CD44) play an important role in invasion and metastasis during tumor progression. The increase in integrin alpha 4 beta 1 expression on primary melanomas has been reported to significantly correlate with the development of metastases. alpha 4 beta 1 is a cell surface heterodimer that mediates cell-cell and cell-extracellular matrix interactions through adhesion to vascular cell adhesion molecule (VCAM)-1 and to the IIICS region of fibronectin. To test the effects of alpha 4 beta 1 expression on tumor cell metastasis, Chinese hamster ovary cells were transfected with human alpha 4 cDNA. Whereas alpha 4-negative Chinese hamster ovary cells developed only pulmonary metastasis, alpha 4-positive Chinese hamster ovary cells developed bone and pulmonary metastasis in 3 to 4 weeks when injected intravenously into nude mice. Bone metastasis was inhibited by antibody against alpha 4 or VCAM-1. Expression of alpha 3 beta 1, alpha 6 beta 1, or alpha V beta 1 did not induce bone metastasis. Expression of alpha 4 beta 1 also induced bone metastasis in K562 human erythroleukemia cells injected into SCID mice. These results demonstrate that alpha 4 beta 1 can induce tumor cell trafficking to bone, probably via interaction with VCAM-1 that is constitutively expressed on bone marrow stromal cells. Images Figure 1 Figure 3 PMID:8546226

Connective Tissue Growth Factor reporter mice label a subpopulation of mesenchymal progenitor cells that reside in the trabecular bone region.

PubMed

Wang, Wen; Strecker, Sara; Liu, Yaling; Wang, Liping; Assanah, Fayekah; Smith, Spenser; Maye, Peter

2015-02-01

Few gene markers selectively identify mesenchymal progenitor cells inside the bone marrow. We have investigated a cell population located in the mouse bone marrow labeled by Connective Tissue Growth Factor reporter expression (CTGF-EGFP). Bone marrow flushed from CTGF reporter mice yielded an EGFP+ stromal cell population. Interestingly, the percentage of stromal cells retaining CTGF reporter expression decreased with age in vivo and was half the frequency in females compared to males. In culture, CTGF reporter expression and endogenous CTGF expression marked the same cell types as those labeled using Twist2-Cre and Osterix-Cre fate mapping approaches, which previously had been shown to identify mesenchymal progenitors in vitro. Consistent with this past work, sorted CTGF+ cells displayed an ability to differentiate into osteoblasts, chondrocytes, and adipocytes in vitro and into osteoblast, adipocyte, and stromal cell lineages after transplantation into a parietal bone defect. In vivo examination of CTGF reporter expression in bone tissue sections revealed that it marked cells highly localized to the trabecular bone region and was not expressed in the perichondrium or periosteum. Mesenchymal cells retaining high CTGF reporter expression were adjacent to, but distinct from mature osteoblasts lining bone surfaces and endothelial cells forming the vascular sinuses. Comparison of CTGF and Osterix reporter expression in bone tissue sections indicated an inverse correlation between the strength of CTGF expression and osteoblast maturation. Down-regulation of CTGF reporter expression also occurred during in vitro osteogenic differentiation. Collectively, our studies indicate that CTGF reporter mice selectively identify a subpopulation of bone marrow mesenchymal progenitor cells that reside in the trabecular bone region. Copyright © 2014 Elsevier Inc. All rights reserved.

Connective Tissue Growth Factor Reporter Mice Label a Subpopulation of Mesenchymal Progenitor Cells that Reside in the Trabecular Bone Region

PubMed Central

Wang, Wen; Strecker, Sara; Liu, Yaling; Wang, Liping; Assanah, Fayekah; Smith, Spenser; Maye, Peter

2014-01-01

Few gene markers selectively identify mesenchymal progenitor cells inside the bone marrow. We have investigated a cell population located in the mouse bone marrow labeled by Connective Tissue Growth Factor reporter expression (CTGF-EGFP). Bone marrow flushed from CTGF reporter mice yielded an EGFP+ stromal cell population. Interestingly, the percentage of stromal cells retaining CTGF reporter expression decreased with age in vivo and was half the frequency in females compared to males. In culture, CTGF reporter expression and endogenous CTGF expression marked the same cell types as those labeled using Twist2-Cre and Osterix-Cre fate mapping approaches, which previously has been shown to identify mesenchymal progenitors in vitro. Consistent with this past work, sorted CTGF+ cells displayed an ability to differentiate into osteoblasts, chondrocytes, and adipocytes in vitro and into osteoblast, adipocyte, and stromal cell lineages after transplantation into a parietal bone defect. In vivo examination of CTGF reporter expression in bone tissue sections revealed it marked cells highly localized to the trabecular bone region and was not expressed in the perichondrium or periosteum. Mesenchymal cells retaining high CTGF reporter expression were adjacent to, but distinct from mature osteoblasts lining bone surfaces and endothelial cells forming the vascular sinuses. Comparison of CTGF and Osterix reporter expression in bone tissue sections indicated an inverse correlation between the strength of CTGF expression and osteoblast maturation. Down-regulation of CTGF reporter expression also occurred during in vitro osteogenic differentiation. Collectively, our studies indicate that CTGF reporter mice selectively identify a subpopulation of bone marrow mesenchymal progenitor cells that reside in the trabecular bone region. PMID:25464947

Polyurethane foam/nano hydroxyapatite composite as a suitable scaffold for bone tissue regeneration.

PubMed

Meskinfam, M; Bertoldi, S; Albanese, N; Cerri, A; Tanzi, M C; Imani, R; Baheiraei, N; Farokhi, M; Farè, S

2018-01-01

In bone tissue regeneration, the use of biomineralized scaffolds to create the 3D porous structure needed for well-fitting with defect size and appropriate cell interactions, is a promising alternative to autologous and heterologous bone grafts. Biomineralized polyurethane (PU) foams are here investigated as scaffold for bone tissue regeneration. Biomineralization of the foams was carried out by activation of PU surface by a two steps procedure performed for different times (1 to 4 weeks). Scaffolds were investigated for morphological, chemico-physical and mechanical properties, as well as for in vitro interaction with rat Bone Marrow Mesenchymal Stem Cells (BMSCs). Untreated and biomineralized PU samples showed a homogenous morphology and regular pore size (average Ø=407μm). Phase and structure of formed calcium phosphates (CaPs) layer onto the PU foam were analyzed by Fourier Transform Infrared spectroscopy and X-ray diffraction, proving the formation of bone-like nano hydroxyapatite. Biomineralization caused a significant increase of mechanical properties of treated foams compared to untreated ones. Biomineralization also affected the PU scaffold cytocompatibility providing a more appropriate surface for cell attachment and proliferation. Considering the obtained results, the proposed scaffold can be considered suitable for bone tissue regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

Additively Manufactured and Surface Biofunctionalized Porous Nitinol.

PubMed

Gorgin Karaji, Z; Speirs, M; Dadbakhsh, S; Kruth, J-P; Weinans, H; Zadpoor, A A; Amin Yavari, S

2017-01-18

Enhanced bone tissue regeneration and improved osseointegration are among the most important goals in design of multifunctional orthopedic biomaterials. In this study, we used additive manufacturing (selective laser melting) to develop multifunctional porous nitinol that combines superelasticity with a rationally designed microarchitecture and biofunctionalized surface. The rational design based on triply periodic minimal surfaces aimed to properly adjust the pore size, increase the surface area (thereby amplifying the effects of surface biofunctionalization), and resemble the curvature characteristics of trabecular bone. The surface of additively manufactured (AM) porous nitinol was biofunctionalized using polydopamine-immobilized rhBMP2 for better control of the release kinetics. The actual morphological properties of porous nitinol measured by microcomputed tomography (e.g., open/close porosity, and surface area) closely matched the design values. The superelasticity originated from the austenite phase formed in the nitinol porous structure at room temperature. Polydopamine and rhBMP2 signature peaks were confirmed by X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy tests. The release of rhBMP2 continued until 28 days. The early time and long-term release profiles were found to be adjustable independent of each other. In vitro cell culture showed improved cell attachment, cell proliferation, cell morphology (spreading, spindle-like shape), and cell coverage as well as elevated levels of ALP activity and increased calcium content for biofunctionalized surfaces as compared to as-manufactured specimens. The demonstrated functionalities of porous nitinol could be used as a basis for deployable orthopedic implants with rationally designed microarchitectures that maximize bone tissue regeneration performance by release of biomolecules with adjustable and well-controlled release profiles.

Adsorption of enamel matrix proteins to a bovine-derived bone grafting material and its regulation of cell adhesion, proliferation, and differentiation.

PubMed

Miron, Richard J; Bosshardt, Dieter D; Hedbom, Erik; Zhang, Yufeng; Haenni, Beat; Buser, Daniel; Sculean, Anton

2012-07-01

The use of various combinations of enamel matrix derivative (EMD) and grafting materials has been shown to promote periodontal wound healing/regeneration. However, the downstream cellular behavior of periodontal ligament (PDL) cells and osteoblasts has not yet been studied. Furthermore, it is unknown to what extent the bleeding during regenerative surgery may influence the adsorption of exogenous proteins to the surface of bone grafting materials and the subsequent cellular behavior. In the present study, the aim is to test EMD adsorption to the surface of natural bone mineral (NBM) particles in the presence of blood and determine the effect of EMD coating to NBM particles on downstream cellular pathways, such as adhesion, proliferation, and differentiation of primary human osteoblasts and PDL cells. NBM particles were precoated in various settings with EMD or human blood and analyzed for protein adsorption patterns via fluorescent imaging and high-resolution immunocytochemistry with an anti-EMD antibody. Cell attachment and cell proliferation were quantified using fluorescent double-stranded DNA-binding dye. Cell differentiation was analyzed using real-time polymerase chain reaction for genes encoding runt-related transcription factor 2, alkaline phosphatase (ALP), osteocalcin (OC), and collagen1α1 (COL1A1), and mineralization was assessed using red dye staining. Analysis of cell attachment and cell proliferation revealed significantly higher osteoblast and PDL cell attachment on EMD-coated surfaces when compared with control and blood-coated surfaces. EMD also stimulated release of growth factors and cytokines, including bone morphogenetic protein 2 and transforming growth factor β1. Moreover, there were significantly higher mRNA levels of osteoblast differentiation markers, including COL1A1, ALP, and OC, in osteoblasts and PDL cells cultured on EMD-coated NBM particles. The present results suggest that 1) EMD enhances osteoblast and PDL cell attachment, proliferation, and differentiation on NBM particles, and 2) blood contamination of the grafting material before mixing with EMD may inhibit EMD adsorption.

Tolerance to MHC class II disparate allografts through genetic modification of bone marrow

PubMed Central

Jindra, Peter T.; Tripathi, Sudipta; Tian, Chaorui; Iacomini, John; Bagley, Jessamyn

2012-01-01

Induction of molecular chimerism through genetic modification of bone marrow is a powerful tool for the induction of tolerance. Here we demonstrate for the first time that expression of an allogeneic MHC class II gene in autologous bone marrow cells, resulting in a state of molecular chimerism, induces tolerance to MHC class II mismatched skin grafts, a stringent test of transplant tolerance. Reconstitution of recipients with syngeneic bone marrow transduced with retrovirus encoding H-2I-Ab (I-Ab) resulted the long-term expression of the retroviral gene product on the surface of MHC class II-expressing bone marrow derived cell types. Mechanistically, tolerance was maintained by the presence of regulatory T cells, which prevented proliferation and cytokine production by alloreactive host T cells. Thus, the introduction of MHC class II genes into bone marrow derived cells through genetic engineering results in tolerance. These results have the potential to extend the clinical applicability of molecular chimerism for tolerance induction. PMID:22833118

From isolation to implantation: a concise review of mesenchymal stem cell therapy in bone fracture repair

PubMed Central

2014-01-01

Compromised bone-regenerating capability following a long bone fracture is often the result of reduced host bone marrow (BM) progenitor cell numbers and efficacy. Without surgical intervention, these malunions result in mobility restrictions, deformities, and disability. The clinical application of BM-derived mesenchymal stem cells (MSCs) is a feasible, minimally invasive therapeutic option to treat non-union fractures. This review focuses on novel, newly identified cell surface markers in both the mouse and human enabling the isolation and purification of osteogenic progenitor cells as well as their direct and indirect contributions to fracture repair upon administration. Furthermore, clinical success to date is summarized with commentary on autologous versus allogeneic cell sources and the methodology of cell administration. Given our clinical success to date in combination with recent advances in the identification, isolation, and mechanism of action of MSCs, there is a significant opportunity to develop improved technologies for defining therapeutic MSCs and potential to critically inform future clinical strategies for MSC-based bone regeneration. PMID:25099622

Analysis of the surface expression of c-kit and occurrence of the c-kit Asp816Val activating mutation in T cells, B cells, and myelomonocytic cells in patients with mastocytosis.

PubMed

Akin, C; Kirshenbaum, A S; Semere, T; Worobec, A S; Scott, L M; Metcalfe, D D

2000-02-01

The Asp816Val c-kit activating mutation is detectable in the peripheral blood cells of some patients with mastocytosis and in lesional skin biopsies obtained from adult patients with urticaria pigmentosa. These observations led to the conclusion that this mutation is present in mast cells and mast cell precursors that express c-kit. However, the distribution of the Asp816Val mutation among hematopoietic lineages is unknown. To determine the distribution of the Asp816Val mutation among hematopoietic lineages and to explore its relationship to clinical disease, we examined cells bearing differentiation markers for myelomonocytic cells as well as T and B lymphocytes, in both peripheral blood and bone marrow obtained from patients with mastocytosis. The presence of Asp816Val c-kit mutation in cells magnetically sorted from peripheral blood or bone marrow according to surface differentiation markers was studied by reverse transcriptase polymerase chain reaction (RT-PCR) restriction fragment length polymorphism (RFLP) analysis. The surface expression of c-kit was determined by flow cytometry. The mutation was detectable by RT-PCR in at least one cell lineage in the bone marrow in 7 of 7 patients examined and in the peripheral blood of 11 of 11 adult patients with urticaria pigmentosa and indolent disease. The mutation was identified most frequently in B cells and myeloid cells. Flow cytometric analysis demonstrated that the differentiated cells expressing mutated c-kit were negative for surface KIT. These results are consistent with the conclusion that the c-kit Asp816Val mutation occurs in an early progenitor cell and is carried by myelomonocytic cells, T cells, and B cells in addition to mast cells. However, unlike mast cells, these myelomonocytic cells, T cells, and B cells do not concomitantly express surface c-kit and thus may be less susceptible to the effects of this mutation.

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.

A Spontaneous 3D Bone-On-a-Chip for Bone Metastasis Study of Breast Cancer Cells.

PubMed

Hao, Sijie; Ha, Laura; Cheng, Gong; Wan, Yuan; Xia, Yiqiu; Sosnoski, Donna M; Mastro, Andrea M; Zheng, Si-Yang

2018-03-01

Bone metastasis occurs at ≈70% frequency in metastatic breast cancer. The mechanisms used by tumors to hijack the skeleton, promote bone metastases, and confer therapeutic resistance are poorly understood. This has led to the development of various bone models to investigate the interactions between cancer cells and host bone marrow cells and related physiological changes. However, it is challenging to perform bone studies due to the difficulty in periodic sampling. Herein, a bone-on-a-chip (BC) is reported for spontaneous growth of a 3D, mineralized, collagenous bone tissue. Mature osteoblastic tissue of up to 85 µm thickness containing heavily mineralized collagen fibers naturally formed in 720 h without the aid of differentiation agents. Moreover, co-culture of metastatic breast cancer cells is examined with osteoblastic tissues. The new bone-on-a-chip design not only increases experimental throughput by miniaturization, but also maximizes the chances of cancer cell interaction with bone matrix of a concentrated surface area and facilitates easy, frequent observation. As a result, unique hallmarks of breast cancer bone colonization, previously confirmed only in vivo, are observed. The spontaneous 3D BC keeps the promise as a physiologically relevant model for the in vitro study of breast cancer bone metastasis. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

Growth promoting in vitro effect of synthetic cyclic RGD-peptides on human osteoblast-like cells attached to cancellous bone.

PubMed

Magdolen, Ursula; Auernheimer, Jörg; Dahmen, Claudia; Schauwecker, Johannes; Gollwitzer, Hans; Tübel, Jutta; Gradinger, Reiner; Kessler, Horst; Schmitt, Manfred; Diehl, Peter

2006-06-01

In tissue engineering, the application of biofunctional compounds on biomaterials such as integrin binding RGD-peptides has gained growing interest. Anchorage-dependent cells like osteoblasts bind to these peptides thus ameliorating the integration of a synthetic implant. In case sterilized bone grafts are used as substitutes for reconstruction of bone defects, the ingrowth of the implanted bone is often disturbed because of severe pretreatment such as irradiation or autoclaving, impairing the biological and mechanical properties of the bone. We report for the first time on the in vitro coating of the surface of freshly resected, cleaned bone discs with synthetic, cyclic RGD-peptides. For this approach, two different RGD-peptides were used, one containing two phosphonate anchors, the other peptide four of these binding moieties to allow efficient association of these reactive RGD-peptides to the inorganic bone matrix. Human osteoblast-like cells were cultured on RGD-coated bone discs and the adherence and growth of the cells were analyzed. Coating of bone discs with RGD-peptides did not improve the adhesion rate of osteoblast-like cells to the discs but significantly (up to 40%) accelerated growth of these cells within 8 days after attachment. This effect points to pretreatment of bone implants, especially at the critical interface area between the implanted bone and the non-resected residual bone structure, before re-implantation in order to stimulate and enhance osteointegration of a bone implant.

Enhanced cell adhesion on bioinert ceramics mediated by the osteogenic cell membrane enzyme alkaline phosphatase.

PubMed

Aminian, Alieh; Shirzadi, Bahareh; Azizi, Zahra; Maedler, Kathrin; Volkmann, Eike; Hildebrand, Nils; Maas, Michael; Treccani, Laura; Rezwan, Kurosch

2016-12-01

Functional bone and dental implant materials are required to guide cell response, offering cues that provide specific instructions to cells at the implant/tissue interface while maintaining full biocompatibility as well as the desired structural requirements and functions. In this work we investigate the influence of covalently immobilized alkaline phosphatase (ALP), an enzyme involved in bone mineralization, on the first contact and initial cell adhesion. To this end, ALP is covalently immobilized by carbodiimide-mediated chemoligation on two highly bioinert ceramics, alpha-alumina (Al2O3) and yttria-stabilized zirconia (Y-TZP) that are well-established for load-bearing applications. The physicochemical surface properties are evaluated by profilometry, zeta potential and water contact angle measurements. The initial cell adhesion of human osteoblasts (HOBs), human osteoblast-like cells (MG-63) and mesenchymal stromal cells (hMSCs) was investigated. Cell adhesion was assessed at serum free condition via quantification of percentage of adherent cells, adhesion area and staining of the focal adhesion protein vinculin. Our findings show that after ALP immobilization, the Al2O3 and Y-TZP surfaces gained a negative charge and their hydrophilicity was increased. In the presence of surface-immobilized ALP, a higher cell adhesion, more pronounced cell spreading and a higher number of focal contact points were found. Thereby, this work gives evidence that surface functionalization with ALP can be utilized to modify inert materials for biological conversion and faster bone regeneration on inert and potentially load-bearing implant materials. Copyright © 2016 Elsevier B.V. All rights reserved.

3D silicon doped hydroxyapatite scaffolds decorated with Elastin-like Recombinamers for bone regenerative medicine.

PubMed

Vila, Mercedes; García, Ana; Girotti, Alessandra; Alonso, Matilde; Rodríguez-Cabello, Jose Carlos; González-Vázquez, Arlyng; Planell, Josep A; Engel, Elisabeth; Buján, Julia; García-Honduvilla, Natalio; Vallet-Regí, María

2016-11-01

The current study reports on the manufacturing by rapid prototyping technique of three-dimensional (3D) scaffolds based on silicon substituted hydroxyapatite with Elastin-like Recombinamers (ELRs) functionalized surfaces. Silicon doped hydroxyapatite (Si-HA), with Ca 10 (PO 4 ) 5.7 (SiO 4 ) 0.3 (OH) 1.7 h 0.3 nominal formula, was surface functionalized with two different types of polymers designed by genetic engineering: ELR-RGD that contain cell attachment specific sequences and ELR-SN A 15/RGD with both hydroxyapatite and cells domains that interact with the inorganic phase and with the cells, respectively. These hybrid materials were subjected to in vitro assays in order to clarify if the ELRs coating improved the well-known biocompatible and bone regeneration properties of calcium phosphates materials. The in vitro tests showed that there was a total and homogeneous colonization of the 3D scaffolds by Bone marrow Mesenchymal Stromal Cells (BMSCs). In addition, the BMSCs were viable and able to proliferate and differentiate into osteoblasts. Bone tissue engineering is an area of increasing interest because its main applications are directly related to the rising life expectancy of the population, which promotes higher rates of several bone pathologies, so innovative strategies are needed for bone tissue regeneration therapies. Here we use the rapid prototyping technology to allow moulding ceramic 3D scaffolds and we use different bio-polymers for the functionalization of their surfaces in order to enhance the biological response. Combining the ceramic material (silicon doped hydroxyapatite, Si-HA) and the Elastin like Recombinamers (ELRs) polymers with the presence of the integrin-mediate adhesion domain alone or in combination with SNA15 peptide that possess high affinity for hydroxyapatite, provided an improved Bone marrow Mesenchymal Stromal Cells (BMSCs) differentiation into osteoblastic linkage. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Characterizing the inorganic/organic interface in cancer bone metastasis

NASA Astrophysics Data System (ADS)

Wu, Fei

Bone metastasis frequently occurs in patients with advanced breast cancer and remains a major source of mortality. At the molecular level, bone is a nanocomposite composed of inorganic bone mineral deposited within an organic extracellular matrix (ECM). Although the exact mechanisms of bone metastasis remain unclear, the nanoscale materials properties of bone mineral have been implicated in this process. Bone apatite is closely related to synthetic hydroxyapatite (HAP, Ca10(PO4)6(OH)2) in terms of structural and mechanical properties. Additionally, although the primary protein content of bone is collagen I, the glycoprotein fibronectin (Fn) is essential in maintaining the overall integrity of the bone matrix. Importantly, in vivo, neither breast cancer cells nor normal bone cells interact directly with the bone mineral but rather with the protein film adsorbed onto the mineral surface. Therefore, we hypothesized that breast cancer cell functions were regulated by differential fibronectin adsorption onto hydroxyapatite, which led to pathological remodeling of the bone matrix and sustained bone metastasis. Three model systems containing HAP and Fn were developed for this thesis. In model system I, a library of synthetic HAP nanoparticles were utilized to investigate the effect of mineral size, shape, and crystallinity on Fn conformation, using Forster resonance energy transfer (FRET) spectroscopy. In model system II, Fn-functionalized large geologic HAP crystals were used instead of HAP nanoparticles to avoid cellular uptake when investigating subsequent cell functions. Overall our FRET analysis (models I and II) revealed that Fn conformation depended on size, surface chemistry, and roughness of underlying HAP. When breast cancer cells were seeded on the Fn-coated HAP crystal facets (model II), our data indicated high secretion levels of proangiogenic and proinflammatory factors associated with the presence of unfolded Fn conformations, likely caused by differential engagement of cell receptors integrins with the underlying Fn. Finally, in model system III, Fn fibrillar structures (mimicking the bone matrix) were fabricated and characterized in presence of HAP nanoparticles, suggesting that the presence of microcalcifications found in tumorous/inflammed tissues affects both the structural and mechanical properties of the surrounding ECM. Collectively, our study of cellular behavior regulated by mineral/ECM interactions provides insights into the pathogenesis of breast cancer bone metastasis as well as other HAP-related inflammation.

Surface micromorphology of cross-linked tetrafunctional polylactide scaffolds inducing vessel growth and bone formation.

PubMed

Kuznetsova, D; Ageykin, A; Koroleva, A; Deiwick, A; Shpichka, A; Solovieva, A; Kostjuk, S; Meleshina, A; Rodimova, S; Akovanceva, A; Butnaru, D; Frolova, A; Zagaynova, E; Chichkov, B; Bagratashvili, V; Timashev, P

2017-04-28

In the presented study, we have developed a synthetic strategy allowing a gradual variation of a polylactide arms' length, which later influences the micromorphology of the scaffold surface, formed by a two-photon polymerization technique. It has been demonstrated that the highest number of cells is present on the scaffolds with the roughest surface made of the polylactide with longer arms (PLA760), and osteogenic differentiation of mesenchymal stem cells is most pronounced on such scaffolds. According to the results of biological testing, the PLA760 scaffolds were implanted into a created cranial defect in a mouse for an in vivo assessment of the bone tissue formation. The in vivo experiments have shown that, by week 10, deposition of calcium phosphate particles occurs in the scaffold at the defect site, as well as, the formation of a new bone and ingrowth of blood vessels from the surrounding tissues. These results demonstrate that the cross-linked microstructured tetrafunctional polylactide scaffolds are promising microstructures for bone regeneration in tissue engineering.

Bone marrow adipocytes: a neglected target tissue for growth hormone.

PubMed

Gevers, Evelien F; Loveridge, Nigel; Robinson, Iain C A F

2002-10-01

Bone marrow (BM) contains numerous adipocytes. These share a common precursor with osteoblasts and chondrocytes, but their function is unknown. It is unclear what regulates the differentiation of these three different cell types, though their subsequent metabolic activity is under hormonal regulation. GH and estrogen stimulate bone growth and mineralization, by direct effects on chondrocytes and osteoblasts. GH also stimulates lipolysis in subcutaneous and visceral adipocytes. However, adipocytes in BM have largely been ignored as potential targets for GH or estrogen action. We have addressed this by measuring BM adipocyte number, perimeter and area as well as bone area and osteoblast activity in GH-deficient dwarf (dw/dw), normal, or ovariectomized (Ovx) rats, with or without GH, IGF-1, PTH, or estrogen treatment or high fat feeding. Marrow adipocyte numbers were increased 5-fold (P < 0.001) in dw/dw rats, and cell size was also increased by 20%. These values returned toward normal in dw/dw rats given GH but not when given IGF-1. Cancellous bone area and osteoblast number were significantly (P < 0.005) lower in dw/dw rats, though alkaline phosphatase (ALP) activity in individual osteoblasts was unchanged. GH treatment increased % osteoblast covered bone surface without affecting individual cell ALP activity. Ovariectomy in normal or dw/dw rats had no affect on marrow adipocyte number nor size, although estrogen treatment in ovariectomized (Ovx) normal rats did increase adipocyte number. Ovx decreased tibial cancellous bone area in normal rats (64%; P < 0.05) and decreased osteoblast ALP-activity (P < 0.01) but did not affect the percentage of osteoblast-covered bone surface. Estrogen replacement reversed these changes. While treatment with PTH by continuous sc infusion decreased cancellous bone (P < 0.05) and high fat feeding increased the size of BM adipocytes (P < 0.01), they did not affect BM adipocyte number. These results suggest that GH has a specific action on BM adipocytes that is not simply due to altered bone or fat metabolism. We conclude that the marrow adipocyte lineage is an important and specific target for GH action. The inverse relationship between adipocyte number and osteoblast covered bone surface, together with the well-known effects of GH on epiphysial chondrocytes leads us to propose that GH plays two important roles on cells of all three lineages. During differentiation, it regulates the numbers of each cell type that are maintained from the common precursor lineage. Subsequently it has cell-specific effects on the metabolic activities of the differentiated cells. In the case of marrow adipocytes, GH-dependent lipolysis could provide an important hormonally regulated local high energy source in bone.

Computational mechanobiology to study the effect of surface geometry on peri-implant tissue differentiation.

PubMed

Andreykiv, A; van Keulen, F; Prendergast, P J

2008-10-01

The geometry of an implant surface to best promote osseointegration has been the subject of several experimental studies, with porous beads and woven mesh surfaces being among the options available. Furthermore, it is unlikely that one surface geometry is optimal for all loading conditions. In this paper, a computational method is used to simulate tissue differentiation and osseointegration on a smooth surface, a surface covered with sintered beads (this simulated the experiment (Simmons, C., and Pilliar, R., 2000, Biomechanical Study of Early Tissue Formation Around Bone-Interface Implants: The Effects of Implant Surface Geometry," Bone Engineering, J. E. Davies, ed., Emsquared, Chap. A, pp. 369-379) and established that the method gives realistic results) and a surface covered by porous tantalum. The computational method assumes differentiation of mesenchymal stem cells in response to fluid flow and shear strain and models cell migration and proliferation as continuum processes. The results of the simulation show a higher rate of bone ingrowth into the surfaces with porous coatings as compared with the smooth surface. It is also shown that a thicker interface does not increase the chance of fixation failure.

In vitro biological evaluation of beta-TCP/HDPE--A novel orthopedic composite: a survey using human osteoblast and fibroblast bone cells.

PubMed

Homaeigohar, S Sh; Shokrgozar, M A; Khavandi, A; Sadi, A Yari

2008-02-01

Beta-tricalcium phosphate reinforced high density polyethylene (beta-TCP/HDPE) was prepared to simulate bone composition and to study its capacity to act as bone tissue. This material was produced by replacing the mineral component and collagen soft tissue of the bone with beta-TCP and HDPE, respectively. The biocompatibility of the composite samples with different volume fractions of TCP (20, 30 and 40 vol %) was examined in vitro using two osteoblast cell lines G-292 and Saos-2, and also a type of fibroblast cell isolated from bone tissue, namely human bone fibroblast (HBF) by proliferation, and cell adhesion assays. Cell-material interaction with the surface of the composite samples was examined by scanning electron microscopy (SEM). The effect of beta-TCP/HDPE on the behavior of osteoblast and fibroblast cells was compared with those of composite and negative control samples; polyethylene (PE) and tissue culture polystyrene (TPS), respectively. In general, the results showed that the composite samples containing beta-TCP as reinforcement supported a higher rate of proliferation by various bone cells after 3, 7, and 14 days of incubation compared to the composite control sample. Furthermore, more osteoblast cells were attached to the surface of the composite samples when compared to the composite control samples after the above incubation periods (p < 0.05), while in the case of HBF an equal or even higher number of cells adhered to PE was observed. The number of adhered osteoblast cells was almost equal and in some days even higher than the number of adhered cells on negative control sample, while in the case of fibroblast this difference was significantly higher than TPS (p < 0.05). Adhered cells presented a normal morphology by SEM and many of the cells were observed to be undergoing cell division. These findings indicate that beta-TCP/HDPE composites are biocompatible, nontoxic, and act to stimulate proliferation and adhesion of the cells, whether osteoblast or fibroblast. (c) 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008.

[Bone remodeling and modeling/mini-modeling.

PubMed

Hasegawa, Tomoka; Amizuka, Norio

Modeling, adapting structures to loading by changing bone size and shapes, often takes place in bone of the fetal and developmental stages, while bone remodeling-replacement of old bone into new bone-is predominant in the adult stage. Modeling can be divided into macro-modeling(macroscopic modeling)and mini-modeling(microscopic modeling). In the cellular process of mini-modeling, unlike bone remodeling, bone lining cells, i.e., resting flattened osteoblasts covering bone surfaces will become active form of osteoblasts, and then, deposit new bone onto the old bone without mediating osteoclastic bone resorption. Among the drugs for osteoporotic treatment, eldecalcitol(a vitamin D3 analog)and teriparatide(human PTH[1-34])could show mini-modeling based bone formation. Histologically, mature, active form of osteoblasts are localized on the new bone induced by mini-modeling, however, only a few cell layer of preosteoblasts are formed over the newly-formed bone, and accordingly, few osteoclasts are present in the region of mini-modeling. In this review, histological characteristics of bone remodeling and modeling including mini-modeling will be introduced.

A Dual-Action Armed Replicating Adenovirus for the Treatment of Osteoblastic Bone Metastases of Prostate Cancer

DTIC Science & Technology

2007-03-01

ligand), a membrane-bound cytokine expressed in osteoblasts/stromal cells, which binds to RANK, a cell- surface protein present on osteoclast...cocultures were incubated for 10 days and then the culture medium was employed in an ELISA to detect the osteoclast marker , tartrate- resistant acid...bone marrow stromal cells and osteoclast precursor cells. Ten days post-infection, expression of the osteoclast marker enzyme TRAP was determined

The Role of Dextran Coatings on the Cytotoxicity Properties of Ceria Nanoparticles Toward Bone Cancer Cells

NASA Astrophysics Data System (ADS)

Yazici, Hilal; Alpaslan, Ece; Webster, Thomas J.

2015-04-01

Cerium oxide nanoparticles have demonstrated great potential as antioxidant and radioprotective agents for nanomedicine applications especially for cancer therapy. The surface chemistry of nanoparticles is an important property that has a significant effect on their performance in biological applications including cancer diagnosis, cancer treatment, and bacterial infection. Recently, various nanosized cerium oxide particles with different types of polymer coatings have been developed to improve aqueous solubility and allow for surface functionalization for distinct applications. In this study, the role of ceria nanoparticles coated with dextran on the cytotoxicity properties of bone cancer cells was shown. Specifically, 0.1 M and 0.01 M dextran-coated, <5-nm ceria nanoparticles, were synthesized. The cytotoxicity of 0.1 M and 0.01 M dextran-coated ceria nanoparticles was evaluated against osteosarcoma cells. A change in cell viability was observed when treating osteosarcoma cells with 0.1 M dextran-coated ceria nanoparticles in the 250 -1000 μg/mL concentration range. In contrast, minimal toxicity to bone cancer cells was observed for the 0.01 M dextran coating after 3 days compared with the 0.1 M dextran coating. These results indicated that surface dextran functionalization had a positive impact on the cytotoxicity of cerium oxide nanoparticles against osteosarcoma cells.

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.

Upregulation of BMSCs Osteogenesis by Positively-Charged Tertiary Amines on Polymeric Implants via Charge/iNOS Signaling Pathway

PubMed Central

Zhang, Wei; Liu, Na; Shi, Haigang; Liu, Jun; Shi, Lianxin; Zhang, Bo; Wang, Huaiyu; Ji, Junhui; Chu, Paul K.

2015-01-01

Positively-charged surfaces on implants have a similar potential to upregulate osteogenesis of bone marrow-derived mesenchymal stem cells (BMSCs) as electromagnetic therapy approved for bone regeneration. Generally, their osteogenesis functions are generally considered to stem from the charge-induced adhesion of extracellular matrix (ECM) proteins without exploring the underlying surface charge/cell signaling molecule pathways. Herein, a positively-charged surface with controllable tertiary amines is produced on a polymer implant by plasma surface modification. In addition to inhibiting the TNF-α expression, the positively-charged surface with tertiary amines exhibits excellent cytocompatibility as well as remarkably upregulated osteogenesis-related gene/protein expressions and calcification of the contacted BMSCs. Stimulated by the charged surface, these BMSCs display high iNOS expressions among the three NOS isoforms. Meanwhile, downregulation of the iNOS by L-Can or siRNA inhibit osteogenic differentiation in the BMSCs. These findings suggest that a positively-charged surface with tertiary amines induces osteogenesis of BMSCs via the surface charge/iNOS signaling pathway in addition to elevated ECM protein adhesion. Therefore, creating a positively-charged surface with tertiary amines is a promising approach to promote osseointegration with bone tissues. PMID:25791957

Extended Culture of Bone Marrow with Granulocyte Macrophage-Colony Stimulating Factor Generates Immunosuppressive Cells

PubMed Central

Na, Hye Young; Sohn, Moah; Ryu, Seul Hye; Choi, Wanho; In, Hyunju; Shin, Hyun Soo

2018-01-01

Bone marrow-derived dendritic cells (BM-DCs) are generated from bone marrow (BM) cells cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) for a week. In this study we investigated the effect of duration on the BM culture with GM-CSF. Within several months, the cells in the BM culture gradually expressed homogeneous levels of CD11c and major histocompatibility complex II on surface, and they became unable to stimulate allogeneic naïve T cells in mixed lymphocyte reaction (MLR). In addition, when the BM culture were sustained for 32 wk or longer, the BM cells acquired ability to suppress the proliferation of allogeneic T cells in MLR as well as the response of ovalbumin-specific OT-I transgenic T cells in antigen-dependent manner. We found that, except for programmed death-ligand 1, most cell surface molecules were expressed lower in the BM cells cultured with GM-CSF for the extended duration. These results indicate that BM cells in the extended culture with GM-CSF undergo 2 distinct steps of functional change; first, they lose the immunostimulatory capacity; and next, they gain the immunosuppressive ability. PMID:29736292

Cells Recognize and Prefer Bone-like Hydroxyapatite: Biochemical Understanding of Ultrathin Mineral Platelets in Bone.

PubMed

Liu, Cuilian; Zhai, Halei; Zhang, Zhisen; Li, Yaling; Xu, Xurong; Tang, Ruikang

2016-11-09

Hydroxyapatite (HAP) nanocrystallites in all types of bones are distinguished by their ultrathin characteristics, which are uniaxially oriented with fibrillar collagen to uniquely expose the (100) faces. We speculate that living organisms prefer the specific crystal morphology and orientation of HAP because of the interactions between cells and crystals at the mineral-cell interface. Here, bone-like platy HAP (p-HAP) and two different rod-like HAPs were synthesized to investigate the ultrathin mineral modulating effect on cell bioactivity and bone generation. Cell viability and osteogenic differentiation of mesenchymal stem cells (MSCs) were significantly promoted by the platy HAP with (100) faces compared to rod-like HAPs with (001) faces as the dominant crystal orientation, which indicated that MSCs can recognize the crystal face and prefer the (100) HAP faces. This face-specific preference is dependent on the selective adsorption of fibronectin (FN), a plasma protein that plays a central role in cell adhesion, on the HAP surface. This selective adsorption is further confirmed by molecule dynamics (MD) simulation. Our results demonstrate that it is an intelligent choice for cells to use ultrathin HAP with a large (100) face as a basic building block in the hierarchical structure of bone, which is crucial to the promotion of MSCs osteoinductions during bone formation.

Use of Hes1-GFP reporter mice to assess activity of the Hes1 promoter in bone cells under chronic inflammation

PubMed Central

Zhang, Hengwei; Sun, Wen; Li, Xing; Wang, Mengmeng; Boyce, Brendan F; Hilton, Matthew J; Xing, Lianping

2016-01-01

Notch signaling plays a critical role in maintaining bone homeostasis partially by controlling the formation of osteoblasts from mesenchymal stem cells (MSCs). We reported that TNF activates Notch signaling in MSCs which inhibits osteoblast differentiation in TNF transgenic (TNF-Tg) mice, a mouse model of chronic inflammatory arthritis. In the current study, we used Hes1-GFP and Hes1-GFP/TNF-Tg mice to study the distribution and dynamic change of Notch active cells in normal and inflammatory bone loss and mechanisms mediating their enhanced proliferation. We found that Hes1-GFP+ cells are composed of cells expressing mesenchymal, hematopoietic and endothelial surface markers. CD45−/Hes1-GFP+ cells express high levels of mesenchymal markers and form CFU-F and CFU-ALP colonies. Expansion of CFU-F colonies is associated with a rapid increase in Hes1-GFP+ cell numbers and their GFP intensity. The GFP signal is lost when a CFU-F colony differentiates into an ALP+ osteoblast colony. TNF increases the numbers of CD45−/Hes1-GFP+ cells, which are stained negatively for osteoblast marker osteocalcin and localized adjacent to endosteal and trabecular bone surfaces. CD45−/Hes1-GFP+ cells in Hes1-GFP/TNF-Tg mice have increased BrdU incorporation and PDGFRβ levels. TNF increases the number of proliferating Hes1-GFP+ cells, which is prevented by a specific PDGFRβ inhibitor. Notch inhibition blocks TNF-mediated PDGFRβ expression and cell proliferation. Thus, TNF-induced MSC proliferation is mediated by PDGFRβ signal, which works at downstream of Notch. Hes1-GFP mice can be used to assess the activation status of Notch in bone cells. PMID:27269414

Use of Hes1-GFP reporter mice to assess activity of the Hes1 promoter in bone cells under chronic inflammation.

PubMed

Zhang, Hengwei; Sun, Wen; Li, Xing; Wang, Mengmeng; Boyce, Brendan F; Hilton, Matthew J; Xing, Lianping

2016-09-01

Notch signaling plays a critical role in maintaining bone homeostasis partially by controlling the formation of osteoblasts from mesenchymal stem cells (MSCs). We reported that TNF activates Notch signaling in MSCs which inhibits osteoblast differentiation in TNF transgenic (TNF-Tg) mice, a mouse model of chronic inflammatory arthritis. In the current study, we used Hes1-GFP and Hes1-GFP/TNF-Tg mice to study the distribution and dynamic change of Notch active cells in normal and inflammatory bone loss and mechanisms mediating their enhanced proliferation. We found that Hes1-GFP+ cells are composed of cells expressing mesenchymal, hematopoietic and endothelial surface markers. CD45-/Hes1-GFP+ cells express high levels of mesenchymal markers and form CFU-F and CFU-ALP colonies. Expansion of CFU-F colonies is associated with a rapid increase in Hes1-GFP+ cell numbers and their GFP intensity. The GFP signal is lost when a CFU-F colony differentiates into an ALP+ osteoblast colony. TNF increases the numbers of CD45-/Hes1-GFP+ cells, which are stained negatively for osteoblast marker osteocalcin and localized adjacent to endosteal and trabecular bone surfaces. CD45-/Hes1-GFP+ cells in Hes1-GFP/TNF-Tg mice have increased BrdU incorporation and PDGFRβ levels. TNF increases the number of proliferating Hes1-GFP+ cells, which is prevented by a specific PDGFRβ inhibitor. Notch inhibition blocks TNF-mediated PDGFRβ expression and cell proliferation. Thus, TNF-induced MSC proliferation is mediated by PDGFRβ signal, which works at downstream of Notch. Hes1-GFP mice can be used to assess the activation status of Notch in bone cells. Copyright © 2016 Elsevier Inc. All rights reserved.

In vivo XCT bone characterization of lattice structured implants fabricated by additive manufacturing.

PubMed

Obaton, A-F; Fain, J; Djemaï, M; Meinel, D; Léonard, F; Mahé, E; Lécuelle, B; Fouchet, J-J; Bruno, G

2017-08-01

Several cylindrical specimens and dental implants, presenting diagonal lattice structures with different cell sizes (600, 900 and 1200 μm) were additively manufactured by selective laser melting process. Then they were implanted for two months in a sheep. After removal, they were studied by Archimedes' method as well as X-ray computed tomography in order to assess the penetration of bone into the lattice. We observed that the additive manufactured parts were geometrically conformed to the theoretical specifications. However, several particles were left adhering to the surface of the lattice, thereby partly or entirely obstructing the cells. Nevertheless, bone penetration was clearly visible. We conclude that the 900 μm lattice cell size is more favourable to bone penetration than the 1200 μm lattice cell size, as the bone penetration is 84% for 900 μm against 54% for 1200 μm cell structures. The lower bone penetration value for the 1200 μm lattice cell could possibly be attributed to the short residence time in the sheep. Our results lead to the conclusion that lattice implants additively manufactured by selective laser melting enable better bone integration.

Prevention of glucocorticoid induced bone changes with beta-ecdysone

PubMed Central

Dai, Weiwei; Jiang, Li; Lay, Yu-An Evan; Chen, Haiyan; Jin, Guoqin; Zhang, Hongliang; Kot, Alex; Ritchie, Robert O.; Lane, Nancy E.; Yao, Wei

2015-01-01

Beta-ecdysone (βEcd) is a phytoecdysteroid found in the dry roots and seeds of the asteraceae and achyranthes plants, and is reported to increase osteogenesis in vitro. Since glucocorticoid (GCs) excess is associated with a decrease in bone formation, the purpose of this study was to determine if treatment with βEcd could prevent GC-induced osteoporosis. Two-month-old male Swiss-Webster mice (n=8-10/group) were randomized to either placebo or slow release prednisolone pellets (3.3mg/kg/d) and treated with vehicle control or βEcd (0.5mg/kg/d) for 21 days. GC treatment inhibited age-dependent trabecular gain and cortical bone expansion and this was accompanied by a 30-50% lower bone formation rate (BFR) at both the endosteal and periosteal surfaces. Mice treated with only βEcd significantly increased bone formation on endosteal and periosteal bone surfaces, and increased cortical bone mass were their controls to compare to GC alone. Concurrent treatment of βEcd and GC completely prevented the GC-induced reduction in BFR, trabecular bone volume and partially prevented cortical bone loss. In vitro studies determined that βEcd prevented the GC increase in autophagy of the bone marrow stromal cells as well as in whole bone. In summary, βEcd prevented GC induced changes in bone formation, bone cell viability and bone mass. Additional studies are warranted of βEcd for the treatment of GC induced bone loss. PMID:25585248

Prevention of glucocorticoid induced bone changes with beta-ecdysone.

PubMed

Dai, Weiwei; Jiang, Li; Lay, Yu-An Evan; Chen, Haiyan; Jin, Guoqin; Zhang, Hongliang; Kot, Alexander; Ritchie, Robert O; Lane, Nancy E; Yao, Wei

2015-05-01

Beta-ecdysone (βEcd) is a phytoecdysteroid found in the dry roots and seeds of the asteraceae and achyranthes plants, and is reported to increase osteogenesis in vitro. Since glucocorticoid (GC) excess is associated with a decrease in bone formation, the purpose of this study was to determine if treatment with βEcd could prevent GC-induced osteoporosis. Two-month-old male Swiss-Webster mice (n=8-10/group) were randomized to either placebo or slow release prednisolone pellets (3.3mg/kg/day) and treated with vehicle control or βEcd (0.5mg/kg/day) for 21days. GC treatment inhibited age-dependent trabecular gain and cortical bone expansion and this was accompanied by a 30-50% lower bone formation rate (BFR) at both the endosteal and periosteal surfaces. Mice treated with only βEcd significantly increased bone formation on the endosteal and periosteal bone surfaces, and increased cortical bone mass were their controls to compare to GC alone. Concurrent treatment of βEcd and GC completely prevented the GC-induced reduction in BFR, trabecular bone volume and partially prevented cortical bone loss. In vitro studies determined that βEcd prevented the GC increase in autophagy of the bone marrow stromal cells as well as in whole bone. In summary, βEcd prevented GC induced changes in bone formation, bone cell viability and bone mass. Additional studies are warranted of βEcd for the treatment of GC induced bone loss. Copyright © 2015 Elsevier Inc. All rights reserved.

The effect of hydrophilic titanium surface modification on macrophage inflammatory cytokine gene expression.

PubMed

Hamlet, Stephen; Alfarsi, Mohammed; George, Roy; Ivanovski, Saso

2012-05-01

Chemical modification of microrough titanium dental implants to produce a hydrophilic surface with increased wettability and improved surface energy has been demonstrated clinically to achieve superior bone wound healing and osseointegration compared to that achieved with a microrough titanium surface alone. As the recruitment of the necessary osseoinductive precursors involved in bone wound healing and osseointegration to the wound site is facilitated by the action of cytokines, this study sought to determine the in vitro effect of hydrophilic surface modification on the expression of pro-inflammatory cytokines from adherent macrophages. The surface topography and composition of the titanium surfaces was characterized by scanning electron microscopy and X-ray photoelectron spectroscopy. Macrophage attachment and proliferation was assessed using an MTT assay. The expression of 84 pro-inflammatory cytokines and chemokines by adherent RAW 264.7 cells, a murine leukaemic monocyte cell line, was assessed by PCR array after 24 h culture on either smooth polished, sand-blasted acid-etched (SLA) or hydrophilic-modified SLA (SLActive) titanium surfaces. Following 24 h culture on titanium, surface microroughness activated pro-inflammatory cytokine gene transcription in RAW 264.7 cells. Although there was no significant difference in the degree of cellular attachment or proliferation of RAW 264.7 cells to the different titanium surfaces, by 24 h the hydrophilic surface elicited a gene expression profile with significant down-regulation of the key pro-inflammatory cytokines Tnfα, IL-1α, IL-1β and the chemokine Ccl-2. Down-regulation of the expression of pro-inflammatory cytokine genes may thus modulate the inflammatory response and may facilitate the enhanced bone wound healing and osseointegration observed clinically using implants with a microrough hydrophilic surface. © 2011 John Wiley & Sons A/S.

Upregulation of cell proliferation via Shc and ERK1/2 MAPK signaling in SaOS-2 osteoblasts grown on magnesium alloy surface coating with tricalcium phosphate.

PubMed

Jiang, Tianlong; Guo, Lei; Ni, Shenghui; Zhao, Yuyan

2015-04-01

Magnesium (Mg) alloys have been demonstrated to be viable orthopedic implants because of mechanical and biocompatible properties similar to natural bone. In order to improve its osteogenic properties, a porous β-tricalcium phosphate (β-TCP) was coated on the Mg-3AI-1Zn alloy by alkali-heat treatment technique. The human bone-derived cells (SaOS-2) were cultured on (β-TCP)-Mg-3AI-1Zn in vitro, and the osteoblast response, the morphology and the elements on this alloy surface were investigated. Also, the regulation of key intracellular signalling proteins was investigated in the SaOS-2 cells cultured on alloy surface. The results from scanning electron microscope and immunofluorescence staining demonstrated that (β-TCP)-Mg-3AI-1Zn induced significant osteogenesis. SaOS-2 cell proliferation was improved by β-TCP coating. Moreover, the (β-TCP)-Mg-3AI-1Zn surface induced activation of key intracellular signalling proteins in SaOS-2 cells. We observed an enhanced activation of Src homology and collagen (Shc), a common point of integration between bone morphogenetic protein 2, and the Ras/mitogen-activated protein kinase (MAPK) pathway. ERK1/2 MAP kinase activation was also upregulated, suggesting a role in mediating osteoblastic cell interactions with biomaterials. The signalling pathway involving c-fos (member of the activated protein-1) was also shown to be upregulated in osteoblasts cultured on the (β-TCP)-Mg-3AI-1Zn. These results suggest that β-TCP coating may contribute to successful osteoblast function on Mg alloy surface. (β-TCP)-Mg-3AI-1Zn may upregulate cell proliferation via Shc and ERK1/2 MAPK signaling in SaOS-2 osteoblasts grown on Mg alloy surface.

Surface bioactivity modification of titanium by CO 2 plasma treatment and induction of hydroxyapatite: In vitro and in vivo studies

NASA Astrophysics Data System (ADS)

Hu, Xixue; Shen, Hong; Shuai, Kegang; Zhang, Enwei; Bai, Yanjie; Cheng, Yan; Xiong, Xiaoling; Wang, Shenguo; Fang, Jing; Wei, Shicheng

2011-01-01

Since metallic biomaterials used for orthopedic and dental implants possess a paucity of reactive functional groups, bioactivity modification of these materials is challenging. In the present work, the titanium discs and rods were treated with carbon dioxide plasma and then incubated in a modified simulated body fluid 1.5SBF to obtain a hydroxyapatite layer. Surface hydrophilicity of samples, changes of surface chemistry, surface morphologies of samples, and structural analysis of formed hydroxyapatite were investigated by contact angle to water, X-ray photoelectron spectrometer (XPS), scanning electron microscopy (SEM), Fourier transform infrared (FTIR) and X-ray diffraction (XRD). The results demonstrated that hydrophilicity of titanium surface was improved and hydroxyl groups increased after modification with carbon dioxide plasma treatment. The hydroxyl groups on the surface of titanium were the richest after carbon dioxide plasma treatment under the condition of 20 W for less than 30 s. The hydroxyapatite formability of titanium surface was enhanced by carbon dioxide plasma pretreatment, which was attributed to the surface chemistry. MC3T3-E1 cell as a model cell was cultured on the Ti, CPT-Ti and CPT/SBF-Ti discs in vitro, and the results of the morphology and differentiation of the cell showed that CPT/SBF-Ti was the highest bioactive. The relative parameters of the new bone around the Ti and CPT/SBF-Ti rods including bone mineral density (BMD), a ratio of bone volume to total volume (BV/TV), trabecular thickness (Tb.Th.) and trabecular number (Tb.N.) were analyzed by a micro-computed tomography (micro-CT) after 4-, 8- and 12-week implantation periods in vivo. The results indicated that the CPT/SBF-Ti was more advantageous for new bone formation.

3D Powder Printed Bioglass and β-Tricalcium Phosphate Bone Scaffolds.

PubMed

Seidenstuecker, Michael; Kerr, Laura; Bernstein, Anke; Mayr, Hermann O; Suedkamp, Norbert P; Gadow, Rainer; Krieg, Peter; Hernandez Latorre, Sergio; Thomann, Ralf; Syrowatka, Frank; Esslinger, Steffen

2017-12-22

The use of both bioglass (BG) and β tricalcium phosphate (β-TCP) for bone replacement applications has been studied extensively due to the materials' high biocompatibility and ability to resorb when implanted in the body. 3D printing has been explored as a fast and versatile technique for the fabrication of porous bone scaffolds. This project investigates the effects of using different combinations of a composite BG and β-TCP powder for 3D printing of porous bone scaffolds. Porous 3D powder printed bone scaffolds of BG, β-TCP, 50/50 BG/β-TCP and 70/30 BG/β-TCP compositions were subject to a variety of characterization and biocompatibility tests. The porosity characteristics, surface roughness, mechanical strength, viability for cell proliferation, material cytotoxicity and in vitro bioactivity were assessed. The results show that the scaffolds can support osteoblast-like MG-63 cells growth both on the surface of and within the scaffold material and do not show alarming cytotoxicity; the porosity and surface characteristics of the scaffolds are appropriate. Of the two tested composite materials, the 70/30 BG/β-TCP scaffold proved to be superior in terms of biocompatibility and mechanical strength. The mechanical strength of the scaffolds makes them unsuitable for load bearing applications. However, they can be useful for other applications such as bone fillers.

Bone regeneration: in vitro evaluation of the behaviour of osteoblast-like MG63 cells placed in contact with polylactic-co-glycolic acid, deproteinized bovine bone and demineralized freeze-dried bone allograft.

PubMed

Pappalardo, S; Mastrangelo, F; Reale Marroccia, D; Cappello, V; Ciampoli, C; Carlino, V; Tanteri, L; Costanzo, M; Sinatra, F; Tetè, S

2008-01-01

Insufficient bone density of the alveolar crests, caused by loss of the dental elements, sometimes impedes the primary stability of an integrated bone implant. The techniques of bone regeneration allow to obtain a sufficient quantity of alveolar bone to permit the implant rehabilitation of the edentulous crests. Today several grafting materials are available and they have different characteristics, according to their structure, which influence the different behaviour of the grafting materials to the bone and the implant surface. The aim of this study is to evaluate the interaction between a human osteosarcoma MG63 cell line and three different biomaterials: polylactic-co-glycolic acid (PLAGA), deproteinized bovine bone and demineralised freeze-dried bone allograft (DFDBA). From this study a different behaviour emerges of the osteoblast-like MG63 cells in relation to the sublayer on which these cells were placed in culture. The results of the study, in fact, demonstrate that the most osteoconductive material of the three analysed is the DFDBA, followed by DPBB. On the contrary, the PLGA, because of its roughness, does not seem to represent a valid support for cell growth, and does not encourage any morphologic modification in tumor cells. Furthermore, deproteinized bovine bone shows a differentiating effect which could lead to hypothesise an osteoconductive capacity of this biomaterial. Further studies should be carried out with the aim of explaining the results obtained.

Micro-Topographies Promote Late Chondrogenic Differentiation Markers in the ATDC5 Cell Line.

PubMed

Le, Bach Q; Vasilevich, Aliaksei; Vermeulen, Steven; Hulshof, Frits; Stamatialis, Dimitrios F; van Blitterswijk, Clemens A; de Boer, Jan

2017-05-01

Chemical and mechanical cues are well-established influencers of in vitro chondrogenic differentiation of ATDC5 cells. Here, we investigate the role of topographical cues in this differentiation process, a study not been explored before. Previously, using a library of surface micro-topographies we found some distinct patterns that induced alkaline phosphatase (ALP) production in human mesenchymal stromal cells. ALP is also a marker for hypertrophy, the end stage of chondrogenic differentiation preceding bone formation. Thus, we hypothesized that these patterns could influence end-stage chondrogenic differentiation of ATDC5 cells. In this study, we randomly selected seven topographies among the ALP influencing hits. Cells grown on these surfaces displayed varying nuclear shape and actin filament structure. When stimulated with insulin-transferrin-selenium (ITS) medium, nodule formation occurred and in some cases showed alignment to the topographical patterns. Gene expression analysis of cells growing on topographical surfaces in the presence of ITS medium revealed a downregulation of early markers and upregulation of late markers of chondrogenic differentiation compared to cells grown on a flat surface. In conclusion, we demonstrated that surface topography in addition to other cues can promote hypertrophic differentiation suitable for bone tissue engineering.

Ti nanorod arrays with a medium density significantly promote osteogenesis and osteointegration

PubMed Central

Ning, Chengyun; Wang, Shuangying; Zhu, Ye; Zhong, Meiling; Lin, Xi; Zhang, Yu; Tan, Guoxin; Li, Mei; Yin, Zhaoyi; Yu, Peng; Wang, Xiaolan; Li, Ying; He, Tianrui; Chen, Wei; Wang, Yingjun; Mao, Chuanbin

2016-01-01

Ti implants are good candidates in bone repair. However, how to promote bone formation on their surface and their consequent perfect integration with the surrounding tissue is still a challenge. To overcome such challenge, we propose to form Ti nanorods on their surface to promote the new bone formation around the implants. Here Ti nanorod arrays (TNrs) with different densities were produced on pure Ti surfaces using an anodizing method. The influence of TNr density on the protein adsorption as well as on the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 pre-osteoblastic cells were assessed. The TNrs were also implanted into the bone defects in rabbits to test their application in promoting bone formation and osteointegration at the implant-bone interface. TNrs with the medium density were found to show the best capability in promoting the protein adsorption from surrounding medium, which in turn efficiently enhanced osteogenic differentiation in vitro and osteointegration in vivo. Our work suggests that growing TNrs with a medium density on the surface of traditional Ti implants is an efficient and facile method for promoting bone formation and osteointegration in bone repair. PMID:26743328

Ti nanorod arrays with a medium density significantly promote osteogenesis and osteointegration

NASA Astrophysics Data System (ADS)

Ning, Chengyun; Wang, Shuangying; Zhu, Ye; Zhong, Meiling; Lin, Xi; Zhang, Yu; Tan, Guoxin; Li, Mei; Yin, Zhaoyi; Yu, Peng; Wang, Xiaolan; Li, Ying; He, Tianrui; Chen, Wei; Wang, Yingjun; Mao, Chuanbin

2016-01-01

Ti implants are good candidates in bone repair. However, how to promote bone formation on their surface and their consequent perfect integration with the surrounding tissue is still a challenge. To overcome such challenge, we propose to form Ti nanorods on their surface to promote the new bone formation around the implants. Here Ti nanorod arrays (TNrs) with different densities were produced on pure Ti surfaces using an anodizing method. The influence of TNr density on the protein adsorption as well as on the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 pre-osteoblastic cells were assessed. The TNrs were also implanted into the bone defects in rabbits to test their application in promoting bone formation and osteointegration at the implant-bone interface. TNrs with the medium density were found to show the best capability in promoting the protein adsorption from surrounding medium, which in turn efficiently enhanced osteogenic differentiation in vitro and osteointegration in vivo. Our work suggests that growing TNrs with a medium density on the surface of traditional Ti implants is an efficient and facile method for promoting bone formation and osteointegration in bone repair.

Mechanotransduction and the functional response of bone to mechanical strain

NASA Technical Reports Server (NTRS)

Duncan, R. L.; Turner, C. H.

1995-01-01

Mechanotransduction plays a crucial role in the physiology of many tissues including bone. Mechanical loading can inhibit bone resorption and increase bone formation in vivo. In bone, the process of mechanotransduction can be divided into four distinct steps: (1) mechanocoupling, (2) biochemical coupling, (3) transmission of signal, and (4) effector cell response. In mechanocoupling, mechanical loads in vivo cause deformations in bone that stretch bone cells within and lining the bone matrix and create fluid movement within the canaliculae of bone. Dynamic loading, which is associated with extracellular fluid flow and the creation of streaming potentials within bone, is most effective for stimulating new bone formation in vivo. Bone cells in vitro are stimulated to produce second messengers when exposed to fluid flow or mechanical stretch. In biochemical coupling, the possible mechanisms for the coupling of cell-level mechanical signals into intracellular biochemical signals include force transduction through the integrin-cytoskeleton-nuclear matrix structure, stretch-activated cation channels within the cell membrane, G protein-dependent pathways, and linkage between the cytoskeleton and the phospholipase C or phospholipase A pathways. The tight interaction of each of these pathways would suggest that the entire cell is a mechanosensor and there are many different pathways available for the transduction of a mechanical signal. In the transmission of signal, osteoblasts, osteocytes, and bone lining cells may act as sensors of mechanical signals and may communicate the signal through cell processes connected by gap junctions. These cells also produce paracrine factors that may signal osteoprogenitors to differentiate into osteoblasts and attach to the bone surface. Insulin-like growth factors and prostaglandins are possible candidates for intermediaries in signal transduction. In the effector cell response, the effects of mechanical loading are dependent upon the magnitude, duration, and rate of the applied load. Longer duration, lower amplitude loading has the same effect on bone formation as loads with short duration and high amplitude. Loading must be cyclic to stimulate new bone formation. Aging greatly reduces the osteogenic effects of mechanical loading in vivo. Also, some hormones may interact with local mechanical signals to change the sensitivity of the sensor or effector cells to mechanical load.

Enhancing the Hydrophilicity and Cell Attachment of 3D Printed PCL/Graphene Scaffolds for Bone Tissue Engineering

PubMed Central

Wang, Weiguang; Caetano, Guilherme; Ambler, William Stephen; Blaker, Jonny James; Frade, Marco Andrey; Mandal, Parthasarathi; Diver, Carl; Bártolo, Paulo

2016-01-01

Scaffolds are physical substrates for cell attachment, proliferation, and differentiation, ultimately leading to the regeneration of tissues. They must be designed according to specific biomechanical requirements, i.e., certain standards in terms of mechanical properties, surface characteristics, porosity, degradability, and biocompatibility. The optimal design of a scaffold for a specific tissue strongly depends on both materials and manufacturing processes, as well as surface treatment. Polymeric scaffolds reinforced with electro-active particles could play a key role in tissue engineering by modulating cell proliferation and differentiation. This paper investigates the use of an extrusion-based additive manufacturing system to produce poly(ε-caprolactone) (PCL)/pristine graphene scaffolds for bone tissue applications and the influence of chemical surface modification on their biological behaviour. Scaffolds with the same architecture but different concentrations of pristine graphene were evaluated from surface property and biological points of view. Results show that the addition of pristine graphene had a positive impact on cell viability and proliferation, and that surface modification leads to improved cell response. PMID:28774112

Enhancing the Hydrophilicity and Cell Attachment of 3D Printed PCL/Graphene Scaffolds for Bone Tissue Engineering.

PubMed

Wang, Weiguang; Caetano, Guilherme; Ambler, William Stephen; Blaker, Jonny James; Frade, Marco Andrey; Mandal, Parthasarathi; Diver, Carl; Bártolo, Paulo

2016-12-07

Scaffolds are physical substrates for cell attachment, proliferation, and differentiation, ultimately leading to the regeneration of tissues. They must be designed according to specific biomechanical requirements, i.e., certain standards in terms of mechanical properties, surface characteristics, porosity, degradability, and biocompatibility. The optimal design of a scaffold for a specific tissue strongly depends on both materials and manufacturing processes, as well as surface treatment. Polymeric scaffolds reinforced with electro-active particles could play a key role in tissue engineering by modulating cell proliferation and differentiation. This paper investigates the use of an extrusion-based additive manufacturing system to produce poly( ε -caprolactone) (PCL)/pristine graphene scaffolds for bone tissue applications and the influence of chemical surface modification on their biological behaviour. Scaffolds with the same architecture but different concentrations of pristine graphene were evaluated from surface property and biological points of view. Results show that the addition of pristine graphene had a positive impact on cell viability and proliferation, and that surface modification leads to improved cell response.

The effects of implant topography on osseointegration under estrogen deficiency induced osteoporotic conditions: Histomorphometric, transcriptional and ultrastructural analysis.

PubMed

Du, Zhibin; Xiao, Yin; Hashimi, Saeed; Hamlet, Stephen M; Ivanovski, Saso

2016-09-15

Compromised bone quality and/or healing in osteoporosis are recognised risk factors for impaired dental implant osseointegration. This study examined the effects of (1) experimentally induced osteoporosis on titanium implant osseointegration and (2) the effect of modified implant surface topography on osseointegration under osteoporosis-like conditions. Machined and micro-roughened surface implants were placed into the maxillary first molar root socket of 64 ovariectomised and sham-operated Sprague-Dawley rats. Subsequent histological and SEM observations showed tissue maturation on the micro-rough surfaced implants in ovariectomised animals as early as 3days post-implantation. The degree of osseointegration was also significantly higher around the micro-rough implants in ovariectomised animals after 14days of healing although by day 28, similar levels of osseointegration were found for all test groups. The micro-rough implants significantly increased the early (day 3) gene expression of alkaline phosphatase, osteocalcin, receptor activator of nuclear factor kappa-B ligand and dentin matrix protein 1 in implant adherent cells. By day 7, the expression of inflammatory genes decreased while the expression of the osteogenic markers increased further although there were few statistically significant differences between the micro-rough and machined surfaces. Osteocyte morphology was also affected by estrogen deficiency with the size of the cells being reduced in trabecular bone. In conclusion, estrogen deficiency induced osteoporotic conditions negatively influenced the early osseointegration of machined implants while micro-rough implants compensated for these deleterious effects by enhancing osteogenic cell differentiation on the implant surface. Lower bone density, poor bone quality and osseous microstructural changes are all features characteristic of osteoporosis that may impair the osseointegration of dental implants. Using a clinically relevant trabecular bone model in the rat maxilla, we demonstrated histologically that the negative effects of surgically-induced osteoporosis on osseointegration could be ameliorated by the biomaterial's surface topography. Furthermore, gene expression analysis suggests this may be a result of enhanced osteogenic cell differentiation on the implant surface. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Fortifying the Bone-Implant Interface Part 1: An In Vitro Evaluation of 3D-Printed and TPS Porous Surfaces

PubMed Central

Lindsey, Derek P.; Bahney, Chelsea S.; Woods, Shane A.; Wolfe, Mark L.; Yerby, Scott A.

2017-01-01

Background An aging society and concomitant rise in the incidence of impaired bone health have led to the need for advanced osteoconductive spinal implant surfaces that promote greater biological fixation (e.g. for interbody fusion cages, sacroiliac joint fusion implants, and artificial disc replacements). Additive manufacturing, i.e. 3D-printing, may improve bone integration by generating biomimetic spinal implant surfaces that mimic bone morphology. Such surfaces may foster an enhanced cellular response compared to traditional implant surfacing processes. Methods This study investigated the response of human osteoblasts to additive manufactured (AM) trabecular-like titanium implant surfaces compared to traditionally machined base material with titanium plasma spray (TPS) coated surfaces, with and without a nanocrystalline hydroxyapatite (HA) coating. For TPS-coated discs, wrought Ti6Al4V ELI was machined and TPS-coating was applied. For AM discs, Ti6Al4V ELI powder was 3D-printed to form a solid base and trabecular-like porous surface. The HA-coating was applied via a precipitation dip-spin method. Surface porosity, pore size, thickness, and hydrophilicity were characterized. Initial cell attachment, proliferation, alkaline phosphatase (ALP) activity, and calcium production of hFOB cells (n=5 per group) were measured. Results Cells on AM discs exhibited expedited proliferative activity. While there were no differences in mean ALP expression and calcium production between TPS and AM discs, calcium production on the AM discs trended 48% higher than on TPS discs (p=0.07). Overall, HA-coating did not further enhance results compared to uncoated TPS and AM discs. Conclusions Results demonstrate that additive manufacturing allows for controlled trabecular-like surfaces that promote earlier cell proliferation and trends toward higher calcium production than TPS coating. Results further showed that nanocrystalline HA may not provide an advantage on porous titanium surfaces. Clinical Relevance Additive manufactured porous titanium surfaces may induce a more osteogenic environment compared to traditional TPS, and thus present as an attractive alternative to TPS-coating for orthopedic spinal implants. PMID:28765799

Cell-specific paracrine actions of IL-6 family cytokines from bone, marrow and muscle that control bone formation and resorption.

PubMed

Sims, Natalie A

2016-10-01

Bone renews itself and changes shape throughout life to account for the changing needs of the body; this requires co-ordinated activities of bone resorbing cells (osteoclasts), bone forming cells (osteoblasts) and bone's internal cellular network (osteocytes). This review focuses on paracrine signaling by the IL-6 family of cytokines between bone cells, bone marrow, and skeletal muscle in normal physiology and in pathological states where their levels may be locally or systemically elevated. These functions include the support of osteoclast formation by osteoblast lineage cells in response to interleukin 6 (IL-6), interleukin 11 (IL-11), oncostatin M (OSM) and cardiotrophin 1 (CT-1). In addition it will discuss how bone-resorbing osteoclasts promote osteoblast activity by secreting CT-1, which acts as a "coupling factor" on osteocytes, osteoblasts, and their precursors to promote bone formation. OSM, produced by osteoblast lineage cells and macrophages, stimulates bone formation via osteocytes. IL-6 family cytokines also mediate actions of other bone formation stimuli like parathyroid hormone (PTH) and mechanical loading. CT-1, OSM and LIF suppress marrow adipogenesis by shifting commitment of pluripotent precursors towards osteoblast differentiation. Ciliary neurotrophic factor (CNTF) is released as a myokine from skeletal muscle and suppresses osteoblast differentiation and bone formation on the periosteum (outer bone surface in apposition to muscle). Finally, IL-6 acts directly on marrow-derived osteoclasts to stimulate release of "osteotransmitters" that act through the cortical osteocyte network to stimulate bone formation on the periosteum. Each will be discussed as illustrations of how the extended family of IL-6 cytokines acts within the skeleton in physiology and may be altered in pathological conditions or by targeted therapies. Copyright © 2016 Elsevier Ltd. All rights reserved.

The cell biology and role of resorptive cells in diseases: A review.

PubMed

Babaji, Prashant; Devanna, Raghu; Jagtap, Kiran; Chaurasia, Vishwajit Rampratap; Jerry, Jeethu John; Choudhury, Basanta Kumar; Duhan, Dinesh

2017-01-01

Resorptive cells are responsible for the resorption of mineralized matrix of hard tissues. Bone-resorbing cells are called osteoclasts; however, they can resorb mineralized dental tissues or calcified cartilage and then they are called odontoclasts and chondroclasts, respectively. Resorptive cells form when mononuclear precursors derived from a monocyte-macrophage cell lineage are attracted to certain mineralized surfaces and subsequently fuse and adhere onto them for exerting their resorbing activity. These cells are responsible for degradation of calcified extracellular matrix composed of organic molecules and hydroxyapatite. The activity of these cells can be observed in both physiological and pathological processes throughout life and their activity is mainly required in bone turnover and growth, spontaneous and induced (orthodontic) tooth movement, tooth eruption, and bone fracture healing, as well as in pathological conditions such as osteoporosis, osteoarthritis, and bone metastasis. In addition, they are responsible for daily control of calcium homeostasis. Clastic cells also resorb the primary teeth for shedding before the permanent teeth erupt into the oral cavity.

Inorganic materials for bone repair or replacement applications.

PubMed

Hertz, Audrey; Bruce, Ian J

2007-12-01

In recent years, excipient systems have been used increasingly in biomedicine in reconstructive and replacement surgery, as bone cements, drug-delivery vehicles and contrast agents. Particularly, interest has been growing in the development and application of controlled pore inorganic ceramic materials for use in bone-replacement and bone-repair roles and, in this context, attention has been focused on calcium-phosphate, bioactive glasses and SiO2- and TiO2-based materials. It has been shown that inorganic materials that most closely mimic bone structure and surface chemistry most closely function best in bone replacement/repair and, in particular, if a substance possesses a macroporous structure (pores and interconnections >100 microm diameter), then cell infiltration, bone growth and vascularization can all be promoted. The surface roughness and micro/mesoporosity of a material have also been observed to significantly influence its ability to promote apatite nucleation and cell attachment significantly. Pores (where present) can also be packed with pharmaceuticals and biomolecules (e.g., bone morphogenetic proteins [BMPs], which can stimulate bone formation). Finally, the most bio-efficient - in terms of collagen formation and apatite nucleation - materials are those that are able to provide soluble mineralizing species (Si, Ca, PO(4)) at their implant sites and/or are doped or have been surface-activated with specific functional groups. This article presents the context and latest advances in the field of bone-repair materials, especially with respect to the development of bioactive glasses and micro/mesoporous and macroporous inorganic scaffolds. It deals with the possible methods of preparing porous pure/doped or functionalized silicas or their composites, the studies that have been undertaken to evaluate their abilities to act as bone repair scaffolds and also presents future directions for work in that context.

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

Independent effects of the chemical and microstructural surface properties of polymer/ceramic composites on proliferation and osteogenic differentiation of human MSCs.

PubMed

Sun, Lanying; Danoux, Charlène B; Wang, Qibao; Pereira, Daniel; Barata, David; Zhang, Jingwei; LaPointe, Vanessa; Truckenmüller, Roman; Bao, Chongyun; Xu, Xin; Habibovic, Pamela

2016-09-15

Within the general aim of finding affordable and sustainable regenerative solutions for damaged and diseased tissues and organs, significant efforts have been invested in developing synthetic alternatives to natural bone grafts, such as autografts. Calcium phosphate (CaP) ceramics are among widely used synthetic bone graft substitutes, but their mechanical properties and bone regenerative capacity are still outperformed by their natural counterparts. In order to improve the existing synthetic bone graft substitutes, it is imperative to understand the effects of their individual properties on a biological response, and to find a way to combine the desired properties into new, improved functional biomaterials. To this end, we studied the independent effects of the chemical composition and surface microstructure of a poly(lactic acid)/hydroxyapatite (PLA/HA) composite material on the proliferation and osteogenic differentiation of clinically relevant bone marrow-derived human mesenchymal stromal cells (hMSCs). While the molecular weight of the polymer and presence/absence of the ceramic phase were used as the chemical variables, a soft embossing technique was used to pattern the surfaces of all materials with either pits or pillars with identical microscale dimensions. The results indicated that, while cell morphology was affected by both the presence and availability of HA and by the surface microstructure, the effect of the latter parameter on cell proliferation was negligible. The osteogenic differentiation of hMSCs, and in particular the expression of bone morphogenetic protein 2 (BMP-2) and osteopontin (OP) were significantly enhanced when cells were cultured on the composite based on low-molecular-weight PLA, as compared to the high-molecular-weight PLA-based composite and the two pure polymers. The OP expression on the low-molecular-weight PLA-based composite was further enhanced when the surface was patterned with pits. Taken together, within this experimental set up, the individual effect of the chemistry, and in particular of the presence of CaP, was more pronounced than the individual effect of the surface microstructure, although their combined effects were, in some cases, synergistic. The approach presented here opens new routes to study the interactions of biomaterials with the biological environment in greater depths, which can serve as a starting point for developing biomaterials with improved bioactivity. The aim of the this study was to obtain insight into independent effects of the chemical composition and surface microstructure of a poly(lactic acid)/hydroxyapatite (PLA/HA) composite material on the morphology, proliferation and osteogenic differentiation of clinically relevant bone marrow-derived human mesenchymal stromal cells (hMSCs). While the need for synthetic alternatives for natural bone in bone regenerative strategies is rapidly increasing, the clinical performance of synthetic biomaterials needs to be further improved. To do this successfully, we believe that a better understanding of the relationship between a property of a material and a biological response is imperative. This study is a step forward in this direction, and we are therefore convinced that it will be of interest to the readers of Acta Biomaterialia. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Preparation of Laponite Bioceramics for Potential Bone Tissue Engineering Applications

PubMed Central

Li, Kai; Ju, Yaping; Li, Jipeng; Zhang, Yongxing; Li, Jinhua; Liu, Xuanyong; Shi, Xiangyang; Zhao, Qinghua

2014-01-01

We report a facile approach to preparing laponite (LAP) bioceramics via sintering LAP powder compacts for bone tissue engineering applications. The sintering behavior and mechanical properties of LAP compacts under different temperatures, heating rates, and soaking times were investigated. We show that LAP bioceramic with a smooth and porous surface can be formed at 800°C with a heating rate of 5°C/h for 6 h under air. The formed LAP bioceramic was systematically characterized via different methods. Our results reveal that the LAP bioceramic possesses an excellent surface hydrophilicity and serum absorption capacity, and good cytocompatibility and hemocompatibility as demonstrated by resazurin reduction assay of rat mesenchymal stem cells (rMSCs) and hemolytic assay of pig red blood cells, respectively. The potential bone tissue engineering applicability of LAP bioceramic was explored by studying the surface mineralization behavior via soaking in simulated body fluid (SBF), as well as the surface cellular response of rMSCs. Our results suggest that LAP bioceramic is able to induce hydroxyapatite deposition on its surface when soaked in SBF and rMSCs can proliferate well on the LAP bioceramic surface. Most strikingly, alkaline phosphatase activity together with alizarin red staining results reveal that the produced LAP bioceramic is able to induce osteoblast differentiation of rMSCs in growth medium without any inducing factors. Finally, in vivo animal implantation, acute systemic toxicity test and hematoxylin and eosin (H&E)-staining data demonstrate that the prepared LAP bioceramic displays an excellent biosafety and is able to heal the bone defect. Findings from this study suggest that the developed LAP bioceramic holds a great promise for treating bone defects in bone tissue engineering. PMID:24955961

Novel Approach in the Use of Plasma Spray: Preparation of Bulk Titanium for Bone Augmentations

PubMed Central

Fousova, Michaela; Vojtech, Dalibor; Jablonska, Eva; Fojt, Jaroslav; Lipov, Jan

2017-01-01

Thermal plasma spray is a common, well-established technology used in various application fields. Nevertheless, in our work, this technology was employed in a completely new way; for the preparation of bulk titanium. The aim was to produce titanium with properties similar to human bone to be used for bone augmentations. Titanium rods sprayed on a thin substrate wire exerted a porosity of about 15%, which yielded a significant decrease of Young′s modulus to the bone range and provided rugged topography for enhanced biological fixation. For the first verification of the suitability of the selected approach, tests of the mechanical properties in terms of compression, bending, and impact were carried out, the surface was characterized, and its compatibility with bone cells was studied. While preserving a high enough compressive strength of 628 MPa, the elastic modulus reached 11.6 GPa, thus preventing a stress-shielding effect, a generally known problem of implantable metals. U-2 OS and Saos-2 cells derived from bone osteosarcoma grown on the plasma-sprayed surface showed good viability. PMID:28837101

Role of geometrical cues in bone marrow-derived mesenchymal stem cell survival, growth and osteogenic differentiation.

PubMed

Gupta, Dhanak; Grant, David M; Zakir Hossain, Kazi M; Ahmed, Ifty; Sottile, Virginie

2018-02-01

Mesenchymal stem cells play a vital role in bone formation process by differentiating into osteoblasts, in a tissue that offers not a flat but a discontinuous three-dimensional (3D) topography in vivo. In order to understand how geometry may be affecting mesenchymal stem cells, this study explored the influence of 3D geometry on mesenchymal stem cell-fate by comparing cell growth, viability and osteogenic potential using monolayer (two-dimensional, 2D) with microsphere (3D) culture systems normalised to surface area. The results suggested lower cell viability and reduced cell growth in 3D. Alkaline phosphatase activity was higher in 3D; however, both collagen and mineral deposition appeared significantly lower in 3D, even after osteogenic supplementation. Also, there were signs of patchy mineralisation in 3D with or without osteogenic supplementation as early as day 7. These results suggest that the convex surfaces on microspheres and inter-particulate porosity may have led to variable cell morphology and fate within the 3D culture. This study provides deeper insights into geometrical regulation of mesenchymal stem cell responses applicable for bone tissue engineering.

CD146/MCAM defines functionality of human bone marrow stromal stem cell populations.

PubMed

Harkness, Linda; Zaher, Walid; Ditzel, Nicholas; Isa, Adiba; Kassem, Moustapha

2016-01-11

Identification of surface markers for prospective isolation of functionally homogenous populations of human skeletal (stromal, mesenchymal) stem cells (hMSCs) is highly relevant for cell therapy protocols. Thus, we examined the possible use of CD146 to subtype a heterogeneous hMSC population. Using flow cytometry and cell sorting, we isolated two distinct hMSC-CD146(+) and hMSC-CD146(-) cell populations from the telomerized human bone marrow-derived stromal cell line (hMSC-TERT). Cells were examined for differences in their size, shape and texture by using high-content analysis and additionally for their ability to differentiate toward osteogenesis in vitro and form bone in vivo, and their migrational ability in vivo and in vitro was investigated. In vitro, the two cell populations exhibited similar growth rate and differentiation capacity to osteoblasts and adipocytes on the basis of gene expression and protein production of lineage-specific markers. In vivo, hMSC-CD146(+) and hMSC-CD146(-) cells formed bone and bone marrow organ when implanted subcutaneously in immune-deficient mice. Bone was enriched in hMSC-CD146(-) cells (12.6 % versus 8.1 %) and bone marrow elements enriched in implants containing hMSC-CD146(+) cells (0.5 % versus 0.05 %). hMSC-CD146(+) cells exhibited greater chemotactic attraction in a transwell migration assay and, when injected intravenously into immune-deficient mice following closed femoral fracture, exhibited wider tissue distribution and significantly increased migration ability as demonstrated by bioluminescence imaging. Our studies demonstrate that CD146 defines a subpopulation of hMSCs capable of bone formation and in vivo trans-endothelial migration and thus represents a population of hMSCs suitable for use in clinical protocols of bone tissue regeneration.

Osteoinductive activity of insulin-functionalized cell culture surfaces obtained using diazonium chemistry

PubMed Central

Mikulska, Anna; Filipowska, Joanna; Osyczka, Anna M.; Nowakowska, Maria; Szczubiałka, Krzysztof

2015-01-01

Polymeric surfaces suitable for cell culture (DR/Pec) were constructed from diazoresin (DR) and pectin (Pec) in a form of ultrathin films using the layer-by-layer (LbL) technique. The surfaces were functionalized with insulin using diazonium chemistry. Such functionalized surfaces were used to culture human mesenchymal stem cells (hMSCs) to assess their suitability for bone tissue engineering and regeneration. The activity of insulin immobilized on the surfaces (DR/Pec/Ins) was compared to that of insulin dissolved in the culture medium. Human MSC grown on insulin-immobilized DR/Pec surfaces displayed increased proliferation and higher osteogenic activity. The latter was determined by means of alkaline phosphatase (ALP) activity, which increases at early stages of osteoblasts differentiation. Insulin dissolved in the culture medium did not stimulate cell proliferation and its osteogenic activity was significantly lower. Addition of recombinant human bone morphogenetic protein 2 (rhBMP-2) to the culture medium further increased ALP activity in hMSCs indicating additive osteogenic action of immobilized insulin and rhBMP-2. PMID:25629028

Biological response of laser macrostructured and oxidized titanium alloy: an in vitro and in vivo study.

PubMed

Paz, María Dolores; Álava, J Iñaki; Goikoetxea, Leire; Chiussi, Stefano; Díaz-Güemes, Idoia; Usón, Jesus; Sánchez, Francisco; León, Betty

2011-01-01

To assess both the in vitro and in vivo biological response of a laser modified surface in an integrated manner. A combined innovative approach applies lasers to macrostructure as well as to oxidize the surface of titanium alloy implants. A Nd:YAG marking and ArF excimer lasers were used for macrostructuring and UV-oxidizing the surface of Ti6Al4V discs, respectively. Human fetal osteoblastic cell culture and a sheep tibia model were used to assess the cell response and the osseogeneration capability of as-machined, laser macrostructured and laser macrostructured and oxidized surfaces. In vitro: Laser macrostructuration alone did not promote cell response. Cellular proliferation was enhanced by the additional UV laser oxidation. In vivo: A greater significant percentage of bone-implant contact was obtained for both laser treated surfaces compared to machine-turned control samples, three months after implantation, in spite of the low cellular response for macrostructured samples. The use of sheep model for six months appears to be less adequate for a comparison because of the high level of bone integration in all samples. In spite of the often reported positive effect of titanium oxidation on the triggering of faster osseointegration, in this experiment the additional UV laser oxidation did not lead to a significant in vivo improvement. Laser macrostructuration of titanium alloy surfaces appears to promote bone apposition and may therefore constitute a promising surface modification strategy. In animal models, the natural process of titanium surface oxidation, because of physiologic fluids, alters properties observed in vitro with cells.

Covalent Binding of BMP-2 on Surfaces Using a Self-assembled Monolayer Approach

PubMed Central

Pohl, Theresa L. M.; Schwab, Elisabeth H.; Cavalcanti-Adam, Elisabetta A.

2013-01-01

Bone morphogenetic protein 2 (BMP-2) is a growth factor embedded in the extracellular matrix of bone tissue. BMP-2 acts as trigger of mesenchymal cell differentiation into osteoblasts, thus stimulating healing and de novo bone formation. The clinical use of recombinant human BMP-2 (rhBMP-2) in conjunction with scaffolds has raised recent controversies, based on the mode of presentation and the amount to be delivered. The protocol presented here provides a simple and efficient way to deliver BMP-2 for in vitro studies on cells. We describe how to form a self-assembled monolayer consisting of a heterobifunctional linker, and show the subsequent binding step to obtain covalent immobilization of rhBMP-2. With this approach it is possible to achieve a sustained presentation of BMP-2 while maintaining the biological activity of the protein. In fact, the surface immobilization of BMP-2 allows targeted investigations by preventing unspecific adsorption, while reducing the amount of growth factor and, most notably, hindering uncontrolled release from the surface. Both short- and long-term signaling events triggered by BMP-2 are taking place when cells are exposed to surfaces presenting covalently immobilized rhBMP-2, making this approach suitable for in vitro studies on cell responses to BMP-2 stimulation. PMID:24021994

Novel β-TCP Coated Titanium Nanofiber Surface for Enhanced Bone Growth.

PubMed

Lim, Hyun-Pil; Park, Sang-Won; Yun, Kwi-Dug; Park, Chan; Ji, Min-Kyung; Oh, Gye-Jeong; Lee, Jong-Tak; Lee, Kwangmin

2018-02-01

In this study, we examined the effect of β-tricalcium phosphate (β-TCP) coating on alkali-treated CP Grade II titanium surface via RF magnetron sputtering on osteoblast like cell (MC3T3-E1) viability and bone formation in rat tibia. The specimens were divided into three groups; commercially pure titanium (control group), alkali-treated titanium with nanofiber structure (NF group) and β-TCP coating on alkali-treated titanium with nanofiber structure (TNF group). The surface characteristics of specimens were observed under a field emission scanning electron microscope (FE-SEM), and contact angle was measured. The cell viability was assessed in vitro after 1 day, 3 days and 7 days. Implants of 2.0 mm diameter and 5.0 mm length were inserted into the tibia of rats. After 4 wks, the histomorphometric analysis was performed. Group NF and group TNF showed improved hydrophilicity of Ti. Group TNF showed significantly higher cell viability (P < 0.05) after 7 days. The bone to implant contact (BIC) ratio of the control group, NF group, and TNF group were 32.3%, 35.5%, and 63.9%, respectively. The study results suggested that β-TCP coated alkali-treated titanium surface via RF magnetron sputtering might be effective in implant dentistry due to enhanced hydrophilicity, improved cell response, and better osseointegration.

Electrospun silk fibroin/poly(lactide-co-ε-caprolactone) nanofibrous scaffolds for bone regeneration

PubMed Central

Wang, Zi; Lin, Ming; Xie, Qing; Sun, Hao; Huang, Yazhuo; Zhang, DanDan; Yu, Zhang; Bi, Xiaoping; Chen, Junzhao; Wang, Jing; Shi, Wodong; Gu, Ping; Fan, Xianqun

2016-01-01

Background Tissue engineering has become a promising therapeutic approach for bone regeneration. Nanofibrous scaffolds have attracted great interest mainly due to their structural similarity to natural extracellular matrix (ECM). Poly(lactide-co-ε-caprolactone) (PLCL) has been successfully used in bone regeneration, but PLCL polymers are inert and lack natural cell recognition sites, and the surface of PLCL scaffold is hydrophobic. Silk fibroin (SF) is a kind of natural polymer with inherent bioactivity, and supports mesenchymal stem cell attachment, osteogenesis, and ECM deposition. Therefore, we fabricated hybrid nanofibrous scaffolds by adding different weight ratios of SF to PLCL in order to find a scaffold with improved properties for bone regeneration. Methods Hybrid nanofibrous scaffolds were fabricated by blending different weight ratios of SF with PLCL. Human adipose-derived stem cells (hADSCs) were seeded on SF/PLCL nanofibrous scaffolds of various ratios for a systematic evaluation of cell adhesion, proliferation, cytotoxicity, and osteogenic differentiation; the efficacy of the composite of hADSCs and scaffolds in repairing critical-sized calvarial defects in rats was investigated. Results The SF/PLCL (50/50) scaffold exhibited favorable tensile strength, surface roughness, and hydrophilicity, which facilitated cell adhesion and proliferation. Moreover, the SF/PLCL (50/50) scaffold promoted the osteogenic differentiation of hADSCs by elevating the expression levels of osteogenic marker genes such as BSP, Ocn, Col1A1, and OPN and enhanced ECM mineralization. In vivo assays showed that SF/PLCL (50/50) scaffold improved the repair of the critical-sized calvarial defect in rats, resulting in increased bone volume, higher trabecular number, enhanced bone mineral density, and increased new bone areas, compared with the pure PLCL scaffold. Conclusion The SF/PLCL (50/50) nanofibrous scaffold facilitated hADSC proliferation and osteogenic differentiation in vitro and further promoted new bone formation in vivo, suggesting that the SF/PLCL (50/50) nanofibrous scaffold holds great potential in bone tissue regeneration. PMID:27114708

Surface topography of hydroxyapatite promotes osteogenic differentiation of human bone marrow mesenchymal stem cells.

PubMed

Yang, Wanlei; Han, Weiqi; He, Wei; Li, Jianlei; Wang, Jirong; Feng, Haotian; Qian, Yu

2016-03-01

Effective and safe induction of osteogenic differentiation is one of the key elements of bone tissue engineering. Surface topography of scaffold materials was recently found to promote osteogenic differentiation. Utilization of this topography may be a safer approach than traditional induction by growth factors or chemicals. The aim of this study is to investigate the enhancement of osteogenic differentiation by surface topography and its mechanism of action. Hydroxyapatite (HA) discs with average roughness (Ra) of surface topography ranging from 0.2 to 1.65 μm and mean distance between peaks (RSm) ranging from 89.7 to 18.6 μm were prepared, and human bone-marrow mesenchymal stem cells (hBMSCs) were cultured on these discs. Optimal osteogenic differentiation was observed on discs with surface topography characterized by Ra ranging from 0.77 to 1.09 μm and RSm ranging from 53.9 to 39.3 μm. On this surface configuration of HA, hBMSCs showed oriented attachment, F-actin arrangement, and a peak in the expression of Yes-associated protein (YAP) and PDZ binding motif (TAZ) (YAP/TAZ). These results indicated that the surface topography of HA promoted osteogenic differentiation of hBMSCs, possibly by increasing cell attachment and promoting the YAP/TAZ signaling pathway. Copyright © 2015 Elsevier B.V. All rights reserved.

Cellular dosimetry calculations for Strontium-90 using Monte Carlo code PENELOPE.

PubMed

Hocine, Nora; Farlay, Delphine; Boivin, Georges; Franck, Didier; Agarande, Michelle

2014-11-01

To improve risk assessments associated with chronic exposure to Strontium-90 (Sr-90), for both the environment and human health, it is necessary to know the energy distribution in specific cells or tissue. Monte Carlo (MC) simulation codes are extremely useful tools for calculating deposition energy. The present work was focused on the validation of the MC code PENetration and Energy LOss of Positrons and Electrons (PENELOPE) and the assessment of dose distribution to bone marrow cells from punctual Sr-90 source localized within the cortical bone part. S-values (absorbed dose per unit cumulated activity) calculations using Monte Carlo simulations were performed by using PENELOPE and Monte Carlo N-Particle eXtended (MCNPX). Cytoplasm, nucleus, cell surface, mouse femur bone and Sr-90 radiation source were simulated. Cells are assumed to be spherical with the radii of the cell and cell nucleus ranging from 2-10 μm. The Sr-90 source is assumed to be uniformly distributed in cell nucleus, cytoplasm and cell surface. The comparison of S-values calculated with PENELOPE to MCNPX results and the Medical Internal Radiation Dose (MIRD) values agreed very well since the relative deviations were less than 4.5%. The dose distribution to mouse bone marrow cells showed that the cells localized near the cortical part received the maximum dose. The MC code PENELOPE may prove useful for cellular dosimetry involving radiation transport through materials other than water, or for complex distributions of radionuclides and geometries.

The gut microbiota regulates bone mass in mice

PubMed Central

Sjögren, Klara; Engdahl, Cecilia; Henning, Petra; Lerner, Ulf H; Tremaroli, Valentina; Lagerquist, Marie K; Bäckhed, Fredrik; Ohlsson, Claes

2012-01-01

The gut microbiota modulates host metabolism and development of immune status. Here we show that the gut microbiota is also a major regulator of bone mass in mice. Germ-free (GF) mice exhibit increased bone mass associated with reduced number of osteoclasts per bone surface compared with conventionally raised (CONV-R) mice. Colonization of GF mice with a normal gut microbiota normalizes bone mass. Furthermore, GF mice have decreased frequency of CD4+ T cells and CD11b+/GR 1 osteoclast precursor cells in bone marrow, which could be normalized by colonization. GF mice exhibited reduced expression of inflammatory cytokines in bone and bone marrow compared with CONV-R mice. In summary, the gut microbiota regulates bone mass in mice, and we provide evidence for a mechanism involving altered immune status in bone and thereby affected osteoclast-mediated bone resorption. Further studies are required to evaluate the gut microbiota as a novel therapeutic target for osteoporosis. © 2012 American Society for Bone and Mineral Research. PMID:22407806

Effect of Different Titanium Surfaces on Maturation of Murine Bone Marrow-Derived Dendritic Cells

NASA Astrophysics Data System (ADS)

Zheng, Xiaofei; Zhou, Fengjuan; Gu, Yifei; Duan, Xiaobo; Mo, Anchun

2017-02-01

Dendritic cells (DCs) play a pivotal role in the host response to implanted biomaterials. Osseointegration of titanium (Ti) implant is an immunological and inflammatory-driven process. However, the role of DCs in this complex process is largely unknown. This study aimed to investigate the effect of different Ti surfaces on DC maturation, and evaluate its subsequent potential on osteogenic differentiation of preosteoblasts. Murine bone marrow-derived DCs were seeded on Ti disks with different surface treatments, including pretreatment (PT), sandblasted/acid-etched (SLA) and modified SLA (modSLA) surface. Compared with DCs cultured on PT and SLA surfaces, the cells seeded on modSLA surface demonstrated a more round morphology with lower expression of CD86 and MHC-II, the DC maturation markers. Those cells also secreted high levels of anti-inflammatory cytokine IL-10 and TGF-β. Notably, addition of conditioned medium (CM) from modSLA-induced DCs significantly increased the mRNA expression of Runx2 and ALP as well as ALP activity by murine preosteoblast MC3T3-E1 cells. Our data demonstrated that Ti disks with different surfaces lead to differential DCs responses. PT and SLA surfaces induce DCs mature, while DCs seeded on modSLA-Ti surface maintain an immature phenotype and exhibit a potential of promoting osteogenic differentiation of MC3T3-E1 cells.

Surface biofunctionalization of three-dimensional porous poly(lactic acid) scaffold using chitosan/OGP coating for bone tissue engineering.

PubMed

Zeng, Sen; Ye, Jianhua; Cui, Zhixiang; Si, Junhui; Wang, Qianting; Wang, Xiaofeng; Peng, Kaiping; Chen, Wenzhe

2017-08-01

As one of the stimulators on bone formation, osteogenic growth peptide (OGP) improves both proliferation and differentiation of the bone cells in vitro and in vivo. The aim of this work was the preparation of three dimensional porous poly(lactic acid) (PLA) scaffold with high porosity from PLA-dioxane-water ternary system with the use of vacuum-assisted solvent casting, phase separation, solvent extraction and particle leaching methods. Then, by surface coating of PLA scaffold with chitosan (CS)/OGP solution, biofunctionalization of PLA scaffold had been completed for application in bone regeneration. The effects of frozen temperature (-20, -50, -80°C) and PLA solution concentration (10, 12, 14wt%) on the microstructure, water absorption, porosity, hydrophilicity, mechanical properties, and biocompatibility of PLA and CS/OGP/PLA scaffold were investigated. Results showed that both PLA and CS/OGP/PLA scaffolds have an interconnected network structure and a porosity of up to 96.1% and 91.5%, respectively. The CS/OGP/PLA scaffold exhibited better hydrophilicity and mechanical properties than that of uncoated PLA scaffold. Moreover, the results of cell culture test showed that CS/OGP coating could stimulate the proliferation and growth of osteoblast cells on CS/OGP/PLA scaffold. These finding suggested that the surface biofunctionalization by CS/OGP coating layer could be an effective method on enhancing cell adhesion to synthetic polymer-based scaffolds in tissue engineering application and the developed porous CS/OGP/PLA scaffold should be considered as alternative biomaterials for bone regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

Magnesium ion implantation on a micro/nanostructured titanium surface promotes its bioactivity and osteogenic differentiation function

PubMed Central

Wang, Guifang; Li, Jinhua; Zhang, Wenjie; Xu, Lianyi; Pan, Hongya; Wen, Jin; Wu, Qianju; She, Wenjun; Jiao, Ting; Liu, Xuanyong; Jiang, Xinquan

2014-01-01

As one of the important ions associated with bone osseointegration, magnesium was incorporated into a micro/nanostructured titanium surface using a magnesium plasma immersion ion-implantation method. Hierarchical hybrid micro/nanostructured titanium surfaces followed by magnesium ion implantation for 30 minutes (Mg30) and hierarchical hybrid micro/nanostructured titanium surfaces followed by magnesium ion implantation for 60 minutes (Mg60) were used as test groups. The surface morphology, chemical properties, and amount of magnesium ions released were evaluated by field-emission scanning electron microscopy, energy dispersive X-ray spectroscopy, field-emission transmission electron microscopy, and inductively coupled plasma-optical emission spectrometry. Rat bone marrow mesenchymal stem cells (rBMMSCs) were used to evaluate cell responses, including proliferation, spreading, and osteogenic differentiation on the surface of the material or in their medium extraction. Greater increases in the spreading and proliferation ability of rBMMSCs were observed on the surfaces of magnesium-implanted micro/nanostructures compared with the control plates. Furthermore, the osteocalcin (OCN), osteopontin (OPN), and alkaline phosphatase (ALP) genes were upregulated on both surfaces and in their medium extractions. The enhanced cell responses were correlated with increasing concentrations of magnesium ions, indicating that the osteoblastic differentiation of rBMMSCs was stimulated through the magnesium ion function. The magnesium ion-implanted micro/nanostructured titanium surfaces could enhance the proliferation, spreading, and osteogenic differentiation activity of rBMMSCs, suggesting they have potential application in improving bone-titanium integration. PMID:24940056

Reactions and Surface Transformations of a Bone-Bioactive Material in a Simulated Microgravity Environment

NASA Technical Reports Server (NTRS)

Radin, S.; Ducheyne, P.; Ayyaswamy, P. S.

1999-01-01

A comprehensive program to investigate the expeditious in vitro formation of three-dimensional bone-like tissue is currently underway at the University of Pennsylvania. The study reported here forms a part of that program. Three-dimensional bone-like tissue structures may be grown under the simulated microgravity conditions of NASA designed Rotating Wall Bioreactor Vessels (RWV's). Such tissue growth will have wide clinical applications. In addition, an understanding of the fundamental changes that occur to bone cells under simulated microgravity would yield important information that will help in preventing or minimizing astronaut bone loss, a major health issue with travel or stay in space over long periods of time. The growth of three-dimensional bone-like tissue structures in RWV's is facilitated by the use of microcarriers which provide structural support. If the microcarrier material additionally promotes bone cell growth, then it is particularly advantageous to employ such microcarriers. We have found that reactive, bone-bioactive glass (BBG) is an attractive candidate for use as microcarrier material. Specifically, it has been found that BBG containing Ca- and P- oxides upregulates osteoprogenitor cells to osteoblasts. This effect on cells is preceded by BBG reactions in solution which result in the formation of a Ca-P surface layer. This surface further transforms to a bone-like mineral (i.e., carbonated crystalline hydroxyapatite (c-HA)). At normal gravity, time-dependent, immersion-induced BBG reactions and transformations are greatly affected both by variations in the composition of the milieu in which the glass is immersed and on the immersion conditions. However, the nature of BBG reactions and phase transformations under the simulated microgravity conditions of RWV's are unknown, and must be understood in order to successfully use BBG as microcarrier material in RWV'S. In this paper, we report some of our recent findings in this regard using experimental and numerical methods. BBG composition 45S5, the most reactive among known bone-bioactive glasses, was chosen for the study. BBG 45S5 behavior in physiological solutions was tested in simulated microgravity and compared with that at normal gravity. On the basis of our numerical study, we have chosen the BBG granule size to be in the range 40-70 microns, and a RWV rotational speed of 10 rpm. Our numerical study has shown that these parameters enable the microcarrier to remain suspended in the medium without experiencing collisions with the wall of the vessel. Immersion-induced changes in the solution composition and the material surface were analyzed after immersion.

A novel three-dimensional bone chip organ culture.

PubMed

Kuttenberger, Johannes; Polska, Elzbieta; Schaefer, Birgit M

2013-07-01

The objective of this study was to develop a 3D bone chip organ culture model. We aimed to collect in vitro evidence of the ability of vital bone chips to promote new bone formation. We developed a 3D in vitro hypoxic bone chip organ culture model. Histology of the bone chips was performed before and after culture and immunohistochemistry after 3-week culture. The 3D culture supernatants were tested for the presence of pro-angiogenic growth factors, TGFβ1, GADPH, bone alkaline phosphatase, osteocalcin, osteonectin, osteopontin, bone sialoprotein and collagen type I. Histology after culture revealed bone chips in a matrix of fibrin remnants and a fibrous-appearing matter. Collagen type I- and IV-positive structures were also identified. Cells could be seen on the surface of the bone chips, with spindle-shaped cells bridging the bone chip particles. Pro-angiogenic growth factors and TGFβ1were detected in the 3D cell culture supernatants. The transcripts for osteocalcin, bone sialoprotein and collagen type I were revealed only via PCR. Our results indicate that bone chips in our 3D organ culture remain vital and may stimulate the growth of a bone-forming matrix. The use of autogenous bone chips for oral and maxillofacial bone augmentation procedures is widespread in clinical practice. The rationale for this is that if bone chips remain vital in vivo, they could provide an environment promoting new bone formation through growth factors and cells. This 3D culture method is an essential tool for investigating the behaviour of bone chips.

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.

Surface modifications of dental implants.

PubMed

Stanford, C M

2008-06-01

Dental implant surface technologies have been evolving rapidly to enhance a more rapid bone formation on their surface and hold a potential to increase the predictability of expedited implant therapy. While implant outcomes have become highly predictable, there are sites and conditions that result in elevated implant loss. This paper reviews the impact of macro-retentive features which includes approaches to surface oxide modification, thread design, press-fit and sintered-bead technologies to increase predictability of outcomes. Implant designs that lead to controlled lateral compression of the bone can improve primary stability as long as the stress does not exceed the localized yield strength of the cortical bone. Some implant designs have reduced crestal bone loss by use of multiple cutting threads that are closely spaced, smoothed on the tip but designed to create a hoop-stress stability of the implant as it is completely seated in the osteotomy. Following the placement of the implant, there is a predictable sequence of bone turnover and replacement at the interface that allows the newly formed bone to adapt to microscopic roughness on the implant surface, and on some surfaces, a nanotopography (<10(-9) m scale) that has been shown to preferably influence the formation of bone. Newly emerging studies show that bone cells are exquisitely sensitive to these topographical features and will upregulate the expression of bone related genes for new bone formation when grown on these surfaces. We live in an exciting time of rapid changes in the modalities we can offer patients for tooth replacement therapy. Given this, it is our responsibility to be critical when claims are made, incorporate into our practice what is proven and worthwhile, and to continue to support and provide the best patient care possible.

RGD-conjugated rod-like viral nanoparticles on 2D scaffold improved bone differentiation of mesenchymal stem cells

NASA Astrophysics Data System (ADS)

Wang, Qian; Pongkwan, Sitasuwan; Lee, L.; Li, Kai; Nguyen, Huong

2014-05-01

Viral nanoparticles have uniform and well-defined nano-structures and can be produced in large quantities. Several plant viral nanoparticles have been tested in biomedical applications due to the lack of mammalian cell infectivity. We are particularly interested in using Tobacco mosaic virus (TMV), which has been demonstrated to enhance bone tissue regeneration, as a tuneable nanoscale building block for biomaterials development. Unmodified TMV particles have been shown to accelerate osteogenic differentiation of adult stem cells by synergistically upregulating BMP2 and IBSP expression with dexamethasone. However, the lack of affinity to mammalian cell surface resulted in low initial cell adhesion. In this study, to increase cell binding capacity of TMV based material the chemical functionalization of TMV with arginine-glycine-aspartic acid (RGD) peptide was explored. An azide-derivatized RGD peptide was “clicked” to tyrosine residues on TMV outer surface via an efficient copper(I) catalysed azide-alkyne cycloaddition reaction. The ligand spacing is calculated to be 2-4 nm, which could offer a polyvalent ligand clustering effect for enhanced cell receptor signalling, further promoting the proliferation and osteogenic differentiation of bone marrow derived mesenchymal stem cells.

Betulinic acid, a bioactive pentacyclic triterpenoid, inhibits skeletal-related events induced by breast cancer bone metastases and treatment

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

Park, Se Young; Kim, Hyun-Jeong; Kim, Ki Rim

Many breast cancer patients experience bone metastases and suffer skeletal complications. The present study provides evidence on the protective and therapeutic potential of betulinic acid on cancer-associated bone diseases. Betulinic acid is a naturally occurring triterpenoid with the beneficial activity to limit the progression and severity of cancer, diabetes, cardiovascular diseases, atherosclerosis, and obesity. We first investigated its effect on breast cancer cells, osteoblastic cells, and osteoclasts in the vicious cycle of osteolytic bone metastasis. Betulinic acid reduced cell viability and the production of parathyroid hormone-related protein (PTHrP), a major osteolytic factor, in MDA-MB-231 human metastatic breast cancer cells stimulatedmore » with or without tumor growth factor-β. Betulinic acid blocked an increase in the receptor activator of nuclear factor-kappa B ligand (RANKL)/osteoprotegerin ratio by downregulating RANKL protein expression in PTHrP-treated human osteoblastic cells. In addition, betulinic acid inhibited RANKL-induced osteoclastogenesis in murine bone marrow macrophages and decreased the production of resorbed area in plates with a bone biomimetic synthetic surface by suppressing the secretion of matrix metalloproteinase (MMP)-2, MMP-9, and cathepsin K in RANKL-induced osteoclasts. Furthermore, oral administration of betulinic acid inhibited bone loss in mice intra-tibially inoculated with breast cancer cells and in ovariectomized mice causing estrogen deprivation, as supported by the restored bone morphometric parameters and serum bone turnover markers. Taken together, these findings suggest that betulinic acid may have the potential to prevent bone loss in patients with bone metastases and cancer treatment-induced estrogen deficiency. - Highlights: • Betulinic acid reduced PTHrP production in human metastatic breast cancer cells. • Betulinic acid blocked RANKL/OPG ratio in PTHrP-stimulated human osteoblastic cells. • Betulinic acid inhibited RANKL-induced osteoclastogenesis in bone marrow macrophages. • Betulinic acid decreased bone resorption by suppressing osteoclast activity. • Orally administered betulinic acid inhibited cancer-associated bone diseases in mice.« less

In vitro adhesion of fibroblastic cells to titanium alloy discs treated with sodium hydroxide.

PubMed

Al Mustafa, Maisa; Agis, Hermann; Müller, Heinz-Dieter; Watzek, Georg; Gruber, Reinhard

2015-01-01

Adhesion of osteogenic cells on titanium surfaces is a prerequisite for osseointegration. Alkali treatment can increase the hydrophilicity of titanium implant surfaces, thereby supporting the adhesion of blood components. However, it is unclear if alkali treatment also supports the adhesion of cells with a fibroblastic morphology to titanium. Here, we have used a titanium alloy (Ti-6AL-4V) processed by alkali treatment to demonstrate the impact of hydrophilicity on the adhesion of primary human gingival fibroblast and bone cells. Also included were the osteosarcoma and fibroblastoma cell lines, MG63 and L929, respectively. Cell adhesion was determined by scanning electron microscopy. We also measured viability, proliferation, and protein synthesis of the adherent cells. Alkali treatment increased the adhesion of gingival fibroblasts, bone cells, and the two cell lines when seeded onto the titanium alloy surface for 1 h. At 3 h, no significant changes in cell adhesion were observed. Cells grown for 1 day on the titanium alloy surfaces processed by alkali treatment behave similarly to untreated controls with regard to viability, proliferation, and protein synthesis. Based on these preliminary In vitro findings, we conclude that alkali treatment can support the early adhesion of cells with fibroblastic characteristics to a titanium alloy surface. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Surface microstructuring of biocompatible bone analogue material HAPEX using LIGA technique and embossing

NASA Astrophysics Data System (ADS)

Schneider, Andreas; Rea, Susan; Huq, Ejaz; Bonfield, William

2003-04-01

HAPEX is an artificial bone analogue composite based on hydroxyapatite and polyethylene, which can be applied for growth of bone cells. Due to its biocompatibility and favourable mechanical properties, HAPEX is used for orthopaedic implants like tympanic (middle ear) bones. The morphology of HAPEX surfaces is of high interest and it is believed that surface structuring on a micron scale might improve the growth conditions for bone cells. A new and simple approach for the microstructuring of HAPEX surfaces has been investigated using LIGA technique. LIGA is a combination of several processes, in particular lithography, electroplating and forming/moulding. For HAPEX surface structuring, arrays of dots, grids and lines with typical lateral dimension ranging from 5 μm to 50 μm were created on a chromium photomask and the patterns were transferred into thick SU-8 photoresist (structure height > 10 μm) by UV lithography. Subsequently, the SU-8 structures served as moulds for electroplating nickel on Si wafers and nickel substrates. The final nickel microstructures were used as embossing master for the HAPEX material. Embossing was carried out using a conventional press (> 500 hPa) with the facility to heat the master and the HAPEX. The temperature ranged from ambient to a few degrees above glass transition temperature (Tg) of HAPEX. The paper will include details of the fabrication process and process tolerances in lateral and vertical directions. Data obtained are correlated to the temperature used during embossing.

Nanostructured magnesium increases bone cell density.

PubMed

Weng, Lucy; Webster, Thomas J

2012-12-07

Magnesium has attracted some attention in orthopedics due to its biodegradability and mechanical properties. Since magnesium is an essential natural mineral for bone growth, it can be expected that as a biomaterial, it would support bone formation. However, upon degradation in the body, magnesium releases OH(-) which results in an alkaline pH that can be detrimental to cell density (for example, osteoblasts or bone forming cells). For this reason, modification of magnesium may be necessary to compensate for such detrimental effects to cells. This study created biologically inspired nanoscale surface features on magnesium by soaking magnesium in various concentrations of NaOH (from 1 to 10 N) and for various periods of time (from 10 to 30 min). The results provided the first evidence of increased roughness, surface energy, and consequently greater osteoblast adhesion, after 4 h as well as density up to 7 days on magnesium treated with any concentration of NaOH for any length of time compared to untreated controls. For these reasons, this study suggests that soaking magnesium in NaOH could be an inexpensive, simple and effective manner to promote osteoblast functions for numerous orthopedic applications and, thus, should be further studied.

Bioactive scaffold for bone tissue engineering: An in vivo study

NASA Astrophysics Data System (ADS)

Livingston, Treena Lynne

Massive bone loss of the proximal femur is a common problem in revision cases of total hip implants. Allograft is typically used to reconstruct the site for insertion of the new prosthesis. However, for long term fixation and function, it is desirable that the allograft becomes fully replaced by bone tissue and aids in the regeneration of bone to that site. However, allograft use is typically associated with delayed incorporation and poor remodeling. Due to these profound limitations, alternative approaches are needed. Tissue engineering is an attractive approach to designing improved graft materials. By combining osteogenic activity with a resorbable scaffold, bone formation can be stimulated while providing structure and stability to the limb during incorporation and remodeling of the scaffold. Porous, surface modified bioactive ceramic scaffolds (pSMC) have been developed which stimulate the expression of the osteoblastic phenotype and production of bone-like tissue in vitro. The scaffold and two tissue-engineered constructs, osteoprogenitor cells seeded onto scaffolds or cells expanded in culture to form bone tissue on the scaffolds prior to implantation, were investigated in a long bone defect model. The rate of incorporation was assessed. Both tissue-engineered constructs stimulated bone formation and comparable repair at 2 weeks. In a rat femoral window defect model, bone formation increased over time for all groups in concert with scaffold resorption, leading to a 40% increase in bone and 40% reduction of the scaffold in the defect by 12 weeks. Both tissue-engineered constructs enhanced the rate of mechanical repair of long bones due to better bony union with the host cortex. Long bones treated with tissue engineered constructs demonstrated a return in normal torsional properties by 4 weeks as compared to 12 weeks for long bones treated with pSMC. Culture expansion of cells to produce bone tissue in vitro did not accelerate incorporation over the treatment with cells seeded at the time of surgery. Porous, surface modified bioactive ceramic is a promising scaffold material for tissue-engineered bone repair. Bone formation and scaffold resorption act in concert for maintenance and improvement of the structural properties of the long bones over time. As determined histomorphometrically and mechanically, the rate of incorporation of the scaffold was enhanced with the tissue-engineered constructs.

Effect of Calcium-Infiltrated Hydroxyapatite Scaffolds on the Hematopoietic Fate of Human Umbilical Vein Endothelial Cells.

PubMed

Zhang, Qinghao; Gerlach, Jörg C; Schmelzer, Eva; Nettleship, Ian

2017-01-01

Foamed hydroxyapatite offers a three-dimensional scaffold for the development of bone constructs, mimicking perfectly the in vivo bone structure. In vivo, calcium release at the surface is assumed to provide a locally increased gradient supporting the maintenance of the hematopoietic stem cells niche. We fabricated hydroxyapatite scaffolds with high surface calcium concentration by infiltration, and used human umbilical vein endothelial cells (HUVECs) as a model to study the effects on hematopoietic lineage direction. HUVECs are umbilical vein-derived and thus possess progenitor characteristics, with a prospective potential to give rise to hematopoietic lineages. HUVECs were cultured for long term on three-dimensional porous hydroxyapatite scaffolds, which were either infiltrated biphasic foams or untreated. Controls were cultured in two-dimensional dishes. The release of calcium into culture medium was determined, and cells were analyzed for typical hematopoietic and endothelial gene expressions, surface markers by flow cytometry, and hematopoietic potential using colony-forming unit assays. Our results indicate that the biphasic foams promoted a hematopoietic lineage direction of HUVECs, suggesting an improved in vivo-like scaffold for hematopoietic bone tissue engineering. © 2017 S. Karger AG, Basel.

Fabrication and Characterization of Carbon Fiber-Reinforced Nano-Hydroxyapatite/Polyamide46 Biocomposite for Bone Substitute.

PubMed

Deng, Zhennan; Han, Hongjuan; Yang, Jingyuan; Li, Yuanyuan; Du, Shengnan; Ma, Jianfeng

2017-05-24

BACKGROUND Ideal bone repair material should be of good biocompatibility and high bioactivity. Besides, their mechanical properties should be equivalent to those of natural bone. The objective of this study was to fabricate a novel biocomposite suitable for load-bearing bone defect repair. MATERIAL AND METHODS A novel biocomposite composed of carbon fiber, hydroxyapatite and polyamide46 (CF/HA/PA46) was fabricated, and its mechanical performances and preliminary cell responses were evaluated to explore its feasibility for load-bearing bone defect repair. RESULTS The resultant CF/HA/PA46 biocomposite showed a bending strength of 159-223 MPa, a tensile strength of 127-199 MPa and a tensile modulus of 7.7-10.8 GPa, when the CF content was 5-20% (mass fraction) in biocomposite. The MG63 cells, showing an osteogenic phenotype, were well adhered and spread on the surface of the CF/HA/PA46 biocomposite. Moreover, the cells vitality and differentiation on the CF/HA/PA46 biocomposite surface were obviously increased during the culture time and there was no significant difference between the CF/HA/PA46 biocomposite and HA/PA (as control) at all the experimental time (P>0.05). CONCLUSIONS The addition of CF into HA/PA46 composite manifest improved the mechanical performances and showed favorable effects on biocompatibility of MG63 cells. The obtained biocomposite has high potential for bone repair in load-bearing sites.

Fabrication and Characterization of Carbon Fiber-Reinforced Nano-Hydroxyapatite/Polyamide46 Biocomposite for Bone Substitute

PubMed Central

Deng, Zhennan; Han, Hongjuan; Yang, Jingyuan; Li, Yuanyuan; Du, Shengnan; Ma, Jianfeng

2017-01-01

Background Ideal bone repair material should be of good biocompatibility and high bioactivity. Besides, their mechanical properties should be equivalent to those of natural bone. The objective of this study was to fabricate a novel biocomposite suitable for load-bearing bone defect repair. Material/Methods A novel biocomposite composed of carbon fiber, hydroxyapatite and polyamide46 (CF/HA/PA46) was fabricated, and its mechanical performances and preliminary cell responses were evaluated to explore its feasibility for load-bearing bone defect repair. Results The resultant CF/HA/PA46 biocomposite showed a bending strength of 159–223 MPa, a tensile strength of 127–199 MPa and a tensile modulus of 7.7–10.8 GPa, when the CF content was 5–20% (mass fraction) in biocomposite. The MG63 cells, showing an osteogenic phenotype, were well adhered and spread on the surface of the CF/HA/PA46 biocomposite. Moreover, the cells vitality and differentiation on the CF/HA/PA46 biocomposite surface were obviously increased during the culture time and there was no significant difference between the CF/HA/PA46 biocomposite and HA/PA (as control) at all the experimental time (P>0.05). Conclusions The addition of CF into HA/PA46 composite manifest improved the mechanical performances and showed favorable effects on biocompatibility of MG63 cells. The obtained biocomposite has high potential for bone repair in load-bearing sites. PMID:28536416

Association of murine lupus and thymic full-length endogenous retroviral expression maps to a bone marrow stem cell

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

Krieg, A.M.; Gourley, M.F.; Steinberg, A.D.

1991-05-01

Recent studies of thymic gene expression in murine lupus have demonstrated 8.4-kb (full-length size) modified polytropic (Mpmv) endogenous retroviral RNA. In contrast, normal control mouse strains do not produce detectable amounts of such RNA in their thymuses. Prior studies have attributed a defect in experimental tolerance in murine lupus to a bone marrow stem cell rather than to the thymic epithelium; in contrast, infectious retroviral expression has been associated with the thymic epithelium, rather than with the bone marrow stem cell. The present study was designed to determine whether the abnormal Mpmv expression associated with murine lupus mapped to thymicmore » epithelium or to a marrow precursor. Lethally irradiated control and lupus-prone mice were reconstituted with T cell depleted bone marrow; one month later their thymuses were studied for endogenous retroviral RNA and protein expression. Recipients of bone marrow from nonautoimmune donors expressed neither 8.4-kb Mpmv RNA nor surface MCF gp70 in their thymuses. In contrast, recipients of bone marrow from autoimmune NZB or BXSB donors expressed thymic 8.4-kb Mpmv RNA and mink cell focus-forming gp70. These studies demonstrate that lupus-associated 8.4-kb Mpmv endogenous retroviral expression is determined by bone marrow stem cells.« less

Bone regeneration: molecular and cellular interactions with calcium phosphate ceramics

PubMed Central

Barrère, Florence; van Blitterswijk, Clemens A; de Groot, Klaas

2006-01-01

Calcium phosphate bioceramics are widely used in orthopedic and dental applications and porous scaffolds made of them are serious candidates in the field of bone tissue engineering. They have superior properties for the stimulation of bone formation and bone bonding, both related to the specific interactions of their surface with the extracellular fluids and cells, ie, ionic exchanges, superficial molecular rearrangement and cellular activity. PMID:17717972

The Effect of Simulated Microgravity Environment of RWV Bioreactors on Surface Reactions and Adsorption of Serum Proteins on Bone-bioactive Microcarriers

NASA Technical Reports Server (NTRS)

Radin, Shula; Ducheyne, P.; Ayyaswamy, P. S.

2003-01-01

Biomimetically modified bioactive materials with bone-like surface properties are attractive candidates for use as microcarriers for 3-D bone-like tissue engineering under simulated microgravity conditions of NASA designed rotating wall vessel (RWV) bioreactors. The simulated microgravity environment is attainable under suitable parametric conditions of the RWV bioreactors. Ca-P containing bioactive glass (BG), whose stimulatory effect on bone cell function had been previously demonstrated, was used in the present study. BG surface modification via reactions in solution, resulting formation of bone-like minerals at the surface and adsorption of serum proteins is critical for obtaining the stimulatory effect. In this paper, we report on the major effects of simulated microgravity conditions of the RWV on the BG reactions surface reactions and protein adsorption in physiological solutions. Control tests at normal gravity were conducted at static and dynamic conditions. The study revealed that simulated microgravity remarkably enhanced reactions involved in the BG surface modification, including BG dissolution, formation of bone-like minerals at the surface and adsorption of serum proteins. Simultaneously, numerical models were developed to simulate the mass transport of chemical species to and from the BG surface under normal gravity and simulated microgravity conditions. The numerical results showed an excellent agreement with the experimental data at both testing conditions.

Double-layered cell transfer technology for bone regeneration

PubMed Central

Akazawa, Keiko; Iwasaki, Kengo; Nagata, Mizuki; Yokoyama, Naoki; Ayame, Hirohito; Yamaki, Kazumasa; Tanaka, Yuichi; Honda, Izumi; Morioka, Chikako; Kimura, Tsuyoshi; Komaki, Motohiro; Kishida, Akio; Izumi, Yuichi; Morita, Ikuo

2016-01-01

For cell-based medicine, to mimic in vivo cellular localization, various tissue engineering approaches have been studied to obtain a desirable arrangement of cells on scaffold materials. We have developed a novel method of cell manipulation called “cell transfer technology”, enabling the transfer of cultured cells onto scaffold materials, and controlling cell topology. Here we show that using this technique, two different cell types can be transferred onto a scaffold surface as stable double layers or in patterned arrangements. Various combinations of adherent cells were transferred to a scaffold, amniotic membrane, in overlapping bilayers (double-layered cell transfer), and transferred cells showed stability upon deformations of the material including folding and trimming. Transplantation of mesenchymal stem cells from periodontal ligaments (PDLSC) and osteoblasts, using double-layered cell transfer significantly enhanced bone formation, when compared to single cell type transplantation. Our findings suggest that this double-layer cell transfer is useful to produce a cell transplantation material that can bear two cell layers. Moreover, the transplantation of an amniotic membrane with PDLSCs/osteoblasts by cell transfer technology has therapeutic potential for bone defects. We conclude that cell transfer technology provides a novel and unique cell transplantation method for bone regeneration. PMID:27624174

Leukemia-associated antigens in man.

PubMed

Brown, G; Capellaro, D; Greaves, M

1975-12-01

Rabbit antisera raised against acute lymphoblastic leukemia (ALL) cells were used to distinguish ALL from other leukemias, to identify rare leukemia cells in the bone marrow of patients in remission, and to define human leukemia-associated antigens. Antibody binding was studied with the use of immunofluorescence reagents and the analytic capacity of the Fluorescence Activated Cell Sorter-1 (FACS-1). The results indicated that most non-T-cell ALL have three leukemia-associated antigens on their surface which are absent from normal lymphoid cells: 1) an antigen shared with myelocytes, myeloblastic leukemia cells, and fetal liver (hematopoietic) cells; 2) an antigen shared with a subset of intermediate normoblasts in normal bone marrow and fetal liver; and 3) an antigen found thus far only on non-T-cell ALL and in some acute undifferentiated leukemias, which we therefore regard as a strong candidate for a leukemia-specific antigen. These antigens are absent from a subgroup of ALL patients in which the lymphoblasta express T-cell surface markers. Preliminary studies on the bone marrow samples of patients in remission indicated that rare leukemia cells were present in some samples. The implications of these findings with respect to the heterogeneity and cell origin(s) of ALL, its diagnosis, and its potential monitoring during treatment were discussed.

Surface nitridation improves bone cell response to melt-derived bioactive silicate/borosilicate glass composite scaffolds.

PubMed

Orgaz, Felipe; Dzika, Alexandra; Szycht, Olga; Amat, Daniel; Barba, Flora; Becerra, José; Santos-Ruiz, Leonor

2016-01-01

Novel bioactive amorphous glass-glass composite scaffolds (ICIE16/BSG) with interconnected porosity have been developed. Hierarchically interconnected porous glass scaffolds were prepared from a mixture of two melt-derived glasses: a ICIE16 bioactive glass that was previously developed by Wu et al. (2011) to prevent crystallization, and a borosilicate glass of composition 73.48 SiO2-11.35 B2O3-15.15 Na2O (wt%). The resulting melt derived glass-glass composite scaffolds (ICIE16/BSG) were subject to surface functionalization to further improve its interaction with biological systems. Surface functionalization was performed by a nitridation process with hot gas N2/ammonia at 550°C for 2h, obtaining the ICIE16/BSG-NITRI. Evaluation of the degradation rate and the conversion to hydroxyapatite after immersion in simulated body fluid predicted a good biological activity of all the scaffolds, but particularly of the nitrided ones. In vitro evaluation of osteoblastic cells cultured onto the nitrided and non-nitrided scaffolds showed cell attachment, proliferation and differentiation on all scaffolds, but both proliferation and differentiation were improved in the nitrided ICIE16/BSG-NITRI. Biomaterials are often required in the clinic to stimulate bone repair. We have developed a novel bioglass (ICIE16/SBG-NITRI) that can be sintered into highly porous 3D scaffolds, and we have further improved its bioactivity by nitridation. ICIE16/SBG-NITRI was synthesized from a mixture of two melt-derived glasses through combined gel casting and foam replication techniques, followed by nitridation. To mimic bone, it presents high-interconnected porosity while being mechanically stable. Nitridation improved its reactivity and bioactivity facilitating its resorption and the deposition of apatite (bone-like mineral) on its surface and increasing its degradation rate. The nitrided surface also improved the bioglass' interaction with bone cells, which were found to attach better to ICIE16/SBG-NITRI and to differentiate earlier on its surface. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Cosmos: 1989 immunology studies

NASA Technical Reports Server (NTRS)

Sonnenfeld, Gerald

1991-01-01

The effects of flight on Cosmos mission 2044 on leukocyte subset distribution and the sensitivity of bone marrow cells to colony stimulating factor-GM were determined. A parallel study with antiorthostatic suspension was also carried out. The study involved repetition and expansion of studies performed on Cosmos 1887. Spleen and bone marrow cells were obtained from flown, vivarium control, synchronous control, and suspended rats. The cells were stained with a series of monoclonal antibodies directed against rat leukocyte cell surface antigens. Control cells were stained with a monoclonal antibody directed against an irrelevant species or were unstained. Cells were then analyzed for fluorescence using a FACSCAN flow cytometer. Bone marrow cells were placed in culture with GM-CSF in McCoy's 5a medium and incubated for 5 days. Cultures were then evaluated for the number of colonies of 50 cells or greater.

In-vitro biocompatibility, bioactivity, and mechanical strength of PMMA-PCL polymer containing fluorapatite and graphene oxide bone cements.

PubMed

Pahlevanzadeh, F; Bakhsheshi-Rad, H R; Hamzah, E

2018-06-01

In this study, a bone cement consisting of poly methyl methacrylate (PMMA)-poly caprolactone (PCL)-fluorapatite (FA)-graphene oxide (GO) was synthesized as bone filler for application in orthopedic surgeries. The FA and GO particulates were homogenously distributed in the PMMA-PCL polymer matrix and no defects and agglomeration were found in the PMMA-PCL/FA/GO bone cement. The in-vitro bioactivity result exhibited that addition of FA and GO to the polymer cement (PMMA-PCL) improved the apatite formation ability on the surface of polymer. The results also showed that addition of FA to the polymer bone cement escalated the compressive strength and elastic modulus while reducing elongation to 8 ± 2%. However, after addition of GO into the PMMA-PCL/FA bone cement, both compressive strength and elongation considerably increased to 101 ± 5 MPa and 35 ± 6%, respectively. Furthermore, tensile tests exhibited that inclusion of GO was favorable in improving the tensile modulus, UTS and elongation of the PMMA-PCL/FA bone cement. The cytotoxicity test pointed out that MG63 osteoblast cells viability increased to 279 ± 15% after addition of FA and GO to the PMMA-PCL polymer bone cement. The DAPI (4',6-diamidino-2-phenylindole) staining demonstrated better spreading and attachment of MG63 cells on PMMA-PCL/FA/GO surface compared to the PMMA-PCL bone cements. These results confirm the suitable mechanical properties and favorable bioactivity along with high cells viability of PMMA-PCL/FA/GO bone cement, indicating its potentials for orthopedic applications. Copyright © 2018 Elsevier Ltd. All rights reserved.

Biomaterial strategies for engineering implants for enhanced osseointegration and bone repair

PubMed Central

Agarwal, Rachit; García, Andrés J.

2015-01-01

Bone tissue has a remarkable ability to regenerate and heal itself. However, large bone defects and complex fractures still present a significant challenge to the medical community. Current treatments center on metal implants for structural and mechanical support and auto- or allo-grafts to substitute long bone defects. Metal implants are associated with several complications such as implant loosening and infections. Bone grafts suffer from donor site morbidity, reduced bioactivity, and risk of pathogen transmission. Surgical implants can be modified to provide vital biological cues, growth factors and cells in order to improve osseointegration and repair of bone defects. Here we review strategies and technologies to engineer metal surfaces to promote osseointegration with the host tissue. We also discuss strategies for modifying implants for cell adhesion and bone growth via integrin signaling and growth factor and cytokine delivery for bone defect repair. PMID:25861724

Differential responses of osteoblast lineage cells to nanotopographically-modified, microroughened titanium-aluminum-vanadium alloy surfaces.

PubMed

Gittens, Rolando A; Olivares-Navarrete, Rene; McLachlan, Taylor; Cai, Ye; Hyzy, Sharon L; Schneider, Jennifer M; Schwartz, Zvi; Sandhage, Kenneth H; Boyan, Barbara D

2012-12-01

Surface structural modifications at the micrometer and nanometer scales have driven improved success rates of dental and orthopaedic implants by mimicking the hierarchical structure of bone. However, how initial osteoblast-lineage cells populating an implant surface respond to different hierarchical surface topographical cues remains to be elucidated, with bone marrow mesenchymal stem cells (MSCs) or immature osteoblasts as possible initial colonizers. Here we show that in the absence of any exogenous soluble factors, osteoblastic maturation of primary human osteoblasts (HOBs) but not osteoblastic differentiation of MSCs is strongly influenced by nanostructures superimposed onto a microrough Ti6Al4V (TiAlV) alloy. The sensitivity of osteoblasts to both surface microroughness and nanostructures led to a synergistic effect on maturation and local factor production. Osteoblastic differentiation of MSCs was sensitive to TiAlV surface microroughness with respect to production of differentiation markers, but no further enhancement was found when cultured on micro/nanostructured surfaces. Superposition of nanostructures to microroughened surfaces affected final MSC numbers and enhanced production of vascular endothelial growth factor (VEGF) but the magnitude of the response was lower than for HOB cultures. Our results suggest that the differentiation state of osteoblast-lineage cells determines the recognition of surface nanostructures and subsequent cell response, which has implications for clinical evaluation of new implant surface nanomodifications. Copyright © 2012 Elsevier Ltd. All rights reserved.

Differential Responses of Osteoblast Lineage Cells to Nanotopographically-Modified, Microroughened Titanium-Aluminum-Vanadium Alloy Surfaces

PubMed Central

Gittens, Rolando A.; Olivares-Navarrete, Rene; McLachlan, Taylor; Cai, Ye; Hyzy, Sharon L.; Schneider, Jennifer M.; Schwartz, Zvi; Sandhage, Kenneth H.; Boyan, Barbara D.

2013-01-01

Surface structural modifications at the micrometer and nanometer scales have driven improved success rates of dental and orthopaedic implants by mimicking the hierarchical structure of bone. However, how initial osteoblast-lineage cells populating an implant surface respond to different hierarchical surface topographical cues remains to be elucidated, with bone marrow mesenchymal stem cells (MSCs) or immature osteoblasts as possible initial colonizers. Here we show that in the absence of any exogenous soluble factors, osteoblastic maturation of primary human osteoblasts (HOBs) but not osteoblastic differentiation of MSCs is strongly influenced by nanostructures superimposed onto a microrough Ti6Al4V (TiAlV) alloy. The sensitivity of osteoblasts to both surface microroughness and nanostructures led to a synergistic effect on maturation and local factor production. Osteoblastic differentiation of MSCs was sensitive to TiAlV surface microroughness with respect to production of differentiation markers, but no further enhancement was found when cultured on micro/nanostructured surfaces. Superposition of nanostructures to microroughened surfaces affected final MSC numbers and enhanced production of vascular endothelial growth factor (VEGF) but the magnitude of the response was lower than for HOB cultures. Our results suggest that the differentiation state of osteoblast-lineage cells determines the recognition of surface nanostructures and subsequent cell response, which has implications for clinical evaluation of new implant surface nanomodifications. PMID:22989383

Immunohistochemical study of bone sialoprotein and osteopontin in healthy and diseased root surfaces.

PubMed

Lao, Martin; Marino, Victor; Bartold, P Mark

2006-10-01

Periodontal disease is marked by inflammation and damage to tooth-supporting tissues. In particular, damage occurs to factors present in cementum that are thought to have the ability to influence the regeneration of surrounding tissues. Bone sialoprotein and osteopontin are major non-collagenous proteins in mineralized connective tissues associated with precementoblast chemo-attraction, adhesion to the root surface, and cell differentiation. The purpose of this investigation was to determine whether the expression and distribution of bone sialoprotein and osteopontin on root surfaces affected by periodontitis are altered compared to healthy, non-diseased root surfaces. Thirty healthy and 30 periodontitis-affected teeth were collected. Following fixation and demineralization, specimens were embedded in paraffin, sectioned, and exposed to antibodies against bone sialoprotein and osteopontin. Stained sections were assessed using light microscopy. Bone sialoprotein was not detected in the exposed cementum (absence of overlying periodontal ligament) of diseased teeth. In most areas where the periodontal ligament was intact, bone sialoprotein was detected for healthy and diseased teeth. For teeth reactive for bone sialoprotein, the matrix of the cementum just below the periodontal ligament was moderately stained. A similar immunoreactivity pattern for osteopontin was observed. The absence of bone sialoprotein and osteopontin staining along exposed cementum surfaces may be due to structural and compositional changes in matrix components associated with periodontal disease. This may influence the ability for regeneration and new connective tissue attachment onto previously diseased root surfaces.

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.

Textural versus electrostatic exclusion-enrichment effects in the effective chemical transport within the cortical bone: a numerical investigation.

PubMed

Lemaire, T; Kaiser, J; Naili, S; Sansalone, V

2013-11-01

Interstitial fluid within bone tissue is known to govern the remodelling signals' expression. Bone fluid flow is generated by skeleton deformation during the daily activities. Due to the presence of charged surfaces in the bone porous matrix, the electrochemical phenomena occurring in the vicinity of mechanosensitive bone cells, the osteocytes, are key elements in the cellular communication. In this study, a multiscale model of interstitial fluid transport within bone tissues is proposed. Based on an asymptotic homogenization method, our modelling takes into account the physicochemical properties of bone tissue. Thanks to this multiphysical approach, the transport of nutrients and waste between the blood vessels and the bone cells can be quantified to better understand the mechanotransduction of bone remodelling. In particular, it is shown that the electrochemical tortuosity may have stronger implications in the mass transport within the bone than the purely morphological one. Copyright © 2013 John Wiley & Sons, Ltd.

Role of Cbl-PI3K Interaction during Skeletal Remodeling in a Murine Model of Bone Repair.

PubMed

Scanlon, Vanessa; Soung, Do Yu; Adapala, Naga Suresh; Morgan, Elise; Hansen, Marc F; Drissi, Hicham; Sanjay, Archana

2015-01-01

Mice in which Cbl is unable to bind PI3K (YF mice) display increased bone volume due to enhanced bone formation and repressed bone resorption during normal bone homeostasis. We investigated the effects of disrupted Cbl-PI3K interaction on fracture healing to determine whether this interaction has an effect on bone repair. Mid-diaphyseal femoral fractures induced in wild type (WT) and YF mice were temporally evaluated via micro-computed tomography scans, biomechanical testing, histological and histomorphometric analyses. Imaging analyses revealed no change in soft callus formation, increased bony callus formation, and delayed callus remodeling in YF mice compared to WT mice. Histomorphometric analyses showed significantly increased osteoblast surface per bone surface and osteoclast numbers in the calluses of YF fractured mice, as well as increased incorporation of dynamic bone labels. Furthermore, using laser capture micro-dissection of the fracture callus we found that cells lacking Cbl-PI3K interaction have higher expression of Osterix, TRAP, and Cathepsin K. We also found increased expression of genes involved in propagating PI3K signaling in cells isolated from the YF fracture callus, suggesting that the lack of Cbl-PI3K interaction perhaps results in enhanced PI3K signaling, leading to increased bone formation, but delayed remodeling in the healing femora.

A bone-resorption surface-targeting nanoparticle to deliver anti-miR214 for osteoporosis therapy

PubMed Central

Zhang, Shufan; Liu, Jiafan; Sun, Yao; Wang, Xiaogang

2017-01-01

With increasing fracture risks due to fragility, osteoporosis is a global health problem threatening postmenopausal women. In these patients, osteoclasts play leading roles in bone loss and fracture. How to inhibit osteoclast activity is the key issue for osteoporosis treatment. In recent years, miRNA-based gene therapy through gene regulation has been considered a potential therapeutic method. However, in light of the side effects, the use of therapeutic miRNAs in osteoporosis treatment is still limited by the lack of tissue/cell-specific delivery systems. Here, we developed polyurethane (PU) nanomicelles modified by the acidic peptide Asp8. Our data showed that without overt toxicity or eliciting an immune response, this delivery system encapsulated and selectively deliver miRNAs to OSCAR+ osteoclasts at bone-resorption surface in vivo. With the Asp8-PU delivery system, anti-miR214 was delivered to osteoclasts, and bone microarchitecture and bone mass were improved in ovariectomized osteoporosis mice. Therefore, Asp8-PU could be a useful bone-resorption surface-targeting delivery system for treatment of osteoclast-induced bone diseases and aging-related osteoporosis. PMID:29075114

Evidence against the mucosal traction theory in cholesteatoma.

PubMed

Pauna, Henrique F; Monsanto, Rafael C; Schachern, Patricia; Paparella, Michael M; Chole, Richard A; Cureoglu, Sebahattin

2017-10-08

To investigate the distribution of ciliated epithelium in the human middle ear and its potential role in the formation of cholesteatoma. Comparative human temporal bone study. We selected temporal bones from 14 donors with a diagnosis of cholesteatoma, 15 with chronic otitis media without retraction pockets, 14 with chronic otitis media with retraction pockets, 14 with cystic fibrosis (CF), and 16 controls. We mapped the distribution of the ciliated cells in the mucosal lining of the middle ear and tympanic membrane using three-dimensional reconstruction analysis, and counted the number of ciliated cells in the middle ear mucosa. Ciliated cells are extremely sparse in the epithelial lining of the lateral surface of the ossicles in the epitympanum and the medial surface of the tympanic membrane. Furthermore, there is a significant decrease in the number of ciliated cells in these areas in temporal bones with cholesteatoma, chronic otitis media, chronic otitis media with retraction pockets, and CF compared to controls. Ciliated cells most commonly are located at the hypotympanum and the Eustachian tube opening but not the tympanic membrane or epitympanum. The paucity of ciliated epithelial cells on the medial side of the tympanic membrane and the lateral surface of the ossicles in the epitympanum in cases with cholesteatoma and/or chronic otitis media do not support the mucosal migration theory of cholesteatoma formation. NA. Laryngoscope, 2017. © 2017 The American Laryngological, Rhinological and Otological Society, Inc.

Microstructured Titanium Regulates Interleukin Production by Osteoblasts, an Effect Modulated by Exogenous BMP-2

PubMed Central

Hyzy, Sharon; Olivares-Navarrete, Rene; Hutton, Daphne L.; Tan, Christian; Boyan, Barbara D.; Schwartz, Zvi

2013-01-01

Microtextured implant surfaces increase osteoblast differentiation in vitro and enhance bone-to-implant contact in vivo and clinically. These implants may be used in combination with recombinant human bone morphogenetic protein 2 (rhBMP-2) to enhance peri-implant bone formation. However, the effect of surface modifications alone or in combination with rhBMP-2 on osteoblast-produced inflammatory microenvironment is unknown. MG63 cells were cultured on tissue culture polystyrene or titanium substrates: smooth pretreated (PT, Ra=0.2μm), sandblasted/acid-etched (SLA, Ra=3.2μm), or hydrophilic-SLA (modSLA). Expression and protein production of pro-inflammatory interleukins (IL1b, IL6, IL8, IL17) and anti-inflammatory interleukins (IL10) were measured in cells with or without rhBMP-2. To determine which BMP signaling pathways were involved, cultures were incubated with BMP pathway inhibitors to blocking Smad (dorsomorphin), TAB/TAK1 ((5Z)-7-oxozeaenol), or PKA (H-8) signaling. Culture on rough SLA and modSLA surfaces decreased pro-inflammatory interleukins and increased anti-inflammatory IL10. This effect was negated in cells treated with rhBMP-2, which caused an increase in pro-inflammatory interleukins and a decrease in anti-inflammatory interleukins through TAB/TAK signaling. The results suggest that surface microtexture modulates the inflammatory process during osseointegration, an effect that may enhance healing. However, rhBMP-2 in combination with microtextured titanium implants can influence the effect of cells on these surfaces, and may adversely affect cells involved in osseointegration. PMID:23123301

The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status.

PubMed

Horváthová, Mira; Ilavská, Silvia; Štefíková, Kornélia; Szabová, Michaela; Krivošíková, Zora; Jahnová, Eva; Tulinská, Jana; Spustová, Viera; Gajdoš, Martin

2017-07-11

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs) were in postmenopausal women versus fertile women. Body mass index (BMI) affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells ( p < 0.05). The confounding factors such as women age, BMI, bone mineral density (BMD), waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause.

The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status

PubMed Central

Horváthová, Mira; Ilavská, Silvia; Štefíková, Kornélia; Szabová, Michaela; Krivošíková, Zora; Jahnová, Eva; Tulinská, Jana; Spustová, Viera; Gajdoš, Martin

2017-01-01

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs) were in postmenopausal women versus fertile women. Body mass index (BMI) affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells (p < 0.05). The confounding factors such as women age, BMI, bone mineral density (BMD), waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause. PMID:28696349

Identification of a constitutive law for trabecular bone samples under remodeling in the framework of irreversible thermodynamics

NASA Astrophysics Data System (ADS)

Louna, Zineeddine; Goda, Ibrahim; Ganghoffer, Jean-François

2018-01-01

We construct in the present paper constitutive models for bone remodeling based on micromechanical analyses at the scale of a representative unit cell (RUC) including a porous trabecular microstructure. The time evolution of the microstructure is simulated as a surface remodeling process by relating the surface growth remodeling velocity to a surface driving force incorporating a (surface) Eshelby tensor. Adopting the framework of irreversible thermodynamics, a 2D constitutive model based on the setting up of the free energy density and a dissipation potential is identified from FE simulations performed over a unit cell representative of the trabecular architecture obtained from real bone microstructures. The static and evolutive effective properties of bone at the scale of the RUC are obtained by combining a methodology for the evaluation of the average kinematic and static variables over a prototype unit cell and numerical simulations with controlled imposed first gradient rates. The formulated effective growth constitutive law at the scale of the homogenized set of trabeculae within the RUC is of viscoplastic type and relates the average growth strain rate to the homogenized stress tensor. The postulated model includes a power law function of an effective stress chosen to depend on the first and second stress invariants. The model coefficients are calibrated from a set of virtual testing performed over the RUC subjected to a sequence of loadings. Numerical simulations show that overall bone growth does not show any growth kinematic hardening. The obtained results quantify the strength and importance of different types of external loads (uniaxial tension, simple shear, and biaxial loading) on the overall remodeling process and the development of elastic deformations within the RUC.

Immobilization of cross linked Col-I-OPN bone matrix protein on aminolysed PCL surfaces enhances initial biocompatibility of human adipogenic mesenchymal stem cells (hADMSC)

NASA Astrophysics Data System (ADS)

Kim, Young-Hee; Jyoti, Md. Anirban; Song, Ho-Yeon

2014-06-01

In bone tissue engineering surface modification is considered as one of the important ways of fabricating successful biocompatible material. Addition of biologically active functionality on the surfaces has been tried for improving the overall biocompatibility of the system. In this study poly-ɛ-caprolactone film surfaces have been modified through aminolysis and immobilization process. Collagen type I (COL-I) and osteopontin (OPN), which play an important role in osteogenesis, was immobilized onto PCL films followed by aminolysis treatment using 1,6-hexanediamine. Characterization of animolysed and immobilized surfaces were done by a number techniques using scanning electron microscopy (SEM), FT-IR, XPS, ninhydrin staining, SDS-PAGE and confocal microscopy and compared between the modified and un-modified surfaces. Results of the successive experiments showed that aminolysis treatment was homogeneously achieved which helped to entrap or immobilize Col-I-OPN proteins on surfaces of PCL film. In vitro studies with human adipogenic mesenchymal stem cells (hADMSC) also confirmed the attachment and proliferation of cells was better in modified PCL surfaces than the unmodified surfaces. SEM, confocal microscopy and MTT assay showed a significant increase in cell spreading, attachment and proliferations on the biofunctionalized surfaces compared to the unmodified PCL surfaces at all-time points indicating the success of surface biofunctionalization.

Evaluate the growth and adhesion of osteoblast cells on nanocomposite scaffold of hydroxyapatite/titania coated with poly hydroxybutyrate

PubMed Central

Pourmollaabbassi, Babak; Karbasi, Saeed; Hashemibeni, Batool

2016-01-01

Background: The generation of bioartificial bone tissues may help to overcome the problems related to donor site morbidity and size limitations. Materials and Methods: In this paper, hydroxyapatite (HA) powder was made out of bovine bone by thermal analysis at 900°C and first, and then, porous HA (50 weight percentage) was produced by polyurethane sponge replication method. In order to improve the scaffold mechanical properties, they have been coated with poly hydroxybutyrate. In terms of phase studies, morphology, and specifying agent groups, the specific characterization devices such as X-ray diffraction and Fourier transform infrared, were employed. To compare the behavior of cellular scaffolds, they were divided into four groups of scaffolds. The osteoblast cells were cultured. To perform phase studies, analysis of Methylthiazole tetrazolium (MTT) and Trypan blue were carried out for the viability and attachment on the surface of the scaffold, and the specification of Scanning electron microscopy was employed for the morphology of the cells. Results: The results of MTT analysis performed on four groups of scaffolds have shown that Titanium oxide (Tio2) had no effect on cell growth alone and HA was the main factor of growth and cell osteoblast adhesion on the scaffold. Moreover, the results showed that the use of coating with poly-3-hydroxybutyrate saved the factors and placed the osteoblasts within the pore. Since the main part of bone consists of HA, the TiO2 accelerates the formation of apatite crystals at the scaffold surface which is the evidence for bone tissue regeneration. Conclusions: It is likely that the relation between HA and TiO2 leads to an increase in osteoblast adhesion and growth of cells on the scaffold surface. PMID:27761431

Evaluate the growth and adhesion of osteoblast cells on nanocomposite scaffold of hydroxyapatite/titania coated with poly hydroxybutyrate.

PubMed

Pourmollaabbassi, Babak; Karbasi, Saeed; Hashemibeni, Batool

2016-01-01

The generation of bioartificial bone tissues may help to overcome the problems related to donor site morbidity and size limitations. In this paper, hydroxyapatite (HA) powder was made out of bovine bone by thermal analysis at 900°C and first, and then, porous HA (50 weight percentage) was produced by polyurethane sponge replication method. In order to improve the scaffold mechanical properties, they have been coated with poly hydroxybutyrate. In terms of phase studies, morphology, and specifying agent groups, the specific characterization devices such as X-ray diffraction and Fourier transform infrared, were employed. To compare the behavior of cellular scaffolds, they were divided into four groups of scaffolds. The osteoblast cells were cultured. To perform phase studies, analysis of Methylthiazole tetrazolium (MTT) and Trypan blue were carried out for the viability and attachment on the surface of the scaffold, and the specification of Scanning electron microscopy was employed for the morphology of the cells. The results of MTT analysis performed on four groups of scaffolds have shown that Titanium oxide (Tio 2 ) had no effect on cell growth alone and HA was the main factor of growth and cell osteoblast adhesion on the scaffold. Moreover, the results showed that the use of coating with poly-3-hydroxybutyrate saved the factors and placed the osteoblasts within the pore. Since the main part of bone consists of HA, the TiO 2 accelerates the formation of apatite crystals at the scaffold surface which is the evidence for bone tissue regeneration. It is likely that the relation between HA and TiO 2 leads to an increase in osteoblast adhesion and growth of cells on the scaffold surface.

Characterization of bone marrow mononuclear cells on biomaterials for bone tissue engineering in vitro.

PubMed

Henrich, Dirk; Verboket, René; Schaible, Alexander; Kontradowitz, Kerstin; Oppermann, Elsie; Brune, Jan C; Nau, Christoph; Meier, Simon; Bonig, Halvard; Marzi, Ingo; Seebach, Caroline

2015-01-01

Bone marrow mononuclear cells (BMCs) are suitable for bone tissue engineering. Comparative data regarding the needs of BMC for the adhesion on biomaterials and biocompatibility to various biomaterials are lacking to a large extent. Therefore, we evaluated whether a surface coating would enhance BMC adhesion and analyze the biocompatibility of three different kinds of biomaterials. BMCs were purified from human bone marrow aspirate samples. Beta tricalcium phosphate (β-TCP, without coating or coated with fibronectin or human plasma), demineralized bone matrix (DBM), and bovine cancellous bone (BS) were assessed. Seeding efficacy on β-TCP was 95% regardless of the surface coating. BMC demonstrated a significantly increased initial adhesion on DBM and β-TCP compared to BS. On day 14, metabolic activity was significantly increased in BMC seeded on DBM in comparison to BMC seeded on BS. Likewise increased VEGF-synthesis was observed on day 2 in BMC seeded on DBM when compared to BMC seeded on BS. The seeding efficacy of BMC on uncoated biomaterials is generally high although there are differences between these biomaterials. Beta-TCP and DBM were similar and both superior to BS, suggesting either as suitable materials for spatial restriction of BMC used for regenerative medicine purposes in vivo.

Characterization of Bone Marrow Mononuclear Cells on Biomaterials for Bone Tissue Engineering In Vitro

PubMed Central

Verboket, René; Kontradowitz, Kerstin; Oppermann, Elsie; Brune, Jan C.; Nau, Christoph; Meier, Simon; Bonig, Halvard; Marzi, Ingo; Seebach, Caroline

2015-01-01

Bone marrow mononuclear cells (BMCs) are suitable for bone tissue engineering. Comparative data regarding the needs of BMC for the adhesion on biomaterials and biocompatibility to various biomaterials are lacking to a large extent. Therefore, we evaluated whether a surface coating would enhance BMC adhesion and analyze the biocompatibility of three different kinds of biomaterials. BMCs were purified from human bone marrow aspirate samples. Beta tricalcium phosphate (β-TCP, without coating or coated with fibronectin or human plasma), demineralized bone matrix (DBM), and bovine cancellous bone (BS) were assessed. Seeding efficacy on β-TCP was 95% regardless of the surface coating. BMC demonstrated a significantly increased initial adhesion on DBM and β-TCP compared to BS. On day 14, metabolic activity was significantly increased in BMC seeded on DBM in comparison to BMC seeded on BS. Likewise increased VEGF-synthesis was observed on day 2 in BMC seeded on DBM when compared to BMC seeded on BS. The seeding efficacy of BMC on uncoated biomaterials is generally high although there are differences between these biomaterials. Beta-TCP and DBM were similar and both superior to BS, suggesting either as suitable materials for spatial restriction of BMC used for regenerative medicine purposes in vivo. PMID:25802865

ARQ-197, a small-molecule inhibitor of c-Met, reduces tumour burden and prevents myeloma-induced bone disease in vivo.

PubMed

Lath, Darren L; Buckle, Clive H; Evans, Holly R; Fisher, Matthew; Down, Jenny M; Lawson, Michelle A; Chantry, Andrew D

2018-01-01

The receptor tyrosine kinase c-Met, its ligand HGF, and components of the downstream signalling pathway, have all been implicated in the pathogenesis of myeloma, both as modulators of plasma cell proliferation and as agents driving osteoclast differentiation and osteoblast inhibition thus, all these contribute substantially to the bone destruction typically caused by myeloma. Patients with elevated levels of HGF have a poor prognosis, therefore, targeting these entities in such patients may be of substantial benefit. We hypothesized that ARQ-197 (Tivantinib), a small molecule c-Met inhibitor, would reduce myeloma cell growth and prevent myeloma-associated bone disease in a murine model. In vitro we assessed the effects of ARQ-197 on myeloma cell proliferation, cytotoxicity and c-Met protein expression in human myeloma cell lines. In vivo we injected NOD/SCID-γ mice with PBS (non-tumour bearing) or JJN3 cells and treated them with either ARQ-197 or vehicle. In vitro exposure of JJN3, U266 or NCI-H929 cells to ARQ-197 resulted in a significant inhibition of cell proliferation and an induction of cell death by necrosis, probably caused by significantly reduced levels of phosphorylated c-Met. In vivo ARQ-197 treatment of JJN3 tumour-bearing mice resulted in a significant reduction in tumour burden, tumour cell proliferation, bone lesion number, trabecular bone loss and prevented significant decreases in the bone formation rate on the cortico-endosteal bone surface compared to the vehicle group. However, no significant differences on bone parameters were observed in non-tumour mice treated with ARQ-197 compared to vehicle, implying that in tumour-bearing mice the effects of ARQ-197 on bone cells was indirect. In summary, these res ults suggest that ARQ-197 could be a promising therapeutic in myeloma patients, leading to both a reduction in tumour burden and an inhibition of myeloma-induced bone disease.

Fabrication of Trabecular Bone-Templated Tissue-Engineered Constructs by 3D Inkjet Printing.

PubMed

Vanderburgh, Joseph P; Fernando, Shanik J; Merkel, Alyssa R; Sterling, Julie A; Guelcher, Scott A

2017-11-01

3D printing enables the creation of scaffolds with precisely controlled morphometric properties for multiple tissue types, including musculoskeletal tissues such as cartilage and bone. Computed tomography (CT) imaging has been combined with 3D printing to fabricate anatomically scaled patient-specific scaffolds for bone regeneration. However, anatomically scaled scaffolds typically lack sufficient resolution to recapitulate the <100 micrometer-scale trabecular architecture essential for investigating the cellular response to the morphometric properties of bone. In this study, it is hypothesized that the architecture of trabecular bone regulates osteoblast differentiation and mineralization. To test this hypothesis, human bone-templated 3D constructs are fabricated via a new micro-CT/3D inkjet printing process. It is shown that this process reproducibly fabricates bone-templated constructs that recapitulate the anatomic site-specific morphometric properties of trabecular bone. A significant correlation is observed between the structure model index (a morphometric parameter related to surface curvature) and the degree of mineralization of human mesenchymal stem cells, with more concave surfaces promoting more extensive osteoblast differentiation and mineralization compared to predominately convex surfaces. These findings highlight the significant effects of trabecular architecture on osteoblast function. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A stem cell apostasy: A tale of 4 H words

PubMed Central

Quesenberry, Peter J.; Goldberg, Laura R.; Dooner, Mark S.

2014-01-01

The field of hematopoietic stem cell biology has become increasingly dominated by the pursuit and study of highly purified populations of hematopoietic stem cells (HSCs). Such HSCs are typically isolated based on their cell surface marker expression patterns and ultimately defined by their multipotency and capacity for self-generation. However, even with progressively more stringent stem cell separation techniques, the resultant HSC population remains heterogeneous with respect to both self-renewal and differentiation capacity. Critical studies on un-separated whole bone marrow (WBM) have definitively shown that long-term engraftable hematopoietic stem cells are in active cell cycle and thus continually changing phenotype. Therefore, they cannot be purified by current approaches dependent on stable surface epitope expression because the surface markers are continually changing as well. These critical cycling cells are discarded with current stem cell purifications. Despite this, research defining such characteristics as self-renewal capacity, lineage-commitment, bone marrow niches, and proliferative state of HSCs continues to focus predominantly on this small sub-population of purified marrow cells. This review discusses the research leading to the hierarchical model of hematopoiesis and questions the dogmas pertaining to HSC quiescence and purification. PMID:25183450

The structurally effect of surface coated rhamnogalacturonan I on response of the osteoblast-like cell line SaOS-2.

PubMed

Svava, Rikke; Gurzawska, Katarzyna; Yihau, Yu; Haugshøj, Kenneth Brian; Dirscherl, Kai; Levery, Steven B; Jørgensen, Niklas Rye; Gotfredsen, Klaus; Damager, Iben; Ulvskov, Peter; Jørgensen, Bodil

2014-06-01

Osseointegration is important when implants are inserted into the bone and can be improved by biochemical surface coating of the implant. In this paper enzymatically modified rhamnogalacturonan I (RG-I) from apple and lupin was used for biochemical coating of aminated surfaces and the importance of the quality of RG-I, the nature of the binding, the fine structure of RG-I, and its effect on SaOS-2 cell line cultured on coated surfaces was investigated. SaOS-2 cells are osteoblast-like cells and a well-established in vitro model of bone-matrix forming osteoblasts. Purification by gel filtration could remove small fragments of galacturonic acid (GalA) and binding studies showed that the purity of the RG-I molecules was important for the quality of the coating. The structure of RG-I and osteoblast-like cells' viability were positively correlated so that high content of 1,4-linked galactose (Gal) and a low content of arabinose in the RG-I molecules favored cell viability. These results indicate that coating of implants with RG-I affect osseointegration positively. Copyright © 2013 Wiley Periodicals, Inc.

3D-Printed Bioactive Ca3SiO5 Bone Cement Scaffolds with Nano Surface Structure for Bone Regeneration.

PubMed

Yang, Chen; Wang, Xiaoya; Ma, Bing; Zhu, Haibo; Huan, Zhiguang; Ma, Nan; Wu, Chengtie; Chang, Jiang

2017-02-22

Silicate bioactive materials have been widely studied for bone regeneration because of their eminent physicochemical properties and outstanding osteogenic bioactivity, and different methods have been developed to prepare porous silicate bioactive ceramics scaffolds for bone-tissue engineering applications. Among all of these methods, the 3D-printing technique is obviously the most efficient way to control the porous structure. However, 3D-printed bioceramic porous scaffolds need high-temperature sintering, which will cause volume shrinkage and reduce the controllability of the pore structure accuracy. Unlike silicate bioceramic, bioactive silicate cements such as tricalcium silicate (Ca 3 SiO 5 and C 3 S) can be self-set in water to obtain high mechanical strength under mild conditions. Another advantage of using C 3 S to prepare 3D scaffolds is the possibility of simultaneous drug loading. Herein, we, for the first time, demonstrated successful preparation of uniform 3D-printed C 3 S bone cement scaffolds with controllable 3D structure at room temperature. The scaffolds were loaded with two model drugs and showed a loading location controllable drug-release profile. In addition, we developed a surface modification process to create controllable nanotopography on the surface of pore wall of the scaffolds, which showed activity to enhance rat bone-marrow stem cells (rBMSCs) attachment, spreading, and ALP activities. The in vivo experiments revealed that the 3D-printed C 3 S bone cement scaffolds with nanoneedle-structured surfaces significantly improved bone regeneration, as compared to pure C 3 S bone cement scaffolds, suggesting that 3D-printed C 3 S bone cement scaffolds with controllable nanotopography surface are bioactive implantable biomaterials for bone repair.

Human dental pulp stem cells: from biology to clinical applications.

PubMed

d'Aquino, Riccardo; De Rosa, Alfredo; Laino, Gregorio; Caruso, Filippo; Guida, Luigi; Rullo, Rosario; Checchi, Vittorio; Laino, Luigi; Tirino, Virginia; Papaccio, Gianpaolo

2009-07-15

Dental pulp stem cells (DPSCs) can be found within the "cell rich zone" of dental pulp. Their embryonic origin, from neural crests, explains their multipotency. Up to now, two groups have studied these cells extensively, albeit with different results. One group claims that these cells produce a "dentin-like tissue", whereas the other research group has demonstrated that these cells are capable of producing bone, both in vitro and in vivo. In addition, it has been reported that these cells can be easily cryopreserved and stored for long periods of time and still retain their multipotency and bone-producing capacity. Moreover, recent attention has been focused on tissue engineering and on the properties of these cells: several scaffolds have been used to promote 3-D tissue formation and studies have demonstrated that DPSCs show good adherence and bone tissue formation on microconcavity surface textures. In addition, adult bone tissue with good vascularization has been obtained in grafts. These results enforce the notion that DPSCs can be used successfully for tissue engineering. (c) 2008 Wiley-Liss, Inc.

Plasma Surface Modification for Immobilization of Bone Morphogenic Protein-2 on Polycaprolactone Scaffolds

NASA Astrophysics Data System (ADS)

Kim, Byung Hoon; Myung, Sung Woon; Jung, Sang Chul; Ko, Yeong Mu

2013-11-01

The immobilization of recombinant human bone formation protein-2 (rhBMP-2) on polycaprolactone (PCL) scaffolds was performed by plasma polymerization. RhBMP-2, which induces osteoblast differentiation in various cell types, is a growth factor that plays an important role in bone formation and repair. The surface of the PCL scaffold was functionalized with the carboxyl groups of plasma-polymerized acrylic acid (PPAA) thin films. Plasma polymerization was carried out at a discharge power of 60 W at an acrylic acid flow rate of 7 sccm for 5 min. The PPAA thin film exhibited moderate hydrophilic properties and possessed a high density of carboxyl groups. Carboxyl groups and rhBMP-2 on the PCL scaffolds surface were identified by attenuated total reflection Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, respectively. The alkaline phosphatase activity assay showed that the rhBMP-2 immobilized PCL scaffold increased the level of MG-63 cell differentiation. Plasma surface modification for the preparation of biomaterials, such as biofunctionalized polymer scaffolds, can be used for the binding of bioactive molecules in tissue engineering.

A mechanism of bone tissue loss in monkeys (BION - 11).

NASA Astrophysics Data System (ADS)

Rodionova, N. V.; Oganov, V. S.

The elucidation of mechanisms of bone tissue loss under the spaceflight conditions remains an actual problem until now It was established that primary reactions to a mechanical stress evolve at the cellular level therefore the main attention of the researchers was aimed at studying bone tissue cells and their interactions With the use of electron microscopy we studied osteoblasts osteocytes osteoclasts and stromal cells in bioptats of the iliac bone crest from monkeys flown on board the satellite guillemotleft BION - 11 guillemotright during 2 weeks The flight samples were compared with the vivarium and simulation controls The functional state of cells was evaluated by the degree of development of organelles for specific biosyntheses rough endoplasmic reticulum Golgy complex nucleus state interrelation with a mineralized matrix The analysis of the obtained results and data of other authors Klein -- Nulend et al 2003 etc permits to suppose that the following sequence of cell interactions underlies the bone tissue loss during mechanical stress microgravity reaction of mechano-sensitive osteocytes to a mechanical stimulus consisting in enhancement of osteolytic processes in cells which results in a partial bone tissue loss along the local unloading Simultaneously the modulating signals are transmitted through a system of canals and processes towards active osteoblasts surface osteocytes and bone marrow stromal cells as well As a reply to a mechanical stimulus there occurs a reduction slowing down of proliferation

Cementoblast Delivery for Periodontal Tissue Engineering

PubMed Central

Zhao, Ming; Jin, Qiming; Berry, Janice E.; Nociti, Francisco H.; Giannobile, William V.; Somerman, Martha J.

2008-01-01

Background Predictable periodontal regeneration following periodontal disease is a major goal of therapy. The objective of this proof of concept investigation was to evaluate the ability of cementoblasts and dental follicle cells to promote periodontal regeneration in a rodent periodontal fenestration model. Methods The buccal aspect of the distal root of the first mandibular molar was denuded of its periodontal ligament (PDL), cementum, and superficial dentin through a bony window created bilaterally in 12 athymic rats. Treated defects were divided into three groups: 1) carrier alone (PLGA polymer sponges), 2) carrier + follicle cells, and 3) carrier + cementoblasts. Cultured murine primary follicle cells and immortalized cementoblasts were delivered to the defects via biodegradable PLGA polymer sponges, and mandibulae were retrieved 3 weeks and 6 weeks post-surgery for histological evaluation. In situ hybridization, for gene expression of bone sialoprotein (BSP) and osteocalcin (OCN), and histomorphometric analysis were further done on 3-week specimens. Results Three weeks after surgery, histology of defects treated with carrier alone indicated PLGA particles, fibrous tissue, and newly formed bone scattered within the defect area. Defects treated with carrier + follicle cells had a similar appearance, but with less formation of bone. In contrast, in defects treated with carrier + cementoblasts, mineralized tissues were noted at the healing site with extension toward the root surface, PDL region, and laterally beyond the buccal plate envelope of bone. No PDL-bone fibrous attachment was observed in any of the groups at this point. In situ hybridization showed that the mineralized tissue formed by cementoblasts gave strong signals for both BSP and OCN genes, confirming its nature as cementum or bone. The changes noted at 3 weeks were also observed at 6 weeks. Cementoblast-treated and carrier alone-treated defects exhibited complete bone bridging and PDL formation, whereas follicle cell-treated defects showed minimal evidence of osteogenesis. No new cementum was formed along the root surface in the above two groups. Cementoblast-treated defects were filled with trabeculated mineralized tissue similar to, but more mature, than that seen at 3 weeks. Furthermore, the PDL region was maintained with well-organized collagen fibers connecting the adjacent bone to a thin layer of cementum-like tissue observed on the root surface. Neoplastic changes were observed at the superficial portions of the implants in two of the 6-week cementoblast-treated specimens, possibly due in part to the SV40-transformed nature of the implanted cell line. Conclusions This pilot study demonstrates that cementoblasts have a marked ability to induce mineralization in periodontal wounds when delivered via polymer sponges, while implanted dental follicle cells seem to inhibit periodontal healing. These results confirm the selective behaviors of different cell types in vivo and support the role of cementoblasts as a tool to better understand periodontal regeneration and cementogenesis. PMID:15025227

Microgrooved Polymer Substrates Promote Collective Cell Migration To Accelerate Fracture Healing in an in Vitro Model.

PubMed

Zhang, Qing; Dong, Hua; Li, Yuli; Zhu, Ye; Zeng, Lei; Gao, Huichang; Yuan, Bo; Chen, Xiaofeng; Mao, Chuanbin

2015-10-21

Surface topography can affect cell adhesion, morphology, polarity, cytoskeleton organization, and osteogenesis. However, little is known about the effect of topography on the fracture healing in repairing nonunion and large bone defects. Microgrooved topography on the surface of bone implants may promote cell migration into the fracture gap to accelerate fracture healing. To prove this hypothesis, we used an in vitro fracture (wound) healing assay on the microgrooved polycaprolactone substrates to study the effect of microgroove widths and depths on the osteoblast-like cell (MG-63) migration and the subsequent healing. We found that the microgrooved substrates promoted MG-63 cells to migrate collectively into the wound gap, which serves as a fracture model, along the grooves and ridges as compared with the flat substrates. Moreover, the groove widths did not show obvious influence on the wound healing whereas the smaller groove depths tended to favor the collective cell migration and thus subsequent healing. The microgrooved substrates accelerated the wound healing by facilitating the collective cell migration into the wound gaps but not by promoting the cell proliferation. Furthermore, microgrooves were also found to promote the migration of human mesenchymal stem cells (hMSCs) to heal the fracture model. Though osteogenic differentiation of hMSCs was not improved on the microgrooved substrate, collagen I and minerals deposited by hMSCs were organized in a way similar to those in the extracellular matrix of natural bone. These findings suggest the necessity in using microgrooved implants in enhancing fracture healing in bone repair.

Origins of endothelial and osteogenic cells in the subcutaneous collagen gel implant.

PubMed

Bilic-Curcic, I; Kalajzic, Z; Wang, L; Rowe, D W

2005-11-01

The interdependent relationship between vascular endothelial cells and osteoblasts during bone formation and fracture healing has been long appreciated. This paper reports a heterotopic implant model using FGF-2-expanded bone marrow stromal cells (BMSC) derived from Tie2eGFP (endothelial marker) and pOBCol3.6GFPcyan or topaz (early osteoblast marker) transgenic mice to appreciate the host/donor relationships of cells participating in the process of heterotopic bone formation. The study included various combinations of Tie2eGFP and pOBCol3.6GFPcyan and topaz transgenics as BMSC or whole bone marrow (WBM) donors and also as recipients. Rat tail collagen was used as a carrier of donor cells and implantation was done in lethally irradiated mice rescued with WBM injection. Development of ossicles in the implants was followed weekly during the 4- to 5-week long post-implantation period. By 4-5 weeks after total body irradiation (TBI) and implantation, a well-formed bone spicule had developed that was invested with bone marrow. Experiments showed absolute dominance of donor-derived cells in the formation of endothelial-lined vessels inside the implants as well as the marrow stromal-derived osteogenic cells. Host-derived fibroblasts and osteogenic cells were confined to the fibrous capsule surrounding the implant. In addition, cells lining the endosteal surface of newly formed marrow space carrying a pOBCol3.6GFP marker were observed that were contributed by WBM donor cells and the host. Thus, FGF-2-expanded BMSC appear to be a source of endothelial and osteogenic progenitor cells capable of eliciting heterotopic bone formation independent of cells from the host. This model should be useful for understanding the interactions between these two cell types that control osteogenic differentiation in vivo.

Bone cell-materials interaction on Si microchannels with bioinert coatings.

PubMed

Condie, Russell; Bose, Susmita; Bandyopadhyay, Amit

2007-07-01

Bone implant life is dependent upon integration of biomaterial surfaces with local osteoblasts. This investigation studied the effects of various microchannel parameters and surface chemistry on immortalized osteoblast precursor cell (OPC1) adhesion. Cell-materials interactions were observed within channels of varying length, width, tortuosity, convergence, divergence and chemistry. Si wafers were used to create four distinct 1cm(2) designs of varying channel dimensions. After anisotropic chemical etching to a depth of 120microm, wafers were sputter coated with gold and titanium; and on another surface SiO(2) was grown to vary the surface chemistry of these microchannels. OPC1 cells were seeded in the central cavity of each chip before incubation in tissue culture plates. On days 5, 11 and 16, samples were taken out, fixed and processed for microscopic analysis. Samples were visually characterized, qualitatively scored and analyzed. Channel walls did not contain OPC1 migration, but showed locally interrupted adhesion. Scores for channels of floor widths as narrow as 350microm were significantly reduced. No statistically significant preference was detected for gold, titanium or SiO(2) surfaces. Bands of OPC1 cells appeared to align with nearby channels, suggesting that cell morphology may be controlled by topography of the design to improve osseointegration.

Nanoporous metals for biodegradable implants: Initial bone mesenchymal stem cell adhesion and degradation behavior.

PubMed

Heiden, Michael; Huang, Sabrina; Nauman, Eric; Johnson, David; Stanciu, Lia

2016-07-01

Nanostructured Fe-Mn and Fe-Mn-Zn metal scaffolds were generated through a well-controlled selective leaching process in order to fulfill the growing demand for adjustable degradation rates and improved cellular response of resorbable materials. Mouse bone marrow mesenchymal stem cells (D1 ORL UVA) were seeded onto eleven, carefully chosen nanoporous surfaces for 24 h in vitro. Using a combination of fluorescence microscopy, scanning electron microscopy (SEM), and an MTS assay, it was discovered that scaffolds with nanoscale roughened surfaces had increased cell attachment by up to 123% compared to polished smooth Fe-Mn surfaces. Significant cell spreading and construction of cell multilayers were also apparent after 24 h, suggesting better adhesion. Additionally, static electrochemical polarization experiments revealed an improvement of up to 26% in the actual rate of biodegradation for Fe-Mn surface-modified materials. However, any residual concentration of zinc after leaching was shown to slightly increase corrosion resistance. The results demonstrate that selectively leached, nanostructured Fe-Mn surfaces have the potential of being tailored to a diverse set of transient implant scenarios, while also effectively boosting overall biocompatibility, initial cell attachment, and degradation rate. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1747-1758, 2016. © 2016 Wiley Periodicals, Inc.

Laser-modified titanium surfaces enhance the osteogenic differentiation of human mesenchymal stem cells.

PubMed

Bressel, Tatiana A B; de Queiroz, Jana Dara Freires; Gomes Moreira, Susana Margarida; da Fonseca, Jéssyca T; Filho, Edson A; Guastaldi, Antônio Carlos; Batistuzzo de Medeiros, Silvia Regina

2017-11-28

Titanium surfaces have been modified by various approaches with the aim of improving the stimulation of osseointegration. Laser beam (Yb-YAG) treatment is a controllable and flexible approach to modifying surfaces. It creates a complex surface topography with micro and nano-scaled patterns, and an oxide layer that can improve the osseointegration of implants, increasing their usefulness as bone implant materials. Laser beam irradiation at various fluences (132, 210, or 235 J/cm 2 ) was used to treat commercially pure titanium discs to create complex surface topographies. The titanium discs were investigated by scanning electron microscopy, X-ray diffraction, and measurement of contact angles. The surface generated at a fluence of 235 J/cm 2 was used in the biological assays. The behavior of mesenchymal stem cells from an umbilical cord vein was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, a mineralization assay, and an alkaline phosphatase activity assay and by carrying out a quantitative real-time polymerase chain reaction for osteogenic markers. CHO-k1 cells were also exposed to titanium discs in the MTT assay. The best titanium surface was that produced by laser beam irradiation at 235 J/cm 2 fluence. Cell proliferation analysis revealed that the CHO-k1 and mesenchymal stem cells behaved differently. The laser-processed titanium surface increased the proliferation of CHO-k1 cells, reduced the proliferation of mesenchymal stem cells, upregulated the expression of the osteogenic markers, and enhanced alkaline phosphatase activity. The laser-treated titanium surface modulated cellular behavior depending on the cell type, and stimulated osteogenic differentiation. This evidence supports the potential use of laser-processed titanium surfaces as bone implant materials, and their use in regenerative medicine could promote better outcomes.

A c-fms tyrosine kinase inhibitor, Ki20227, suppresses osteoclast differentiation and osteolytic bone destruction in a bone metastasis model.

PubMed

Ohno, Hiroaki; Kubo, Kazuo; Murooka, Hideko; Kobayashi, Yoshiko; Nishitoba, Tsuyoshi; Shibuya, Masabumi; Yoneda, Toshiyuki; Isoe, Toshiyuki

2006-11-01

In bone metastatic lesions, osteoclasts play a key role in the development of osteolysis. Previous studies have shown that macrophage colony-stimulating factor (M-CSF) is important for the differentiation of osteoclasts. In this study, we investigated whether an inhibitor of M-CSF receptor (c-Fms) suppresses osteoclast-dependent osteolysis in bone metastatic lesions. We developed small molecule inhibitors against ligand-dependent phosphorylation of c-Fms and examined the effects of these compounds on osteolytic bone destruction in a bone metastasis model. We discovered a novel quinoline-urea derivative, Ki20227 (N-{4-[(6,7-dimethoxy-4-quinolyl)oxy]-2-methoxyphenyl}-N'-[1-(1,3-thiazole-2-yl)ethyl]urea), which is a c-Fms tyrosine kinase inhibitor. The IC(50)s of Ki20227 to inhibit c-Fms, vascular endothelial growth factor receptor-2 (KDR), stem cell factor receptor (c-Kit), and platelet-derived growth factor receptor beta were found to be 2, 12, 451, and 217 nmol/L, respectively. Ki20227 did not inhibit other kinases tested, such as fms-like tyrosine kinase-3, epidermal growth factor receptor, or c-Src (c-src proto-oncogene product). Ki20227 was also found to inhibit the M-CSF-dependent growth of M-NFS-60 cells but not the M-CSF-independent growth of A375 human melanoma cells in vitro. Furthermore, in an osteoclast-like cell formation assay using mouse bone marrow cells, Ki20227 inhibited the development of tartrate-resistant acid phosphatase-positive osteoclast-like cells in a dose-dependent manner. In in vivo studies, oral administration of Ki20227 suppressed osteoclast-like cell accumulation and bone resorption induced by metastatic tumor cells in nude rats following intracardiac injection of A375 cells. Moreover, Ki20227 decreased the number of tartrate-resistant acid phosphatase-positive osteoclast-like cells on bone surfaces in ovariectomized (ovx) rats. These findings suggest that Ki20227 inhibits osteolytic bone destruction through the suppression of M-CSF-induced osteoclast accumulation in vivo. Therefore, Ki20227 may be a useful therapeutic agent for osteolytic disease associated with bone metastasis and other bone diseases.

Determination of the Fate and Function of Innate Lymphoid Cells Following Adoptive Transfer of Innate Lymphoid Cell Precursors.

PubMed

O'Sullivan, Timothy E; Sun, Joseph C

2018-01-01

Innate lymphoid cells are a heterogeneous family of tissue-resident and circulating lymphocytes that play an important role in host immunity. Recent studies have profiled the developmental pathways of mature ILCs and have identified ILC progenitors in the bone marrow through the use of transcription factor reporter mice. Here we describe methodology to identify and isolate bone marrow CHILP and ILC2 progenitor (ILC2P) cells based on cell surface marker expression for adoptive transfer into lymphopenic mice to track the fate of developing ILCs.

[Scanning electron microscopy observation of the growth of osteoblasts on Ti-24Nb-4Zr-8Sn modified by micro-arc oxidation and alkali-heat treatment and implant-bone interface].

PubMed

Han, Xue; Liu, Hong-Chen; Wang, Dong-Sheng; Li, Shu-Jun; Yang, Rui

2011-01-01

To observe the efficacy of micro-arc oxidation and alkali-heat treatment (MAH) on Ti-24Nb-4Zr-8Sn (Ti2448). Disks (diameter of 14.5 mm, thickness of 1 mm) and cylinders (diameter of 3 mm, height of 10 mm) were fabricated from Ti2448 alloy. Samples were divided into three groups: polished (Ti2448), micro-arc oxidation(MAO-Ti2448), micro-arc oxidation and alkali-heat treatment (MAH-Ti2448). MC3T3-E1 osteoblastic cells were cultured on the disks and cell morphology was observed with scanning electron microscopy (SEM) aftre 3 days. The cylinder samples were implanted in the tibia of dogs and implant-bone interface was observed with SEM after 3 months. A rough and porous structure was shown in both MAO and MAH group. The MC3T3-E1 cells on the MAH-Ti2448 discs spread fully in intimate contact with the underlying coarse surface through active cytoskeletal extentions. Osseointegration was formed in the implant-bone interface in MAH samples. MAH treatment can provide a more advantageous Ti2448 surface to osteoblastic cells than MAO treatment does, and the former can improve the implant-bone integration.

Bone-repair properties of biodegradable hydroxyapatite nano-rod superstructures

NASA Astrophysics Data System (ADS)

D'Elía, Noelia L.; Mathieu, Colleen; Hoemann, Caroline D.; Laiuppa, Juan A.; Santillán, Graciela E.; Messina, Paula V.

2015-11-01

Nano-hydroxyapatite (nano-HAp) materials show an analogous chemical composition to the biogenic mineral components of calcified tissues and depending on their topography they may mimic the specific arrangement of the crystals in bone. In this work, we have evaluated the potential of four synthesized nano-HAp superstructures for the in vitro conditions of bone-repair. Experiments are underway to investigate the effects of the material microstructure, surface roughness and hydrophilicity on their osseo-integration, osteo-conduction and osteo-induction abilities. Materials were tested in the presence of both, rat primary osteoblasts and rabbit mesenchymal stem cells. The following aspects are discussed: (i) cytotoxicity and material degradation; (ii) rat osteoblast spreading, proliferation and differentiation; and (iii) rabbit mesenchymal stem cell adhesion on nano-HAp and nano-HAp/collagen type I coatings. We effectively prepared a material based on biomimetic HAp nano-rods displaying the appropriate surface topography, hydrophilicity and degradation properties to induce the in vitro desired cellular responses for bone bonding and healing. Cells seeded on the selected material readily attached, proliferated and differentiated, as confirmed by cell viability, mitochondrial metabolic activity, alkaline phosphatase (ALP) activity and cytoskeletal integrity analysis by immunofluorescence localization of alpha-smooth muscle actin (α-SMA) protein. These results highlight the influence of material's surface characteristics to determine their tissue regeneration potential and their future use in engineering osteogenic scaffolds for orthopedic implants.Nano-hydroxyapatite (nano-HAp) materials show an analogous chemical composition to the biogenic mineral components of calcified tissues and depending on their topography they may mimic the specific arrangement of the crystals in bone. In this work, we have evaluated the potential of four synthesized nano-HAp superstructures for the in vitro conditions of bone-repair. Experiments are underway to investigate the effects of the material microstructure, surface roughness and hydrophilicity on their osseo-integration, osteo-conduction and osteo-induction abilities. Materials were tested in the presence of both, rat primary osteoblasts and rabbit mesenchymal stem cells. The following aspects are discussed: (i) cytotoxicity and material degradation; (ii) rat osteoblast spreading, proliferation and differentiation; and (iii) rabbit mesenchymal stem cell adhesion on nano-HAp and nano-HAp/collagen type I coatings. We effectively prepared a material based on biomimetic HAp nano-rods displaying the appropriate surface topography, hydrophilicity and degradation properties to induce the in vitro desired cellular responses for bone bonding and healing. Cells seeded on the selected material readily attached, proliferated and differentiated, as confirmed by cell viability, mitochondrial metabolic activity, alkaline phosphatase (ALP) activity and cytoskeletal integrity analysis by immunofluorescence localization of alpha-smooth muscle actin (α-SMA) protein. These results highlight the influence of material's surface characteristics to determine their tissue regeneration potential and their future use in engineering osteogenic scaffolds for orthopedic implants. Electronic supplementary information (ESI) available: Calculation of roughness parameters Rz, Rz,max, and Rz, prom. Nano-HAp powder degradation after immersion in phosphate buffer (pH = 7.4). Optical phase contrast microphotographs of MSC adhesion on nano-HAp and nano-HAp/Co I coatings at different concentrations. Laser scanning confocal microphotographs of MSCs' α-SMA expression spreading on large amounts of nano-HAp (MI) coatings. Immunofluorescence microphotograph analysis by image software. See DOI: 10.1039/c5nr04850h

Topography of calcium phosphate ceramics regulates primary cilia length and TGF receptor recruitment associated with osteogenesis.

PubMed

Zhang, Jingwei; Dalbay, Melis T; Luo, Xiaoman; Vrij, Erik; Barbieri, Davide; Moroni, Lorenzo; de Bruijn, Joost D; van Blitterswijk, Clemens A; Chapple, J Paul; Knight, Martin M; Yuan, Huipin

2017-07-15

The surface topography of synthetic biomaterials is known to play a role in material-driven osteogenesis. Recent studies show that TGFβ signalling also initiates osteogenic differentiation. TGFβ signalling requires the recruitment of TGFβ receptors (TGFβR) to the primary cilia. In this study, we hypothesize that the surface topography of calcium phosphate ceramics regulates stem cell morphology, primary cilia structure and TGFβR recruitment to the cilium associated with osteogenic differentiation. We developed a 2D system using two types of tricalcium phosphate (TCP) ceramic discs with identical chemistry. One sample had a surface topography at micron-scale (TCP-B, with a bigger surface structure dimension) whilst the other had a surface topography at submicron scale (TCP-S, with a smaller surface structure dimension). In the absence of osteogenic differentiation factors, human bone marrow stromal cells (hBMSCs) were more spread on TCP-S than on TCP-B with alterations in actin organization and increased primary cilia prevalence and length. The cilia elongation on TCP-S was similar to that observed on glass in the presence of osteogenic media and was followed by recruitment of transforming growth factor-β RII (p-TGFβ RII) to the cilia axoneme. This was associated with enhanced osteogenic differentiation of hBMSCs on TCP-S, as shown by alkaline phosphatase activity and gene expression for key osteogenic markers in the absence of additional osteogenic growth factors. Similarly, in vivo after a 12-week intramuscular implantation in dogs, TCP-S induced bone formation while TCP-B did not. It is most likely that the surface topography of calcium phosphate ceramics regulates primary cilia length and ciliary recruitment of p-TGFβ RII associated with osteogenesis and bone formation. This bioengineering control of osteogenesis via primary cilia modulation may represent a new type of biomaterial-based ciliotherapy for orthopedic, dental and maxillofacial surgery applications. The surface topography of synthetic biomaterials plays important roles in material-driven osteogenesis. The data presented herein have shown that the surface topography of calcium phosphate ceramics regulates mesenchymal stromal cells (e.g., human bone marrow mesenchymal stromal cells, hBMSCs) with respect to morphology, primary cilia structure and TGFβR recruitment to the cilium associated with osteogenic differentiation in vitro. Together with bone formation in vivo, our results suggested a new type of biomaterial-based ciliotherapy for orthopedic, dental and maxillofacial surgery by the bioengineering control of osteogenesis via primary cilia modulation. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Mechanical Properties of Optimized Diamond Lattice Structure for Bone Scaffolds Fabricated via Selective Laser Melting.

PubMed

Liu, Fei; Zhang, David Z; Zhang, Peng; Zhao, Miao; Jafar, Salman

2018-03-03

Developments in selective laser melting (SLM) have enabled the fabrication of periodic cellular lattice structures characterized by suitable properties matching the bone tissue well and by fluid permeability from interconnected structures. These multifunctional performances are significantly affected by cell topology and constitutive properties of applied materials. In this respect, a diamond unit cell was designed in particular volume fractions corresponding to the host bone tissue and optimized with a smooth surface at nodes leading to fewer stress concentrations. There were 33 porous titanium samples with different volume fractions, from 1.28 to 18.6%, manufactured using SLM. All of them were performed under compressive load to determine the deformation and failure mechanisms, accompanied by an in-situ approach using digital image correlation (DIC) to reveal stress-strain evolution. The results showed that lattice structures manufactured by SLM exhibited comparable properties to those of trabecular bone, avoiding the effects of stress-shielding and increasing longevity of implants. The curvature of optimized surface can play a role in regulating the relationship between density and mechanical properties. Owing to the release of stress concentration from optimized surface, the failure mechanism of porous titanium has been changed from the pattern of bottom-up collapse by layer (or cell row) to that of the diagonal (45°) shear band, resulting in the significant enhancement of the structural strength.

Mechanical Properties of Optimized Diamond Lattice Structure for Bone Scaffolds Fabricated via Selective Laser Melting

PubMed Central

Zhang, David Z.; Zhang, Peng; Zhao, Miao; Jafar, Salman

2018-01-01

Developments in selective laser melting (SLM) have enabled the fabrication of periodic cellular lattice structures characterized by suitable properties matching the bone tissue well and by fluid permeability from interconnected structures. These multifunctional performances are significantly affected by cell topology and constitutive properties of applied materials. In this respect, a diamond unit cell was designed in particular volume fractions corresponding to the host bone tissue and optimized with a smooth surface at nodes leading to fewer stress concentrations. There were 33 porous titanium samples with different volume fractions, from 1.28 to 18.6%, manufactured using SLM. All of them were performed under compressive load to determine the deformation and failure mechanisms, accompanied by an in-situ approach using digital image correlation (DIC) to reveal stress–strain evolution. The results showed that lattice structures manufactured by SLM exhibited comparable properties to those of trabecular bone, avoiding the effects of stress-shielding and increasing longevity of implants. The curvature of optimized surface can play a role in regulating the relationship between density and mechanical properties. Owing to the release of stress concentration from optimized surface, the failure mechanism of porous titanium has been changed from the pattern of bottom-up collapse by layer (or cell row) to that of the diagonal (45°) shear band, resulting in the significant enhancement of the structural strength. PMID:29510492

The effect of oxygen plasma pretreatment and incubation in modified simulated body fluids on the formation of bone-like apatite on poly(lactide-co-glycolide) (70/30).

PubMed

Qu, Xue; Cui, Wenjin; Yang, Fei; Min, Changchun; Shen, Hong; Bei, Jianzhong; Wang, Shenguo

2007-01-01

In this study, biodegradable poly(lactide-co-glycolide) (PLGA) (70/30) films and scaffolds were first treated with oxygen plasma and then incubated in a modified simulated body fluid 1.5SBF0 to prepare a bone-like apatite layer. The formation of the apatite and its influence on osteoblast-like cells growth were investigated. It was found that the bone-like apatite formability of PLGA(70/30) was enhanced by plasma pretreatment. The changes of surface chemistry and surface topography induced by oxygen plasma treatment were both effective for apatite formation. The apatite formability increased with increasing plasma-treating time. Under a treating condition of 20 W for 30 min, oxygen plasma treatment could penetrate into the inner scaffold. After 6 days incubation, the apatite formed in plasma-treated scaffold was better distributed than in untreated scaffold, and the weight and mechanical strength of the plasma-treated scaffold were both enhanced. Compared with PLGA(70/30), the apatite layer formed on oxygen plasma-treated PLGA(70/30) surface enhanced adhesion and proliferation of OCT-1 osteoblast-like cell, but had no significant effect on cell's ALP activity at day 7. A prolonged investigation is being in process to further verify the bone-like apatite effects on osteogenic differentiation.

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.

Bioactive ceramic coating on orthopedic implants for enhanced bone tissue integration

NASA Astrophysics Data System (ADS)

Aniket

Tissue integration between bone and orthopedic implant is essential for implant fixation and longevity. An immunological response leads to fibrous encapsulation of metallic implants leading to implant instability and failure. Bioactive ceramics have the ability to directly bond to bone; however, they have limited mechanical strength for load bearing applications. Coating bioactive ceramics on metallic implant offers the exciting opportunity to enhance bone formation without compromising the mechanical strength of the implant. In the present study, we have developed a novel bioactive silica-calcium phosphate nanocomposite (SCPC) coating on medical grade Ti-6Al-4V orthopedic implant using electrophoretic deposition (EPD) and evaluated bone tissue response to the coated implant at the cellular level. The effect of SCPC composition and suspending medium pH on the zeta potential of three different SCPC formulations; SCPC25, SCPC50 and SCPC75 were analyzed. The average zeta potential of SCPC50 in pure ethanol was more negative than that of SCPC25 or SCPC75; however the difference was not statistically significant. Ti-6Al-4V discs were passivated, coated with SCPC50 (200 nm - 10 mum) and thermally treated at 600 - 800 ºC to produce a coating thickness in the range of 43.1 +/- 5.7 to 30.1 +/- 4.6 μm. After treatment at 600, 700 and 800 ºC, the adhesion strength at the SCPC50/Ti-6Al-4V interface was 42.6 +/- 3.6, 44.7 +/- 8.7 and 47.2 +/- 4.3 MPa, respectively. XRD analyses of SCPC50 before and after EPD coating indicated no change in the crystallinity of the material. Fracture surface analyses showed that failure occurred within the ceramic layer or at the ceramic/polymer interface; however, the ceramic/metal interface was intact in all samples. The adhesion strength of SCPC50-coated substrates after immersion in PBS for 2 days (11.7 +/- 3.9 MPa) was higher than that measured on commercially available hydroxyapatite (HA) coated substrates (5.5 +/- 2.7 MPa), although the difference was not statistically significant. SEM - EDX analyses of SCPC50-coated Ti-6Al-4V pre-immersed in PBS for 7 days showed the formation of a Ca-deficient HA surface layer. Bone cells attached to the SCPC50-coated implants expressed significantly higher (p < 0.05) alkaline phosphatase activity (82.4 +/- 25.6 nmoles p-NP/mg protein/min) than that expressed by cells attached to HA-coated or uncoated implants. Protein adsorption analyses showed that SCPC50-coated substrates adsorbed significantly more (p < 0.05) serum protein (14.9 +/- 1.2 mug) than control uncoated substrates (8.9 +/- 0.7 mug). Moreover, Western blot analysis showed that the SCPC50 coating has a high affinity for serum fibronectin. Protein conformation analyses by FTIR showed that the ratio of the area under the peak for amide I/amide II bands was significantly higher (p < 0.05) on the surface of SCPC50-coated substrate (5.0 +/- 0.6) than that on the surface of the control uncoated substrates (2.2 +/- 0.3). Moreover, ICP-OES analyses indicated that SCPC50-coated substrates withdrew Ca ions from, and released P and Si ions into, the tissue culture medium, respectively. In conjunction with the favorable protein adsorption and modifications in medium composition, MC3T3-E1 osteoblast-like cells attached to SCPC50-coated substrates expressed 10-fold higher level of mRNA encoding osteocalcin and had significantly higher production of osteopontin and osteocalcin proteins than cells attached to the uncoated Ti-6Al-4V substrate. In addition, osteoblast-like cells attached to the SCPC50-coated substrates produced significantly lower levels of the inflammatory and osteoclastogenic cytokines, IL-6, IL-12p40 and RANKL than those attached to uncoated Ti-6Al-4V. Surface topography analyses using AFM suggested that the SCPC50 particles deposit onto the metal surface in a manner that preferentially fills the grooves on the substrate created during substrate preparation. An increase in the surface roughness of the SCPC50-coated substrate from 217.8 +/- 54.6 nm to 284.3 +/- 37.3 nm was accompanied by enhanced material dissolution, reduced cell proliferation and poor actin cytoskeleton organization, which are characteristics typical of differentiating bone cells on bioactive ceramic surfaces. Results of the study demonstrate that bioactive SCPC50 can efficiently be coated on Ti-6Al-4V using EPD. Moreover, the in vitro bone cell response suggests that SCPC50-coating has the potential to enhance bone integration with orthopedic and maxillofacial implants while minimizing the induction of inflammatory bone cell responses.

Inhibition of bone loss with surface-modulated, drug-loaded nanoparticles in an intraosseous model of prostate cancer.

PubMed

Adjei, Isaac M; Sharma, Blanka; Peetla, Chiranjeevi; Labhasetwar, Vinod

2016-06-28

Advanced-stage prostate cancer usually metastasizes to bone and is untreatable due to poor biodistribution of intravenously administered anticancer drugs to bone. In this study, we modulated the surface charge/composition of biodegradable nanoparticles (NPs) to sustain their blood circulation time and made them small enough to extravasate through the openings of the bone's sinusoidal capillaries and thus localize into marrow. NPs with a neutral surface charge, achieved by modulating the NP surface-associated emulsifier composition, were more effective at localizing to bone marrow than NPs with a cationic or anionic surface charge. These small neutral NPs (~150nm vs. the more usual ~320nm) were also ~7-fold more effective in localizing in bone marrow than large NPs. We hypothesized that NPs that effectively localize to marrow could improve NP-mediated anticancer drug delivery to sites of bone metastasis, thereby inhibiting cancer progression and preventing bone loss. In a PC-3M-luc cell-induced osteolytic intraosseous model of prostate cancer, these small neutral NPs demonstrated greater accumulation in bone within metastatic sites than in normal contralateral bone as well as co-localization with the tumor mass in marrow. Significantly, a single-dose intravenous administration of these small neutral NPs loaded with paclitaxel (PTX-NPs), but not anionic PTX-NPs, slowed the progression of bone metastasis. In addition, neutral PTX-NPs prevented bone loss, whereas animals treated with the rapid-release drug formulation Cremophor EL (PTX-CrEL) or saline (control) showed >50% bone loss. Neutral PTX-NPs did not cause acute toxicity, whereas animals treated with PTX-CrEL experienced weight loss. These results indicate that NPs with appropriate physical and sustained drug-release characteristics could be explored to treat bone metastasis, a significant clinical issue in prostate and other cancers. Copyright © 2016 Elsevier B.V. All rights reserved.

Biological Response of Human Bone Marrow-Derived Mesenchymal Stem Cells to Commercial Tantalum Coatings with Microscale and Nanoscale Surface Topographies

NASA Astrophysics Data System (ADS)

Skoog, Shelby A.; Kumar, Girish; Goering, Peter L.; Williams, Brian; Stiglich, Jack; Narayan, Roger J.

2016-06-01

Tantalum is a promising orthopaedic implant coating material due to its robust mechanical properties, corrosion resistance, and excellent biocompatibility. Previous studies have demonstrated improved biocompatibility and tissue integration of surface-treated tantalum coatings compared to untreated tantalum. Surface modification of tantalum coatings with biologically inspired microscale and nanoscale features may be used to evoke optimal tissue responses. The goal of this study was to evaluate commercial tantalum coatings with nanoscale, sub-microscale, and microscale surface topographies for orthopaedic and dental applications using human bone marrow-derived mesenchymal stem cells (hBMSCs). Tantalum coatings with different microscale and nanoscale surface topographies were fabricated using a diffusion process or chemical vapor deposition. Biological evaluation of the tantalum coatings using hBMSCs showed that tantalum coatings promote cellular adhesion and growth. Furthermore, hBMSC adhesion to the tantalum coatings was dependent on surface feature characteristics, with enhanced cell adhesion on sub-micrometer- and micrometer-sized surface topographies compared to hybrid nano-/microstructures. Nanostructured and microstructured tantalum coatings should be further evaluated to optimize the surface coating features to promote osteogenesis and enhance osseointegration of tantalum-based orthopaedic implants.

Bio-active engineered 50 nm silica nanoparticles with bone anabolic activity: therapeutic index, effective concentration, and cytotoxicity profile in vitro

PubMed Central

Ha, Shin-Woo; Sikorski, James A.; Weitzmann, M. Neale; Beck, George R.

2014-01-01

Silica-based nanomaterials are generally considered to be excellent candidates for therapeutic applications particularly related to skeletal metabolism however the current data surrounding the safety of silica based nanomaterials is conflicting. This may be due to differences in size, shape, incorporation of composite materials, surface properties, as well as the presence of contaminants following synthesis. In this study we performed extensive in vitro safety profiling of ~50 nm spherical silica nanoparticles with OH-terminated or Polyethylene Glycol decorated surface, with and without a magnetic core, and synthesized by the Stöber method. Nineteen different cell lines representing all major organ types were used to investigate an in vitro lethal concentration (LC) and results revealed little toxicity in any cell type analyzed. To calculate an in vitro therapeutic index we quantified the effective concentration at 50% response (EC50) for nanoparticle-stimulated mineral deposition activity using primary bone marrow stromal cells (BMSCs). The EC50 for BMSCs was not substantially altered by surface or magnetic core. The calculated Inhibitory concentration 50% (IC50) for pre-osteoclasts was similar to the osteoblastic cells. These results demonstrate the pharmacological potential of certain silica-based nanomaterial formulations for use in treating bone diseases based on a favorable in vitro therapeutic index. PMID:24333519

Role of hepatocyte growth factor in the development of dendritic cells from CD34+ bone marrow cells.

PubMed

Ovali, E; Ratip, S; Kibaroglu, A; Tekelioglu, Y; Cetiner, M; Karti, S; Aydin, F; Bayik, M; Akoglu, T

2000-05-01

Hepatocyte growth factor (HGF) is known to augment the effects of stem cell factor, interleukin-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), erythropoetin, and granulocyte colony-stimulating factor, all of which are involved in hematopoiesis. HGF is also known to have a role in immune responses. The aim of this study was to investigate whether HGF is involved in the development of dendritic cells (DC) from CD34+ bone marrow cells. CD34+ cells obtained from three healthy donors were incubated in various combinations of HGF, GM-CSF, and tumor necrosis factor (TNF) for 12 days. Developing cell populations were analyzed for surface markers, morphology and functional capacities by flow cytometry, light microscopy and mixed lymphocyte reaction, respectively. Incubation with HGF alone generated greater number of dendritic cells from CD34+ bone marrow cells than incubation with GM-CSF, or a combination of GM-CSF with TNF. HGF was also found to potentiate the effect of GM-CSF on DC and monocyte development. The effects of HGF were inhibited by the concurrent use of TNF. HGF appears to be a significant factor in the development of dendritic cells from CD34+ bone marrow cells.

Laser Sintered Porous Ti-6Al-4V Implants Stimulate Vertical Bone Growth.

PubMed

Cheng, Alice; Cohen, David J; Kahn, Adrian; Clohessy, Ryan M; Sahingur, Kaan; Newton, Joseph B; Hyzy, Sharon L; Boyan, Barbara D; Schwartz, Zvi

2017-08-01

The objective of this study was to examine the ability of 3D implants with trabecular-bone-inspired porosity and micro-/nano-rough surfaces to enhance vertical bone ingrowth. Porous Ti-6Al-4V constructs were fabricated via laser-sintering and processed to obtain micro-/nano-rough surfaces. Male and female human osteoblasts were seeded on constructs to analyze cell morphology and response. Implants were then placed on rat calvaria for 10 weeks to assess vertical bone ingrowth, mechanical stability and osseointegration. All osteoblasts showed higher levels of osteocalcin, osteoprotegerin, vascular endothelial growth factor and bone morphogenetic protein 2 on porous constructs compared to solid laser-sintered controls. Porous implants placed in vivo resulted in an average of 3.1 ± 0.6 mm 3 vertical bone growth and osseointegration within implant pores and had significantly higher pull-out strength values than solid implants. New bone formation and pull-out strength was not improved with the addition of demineralized bone matrix putty. Scanning electron images and histological results corroborated vertical bone growth. This study indicates that Ti-6Al-4V implants fabricated by additive manufacturing to have porosity based on trabecular bone and post-build processing to have micro-/nano-surface roughness can support vertical bone growth in vivo, and suggests that these implants may be used clinically to increase osseointegration in challenging patient cases.

CD31+ Cells From Peripheral Blood Facilitate Bone Regeneration in Biologically Impaired Conditions Through Combined Effects on Immunomodulation and Angiogenesis.

PubMed

Sass, F Andrea; Schmidt-Bleek, Katharina; Ellinghaus, Agnes; Filter, Sebastian; Rose, Alexander; Preininger, Bernd; Reinke, Simon; Geissler, Sven; Volk, Hans-Dieter; Duda, Georg N; Dienelt, Anke

2017-05-01

Controlled revascularization and inflammation are key elements regulating endogenous regeneration after (bone) tissue trauma. Peripheral blood-derived cell subsets, such as regulatory T-helper cells and circulating (endothelial) progenitor cells, respectively, can support endogenous tissue healing, whereas effector T cells that are associated with an aged immune system can hinder bone regeneration. CD31 is expressed by diverse leukocytes and is well recognized as a marker of circulating endothelial (precursor) cells; however, CD31 is absent from the surface of differentiated effector T cells. Thus, we hypothesized that by separating the inhibitory fractions from the supportive fractions of circulating cells within the peripheral blood (PB) using the CD31 marker, bone regeneration in biologically compromised conditions, such as those observed in aged patients, could be improved. In support of our hypothesis, we detected an inverse correlation between CD31+ cells and effector T cells in the hematomas of human fracture patients, dependent on the age of the patient. Furthermore, we demonstrated the regenerative capacity of human PB-CD31+ cells in vitro. These findings were translated to a clinically relevant rat model of impaired bone healing. The transplantation of rat PB-CD31+ cells advanced bone tissue restoration in vivo and was associated with an early anti-inflammatory response, the stimulation of (re)vascularization, and reduced fibrosis. Interestingly, the depletion or enrichment of the highly abundant CD31+/14+ monocytes from the mixed CD31+ cell population diminished tissue regeneration at different levels, suggesting combined effects within the PB-CD31+ subsets. In summary, an intraoperative enrichment of PB-CD31+ cells might be a novel option to facilitate endogenous regeneration under biologically impaired situations by supporting immunomodulation and vascularization. © 2016 American Society for Bone and Mineral Research. © 2016 American Society for Bone and Mineral Research.

Local delivery of controlled-release simvastatin to improve the biocompatibility of polyethylene terephthalate artificial ligaments for reconstruction of the anterior cruciate ligament.

PubMed

Zhang, Peng; Han, Fei; Li, Yunxia; Chen, Jiwu; Chen, Tianwu; Zhi, Yunlong; Jiang, Jia; Lin, Chao; Chen, Shiyi; Zhao, Peng

2016-01-01

The Ligament Advanced Reinforcement System has recently been widely used as the primary graft of choice in anterior cruciate ligament (ACL) reconstruction. But the biological graft-bone healing still remains a problem. Previous studies have shown that simvastatin (SIM) stimulates bone formation. The objective of this study was to investigate whether surface coating with collagen containing low-dose SIM microsphere could enhance the surface biocompatibility of polyethylene terephthalate (PET) artificial ligaments to accelerate graft-to-bone healing. The in vitro studies demonstrated that bone marrow stromal cells on the collagen-coated PET scaffolds (COL/PET) and simvastatin/collagen-coated PET scaffolds (SIM/COL/PET) proliferated vigorously. Compared with the PET group and the COL/PET group, SIM could induce bone marrow stromal cells' osteoblastic differentiation, high alkaline phosphatase activity, more mineralization deposition, and more expression of osteoblast-related genes, such as osteocalcin, runt-related transcription factor 2, bone morphogenetic protein-2, and vascular endothelial growth factor, in the SIM/COL/PET group. In vivo, rabbits received ACL reconstruction with different scaffolds. Histological analysis demonstrated that graft-bone healing was significantly greater with angiogenesis and osteogenesis in the SIM/COL/PET group than the other groups. In addition, biomechanical testing at the eighth week demonstrated a significant increase in the ultimate failure load and stiffness in the SIM/COL/PET group. The low dose of SIM-sustained release from SIM/COL/PET promoted the graft-bone healing via its effect on both angiogenesis and osteogenesis. This study suggested that collagen containing low-dose SIM microsphere coating on the surface of PET artificial ligaments could be potentially applied for ACL reconstruction.

Hierarchically engineered fibrous scaffolds for bone regeneration

PubMed Central

Sachot, Nadège; Castaño, Oscar; Mateos-Timoneda, Miguel A.; Engel, Elisabeth; Planell, Josep A.

2013-01-01

Surface properties of biomaterials play a major role in the governing of cell functionalities. It is well known that mechanical, chemical and nanotopographic cues, for example, influence cell proliferation and differentiation. Here, we present a novel coating protocol to produce hierarchically engineered fibrous scaffolds with tailorable surface characteristics, which mimic bone extracellular matrix. Based on the sol–gel method and a succession of surface treatments, hollow electrospun polylactic acid fibres were coated with a silicon–calcium–phosphate bioactive organic–inorganic glass. Compared with pure polymeric fibres that showed a completely smooth surface, the coated fibres exhibited a nanostructured topography and greater roughness. They also showed improved hydrophilic properties and a Young's modulus sixfold higher than non-coated ones, while remaining fully flexible and easy to handle. Rat mesenchymal stem cells cultured on these fibres showed great cellular spreading and interactions with the material. This protocol can be transferred to other structures and glasses, allowing the fabrication of various materials with well-defined features. This novel approach represents therefore a valuable improvement in the production of artificial matrices able to direct stem cell fate through physical and chemical interactions. PMID:23985738

Effects of hypergravity on immunologic function

NASA Technical Reports Server (NTRS)

Sonnenfeld, G.; Koebel, D. A.; Davis, S.

1995-01-01

The purpose of this study was to compare the effects of hypergravity exposure (2g) with those of exposure to space flight in the Cosmos 2044 flight. To do so, rats were centrifuged continuously for 14 days. Two different experiments were carried out on tissue obtained from the centrifuged rats. In the first experiment, rat bone marrow cells were examined for their response to recombinant murine colony stimulating factor-granulocyte/monocyte (GM-CSF). In the second experiment, rat spleen and bone marrow cells were stained in with a variety of antibodies directed against cell surface antigenic markers. These cells were preserved and analyzed on a flow cytometer. The results of the studies indicated that bone marrow cells from centrifuged rats showed no significant change in response to GM-CSF as compared to bone marrow cells from control rats. Spleen cells from flown rats showed some statistically significant changes in leukocytes subset distribution, but no differences that appeared to be of biological significance. These results indicate that hypergravity did not greatly affect the same immunological parameters affected by space flight in the Cosmos 2044 mission.

Effects of hypergravity on immunologic function

NASA Technical Reports Server (NTRS)

Sonnenfeld, G.; Koebel, D. A.; Davis, S.

1994-01-01

The purpose of this study was to compare the effects of hypergravity exposure (2g) with those of exposure to space flight in the Cosmos 2044 flight. To do so, rats were centrifuged continuously for 14 days. Two different experiments were carried out on tissue obtained from the centrifuged rats. In the first experiment, rat bone marrow cells were examined for their response to recombinant murine colony stimulating factor-granulocyte/monocyte (GM-CSF). In the second experiment, rate spleen and bone marrow cells were stained in with a variety of antibodies directed against cell surface antigenic markers. These cells were preserved and analyzed on a flow cytometer. The results of the studies indicated that bone marrow cells from centrifuged rats showed no significant change in response to GM-CSF as compared to bone marrow cells from control rats. Spleen cells from flown rats showed some statistically significant changes in leukocytes subset distribution, but no differences that appeared to be of biological significance. These results indicate that hypergravity did not greatly affect the same immunological parameters affected by space flight in the Cosmos 2044 mission.

Researching into the cellular shape, volume and elasticity of mesenchymal stem cells, osteoblasts and osteosarcoma cells by atomic force microscopy

PubMed Central

Docheva, Denitsa; Padula, Daniela; Popov, Cvetan; Mutschler, Wolf; Clausen-Schaumann, Hauke; Schieker, Matthias

2008-01-01

Abstract Within the bone lie several different cell types, including osteoblasts (OBs) and mesenchymal stem cells (MSCs). The MSCs are ideal targets for regenerative medicine of bone due to their differentiation potential towards OBs. Human MSCs exhibit two distinct morphologies: rapidly self-renewing cells (RS) and flat cells (FC) with very low proliferation rates. Another cell type found in pathological bone conditions is osteosarcoma. In this study, we compared the topographic and morphometric features of RS and FC cells, human OBs and MG63 osteosarcoma cells by atomic force microscopy (AFM). The results demonstrated clear differences: FC and hOB cells showed similar ruffled topography, whereas RS and MG63 cells exhibited smoother surfaces. Furthermore, we investigated how selected substrates influence cell morphometry. We found that RS and MG63 cells were flatter on fibrous substrates such as polystyrene and collagen I, but much more rounded on glass, the smoothest surface. In contrast, cells with large area, namely FC and hOB cells, did not exhibit pronounced changes in flatness with regards to the different substrates. They were, however, remarkably flatter in comparison to RS and MG63 cells. We could explain the differences in flatness by the extent of adhesion. Indeed, FC and hOB cells showed much higher content of focal adhesions. Finally, we used the AFM to determine the cellular Young's modulus. RS, FC and hOB cells showed comparable stiffness on the three different substrates, while MG63 cells demonstrated the unique feature of increased elasticity on collagen I. In summary, our results show, for the first time, a direct comparison between the morphometric and biophysical features of different human cell types derived from normal and pathological bone. Our study manifests the opinion that along with RNA, proteomic and functional research, morphological and biomechanical characterization of cells also reveals novel cell features and interrelationships. PMID:18419596

Self-assembled monolayers of alendronate on Ti6Al4V alloy surfaces enhance osteogenesis in mesenchymal stem cells

NASA Astrophysics Data System (ADS)

Rojo, Luis; Gharibi, Borzo; McLister, Robert; Meenan, Brian J.; Deb, Sanjukta

2016-07-01

Phosphonates have emerged as an alternative for functionalization of titanium surfaces by the formation of homogeneous self-assembled monolayers (SAMs) via Ti-O-P linkages. This study presents results from an investigation of the modification of Ti6Al4V alloy by chemisorption of osseoinductive alendronate using a simple, effective and clean methodology. The modified surfaces showed a tailored topography and surface chemistry as determined by SEM microscopy and RAMAN spectroscopy. X-ray photoelectron spectroscopy revealed that an effective mode of bonding is created between the metal oxide surface and the phosphate residue of alendronate, leading to formation of homogenous drug distribution along the surface. In-vitro studies showed that alendronate SAMs induce differentiation of hMSC to a bone cell phenotype and promote bone formation on modified surfaces. Here we show that this novel method for the preparation of functional coatings on titanium-based medical devices provides osseoinductive bioactive molecules to promote enhanced integration at the site of implantation.

Self-assembled monolayers of alendronate on Ti6Al4V alloy surfaces enhance osteogenesis in mesenchymal stem cells

PubMed Central

Rojo, Luis; Gharibi, Borzo; McLister, Robert; Meenan, Brian J.; Deb, Sanjukta

2016-01-01

Phosphonates have emerged as an alternative for functionalization of titanium surfaces by the formation of homogeneous self-assembled monolayers (SAMs) via Ti-O-P linkages. This study presents results from an investigation of the modification of Ti6Al4V alloy by chemisorption of osseoinductive alendronate using a simple, effective and clean methodology. The modified surfaces showed a tailored topography and surface chemistry as determined by SEM microscopy and RAMAN spectroscopy. X-ray photoelectron spectroscopy revealed that an effective mode of bonding is created between the metal oxide surface and the phosphate residue of alendronate, leading to formation of homogenous drug distribution along the surface. In-vitro studies showed that alendronate SAMs induce differentiation of hMSC to a bone cell phenotype and promote bone formation on modified surfaces. Here we show that this novel method for the preparation of functional coatings on titanium-based medical devices provides osseoinductive bioactive molecules to promote enhanced integration at the site of implantation. PMID:27468811

Histology of 8 atypical femoral fractures: remodeling but no healing.

PubMed

Schilcher, Jörg; Sandberg, Olof; Isaksson, Hanna; Aspenberg, Per

2014-06-01

The pathophysiology behind bisphosphonate-associated atypical femoral fractures remains unclear. Histological findings at the fracture site itself may provide clues. Between 2008 and 2013, we collected bone biopsies including the fracture line from 4 complete and 4 incomplete atypical femoral fractures. 7 female patients reported continuous bisphosphonate use for 10 years on average. 1 patient was a man who was not using bisphosphonates. Dual-energy X-ray absorptiometry of the hip and spine showed no osteoporosis in 6 cases. The bone biopsies were evaluated by micro-computed tomography, infrared spectroscopy, and qualitative histology. Incomplete fractures involved the whole cortical thickness and showed a continuous gap with a mean width of 180 µm. The gap contained amorphous material and was devoid of living cells. In contrast, the adjacent bone contained living cells, including active osteoclasts. The fracture surfaces sometimes consisted of woven bone, which may have formed in localized defects caused by surface fragmentation or resorption. Atypical femoral fractures show signs of attempted healing at the fracture site. The narrow width of the fracture gap and its necrotic contents are compatible with the idea that micromotion prevents healing because it leads to strains within the fracture gap that preclude cell survival.

The sintered microsphere matrix for bone tissue engineering: in vitro osteoconductivity studies.

PubMed

Borden, Mark; Attawia, Mohamed; Laurencin, Cato T

2002-09-05

A tissue engineering approach has been used to design three-dimensional synthetic matrices for bone repair. The osteoconductivity and degradation profile of a novel polymeric bone-graft substitute was evaluated in an in vitro setting. Using the copolymer poly(lactide-co-glycolide) [PLAGA], a sintering technique based on microsphere technology was used to fabricate three-dimensional porous scaffolds for bone regeneration. Osteoblasts and fibroblasts were seeded onto a 50:50 PLAGA scaffold. Morphologic evaluation through scanning electron microscopy demonstrated that both cell types attached and spread over the scaffold. Cells migrated through the matrix using cytoplasmic extensions to bridge the structure. Cross-sectional images indicated that cellular proliferation had penetrated into the matrix approximately 700 microm from the surface. Examination of the surfaces of cell/matrix constructs demonstrated that cellular proliferation had encompassed the pores of the matrix by 14 days of cell culture. With the aim of optimizing polymer composition and polymer molecular weight, a degradation study was conducted utilizing the matrix. The results demonstrate that degradation of the sintered matrix is dependent on molecular weight, copolymer ratio, and pore volume. From this data, it was determined that 75:25 PLAGA with an initial molecular weight of 100,000 has an optimal degradation profile. These studies show that the sintered microsphere matrix has an osteoconductive structure capable of functioning as a cellular scaffold with a degradation profile suitable for bone regeneration. Copyright 2002 Wiley Periodicals, Inc.

Robotic dispensing of composite scaffolds and in vitro responses of bone marrow stromal cells.

PubMed

Hong, Seok-Jung; Jeong, Ishik; Noh, Kyung-Tae; Yu, Hye-Sun; Lee, Gil-Su; Kim, Hae-Won

2009-09-01

The development of bioactive scaffolds with a designed pore configuration is of particular importance in bone tissue engineering. In this study, bone scaffolds with a controlled pore structure and a bioactive composition were produced using a robotic dispensing technique. A poly(epsilon-caprolactone) (PCL) and hydroxyapatite (HA) composite solution (PCL/HA = 1) was constructed into a 3-dimensional (3D) porous scaffold by fiber deposition and layer-by-layer assembly using a computer-aided robocasting machine. The in vitro tissue cell compatibility was examined using rat bone marrow stromal cells (rBMSCs). The adhesion and growth of cells onto the robotic dispensed scaffolds were observed to be limited by applying the conventional cell seeding technique. However, the initially adhered cells were viable on the scaffold surface. The alkaline phosphatase activity of the cells was significantly higher on the HA-PCL than on the PCL and control culture dish, suggesting that the robotic dispensed HA-PCL scaffold should stimulate the osteogenic differentiation of rBMSCs. Moreover, the expression of a series of bone-associated genes, including alkaline phosphatase and collagen type I, was highly up-regulated on the HA-PCL scaffold as compared to that on the pure PCL scaffold. Overall, the robotic dispensed HA-PCL is considered to find potential use as a bioactive 3D scaffold for bone tissue engineering.

SLM produced porous titanium implant improvements for enhanced vascularization and osteoblast seeding.

PubMed

Matena, Julia; Petersen, Svea; Gieseke, Matthias; Kampmann, Andreas; Teske, Michael; Beyerbach, Martin; Murua Escobar, Hugo; Haferkamp, Heinz; Gellrich, Nils-Claudius; Nolte, Ingo

2015-04-02

To improve well-known titanium implants, pores can be used for increasing bone formation and close bone-implant interface. Selective Laser Melting (SLM) enables the production of any geometry and was used for implant production with 250-µm pore size. The used pore size supports vessel ingrowth, as bone formation is strongly dependent on fast vascularization. Additionally, proangiogenic factors promote implant vascularization. To functionalize the titanium with proangiogenic factors, polycaprolactone (PCL) coating can be used. The following proangiogenic factors were examined: vascular endothelial growth factor (VEGF), high mobility group box 1 (HMGB1) and chemokine (C-X-C motif) ligand 12 (CXCL12). As different surfaces lead to different cell reactions, titanium and PCL coating were compared. The growing into the porous titanium structure of primary osteoblasts was examined by cross sections. Primary osteoblasts seeded on the different surfaces were compared using Live Cell Imaging (LCI). Cross sections showed cells had proliferated, but not migrated after seven days. Although the cell count was lower on titanium PCL implants in LCI, the cell count and cell spreading area development showed promising results for titanium PCL implants. HMGB1 showed the highest migration capacity for stimulating the endothelial cell line. Future perspective would be the incorporation of HMGB1 into PCL polymer for the realization of a slow factor release.

Gene delivery nanocarriers of bioactive glass with unique potential to load BMP2 plasmid DNA and to internalize into mesenchymal stem cells for osteogenesis and bone regeneration

NASA Astrophysics Data System (ADS)

Kim, Tae-Hyun; Singh, Rajendra K.; Kang, Min Sil; Kim, Joong-Hyun; Kim, Hae-Won

2016-04-01

The recent development of bioactive glasses with nanoscale morphologies has spurred their specific applications in bone regeneration, for example as drug and gene delivery carriers. Bone engineering with stem cells genetically modified with this unique class of nanocarriers thus holds great promise in this avenue. Here we report the potential of the bioactive glass nanoparticle (BGN) system for the gene delivery of mesenchymal stem cells (MSCs) targeting bone. The composition of 15% Ca-added silica, proven to be bone-bioactive, was formulated into surface aminated mesoporous nanospheres with enlarged pore sizes, to effectively load and deliver bone morphogenetic protein-2 (BMP2) plasmid DNA. The enlarged mesopores were highly effective in loading BMP2-pDNA with an efficiency as high as 3.5 wt% (pDNA w.r.t. BGN), a level more than twice than for small-sized mesopores. The BGN nanocarriers released the genetic molecules in a highly sustained manner (for as long as 2 weeks). The BMP2-pDNA/BGN complexes were effectively internalized to rat MSCs with a cell uptake level of ~73%, and the majority of cells were transfected to express the BMP2 protein. Subsequent osteogenesis of the transfected MSCs was demonstrated by the expression of bone-related genes, including bone sialoprotein, osteopontin, and osteocalcin. The MSCs transfected with BMP2-pDNA/BGN were locally delivered inside a collagen gel to the target calvarium defects. The results showed significantly improved bone regeneration, as evidenced by the micro-computed tomographic, histomorphometric and immunohistochemical analyses. This study supports the excellent capacity of the BGN system as a pDNA-delivery nanocarrier in MSCs, and the engineered system, BMP2-pDNA/BGN with MSCs, may be considered a new promising candidate to advance the therapeutic potential of stem cells through genetic modification, targeting bone defects and diseases.The recent development of bioactive glasses with nanoscale morphologies has spurred their specific applications in bone regeneration, for example as drug and gene delivery carriers. Bone engineering with stem cells genetically modified with this unique class of nanocarriers thus holds great promise in this avenue. Here we report the potential of the bioactive glass nanoparticle (BGN) system for the gene delivery of mesenchymal stem cells (MSCs) targeting bone. The composition of 15% Ca-added silica, proven to be bone-bioactive, was formulated into surface aminated mesoporous nanospheres with enlarged pore sizes, to effectively load and deliver bone morphogenetic protein-2 (BMP2) plasmid DNA. The enlarged mesopores were highly effective in loading BMP2-pDNA with an efficiency as high as 3.5 wt% (pDNA w.r.t. BGN), a level more than twice than for small-sized mesopores. The BGN nanocarriers released the genetic molecules in a highly sustained manner (for as long as 2 weeks). The BMP2-pDNA/BGN complexes were effectively internalized to rat MSCs with a cell uptake level of ~73%, and the majority of cells were transfected to express the BMP2 protein. Subsequent osteogenesis of the transfected MSCs was demonstrated by the expression of bone-related genes, including bone sialoprotein, osteopontin, and osteocalcin. The MSCs transfected with BMP2-pDNA/BGN were locally delivered inside a collagen gel to the target calvarium defects. The results showed significantly improved bone regeneration, as evidenced by the micro-computed tomographic, histomorphometric and immunohistochemical analyses. This study supports the excellent capacity of the BGN system as a pDNA-delivery nanocarrier in MSCs, and the engineered system, BMP2-pDNA/BGN with MSCs, may be considered a new promising candidate to advance the therapeutic potential of stem cells through genetic modification, targeting bone defects and diseases. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr07933k

Laser bioengineering of glass-titanium implants surface

NASA Astrophysics Data System (ADS)

Lusquiños, F.; Arias-González, F.; Penide, J.; del Val, J.; Comesaña, R.; Quintero, F.; Riveiro, A.; Boutinguiza, M.; Pascual, M. J.; Durán, A.; Pou, J.

2013-11-01

Osseointegration is the mean challenge when surgical treatments fight against load-bearing bone diseases. Absolute bone replacement by a synthetic implant has to be completed not only from the mechanics point of view, but also from a biological approach. Suitable strength, resilience and stress distribution of titanium alloy implants are spoiled by the lack of optimal biological characteristics. The inert quality of extra low interstitial titanium alloy, which make it the most attractive metallic alloy for biomedical applications, oppose to an ideal surface with bone cell affinity, and capable to stimulate bone attachment bone growth. Diverse laser treatments have been proven as effective tools to modify surface properties, such as wettability in contact to physiological fluids, or osteoblast guided and slightly enhanced attachment. The laser surface cladding can go beyond by providing titanium alloy surfaces with osteoconduction and osteoinduction properties. In this research work, the laser radiation is used to produce bioactive glass coatings on Ti6Al4V alloy substrates. Specific silicate bioactive glass compositions has been investigated to achieve suitable surface tension and viscosity temperature behavior during processing, and to provide with the required release of bone growth gene up regulation agents in the course of resorption mediated by physiological fluids. The produced coatings and interfaces, the surface osteoconduction properties, and the chemical species release in simulated physiological fluid were characterized by scanning electron microscopy (SEM), hot stage microscopy (HSM), X-ray diffraction (XRD), X ray fluorescence (XRF), and Fourier transform infrared spectroscopy (FTIR).

Adsorption of Amorphous Silica Nanoparticles onto Hydroxyapatite Surfaces Differentially Alters Surfaces Properties and Adhesion of Human Osteoblast Cells

PubMed Central

Kalia, Priya; Brooks, Roger A.; Kinrade, Stephen D.; Morgan, David J.; Brown, Andrew P.; Rushton, Neil; Jugdaohsingh, Ravin

2016-01-01

Silicon (Si) is suggested to be an important/essential nutrient for bone and connective tissue health. Silicon-substituted hydroxyapatite (Si-HA) has silicate ions incorporated into its lattice structure and was developed to improve attachment to bone and increase new bone formation. Here we investigated the direct adsorption of silicate species onto an HA coated surface as a cost effective method of incorporating silicon on to HA surfaces for improved implant osseointegration, and determined changes in surface characteristics and osteoblast cell adhesion. Plasma-sprayed HA-coated stainless steel discs were incubated in silica dispersions of different concentrations (0–42 mM Si), at neutral pH for 12 h. Adsorbed Si was confirmed by XPS analysis and quantified by ICP-OES analysis following release from the HA surface. Changes in surface characteristics were determined by AFM and measurement of surface wettability. Osteoblast cell adhesion was determined by vinculin plaque staining. Maximum Si adsorption to the HA coated disc occurred after incubation in the 6 mM silica dispersion and decreased progressively with higher silica concentrations, while no adsorption was observed with dispersions below 6 mM Si. Comparison of the Si dispersions that produced the highest and lowest Si adsorption to the HA surface, by TEM-based analysis, revealed an abundance of small amorphous nanosilica species (NSP) of ~1.5 nm in diameter in the 6 mM Si dispersion, with much fewer and larger NSP in the 42 mM Si dispersions. 29Si-NMR confirmed that the NSPs in the 6 mM silica dispersion were polymeric and similar in composition to the larger NSPs in the 42 mM Si dispersion, suggesting that the latter were aggregates of the former. Amorphous NSP adsorbed from the 6 mM dispersion on to a HA-coated disc surface increased the surface’s water contact angle by 53°, whereas that adsorbed from the 42 mM dispersion decreased the contact angle by 18°, indicating increased and decreased hydrophobicity, respectively. AFM showed an increase in surface roughness of the 6 mM Si treated surface, which correlated well with an increase in number of vinculin plaques. These findings suggest that NSP of the right size (relative to charge) adsorb readily to the HA surface, changing the surface characteristics and, thus, improving osteoblast cell adhesion. This treatment provides a simple way to modify plasma-coated HA surfaces that may enable improved osseointegration of bone implants. PMID:26863624

Early osteoblast responses to orthopedic implants: Synergy of surface roughness and chemistry of bioactive ceramic coating.

PubMed

Aniket; Reid, Robert; Hall, Benika; Marriott, Ian; El-Ghannam, Ahmed

2015-06-01

Pro-osteogenic stimulation of bone cells by bioactive ceramic-coated orthopedic implants is influenced by both surface roughness and material chemistry; however, their concomitant impact on osteoblast behavior is not well understood. The aim of this study is to investigate the effects of nano-scale roughness and chemistry of bioactive silica-calcium phosphate nanocomposite (SCPC50) coated Ti-6Al-4V on modulating early bone cell responses. Cell attachment was higher on SCPC50-coated substrates compared to the uncoated controls; however, cells on the uncoated substrate exhibited greater spreading and superior quality of F-actin filaments than cells on the SCPC50-coated substrates. The poor F-actin filament organization on SCPC50-coated substrates is thought to be due to the enhanced calcium uptake by the ceramic surface. Dissolution analyses showed that an increase in surface roughness was accompanied by increased calcium uptake, and increased phosphorous and silicon release, all of which appear to interfere with F-actin assembly and osteoblast morphology. Moreover, cell attachment onto the SCPC50-coated substrates correlated with the known adsorption of fibronectin, and was independent of surface roughness. High-throughput genome sequencing showed enhanced expression of extracellular matrix and cell differentiation related genes. These results demonstrate a synergistic relationship between bioactive ceramic coating roughness and material chemistry resulting in a phenotype that leads to early osteoblast differentiation. © 2014 Wiley Periodicals, Inc.

Effect of heparin and alendronate coating on titanium surfaces on inhibition of osteoclast and enhancement of osteoblast function

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

Moon, Ho-Jin; Yun, Young-Pil; Han, Choong-Wan

2011-09-23

Highlights: {yields} We examine bone metabolism of engineered alendronate attached to Ti surfaces. {yields} Alendronate-immobilized Ti enhances activation of osteoblast differentiation. {yields} Alendronate-immobilized Ti inhibits osteoclast differentiation. {yields} Alendronate-immobilized Ti may be a bioactive implant with dual functions. -- Abstract: The failure of orthopedic and dental implants has been attributed mainly to loosening of the implant from host bone, which may be due to weak bonding of the implant material to bone tissue. Titanium (Ti) is used in the field of orthopedic and dental implants because of its excellent biocompatibility and outstanding mechanical properties. Therefore, in the field of materialsmore » science and tissue engineering, there has been extensive research to immobilize bioactive molecules on the surface of implant materials in order to provide the implants with improved adhesion to the host bone tissue. In this study, chemically active functional groups were introduced on the surface of Ti by a grafting reaction with heparin and then the Ti was functionalized by immobilizing alendronate onto the heparin-grafted surface. In the MC3T3-E1 cell osteogenic differentiation study, the alendronate-immobilized Ti substrates significantly enhanced alkaline phosphatase activity (ALP) and calcium content. Additionally, nuclear factor kappa B ligand (RANKL)-induced osteoclast differentiation of RAW264.7 cells was inhibited with the alendronate-immobilized Ti as confirmed by TRAP analysis. Real time PCR analysis showed that mRNA expressions of osteocalcin and osteopontin, which are markers for osteogenesis, were upregulated in MC3T3-E1 cells cultured on alendronate-immobilized Ti. The mRNA expressions of TRAP and Cathepsin K, markers for osteoclastogenesis, in RAW264.7 cells cultured on alendronate-immobilized Ti were down-regulated. Our study suggests that alendronate-immobilized Ti may be a bioactive implant with dual functions to enhance osteoblast differentiation and to inhibit osteoclast differentiation simultaneously.« less

Is cortical bone hip? What determines cortical bone properties?

PubMed

Epstein, Sol

2007-07-01

Increased bone turnover may produce a disturbance in bone structure which may result in fracture. In cortical bone, both reduction in turnover and increase in hip bone mineral density (BMD) may be necessary to decrease hip fracture risk and may require relatively greater proportionate changes than for trabecular bone. It should also be noted that increased porosity produces disproportionate reduction in bone strength, and studies have shown that increased cortical porosity and decreased cortical thickness are associated with hip fracture. Continued studies for determining the causes of bone strength and deterioration show distinct promise. Osteocyte viability has been observed to be an indicator of bone strength, with viability as the result of maintaining physiological levels of loading and osteocyte apoptosis as the result of a decrease in loading. Osteocyte apoptosis and decrease are major factors in the bone loss and fracture associated with aging. Both the osteocyte and periosteal cell layer are assuming greater importance in the process of maintaining skeletal integrity as our knowledge of these cells expand, as well being a target for pharmacological agents to reduce fracture especially in cortical bone. The bisphosphonate alendronate has been seen to have a positive effect on cortical bone by allowing customary periosteal growth, while reducing the rate of endocortical bone remodeling and slowing bone loss from the endocortical surface. Risedronate treatment effects were attributed to decrease in bone resorption and thus a decrease in fracture risk. Ibandronate has been seen to increase BMD as the spine and femur as well as a reduced incidence of new vertebral fractures and non vertebral on subset post hoc analysis. And treatment with the anabolic agent PTH(1-34) documented modeling and remodelling of quiescent and active bone surfaces. Receptor activator of nuclear factor kappa B ligand (RANKL) plays a key role in bone destruction, and the human monoclonal antibody denosumab binds to RANKL, inhibiting its action and thus improving BMD significantly.

Experiment K-7-23: Effect of Spaceflight on Level and Function of Immune Cells. Part 1; Immunology Studies

NASA Technical Reports Server (NTRS)

Sonnenfeld, G.; Mandel, A.; Konstantinova, I. V.; Berry, W. D.; Taylor, G. R.; Lesnyak, A. T.; Fuchs, B. B.; Rakhmilevich, A. L.

1994-01-01

Two different experiments were carried out in this segment of the immunology protocol for samples received from rats flown on Cosmos 2044. Control groups included vivarium, synchronous and antiorthostatically suspended rats. In the first experiment, rat bone marrow cells were examined in Moscow for their response to recombinant murine colony stimulating factor-granulocyte / monocyte (CSF-GM). In the second experiment, rat spleen and bone marrow cells were stained in Moscow with a variety of antibodies directed against cell surface antigenic markers. These cells were preserved and shipped to the United States for analysis on a flow cytometer. The results of the studies indicated that bone marrow cells from flown and suspended rats showed a decreased response to CSF-GM as compared to bone marrow cells from control rats. Spleen cells from flown rats showed increased percentages of suppressor-cytotoxic-T and helper-T cells amongst the entire cell population. Bone marrow cells showed an increase in the percentage of helper-T cells in the myelogenous population and increased percentages of anti-asialo GM-1 bearing, interleukin-2 receptor bearing, pan-T and helper-T cells in the lymphocytic population. Cell populations from rats suspended antiorthostatically did not follow the same pattern of distribution of leukocytes as cell populations for flown rats. These results are similar, but not identical to, earlier results from Cosmos 1887, and confirm that space flight can have profound effects on immune system components and activities.

Bone marrow mesenchymal stem cell response to nano-structured oxidized and turned titanium surfaces.

PubMed

Annunziata, Marco; Oliva, Adriana; Buosciolo, Antonietta; Giordano, Michele; Guida, Agostino; Guida, Luigi

2012-06-01

The aim of this study was to analyse the topographic features of a novel nano-structured oxidized titanium implant surface and to evaluate its effect on the response of human bone marrow mesenchymal stem cells (BM-MSC) compared with a traditional turned surface. The 10 × 10 × 1 mm turned (control) and oxidized (test) titanium samples (P.H.I. s.r.l.) were examined by scanning electron microscopy (SEM) and atomic force microscopy (AFM) and characterized by height, spatial and hybrid roughness parameters at different dimensional ranges of analysis. Primary cultures of BM-MSC were seeded on titanium samples and cell morphology, adhesion, proliferation and osteogenic differentiation, in terms of alkaline phosphatase activity, osteocalcin synthesis and extracellular matrix mineralization, were evaluated. At SEM and AFM analyses turned samples were grooved, whereas oxidized surfaces showed a more complex micro- and nano-scaled texture, with higher values of roughness parameters. Cell adhesion and osteogenic parameters were greater on oxidized (P<0.05 at least) vs. turned surfaces, whereas the cell proliferation rate was similar on both samples. Although both control and test samples were in the range of average roughness proper of smooth surfaces, they exhibited significantly different topographic properties in terms of height, spatial and, mostly, of hybrid parameters. This different micro- and nano-structure resulted in an enhanced adhesion and differentiation of cells plated onto the oxidized surfaces. © 2011 John Wiley & Sons A/S.

Poly(dopamine) coating to biodegradable polymers for bone tissue engineering.

PubMed

Tsai, Wei-Bor; Chen, Wen-Tung; Chien, Hsiu-Wen; Kuo, Wei-Hsuan; Wang, Meng-Jiy

2014-02-01

In this study, a technique based on poly(dopamine) deposition to promote cell adhesion was investigated for the application in bone tissue engineering. The adhesion and proliferation of rat osteoblasts were evaluated on poly(dopamine)-coated biodegradable polymer films, such as polycaprolactone, poly(l-lactide) and poly(lactic-co-glycolic acid), which are commonly used biodegradable polymers in tissue engineering. Cell adhesion was significantly increased to a plateau by merely 15 s of dopamine incubation, 2.2-4.0-folds of increase compared to the corresponding untreated substrates. Cell proliferation was also greatly enhanced by poly(dopamine) deposition, indicated by shortened cell doubling time. Mineralization was also increased on the poly(dopamine)-deposited surfaces. The potential of poly(dopamine) deposition in bone tissue engineering is demonstrated in this study.

The Cdc42 guanine nucleotide exchange factor FGD6 coordinates cell polarity and endosomal membrane recycling in osteoclasts.

PubMed

Steenblock, Charlotte; Heckel, Tobias; Czupalla, Cornelia; Espírito Santo, Ana Isabel; Niehage, Christian; Sztacho, Martin; Hoflack, Bernard

2014-06-27

The initial step of bone digestion is the adhesion of osteoclasts onto bone surfaces and the assembly of podosomal belts that segregate the bone-facing ruffled membrane from other membrane domains. During bone digestion, membrane components of the ruffled border also need to be recycled after macropinocytosis of digested bone materials. How osteoclast polarity and membrane recycling are coordinated remains unknown. Here, we show that the Cdc42-guanine nucleotide exchange factor FGD6 coordinates these events through its Src-dependent interaction with different actin-based protein networks. At the plasma membrane, FGD6 couples cell adhesion and actin dynamics by regulating podosome formation through the assembly of complexes comprising the Cdc42-interactor IQGAP1, the Rho GTPase-activating protein ARHGAP10, and the integrin interactors Talin-1/2 or Filamin A. On endosomes and transcytotic vesicles, FGD6 regulates retromer-dependent membrane recycling through its interaction with the actin nucleation-promoting factor WASH. These results provide a mechanism by which a single Cdc42-exchange factor controlling different actin-based processes coordinates cell adhesion, cell polarity, and membrane recycling during bone degradation. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

High dietary cholesterol masks type 2 diabetes-induced osteopenia and changes in bone microstructure in rats.

PubMed

Lapmanee, Sarawut; Charoenphandhu, Narattaphol; Aeimlapa, Ratchaneevan; Suntornsaratoon, Panan; Wongdee, Kannikar; Tiyasatkulkovit, Wacharaporn; Kengkoom, Kanchana; Chaimongkolnukul, Khuanjit; Seriwatanachai, Dutmanee; Krishnamra, Nateetip

2014-10-01

Type 2 diabetes mellitus (T2DM) often occurs concurrently with high blood cholesterol or dyslipidemia. Although T2DM has been hypothesized to impair bone microstructure, several investigations showed that, when compared to age-matched healthy individuals, T2DM patients had normal or relatively high bone mineral density (BMD). Since cholesterol and lipids profoundly affect the function of osteoblasts and osteoclasts, it might be cholesterol that obscured the changes in BMD and bone microstructure in T2DM. The present study, therefore, aimed to determine bone elongation, epiphyseal histology, and bone microstructure in non-obese T2DM Goto-Kakizaki rats treated with normal (GK-ND) and high cholesterol diet. We found that volumetric BMD was lower in GK-ND rats than the age-matched wild-type controls. In histomorphometric study of tibial metaphysis, T2DM evidently suppressed osteoblast function as indicated by decreases in osteoblast surface, mineral apposition rate, and bone formation rate in GK-ND rats. Meanwhile, the osteoclast surface and eroded surface were increased in GK-ND rats, thus suggesting an activation of bone resorption. T2DM also impaired bone elongation, presumably by retaining the chondrogenic precursor cells in the epiphyseal resting zone. Interestingly, several bone changes in GK rats (e.g., increased osteoclast surface) disappeared after high cholesterol treatment as compared to wild-type rats fed high cholesterol diet. In conclusion, high cholesterol diet was capable of masking the T2DM-induced osteopenia and changes in several histomorphometric parameters that indicated bone microstructural defect. Cholesterol thus explained, in part, why a decrease in BMD was not observed in T2DM, and hence delayed diagnosis of the T2DM-associated bone disease.

Graphene coating on the surface of CoCrMo alloy enhances the adhesion and proliferation of bone marrow mesenchymal stem cells.

PubMed

Zhang, Qi; Li, Kewen; Yan, Jinhong; Wang, Zhuo; Wu, Qi; Bi, Long; Yang, Min; Han, Yisheng

2018-03-18

The objective was to investigate whether a graphene coating could improve the surface bioactivity of a cobalt-chromium-molybdenum-based alloy (CoCrMo). Graphene was produced by chemical vapor deposition and transferred to the surface of the CoCrMo alloy using an improved wet transfer approach. The morphology of the samples was observed, and the adhesion force and stabilization of graphene coating were analyzed by a nanoscratch test and ultrasonication test. In an in vitro study, the adhesion and proliferation of bone marrow mesenchymal stem cells (BMSCs) cultured on the samples were quantified via an Alamar Blue assay and cell counting kit-8 (CCK-8) assay. The results showed that it is feasible to apply graphene to modify the surface of a CoCrMo alloy, and the enhancement of the adhesion and proliferation of BMSCs was also shown in the present study. In conclusion, graphene exhibits considerable potential for enhancing the surface bioactivity of CoCrMo alloy. Copyright © 2018 Elsevier Inc. 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

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

Implant materials generate different peri-implant inflammatory factors: poly-ether-ether-ketone promotes fibrosis and microtextured titanium promotes osteogenic factors.

PubMed

Olivares-Navarrete, Rene; Hyzy, Sharon L; Slosar, Paul J; Schneider, Jennifer M; Schwartz, Zvi; Boyan, Barbara D

2015-03-15

An in vitro study examining factors produced by human mesenchymal stem cells on spine implant materials. The aim of this study was to examine whether the inflammatory microenvironment generated by cells on titanium-aluminum-vanadium (Ti-alloy, TiAlV) surfaces is affected by surface microtexture and whether it differs from that generated on poly-ether-ether-ketone (PEEK). Histologically, implants fabricated from PEEK have a fibrous connective tissue surface interface whereas Ti-alloy implants demonstrate close approximation with surrounding bone. Ti-alloy surfaces with complex micron/submicron scale roughness promote osteoblastic differentiation and foster a specific cellular environment that favors bone formation whereas PEEK favors fibrous tissue formation. Human mesenchymal stem cells were cultured on tissue culture polystyrene, PEEK, smooth TiAlV, or macro-/micro-/nano-textured rough TiAlV (mmnTiAlV) disks. Osteoblastic differentiation and secreted inflammatory interleukins were assessed after 7 days. Fold changes in mRNAs for inflammation, necrosis, DNA damage, or apoptosis with respect to tissue culture polystyrene were measured by low-density polymerase chain reaction array. Data were analyzed by analysis of variance, followed by Bonferroni's correction of Student's t-test. Cells on PEEK upregulated mRNAs for chemokine ligand-2, interleukin (IL) 1β, IL6, IL8, and tumor necrosis factor. Cells grown on the mmnTiAlV had an 8-fold reduction in mRNAs for toll-like receptor-4. Cells grown on mmnTiAlV had reduced levels of proinflammatory interleukins. Cells on PEEK had higher mRNAs for factors strongly associated with cell death/apoptosis, whereas cells on mmnTiAlV exhibited reduced cytokine factor levels. All results were significant (P < 0.05). These results suggest that fibrous tissue around PEEK implants may be due to several factors: reduced osteoblastic differentiation of progenitor cells and production of an inflammatory environment that favors cell death via apoptosis and necrosis. Ti alloy surfaces with complex macro/micro/nanoscale roughness promote osteoblastic differentiation and foster a specific cellular environment that favors bone formation. N/A.

In vitro characterization of MG-63 osteoblast-like cells cultured on organic-inorganic lyophilized gelatin sponges for early bone healing.

PubMed

Rodriguez, Isaac A; Saxena, Gunjan; Hixon, Katherine R; Sell, Scott A; Bowlin, Gary L

2016-08-01

The development of three-dimensional porous scaffolds with enhanced osteogenic and angiogenic potential would be beneficial for inducing early-stage bone regeneration. Previous studies have demonstrated the advantages of mineralized and nonmineralized acellular 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC) cross-linked gelatin sponges enhanced with preparations rich in growth factors, hydroxyapatite, and chitin whiskers. In this study, those same scaffolds were mineralized and dynamically seeded with MG-63 cells. Cell proliferation, protein/cytokine secretion, and compressive mechanical properties of scaffolds were evaluated. It was found that mineralization and the addition of growth factors increased cell proliferation compared to gelatin controls. Cells on all scaffolds responded in an appropriate bone regenerative fashion as shown through osteocalcin secretion and little to no secretion of bone resorbing markers. However, compressive mechanical properties of cellularized scaffolds were not significantly different from acellular scaffolds. The combined results of increased cellular attachment, infiltration, and bone regenerative protein/cytokine secretion on scaffolds support the need for the addition of a bone-like mineral surface. Cellularized scaffolds containing growth factors reported similar advantages and mechanical values in the range of native tissues present in the early stages of bone healing. These results suggest that the developed composite sponges exhibited cellular responses and mechanical properties appropriate for promoting early bone healing in various applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2011-2019, 2016. © 2016 Wiley Periodicals, Inc.

Numerical simulation of fluid field and in vitro three-dimensional fabrication of tissue-engineered bones in a rotating bioreactor and in vivo implantation for repairing segmental bone defects.

PubMed

Song, Kedong; Wang, Hai; Zhang, Bowen; Lim, Mayasari; Liu, Yingchao; Liu, Tianqing

2013-03-01

In this paper, two-dimensional flow field simulation was conducted to determine shear stresses and velocity profiles for bone tissue engineering in a rotating wall vessel bioreactor (RWVB). In addition, in vitro three-dimensional fabrication of tissue-engineered bones was carried out in optimized bioreactor conditions, and in vivo implantation using fabricated bones was performed for segmental bone defects of Zelanian rabbits. The distribution of dynamic pressure, total pressure, shear stress, and velocity within the culture chamber was calculated for different scaffold locations. According to the simulation results, the dynamic pressure, velocity, and shear stress around the surface of cell-scaffold construction periodically changed at different locations of the RWVB, which could result in periodical stress stimulation for fabricated tissue constructs. However, overall shear stresses were relatively low, and the fluid velocities were uniform in the bioreactor. Our in vitro experiments showed that the number of cells cultured in the RWVB was five times higher than those cultured in a T-flask. The tissue-engineered bones grew very well in the RWVB. This study demonstrates that stress stimulation in an RWVB can be beneficial for cell/bio-derived bone constructs fabricated in an RWVB, with an application for repairing segmental bone defects.

Deletion of Adseverin in Osteoclasts Affects Cell Structure But Not Bone Metabolism.

PubMed

Cao, Yixuan; Wang, Yongqiang; Sprangers, Sara; Picavet, Daisy I; Glogauer, Michael; McCulloch, Christopher A; Everts, Vincent

2017-08-01

Adseverin is an actin-severing/capping protein that may contribute to osteoclast differentiation in vitro but its role in bone remodeling of healthy animals is not defined. We analyzed bone and osteoclast structure in adseverin conditional null mice at alveolar and long bone sites. In wild-type and adseverin null mice, as measured by dual-energy X-ray absorptiometry, there were no differences of bone mineral content or bone mineral density, indicating no change of bone metabolism. In tibiae, TRAcP + osteoclasts were formed in comparable numbers in adseverin null and wild-type mice. Ultrastructural analysis showed normal and similar abundance of ruffled borders, sealing zones, and mitochondria, and with no difference of osteoclast nuclear numbers. In contrast, analyses of long bone showed that in the absence of adseverin osteoclasts were smaller (120 ± 13 vs. 274 ± 19 µm 2 ; p < 0.05), as were nuclear size and the surface area of cytoplasm. The nuclei of adseverin null osteoclasts exhibited more heterochromatin (31 ± 3%) than wild-type cells (8 ± 1%), suggesting that adseverin affects cell differentiation. The data indicate that in healthy, developing tissues, adseverin contributes to the regulation of osteoclast structure but not to bone metabolism in vivo.

Biomimetic Multispiked Connecting Ti-Alloy Scaffold Prototype for Entirely-Cementless Resurfacing Arthroplasty Endoprostheses-Exemplary Results of Implantation of the Ca-P Surface-Modified Scaffold Prototypes in Animal Model and Osteoblast Culture Evaluation.

PubMed

Uklejewski, Ryszard; Rogala, Piotr; Winiecki, Mariusz; Tokłowicz, Renata; Ruszkowski, Piotr; Wołuń-Cholewa, Maria

2016-06-29

We present here-designed, manufactured, and tested by our research team-the Ti-alloy prototype of the multispiked connecting scaffold (MSC-Scaffold) interfacing the components of resurfacing arthroplasty (RA) endoprostheses with bone. The spikes of the MSC-Scaffold prototype mimic the interdigitations of the articular subchondral bone, which is the natural biostructure interfacing the articular cartilage with the periarticular trabecular bone. To enhance the osteoinduction/osteointegration potential of the MSC-Scaffold, the attempts to modify its bone contacting surfaces by the process of electrochemical cathodic deposition of Ca-P was performed with further immersion of the MSC-Scaffold prototypes in SBF in order to transform the amorphous calcium-phosphate coating in hydroxyapatite-like (HA-like) coating. The pilot experimental study of biointegration of unmodified and Ca-P surface-modified MSC-Scaffold prototypes was conducted in an animal model (swine) and in osteoblast cell culture. On the basis of a microscope-histological method the biointegration was proven by the presence of trabeculae in the interspike spaces of the MSC-Scaffold prototype on longitudinal and cross-sections of bone-implant specimens. The percentage of trabeculae in the area between the spikes of specimen containing Ca-P surface modified scaffold prototype observed in microCT reconstructions of the explanted joints was visibly higher than in the case of unmodified MSC-Scaffold prototypes. Significantly higher Alkaline Phosphatase (ALP) activity and the cellular proliferation in the case of Ca-P-modified MSC-Scaffold pre-prototypes, in comparison with unmodified pre-prototypes, was found in osteoblast cell cultures. The obtained results of experimental implantation in an animal model and osteoblast cell culture evaluations of Ca-P surface-modified and non-modified biomimetic MSC-Scaffold prototypes for biomimetic entirely-cementless RA endoprostheses indicate the enhancement of the osteoinduction/osteointegration potential by the Ca-P surface modification of the Ti-alloy MSC-Scaffold prototype. Planned further research on the prototype of this biomimetic MSC-Scaffold for a new generation of RA endoprostheses is also given.

Biomimetic Multispiked Connecting Ti-Alloy Scaffold Prototype for Entirely-Cementless Resurfacing Arthroplasty Endoprostheses—Exemplary Results of Implantation of the Ca-P Surface-Modified Scaffold Prototypes in Animal Model and Osteoblast Culture Evaluation

PubMed Central

Uklejewski, Ryszard; Rogala, Piotr; Winiecki, Mariusz; Tokłowicz, Renata; Ruszkowski, Piotr; Wołuń-Cholewa, Maria

2016-01-01

We present here—designed, manufactured, and tested by our research team—the Ti-alloy prototype of the multispiked connecting scaffold (MSC-Scaffold) interfacing the components of resurfacing arthroplasty (RA) endoprostheses with bone. The spikes of the MSC-Scaffold prototype mimic the interdigitations of the articular subchondral bone, which is the natural biostructure interfacing the articular cartilage with the periarticular trabecular bone. To enhance the osteoinduction/osteointegration potential of the MSC-Scaffold, the attempts to modify its bone contacting surfaces by the process of electrochemical cathodic deposition of Ca-P was performed with further immersion of the MSC-Scaffold prototypes in SBF in order to transform the amorphous calcium-phosphate coating in hydroxyapatite-like (HA-like) coating. The pilot experimental study of biointegration of unmodified and Ca-P surface-modified MSC-Scaffold prototypes was conducted in an animal model (swine) and in osteoblast cell culture. On the basis of a microscope-histological method the biointegration was proven by the presence of trabeculae in the interspike spaces of the MSC-Scaffold prototype on longitudinal and cross-sections of bone-implant specimens. The percentage of trabeculae in the area between the spikes of specimen containing Ca-P surface modified scaffold prototype observed in microCT reconstructions of the explanted joints was visibly higher than in the case of unmodified MSC-Scaffold prototypes. Significantly higher Alkaline Phosphatase (ALP) activity and the cellular proliferation in the case of Ca-P-modified MSC-Scaffold pre-prototypes, in comparison with unmodified pre-prototypes, was found in osteoblast cell cultures. The obtained results of experimental implantation in an animal model and osteoblast cell culture evaluations of Ca-P surface-modified and non-modified biomimetic MSC-Scaffold prototypes for biomimetic entirely-cementless RA endoprostheses indicate the enhancement of the osteoinduction/osteointegration potential by the Ca-P surface modification of the Ti-alloy MSC-Scaffold prototype. Planned further research on the prototype of this biomimetic MSC-Scaffold for a new generation of RA endoprostheses is also given. PMID:28773652

In vitro electromagnetically stimulated SAOS-2 osteoblasts inside porous hydroxyapatite

PubMed Central

Fassina, Lorenzo; Saino, Enrica; Sbarra, Maria Sonia; Visai, Livia; De Angelis, Maria Gabriella Cusella; Magenes, Giovanni; Benazzo, Francesco

2009-01-01

One of the key challenges in reconstructive bone surgery is to provide living constructs that possess the ability to integrate in the surrounding tissue. Bone graft substitutes, such as autografts, allografts, xenografts, and biomaterials have been widely used to heal critical-size long bone defects due to trauma, tumor resection, congenital deformity, and tissue degeneration. In particular, porous hydroxyapatite is widely used in reconstructive bone surgery owing to its biocompatibility. In addition, the in vitro modification of hydroxyapatite with osteogenic signals enhances the tissue regeneration in vivo, suggesting that the biomaterial modification could play an important role in tissue engineering. In this study we have followed a biomimetic strategy where electromagnetically stimulated SAOS-2 human osteoblasts proliferated and built their extracellular matrix inside a porous hydroxyapatite scaffold. The electromagnetic stimulus had the following parameters: intensity of the magnetic field equal to 2 mT, amplitude of the induced electric tension equal to 5 mV, frequency of 75 Hz, and pulse duration of 1.3 ms. In comparison with control conditions, the electromagnetic stimulus increased the cell proliferation and the surface coating with bone proteins (decorin, osteocalcin, osteopontin, type-I collagen, and type-III collagen). The physical stimulus aimed at obtaining a better modification of the biomaterial internal surface in terms of cell colonization and coating with bone matrix. PMID:19827111

Surface modification of nano-silica on the ligament advanced reinforcement system for accelerated bone formation: primary human osteoblasts testing in vitro and animal testing in vivo

NASA Astrophysics Data System (ADS)

Li, Mengmeng; Wang, Shiwen; Jiang, Jia; Sun, Jiashu; Li, Yuzhuo; Huang, Deyong; Long, Yun-Ze; Zheng, Wenfu; Chen, Shiyi; Jiang, Xingyu

2015-04-01

The Ligament Advanced Reinforcement System (LARS) has been considered as a promising graft for ligament reconstruction. To improve its biocompatibility and effectiveness on new bone formation, we modified the surface of a polyethylene terephthalate (PET) ligament with nanoscale silica using atom transfer radical polymerization (ATRP) and silica polymerization. The modified ligament is tested by both in vitro and in vivo experiments. Human osteoblast testing in vitro exhibits an ~21% higher value in cell viability for silica-modified grafts compared with original grafts. Animal testing in vivo shows that there is new formed bone in the case of a nanoscale silica-coated ligament. These results demonstrate that our approach for nanoscale silica surface modification on LARS could be potentially applied for ligament reconstruction.The Ligament Advanced Reinforcement System (LARS) has been considered as a promising graft for ligament reconstruction. To improve its biocompatibility and effectiveness on new bone formation, we modified the surface of a polyethylene terephthalate (PET) ligament with nanoscale silica using atom transfer radical polymerization (ATRP) and silica polymerization. The modified ligament is tested by both in vitro and in vivo experiments. Human osteoblast testing in vitro exhibits an ~21% higher value in cell viability for silica-modified grafts compared with original grafts. Animal testing in vivo shows that there is new formed bone in the case of a nanoscale silica-coated ligament. These results demonstrate that our approach for nanoscale silica surface modification on LARS could be potentially applied for ligament reconstruction. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr01439e

TiO2 -enriched polymeric powder coatings support human mesenchymal cell spreading and osteogenic differentiation.

PubMed

Mozumder, Mohammad Sayem; Zhu, Jesse; Perinpanayagam, Hiran

2011-06-01

Novel polymeric powder coatings (PPC) were prepared by ultrafine powder coating technology and shown to support human mesenchymal cell attachment and growth. PPC surfaces enriched with nano-TiO(2) (nTiO(2)) showed enhanced cellular responses, and were compared to commercially pure titanium (cpTi). After cell attachment and growth, osteogenic differentiation and bone matrix formation ensures osseointegration for implantable biomaterials. Therefore, the objective of this study was to determine if mesenchymal cells grown on PPC could undergo osteogenic differentiation by inducing Runx2 and bone matrix proteins, and then initiate mineralization. Atomic force microscopy revealed intricate three-dimensional micro-topographies, and the measures of nano-roughness and porosity were similar for all PPC surfaces. Scanning electron microscopy showed that the cells attached and spread out over all of the surfaces. After 1 week in osteogenic media, RT-PCR analysis showed the induction of Runx2, the up-regulation of type I collagen, and the initial detection of alkaline phosphatase and bone sialoprotein. After 4 weeks, Alizarin Red staining showed mineral deposition. However, cell spreading and osteogenic differentiation were significantly (P < 0.05) higher on the cpTi controls than on the PPC surfaces. Furthermore, spreading and differentiation were consistently higher on the titanium-enriched PPC-2, -3 and -4 than on the titanium-free PPC-1. Therefore, despite the presence of complex micro-topographies and nano-features, titanium-enrichment enhanced the cellular response, and pure titanium still provided the best substrate. These findings confirm the cytocompatibility of these novel polymeric coatings and suggest that titanium-enrichment and nTiO(2) additives may enhance their performance.

Bone tissue engineering with human mesenchymal stem cell sheets constructed using magnetite nanoparticles and magnetic force.

PubMed

Shimizu, Kazunori; Ito, Akira; Yoshida, Tatsuro; Yamada, Yoichi; Ueda, Minoru; Honda, Hiroyuki

2007-08-01

An in vitro reconstruction of three-dimensional (3D) tissues without the use of scaffolds may be an alternative strategy for tissue engineering. We have developed a novel tissue engineering strategy, termed magnetic force-based tissue engineering (Mag-TE), in which magnetite cationic liposomes (MCLs) with a positive charge at the liposomal surface, and magnetic force were used to construct 3D tissue without scaffolds. In this study, human mesenchymal stem cells (MSCs) magnetically labeled with MCLs were seeded onto an ultra-low attachment culture surface, and a magnet (4000 G) was placed on the reverse side. The MSCs formed multilayered sheet-like structures after a 24-h culture period. MSCs in the sheets constructed by Mag-TE maintained an in vitro ability to differentiate into osteoblasts, adipocytes, or chondrocytes after a 21-day culture period using each induction medium. Using an electromagnet, MSC sheets constructed by Mag-TE were harvested and transplanted into the bone defect in the crania of nude rats. Histological observation revealed that new bone surrounded by osteoblast-like cells was formed in the defect area 14 days after transplantation with MSC sheets, whereas no bone formation was observed in control rats without the transplant. These results indicated that Mag-TE could be used for the transplantation of MSC sheets using magnetite nanoparticles and magnetic force, providing novel methodology for bone tissue engineering.

The biocompatibility of dense and porous Nickel-Titanium produced by selective laser melting.

PubMed

Habijan, T; Haberland, C; Meier, H; Frenzel, J; Wittsiepe, J; Wuwer, C; Greulich, C; Schildhauer, T A; Köller, M

2013-01-01

Nickel-Titanium shape memory alloys (NiTi-SMA) are of biomedical interest due to their unusual range of pure elastic deformability and their elastic modulus, which is closer to that of bone than any other metallic or ceramic material. Newly developed porous NiTi, produced by Selective Laser Melting (SLM), is currently under investigation as a potential carrier material for human mesenchymal stem cells (hMSC). SLM enables the production of highly complex and tailor-made implants for patients on the basis of CT data. Such implants could be used for the reconstruction of the skull, face, or pelvis. hMSC are a promising cell type for regenerative medicine and tissue engineering due to their ability to support the regeneration of critical size bone defects. Loading porous SLM-NiTi implants with autologous hMSC may enhance bone growth and healing for critical bone defects. The purpose of this study was to assess whether porous SLM-NiTi is a suitable carrier for hMSC. Specimens of varying porosity and surface structure were fabricated via SLM. hMSC were cultured for 8 days on NiTi specimens, and cell viability was analyzed using two-color fluorescence staining. Viable cells were detected on all specimens after 8 days of cell culture. Cell morphology and surface topography were analyzed by scanning electron microscopy (SEM). Cell morphology and surface topology were dependent on the orientation of the specimens during SLM production. The Nickel ion release can be reduced significantly by aligned laser processing conditions. The presented results clearly attest that both dense SLM-NiTi and porous SLM-NiTi are suitable carriers for hMSC. Nevertheless, before carrying out in vivo studies, some work on optimization of the manufacturing process and post-processing is required. Copyright © 2012 Elsevier B.V. All rights reserved.

Demineralized dentin matrix composite collagen material for bone tissue regeneration.

PubMed

Li, Jianan; Yang, Juan; Zhong, Xiaozhong; He, Fengrong; Wu, Xiongwen; Shen, Guanxin

2013-01-01

Demineralized dentin matrix (DDM) had been successfully used in clinics as bone repair biomaterial for many years. However, particle morphology of DDM limited it further applications. In this study, DDM and collagen were prepared to DDM composite collagen material. The surface morphology of the material was studied by scanning electron microscope (SEM). MC3T3-E1 cells responses in vitro and tissue responses in vivo by implantation of DDM composite collagen material in bone defect of rabbits were also investigated. SEM analysis showed that DDM composite collagen material evenly distributed and formed a porous scaffold. Cell culture and animal models results indicated that DDM composite collagen material was biocompatible and could support cell proliferation and differentiation. Histological evaluation showed that DDM composite collagen material exhibited good biocompatibility, biodegradability and osteoconductivity with host bone in vivo. The results suggested that DDM composite collagen material might have a significant clinical advantage and potential to be applied in bone and orthopedic surgery.

Impact of non-thermal plasma surface modification on porous calcium hydroxyapatite ceramics for bone regeneration

PubMed Central

Moriguchi, Yu; Lee, Dae-Sung; Thamina, Khair; Masuda, Kazuto; Itsuki, Dai; Yoshikawa, Hideki; Hamaguchi, Satoshi; Myoui, Akira

2018-01-01

In the physiochemical sciences, plasma is used to describe an ionized gas. Previous studies have implicated plasma surface treatment in the enhancement of hydrophilicity of implanted musculoskeletal reconstructive materials. Hydroxyapatite (HA) ceramics, widely used in bone tissue regeneration, have made great advancements to skeletal surgery. In the present study, we investigate the impact of low-pressure plasma on the interconnected porous calcium hydroxyapatite (IP-CHA) both in vitro and in vivo. Our results indicate that dielectric barrier discharge (DBD) plasma, when used with oxygen, can augment the hydrophilicity of non-porous HA surfaces and the osteoconductivity of the IP-CHA disc via increased water penetration of inner porous structures, as demonstrated through microfocus computed tomography (μCT) assay. In vivo implantation of plasma-treated IP-CHA displayed superior bone ingrowth than untreated IP-CHA. Though plasma-treated IP-CHA did not alter osteoblast cell proliferation, it accelerated osteogenic differentiation of seeded marrow mesenchymal stem cells. In vitro X-ray photoelectron spectroscopy (XPS) revealed that this plasma treatment increases levels of oxygen, rather than nitrogen, on the plasma-treated IP-CHA surface. These findings suggest that plasma treatment, an easy and simple processing, can significantly improve the osteoconductive potential of commonly used artificial bones such as IP-CHA. Further optimization of plasma treatment and longer-term follow-up of in vivo application are required toward its clinical application. PMID:29538457

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.

Three-Dimensional Printing of Hollow-Struts-Packed Bioceramic Scaffolds for Bone Regeneration.

PubMed

Luo, Yongxiang; Zhai, Dong; Huan, Zhiguang; Zhu, Haibo; Xia, Lunguo; Chang, Jiang; Wu, Chengtie

2015-11-04

Three-dimensional printing technologies have shown distinct advantages to create porous scaffolds with designed macropores for application in bone tissue engineering. However, until now, 3D-printed bioceramic scaffolds only possessing a single type of macropore have been reported. Generally, those scaffolds with a single type of macropore have relatively low porosity and pore surfaces, limited delivery of oxygen and nutrition to surviving cells, and new bone tissue formation in the center of the scaffolds. Therefore, in this work, we present a useful and facile method for preparing hollow-struts-packed (HSP) bioceramic scaffolds with designed macropores and multioriented hollow channels via a modified coaxial 3D printing strategy. The prepared HSP scaffolds combined high porosity and surface area with impressive mechanical strength. The unique hollow-struts structures of bioceramic scaffolds significantly improved cell attachment and proliferation and further promoted formation of new bone tissue in the center of the scaffolds, indicating that HSP ceramic scaffolds can be used for regeneration of large bone defects. In addition, the strategy can be used to prepare other HSP ceramic scaffolds, indicating a universal application for tissue engineering, mechanical engineering, catalysis, and environmental materials.

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

Surface Modification of Porous Titanium Granules for Improving Bioactivity.

PubMed

Karaji, Zahra Gorgin; Houshmand, Behzad; Faghihi, Shahab

The highly porous titanium granules are currently being used as bone substitute material and for bone tissue augmentation. However, they suffer from weak bone bonding ability. The aim of this study was to create a nanostructured surface oxide layer on irregularly shaped titanium granules to improve their bioactivity. This could be achieved using optimized electrochemical anodic oxidation (anodizing) and heat treatment processes. The anodizing process was done in an ethylene glycol-based electrolyte at an optimized condition of 60 V for 3 hours. The anodized granules were subsequently annealed at 450°C for 1 hour. Scanning electron microscopy (SEM), energy-dispersive x-ray spectroscopy (EDS), and x-ray diffraction (XRD) were used to characterize the surface structure and morphology of the granules. The in vitro bioactivity of the samples was evaluated by immersion of specimens in simulated body fluid (SBF) for 1, 2, and 3 weeks. The human osteoblastic sarcoma cell line, MG63, was used to evaluate cell viability on the samples using dimethylthiazol-diphenyl tetrazolium bromide (MTT) assay. The results demonstrated the formation of amorphous nanostructured titanium oxide after anodizing, which transformed to crystalline anatase and rutile phases upon heat treatment. After immersion in SBF, spherical aggregates of amorphous calcium phosphate were formed on the surface of the anodized sample, which turned into crystalline hydroxyapatite on the surface of the anodized annealed sample. No cytotoxicity was detected among the samples. It is suggested that anodic oxidation followed by heat treatment could be used as an effective surface treatment procedure to improve bioactivity of titanium granules implemented for bone tissue repair and augmentation.

Reprogramming hMSCs morphology with silicon/porous silicon geometric micro-patterns.

PubMed

Ynsa, M D; Dang, Z Y; Manso-Silvan, M; Song, J; Azimi, S; Wu, J F; Liang, H D; Torres-Costa, V; Punzon-Quijorna, E; Breese, M B H; Garcia-Ruiz, J P

2014-04-01

Geometric micro-patterned surfaces of silicon combined with porous silicon (Si/PSi) have been manufactured to study the behaviour of human Mesenchymal Stem Cells (hMSCs). These micro-patterns consist of regular silicon hexagons surrounded by spaced columns of silicon equilateral triangles separated by PSi. The results show that, at an early culture stage, the hMSCs resemble quiescent cells on the central hexagons with centered nuclei and actin/β-catenin and a microtubules network denoting cell adhesion. After 2 days, hMSCs adapted their morphology and cytoskeleton proteins from cell-cell dominant interactions at the center of the hexagonal surface. This was followed by an intermediate zone with some external actin fibres/β-catenin interactions and an outer zone where the dominant interactions are cell-silicon. Cells move into silicon columns to divide, migrate and communicate. Furthermore, results show that Runx2 and vitamin D receptors, both specific transcription factors for skeleton-derived cells, are expressed in cells grown on micropatterned silicon under all observed circumstances. On the other hand, non-phenotypic alterations are under cell growth and migration on Si/PSi substrates. The former consideration strongly supports the use of micro-patterned silicon surfaces to address pending questions about the mechanisms of human bone biogenesis/pathogenesis and the study of bone scaffolds.

Multiphysics of bone remodeling: A 2D mesoscale activation simulation.

PubMed

Spingarn, C; Wagner, D; Rémond, Y; George, D

2017-01-01

In this work, we present an evolutive trabecular model for bone remodeling based on a boundary detection algorithm accounting for both biology and applied mechanical forces, known to be an important factor in bone evolution. A finite element (FE) numerical model using the Abaqus/Standard® software was used with a UMAT subroutine to solve the governing coupled mechanical-biological non-linear differential equations of the bone evolution model. The simulations present cell activation on a simplified trabeculae configuration organization with trabecular thickness of 200µm. For this activation process, the results confirm that the trabeculae are mainly oriented in the active direction of the principal mechanical stresses and according to the principal applied mechanical load directions. The trabeculae surface activation is clearly identified and can provide understanding of the different bone cell activations in more complex geometries and load conditions.

Ultrastructural analysis of bone nodules formed in vitro by isolated fetal rat calvaria cells

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

Bhargava, U.; Bar-Lev, M.; Bellows, C.G.

When cells enzymatically digested from 21 d fetal rat calvaria are grown in ascorbic acid and Na beta-glycerophosphate, they form discrete three-dimensional nodular structures with the histological and immunohistochemical appearance of woven bone. The present investigation was undertaken to verify that bone-like features were identifiable at the ultrastructural level. The nodules formed on top of a fibroblast-like multilayer of cells. The upper surface of the nodules was lined by a continuous layer of cuboidal osteoblastic cells often seen to be joined by adherens junctions. Numerous microvilli, membrane protrusions, and coated pits could be seen on the upper surface of thesemore » cells, their cytoplasm contained prominent RER and Golgi membranes, and processes extended from their lower surfaces into a dense, highly organized collagenous matrix. Some osteocyte-like cells were completely embedded within this matrix; they also displayed RER and prominent processes which extended through the matrix and often made both adherens and gap junctional contacts with the processes of other cells. The fibroblastic cells not participating in nodule formation were surrounded by a less dense collagenous matrix and, in contrast to the matrix of the nodules, it did not mineralize. An unmineralized osteoid-like layer was seen directly below the cuboidal top layer of cells. A mineralization front was detectable below this in which small, discrete structures resembling matrix vesicles and feathery mineral crystals were evident and frequently associated with the collagen fibrils. More heavily mineralized areas were seen further into the nodule. Electron microprobe and electron and X-ray diffraction analysis confirmed the mineral to be hydroxyapatite.« less

A Bone-Implant Interaction Mouse Model for Evaluating Molecular Mechanism of Biomaterials/Bone Interaction.

PubMed

Liu, Wenlong; Dan, Xiuli; Wang, Ting; Lu, William W; Pan, Haobo

2016-11-01

The development of an optimal animal model that could provide fast assessments of the interaction between bone and orthopedic implants is essential for both preclinical and theoretical researches in the design of novel biomaterials. Compared with other animal models, mice have superiority in accessing the well-developed transgenic modification techniques (e.g., cell tracing, knockoff, knockin, and so on), which serve as powerful tools in studying molecular mechanisms. In this study, we introduced the establishment of a mouse model, which was specifically tailored for the assessment of bone-implant interaction in a load-bearing bone marrow microenvironment and could potentially allow the molecular mechanism study of biomaterials by using transgenic technologies. The detailed microsurgery procedures for developing a bone defect (Φ = 0.8 mm) at the metaphysis region of the mouse femur were recorded. According to our results, the osteoconductive and osseointegrative properties of a well-studied 45S5 bioactive glass were confirmed by utilizing our mouse model, verifying the reliability of this model. The feasibility and reliability of the present model were further checked by using other materials as objects of study. Furthermore, our results indicated that this animal model provided a more homogeneous tissue-implant interacting surface than the rat at the early stage of implantation and this is quite meaningful for conducting quantitative analysis. The availability of transgenic techniques to mechanism study of biomaterials was further testified by establishing our model on Nestin-GFP transgenic mice. Intriguingly, the distribution of Nestin + cells was demonstrated to be recruited to the surface of 45S5 glass as early as 3 days postsurgery, indicating that Nestin + lineage stem cells may participate in the subsequent regeneration process. In summary, the bone-implant interaction mouse model could serve as a potential candidate to evaluate the early stage tissue response near the implant surface in a bone marrow microenvironment, and it also shows great potential in making transgenic animal resource applicable to biomaterial studies, so that the design of novel biomaterials could be better guided.

Production of new 3D scaffolds for bone tissue regeneration by rapid prototyping.

PubMed

Fradique, R; Correia, T R; Miguel, S P; de Sá, K D; Figueira, D R; Mendonça, A G; Correia, I J

2016-04-01

The incidence of bone disorders, whether due to trauma or pathology, has been trending upward with the aging of the worldwide population. The currently available treatments for bone injuries are rather limited, involving mainly bone grafts and implants. A particularly promising approach for bone regeneration uses rapid prototyping (RP) technologies to produce 3D scaffolds with highly controlled structure and orientation, based on computer-aided design models or medical data. Herein, tricalcium phosphate (TCP)/alginate scaffolds were produced using RP and subsequently their physicochemical, mechanical and biological properties were characterized. The results showed that 60/40 of TCP and alginate formulation was able to match the compression and present a similar Young modulus to that of trabecular bone while presenting an adequate biocompatibility. Moreover, the biomineralization ability, roughness and macro and microporosity of scaffolds allowed cell anchoring and proliferation at their surface, as well as cell migration to its interior, processes that are fundamental for osteointegration and bone regeneration.

Engineering bone tissue substitutes from human induced pluripotent stem cells.

PubMed

de Peppo, Giuseppe Maria; Marcos-Campos, Iván; Kahler, David John; Alsalman, Dana; Shang, Linshan; Vunjak-Novakovic, Gordana; Marolt, Darja

2013-05-21

Congenital defects, trauma, and disease can compromise the integrity and functionality of the skeletal system to the extent requiring implantation of bone grafts. Engineering of viable bone substitutes that can be personalized to meet specific clinical needs represents a promising therapeutic alternative. The aim of our study was to evaluate the utility of human-induced pluripotent stem cells (hiPSCs) for bone tissue engineering. We first induced three hiPSC lines with different tissue and reprogramming backgrounds into the mesenchymal lineages and used a combination of differentiation assays, surface antigen profiling, and global gene expression analysis to identify the lines exhibiting strong osteogenic differentiation potential. We then engineered functional bone substitutes by culturing hiPSC-derived mesenchymal progenitors on osteoconductive scaffolds in perfusion bioreactors and confirmed their phenotype stability in a subcutaneous implantation model for 12 wk. Molecular analysis confirmed that the maturation of bone substitutes in perfusion bioreactors results in global repression of cell proliferation and an increased expression of lineage-specific genes. These results pave the way for growing patient-specific bone substitutes for reconstructive treatments of the skeletal system and for constructing qualified experimental models of development and disease.

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

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.

Diatomite reinforced chitosan composite membrane as potential scaffold for guided bone regeneration.

PubMed

Tamburaci, Sedef; Tihminlioglu, Funda

2017-11-01

In this study, natural silica source, diatomite, incorporated novel chitosan based composite membranes were fabricated and characterized for bone tissue engineering applications as possible bone regeneration membrane. The effect of diatomite loading on the mechanical, morphological, chemical, thermal and surface properties, wettability and in vitro cytotoxicity and cell proliferation on of composite membranes were investigated and observed by tensile test, atomic force microscopy (AFM), Fourier transform infrared spectroscopy (FTIR), thermal gravimetric analysis (TGA), protein adsorption assay, air/water contact angle analysis and WST-1 respectively. Swelling studies were also performed by water absorption capacity determination. Results showed that incorporation of diatomite to the chitosan matrix increased the surface roughness, swelling capacity and tensile modulus of membranes. An increase of about 52% in Young's modulus was achieved for 10wt% diatomite composite membranes compared with chitosan membranes. High cell viability results were obtained with indirect extraction method. Besides, in vitro cell proliferation and ALP activity results showed that diatom incorporation significantly increased the ALP activity of Saos-2 cells cultured on chitosan membranes. The novel composite membranes prepared in the present study with tunable properties can be considered as a potential candidate as a scaffold in view of its enhanced physical & chemical properties as well as biological activities for bone tissue engineering applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Physical characterization and osteogenic activity of the quaternized chitosan-loaded PMMA bone cement.

PubMed

Tan, Honglue; Guo, Shengrong; Yang, Shengbing; Xu, Xiaofen; Tang, Tingting

2012-07-01

Gentamicin-loaded polymethylmethacrylate (PMMA), widely used for primary cemented arthroplasty and revision surgery for preventing or treating infections, may lead to the evolution of antibiotic-resistant bacteria and dysfunction of osteogenic cells, which further influence the osteointegration of bone cement. In a previous study, we reported that a new quaternized chitosan derivative (hydroxypropyltrimethyl ammonium chloride chitosan, HACC) that was loaded into PMMA significantly inhibited the formation of biofilms caused by methicillin-resistant Staphylococcus strains. In the present study, we further investigated the surface morphology, hydrophilicity, apatite formation ability and osteogenic activity of HACC-loaded PMMA. Chitosan-loaded PMMA, gentamicin-loaded PMMA and PMMA without antibiotic were also investigated and compared. The results showed that, compared to other PMMA-based cements, HACC-loaded PMMA had improved properties such as a lower polymerization temperature, prolonged setting time, porous structures after immersion in phosphate-buffered saline, higher hydrophilicity, more apatite formation on the surface after immersion in simulated body fluid, and better attachment and spreading of the human-marrow-derived mesenchymal stem cells. We also found better stem cell proliferation, osteogenic differentiation, and osteogenesis-associated genes expression on the surface of the HACC-loaded PMMA compared to the gentamicin-loaded PMMA. Therefore, this new anti-infective bone cement had improved physical properties and osteogenic activity, which may lead to better osteointegration of the bone cement in cemented arthroplasty. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Periodontal regeneration with multi-layered periodontal ligament-derived cell sheets in a canine model.

PubMed

Iwata, Takanori; Yamato, Masayuki; Tsuchioka, Hiroaki; Takagi, Ryo; Mukobata, Shigeki; Washio, Kaoru; Okano, Teruo; Ishikawa, Isao

2009-05-01

Periodontal regeneration has been challenged with chemical reagents and/or biological approaches, however, there is still no sufficient technique that can regenerate complete periodontium, including alveolar bone, cementum, and well-oriented collagen fibers. The purpose of this study was to examine multi-layered sheets of periodontal ligament (PDL)-derived cells for periodontal regeneration. Canine PDL cells were isolated enzymatically and expanded in vitro. The cell population contained cells capable of making single cell-derived colonies at an approximately 20% frequency. Expression of mRNA of periodontal marker genes, S100 calcium binding protein A4 and periostin, was observed. Alkaline phosphatase activity and gene expression of both osteoblastic/cementoblastic and periodontal markers were upregulated by osteoinductive medium. Then, three-layered PDL cell sheets supported with woven polyglycolic acid were transplanted to dental root surfaces having three-wall periodontal defects in an autologous manner, and bone defects were filled with porous beta-tricalcium phosphate. Cell sheet transplantation regenerated both new bone and cementum connecting with well-oriented collagen fibers, while only limited bone regeneration was observed in control group where cell sheet transplantation was eliminated. These results suggest that PDL cells have multiple differentiation properties to regenerate periodontal tissues comprising hard and soft tissues. PDL cell sheet transplantation should prove useful for periodontal regeneration in clinical settings.

Transplantation of human placenta-derived mesenchymal stem cells in a silk fibroin/hydroxyapatite scaffold improves bone repair in rabbits.

PubMed

Jin, Jun; Wang, Jun; Huang, Jian; Huang, Fang; Fu, Jianhong; Yang, Xinjing; Miao, Zongning

2014-11-01

The main requirements for successful tissue engineering of the bone are non-immunogenic cells with osteogenic potential and a porous biodegradable scaffold. The purpose of this study is to evaluate the potential of a silk fibroin/hydroxyapatite (SF/HA) porous material as a delivery vehicle for human placenta-derived mesenchymal stem cells (PMSCs) in a rabbit radius defect model. In this study, we randomly assigned 16 healthy adult New Zealand rabbits into two groups, subjected to transplantation with either SF/HA and PMSCs (experimental group) or SF/HA alone (control group). To evaluate fracture healing, we assessed the extent of graft absorption, the quantity of newly formed bone, and re-canalization of the cavitas medullaris using radiographic and histological tools. We performed flow cytometric analysis to characterize PMSCs, and found that while they express CD90, CD105 and CD73, they stain negative for HLA-DR and the hematopoietic cell surface markers CD34 and CD45. When PMSCs were exposed to osteogenic induction medium, they secreted calcium crystals that were identified by von Kossa staining. Furthermore, when seeded on the surface of SF/HA scaffold, they actively secreted extracellular matrix components. Here, we show, through radiographic and histological analyses, that fracture healing in the experimental group is significantly improved over the control group. This strongly suggests that transplantation of human PMSCs grown in an SF/HA scaffold into injured radius segmental bone in rabbits, can markedly enhance tissue repair. Our finding provides evidence supporting the utility of human placenta as a potential source of stem cells for bone tissue engineering. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Cathodic Polarization Coats Titanium Based Implant Materials with Enamel Matrix Derivate (EMD)

PubMed Central

Frank, Matthias J.; Walter, Martin S.; Rubert, Marina; Thiede, Bernd; Monjo, Marta; Reseland, Janne E.; Haugen, Håvard J.; Lyngstadaas, Ståle Petter

2014-01-01

The idea of a bioactive surface coating that enhances bone healing and bone growth is a strong focus of on-going research for bone implant materials. Enamel matrix derivate (EMD) is well documented to support bone regeneration and activates growth of mesenchymal tissues. Thus, it is a prime candidate for coating of existing implant surfaces. The aim of this study was to show that cathodic polarization can be used for coating commercially available implant surfaces with an immobilized but functional and bio-available surface layer of EMD. After coating, XPS revealed EMD-related bindings on the surface while SIMS showed incorporation of EMD into the surface. The hydride layer of the original surface could be activated for coating in an integrated one-step process that did not require any pre-treatment of the surface. SEM images showed nano-spheres and nano-rods on coated surfaces that were EMD-related. Moreover, the surface roughness remained unchanged after coating, as it was shown by optical profilometry. The mass peaks observed in the matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS) analysis confirmed the integrity of EMD after coating. Assessment of the bioavailability suggested that the modified surfaces were active for osteoblast like MC3M3-E1 cells in showing enhanced Coll-1 gene expression and ALP activity. PMID:28788564

Histopathological Comparison between Bone Marrow- and Periodontium-derived Stem Cells for Bone Regeneration in Rabbit Calvaria.

PubMed

Kadkhoda, Z; Safarpour, A; Azmoodeh, F; Adibi, S; Khoshzaban, A; Bahrami, N

2016-01-01

Periodontitis is an important oral disease. Stem cell therapy has found its way in treatment of many diseases. To evaluate the regenerative potential of periodontal ligament-derived stem cells (PDLSCs) and osteoblast differentiated from PDLSC in comparison with bone marrow-derived mesenchymal stem cells (BM-MSCs) and pre-osteoblasts in calvarial defects. After proving the existence of surface markers by flow cytometry, BM-MSCs were differentiated into osteoblasts. 5 defects were made on rabbit calvaria. 3 of them were first covered with collagen membrane and then with BM-MSCs, PDLSCs, and pre-osteoblasts. The 4(th) defect was filled with collagen membrane and the 5(th) one was served as control. After 4 weeks, histological (quantitative) and histomorphological (qualitative) surveys were performed. Both cell lineages were positive for CD-90 cell marker, which was specifically related to stem cells. Alizarin red staining was done for showing mineral material. RT-PCR set up for the expression of Cbfa1 gene, BMP4 gene, and PGLAP gene, confirmed osteoblast differentiation. The findings indicated that although PDLSCs and pre-osteoblasts could be used for bone regeneration, the rate of regeneration in BM-MSCs-treated cavities was more significant (p<0.0001). The obtained results are probably attributable to the effective micro-environmental signals caused by different bone types and the rate of cell maturation.

Bio-active engineered 50 nm silica nanoparticles with bone anabolic activity: therapeutic index, effective concentration, and cytotoxicity profile in vitro.

PubMed

Ha, Shin-Woo; Sikorski, James A; Weitzmann, M Neale; Beck, George R

2014-04-01

Silica-based nanomaterials are generally considered to be excellent candidates for therapeutic applications particularly related to skeletal metabolism however the current data surrounding the safety of silica based nanomaterials is conflicting. This may be due to differences in size, shape, incorporation of composite materials, surface properties, as well as the presence of contaminants following synthesis. In this study we performed extensive in vitro safety profiling of ∼ 50 nm spherical silica nanoparticles with OH-terminated or Polyethylene Glycol decorated surface, with and without a magnetic core, and synthesized by the Stöber method. Nineteen different cell lines representing all major organ types were used to investigate an in vitro lethal concentration (LC) and results revealed little toxicity in any cell type analyzed. To calculate an in vitro therapeutic index we quantified the effective concentration at 50% response (EC50) for nanoparticle-stimulated mineral deposition activity using primary bone marrow stromal cells (BMSCs). The EC50 for BMSCs was not substantially altered by surface or magnetic core. The calculated Inhibitory concentration 50% (IC50) for pre-osteoclasts was similar to the osteoblastic cells. These results demonstrate the pharmacological potential of certain silica-based nanomaterial formulations for use in treating bone diseases based on a favorable in vitro therapeutic index. Copyright © 2013 Elsevier Ltd. All rights reserved.

Influence of surfaces modified with biomimetic extracellular matrices on adhesion and proliferation of mesenchymal stem cells and osteosarcoma cells.

PubMed

Cai, Rong; Kawazoe, Naoki; Chen, Guoping

2015-02-01

Preparation of surfaces modified with biomimetic extracellular matrices (ECMs) is important for investigation of the interaction between ECMs and cells. In the present study, surfaces modified with ECMs from normal somatic cells, stem cells and tumor cells were prepared by cell culture method. The ECMs derived from bone marrow-derived mesenchymal stem cells (MSCs), dermal fibroblasts (FBs), osteoblasts (OBs) and MG63 osteosarcoma cells were deposited on the surfaces of cell-culture polystyrene plates (TCPS). The ECMs from different cell types had different compositions. The effects of the ECM-deposited surfaces on the adhesion, spreading and proliferation of MSCs and MG63 human osteosarcoma cells were dependent on the type of both ECMs and cells. The surfaces deposited with ECMs from MSCs, FBs and OBs promoted cell adhesion more strongly than surfaces deposited with ECMs from MG63 cells and TCPS. Compared to TCPS, the ECM-deposited surfaces promoted proliferation of MSCs while they inhibited the proliferation of MG63 cells. Copyright © 2014 Elsevier B.V. All rights reserved.

Bone regeneration by means of a three-dimensional printed scaffold in a rat cranial defect.

PubMed

Kwon, Doo Yeon; Park, Ji Hoon; Jang, So Hee; Park, Joon Yeong; Jang, Ju Woong; Min, Byoung Hyun; Kim, Wan-Doo; Lee, Hai Bang; Lee, Junhee; Kim, Moon Suk

2018-02-01

Recently, computer-designed three-dimensional (3D) printing techniques have emerged as an active research area with almost unlimited possibilities. In this study, we used a computer-designed 3D scaffold to drive new bone formation in a bone defect. Poly-L-lactide (PLLA) and bioactive β-tricalcium phosphate (TCP) were simply mixed to prepare ink. PLLA + TCP showed good printability from the micronozzle and solidification within few seconds, indicating that it was indeed printable ink for layer-by-layer printing. In the images, TCP on the surface of (and/or inside) PLLA in the printed PLLA + TCP scaffold looked dispersed. MG-63 cells (human osteoblastoma) adhered to and proliferated well on the printed PLLA + TCP scaffold. To assess new bone formation in vivo, the printed PLLA + TCP scaffold was implanted into a full-thickness cranial bone defect in rats. The new bone formation was monitored by microcomputed tomography and histological analysis of the in vivo PLLA + TCP scaffold with or without MG-63 cells. The bone defect was gradually spontaneously replaced with new bone tissues when we used both bioactive TCP and MG-63 cells in the PLLA scaffold. Bone formation driven by the PLLA + TCP30 scaffold with MG-63 cells was significantly greater than that in other experimental groups. Furthermore, the PLLA + TCP scaffold gradually degraded and matched well the extent of the gradual new bone formation on microcomputed tomography. In conclusion, the printed PLLA + TCP scaffold effectively supports new bone formation in a cranial bone defect. Copyright © 2017 John Wiley & Sons, Ltd.

Development and application of biomimetic electrospun nanofibers in total joint replacement

NASA Astrophysics Data System (ADS)

Song, Wei

Failure of osseointegration (direct anchorage of an implant by bone formation at the bone-implant surface) and implant infection (such as that caused by Staphylococcus aureus, S. aureus) are the two main causes of implant failure and loosening. There is a critical need for orthopedic implants that promote rapid osseointegration and prevent bacterial colonization, particularly when placed in bone compromised by disease or physiology of the patients. A better understanding of the key factors that influence cell fate decisions at the bone-implant interface is required. Our study is to develop a class of "bone-like" nanofibers (NFs) that promote osseointegration while preventing bacterial colonization and subsequent infections. This research goal is supported by our preliminary data on the preparation of coaxial electrospun NFs composed of polycaprolactone (PCL) and polyvinyl alcohol (PVA) polymers arranged in a core-sheath shape. The PCL/PVA NFs are biocompatible and biodegradable with appropriate fiber diameter, pore size and mechanical strength, leading to enhanced cell adhesion, proliferation and differentiation of osteoblast precursor cells. The objective is to develop functionalized "bone-like" PCL/PVA NFs matrix embedded with antibiotics (doxycycline (Doxy), bactericidal and anti-osteoclastic) on prosthesis surface. Through a rat tibia implantation model, the Doxy incorporated coaxial NFs has demonstrated excellent in promoting osseointegration and bacteria inhibitory efficacy. NFs coatings significantly enhanced the bonding between implant and bone remodeling within 8 weeks. The SA-induced osteomyelitis was prevented by the sustained release of Doxy from NFs. The capability of embedding numerous bio-components including proteins, growth factors, drugs, etc. enables NFs an effective solution to overcome the current challenged issue in Total joint replacement. In summary, we proposed PCL/PVA electrospun nanofibers as promising biomaterials that can be applied on joint replacement prosthesis to improve osseointegration and prevent osteomyelitis.

Controlling the Biodegradation of Magnesium Implants Through Nanostructured Calcium-Phosphate Coating

NASA Astrophysics Data System (ADS)

Iskandar, Maria Emil

Magnesium (Mg) alloys, a novel class of degradable, metallic biomaterials, have attracted growing interest as a promising alternative for medical implant and device applications due to their advantageous mechanical and biological properties. Moreover, Mg is biodegradable in the physiological environments. However, the major obstacle for Mg to be used as medical implants is its rapid degradation in physiological fluids. Therefore, the present key challenge lies in controlling Mg degradation rate in the physiological environment. The objective of this study was to develop a nanostructured-hydroxyapatite (nHA) coating on polished Mg implants to control the degradation and bone tissue integration of the implants. The nHA coatings were deposited on Mg using the Spire's patented TPA process to moderate the aggressive degradation of Mg and to improve quick osteointegration between Mg and natural bone. Nanostructured-HA coatings mimic the nanostructure and chemistry of natural bone, which will provide a desirable environment for bone tissue regeneration. Surface morphology, element compositions, and crystal structures were characterized using scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS), and x-ray diffractometry (XRD), respectively. SEM images of the deposited nHA-coating was analyzed using ImageJ's quantitative image analysis tool, to determine the nHA-coating particle size and thickness. The degradation of nHA-coated and non-coated Mg samples was investigated by incubating samples in phosphate buffered saline (PBS) and revised simulated body fluid (r-SBF), under standard cell culture conditions. To mimic the in vivo cell response in the physiological environment, rat bone marrow stromal cells (BMSC) were harvested and cultured with nHA-coated and non-coated polished Mg samples to determine cytocompatibilty. The degradation results suggested that the nanocoatings positively mediated Mg degradation. It can therefore be concluded that nHA-coatings show promise for controlling the biodegradation of Mg-based orthopedic implants and devices. Cell studies indicated significantly improved BMSC adhesion on the surfaces of the nHA-coated and non-coated Mg samples, in comparison to the cells surrounding the Mg samples. These results indicated that the nHA-coated and non-coated Mg samples promote cell activity on the surface. However, cell experiments must be repeated on a larger number of samples with extensive short and long term cell studies, to achieve more verifiable results.

Fabrication of Bioceramic Bone Scaffolds for Tissue Engineering

NASA Astrophysics Data System (ADS)

Liu, Fwu-Hsing

2014-10-01

In this study, microhydroxyapatite and nanosilica sol were used as the raw materials for fabrication of bioceramic bone scaffold using selective laser sintering technology in a self-developed 3D Printing apparatus. When the fluidity of ceramic slurry is matched with suitable laser processing parameters, a controlled pore size of porous bone scaffold can be fabricated under a lower laser energy. Results shown that the fabricated scaffolds have a bending strength of 14.1 MPa, a compressive strength of 24 MPa, a surface roughness of 725 nm, a pore size of 750 μm, an apparent porosity of 32%, and a optical density of 1.8. Results indicate that the mechanical strength of the scaffold can be improved after heat treatment at 1200 °C for 2 h, while simultaneously increasing surface roughness conducive to osteoprogenitor cell adhesion. MTT method and SEM observations confirmed that bone scaffolds fabricated under the optimal manufacturing process possess suitable biocompatibility and mechanical properties, allowing smooth adhesion and proliferation of osteoblast-like cells. Therefore, they have great potential for development in the field of tissue engineering.

[The use of structural proximal tibial allografts coated with human albumin in treating extensive periprosthetic knee-joint bone deficiency and averting late complications. Case report].

PubMed

Klára, Tamás; Csönge, Lajos; Janositz, Gábor; Pap, Károly; Lacza, Zsombor

2015-01-11

The authors report the history of a 74-year-old patient who underwent surgical treatment for segmental knee-joint periprosthetic bone loss using structural proximal tibial allografts coated with serum albumin. Successful treatment of late complications which occurred in the postoperative period is also described. The authors emphasize that bone replacement with allografts is a physiological process that enables the stable positioning of the implant and the reconstruction of the soft tissues, the replacement of extensive bone loss, and also it is a less expensive operation. It has been already confirmed that treatment of lyophilised allografts with albumin improves the ability of bone marrow-derived mesenchymal stem cells to adhere and proliferate the surface of the allografts, penetrate the pores and reach deeper layers of the graft. Earlier studies have shown osteoblast activity on the surface and interior of the graft.

Preparation of a non-woven poly(ε-caprolactone) fabric with partially embedded apatite surface for bone tissue engineering applications by partial surface melting of poly(ε-caprolactone) fibers.

PubMed

Kim, In Ae; Rhee, Sang-Hoon

2017-07-01

This article describes a novel method for the preparation of a biodegradable non-woven poly(ε-caprolactone) fabric with a partially embedded apatite surface designed for application as a scaffold material for bone tissue engineering. The non-woven poly(ε-caprolactone) fabric was generated by the electro-spinning technique and then apatite was coated in simulated body fluid after coating the PVA solution containing CaCl 2 ·2H 2 O. The apatite crystals were partially embedded or fully embedded into the thermoplastic poly(ε-caprolactone) fibers by controlling the degree of poly(ε-caprolactone) fiber surface melting in a convection oven. Identical apatite-coated poly(ε-caprolactone) fabric that did not undergo heat-treatment was used as a control. The features of the embedded apatite crystals were evaluated by FE-SEM, AFM, EDS, and XRD. The adhesion strengths of the coated apatite layers and the tensile strengths of the apatite coated fabrics with and without heat-treatment were assessed by the tape-test and a universal testing machine, respectively. The degree of water absorbance was assessed by adding a DMEM droplet onto the fabrics. Moreover, cell penetrability was assessed by seeding preosteoblastic MC3T3-E1 cells onto the fabrics and observing the degrees of cell penetration after 1 and 4 weeks by staining nuclei with DAPI. The non-woven poly(ε-caprolactone) fabric with a partially embedded apatite surface showed good water absorbance, cell penetrability, higher apatite adhesion strength, and higher tensile strength compared with the control fabric. These results show that the non-woven poly(ε-caprolactone) fabric with a partially embedded apatite surface is a potential candidate scaffold for bone tissue engineering due to its strong apatite adhesion strength and excellent cell penetrability. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1973-1983, 2017. © 2017 Wiley Periodicals, Inc.

In vitro study of 3D PLGA/n-HAp/β-TCP composite scaffolds with etched oxygen plasma surface modification in bone tissue engineering

NASA Astrophysics Data System (ADS)

Roh, Hee-Sang; Jung, Sang-Chul; Kook, Min-Suk; Kim, Byung-Hoon

2016-12-01

Three-dimensional (3D) scaffolds have many advantageous properties for bone tissue engineering application, due to its controllable properties such as pore size, structural shape and interconnectivity. In this study, effects on oxygen plasma surface modification and adding of nano-hydroxyapatite (n-HAp) and β-tricalcium phosphate (β-TCP) on the 3D PLGA/n-HAp/β-TCP scaffolds for improving preosteoblast cell (MC3T3-E1) adhesion, proliferation and differentiation were investigated. The 3D PLGA/n-HAp/β-TCP scaffolds were fabricated by 3D Bio-Extruder equipment. The 3D scaffolds were prepared with 0°/90° architecture and pore size of approximately 300 μm. In addition 3D scaffolds surface were etched by oxygen plasma to enhance the hydrophilic property and surface roughness. After oxygen plasma treatment, the surface chemistry and morphology were investigated by Fourier transform infrared spectroscopy, scanning electron microscopy, and atomic force microscopy. And also hydrophilic property was measured by contact angle. The MC3T3-E1 cell proliferation and differentiation were investigated by MTT assay and ALP activity. In present work, the 3D PLGA/HAp/beta-TCP composite scaffold with suitable structure for the growth of osteoblast cells was successfully fabricated by 3D rapid prototyping technique. The surface hydrophilicity and roughness of 3D scaffold increased by oxygen plasma treatment had a positive effect on cell adhesion, proliferation, and differentiation. Furthermore, the differentiation of MC3T3-E1 cell was significantly enhanced by adding of n-HAp and β-TCP on 3D PLGA scaffold. As a result, combination of bioceramics and oxygen plasma treatment showed a synergistic effect on biocompatibility of 3D scaffolds. This result confirms that this technique was useful tool for improving the biocompatibility in bone tissue engineering application.

Surface biofunctionalization of β-TCP blocks using aptamer 74 for bone tissue engineering.

PubMed

Ardjomandi, N; Huth, J; Stamov, D R; Henrich, A; Klein, C; Wendel, H-P; Reinert, S; Alexander, D

2016-10-01

Successful bone regeneration following oral and maxillofacial surgeries depends on efficient functionalization strategies that allow the recruitment of osteogenic progenitor cells at the tissue/implant interface. We have previously identified aptamer 74, which exhibited a binding affinity for osteogenically induced jaw periosteal cells (JPCs). In the present study, this aptamer was used for the surface biofunctionalization of β-tricalcium phosphate (β-TCP) blocks. Atomic force microscopy (AFM) measurements showed increased binding activity of aptamer 74 towards osteogenically induced JPCs compared to untreated controls. The immobilization efficiency of aptamer 74 was analyzed using the QuantiFluor ssDNA assay for 2D surfaces and by amino acid analysis for 3D β-TCP constructs. Following the successful immobilization of aptamer 74 in 2D culture wells and on 3D constructs, in vitro assays showed no significant differences in cell proliferation compared to unmodified surfaces. Interestingly, JPC mineralization was significantly higher on the 2D surfaces and higher cell adhesion was detected on the 3D constructs with immobilized aptamer. Herein, we report an established, biocompatible β-TCP matrix with surface immobilization of aptamer 74, which enhances properties such as cell adhesion on 3D constructs and mineralization on 2D surfaces. Further studies need to be performed to improve the immobilization efficiency and to develop a suitable approach for JPC mineralization growing within 3D β-TCP constructs. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of surface roughness on osteogenesis in vitro and osseointegration in vivo of carbon fiber-reinforced polyetheretherketone–nanohydroxyapatite composite

PubMed Central

Deng, Yi; Liu, Xiaochen; Xu, Anxiu; Wang, Lixin; Luo, Zuyuan; Zheng, Yunfei; Deng, Feng; Wei, Jie; Tang, Zhihui; Wei, Shicheng

2015-01-01

As United States Food and Drug Administration-approved implantable material, carbon fiber-reinforced polyetheretherketone (CFRPEEK) possesses an adjustable elastic modulus similar to cortical bone and is a prime candidate to replace surgical metallic implants. The bioinertness and inferior osteogenic properties of CFRPEEK, however, limit its clinical application as orthopedic/dental implants. In this study, CFRPEEK–nanohydroxyapatite ternary composites (PEEK/n-HA/CF) with variable surface roughness have been successfully fabricated. The effect of surface roughness on their in vitro cellular responses of osteoblast-like MG-63 cells (attachment, proliferation, apoptosis, and differentiation) and in vivo osseointegration is evaluated. The results show that the hydrophilicity and the amount of Ca ions on the surface are significantly improved as the surface roughness of composite increases. In cell culture tests, the results reveal that the cell proliferation rate and the extent of osteogenic differentiation of cells are a function of the size of surface roughness. The composite with moderate surface roughness significantly increases cell attachment/proliferation and promotes the production of alkaline phosphatase (ALP) activity and calcium nodule formation compared with the other groups. More importantly, the PEEK/n-HA/CF implant with appropriate surface roughness exhibits remarkably enhanced bioactivity and osseointegration in vivo in the animal experiment. These findings will provide critical guidance for the design of CFRPEEK-based implants with optimal roughness to regulate cellular behaviors, and to enhance biocompability and osseointegration. Meanwhile, the PEEK/n-HA/CF ternary composite with optimal surface roughness might hold great potential as bioactive biomaterial for bone grafting and tissue engineering applications. PMID:25733834

Ultrastructural evaluation of mesenchymal stem cells from inflamed periodontium in different in vitro conditions.

PubMed

Zaganescu, Raluca; Barbu Tudoran, Lucian; Pall, Emoke; Florea, Adrian; Roman, Alexandra; Soanca, Andrada; Mihaela Mihu, Carmen

2015-09-01

This research aimed to observe the behavior of mesenchymal stem cells (MSCs) isolated from periodontal granulation tissue (gt) when manipulated ex vivo to induce three-dimensional (3D) spheroid (aggregates) formation as well as when seeded on two bone scaffolds of animal origin. Periodontal gt was chosen as a MSC source because of its availability, considering that it is eliminated as a waste material during conventional surgical therapies. 3D aggregates of cells were generated; they were grown for 3 and 7 days, respectively, and then prepared for transmission electron microscopic analysis. The two biomaterials were seeded for 72 h with gtMSCs and prepared for scanning electronic microscopic observation. The ultrastructural analysis of 3D spheroids remarked some differences between the inner and the outer cell layers, with a certain commitment observed at the inner cells. Both scaffolds showed a relatively smooth surface at low magnification. Macro- and micropores having a scarce distribution were observed on both bone substitutes. gtMSCs grew with relative difficulty on the biomaterials. After 72 h of proliferation, gtMSCs scarcely covered the surface of bovine bone scaffolds, demonstrating fibroblast-like or star-like shapes with elongated filiform extensions. Our results add other data on the possible usefulness of gtMSC and could question the current paradigm regarding the complete removal of chronically inflamed gts from the defects during periodontal surgeries. Until optimal protocols for ex vivo manipulation of MSCs are available for clinical settings, it is advisable to use biocompatible bone substitutes that allow the development of progenitor cells. © 2015 Wiley Periodicals, Inc.

Periodontal Bioengineering: A Discourse in Surface Topographies, Progenitor Cells and Molecular Profiles

NASA Astrophysics Data System (ADS)

Dangaria, Smit J.

2011-12-01

Stem/progenitor cells are a population of cells capable of providing replacement cells for a given differentiated cell type. We have applied progenitor cell-based technologies to generate novel tissue-engineered implants that use biomimetic strategies with the ultimate goal of achieving full regeneration of lost periodontal tissues. Mesenchymal periodontal tissues such as cementum, alveolar bone (AB), and periodontal ligament (PDL) are neural crest-derived entities that emerge from the dental follicle (DF) at the onset of tooth root formation. Using a systems biology approach we have identified key differences between these periodontal progenitors on the basis of global gene expression profiles, gene cohort expression levels, and epigenetic modifications, in addition to differences in cellular morphologies. On an epigenetic level, DF progenitors featured high levels of the euchromatin marker H3K4me3, whereas PDL cells, AB osteoblasts, and cementoblasts contained high levels of the transcriptional repressor H3K9me3. Secondly, we have tested the influence of natural extracellular hydroxyapatite matrices on periodontal progenitor differentiation. Dimension and structure of extracellular matrix surfaces have powerful influences on cell shape, adhesion, and gene expression. Here we show that natural tooth root topographies induce integrin-mediated extracellular matrix signaling cascades in tandem with cell elongation and polarization to generate physiological periodontium-like tissues. In this study we replanted surface topography instructed periodontal ligament progenitors (PDLPs) into rat alveolar bone sockets for 8 and 16 weeks, resulting in complete attachment of tooth roots to the surrounding alveolar bone with a periodontal ligament fiber apparatus closely matching physiological controls along the entire root surface. Displacement studies and biochemical analyses confirmed that progenitor-based engineered periodontal tissues were similar to control teeth and uniquely derived from pre-implantation green fluorescent protein (GFP)-labeled progenitors. Together, these studies illustrate the capacity of natural extracellular surface topographies to instruct PDLPs to fully regenerate complex cellular and structural morphologies of tissues once lost to disease. We suggest that our strategy could be used for the replantation of teeth lost due to trauma or as a novel approach for tooth replacement using tooth-shaped replicas.

The synergistic effect of VEGF and biomorphic silicon carbides topography on in vivo angiogenesis and human bone marrow derived mesenchymal stem cell differentiation.

PubMed

Díaz-Rodríguez, P; Gómez-Amoza, J L; Landin, M

2015-08-04

Topographical features of biomaterials are able to modulate cell attachment, spreading and differentiation. The addition of growth factors to implantable biomaterials can modify these cellular responses, enhancing their therapeutic potential. The aim of this research is to establish the influence of biomorphic silicon carbide ceramics (bioSiCs) surface topography on the proliferation and osteoblastic differentiation of mesenchymal stem cells and the potential synergistic effect of the ceramic porous structure together with vascular endothelial growth factor loading (VEGF) on the surface mediated osteoblastic differentiation. Three porous bioSiCs with important differences in their microstructure were obtained from different natural precursors. Samples loaded with or without VEGF through ionic interactions were cultured with human umbilical vein endothelial cells (HUVEC) or bone marrow derived mesenchymal stem cells (hMSCs). Cell behaviour and protein activity with regard to bioSiC porous structure and surface properties were analysed. An in vivo model (Chick Chorioallantoic Membrane; CAM) was used to assess the capability of the VEGF loaded systems to promote angiogenesis. Experimental data show that loaded systems were able to control the release of VEGF for up to 15 d ensuring the activity of the protein, increasing the proliferation of HUVECs and the formation of new blood vessels in the CAM. It was found that the selection of bioSiCs with a higher pore size promoted a higher concentration of osteoblastic differentiation markers of MSCs cultured on the surface of bioSiCs. Furthermore, the addition of VEGF to the systems was able to promote a faster osteoblastic differentiation according to the qPCR results, suggesting a synergy between both the surface properties and the controlled release of the growth factor. The VEGF loaded sapelli bioSiC was found to be the most promising material for bone tissue engineering applications.

Immunohistochemical characterization of nanocrystalline hydroxyapatite silica gel (NanoBone(r)) osteogenesis: a study on biopsies from human jaws.

PubMed

Götz, Werner; Gerber, Thomas; Michel, Barbara; Lossdörfer, Stefan; Henkel, Kai-Olaf; Heinemann, Friedhelm

2008-10-01

Bone substitute biomaterials may be osteogenic, osteoconductive or osteoinductive. To test for these probable characteristics in a new nanoporous grafting material consisting of nanocrystalline hydroxyapatite embedded in a porous silica gel matrix (NanoBone(s)), applied in humans, we studied biopsies from 12 patients before dental implantation following various orofacial augmentation techniques with healing times of between 3.5 and 12 months. Sections from decalcified specimens were investigated using histology, histochemistry [periodic acid Schiff, alcian blue staining and tartrate-resistant acid phosphatase (TRAP)] and immunohistochemistry, with markers for osteogenesis, bone remodelling, resorption and vessel walls (alkaline phosphatase, bone morphogenetic protein-2, collagen type I, ED1, osteocalcin, osteopontin, runx2 and Von-Willebrand factor). Histologically, four specific stages of graft transformation into lamellar bone could be characterized. During early stages of healing, bone matrix proteins were absorbed by NanoBone(s) granules, forming a proteinaceous matrix, which was invaded by small vessels and cells. We assume that the deposition of these molecules promotes early osteogenesis in and around NanoBone(s) and supports the concomitant degradation probably by osteoclast-like cells. TRAP-positive osteoclast-like cells were localized directly on the granular surfaces. Runx2-immunoreactive pre-osteoblasts, which are probably involved in direct osteogenesis forming woven bone that is later transformed into lamellar bone, were attracted. Graft resorption and bone apposition around the graft granules appear concomitantly. We postulate that NanoBone(s) has osteoconductive and biomimetic properties and is integrated into the host's physiological bone turnover at a very early stage.

Synthesis of three-dimensional calcium carbonate nanofibrous structure from eggshell using femtosecond laser ablation

PubMed Central

2011-01-01

Background Natural biomaterials from bone-like minerals derived from avian eggshells have been considered as promising bone substitutes owing to their biodegradability, abundance, and lower price in comparison with synthetic biomaterials. However, cell adhesion to bulk biomaterials is poor and surface modifications are required to improve biomaterial-cell interaction. Three-dimensional (3D) nanostructures are preferred to act as growth support platforms for bone and stem cells. Although there have been several studies on generating nanoparticles from eggshells, no research has been reported on synthesizing 3D nanofibrous structures. Results In this study, we propose a novel technique to synthesize 3D calcium carbonate interwoven nanofibrous platforms from eggshells using high repetition femtosecond laser irradiation. The eggshell waste is value engineered to calcium carbonate nanofibrous layer in a single step under ambient conditions. Our striking results demonstrate that by controlling the laser pulse repetition, nanostructures with different nanofiber density can be achieved. This approach presents an important step towards synthesizing 3D interwoven nanofibrous platforms from natural biomaterials. Conclusion The synthesized 3D nanofibrous structures can promote biomaterial interfacial properties to improve cell-platform surface interaction and develop new functional biomaterials for a variety of biomedical applications. PMID:21251288

SLM Produced Porous Titanium Implant Improvements for Enhanced Vascularization and Osteoblast Seeding

PubMed Central

Matena, Julia; Petersen, Svea; Gieseke, Matthias; Kampmann, Andreas; Teske, Michael; Beyerbach, Martin; Murua Escobar, Hugo; Haferkamp, Heinz; Gellrich, Nils-Claudius; Nolte, Ingo

2015-01-01

To improve well-known titanium implants, pores can be used for increasing bone formation and close bone-implant interface. Selective Laser Melting (SLM) enables the production of any geometry and was used for implant production with 250-µm pore size. The used pore size supports vessel ingrowth, as bone formation is strongly dependent on fast vascularization. Additionally, proangiogenic factors promote implant vascularization. To functionalize the titanium with proangiogenic factors, polycaprolactone (PCL) coating can be used. The following proangiogenic factors were examined: vascular endothelial growth factor (VEGF), high mobility group box 1 (HMGB1) and chemokine (C-X-C motif) ligand 12 (CXCL12). As different surfaces lead to different cell reactions, titanium and PCL coating were compared. The growing into the porous titanium structure of primary osteoblasts was examined by cross sections. Primary osteoblasts seeded on the different surfaces were compared using Live Cell Imaging (LCI). Cross sections showed cells had proliferated, but not migrated after seven days. Although the cell count was lower on titanium PCL implants in LCI, the cell count and cell spreading area development showed promising results for titanium PCL implants. HMGB1 showed the highest migration capacity for stimulating the endothelial cell line. Future perspective would be the incorporation of HMGB1 into PCL polymer for the realization of a slow factor release. PMID:25849656

Quercetin potentiates transdifferentiation of bone marrow mesenchymal stem cells into the beta cells in vitro.

PubMed

Miladpour, B; Rasti, M; Owji, A A; Mostafavipour, Z; Khoshdel, Z; Noorafshan, A; Zal, F

2017-05-01

Type 1 diabetes is an autoimmune disease caused by the destruction of β-cells in the pancreas. Bone marrow mesenchymal stem cells are multipotent and easy accessible adult stem cells that may provide options in the treatment of type 1 diabetes. Injured pancreatic extract can promote the differentiation of rat bone marrow mesenchymal stem cells into β-cells. We aimed to observe the effect of quercetin in differentiation and insulin secretion in β-cells. Bone marrow mesenchymal stem cells were obtained from the tibiae of rats. Cell surface markers were analyzed by flow cytometry. The cells were treated with rat injured pancreatic extract and quercetin for 2 weeks. Insulin secretion was measured by ELISA. Insulin expression and some islet factors were evaluated by RT-PCR. PDX1, a marker for β-cell function and differentiation, was evaluated by both immunocytochemistry and Western blot. β-cell count was determined by stereology and cell count assay. ELISA showed significant differences in insulin secretion in the cells treated with RIPE + 20 μM quercetin (0.55 ± 0.01 µg/L) compared with the cells treated with RIPE alone (0.48 ± 0.01 µg/L) (P = 0.026). RT-PCR results confirmed insulin expression in both groups. PDX1 protein was detected in both groups by Western blot and immunocytochemistry. Stereology results showed a significant increase in β-cell number in the RIPE + quercetin-treated cells (47 ± 2.0) when compared with RIPE treatment alone (44 ± 2.5) (P = 0.015). Quercetin has a strengthening effect on the differentiation of rat bone marrow mesenchymal stem cells into β-cells and increases insulin secretion from the differentiated β-cells in vitro.

Programmable 3D silk bone marrow niche for platelet generation ex vivo and modeling of megakaryopoiesis pathologies

PubMed Central

Di Buduo, Christian A.; Wray, Lindsay S.; Tozzi, Lorenzo; Malara, Alessandro; Chen, Ying; Ghezzi, Chiara E.; Smoot, Daniel; Sfara, Carla; Antonelli, Antonella; Spedden, Elise; Bruni, Giovanna; Staii, Cristian; De Marco, Luigi; Magnani, Mauro; Kaplan, David L.

2015-01-01

We present a programmable bioengineered 3-dimensional silk-based bone marrow niche tissue system that successfully mimics the physiology of human bone marrow environment allowing us to manufacture functional human platelets ex vivo. Using stem/progenitor cells, megakaryocyte function and platelet generation were recorded in response to variations in extracellular matrix components, surface topography, stiffness, coculture with endothelial cells, and shear forces. Millions of human platelets were produced and showed to be functional based on multiple activation tests. Using adult hematopoietic progenitor cells our system demonstrated the ability to reproduce key steps of thrombopoiesis, including alterations observed in diseased states. A critical feature of the system is the use of natural silk protein biomaterial allowing us to leverage its biocompatibility, nonthrombogenic features, programmable mechanical properties, and surface binding of cytokines, extracellular matrix components, and endothelial-derived proteins. This in turn offers new opportunities for the study of blood component production ex vivo and provides a superior tissue system for the study of pathologic mechanisms of human platelet production. PMID:25575540

Use of Adipose Derived Stem Cells to Treat Large Bone Defects. Addendum

DTIC Science & Technology

2009-07-01

optimal delivery . We have also completed characterization of our segmental defect model, including analysis of vascular ingrowth during defect healing...cells seeded in 1.2% Keltone alginate at a density of 12-15x106cells/ml were loaded on 24-well transwell insert membranes [6]. Once hydrogel discs...process from tissue culture plates and hydrogels does not alter the surface phenotype. Gene expression of surface markers and proteins associated with

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.

In vivo response to starch-based scaffolds designed for bone tissue engineering applications.

PubMed

Salgado, A J; Coutinho, O P; Reis, R L; Davies, J E

2007-03-15

Our purpose was to evaluate the in vivo endosseous response to three starch-based scaffolds implanted in rats (n = 54). We implanted the three scaffold groups; a 50/50 (wt %) blend of corn starch and ethylene-vinyl alcohol (SEVA-C), the same composition coated with a biomimetic calcium phosphate (Ca-P) layer (SEVA-C/CaP), and a 50/50 (wt %) blend of corn starch and cellulose acetate (SCA), all produced by extrusion with blowing agents, into distal femurs proximal to the epiphyseal plate, for 1, 3, or 6 weeks. Our results showed that at 1 week considerable reparative bone formed around all scaffold groups, although the bone was separated from the scaffold by an intervening soft tissue interfacial zone that comprised two distinct compartments: the surface of the scaffold was occupied by multinucleate giant cells and the compartment between these cells and the surrounding bone was occupied by a streaming fibrous-like tissue. The extracellular matrix of the latter was continuous with the extracellular bone matrix itself, labeled positively for osteocalcin and appeared mineralized by back-scattered electron imaging. All three scaffolds showed a similar tissue response, with the soft tissue interface diminishing with time. No bone contact was observed with SEVA-C at any time point, only transitory bone contact was observed with SEVA-C/CaP at 3 weeks, but SCA exhibited direct bone contact at 6 weeks where 56.23 +/- 6.46% of the scaffold surface was occupied by bone. We conclude that all materials exhibited a favorable bony response and that the rapidly forming initial "connective tissue" seen around all scaffolds was a very early form of bone formation.

Rapid prototyped porous nickel–titanium scaffolds as bone substitutes

PubMed Central

Hoffmann, Waldemar; Bormann, Therese; Rossi, Antonella; Müller, Bert; Schumacher, Ralf; Martin, Ivan; Wendt, David

2014-01-01

While calcium phosphate–based ceramics are currently the most widely used materials in bone repair, they generally lack tensile strength for initial load bearing. Bulk titanium is the gold standard of metallic implant materials, but does not match the mechanical properties of the surrounding bone, potentially leading to problems of fixation and bone resorption. As an alternative, nickel–titanium alloys possess a unique combination of mechanical properties including a relatively low elastic modulus, pseudoelasticity, and high damping capacity, matching the properties of bone better than any other metallic material. With the ultimate goal of fabricating porous implants for spinal, orthopedic and dental applications, nickel–titanium substrates were fabricated by means of selective laser melting. The response of human mesenchymal stromal cells to the nickel–titanium substrates was compared to mesenchymal stromal cells cultured on clinically used titanium. Selective laser melted titanium as well as surface-treated nickel–titanium and titanium served as controls. Mesenchymal stromal cells had similar proliferation rates when cultured on selective laser melted nickel–titanium, clinically used titanium, or controls. Osteogenic differentiation was similar for mesenchymal stromal cells cultured on the selected materials, as indicated by similar gene expression levels of bone sialoprotein and osteocalcin. Mesenchymal stromal cells seeded and cultured on porous three-dimensional selective laser melted nickel–titanium scaffolds homogeneously colonized the scaffold, and following osteogenic induction, filled the scaffold’s pore volume with extracellular matrix. The combination of bone-related mechanical properties of selective laser melted nickel–titanium with its cytocompatibility and support of osteogenic differentiation of mesenchymal stromal cells highlights its potential as a superior bone substitute as compared to clinically used titanium. PMID:25383165

Periodontal ligament stem/progenitor cells with protein-releasing scaffolds for cementum formation and integration on dentin surface.

PubMed

Cho, Hankyu; Tarafder, Solaiman; Fogge, Michael; Kao, Kristy; Lee, Chang H

2016-11-01

Purpose/Aim: Cementogenesis is a critical step in periodontal tissue regeneration given the essential role of cementum in anchoring teeth to the alveolar bone. This study is designed to achieve integrated cementum formation on the root surfaces of human teeth using growth factor-releasing scaffolds with periodontal ligament stem/progenitor cells (PDLSCs). Human PDLSCs were sorted by CD146 expression, and characterized using CFU-F assay and induced multi-lineage differentiation. Polycaprolactone scaffolds were fabricated using 3D printing, embedded with poly(lactic-co-glycolic acids) (PLGA) microspheres encapsulating connective tissue growth factor (CTGF), bone morphogenetic protein-2 (BMP-2), or bone morphogenetic protein-7 (BMP-7). After removing cementum on human tooth roots, PDLSC-seeded scaffolds were placed on the exposed dentin surface. After 6-week culture with cementogenic/osteogenic medium, cementum formation and integration were evaluated by histomorphometric analysis, immunofluorescence, and qRT-PCR. Periodontal ligament (PDL) cells sorted by CD146 and single-cell clones show a superior clonogenecity and multipotency as compared with heterogeneous populations. After 6 weeks, all the growth factor-delivered groups showed resurfacing of dentin with a newly formed cementum-like layer as compared with control. BMP-2 and BMP-7 showed de novo formation of tissue layers significantly thicker than all the other groups, whereas CTGF and BMP-7 resulted in significantly improved integration on the dentin surface. The de novo mineralized tissue layer seen in BMP-7-treated samples expressed cementum matrix protein 1 (CEMP1). Consistently, BMP-7 showed a significant increase in CEMP1 mRNA expression. Our findings represent important progress in stem cell-based cementum regeneration as an essential part of periodontium regeneration.

Role of Complement on Broken Surfaces After Trauma.

PubMed

Huber-Lang, Markus; Ignatius, Anita; Brenner, Rolf E

2015-01-01

Activation of both the complement and coagulation cascade after trauma and subsequent local and systemic inflammatory response represent a major scientific and clinical problem. After severe tissue injury and bone fracture, exposure of innate immunity to damaged cells and molecular debris is considered a main trigger of the posttraumatic danger response. However, the effects of cellular fragments (e.g., histones) on complement activation remain enigmatic. Furthermore, direct effects of "broken" bone and cartilage surfaces on the fluid phase response of complement and its interaction with key cells of connective tissues are still unknown. Here, we summarize data suggesting direct and indirect complement activation by extracellular and cellular danger associated molecular patterns. In addition, key complement components and the corresponding receptors (such as C3aR, C5aR) have been detected on "exposed surfaces" of the damaged regions. On a cellular level, multiple effects of complement activation products on osteoblasts, osteoclasts, chondrocytes and mesenchymal stem cells have been found.In conclusion, the complement system may be activated by trauma-altered surfaces and is crucially involved in connective tissue healing and posttraumatic systemic inflammatory response.

Surface nanoporosity has a greater influence on osteogenic and bacterial cell adhesion than crystallinity and wettability

NASA Astrophysics Data System (ADS)

Rodriguez-Contreras, Alejandra; Guadarrama Bello, Dainelys; Nanci, Antonio

2018-07-01

There has been much emphasis on the influence of crystallinity and wettability for modulating cell activity, particularly for bone biomaterials. In this context, we have generated titanium oxide layers with similar mesoporous topography and surface roughness but with amorphous or crystalline oxide layers and differential wettability. We then investigated their influence on the behavior of MC3T3 osteoblastic and bacterial cells. There was no difference in cell adhesion, spreading and growth on amorphous and crystalline surfaces. The number of focal adhesions was similar, however, cells on the amorphous surface exhibited a higher frequency of mature adhesions. The crystallinity of the surface layers also had no bearing on bacterial adhesion. While it cannot be excluded that surface crystallinity, roughness and wettability contribute to some degree to determining cell behavior, our data suggest that physical characteristics of surfaces represent the major determinant.

Endochondral ossification is required for haematopoietic stem-cell niche formation.

PubMed

Chan, Charles K F; Chen, Ching-Cheng; Luppen, Cynthia A; Kim, Jae-Beom; DeBoer, Anthony T; Wei, Kevin; Helms, Jill A; Kuo, Calvin J; Kraft, Daniel L; Weissman, Irving L

2009-01-22

Little is known about the formation of niches, local micro-environments required for stem-cell maintenance. Here we develop an in vivo assay for adult haematopoietic stem-cell (HSC) niche formation. With this assay, we identified a population of progenitor cells with surface markers CD45(-)Tie2(-)alpha(V)(+)CD105(+)Thy1.1(-) (CD105(+)Thy1(-)) that, when sorted from 15.5 days post-coitum fetal bones and transplanted under the adult mouse kidney capsule, could recruit host-derived blood vessels, produce donor-derived ectopic bones through a cartilage intermediate and generate a marrow cavity populated by host-derived long-term reconstituting HSC (LT-HSC). In contrast, CD45(-)Tie2(-)alpha(V)(+)CD105(+)Thy1(+) (CD105(+)Thy1(+)) fetal bone progenitors form bone that does not contain a marrow cavity. Suppressing expression of factors involved in endochondral ossification, such as osterix and vascular endothelial growth factor (VEGF), inhibited niche generation. CD105(+)Thy1(-) progenitor populations derived from regions of the fetal mandible or calvaria that do not undergo endochondral ossification formed only bone without marrow in our assay. Collectively, our data implicate endochondral ossification, bone formation that proceeds through a cartilage intermediate, as a requirement for adult HSC niche formation.

Development of bioactive porous α-TCP/HAp beads for bone tissue engineering.

PubMed

Asaoka, Teruo; Ohtake, Shoji; Furukawa, Katsuko S; Tamura, Akito; Ushida, Takashi

2013-11-01

Porous beads of bioactive ceramics such as hydroxyapatite (HAp) and tribasic calcium phosphate (TCP) are considered a promising scaffold for cultivating bone cells. To realize this, α-TCP/HAp functionally graded porous beads are fabricated with two main purposes: to maintain the function of the scaffold with sufficient strength up to the growth of new bone, and is absorbed completely after the growth. HAp is a bioactive material that has both high strength and strong tissue-adhesive properties, but is not readily absorbed by the human body. On the contrary, α-TCP is highly bioabsorbable, resulting in a scaffold that is absorbed before it is completely replaced by bone. In this study, we produced porous, bead-shaped carriers as scaffolds for osteoblast culture. To control the solubility in vivo, the fabricated beads contained α-TCP at the center and HAp at the surface. Cell adaptability of these beads for bone tissue engineering was confirmed in vitro. It was found that α-TCP/HAp bead carriers exhibit low toxicity in the initial stages of cell seeding and cell adhesion. The presence of HAp in the composite bead form effectively increased ALP activity. In conclusion, it is suggested that these newly developed α-TCP/HAp beads are a promising tool for bone tissue engineering. Copyright © 2013 Wiley Periodicals, Inc.

Controlled release of drugs in electrosprayed nanoparticles for bone tissue engineering.

PubMed

Jayaraman, Praveena; Gandhimathi, Chinnasamy; Venugopal, Jayarama Reddy; Becker, David Laurence; Ramakrishna, Seeram; Srinivasan, Dinesh Kumar

2015-11-01

Generating porous topographic substrates, by mimicking the native extracellular matrix (ECM) to promote the regeneration of damaged bone tissues, is a challenging process. Generally, scaffolds developed for bone tissue regeneration support bone cell growth and induce bone-forming cells by natural proteins and growth factors. Limitations are often associated with these approaches such as improper scaffold stability, and insufficient cell adhesion, proliferation, differentiation, and mineralization with less growth factor expression. Therefore, the use of engineered nanoparticles has been rapidly increasing in bone tissue engineering (BTE) applications. The electrospray technique is advantageous over other conventional methods as it generates nanomaterials of particle sizes in the micro/nanoscale range. The size and charge of the particles are controlled by regulating the polymer solution flow rate and electric voltage. The unique properties of nanoparticles such as large surface area-to-volume ratio, small size, and higher reactivity make them promising candidates in the field of biomedical engineering. These nanomaterials are extensively used as therapeutic agents and for drug delivery, mimicking ECM, and restoring and improving the functions of damaged organs. The controlled and sustained release of encapsulated drugs, proteins, vaccines, growth factors, cells, and nucleotides from nanoparticles has been well developed in nanomedicine. This review provides an insight into the preparation of nanoparticles by electrospraying technique and illustrates the use of nanoparticles in drug delivery for promoting bone tissue regeneration. Copyright © 2015 Elsevier B.V. All rights reserved.

The effect of cationically-modified phosphorylcholine polymers on human osteoblasts in vitro and their effect on bone formation in vivo.

PubMed

Lawton, Jonathan M; Habib, Mariam; Ma, Bingkui; Brooks, Roger A; Best, Serena M; Lewis, Andrew L; Rushton, Neil; Bonfield, William

2017-08-17

The effect of introducing cationic charge into phosphorylcholine (PC)-based polymers has been investigated in this study with a view to using these materials as coatings to improve bone formation and osseointegration at the bone-implant interface. PC-based polymers, which have been used in a variety of medical devices to improve biocompatibility, are associated with low protein adsorption resulting in reduced complement activation, inflammatory response and cell adhesion. However, in some applications, such as orthopaedics, good integration between the implant and bone is needed to allow the distribution of loading stresses and a bioactive response is required. It has previously been shown that the incorporation of cationic charge into PC-based polymers may increase protein adsorption that stimulates subsequent cell adhesion. In this paper, the effect of cationic charge in PC-based polymers on human osteoblasts (HObs) in vitro and the effect of these polymers on bone formation in the rat tibia was assessed. Increasing PC positive surface charge increased HOb cell adhesion and stimulated increased cell differentiation and the production of calcium phosphate deposits. However, when implanted in bone these materials were at best biotolerant, stimulating the production of fibrous tissue and areas of loosely associated matrix (LAM) around the implant. Their development, as formulated in this study, as bone interfacing implant coatings is therefore not warranted.

The physiopathology of avascular necrosis of the femoral head: an update.

PubMed

Guerado, Enrique; Caso, Enrique

2016-12-01

The physiopathology of the femoral head bone necrosis is similar for children and for adults. The disease is characterized by apoptosis of bone cells - bone marrow and bone forming cells-resulting in head collapse with a subsequent lesion of the overlying cartilage, and therefore flattening of the rounded surface shape of the head articulating with the acetabulum, provoking, eventually, secondary osteoarthritis. When the disease becomes clinically evident already destructive phenomena have occurred and collapse will eventually ensue. In children, because epiphyseal cartilage has growth capabilities, lost epiphyseal height can be recovered, however in adults collapse is irreversible. In this paper the physiopathology of this disease is examined as well as its implication for treatment. Prevention by genetic studies is discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Plasma-deposited fluorocarbon polymer films on titanium for preventing cell adhesion: a surface finishing for temporarily used orthopaedic implants

NASA Astrophysics Data System (ADS)

Finke, B.; Testrich, H.; Rebl, H.; Walschus, U.; Schlosser, M.; Zietz, C.; Staehlke, S.; Nebe, J. B.; Weltmann, K. D.; Meichsner, J.; Polak, M.

2016-06-01

The design of a titanium implant surface should ideally support its later application in clinical use. Temporarily used implants have to fulfil requirements different from permanent implants: they should ensure the mechanical stabilization of the bone stock but in trauma surgery they should not be integrated into the bone because they will be removed after fracture healing. Finishing of the implant surface by a plasma-fluorocarbon-polymer (PFP) coating is a possible approach for preventing cell adhesion of osteoblasts. Two different low pressure gas-discharge plasma processes, microwave (MW 2.45 GHz) and capacitively coupled radio frequency (RF 13.56 MHz) plasma, were applied for the deposition of the PFP film using a mixture of the precursor octafluoropropane (C3F8) and hydrogen (H2). The thin films were characterized by x-ray photoelectron spectroscopy, Fourier transform infrared reflection absorption spectroscopy, and water contact angle measurements. Cell culture experiments show that cell adhesion and spreading of MG-63 osteoblasts were clearly reduced or nonexistent on these surfaces, also after 24 h of storage in the cell culture medium. In vivo data demonstrated that the local inflammatory tissue response for the PFP films deposited in MW and RF plasma were comparable to uncoated controls.

Hydrocarbon Deposition Attenuates Osteoblast Activity on Titanium

PubMed Central

Hayashi, R.; Ueno, T.; Migita, S.; Tsutsumi, Y.; Doi, H.; Ogawa, T.; Hanawa, T.; Wakabayashi, N.

2014-01-01

Although the reported percentage of bone-implant contact is far lower than 100%, the cause of such low levels of bone formation has rarely been investigated. This study tested the negative biological effect of hydrocarbon deposition onto titanium surfaces, which has been reported to be inevitable. Osteogenic MC3T3-E1 cells were cultured on titanium disks on which the carbon concentration was experimentally regulated to achieve carbon/titanium (C/Ti) ratios of 0.3, 0.7, and 1.0. Initial cellular activities such as cell attachment and cell spreading were concentration-dependently suppressed by the amount of carbon on the titanium surface. The osteoblastic functions of alkaline phosphatase activity and calcium mineralization were also reduced by more than 40% on the C/Ti (1.0) surface. These results indicate that osteoblast activity is influenced by the degree of hydrocarbon contamination on titanium implants and suggest that hydrocarbon decomposition before implant placement may increase the biocompatibility of titanium. PMID:24868012

Computational modelling of biomaterial surface interactions with blood platelets and osteoblastic cells for the prediction of contact osteogenesis.

PubMed

Amor, N; Geris, L; Vander Sloten, J; Van Oosterwyck, H

2011-02-01

Surface microroughness can induce contact osteogenesis (bone formation initiated at the implant surface) around oral implants, which may result from different mechanisms, such as blood platelet-biomaterial interactions and/or interaction with (pre-)osteoblast cells. We have developed a computational model of implant endosseous healing that takes into account these interactions. We hypothesized that the initial attachment and growth factor release from activated platelets is crucial in achieving contact osteogenesis. In order to investigate this, a computational model was applied to an animal experiment [7] that looked at the effect of surface microroughness on endosseous healing. Surface-specific model parameters were implemented based on in vitro data (Lincks et al. Biomaterials 1998;19:2219-32). The predicted spatio-temporal patterns of bone formation correlated with the histological data. It was found that contact osteogenesis could not be predicted if only the osteogenic response of cells was up-regulated by surface microroughness. This could only be achieved if platelet-biomaterial interactions were sufficiently up-regulated as well. These results confirmed our hypothesis and demonstrate the added value of the computational model to study the importance of surface-mediated events for peri-implant endosseous healing. Copyright © 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Effects of titanium surface anodization with CaP incorporation on human osteoblastic response

PubMed Central

OLIVEIRA, Natássia Cristina Martins; MOURA, Camilla Christian Gomes; ZANETTA-BARBOSA, Darceny; MENDONÇA, Daniela Baccelli Silveira; MENDONÇA, Gustavo; DECHICHI, Paula

2015-01-01

In this study we investigated whether anodization with calcium phosphate (CaP) incorporation (Vulcano®) enhances growth factors secretion, osteoblast-specific gene expression, and cell viability, when compared to acid etched surfaces (Porous®) and machined surfaces (Screw®) after 3 and 7 days. Results showed significant cell viability for Porous and Vulcano at day 7, when compared with Screw (p=0.005). At the same time point, significant differences regarding runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP) and bone sialoprotein (BSP) expression were found for all surfaces (p<0.05), but with greater fold induction for Porous and Vulcano. The secretion of transforming growth factor β1 (TGF-β1) and bone morphogenetic protein 2 (BMP-2) was not significantly affected by surface treatment in any experimental time (p>0.05). Although no significant correlation was found for growth factors secretion and Runx2 expression, a significant positive correlation between this gene and ALP/BSP expression showed that their strong association is independent on the type of surface. The incorporation of CaP affected the biological parameters evaluated similar to surfaces just acid etched. The results presented here support the observations that roughness also may play an important role in determining cell response. PMID:23498218

Crystal structures of CaSiO3 polymorphs control growth and osteogenic differentiation of human mesenchymal stem cells on bioceramic surfaces.

PubMed

Zhang, Nianli; Molenda, James A; Mankoci, Steven; Zhou, Xianfeng; Murphy, William L; Sahai, Nita

2013-10-01

The repair and replacement of damaged or diseased human bone tissue requires a stable interface between the orthopedic implant and living tissue. The ideal material should be both osteoconductive (promote bonding to bone) and osteoinductive (induce osteogenic differentiation of cells and generate new bone). Partially resorbable bioceramic materials with both properties are developed by expensive trial-and-error methods. Structure-reactivity relationships for predicting the osteoinductive properties of ceramics would significantly increase the efficiency of developing materials for bone tissue engineering. Here we propose the novel hypothesis that the crystal structure of a bioceramic controls the release rates, subsequent surface modifications due to precipitation of new phases, and thus, the concentrations of soluble factors, and ultimately, the attachment, viability and osteogenic differentiation of human Mesenchymal Stem Cells (hMSCs). To illustrate our hypothesis, we used two CaSiO 3 polymorphs, pseudo-wollastonite (psw, β-CaSiO 3 ) and wollastonite (wol, α-CaSiO 3 ) as scaffolds for hMSC culture. Polymorphs are materials which have identical chemical composition and stoichiometry, but different crystal structures. We combined the results of detailed surface characterizations, including environmental Scanning Electron Microscopy (SEM) back-scattered imaging, and spot-analysis and 2D elemental mapping by SEM-Energy Dispersive X-ray (SEM-EDX), High Resolution Transmission Electron Microscopy (HRTEM) and surface roughness analysis; culture medium solution analyses; and molecular/genetic assays from cell culture. Our results confirmed the hypothesis that the psw polymorph, which has a strained silicate ring structure, is more osteoinductive than the wol polymorph, which has a more stable, open silicate chain structure. The observations could be attributed to easier dissolution (resorption) of psw compared to wol, which resulted in concentration profiles that were more osteoinductive for the former. Thus, we showed that crystal structure is a fundamental parameter to be considered in the intelligent design of pro-osteogenic, partially resorbable bioceramics.

Role of Bruton's tyrosine kinase (BTK) in growth and metastasis of INA6 myeloma cells

PubMed Central

Bam, R; Venkateshaiah, S U; Khan, S; Ling, W; Randal, S S; Li, X; Zhang, Q; van Rhee, F; Barlogie, B; Epstein, J; Yaccoby, S

2014-01-01

Bruton's tyrosine kinase (BTK) and the chemokine receptor CXCR4 are linked in various hematologic malignancies. The aim of the study was to understand the role of BTK in myeloma cell growth and metastasis using the stably BTK knockdown luciferase-expressing INA6 myeloma line. BTK knockdown had reduced adhesion to stroma and migration of myeloma cells toward stromal cell-derived factor-1. BTK knockdown had no effect on short-term in vitro growth of myeloma cells, although clonogenicity was inhibited and myeloma cell growth was promoted in coculture with osteoclasts. In severe combined immunodeficient-rab mice with contralaterally implanted pieces of bones, BTK knockdown in myeloma cells promoted their proliferation and growth in the primary bone but suppressed metastasis to the contralateral bone. BTK knockdown myeloma cells had altered the expression of genes associated with adhesion and proliferation and increased mammalian target of rapamycin signaling. In 176 paired clinical samples, BTK and CXCR4 expression was lower in myeloma cells purified from a focal lesion than from a random site. BTK expression in random-site samples was correlated with proportions of myeloma cells expressing cell surface CXCR4. Our findings highlight intratumoral heterogeneity of myeloma cells in the bone marrow microenvironment and suggest that BTK is involved in determining proliferative, quiescent or metastatic phenotypes of myeloma cells. PMID:25083818

Morphological and histochemical studies of bone and cartilage during periods of stimulated weightlessness

NASA Technical Reports Server (NTRS)

Doty, S. B.

1984-01-01

Rats which were subjected to spaceflight for 2-4 weeks showed considerable loss in ability to form new bone. Animals which are placed into nonweight bearing positions, as a model to simulate the absence of gravity here on the Earth's surface. Show a similar decline in new bone formation. It is suggested that the mechanisms underlying these changes may be the result of reduced transmission of gravitational force to the skeletal cells.

Platelet Dysfunction and a High Bone Mass Phenotype in a Murine Model of Platelet-Type von Willebrand Disease

PubMed Central

Suva, Larry J.; Hartman, Eric; Dilley, Joshua D.; Russell, Susan; Akel, Nisreen S.; Skinner, Robert A.; Hogue, William R.; Budde, Ulrich; Varughese, Kottayil I.; Kanaji, Taisuke; Ware, Jerry

2008-01-01

The platelet glycoprotein Ib-IX receptor binds surface-bound von Willebrand factor and supports platelet adhesion to damaged vascular surfaces. A limited number of mutations within the glycoprotein Ib-IX complex have been described that permit a structurally altered receptor to interact with soluble von Willebrand factor, and this is the molecular basis of platelet-type von Willebrand disease. We have developed and characterized a mouse model of platelet-type von Willebrand disease (G233V) and have confirmed a platelet phenotype mimicking the human disorder. The mice have a dramatic increase in splenic megakaryocytes and splenomegaly. Recent studies have demonstrated that hematopoetic cells can influence the differentiation of osteogenic cells. Thus, we examined the skeletal phenotype of mice expressing the G233V variant complex. At 6 months of age, G233V mice exhibit a high bone mass phenotype with an approximate doubling of trabecular bone volume in both the tibia and femur. Serum measures of bone resorption were significantly decreased in G233V animals. With decreased bone resorption, cortical thickness was increased, medullary area decreased, and consequently, the mechanical strength of the femur was significantly increased. Using ex vivo bone marrow cultures, osteoclast-specific staining in the G233V mutant marrow was diminished, whereas osteoblastogenesis was unaffected. These studies provide new insights into the relationship between the regulation of megakaryocytopoiesis and bone mass. PMID:18187573

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

Bone-targeted cabazitaxel nanoparticles for metastatic prostate cancer skeletal lesions and pain.

PubMed

Gdowski, Andrew S; Ranjan, Amalendu; Sarker, Marjana R; Vishwanatha, Jamboor K

2017-09-01

The aim of this study was to develop a novel cabazitaxel bone targeted nanoparticle (NP) system for improved drug delivery to the bone microenvironment. Nanoparticles were developed using poly(D,L-lactic-co-glycolic acid) and cabazitaxel as the core with amino-bisphosphonate surface conjugation. Optimization of nanoparticle physiochemical properties, in vitro evaluation in prostate cancer cell lines and in vivo testing in an intraosseous model of metastatic prostate cancer was performed. This bone targeted cabazitaxel nanocarrier system showed significant reduction in tumor burden, while at the same time maintaining bone structure integrity and reducing pain in the mouse tumor limb. This bone microenvironment targeted nanoparticle system and clinically relevant approach of evaluation represents a promising advancement for treating bone metastatic cancer.

Abnormal arrangement of a collagen/apatite extracellular matrix orthogonal to osteoblast alignment is constructed by a nanoscale periodic surface structure.

PubMed

Matsugaki, Aira; Aramoto, Gento; Ninomiya, Takafumi; Sawada, Hiroshi; Hata, Satoshi; Nakano, Takayoshi

2015-01-01

Morphological and directional alteration of cells is essential for structurally appropriate construction of tissues and organs. In particular, osteoblast alignment is crucial for the realization of anisotropic bone tissue microstructure. In this article, the orientation of a collagen/apatite extracellular matrix (ECM) was established by controlling osteoblast alignment using a surface geometry with nanometer-sized periodicity induced by laser ablation. Laser irradiation induced self-organized periodic structures (laser-induced periodic surface structures; LIPSS) with a spatial period equal to the wavelength of the incident laser on the surface of biomedical alloys of Ti-6Al-4V and Co-Cr-Mo. Osteoblast orientation was successfully induced parallel to the grating structure. Notably, both the fibrous orientation of the secreted collagen matrix and the c-axis of the produced apatite crystals were orientated orthogonal to the cell direction. To the best of our knowledge, this is the first report demonstrating that bone tissue anisotropy is controllable, including the characteristic organization of a collagen/apatite composite orthogonal to the osteoblast orientation, by controlling the cell alignment using periodic surface geometry. Copyright © 2014 Elsevier Ltd. All rights reserved.

Bisphosphonate action. Alendronate localization in rat bone and effects on osteoclast ultrastructure.

PubMed Central

Sato, M; Grasser, W; Endo, N; Akins, R; Simmons, H; Thompson, D D; Golub, E; Rodan, G A

1991-01-01

Studies of the mode of action of the bisphosphonate alendronate showed that 1 d after the injection of 0.4 mg/kg [3H]alendronate to newborn rats, 72% of the osteoclastic surface, 2% of the bone forming, and 13% of all other surfaces were densely labeled. Silver grains were seen above the osteoclasts and no other cells. 6 d later the label was 600-1,000 microns away from the epiphyseal plate and buried inside the bone, indicating normal growth and matrix deposition on top of alendronate-containing bone. Osteoclasts from adult animals, infused with parathyroid hormone-related peptide (1-34) and treated with 0.4 mg/kg alendronate subcutaneously for 2 d, all lacked ruffled border but not clear zone. In vitro alendronate bound to bone particles with a Kd of approximately 1 mM and a capacity of 100 nmol/mg at pH 7. At pH 3.5 binding was reduced by 50%. Alendronate inhibited bone resorption by isolated chicken or rat osteoclasts when the amount on the bone surface was around 1.3 x 10(-3) fmol/microns 2, which would produce a concentration of 0.1-1 mM in the resorption space if 50% were released. At these concentrations membrane leakiness to calcium was observed. These findings suggest that alendronate binds to resorption surfaces, is locally released during acidification, the rise in concentration stops resorption and membrane ruffling, without destroying the osteoclasts. Images PMID:1661297

A bioactive metallurgical grade porous silicon-polytetrafluoroethylene sheet for guided bone regeneration applications.

PubMed

Chadwick, E G; Clarkin, O M; Raghavendra, R; Tanner, D A

2014-01-01

The properties of porous silicon make it a promising material for a host of applications including drug delivery, molecular and cell-based biosensing, and tissue engineering. Porous silicon has previously shown its potential for the controlled release of pharmacological agents and in assisting bone healing. Hydroxyapatite, the principle constituent of bone, allows osteointegration in vivo, due to its chemical and physical similarities to bone. Synthetic hydroxyapatite is currently applied as a surface coating to medical devices and prosthetics, encouraging bone in-growth at their surface and improving osseointegration. This paper examines the potential for the use of an economically produced porous silicon particulate-polytetrafluoroethylene sheet for use as a guided bone regeneration device in periodontal and orthopaedic applications. The particulate sheet is comprised of a series of microparticles in a polytetrafluoroethylene matrix and is shown to produce a stable hydroxyapatite on its surface under simulated physiological conditions. The microstructure of the material is examined both before and after simulated body fluid experiments for a period of 1, 7, 14 and 30 days using Scanning Electron Microscopy. The composition is examined using a combination of Energy Dispersive X-ray Spectroscopy, Thin film X-ray diffraction, Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy and the uptake/release of constituents at the fluid-solid interface is explored using Inductively Coupled Plasma-Optical Emission Spectroscopy. Microstructural and compositional analysis reveals progressive growth of crystalline, 'bone-like' apatite on the surface of the material, indicating the likelihood of close bony apposition in vivo.

Effects of Build Orientation on Surface Morphology and Bone Cell Activity of Additively Manufactured Ti6Al4V Specimens.

PubMed

Weißmann, Volker; Drescher, Philipp; Seitz, Hermann; Hansmann, Harald; Bader, Rainer; Seyfarth, Anika; Klinder, Annett; Jonitz-Heincke, Anika

2018-05-29

Additive manufacturing of lightweight or functional structures by selective laser beam (SLM) or electron beam melting (EBM) is widespread, especially in the field of medical applications. SLM and EBM processes were applied to prepare Ti6Al4V test specimens with different surface orientations (0°, 45° and 90°). Roughness measurements of the surfaces were conducted and cell behavior on these surfaces was analyzed. Hence, human osteoblasts were seeded on test specimens to determine cell viability (metabolic activity, live-dead staining) and gene expression of collagen type 1 (Col1A1), matrix metalloprotease (MMP) 1 and its natural inhibitor, TIMP1, after 3 and 7 days. The surface orientation of specimens during the manufacturing process significantly influenced the roughness. Surface roughness showed significant impact on cellular viability, whereas differences between the time points day 3 and 7 were not found. Collagen type 1 mRNA synthesis rates in human osteoblasts were enhanced with increasing roughness. Both manufacturing techniques further influenced the induction of bone formation process in the cell culture. Moreover, the relationship between osteoblastic collagen type 1 mRNA synthesis rates and specimen orientation during the building process could be characterized by functional formulas. These findings are useful in the designing of biomedical applications and medical devices.

A COMPUTATIONAL ANALYSIS OF BONE FORMATION IN THE CRANIAL VAULT USING A COUPLED REACTION-DIFFUSION-STRAIN MODEL

PubMed Central

LEE, CHANYOUNG; RICHTSMEIER, JOAN T.; KRAFT, REUBEN H.

2017-01-01

Bones of the murine cranial vault are formed by differentiation of mesenchymal cells into osteoblasts, a process that is primarily understood to be controlled by a cascade of reactions between extracellular molecules and cells. We assume that the process can be modeled using Turing’s reaction-diffusion equations, a mathematical model describing the pattern formation controlled by two interacting molecules (activator and inhibitor). In addition to the processes modeled by reaction-diffusion equations, we hypothesize that mechanical stimuli of the cells due to growth of the underlying brain contribute significantly to the process of cell differentiation in cranial vault development. Structural analysis of the surface of the brain was conducted to explore the effects of the mechanical strain on bone formation. We propose a mechanobiological model for the formation of cranial vault bones by coupling the reaction-diffusion model with structural mechanics. The mathematical formulation was solved using the finite volume method. The computational domain and model parameters are determined using a large collection of experimental data that provide precise three dimensional (3D) measures of murine cranial geometry and cranial vault bone formation for specific embryonic time points. The results of this study suggest that mechanical strain contributes information to specific aspects of bone formation. Our mechanobiological model predicts some key features of cranial vault bone formation that were verified by experimental observations including the relative location of ossification centers of individual vault bones, the pattern of cranial vault bone growth over time, and the position of cranial vault sutures. PMID:29225392

Bioactivity and cytocompatibility of zirconia (ZrO(2)) films fabricated by cathodic arc deposition.

PubMed

Liu, Xuanyong; Huang, Anping; Ding, Chuanxian; Chu, Paul K

2006-07-01

Zirconium oxide thin films were fabricated on silicon wafers using a filtered cathodic arc system in concert with oxygen plasma. The structure and phase composition of the zirconium oxide thin films were characterized by atomic force microscopy (AFM), X-ray diffraction (XRD), Rutherford backscattering spectrometry (RBS), and transmission electron microscopy (TEM). The bioactivity was assessed by investigating the formation of apatite on the film surface after soaking in simulated body fluids. Bone marrow mesenchymal stem cells (BMMSC) were used to further evaluate the cytocompatibility of the materials. The results indicate that the films are composed of stoichiometric ZrO(2) and the composition is quite uniform throughout the thickness. Bone-like apatite can be formed on the surface of the ZrO(2) thin film in our SBF immersion experiments, suggesting that the surface is bioactive. The outermost layer of the ZrO(2) thin film comprises nano-sized particles that can be identified by AFM images taken on the thin film surface and TEM micrographs obtained from the interface between the ZrO(2) thin film and apatite layer. The nanostructured surface is believed to be the key factor that apatite is induced to precipitate on the surface. Bone marrow mesenchymal stem cells are observed to grow and proliferate in good states on the film surface. Our results show that ZrO(2) thin films fabricated by cathodic arc deposition exhibit favorable bioactivity and cytocompatibility.

Placenta Derived Mesenchymal Stem Cells Hosted on RKKP Glass-Ceramic: A Tissue Engineering Strategy for Bone Regenerative Medicine Applications

PubMed Central

Fosca, Marco; De Bonis, Angela; Curcio, Mariangela; Lolli, Maria Grazia; De Stefanis, Adriana; Marchese, Rodolfo; Rau, Julietta V.

2016-01-01

In tissue engineering protocols, the survival of transplanted stem cells is a limiting factor that could be overcome using a cell delivery matrix able to support cell proliferation and differentiation. With this aim, we studied the cell-friendly and biocompatible behavior of RKKP glass-ceramic coated Titanium (Ti) surface seeded with human amniotic mesenchymal stromal cells (hAMSCs) from placenta. The sol-gel synthesis procedure was used to prepare the RKKP glass-ceramic material, which was then deposited onto the Ti surface by Pulsed Laser Deposition method. The cell metabolic activity and proliferation rate, the cytoskeletal actin organization, and the cell cycle phase distribution in hAMSCs seeded on the RKKP coated Ti surface revealed no significant differences when compared to the cells grown on the treated plastic Petri dish. The health of of hAMSCs was also analysed studying the mRNA expressions of MSC key genes and the osteogenic commitment capability using qRT-PCR analysis which resulted in being unchanged in both substrates. In this study, the combination of the hAMSCs' properties together with the bioactive characteristics of RKKP glass-ceramics was investigated and the results obtained indicate its possible use as a new and interesting cell delivery system for bone tissue engineering and regenerative medicine applications. PMID:28078286

Polydopamine mediated assembly of hydroxyapatite nanoparticles and bone morphogenetic protein-2 on magnesium alloys for enhanced corrosion resistance and bone regeneration.

PubMed

Jiang, Yanan; Wang, Bi; Jia, Zhanrong; Lu, Xiong; Fang, Liming; Wang, Kefeng; Ren, Fuzeng

2017-10-01

Magnesium alloys have the great potential to be used as orthopedic implants due to their biodegradability and mechanical resemblance to human cortical bone. However, the rapid degradation in physiological environment with the evolution of hydrogen gas release hinders their clinical applications. In this study, we developed a novel functional and biocompatible coating strategy through polydopamine mediated assembly of hydroxyapatite nanoparticles and growth factor, bone morphogenetic protein-2 (BMP-2), onto the surface of AZ31 Mg alloys. Such functional coating has strong bonding with the substrate and can increase surface hydrophilicity of magnesium alloys. In vitro electrochemical corrosion and hydrogen evolution tests demonstrate that the coating can significantly enhance the corrosion resistance and therefore slow down the degradation of AZ31 Mg alloys. In vitro cell culture reveals that immobilization of HA nanoparticles and BMP-2 can obviously promote cell adhesion and proliferation. Furthermore, in vivo implantation tests indicate that with the synergistic effects of HA nanoparticles and BMP-2, the coating does not cause obvious inflammatory response and can significantly reduce the biodegradation rate of the magnesium alloys and induce the new bone formation adjacent to the implants. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2750-2761, 2017. © 2017 Wiley Periodicals, Inc.

Interleukin-3 Does Not Affect the Differentiation of Mast Cells Derived from Human Bone Marrow Progenitors

PubMed Central

Shimizu, Yuji; Matsumoto, Kenji; Okayama, Yoshimichi; Kentaro, Sakai; Maeno, Toshitaka; Suga, Tatsuo; Miura, Toru; Takai, Shinji; Kurabayashi, Masahiko; Saito, Hirohisa

2008-01-01

Although IL-3 is commonly used for culture of human progenitor-derived mast cells together with Stem cell factor (SCF) and IL-6, the effect of IL-3 on human mast cell differentiation has not been well elucidated. Human bone marrow CD34+ progenitors were cultured for up to 12 weeks in the presence of rhSCF and rhIL-6 either with rhIL-3 (IL-3 (+)) or without rhIL-3 (IL-3 (−)) for the initial 1-week of culture. Total cell number increased at 2 weeks in IL-3 (+), as compared to IL-3 (−), but changes in the appearance of mast cells were delayed. When IL-3 was present for the initial 1-week culture, granules looked more mature with IL-3 than without IL-3. However, tryptase and chymase contents, and surface antigen expression (CD18, CD51, CD54, and CD117) were not altered by IL-3. Surface expression and mRNA level of FcεRIα and histamine release by crosslinking of FcεRIα did not differ from one preparation to the next. GeneChip analysis revealed that no significant differences were observed between IL-3 (+) and IL-3 (−) cells either when inactivated or activated by aggregation of FcεRIα. These findings indicate that initial incubation of human bone marrow CD34+ progenitors with IL-3 does not affect the differentiation of mast cells. PMID:18214796

Surface modification of nano-silica on the ligament advanced reinforcement system for accelerated bone formation: primary human osteoblasts testing in vitro and animal testing in vivo.

PubMed

Li, Mengmeng; Wang, Shiwen; Jiang, Jia; Sun, Jiashu; Li, Yuzhuo; Huang, Deyong; Long, Yun-Ze; Zheng, Wenfu; Chen, Shiyi; Jiang, Xingyu

2015-05-07

The Ligament Advanced Reinforcement System (LARS) has been considered as a promising graft for ligament reconstruction. To improve its biocompatibility and effectiveness on new bone formation, we modified the surface of a polyethylene terephthalate (PET) ligament with nanoscale silica using atom transfer radical polymerization (ATRP) and silica polymerization. The modified ligament is tested by both in vitro and in vivo experiments. Human osteoblast testing in vitro exhibits an ∼21% higher value in cell viability for silica-modified grafts compared with original grafts. Animal testing in vivo shows that there is new formed bone in the case of a nanoscale silica-coated ligament. These results demonstrate that our approach for nanoscale silica surface modification on LARS could be potentially applied for ligament reconstruction.

Selective Laser Melting: a regular unit cell approach for the manufacture of porous, titanium, bone in-growth constructs, suitable for orthopedic applications.

PubMed

Mullen, Lewis; Stamp, Robin C; Brooks, Wesley K; Jones, Eric; Sutcliffe, Christopher J

2009-05-01

In this study, a novel porous titanium structure for the purpose of bone in-growth has been designed, manufactured and evaluated. The structure was produced by Selective Laser Melting (SLM); a rapid manufacturing process capable of producing highly intricate, functionally graded parts. The technique described utilizes an approach based on a defined regular unit cell to design and produce structures with a large range of both physical and mechanical properties. These properties can be tailored to suit specific requirements; in particular, functionally graded structures with bone in-growth surfaces exhibiting properties comparable to those of human bone have been manufactured. The structures were manufactured and characterized by unit cell size, strand diameter, porosity, and compression strength. They exhibited a porosity (10-95%) dependant compression strength (0.5-350 Mpa) comparable to the typical naturally occurring range. It is also demonstrated that optimized structures have been produced that possesses ideal qualities for bone in-growth applications and that these structures can be applied in the production of orthopedic devices. (c) 2008 Wiley Periodicals, Inc.

Strontium (Sr) and silver (Ag) loaded nanotubular structures with combined osteoinductive and antimicrobial activities.

PubMed

Cheng, Hao; Xiong, Wei; Fang, Zhong; Guan, Hanfeng; Wu, Wei; Li, Yong; Zhang, Yong; Alvarez, Mario Moisés; Gao, Biao; Huo, Kaifu; Xu, Jiangwen; Xu, Na; Zhang, Chengcheng; Fu, Jijiang; Khademhosseini, Ali; Li, Feng

2016-02-01

Two frequent problems are associated with the titanium surfaces of bone/dental implants: lack of native tissue integration and associated infection. These problems have prompted a significant body of research regarding the modification of these surfaces. The present study describes a hydrothermal treatment for the fabrication of strontium (Sr) and silver (Ag) loaded nanotubular structures with different tube diameters on titanium surfaces. The Sr loading from a Sr(OH)2 solution was regulated by the size of the inner diameter of the titanium nanotubes (NT) (30nm or 80nm, formed at 10V or 40V, respectively). The quantity of Ag was adjusted by immersing the samples in 1.5 or 2.0M AgNO3 solutions. Sr and Ag were released in a controllable and prolonged matter from the NT-Ag.Sr samples, with negligible cytotoxicity. Prominent antibacterial activity was observed due to the release of Ag. Sr incorporation enhanced the initial cell adhesion, migration, and proliferation of preosteoblast MC3T3-E1 cells. Sr release also up-regulated the expression of osteogenic genes and induced mineralization, as suggested by the presence of more mineralized calcium nodules in cells cultured on NT-Ag.Sr surfaces. In vivo experiments showed that the Sr-loaded samples accelerated the formation of new bone in both osteoporosis and bone defect models, as confirmed by X-ray, Micro-CT evaluation, and histomorphometric analysis of rats implanted with NT-Ag.Sr samples. The antibacterial activity and outstanding osteogenic properties of NT-Ag.Sr samples highlight their excellent potential for use in clinical applications. Two frequent problems associated with Ti surfaces, widely used in orthopedic and dental arenas, are their lack of native tissue integration and risk of infection. We describe a novel approach for the fabrication of strontium (Sr) and silver (Ag) loaded nanotubular structures on titanium surfaces. A relevant aspect of this work is the demonstration of long-lasting and controllable Ag release, leading to excellent antibacterial and anti-adherent properties against methicillin-resistant Staphylococcus aureus (MRSA), and Gram-negative bacteria such as Escherichia coli. The extended release of Sr accelerates the filling of bone defects by improving the repair of damaged cortical bone and increasing trabecular bone microarchitecture. Our results highlight the potential of Sr and Ag loaded nanotubular structures for use in clinical applications. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Rapid isolation of bone marrow mesenchymal stromal cells using integrated centrifuge-based technology.

PubMed

Meppelink, Amanda M; Wang, Xing-Hua; Bradica, Gino; Barron, Kathryn; Hiltz, Kathleen; Liu, Xiang-Hong; Goldman, Scott M; Vacanti, Joseph P; Keating, Armand; Hoganson, David M

2016-06-01

The use of bone marrow-derived mesenchymal stromal cells (MSCs) in cell-based therapies is currently being developed for a number of diseases. Thus far, the clinical results have been inconclusive and variable, in part because of the variety of cell isolation procedures and culture conditions used in each study. A new isolation technique that streamlines the method of concentration and demands less time and attention could provide clinical and economic advantages compared with current methodologies. In this study, we evaluated the concentrating capability of an integrated centrifuge-based technology compared with standard Ficoll isolation. MSCs were concentrated from bone marrow aspirate using the new device and the Ficoll method. The isolation capabilities of the device and the growth characteristics, secretome production, and differentiation capacity of the derived cells were determined. The new MSC isolation device concentrated the bone marrow in 90 seconds and resulted in a mononuclear cell yield 10-fold higher and with a twofold increase in cell retention compared with Ficoll. The cells isolated using the device were shown to exhibit similar morphology and functional activity as assessed by growth curves and secretome production compared to the Ficoll-isolated cells. The surface marker and trilineage differentiation profile of the device-isolated cells was consistent with the known profile of MSCs. The faster time to isolation and greater cell yield of the integrated centrifuge-based technology may make this an improved approach for MSC isolation from bone marrow aspirates. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Surface-mediated delivery of siRNA from fibrin hydrogels for knockdown of the BMP-2 binding antagonist noggin.

PubMed

Kowalczewski, Christine J; Saul, Justin M

2015-10-01

Antagonists and inhibitory molecules responsible for maintaining tissue homeostasis can present a significant barrier to healing when tissue engineering/regenerative medicine strategies are employed. One example of this situation is the up-regulation of antagonists such as noggin in response to increasing concentrations of bone morphogenetic protein-2 (BMP-2) present from endogenous bone repair processes or delivered exogenously from biomaterials (synthetic bone grafts). While recombinant human (rh)BMP-2 delivered from synthetic bone grafts has been shown to be an effective alternative to autografts and allografts, the supraphysiological doses of rhBMP-2 have led to clinically-adverse side effects. The high rhBMP-2 dosage may be required, in part, to overcome the presence of antagonists such as noggin. Small interfering RNA (siRNA) is an appealing approach to overcome this problem because it can knock-down antagonists or inhibitory molecules in a temporary manner. Here, we conducted fundamental studies on the delivery of siRNA from material surfaces as a means to knock-down antagonists like noggin. Non-viral cationic lipid (Lipofectamine)-siRNA complexes were delivered from a fibrin hydrogel surface to MC3T3-E1 preosteoblasts that were treated with a supraphysiological dose of rhBMP-2 to achieve noggin mRNA expression levels higher than cells naïve to rhBMP-2. Confocal microscopy and flow cytometry showed intracellular uptake of siRNA in over 98% of MC3T3-E1 cells after 48 h. Doses of 0.5 and 1 μg noggin siRNA were able to significantly reduce noggin mRNA to levels equivalent to those in MC3T3-E1 cells not exposed to rhBMP-2 with no effects on cell viability. Small interfering RNA (siRNA) has been considered for treatment of diseases ranging from Alzheimer's to cancer. However, the ability to use siRNA in conjunction with biomaterials to direct tissue regeneration processes has received relatively little attention. Using the bone morphogenetic protein 2 antagonist, noggin, as a model, this research describes an approach to knock-down molecules that are inhibitory to desired regenerative pathways at the mRNA level via siRNA delivery from a hydrogel surface. Interactions between the material (fibrin) surface and polycation-siRNA complexes, release of the siRNA from the material surface, high levels of cellular uptake/internalization of siRNA, and significant knockdown of the targeting (noggin) mRNA are demonstrated. Broader future applications include those to nerve regeneration, cardiovascular tissue engineering, directing (stem) cell behavior, and mitigating inflammatory responses to materials. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

In vitro biocompatibility of the surface ion modified NiTi alloy

NASA Astrophysics Data System (ADS)

Gudimova, Ekaterina Yu.; Meisner, Ludmila L.; Lotkov, Aleksander I.; Matveeva, Vera A.; Meisner, Stanislav N.; Matveev, Andrey L.; Shabalina, Olga I.

2016-11-01

This paper presents the results of the chemical, topographic and structural properties of the NiTi alloy surface and their changes after surface treatments by ion implantation techniques with use of ions Ta+ and Si+. The influence of physicochemical properties of the surface ion modified NiTi alloy was studied on in vitro cultured mesenchymal stem cells of the rats' bone marrow. It is shown that the ion surface modification improves histocompatibility of the NiTi alloy and leads to increase of proliferative activity of mesenchymal stem cells on its surface. It was experimentally found that a major contribution to viability improvement mesenchymal stem cells of rat marrow has the chemical composition and the microstructure of the surface area.

Research trends in biomimetic medical materials for tissue engineering: 3D bioprinting, surface modification, nano/micro-technology and clinical aspects in tissue engineering of cartilage and bone.

PubMed

Chen, Cen; Bang, Sumi; Cho, Younghak; Lee, Sahnghoon; Lee, Inseop; Zhang, ShengMin; Noh, Insup

2016-01-01

This review discusses about biomimetic medical materials for tissue engineering of bone and cartilage, after previous scientific commentary of the invitation-based, Korea-China joint symposium on biomimetic medical materials, which was held in Seoul, Korea, from October 22 to 26, 2015. The contents of this review were evolved from the presentations of that symposium. Four topics of biomimetic medical materials were discussed from different research groups here: 1) 3D bioprinting medical materials, 2) nano/micro-technology, 3) surface modification of biomaterials for their interactions with cells and 4) clinical aspects of biomaterials for cartilage focusing on cells, scaffolds and cytokines.

Osteocytes mediate the anabolic actions of canonical Wnt/β-catenin signaling in bone

PubMed Central

Tu, Xiaolin; Delgado-Calle, Jesus; Condon, Keith W.; Maycas, Marta; Zhang, Huajia; Carlesso, Nadia; Taketo, Makoto M.; Burr, David B.; Plotkin, Lilian I.; Bellido, Teresita

2015-01-01

Osteocytes, >90% of the cells in bone, lie embedded within the mineralized matrix and coordinate osteoclast and osteoblast activity on bone surfaces by mechanisms still unclear. Bone anabolic stimuli activate Wnt signaling, and human mutations of components along this pathway underscore its crucial role in bone accrual and maintenance. However, the cell responsible for orchestrating Wnt anabolic actions has remained elusive. We show herein that activation of canonical Wnt signaling exclusively in osteocytes [dominant active (da)βcatOt mice] induces bone anabolism and triggers Notch signaling without affecting survival. These features contrast with those of mice expressing the same daß-catenin in osteoblasts, which exhibit decreased resorption and perinatal death from leukemia. daßcatOt mice exhibit increased bone mineral density in the axial and appendicular skeleton, and marked increase in bone volume in cancellous/trabecular and cortical compartments compared with littermate controls. daßcatOt mice display increased resorption and formation markers, high number of osteoclasts and osteoblasts in cancellous and cortical bone, increased bone matrix production, and markedly elevated periosteal bone formation rate. Wnt and Notch signaling target genes, osteoblast and osteocyte markers, and proosteoclastogenic and antiosteoclastogenic cytokines are elevated in bones of daßcatOt mice. Further, the increase in RANKL depends on Sost/sclerostin. Thus, activation of osteocytic β-catenin signaling increases both osteoclasts and osteoblasts, leading to bone gain, and is sufficient to activate the Notch pathway. These findings demonstrate disparate outcomes of β-catenin activation in osteocytes versus osteoblasts and identify osteocytes as central target cells of the anabolic actions of canonical Wnt/β-catenin signaling in bone. PMID:25605937

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

An image-based skeletal dosimetry model for the ICRP reference newborn—internal electron sources

NASA Astrophysics Data System (ADS)

Pafundi, Deanna; Rajon, Didier; Jokisch, Derek; Lee, Choonsik; Bolch, Wesley

2010-04-01

In this study, a comprehensive electron dosimetry model of newborn skeletal tissues is presented. The model is constructed using the University of Florida newborn hybrid phantom of Lee et al (2007 Phys. Med. Biol. 52 3309-33), the newborn skeletal tissue model of Pafundi et al (2009 Phys. Med. Biol. 54 4497-531) and the EGSnrc-based Paired Image Radiation Transport code of Shah et al (2005 J. Nucl. Med. 46 344-53). Target tissues include the active bone marrow (surrogate tissue for hematopoietic stem cells), shallow marrow (surrogate tissue for osteoprogenitor cells) and unossified cartilage (surrogate tissue for chondrocytes). Monoenergetic electron emissions are considered over the energy range 1 keV to 10 MeV for the following source tissues: active marrow, trabecular bone (surfaces and volumes), cortical bone (surfaces and volumes) and cartilage. Transport results are reported as specific absorbed fractions according to the MIRD schema and are given as skeletal-averaged values in the paper with bone-specific values reported in both tabular and graphic format as electronic annexes (supplementary data). The method utilized in this work uniquely includes (1) explicit accounting for the finite size and shape of newborn ossification centers (spongiosa regions), (2) explicit accounting for active and shallow marrow dose from electron emissions in cortical bone as well as sites of unossified cartilage, (3) proper accounting of the distribution of trabecular and cortical volumes and surfaces in the newborn skeleton when considering mineral bone sources and (4) explicit consideration of the marrow cellularity changes for active marrow self-irradiation as applicable to radionuclide therapy of diseased marrow in the newborn child.

Bone-Inspired Spatially Specific Piezoelectricity Induces Bone Regeneration

PubMed Central

Yu, Peng; Ning, Chengyun; Zhang, Yu; Tan, Guoxin; Lin, Zefeng; Liu, Shaoxiang; Wang, Xiaolan; Yang, Haoqi; Li, Kang; Yi, Xin; Zhu, Ye; Mao, Chuanbin

2017-01-01

The extracellular matrix of bone can be pictured as a material made of parallel interspersed domains of fibrous piezoelectric collagenous materials and non-piezoelectric non-collagenous materials. To mimic this feature for enhanced bone regeneration, a material made of two parallel interspersed domains, with higher and lower piezoelectricity, respectively, is constructed to form microscale piezoelectric zones (MPZs). The MPZs are produced using a versatile and effective laser-irradiation technique in which K0.5Na0.5NbO3 (KNN) ceramics are selectively irradiated to achieve microzone phase transitions. The phase structure of the laser-irradiated microzones is changed from a mixture of orthorhombic and tetragonal phases (with higher piezoelectricity) to a tetragonal dominant phase (with lower piezoelectricity). The microzoned piezoelectricity distribution results in spatially specific surface charge distribution, enabling the MPZs to bear bone-like microscale electric cues. Hence, the MPZs induce osteogenic differentiation of stem cells in vitro and bone regeneration in vivo even without being seeded with stem cells. The concept of mimicking the spatially specific piezoelectricity in bone will facilitate future research on the rational design of tissue regenerative materials. PMID:28900517

Current knowledge about the hydrophilic and nanostructured SLActive surface

PubMed Central

Wennerberg, Ann; Galli, Silvia; Albrektsson, Tomas

2011-01-01

This review summarizes the present documentation for the SLActive surface, a hydrophilic and nanostructured surface produced by Straumann Company in Switzerland, and covers the results from 15 in vitro, 17 in vivo, and 16 clinical studies. The SLActive surface is a development of the large grit-blasted and acid-etched SLA surface, and is further processed to a high degree of hydrophilicity. In general, the in vitro and in vivo studies of the SLActive surface demonstrate a stronger cell and bone tissue response than for the predecessor, the SLA surface, produced by the same company. However, in most studies, this difference disappears after 6–8 weeks. In the clinical studies, a stronger bone response was reported for the SLActive surface during the early healing phase when compared with the SLA surface. However, the later biological response was quite similar for the two surfaces and both demonstrated very good clinical results. PMID:23674916

Early detection of disease program: Evaluation of the cellular immune response

NASA Technical Reports Server (NTRS)

Criswell, B. S.; Knight, V.; Martin, R. R.; Kasel, J. A.

1975-01-01

Surfaces of normal, cultured, and mitogen-stimulated mouse lymphoid cells were examined by scanning electron microscopy (SEM). Lymphocytes with smooth, highly villous and intermediate surfaces were observed in cell suspensions from both spleens and thymuses of normal mice and from spleens of congenitally athymic (nude) mice. Several strain-specific surface features were noted, including the spine-like appearance of microvilli on C57B1/6 lymphocytes. Although thymus cell suspensions contained somewhat more smooth cells than did spleen cell preparations, lymphocyte derivation could not be inferred from SEM examination. Studies of cells stimulated with mitogenic agents for thymus-derived lymphocytes (concanavalin A) or for bone marrow-derived lymphocytes (lipopolysaccharide) suggested that, in the mouse, development of a complex villous surface is a general concomitant of lymphocyte activation and transformation.

Low temperature setting polymer-ceramic composites for bone tissue engineering

NASA Astrophysics Data System (ADS)

Sethuraman, Swaminathan

Tissue engineering is defined as "the application of biological, chemical and engineering principles towards the repair, restoration or regeneration of tissues using scaffolds, cells, factors alone or in combination". The hypothesis of this thesis is that a matrix made of a synthetic biocompatible, biodegradable composite can be designed to mimic the properties of bone, which itself is a composite. The overall goal was to design and develop biodegradable, biocompatible polymer-ceramic composites that will be a practical alternative to current bone repair materials. The first specific aim was to develop and evaluate the osteocompatibility of low temperature self setting calcium deficient apatites for bone tissue engineering. The four different calcium deficient hydroxyapatites evaluated were osteocompatible and expressed the characteristic genes for osteoblast proliferation, maturation, and differentiation. Our next objective was to develop and evaluate the osteocompatibility of biodegradable amino acid ester polyphosphazene in vitro as candidates for forming composites with low temperature apatites. We determined the structure-property relationship, the cellular adhesion, proliferation, and differentiation of primary rat osteoblast cells on two dimensional amino acid ester based polyphosphazene films. Our next goal was to evaluate the amino acid ester based polyphosphazenes in a subcutaneous rat model and our results demonstrated that the polyphosphazenes evaluated in the study were biocompatible. The physio-chemical property characterization, cellular response and gene expression on the composite surfaces were evaluated. The results demonstrated that the precursors formed calcium deficient hydroxyapatite in the presence of biodegradable polyphosphazenes. In addition, cells on the surface of the composites expressed normal phenotype and characteristic genes such as type I collagen, alkaline phosphatase, osteocalcin, osteopontin, and bone sialoprotein. The in vivo study of these novel bone cements in a 5mm unicortical defect in New Zealand white rabbits showed that the implants were osteoconductive, and osteointegrative. In conclusion, the various studies that have been carried out in this thesis to study the feasibility of a bone cement system have shown that these materials are promising candidates for various orthopaedic applications. Overall I believe that these next generation bone cements are promising bone graft substitutes in the armamentarium to treat bone defects.

Development of model hydroxyapatite bone scaffolds with multiscale porosity for potential load bearing applications

NASA Astrophysics Data System (ADS)

Dellinger, Jennifer Gwynne

2005-11-01

Model hydroxyapatite (HA) bone scaffolds consisting of a latticed pattern of rods were fabricated by a solid freeform fabrication (SFF) technique based on the robotic deposition of colloidal pastes. An optimal HA paste formulation for this method was developed. Local porosity, i.e. microporosity (1--30 mum) and sintering porosity (less than 1 mum), were produced by including polymer microsphere porogens in the HA pastes and by controlling the sintering of the scaffolds. Scaffolds with and without local porosity were evaluated with and without in vitro accelerated degradation. Percent weight loss of the scaffolds and calcium and phosphorus concentrations in solution increased with degradation time. After degradation, compressive strength and modulus decreased significantly for scaffolds with local porosity, but did not change significantly for scaffolds without local porosity. The compressive strength and modulus of scaffolds without local porosity were comparable to human cortical bone and were significantly greater than the scaffolds with local porosity. Micropores in HA disks caused surface pits that increased the surface roughness as compared to non-microporous HA disks. Mouse mesenchymal stem cells extended their cell processes into these microporous pits on HA disks in vitro. ALP expression was prolonged, cell attachment strength increased, and ECM production appeared greater on microporous HA disks compared to non-microporous HA disks and tissue culture treated polystyrene controls. Scaffolds with and without microporosity were implanted in goats bones. Microporous scaffolds with rhBMP-2 increased the percent of the scaffold filled with bone tissue compared to microporous scaffolds without rhBMP-2. Lamellar bone inside scaffolds was aligned near the rods junctions whereas lamellar bone was aligned in a more random configuration away from the rod junctions. Microporous scaffolds stained darkly with toluidine blue beneath areas of contact with new bone. This staining might indicate either extracellular matrix (ECM) in the rods or dye bound to the degrading scaffold. Although the presence of microporous topography alone did not influence bone healing in vivo, micropores were shown to provide tailorability of scaffold mechanical properties, provide a location for the storage and controlled release of a growth factor, and provide a location for bone integration inside the scaffold rods.

IGFBP-4 regulates adult skeletal growth in a sex-specific manner.

PubMed

Maridas, David E; DeMambro, Victoria E; Le, Phuong T; Nagano, Kenichi; Baron, Roland; Mohan, Subburaman; Rosen, Clifford J

2017-04-01

Insulin-like growth factor-1 (IGF-1) and its binding proteins are critical mediators of skeletal growth. Insulin-like growth factor-binding protein 4 (IGFBP-4) is highly expressed in osteoblasts and inhibits IGF-1 actions in vitro Yet, in vivo studies suggest that it could potentiate IGF-1 and IGF-2 actions. In this study, we hypothesized that IGFBP-4 might potentiate the actions of IGF-1 on the skeleton. To test this, we comprehensively studied 8- and 16-week-old Igfbp4 -/- mice. Both male and female adult Igfbp4 -/- mice had marked growth retardation with reductions in body weight, body and femur lengths, fat proportion and lean mass at 8 and 16 weeks. Marked reductions in aBMD and aBMC were observed in 16-week-old Igfbp4 -/- females, but not in males. Femoral trabecular BV/TV and thickness, cortical fraction and thickness in 16-week-old Igfbp4 -/- females were significantly reduced. However, surprisingly, males had significantly more trabeculae with higher connectivity density than controls. Concordantly, histomorphometry revealed higher bone resorption and lower bone formation in Igfbp4 -/- females. In contrast, Igfbp4 -/- males had lower mineralized surface/bone surface. Femoral expression of Sost and circulating levels of sclerostin were reduced but only in Igfbp4 -/- males. Bone marrow stromal cultures from mutants showed increased osteogenesis, whereas osteoclastogenesis was markedly increased in cells from Igfbp4 -/- females but decreased in males. In sum, our results indicate that loss of Igfbp4 affects mesenchymal stromal cell differentiation, regulates osteoclastogenesis and influences both skeletal development and adult bone maintenance. Thus, IGFBP-4 modulates the skeleton in a gender-specific manner, acting as both a cell autonomous and cell non-autonomous factor. © 2017 The authors.

Peculiarities of the bone tissue resorption under microgravity conditions

NASA Astrophysics Data System (ADS)

Rodionova, N.; Oganov, V.; Polkovenko, O.; Nitsevich, T.

The actual problem - peculiarities of resorptive processes in the spongiose of thingbones - we studied with the use of tranmissive electron microscopy in experiments on rats (American space station SLS-2) and on monkeys Macaca mulatt? (BION-11). Animals were onboard during 2 weeks. There was established, that the resorption happen with osteoclasts participation. They can create groups of cells. In the osteoclasts population we indicated not typical for the control (ground experiment) "giant" cells, which have on ultrathin sections 5-6 nuclei, many lysosomes, well developed "light" zone and "brush-border". The destruction of minera lized matrix in bone lacunas also happens by the way of osteolytic activity of osteocytes. Lysosome ferments of osteocytes are secreted by the eczocytosis. The osteocytic osteolysis, as well as the osteoclastic one can be seen as a physiological, gormon-dependent mechanism of resorption. The presence of a considerable number of neutrophiles, which enter in some zones of resorption is also typical. When these neutrophiles destruct, they release lysosomic ferments that dissolve the bone matrix. In some zones of resorption we noted the presence of the row from collagen fibrils, which loosed crystals , on mineralized matrix borders. The cell detritus is noted in zones of surface dissolving among crystallic conglomerates. It certificates the processes of osteogenic cells destruction that happen here. So, under the microgravity conditions in zones of adaptive remodeling of the spongiose the processes of the bone tissue resorption happen by some ways, namely: by the functional activization of osteoclasts; by the osteocytic osteolysis increasing; as a result of hydrolytic activity of neutrophiles, entering in these zones, and also by the local demineralization and further destruction of bone matrix surface zones.

Human reaming debris: a source of multipotent stem cells.

PubMed

Wenisch, Sabine; Trinkaus, Katja; Hild, Anne; Hose, Dirk; Herde, Katja; Heiss, Christian; Kilian, Olaf; Alt, Volker; Schnettler, Reinhard

2005-01-01

The biological characteristics of human reaming debris (HRD) generated in the course of surgical treatment of long bone diaphyseal fractures and nonunions are still a matter of dispute. Therefore, the objective of the present investigation has been to characterize the intrinsic properties of human reaming debris in vitro. Samples of reaming debris harvested from 12 patients with closed diaphyseal fractures were examined ultrastucturally and were cultured under standard conditions. After a lag phase of 4-7 days, cells started to grow out from small bone fragments and established a confluent monolayer within 20-22 days. The cells were characterized according to morphology, proliferation capacity, cell surface antigen profile, and differentiation repertoire. The results reveal that human reaming debris is a source of multipotent stem cells which are able to grow and proliferate in vitro. The cells differentiate along the osteogenic pathway after induction and can be directed toward a neuronal phenotype, as has been shown morphologically and by the expression of neuronal markers after DMSO induction. These findings have prompted interest in the use of reaming debris-derived stem cells in cell and bone replacement therapies.

Ultra-Porous Nanoparticle Networks: A Biomimetic Coating Morphology for Enhanced Cellular Response and Infiltration

PubMed Central

Nasiri, Noushin; Ceramidas, Anthony; Mukherjee, Shayanti; Panneerselvan, Anitha; Nisbet, David R.; Tricoli, Antonio

2016-01-01

Orthopedic treatments are amongst the most common cause of surgery and are responsible for a large share of global healthcare expenditures. Engineering materials that can hasten bone integration will improve the quality of life of millions of patients per year and reduce associated medical costs. Here, we present a novel hierarchical biomimetic coating that mimics the inorganic constituent of mammalian bones with the aim of improving osseointegration of metallic implants. We exploit the thermally-driven self-organization of metastable core-shell nanoparticles during their aerosol self-assembly to rapidly fabricate robust, ultra-porous nanoparticle networks (UNN) of crystalline hydroxyapatite (HAp). Comparative analysis of the response of osteoblast cells to the ultra-porous nanostructured HAp surfaces and to the spin coated HAp surfaces revealed superior osseointegrative properties of the UNN coatings with significant cell and filopodia infiltration. This flexible synthesis approach for the engineering of UNN HAp coatings on titanium implants provides a platform technology to study the bone-implant interface for improved osseointegration and osteoconduction. PMID:27076035

Lightweight Open-Cell Scaffolds from Sea Urchin Spines with Superior Material Properties for Bone Defect Repair.

PubMed

Cao, Lei; Li, Xiaokang; Zhou, Xiaoshu; Li, Yong; Vecchio, Kenneth S; Yang, Lina; Cui, Wei; Yang, Rui; Zhu, Yue; Guo, Zheng; Zhang, Xing

2017-03-22

Sea urchin spines (Heterocentrotus mammillatus), with a hierarchical open-cell structure similar to that of human trabecular bone and superior mechanical property (compressive strength ∼43.4 MPa) suitable for machining to shape, were explored for potential applications of bone defect repair. Finite element analyses reveal that the compressive stress concentrates along the dense growth rings and dissipates through strut structures of the stereoms, indicating that the exquisite mesostructures play an important role in high strength-to-weight ratios. The fracture strength of magnesium-substituted tricalcium phosphate (β-TCMP) scaffolds produced by hydrothermal conversion of urchin spines is about 9.3 MPa, comparable to that of human trabecular bone. New bone forms along outer surfaces of β-TCMP scaffolds after implantation in rabbit femoral defects for one month and grows into the majority of the inner open-cell spaces postoperation in three months, showing tight interface between the scaffold and regenerative bone tissue. Fusion of beagle lumbar facet joints using a Ti-6Al-4V cage and β-TCMP scaffold can be completed within seven months with obvious biodegradation of the β-TCMP scaffold, which is nearly completely degraded and replaced by newly formed bone ten months after implantation. Thus, sea urchin spines suitable for machining to shape have advantages for production of biodegradable artificial grafts for bone defect repair.

Synergetic topography and chemistry cues guiding osteogenic differentiation in bone marrow stromal cells through ERK1/2 and p38 MAPK signaling pathway.

PubMed

Zhang, Xinran; Li, Haotian; Lin, Chucheng; Ning, Congqin; Lin, Kaili

2018-01-30

Both the topographic surface and chemical composition modification can enhance rapid osteogenic differentiation and bone formation. Till now, the synergetic effects of topography and chemistry cues guiding biological responses have been rarely reported. Herein, the ordered micro-patterned topography and classically essential trace element of strontium (Sr) ion doping were selected to imitate topography and chemistry cues, respectively. The ordered micro-patterned topography on Sr ion-doped bioceramics was successfully duplicated using the nylon sieve as the template. Biological response results revealed that the micro-patterned topography design or Sr doping could promote cell attachment, ALP activity, and osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). Most importantly, the samples both with micro-patterned topography and Sr doping showed the highest promotion effects, and could synergistically activate the ERK1/2 and p38 MAPK signaling pathways. The results suggested that the grafts with both specific topography and chemistry cues have synergetic effects on osteogenic activity of BMSCs and provide an effective approach to design functional bone grafts and cell culture substrates.

Investigation of biomineralization by Raman spectroscopy

NASA Astrophysics Data System (ADS)

Fatscher, Robert William

Biomineralization is a process in which living organism grow composite materials consisting of inorganic and organic materials. This produces a composite material consisting of both inorganic and organic components, with superior mechanical properties. In the human body bone and dentin are both examples of biominerals. In this research Raman spectroscopy was used to characterize dentin from mice and human teeth, to determine composition. In the mouse tooth samples areas of irregular dentin were found, along the inside of the tooth, to be in the process of mineralization. By analyzing the samples along these areas we were able to determine the composition of dentin and track how it changed in these area. By analysis of the mineral to matrix ratio the areas of irregular dentin were determined to have less mineral present. Observations of other organic components and collagen in increased concentrations in this area suggested these area were in the process of biomineralization. The understanding of the structure of dentin and its biomineralization process is of crucial importance when trying reproduce dentin. Scientists and engineers are able to produce dentin minerals in vitro by culturing various dental stem cells. The ability to create dentin mineral from cells could lead to methods of repairing dentin in patients, or even lead to the creation of a completely engineered tooth. While dentin-like materials can be produced in a laboratory environment, analysis and comparison of the composition of these materials must be performed to ensure the mineral produced is consistent with dentin. Mineralized nodules from six different dental stem cell lines were cultured to produce a mineralized deposit. Utilizing Raman spectroscopy, we were able to determine cell source dependent differences in a variety of dental stem cells, and compare the mineral produced to native dentin. Orthopedic implants are implants used to replace damaged bone, examples include knee, hip and dental implants. These implants are designed to osteointegrate with the native healthy tissues in order to create a functionally stable and structural interface. Biomaterials such as hydroxyapatite and titania are known to increase the rate of bone regeneration in vivo.1 By accelerating the early response of bone forming cells to these implants, better fixation is achieved between the implant and the bone, shortening recovery times and increasing the viability of these implants. In the last part of this research an investigation of osteoblasts cultured at 14 days on five different heat-treated titania substrates was investigated by Raman spectroscopy, in order to observe the initial cellular response to the titania substrates. The heat-treatment of titania changes the amount of oxygen on it's surface which in turn effects the surface energy. A change in the surface energy of a material will affect the cellular response, by culturing cells on various heat-treated titania substrates a relationship between the surface energy and cellular response can be investigated. A faster cellular response would lead to an increased rate of bone regeneration shortening healing times and allowing for better fixation of the implant.

The effects of titanium nitride-coating on the topographic and biological features of TPS implant surfaces.

PubMed

Annunziata, Marco; Oliva, Adriana; Basile, Maria Assunta; Giordano, Michele; Mazzola, Nello; Rizzo, Antonietta; Lanza, Alessandro; Guida, Luigi

2011-11-01

Titanium nitride (TiN) coating has been proposed as an adjunctive surface treatment aimed to increase the physico-mechanical and aesthetic properties of dental implants. In this study we investigated the surface characteristics of TiN-coated titanium plasma sprayed (TiN-TPS) and uncoated titanium plasma sprayed (TPS) surfaces and their biological features towards both primary human bone marrow mesenchymal stem cells (BM-MSC) and bacterial cultures. 15 mm×1 mm TPS and TiN-TPS disks (P.H.I. s.r.l., San Vittore Olona, Milano, Italy) were topographically analysed by confocal optical profilometry. Primary human BM-MSC were obtained from healthy donors, isolated and expanded. Cells were seeded on the titanium disks and cell adhesion, proliferation, protein synthesis and osteoblastic differentiation in terms of alkaline phosphatase activity, osteocalcin synthesis and extracellular mineralization, were evaluated. Furthermore, adhesion and proliferation of Streptococcus pyogenes and Streptococcus sanguinis on both surfaces were also analysed. TiN-TPS disks showed a decreased roughness (about 50%, p < 0.05) and a decreased bacterial adhesion and proliferation compared to TPS ones. No difference (p > 0.05) in terms of BM-MSC adhesion, proliferation and osteoblastic differentiation between TPS and TiN-TPS surfaces was found. TiN coating showed to modify the topographical characteristics of TPS titanium surfaces and to significantly reduce bacterial adhesion and proliferation, although maintaining their biological affinity towards bone cell precursors. Copyright © 2011 Elsevier Ltd. All rights reserved.

In vitro bioactivity of akermanite ceramics.

PubMed

Wu, Chengtie; Chang, Jiang; Ni, Siyu; Wang, Junying

2006-01-01

In this study, the bone-like apatite-formation ability of akermanite ceramics (Ca2MgSi2O7) in simulated body fluid (SBF) and the effects of ionic products from akermanite dissolution on osteoblasts and mouse fibroblasts (cell line L929) were investigated. In addition, osteoblast morphology and proliferation on the ceramics were evaluated. The results showed that akermanite ceramics possessed bone-like apatite-formation ability comparable with bioactive wollastonite ceramics (CaSiO3) after 20 days of soaking in SBF and the mechanism of bone-like apatite formation on akermanite ceramics is similar to that of wollastonite ceramics. The Ca, Si, and Mg ions from akermanite dissolution at certain ranges of concentration significantly stimulated osteoblast and L929 cell proliferation. Furthermore, osteoblasts spread well on the surface of akermanite ceramics, and proliferated with increasing the culture time. The results showed that akermanite ceramics possess bone-like apatite-formation ability and can release soluble ionic products to stimulate cell proliferation, which indicated good bioactivity. (c) 2005 Wiley Periodicals, Inc

Fabrication and characterization of carboxymethyl cellulose novel microparticles for bone tissue engineering.

PubMed

Gaihre, Bipin; Jayasuriya, Ambalangodage C

2016-12-01

In this study we developed carboxymethyl cellulose (CMC) microparticles through ionic crosslinking with the aqueous ion complex of zirconium (Zr) and further complexing with chitosan (CS) and determined the physio-chemical and biological properties of these novel microparticles. In order to assess the role of Zr, microparticles were prepared in 5% and 10% (w/v) zirconium tetrachloride solution. Scanning electron microscopy (SEM) with energy dispersive X-ray spectrometer (EDS) results showed that Zr was uniformly distributed on the surface of the microparticles as a result of which uniform groovy surface was obtained. We found that Zr enhances the surface roughness of the microparticles and stability studies showed that it also increases the stability of microparticles in phosphate buffered saline. The crosslinking of anionic CMC with cationic Zr and CS was confirmed by Fourier transform infrared spectroscopy (FTIR) results. The response of murine pre-osteoblasts (OB-6) when cultured with microparticles was investigated. Live/dead cell assay showed that microparticles did not induce any cytotoxic effects as cells were attaching and proliferating on the well plate as well as along the surface of microparticles. In addition, SEM images showed that microparticles support the attachment of cells and they appeared to be directly interacting with the surface of microparticle. Within 10days of culture most of the top surface of microparticles was covered with a layer of cells indicating that they were proliferating well throughout the surface of microparticles. We observed that Zr enhances the cell attachment and proliferation as more cells were present on microparticles with 10% Zr. These promising results show the potential applications of CMC-Zr microparticles in bone tissue engineering. Copyright © 2016 Elsevier B.V. All rights reserved.

Lack of CD47 Impairs Bone Cell Differentiation and Results in an Osteopenic Phenotype in Vivo due to Impaired Signal Regulatory Protein α (SIRPα) Signaling*

PubMed Central

Koskinen, Cecilia; Persson, Emelie; Baldock, Paul; Stenberg, Åsa; Boström, Ingrid; Matozaki, Takashi; Oldenborg, Per-Arne; Lundberg, Pernilla

2013-01-01

Here, we investigated whether the cell surface glycoprotein CD47 was required for normal formation of osteoblasts and osteoclasts and to maintain normal bone formation activity in vitro and in vivo. In parathyroid hormone or 1α,25(OH)2-vitamin D3 (D3)-stimulated bone marrow cultures (BMC) from CD47−/− mice, we found a strongly reduced formation of multinuclear tartrate-resistant acid phosphatase (TRAP)+ osteoclasts, associated with reduced expression of osteoclastogenic genes (nfatc1, Oscar, Trap/Acp, ctr, catK, and dc-stamp). The production of M-CSF and RANKL (receptor activator of nuclear factor κβ ligand) was reduced in CD47−/− BMC, as compared with CD47+/+ BMC. The stromal cell phenotype in CD47−/− BMC involved a blunted expression of the osteoblast-associated genes osterix, Alp/Akp1, and α-1-collagen, and reduced mineral deposition, as compared with that in CD47+/+ BMC. CD47 is a ligand for SIRPα (signal regulatory protein α), which showed strongly reduced tyrosine phosphorylation in CD47−/− bone marrow stromal cells. In addition, stromal cells lacking the signaling SIRPα cytoplasmic domain also had a defect in osteogenic differentiation, and both CD47−/− and non-signaling SIRPα mutant stromal cells showed a markedly reduced ability to support osteoclastogenesis in wild-type bone marrow macrophages, demonstrating that CD47-induced SIRPα signaling is critical for stromal cell support of osteoclast formation. In vivo, femoral bones of 18- or 28-week-old CD47−/− mice showed significantly reduced osteoclast and osteoblast numbers and exhibited an osteopenic bone phenotype. In conclusion, lack of CD47 strongly impairs SIRPα-dependent osteoblast differentiation, deteriorate bone formation, and cause reduced formation of osteoclasts. PMID:23990469

Extracellular matrix production by human osteoblasts cultured on biodegradable polymers applicable for tissue engineering.

PubMed

El-Amin, S F; Lu, H H; Khan, Y; Burems, J; Mitchell, J; Tuan, R S; Laurencin, C T

2003-03-01

The nature of the extracellular matrix (ECM) is crucial in regulating cell functions via cell-matrix interactions, cytoskeletal organization, and integrin-mediated signaling. In bone, the ECM is composed of proteins such as collagen (CO), fibronectin (FN), laminin (LM), vitronectin (VN), osteopontin (OP) and osteonectin (ON). For bone tissue engineering, the ECM should also be considered in terms of its function in mediating cell adhesion to biomaterials. This study examined ECM production, cytoskeletal organization, and adhesion of primary human osteoblastic cells on biodegradable matrices applicable for tissue engineering, namely polylactic-co-glycolic acid 50:50 (PLAGA) and polylactic acid (PLA). We hypothesized that the osteocompatible, biodegradable polymer surfaces promote the production of bone-specific ECM proteins in a manner dependent on polymer composition. We first examined whether the PLAGA and PLA matrices could support human osteoblastic cell growth by measuring cell adhesion at 3, 6 and 12h post-plating. Adhesion on PLAGA was consistently higher than on PLA throughout the duration of the experiment, and comparable to tissue culture polystyrene (TCPS). ECM components, including CO, FN, LM, ON, OP and VN, produced on the surface of the polymers were quantified by ELISA and localized by immunofluorescence staining. All of these proteins were present at significantly higher levels on PLAGA compared to PLA or TCPS surfaces. On PLAGA, OP and ON were the most abundant ECM components, followed by CO, FN, VN and LN. Immunofluorescence revealed an extracellular distribution for CO and FN, whereas OP and ON were found both intracellularly as well as extracellularly on the polymer. In addition, the actin cytoskeletal network was more extensive in osteoblasts cultured on PLAGA than on PLA or TCPS. In summary, we found that osteoblasts plated on PLAGA adhered better to the substrate, produced higher levels of ECM molecules, and showed greater cytoskeletal organization than on PLA and TCPS. We propose that this difference in ECM composition is functionally related to the enhanced cell adhesion observed on PLAGA. There is initial evidence that specific composition of the PLAGA polymer favors the ECM. Future studies will seek to optimize ECM production on these matrices for bone tissue engineering applications.

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.

BMI-1 Mediates Estrogen-Deficiency-Induced Bone Loss by Inhibiting Reactive Oxygen Species Accumulation and T Cell Activation.

PubMed

Li, Jinbo; Wang, Qian; Yang, Renlei; Zhang, Jiaqi; Li, Xing; Zhou, Xichao; Miao, Dengshun

2017-05-01

Previous studies have shown that estrogen regulates bone homeostasis through regulatory effects on oxidative stress. However, it is unclear how estrogen deficiency triggers reactive oxygen species (ROS) accumulation. Recent studies provide evidence that the B lymphoma Mo-MLV insertion region 1 (BMI-1) plays a critical role in protection against oxidative stress and that this gene is directly regulated by estrogen via estrogen receptor (ER) at the transcriptional level. In this study, ovariectomized mice were given drinking water with/without antioxidant N-acetyl-cysteine (NAC, 1 mg/mL) supplementation, and compared with each other and with sham mice. Results showed that ovariectomy resulted in bone loss with increased osteoclast surface, increased ROS levels, T cell activation, and increased TNF and RANKL levels in serum and in CD4 T cells; NAC supplementation largely prevented these alterations. BMI-1 expression levels were dramatically downregulated in CD4 T cells from ovariectomized mice. We supplemented drinking water to BMI-1-deficient mice with/without NAC and compared them with each other and with wild-type (WT) mice. We found that BMI-1 deficiency mimicked alterations observed in ovariectomy whereas NAC supplementation reversed all alterations induced by BMI-1 deficiency. Because T cells are critical in mediating ovariectomy-induced bone loss, we further assessed whether BMI-1 overexpression in lymphocytes can protect against estrogen deficiency-induced osteoclastogenesis and bone loss by inhibiting oxidative stress, T cell activation, and RANKL production. When WT and Eμ-BMI-1 transgenic mice with BMI-1 specifically overexpressed in lymphocytes were ovariectomized and compared with each other and with WT sham mice, we found that BMI-1 overexpression in lymphocytes clearly reversed all alterations induced by ovariectomy. Results from this study indicate that estrogen deficiency downregulates BMI-1 and subsequently increases ROS, T cell activation, and RANKL production in T cells, thus enhancing osteoclastogenesis and accelerating bone loss. This study clarifies a novel mechanism regulating estrogen deficiency-induced bone loss. © 2016 American Society for Bone and Mineral Research. © 2016 American Society for Bone and Mineral Research.

Development of multi-walled carbon nanotubes reinforced monetite bionanocomposite cements for orthopedic applications.

PubMed

Boroujeni, Nariman Mansoori; Zhou, Huan; Luchini, Timothy J F; Bhaduri, Sarit B

2013-10-01

In this study, we present results of our research on biodegradable monetite (DCPA, CaHPO4) cement with surface-modified multi-walled carbon nanotubes (mMWCNTs) as potential bone defect repair material. The cement pastes showed desirable handling properties and possessed a suitable setting time for use in surgical setting. The incorporation of mMWCNTs shortened the setting time of DCPA and increased the compressive strength of DCPA cement from 11.09±1.85 MPa to 21.56±2.47 MPa. The cytocompatibility of the materials was investigated in vitro using the preosteoblast cell line MC3T3-E1. An increase of cell numbers was observed on both DCPA and DCPA-mMWCNTs. Scanning electron microscopy (SEM) results also revealed an obvious cell growth on the surface of the cements. Based on these results, DCPA-mMWCNTs composite cements can be considered as potential bone defect repair materials. © 2013.

Cell-based cartilage repair strategies in the horse.

PubMed

Ortved, Kyla F; Nixon, Alan J

2016-02-01

Damage to the articular cartilage surface is common in the equine athlete and, due to the poor intrinsic healing capabilities of cartilage, can lead to osteoarthritis (OA). Joint disease and OA are the leading cause of retirement in equine athletes and currently there are no effective treatments to stop the progression of OA. Several different cell-based strategies have been investigated to bolster the weak regenerative response of chondrocytes. Such techniques aim to restore the articular surface and prevent further joint degradation. Cell-based cartilage repair strategies include enhancement of endogenous repair mechanisms by recruitment of stem cells from the bone marrow following perforation of the subchondral bone plate; osteochondral implantation; implantation of chondrocytes that are maintained in defects by either a membrane cover or scaffold, and transplantation of mesenchymal stem cells into cartilage lesions. More recently, bioengineered cartilage and scaffoldless cartilage have been investigated for enhancing repair. This review article focuses on the multitude of cell-based repair techniques for cartilage repair across several species, with special attention paid to the horse. Copyright © 2015 Elsevier Ltd. All rights reserved.

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.

Zirconia toughened alumina ceramic foams for potential bone graft applications: fabrication, bioactivation, and cellular responses.

PubMed

He, X; Zhang, Y Z; Mansell, J P; Su, B

2008-07-01

Zirconia toughened alumina (ZTA) has been regarded as the next generation orthopedic graft material due to its excellent mechanical properties and biocompatibility. Porous ZTA ceramics with good interconnectivity can potentially be used as bone grafts for load-bearing applications. In this work, three-dimensional (3D) interconnected porous ZTA ceramics were fabricated using a direct foaming method with egg white protein as binder and foaming agent. The results showed that the porous ZTA ceramics possessed a bimodal pore size distribution. Their mechanical properties were comparable to those of cancellous bone. Due to the bio-inertness of alumina and zirconia ceramics, surface bioactivation of the ZTA foams was carried out in order to improve their bioactivity. A simple NaOH soaking method was employed to change the surface chemistry of ZTA through hydroxylation. Treated samples were tested by conducting osteoblast-like cell culture in vitro. Improvement on cells response was observed and the strength of porous ZTA has not been deteriorated after the NaOH treatment. The porous 'bioactivated' ZTA ceramics produced here could be potentially used as non-degradable bone grafts for load-bearing applications.

Mineralization and bone regeneration using a bioactive elastin-like recombinamer membrane.

PubMed

Tejeda-Montes, Esther; Klymov, Alexey; Nejadnik, M Reza; Alonso, Matilde; Rodriguez-Cabello, J Carlos; Walboomers, X Frank; Mata, Alvaro

2014-09-01

The search for alternative therapies to improve bone regeneration continues to be a major challenge for the medical community. Here we report on the enhanced mineralization, osteogenesis, and in vivo bone regeneration properties of a bioactive elastin-like recombinamer (ELR) membrane. Three bioactive ELRs exhibiting epitopes designed to promote mesenchymal stem cell adhesion (RGDS), mineralization (DDDEEKFLRRIGRFG), and both cell adhesion and mineralization were synthesized using standard recombinant protein techniques. The ELR materials were then used to fabricate membranes comprising either a smooth surface (Smooth) or channel microtopographies (Channels). Mineralization and osteoblastic differentiation of primary rat mesenchymal stem cells (rMSCs) were analyzed in both static and dynamic (uniaxial strain of 8% at 1 Hz frequency) conditions. Smooth mineralization membranes in static condition exhibited the highest quantity of calcium phosphate (Ca/P of 1.78) deposition with and without the presence of cells, the highest Young's modulus, and the highest production of alkaline phosphatase on day 10 in the presence of cells growing in non-osteogenic differentiation medium. These membranes were tested in a 5 mm-diameter critical-size rat calvarial defect model and analyzed for bone formation on day 36 after implantation. Animals treated with the mineralization membranes exhibited the highest bone volume within the defect as measured by micro-computed tomography and histology with no significant increase in inflammation. This study demonstrates the possibility of using bioactive ELR membranes for bone regeneration applications. Copyright © 2014 Elsevier Ltd. All rights reserved.

Micro/Nano Structural Tantalum Coating for Enhanced Osteogenic Differentiation of Human Bone Marrow Stem Cells

PubMed Central

Ding, Ding; Xie, Youtao; Li, Kai; Huang, Liping; Zheng, Xuebin

2018-01-01

Recently, tantalum has been attracting much attention for its anticorrosion resistance and biocompatibility, and it has been widely used in surface modification for implant applications. To improve its osteogenic differentiation of human bone marrow stem cells (hBMSCs), a micro/nano structure has been fabricated on the tantalum coating surface through the combination of anodic oxidation and plasma spraying method. The morphology, composition, and microstructure of the modified coating were comprehensively studied by employing scanning electron microscopy (SEM), X-ray diffraction (XRD) as well as transmission electron microscopy (TEM). The effects of hierarchical structures as well as micro-porous structure of tantalum coating on the behavior for human bone marrow stem cells (hBMSCs) were evaluated and compared at both cellular and molecular levels in vitro. The experimental results show that a hierarchical micro/nano structure with Ta2O5 nanotubes spread onto a micro-scale tantalum coating has been fabricated successfully, which is confirmed to promote cell adhesion and spreading. Besides, the hierarchical micro/nano tantalum coating can provide 1.5~2.1 times improvement in gene expression, compared with the micro-porous tantalum coating. It demonstrates that it can effectively enhance the proliferation and differentiation of hBMSCs in vitro. PMID:29614022

Micro/Nano Structural Tantalum Coating for Enhanced Osteogenic Differentiation of Human Bone Marrow Stem Cells.

PubMed

Ding, Ding; Xie, Youtao; Li, Kai; Huang, Liping; Zheng, Xuebin

2018-04-03

Recently, tantalum has been attracting much attention for its anticorrosion resistance and biocompatibility, and it has been widely used in surface modification for implant applications. To improve its osteogenic differentiation of human bone marrow stem cells (hBMSCs), a micro/nano structure has been fabricated on the tantalum coating surface through the combination of anodic oxidation and plasma spraying method. The morphology, composition, and microstructure of the modified coating were comprehensively studied by employing scanning electron microscopy (SEM), X-ray diffraction (XRD) as well as transmission electron microscopy (TEM). The effects of hierarchical structures as well as micro-porous structure of tantalum coating on the behavior for human bone marrow stem cells (hBMSCs) were evaluated and compared at both cellular and molecular levels in vitro. The experimental results show that a hierarchical micro/nano structure with Ta₂O₅ nanotubes spread onto a micro-scale tantalum coating has been fabricated successfully, which is confirmed to promote cell adhesion and spreading. Besides, the hierarchical micro/nano tantalum coating can provide 1.5~2.1 times improvement in gene expression, compared with the micro-porous tantalum coating. It demonstrates that it can effectively enhance the proliferation and differentiation of hBMSCs in vitro.

Alginate/nanohydroxyapatite scaffolds with designed core/shell structures fabricated by 3D plotting and in situ mineralization for bone tissue engineering.

PubMed

Luo, Yongxiang; Lode, Anja; Wu, Chengtie; Chang, Jiang; Gelinsky, Michael

2015-04-01

Composite scaffolds, especially polymer/hydroxyapatite (HAP) composite scaffolds with predesigned structures, are promising materials for bone tissue engineering. Various methods including direct mixing of HAP powder with polymers or incubating polymer scaffolds in simulated body fluid for preparing polymer/HAP composite scaffolds are either uncontrolled or require long times of incubation. In this work, alginate/nano-HAP composite scaffolds with designed pore parameters and core/shell structures were fabricated using 3D plotting technique and in situ mineralization under mild conditions (at room temperature and without the use of any organic solvents). Light microscopy, scanning electron microscopy, microcomputer tomography, X-ray diffraction, and Fourier transform infrared spectroscopy were applied to characterize the fabricated scaffolds. Mechanical properties and protein delivery of the scaffolds were evaluated, as well as the cell response to the scaffolds by culturing human bone-marrow-derived mesenchymal stem cells (hBMSC). The obtained data indicate that this method is suitable to fabricate alginate/nano-HAP composite scaffolds with a layer of nano-HAP, coating the surface of the alginate strands homogeneously and completely. The surface mineralization enhanced the mechanical properties and improved the cell attachment and spreading, as well as supported sustaining protein release, compared to pure alginate scaffolds without nano-HAP shell layer. The results demonstrated that the method provides an interesting option for bone tissue engineering application.

Effect of trehalose coating on basic fibroblast growth factor release from tailor-made bone implants.

PubMed

Choi, Sungjin; Lee, Jongil; Igawa, Kazuyo; Suzuki, Shigeki; Mochizuki, Manabu; Nishimura, Ryohei; Chung, Ung-il; Sasaki, Nobuo

2011-12-01

Artificial bone implants are often incorporated with osteoinductive factors to facilitate early bone regeneration. Calcium phosphate, the main component in artificial bone implants, strongly binds these factors, and in a few cases, the incorporated proteins are not released from the implant under conditions of physiological pH, thereby leading to reduction in their osteoinductivity. In this study, we coated tailor-made bone implants with trehalose to facilitate the release of basic fibroblast growth factor (bFGF). In an in vitro study, mouse osteoblastic cells were separately cultured for 48 hr in a medium with a untreated implant (T-), trehalose-coated implant (T+), bFGF-incorporated implant (FT-), and bFGF-incorporated implant with trehalose coating (FT+). In the FT+ group, cell viability was significantly higher than that in the other groups (P<0.05). Scanning electron microscopy (SEM) and X-ray diffraction (XRD) revealed that trehalose effectively covered the surface of the artificial bone implant without affecting the crystallinity or the mechanical strength of the artificial bone implant. These results suggest that coating artificial bone implants with trehalose could limit the binding of bFGF to calcium phosphate.

Polymer-ceramic nanocomposites for applications in the bone surgery

NASA Astrophysics Data System (ADS)

Stodolak, E.; Gadomska, K.; Lacz, A.; Bogun, M.

2009-01-01

The subject of this work was preparation and investigation of properties of a nanocomposite material based on polymer matrix modified with nanometric silica particles (SiO2). The composite matrix consisted of resorbable P(L/DL)LA polymer with certified biocompatibility. Nanometric silica was introduced into the matrix by means of ultrasonic homogenisation and/or mechanical stirring. The silica was introduced directly e.g. as nanoparticles or inside calcium alginate fibres which contained 3 wt.% of amorphous SiO2. Proper dispersion of nano-filliers was confirmed by means of thermal analysis (TG/DTA, DSC). It was observed, that the presence of inorganic nanoparticles influenced several surface parameters of the nanocomposites i.e. hydrophility (a decrease of surface energy) and topography (both in micro- and nano-scale). Additionally, the nanocomposites exhibited enhanced mechanical properties (Young's modulus, tensile strength) compared to the pure polymer. The nanocomposites were bioactive materials (SBF/3 days/37oC). Biological tests (MTT test) showed a good viability of human osteoblasts (hFOB 1.19) in contact with the nanocomposites surface. Results of preliminary biological tests carried out with the use of mother cells extracted from human bone marrow showed that the nanocomposites may provide differenation of bone cells.

Control of surface topography in biomimetic calcium phosphate coatings.

PubMed

Costa, Daniel O; Allo, Bedilu A; Klassen, Robert; Hutter, Jeffrey L; Dixon, S Jeffrey; Rizkalla, Amin S

2012-02-28

The behavior of cells responsible for bone formation, osseointegration, and bone bonding in vivo are governed by both the surface chemistry and topography of scaffold matrices. Bone-like apatite coatings represent a promising method to improve the osteoconductivity and bonding of synthetic scaffold materials to mineralized tissues for regenerative procedures in orthopedics and dentistry. Polycaprolactone (PCL) films were coated with calcium phosphates (CaP) by incubation in simulated body fluid (SBF). We investigated the effect of SBF ion concentration and soaking time on the surface properties of the resulting apatite coatings. CaP coatings were examined by scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transform infrared spectrometry (FTIR), and energy dispersive X-ray spectrometry (EDX). Young's modulus (E(s)) was determined by nanoindentation, and surface roughness was assessed by atomic force microscopy (AFM) and mechanical stylus profilometry. CaP such as carbonate-substituted apatite were deposited onto PCL films. SEM and AFM images of the apatite coatings revealed an increase in topographical complexity and surface roughness with increasing ion concentration of SBF solutions. Young's moduli (E(s)) of various CaP coatings were not significantly different, regardless of the CaP phase or surface roughness. Thus, SBF with high ion concentrations may be used to coat synthetic polymers with CaP layers of different surface topography and roughness to improve the osteoconductivity and bone-bonding ability of the scaffold. © 2012 American Chemical Society

Cosmos 2229 immunology study (Experiment K-8-07)

NASA Technical Reports Server (NTRS)

Sonnenfeld, Gerald

1993-01-01

The purpose of the current study was to further validate use of the rhesus monkey as a model for humans in future space flight testing. The areas of immunological importance examined in the Cosmos 2229 flight were represented by two sets of studies. The first set of studies determined the effect of space flight on the ability of bone marrow cells to respond to granulocyte/monocyte colony stimulating factor (GM-CSF). GM-CSF is an important regulator in the differentiation of bone marrow cells of both monocyte/macrophage and granulocyte lineages and any change in the ability of these cells to respond to GM-CSF can result in altered immune function. A second set of studies determined space flight effects on the expression of cell surface markers on both spleen and bone marrow cells. Immune cell markers included in this study were those for T-cell, B-cell, natural killer cell, and interleukin-2 populations. Variations from a normal cell population percentage, as represented by these markers, can be correlated with alterations in immunological function. Cells were stained with fluorescein-labelled antibodies directed against the appropriate antigens, and then analyzed using a flow cytometer.

Assessment of a new biomimetic scaffold and its effects on bone formation by OCT

NASA Astrophysics Data System (ADS)

Yang, Ying; Aydin, Halil M.; Piskin, Erhan; El Haj, Alicia J.

2009-02-01

The ultimate target of bone tissue engineering is to generate functional load bearing bone. By nature, the porous volume in the trabecular bone is occupied by osseous medulla. The natural bone matrix consists of hydroxyapatite (HA) crystals precipitated along the collagen type I fibres. The mineral phase renders bone strength while collagen provides flexibility. Without mineral component, bone is very flexible and can not bear loads, whereas it is brittle in the case of mineral phase without the collagen presence. In this study, we designed and prepared a new type of scaffold which mimics the features of natural bone. The scaffold consists of three different components, a biphasic polymeric base composed of two different biodegradable polymers prepared by using dual porogen approach and bioactive agents, i.e., collagen and HA particles which are distributed throughout the matrix only in the pore surfaces. Interaction of the bioactive scaffolds possessing very high porosity and interconnected pore structures with cells were investigated in a prolonged culture period by using an osteoblastic cell line. The mineral HA particles have a slight different refractive index from the other elements such as polymeric scaffolds and cell/matrix in a tissue engineering constructs, exhibiting brighter images in OCT. Thus, OCT renders a convenient means to assess the morphology and architecture of the blank biomimetic scaffolds. This study also takes a close observation of OCT images for the cultured cell-scaffold constructs in order to assess neo-formed minerals and matrix. The OCT assessments have been compared with the results from confocal and SEM analysis.

Biomechanical properties of 3D-printed bone scaffolds are improved by treatment with CRFP.

PubMed

Helguero, Carlos G; Mustahsan, Vamiq M; Parmar, Sunjit; Pentyala, Sahana; Pfail, John L; Kao, Imin; Komatsu, David E; Pentyala, Srinivas

2017-12-22

One of the major challenges in orthopedics is to develop implants that overcome current postoperative problems such as osteointegration, proper load bearing, and stress shielding. Current implant techniques such as allografts or endoprostheses never reach full bone integration, and the risk of fracture due to stress shielding is a major concern. To overcome this, a novel technique of reverse engineering to create artificial scaffolds was designed and tested. The purpose of the study is to create a new generation of implants that are both biocompatible and biomimetic. 3D-printed scaffolds based on physiological trabecular bone patterning were printed. MC3T3 cells were cultured on these scaffolds in osteogenic media, with and without the addition of Calcitonin Receptor Fragment Peptide (CRFP) in order to assess bone formation on the surfaces of the scaffolds. Integrity of these cell-seeded bone-coated scaffolds was tested for their mechanical strength. The results show that cellular proliferation and bone matrix formation are both supported by our 3D-printed scaffolds. The mechanical strength of the scaffolds was enhanced by trabecular patterning in the order of 20% for compression strength and 60% for compressive modulus. Furthermore, cell-seeded trabecular scaffolds modulus increased fourfold when treated with CRFP. Upon mineralization, the cell-seeded trabecular implants treated with osteo-inductive agents and pretreated with CRFP showed a significant increase in the compressive modulus. This work will lead to creating 3D structures that can be used in the replacement of not only bone segments, but entire bones.

Biological response of human bone marrow mesenchymal stem cells to fluoride-modified titanium surfaces.

PubMed

Guida, Luigi; Annunziata, Marco; Rocci, Antonio; Contaldo, Maria; Rullo, Rosario; Oliva, Adriana

2010-11-01

The aim of the present study was to examine the behaviour of human bone marrow-derived mesenchymal stem cells (BM-MSC) to fluoride-modified grit-blasted (F-TiO) titanium surfaces compared with grit-blasted ones (TiO). Implant surfaces were analysed by atomic force microscopy (AFM) and scanning electron microscopy (SEM). BM-MSC were isolated from healthy donors and grown on the implant surfaces. Cell adhesion and proliferation, type I collagen (Col I) synthesis, osteoblastic differentiation (in terms of alkaline phosphatase activity, osteocalcin synthesis and extracellular matrix mineralization) were assessed. Furthermore, the ability to affect the osteoblastic/osteoclastic balance in terms of osteoprotegerin (OPG) and activator of nuclear factor κ B ligand (RANKL) ratio was investigated. F-TiO surface showed higher S(a) values (P<0.05) and the presence of nano-scale structures at the AFM and SEM analysis. Comparable cell morphology and similar adhesion values on both surfaces were detected at early time, whereas higher proliferation values on F-TiO samples were observed at 7 and 10 days. Increased Col I and OPG levels for cells grown on F-TiO were found, whereas RANKL was not detectable in any of the conditioned media. BM-MSC showed a similar expression of early and late osteogenic markers on both TiO and F-TiO surfaces. The results of the present study show that the chemical and micro/nano-scale modifications induced by fluoride treatment of TiO-grit blasted surfaces stimulate the proliferation and the extracellular matrix synthesis by BM-MSC, as well as the increase of OPG synthesis, thus preventing osteoclast activation and differentiation. © 2010 John Wiley & Sons A/S.

Extended flow cytometry characterization of normal bone marrow progenitor cells by simultaneous detection of aldehyde dehydrogenase and early hematopoietic antigens: implication for erythroid differentiation studies

PubMed Central

Mirabelli, Peppino; Di Noto, Rosa; Lo Pardo, Catia; Morabito, Paolo; Abate, Giovanna; Gorrese, Marisa; Raia, Maddalena; Pascariello, Caterina; Scalia, Giulia; Gemei, Marica; Mariotti, Elisabetta; Del Vecchio, Luigi

2008-01-01

Background Aldehyde dehydrogenase (ALDH) is a cytosolic enzyme highly expressed in hematopoietic precursors from cord blood and granulocyte-colony stimulating factor mobilized peripheral blood, as well as in bone marrow from patients with acute myeloblastic leukemia. As regards human normal bone marrow, detailed characterization of ALDH+ cells has been addressed by one single study (Gentry et al, 2007). The goal of our work was to provide new information about the dissection of normal bone marrow progenitor cells based upon the simultaneous detection by flow cytometry of ALDH and early hematopoietic antigens, with particular attention to the expression of ALDH on erythroid precursors. To this aim, we used three kinds of approach: i) multidimensional analytical flow cytometry, detecting ALDH and early hematopoietic antigens in normal bone marrow; ii) fluorescence activated cell sorting of distinct subpopulations of progenitor cells, followed by in vitro induction of erythroid differentiation; iii) detection of ALDH+ cellular subsets in bone marrow from pure red cell aplasia patients. Results In normal bone marrow, we identified three populations of cells, namely ALDH+CD34+, ALDH-CD34+ and ALDH+CD34- (median percentages were 0.52, 0.53 and 0.57, respectively). As compared to ALDH-CD34+ cells, ALDH+CD34+ cells expressed the phenotypic profile of primitive hematopoietic progenitor cells, with brighter expression of CD117 and CD133, accompanied by lower display of CD38 and CD45RA. Of interest, ALDH+CD34- population disclosed a straightforward erythroid commitment, on the basis of three orders of evidences. First of all, ALDH+CD34- cells showed a CD71bright, CD105+, CD45- phenotype. Secondly, induction of differentiation experiments evidenced a clear-cut expression of glycophorin A (CD235a). Finally, ALDH+CD34- precursors were not detectable in patients with pure red cell aplasia (PRCA). Conclusion Our study, comparing surface antigen expression of ALDH+/CD34+, ALDH-/CD34+ and ALDH+/CD34- progenitor cell subsets in human bone marrow, clearly indicated that ALDH+CD34- cells are mainly committed towards erythropoiesis. To the best of our knowledge this finding is new and could be useful for basic studies about normal erythropoietic differentiation as well as for enabling the employment of ALDH as a red cell marker in polychromatic flow cytometry characterization of bone marrow from patients with aplastic anemia and myelodysplasia. PMID:18510759

Directed Differentiation of Human-Induced Pluripotent Stem Cells to Mesenchymal Stem Cells.

PubMed

Lian, Qizhou; Zhang, Yuelin; Liang, Xiaoting; Gao, Fei; Tse, Hung-Fat

2016-01-01

Multipotent stromal cells, also known as mesenchymal stem cells (MSCs), possess great potential to generate a wide range of cell types including endothelial cells, smooth muscle cells, bone, cartilage, and lipid cells. This protocol describes in detail how to perform highly efficient, lineage-specific differentiation of human-induced pluripotent stem cells (iPSCs) with an MSCs fate. The approach uses a clinically compliant protocol with chemically defined media, feeder-free conditions, and a CD105 positive and CD24 negative selection to achieve a single cell-based MSCs derivation from differentiating human pluripotent cells in approximately 20 days. Cells generated with this protocol express typical MSCs surface markers and undergo adipogenesis, osteogenesis, and chondrogenesis similar to adult bone marrow-derived MSCs (BM-MSCs). Nonetheless, compared with adult BM-MSCs, iPSC-MSCs display a higher proliferative capacity, up to 120 passages, without obvious loss of self-renewal potential and constitutively express MSCs surface antigens. MSCs generated with this protocol have numerous applications, including expansion to large scale cell numbers for tissue engineering and the development of cellular therapeutics. This approach has been used to rescue limb ischemia, allergic disorders, and cigarette smoke-induced lung damage and to model mesenchymal and vascular disorders of Hutchinson-Gilford progeria syndrome (HGPS).

Bone marrow mesenchymal stem cells ameliorate inflammatory factor-induced dysfunction of INS-1 cells on chip.

PubMed

Sun, Yu; Yao, Zhina; Lin, Peng; Hou, Xinguo; Chen, Li

2014-05-01

Using a microfluidic chip, we have investigated whether bone marrow mesenchymal stem cells (BM-MSCs) could ameliorate IL-1β/IFN-γ-induced dysfunction of INS-1 cells. BM-MSCs were obtained from diabetes mellitus patients and their cell surface antigen expression profiles were analyzed by flow cytometric. INS-1 cells were cocultured with BM-MSCs on a microfluidic chip with persistent perfusion of medium containing 1 ng/mL IL-1β and 2.5 U/mL IFN-γ for 72 h. BM-MSCs could partially rescue INS-1 cells from cytokine-induced dysfunction and ameliorate the expression of insulin and PDX-1 gene in INS-1 cells. Thus BM-MSCs can be viewed as a promising stem cell source to depress inflammatory factor-induced dysfunction of pancreatic β cells in diabetic patients. © 2014 International Federation for Cell Biology.

Influence of silicon doping of titanium nickelide near-surface layers on alloy cytocompatibility

NASA Astrophysics Data System (ADS)

Lotkov, A. I.; Matveev, A. L.; Artemyeva, L. V.; Meysner, S. N.; Matveeva, V. A.; Kudryashov, A. N.

2017-12-01

The cytocompatibility of titanium nickelide (TiNi) with near-surface layers doped with silicon ions was studied on mesenchymal stem cells of rat bone marrow cultivated in vitro. The cytotoxic effect of eluted components of material on the mesenchymal stem cells was determined using a RTCA iCELLigence cellular analyzer. The proliferative activity of mesenchymal stem cells cultivated in the presence or on the surfaces of titanium nickelide samples was estimated from the cell mitochondrial respiration rate in MTT tests using [2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium] tetrazolium salt. It is shown that ion plasma modification of near-surface layers of titanium nickelide with silicon improves the cytocompatibility of the alloy.

Fabrication of 13-93 bioactive glass scaffolds for bone tissue engineering using indirect selective laser sintering.

PubMed

Kolan, Krishna C R; Leu, Ming C; Hilmas, Gregory E; Brown, Roger F; Velez, Mariano

2011-06-01

Bioactive glasses are promising materials for bone scaffolds due to their ability to assist in tissue regeneration. When implanted in vivo, bioactive glasses can convert into hydroxyapatite, the main mineral constituent of human bone, and form a strong bond with the surrounding tissues, thus providing an advantage over polymer scaffold materials. Bone scaffold fabrication using additive manufacturing techniques can provide control over pore interconnectivity during fabrication of the scaffold, which helps in mimicking human trabecular bone. 13-93 glass, a third-generation bioactive material designed to accelerate the body's natural ability to heal itself, was used in the research described herein to fabricate bone scaffolds using the selective laser sintering (SLS) process. 13-93 glass mixed with stearic acid (as the polymer binder) by ball milling was used as the powder feedstock for the SLS machine. The fabricated green scaffolds underwent binder burnout to remove the stearic acid binder and were then sintered at temperatures between 675 °C and 695 °C. The sintered scaffolds had pore sizes ranging from 300 to 800 µm with 50% apparent porosity and an average compressive strength of 20.4 MPa, which is excellent for non-load bearing applications and among the highest reported for an interconnected porous scaffold fabricated with bioactive glasses using the SLS process. The MTT labeling experiment and measurements of MTT formazan formation are evidence that the rough surface of SLS scaffolds provides a cell-friendly surface capable of supporting robust cell growth.

Hierarchical bioceramic scaffolds with 3D-plotted macropores and mussel-inspired surface nanolayers for stimulating osteogenesis

NASA Astrophysics Data System (ADS)

Xu, Mengchi; Zhai, Dong; Xia, Lunguo; Li, Hong; Chen, Shiyi; Fang, Bing; Chang, Jiang; Wu, Chengtie

2016-07-01

The hierarchical structure of biomaterials plays an important role in the process of tissue reconstruction and regeneration. 3D-plotted scaffolds have been widely used for bone tissue engineering due to their controlled macropore structure and mechanical properties. However, the lack of micro- or nano-structures on the strut surface of 3D-plotted scaffolds, especially for bioceramic scaffolds, limits their biological activity. Inspired by the adhesive versatility of mussels and the active ion-chelating capacity of polydopamine, we set out to prepare a hierarchical bioceramic scaffold with controlled macropores and mussel-inspired surface nanolayers by combining the 3D-plotting technique with the polydopamine/apatite hybrid strategy in order to synergistically accelerate the osteogenesis and angiogenesis. β-Tricalcium phosphate (TCP) scaffolds were firstly 3D-plotted and then treated in dopamine-Tris/HCl and dopamine-SBF solutions to obtain TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds, respectively. It was found that polydopamine/apatite hybrid nanolayers were formed on the surface of both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds and TCP-DOPA-SBF scaffolds induced apatite mineralization for the second time during the cell culture. As compared to TCP scaffolds, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly promoted the osteogenesis of bone marrow stromal cells (BMSCs) as well as the angiogenesis of human umbilical vein endothelial cells (HUVECs), and the TCP-DOPA-SBF group presented the highest in vitro osteogenic/angiogenic activity among the three groups. Furthermore, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly improved the formation of new bone in vivo as compared to TCP scaffolds without a nanostructured surface. Our results suggest that the utilization of a mussel-inspired Ca, P-chelated polydopamine nanolayer on 3D-plotted bioceramic scaffolds is a viable and effective strategy to construct a hierarchical structure for synergistically accelerating osteogenesis.The hierarchical structure of biomaterials plays an important role in the process of tissue reconstruction and regeneration. 3D-plotted scaffolds have been widely used for bone tissue engineering due to their controlled macropore structure and mechanical properties. However, the lack of micro- or nano-structures on the strut surface of 3D-plotted scaffolds, especially for bioceramic scaffolds, limits their biological activity. Inspired by the adhesive versatility of mussels and the active ion-chelating capacity of polydopamine, we set out to prepare a hierarchical bioceramic scaffold with controlled macropores and mussel-inspired surface nanolayers by combining the 3D-plotting technique with the polydopamine/apatite hybrid strategy in order to synergistically accelerate the osteogenesis and angiogenesis. β-Tricalcium phosphate (TCP) scaffolds were firstly 3D-plotted and then treated in dopamine-Tris/HCl and dopamine-SBF solutions to obtain TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds, respectively. It was found that polydopamine/apatite hybrid nanolayers were formed on the surface of both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds and TCP-DOPA-SBF scaffolds induced apatite mineralization for the second time during the cell culture. As compared to TCP scaffolds, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly promoted the osteogenesis of bone marrow stromal cells (BMSCs) as well as the angiogenesis of human umbilical vein endothelial cells (HUVECs), and the TCP-DOPA-SBF group presented the highest in vitro osteogenic/angiogenic activity among the three groups. Furthermore, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly improved the formation of new bone in vivo as compared to TCP scaffolds without a nanostructured surface. Our results suggest that the utilization of a mussel-inspired Ca, P-chelated polydopamine nanolayer on 3D-plotted bioceramic scaffolds is a viable and effective strategy to construct a hierarchical structure for synergistically accelerating osteogenesis. Electronic supplementary information (ESI) available. See DOI: 10.1039/c6nr01952h

Disruption of the Cx43/miR21 pathway leads to osteocyte apoptosis and increased osteoclastogenesis with aging.

PubMed

Davis, Hannah M; Pacheco-Costa, Rafael; Atkinson, Emily G; Brun, Lucas R; Gortazar, Arancha R; Harris, Julia; Hiasa, Masahiro; Bolarinwa, Surajudeen A; Yoneda, Toshiyuki; Ivan, Mircea; Bruzzaniti, Angela; Bellido, Teresita; Plotkin, Lilian I

2017-06-01

Skeletal aging results in apoptosis of osteocytes, cells embedded in bone that control the generation/function of bone forming and resorbing cells. Aging also decreases connexin43 (Cx43) expression in bone; and osteocytic Cx43 deletion partially mimics the skeletal phenotype of old mice. Particularly, aging and Cx43 deletion increase osteocyte apoptosis, and osteoclast number and bone resorption on endocortical bone surfaces. We examined herein the molecular signaling events responsible for osteocyte apoptosis and osteoclast recruitment triggered by aging and Cx43 deficiency. Cx43-silenced MLO-Y4 osteocytic (Cx43 def ) cells undergo spontaneous cell death in culture through caspase-3 activation and exhibit increased levels of apoptosis-related genes, and only transfection of Cx43 constructs able to form gap junction channels reverses Cx43 def cell death. Cx43 def cells and bones from old mice exhibit reduced levels of the pro-survival microRNA miR21 and, consistently, increased levels of the miR21 target phosphatase and tensin homolog (PTEN) and reduced phosphorylated Akt, whereas PTEN inhibition reduces Cx43 def cell apoptosis. miR21 reduction is sufficient to induce apoptosis of Cx43-expressing cells and miR21 deletion in miR21 fl/fl bones increases apoptosis-related gene expression, whereas a miR21 mimic prevents Cx43 def cell apoptosis, demonstrating that miR21 lies downstream of Cx43. Cx43 def cells release more osteoclastogenic cytokines [receptor activator of NFκB ligand (RANKL)/high-mobility group box-1 (HMGB1)], and caspase-3 inhibition prevents RANKL/HMGB1 release and the increased osteoclastogenesis induced by conditioned media from Cx43 def cells, which is blocked by antagonizing HMGB1-RAGE interaction. These findings identify a novel Cx43/miR21/HMGB1/RANKL pathway involved in preventing osteocyte apoptosis that also controls osteoclast formation/recruitment and is impaired with aging. © 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Sol-Gel Derived Hydroxyapatite Coating on Mg-3Zn Alloy for Orthopedic Application

NASA Astrophysics Data System (ADS)

Singh, Sanjay; Manoj Kumar, R.; Kuntal, Kishor Kumar; Gupta, Pallavi; Das, Snehashish; Jayaganthan, R.; Roy, Partha; Lahiri, Debrupa

2015-04-01

In recent years, magnesium and its alloys have gained a lot of interest as orthopedic implant constituents because their biodegradability and mechanical properties are closer to that of human bone. However, one major concern with Mg in orthopedics is its high corrosion rate that results in the reduction of mechanical integrity before healing the bone tissue. The current study evaluates the sol-gel-derived hydroxyapatite (HA) coating on a selected Mg alloy (Mg-3Zn) for decreasing the corrosion rate and increasing the bioactivity of the Mg surface. The mechanical integrity of the coating is established as a function of the surface roughness of the substrate and the sintering temperature of the coating. Coating on a substrate roughness of 15-20 nm and sintering at 400°C shows the mechanical properties in similar range of bone, thus making it suitable to avoid the stress-shielding effect. The hydroxyapatite coating on the Mg alloy surface also increases corrosion resistance very significantly by 40 times. Bone cells are also found proliferating better in the HA-coated surface. All these benefits together establish the candidature of sol-gel HA-coated Mg-3Zn alloy in orthopedic application.

Increased bone density in mice lacking the proton receptor, OGR1

PubMed Central

Krieger, Nancy S.; Yao, Zhenqiang; Kyker-Snowman, Kelly; Kim, Min Ho; Boyce, Brendan F.; Bushinsky, David A.

2016-01-01

Chronic metabolic acidosis stimulates cell-mediated calcium efflux from bone through osteoblastic prostaglandin E2-induced stimulation of RANKL leading to increased osteoclastic bone resorption. Osteoblasts express the proton-sensing G-protein coupled receptor, OGR1, which activates IP3-mediated intracellular calcium. Proton-induced osteoblastic intracellular calcium signaling requires OGR1, suggesting OGR1 is the sensor activated during acidosis to cause bone resorption. Growing mice produce large amounts of metabolic acids which must be buffered, primarily by bone, prior to excretion by the kidney. Here we tested whether lack of OGR1 inhibits proton-induced bone resorption by measuring bone mineral density by μCT and histomorphometry in 8 week old male OGR1−/− and C57/Bl6 wild type mice. OGR1−/− mice have normal skeletal development with no atypical gross phenotype. Trabecular and cortical bone volume was increased in tibiae and vertebrae from OGR1−/−. There were increased osteoblast numbers on the cortical and trabecular surfaces of tibiae from OGR1−/− mice, increased endocortical and trabecular bone formation rates, and osteoblastic gene expression. Osteoclast numbers and surface were increased in tibiae of OGR1−/− mice. Thus, in rapidly growing mice, lack of OGR1 leads to increased bone mass with increased bone turnover and a greater increase in bone formation than resorption. This supports the important role of the proton receptor, OGR1, in the response of bone to protons. PMID:26880453

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.

Distinctive Capillary Action by Micro-channels in Bone-like Templates can Enhance Recruitment of Cells for Restoration of Large Bony Defect.

PubMed

Oh, Daniel S; Koch, Alia; Eisig, Sidney; Kim, Sahng Gyoon; Kim, Yoon Hyuk; Kim, Do-Gyoon; Shim, Jae Hyuck

2015-09-11

Without an active, thriving cell population that is well-distributed and stably anchored to the inserted template, exceptional bone regeneration does not occur. With conventional templates, the absence of internal micro-channels results in the lack of cell infiltration, distribution, and inhabitance deep inside the templates. Hence, a highly porous and uniformly interconnected trabecular-bone-like template with micro-channels (biogenic microenvironment template; BMT) has been developed to address these obstacles. The novel BMT was created by innovative concepts (capillary action) and fabricated with a sponge-template coating technique. The BMT consists of several structural components: inter-connected primary-pores (300-400 µm) that mimic pores in trabecular bone, micro-channels (25-70 µm) within each trabecula, and nanopores (100-400 nm) on the surface to allow cells to anchor. Moreover, the BMT has been documented by mechanical test study to have similar mechanical strength properties to those of human trabecular bone (~3.8 MPa)12. The BMT exhibited high absorption, retention, and habitation of cells throughout the bridge-shaped (Π) templates (3 cm height and 4 cm length). The cells that were initially seeded into one end of the templates immediately mobilized to the other end (10 cm distance) by capillary action of the BMT on the cell media. After 4 hr, the cells homogenously occupied the entire BMT and exhibited normal cellular behavior. The capillary action accounted for the infiltration of the cells suspended in the media and the distribution (active migration) throughout the BMT. Having observed these capabilities of the BMT, we project that BMTs will absorb bone marrow cells, growth factors, and nutrients from the periphery under physiological conditions. The BMT may resolve current limitations via rapid infiltration, homogenous distribution and inhabitance of cells in large, volumetric templates to repair massive skeletal defects.

In vitro characterizations of mesoporous hydroxyapatite as a controlled release delivery device for VEGF in orthopedic applications.

PubMed

Poh, Chye Khoon; Ng, Suxiu; Lim, Tee Yong; Tan, Hark Chuan; Loo, Joachim; Wang, Wilson

2012-11-01

Following bone implant surgery, prolonged ischemic conditions at the implant site often result in postsurgical complications like failure of osseointegration at the bone-implant interface which can lead to implant failure. Thus, restoration of the vascular supply is paramount to the proper development of the bone. High surface area mesostructured materials have been shown to be attractive candidates for bone regeneration to enhance cell adhesion and cell proliferation. This study uses hydroxyapatite, a naturally occurring mineral in the bone, fabricated to a range of suitable pore sizes, infused with vascular endothelial growth factor (VEGF), to be progressively released to stimulate revascularization. In this study, several characterizations including nitrogen adsorption analysis, Fourier-transformed infrared spectroscopy, X-ray diffraction, field emission scanning electron microscope, and transmission electron microscope were used to evaluate the synthesized mesoporous hydroxyapatite (MHA). The results showed that MHA can gradually release VEGF for enhancing revascularization, which is beneficial for orthopedic applications. Copyright © 2012 Wiley Periodicals, Inc.

[Comperative study of implant surface characteristics].

PubMed

Katona, Bernadett; Daróczi, Lajos; Jenei, Attila; Bakó, József; Hegedus, Csaba

2013-12-01

The osseointegration between the implant and its' bone environment is very important. The implants shall meet the following requirements: biocompatibility, rigidity, resistance against corrosion and technical producibility. In our present study surface morphology and material characteristics of different implants (Denti Bone Level, Denti Zirconium C, Bionika CorticaL, Straumann SLA, Straumann SLA Active, Dentsply Ankylos and Biotech Kontact implant) were investigated with scanning electron microscopy and energy-dispersive X-ray spectroscopy. The possible surface alterations caused by the manufacturing technology were also investigated. During grit-blasting the implants' surface is blasted with hard ceramic particles (titanium oxide, alumina, calcium phosphate). Properties of blasting material are critical because the osseointegration of dental implants should not be hampered. The physical and chemical features of blasting particles could importantly affect the produced surfaces of implants. Titanium surfaces with micro pits are created after immersion in mixtures of strong acids. On surfaces after dual acid-etching procedures the crosslinking between fibrin and osteogenetic cells could be enhanced therefore bone formation could be directly facilitated on the surface of the implant. Nowadays there are a number of surface modification techniques available. These can be used as a single method or in combination with each other. The effect of the two most commonly used surface modifications (acid-etching and grit-blasting) on different implants are demonstrated in our investigation.

Bioactivation of calcium deficient hydroxyapatite with foamed gelatin gel. A new injectable self-setting bone analogue.

PubMed

Dessì, M; Alvarez-Perez, M A; De Santis, R; Ginebra, M P; Planell, J A; Ambrosio, L

2014-02-01

An alternative approach to bone repair for less invasive surgical techniques, involves the development of biomaterials directly injectable into the injury sites and able to replicate a spatially organized platform with features of bone tissue. Here, the preparation and characterization of an innovative injectable bone analogue made of calcium deficient hydroxyapatite and foamed gelatin is presented. The biopolymer features and the cement self-setting reaction were investigated by rheological analysis. The porous architecture, the evolution of surface morphology and the grains dimension were analyzed with electron microscopy (SEM/ESEM/TEM). The physico-chemical properties were characterized by X-ray diffraction and FTIR analysis. Moreover, an injection test was carried out to prove the positive effect of gelatin on the flow ensuing that cement is fully injectable. The cement mechanical properties are adequate to function as temporary substrate for bone tissue regeneration. Furthermore, MG63 cells and bone marrow-derived human mesenchymal stem cells (hMSCs) were able to migrate and proliferate inside the pores, and hMSCs differentiated to the osteoblastic phenotype. The results are paving the way for an injectable bone substitute with properties that mimic natural bone tissue allowing the successful use as bone filler for craniofacial and orthopedic reconstructions in regenerative medicine.

Mesoporous bioactive glasses: structure characteristics, drug/growth factor delivery and bone regeneration application

PubMed Central

Wu, Chengtie; Chang, Jiang

2012-01-01

The impact of bone diseases and trauma in the whole world has increased significantly in the past decades. Bioactive glasses are regarded as an important bone regeneration material owing to their generally excellent osteoconductivity and osteostimulativity. A new class of bioactive glass, referred to as mesoporous bioglass (MBG), was developed 7 years ago, which possess a highly ordered mesoporous channel structure and a highly specific surface area. The study of MBG for drug/growth factor delivery and bone tissue engineering has grown significantly in the past several years. In this article, we review the recent advances of MBG materials, including the preparation of different forms of MBG, composition–structure relationship, efficient drug/growth factor delivery and bone tissue engineering application. By summarizing our recent research, the interaction of MBG scaffolds with bone-forming cells, the effect of drug/growth factor delivery on proliferation and differentiation of tissue cells and the in vivo osteogenesis of MBG scaffolds are highlighted. The advantages and limitations of MBG for drug delivery and bone tissue engineering have been compared with microsize bioactive glasses and nanosize bioactive glasses. The future perspective of MBG is discussed for bone regeneration application by combining drug delivery with bone tissue engineering and investigating the in vivo osteogenesis mechanism in large animal models. PMID:23741607

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.

Deficiency of bone marrow beta3-integrin enhances non-functional neovascularization.

PubMed

Watson, Alan R; Pitchford, Simon C; Reynolds, Louise E; Direkze, Natalie; Brittan, Mairi; Alison, Malcolm R; Rankin, Sara; Wright, Nicholas A; Hodivala-Dilke, Kairbaan M

2010-03-01

beta3-Integrin is a cell surface adhesion and signalling molecule important in the regulation of tumour angiogenesis. Mice with a global deficiency in beta3-integrin show increased pathological angiogenesis, most likely due to increased vascular endothelial growth factor receptor 2 expression on beta3-null endothelial cells. Here we transplanted beta3-null bone marrow (BM) into wild-type (WT) mice to dissect the role of BM beta3-integrin deficiency in pathological angiogenesis. Mice transplanted with beta3-null bone marrow show significantly enhanced angiogenesis in subcutaneous B16F0 melanoma and Lewis lung carcinoma (LLC) cell models and in B16F0 melanoma lung metastasis when compared with tumours grown in mice transplanted with WT bone marrow. The effect of bone marrow beta3-integrin deficiency was also assessed in the RIPTAg mouse model of pancreatic tumour growth. Again, angiogenesis in mice lacking BM beta3-integrin was enhanced. However, tumour weight between the groups was not significantly altered, suggesting that the enhanced blood vessel density in the mice transplanted with beta3-null bone marrow was not functional. Indeed, we demonstrate that in mice transplanted with beta3-null bone marrow a significant proportion of tumour blood vessels are non-functional when compared with tumour blood vessels in WT-transplanted controls. Furthermore, beta3-null-transplanted mice showed an increased angiogenic response to VEGF in vivo when compared with WT-transplanted animals. BM beta3-integrin deficiency affects the mobilization of progenitor cells to the peripheral circulation. We show that VEGF-induced mobilization of endothelial progenitor cells is enhanced in mice transplanted with beta3-null bone marrow when compared with WT-transplanted controls, suggesting a possible mechanism underlying the increased blood vessel density seen in beta3-null-transplanted mice. In conclusion, although BM beta3-integrin is not required for pathological angiogenesis, our studies demonstrate a role for BM beta3-integrin in VEGF-induced mobilization of bone marrow-derived cells to the peripheral circulation and for the functionality of those vessels in which BM-derived cells become incorporated.

Change in Mouse Bone Turnover in Response to Microgravity on RR-1

NASA Technical Reports Server (NTRS)

Cheng-Campbell, Margareth A.; Blaber, Elizabeth A.; Almeida, Eduardo A. C.

2016-01-01

Mechanical unloading during spaceflight is known to adversely affect mammalian physiology. Our previous studies using the Animal Enclosure Module on short duration Shuttle missions enabled us to identify a deficit in stem cell based-tissue regeneration as being a significant concern for long-duration spaceflight. Specifically, we found that mechanical unloading in microgravity resulted in inhibition of differentiation of mesenchymal and hematopoietic stem cells in the bone marrow compartment. Also, we observed overexpression of a cell cycle arrest molecule, CDKN1ap21, in osteoprecursor cells on the bone surface, chondroprogenitors in the articular cartilage, and in myofibers attached to bone tissue. Specifically in bone tissue during both short (15-day) and long (30-day) microgravity experiments, we observed significant loss of bone tissue and structure in both the pelvis and the femur. After 15-days of microgravity on STS-131, pelvic ischium displayed a 6.23 decrease in bone fraction (p0.005) and 11.91 decrease in bone thickness (p0.002). Furthermore, during long-duration spaceflight we observed onset of an accelerated aging-like phenotype and osteoarthritic disease state indicating that stem cells within the bone tissue fail to repair and regenerate tissues in a normal manner, leading to drastic tissue alterations in response to microgravity. The Rodent Research Hardware System provides the capability to investigate these effects during long-duration experiments on the International Space Station. During the Rodent Research-1 mission 10 16-week-old female C57Bl6J mice were exposed to 37-days of microgravity. All flight animals were euthanized and frozen on orbit for future dissection. Ground (n10) and vivarium controls (n10) were housed and processed to match the flight animal timeline. During this study we collected pelvis, femur, and tibia from all animal groups to test the hypothesis that stem cell-based tissue regeneration is significantly altered after 37-days of spaceflight. To do this, we will analyze differences in bone morphometric parameters using MicroCT. The pelvis, femur, and tibia are key in supporting and distributing weight under normal conditions. Therefore, we expect to see altered remodeling in flight animals in response to microgravity with respect to ground controls. In combination with histomorphometry, these results will help elucidate the complex mechanisms underlying bone tissue maintenance and stem cell regeneration.

Changes in Mouse Bone Turnover in Response to Microgravity

NASA Technical Reports Server (NTRS)

Cheng-Campbell, M.; Blaber, E.; Almeida, E.

2016-01-01

Mechanical unloading during spaceflight is known to adversely affect mammalian physiology. Our previous studies using the Animal Enclosure Module on short duration Shuttle missions enabled us to identify a deficit in stem cell based-tissue regeneration as being a significant concern for long-duration spaceflight. Specifically, we found that mechanical unloading in microgravity resulted in inhibition of differentiation of mesenchymal and hematopoietic stem cells in the bone marrow compartment. Also, we observed overexpression of a cell cycle arrest molecule, CDKN1a/p21, in osteoprecursor cells on the bone surface, chondroprogenitors in the articular cartilage, and in myofibers attached to bone tissue. Specifically in bone tissue during both short (15-day) and long (30-day) microgravity experiments, we observed significant loss of bone tissue and structure in both the pelvis and the femur. After 15-days of microgravity on STS-131, pelvic ischium displayed a 6.23% decrease in bone fraction (p=0.005) and 11.91% decrease in bone thickness (p=0.002). Furthermore, during long-duration spaceflight we observed onset of an accelerated aging-like phenotype and osteoarthritic disease state indicating that stem cells within the bone tissue fail to repair and regenerate tissues in a normal manner, leading to drastic tissue alterations in response to microgravity. The Rodent Research Hardware System provides the capability to investigate these effects during long-duration experiments on the International Space Station. During the Rodent Research-1 mission 10 16-week-old female C57Bl/6J mice were exposed to 37-days of microgravity. All flight animals were euthanized and frozen on orbit for future dissection. Ground (n=10) and vivarium controls (n=10) were housed and processed to match the flight animal timeline. During this study we collected pelvis, femur, and tibia from all animal groups to test the hypothesis that stem cell-based tissue regeneration is significantly altered after 37-days of spaceflight. To do this, we will analyze differences in bone morphometric parameters using MicroCT. The pelvis, femur, and tibia are key in supporting and distributing weight under normal conditions. Therefore, we expect to see altered remodeling in flight animals in response to microgravity with respect to ground controls. In combination with histomorphometry, these results will help elucidate the complex mechanisms underlying bone tissue maintenance and stem cell regeneration.

Bone substitute material composition and morphology differentially modulate calcium and phosphate release through osteoclast-like cells.

PubMed

Konermann, A; Staubwasser, M; Dirk, C; Keilig, L; Bourauel, C; Götz, W; Jäger, A; Reichert, C

2014-04-01

The aim of this study was to determine the material composition and cell-mediated remodelling of different calcium phosphate-based bone substitutes. Osteoclasts were cultivated on bone substitutes (Cerabone, Maxresorb, and NanoBone) for up to 5 days. Bafilomycin A1 addition served as the control. To determine cellular activity, the supernatant content of calcium and phosphate was measured by inductively coupled plasma optical emission spectrometry. Cells were visualized on the materials by scanning electron microscopy. Material composition and surface characteristics were assessed by energy-dispersive X-ray spectroscopy. Osteoclast-induced calcium and phosphate release was material-specific. Maxresorb exhibited the highest ion release to the medium (P = 0.034; calcium 40.25mg/l day 5, phosphate 102.08 mg/l day 5) and NanoBone the lowest (P = 0.021; calcium 8.43 mg/l day 5, phosphate 15.15 mg/l day 5); Cerabone was intermediate (P = 0.034; calcium 16.34 mg/l day 5, phosphate 30.6 mg/l day 5). All investigated materials showed unique resorption behaviours. The presented methodology provides a new perspective on the investigation of bone substitute biodegradation, maintaining the material-specific micro- and macrostructure. Copyright © 2013 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

Surface grafting of a thermoplastic polyurethane with methacrylic acid by previous plasma surface activation and by ultraviolet irradiation to reduce cell adhesion.

PubMed

Alves, P; Pinto, S; Kaiser, Jean-Pierre; Bruinink, Arie; de Sousa, Hermínio C; Gil, M H

2011-02-01

The material performance, in a biological environment, is mainly mediated by its surface properties and by the combination of chemical, physical, biological, and mechanical properties required, for a specific application. In this study, the surface of a thermoplastic polyurethane (TPU) material (Elastollan(®)1180A50) was activated either by plasma or by ultra-violet (UV) irradiation. After surface activation, methacrylic acid (MAA) was linked to the surface of TPU in order to improve its reactivity and to reduce cell adhesion. Grafted surfaces were evaluated by X-ray photoelectron spectroscopy (XPS), by atomic force microscopy (AFM) and by contact angle measurements. Blood compatibility studies and cell adhesion tests with human bone marrow cells (HBMC) were also performed. If was found that UV grafting method led to better results than the plasma activation method, since cell adhesion was reduced when methacrylic acid was grafted to the TPU surface by UV. Copyright © 2010 Elsevier B.V. All rights reserved.

Investigation of early cell–surface interactions of human mesenchymal stem cells on nanopatterned β-type titanium–niobium alloy surfaces

PubMed Central

Medda, Rebecca; Helth, Arne; Herre, Patrick; Pohl, Darius; Rellinghaus, Bernd; Perschmann, Nadine; Neubauer, Stefanie; Kessler, Horst; Oswald, Steffen; Eckert, Jürgen; Spatz, Joachim P.; Gebert, Annett; Cavalcanti-Adam, Elisabetta A.

2014-01-01

Multi-potent adult mesenchymal stem cells (MSCs) derived from bone marrow have therapeutic potential for bone diseases and regenerative medicine. However, an intrinsic heterogeneity in their phenotype, which in turn results in various differentiation potentials, makes it difficult to predict the response of these cells. The aim of this study is to investigate initial cell–surface interactions of human MSCs on modified titanium alloys. Gold nanoparticles deposited on β-type Ti–40Nb alloys by block copolymer micelle nanolithography served as nanotopographical cues as well as specific binding sites for the immobilization of thiolated peptides present in several extracellular matrix proteins. MSC heterogeneity persists on polished and nanopatterned Ti–40Nb samples. However, cell heterogeneity and donor variability decreased upon functionalization of the gold nanoparticles with cyclic RGD peptides. In particular, the number of large cells significantly decreased after 24 h owing to the arrangement of cell anchorage sites, rather than peptide specificity. However, the size and number of integrin-mediated adhesion clusters increased in the presence of the integrin-binding peptide (cRGDfK) compared with the control peptide (cRADfK). These results suggest that the use of integrin ligands in defined patterns could improve MSC-material interactions, not only by regulating cell adhesion locally, but also by reducing population heterogeneity. PMID:24501674

In vitro and in vivo characterization of anodised zirconium as a potential material for biomedical applications.

PubMed

Katunar, Maria R; Gomez Sanchez, Andrea; Santos Coquillat, Ana; Civantos, Ana; Martinez Campos, Enrique; Ballarre, Josefina; Vico, Tamara; Baca, Matias; Ramos, Viviana; Cere, Silvia

2017-06-01

In vitro studies offer the insights for the understanding of the mechanisms at the tissue-implant interface that will provide an effective functioning in vivo. The good biocompatibility of zirconium makes a good candidate for biomedical applications and the attractive in vivo performance is mainly due to the presence of a protective oxide layer. The aim of this study is to evaluate by in vitro and in vivo approach, the influence of surface modification achieved by anodisation at 30 and 60V on zirconium implants on the first steps of the osseointegration process. In this study cell attachment, proliferation and morphology of mouse myoblast C2C12-GFP and in mouse osteoprogenitor MC3T3-E1 cells was evaluated. Also, together with the immune system response, osteoclast differentiation and morphology with RAW 264.7 murine cell line were analysed. It was found that anodisation treatment at 60V enhanced cell spreading and the osteoblastic and osteoclastic cells morphology, showing a strong dependence on the surface characteristics. In vivo tests were performed in a rat femur osteotomy model. Dynamical and static histological and histomorphometric analyses were developed 15 and 30days after surgery. Newly formed bone around Zr60V implants showed a continuous newly compact and homogeneous bone just 15 after surgery, as judged by the enhanced thickness and mineralization rate. The results indicate that anodising treatment at 60V could be an effective improvement in the osseointegration of zirconium by stimulating adhesion, proliferation, morphology, new bone thickness and bone mineral apposition, making zirconium an emerging candidate material for biomedical applications. Copyright © 2017 Elsevier B.V. All rights reserved.

The effects of simulated hypogravity on murine bone marrow cells

NASA Technical Reports Server (NTRS)

Lawless, Desales

1989-01-01

Mouse bone marrow cells grown in complete medium at unit gravity were compared with a similar population cultured in conditions that mimic some aspects of microgravity. After the cells adjusted to the conditions that simulated microgravity, they proliferated as fetal or oncogenic populations; their numbers doubled in twelve hour periods. Differentiated subpopulations were depleted from the heterogeneous mixture with time and the undifferentiated hematopoietic stem cells increased in numbers. The cells in the control groups in unit gravity and those in the bioreactors in conditions of microgravity were monitored under a number of parameters. Each were phenotyped as to cell surface antigens using a panel of monoclonal antibodies and flow cytometry. Other parameters compared included: pH, glucose uptake, oxygen consumption and carbon-dioxide production. Nuclear DNA was monitored by flow cytometry. Functional responses were studied by mitogenic stimulation by various lectins. The importance of these findings should have relevance to the space program. Cells should behave predictably in zero gravity; specific populations can be eliminated from diverse populations and other populations isolated. The availability of stem cell populations will enhance both bone marrow and gene transplant programs. Stem cells will permit developmental biologists study the paths of hematopoiesis.

Osteoblast response to magnesium ion-incorporated nanoporous titanium oxide surfaces.

PubMed

Park, Jin-Woo; Kim, Youn-Jeong; Jang, Je-Hee; Song, Hwangjun

2010-11-01

This study investigated the surface characteristics and in vitro osteoconductivity of a titanium (Ti) surface incorporated with the magnesium ions (Mg) produced by hydrothermal treatment for future application as an endosseous implant surface. Mg-incorporated Ti oxide surfaces were produced by hydrothermal treatment using Mg-containing solution on two different microstructured surfaces--abraded minimally rough (Ma) or grit-blasted moderately rough (RBM) samples. The surface characteristics were evaluated using scanning electron microscopy, thin-film X-ray diffractometry, X-ray photoelectron spectroscopy, optical profilometry, and inductively coupled plasma atomic emission spectroscopy (ICP-AES). MC3T3-E1 pre-osteoblast cell attachment, proliferation, alkaline phosphatase (ALP) activity, and quantitative analysis of osteoblastic gene expression on Ma, RBM, Mg-incorporated Ma (Mg), and Mg-incorporated grit-blasted (RBM/Mg) Ti surfaces were evaluated. Hydrothermal treatment produced an Mg-incorporated Ti oxide layer with nanoporous surface structures. Mg-incorporated surfaces showed surface morphologies and surface roughness values almost identical to those of untreated smooth or micro-rough surfaces at the micron scale. ICP-AES analysis showed Mg ions released from treated surfaces into the solution. Mg incorporation significantly increased cellular attachment (P=0 at 0.5 h, P=0.01 at 1 h) on smooth surfaces, but no differences were found on micro-rough surfaces. Mg incorporation further increased ALP activity in cells grown on both smooth and micro-rough surfaces at 7 and 14 days of culture (P=0). Real-time polymerase chain reaction analysis showed higher mRNA expressions of the osteoblast transcription factor gene (Dlx5), various integrins, and the osteoblast phenotype genes (ALP, bone sialoprotein and osteocalcin) in cells grown on micro-rough (RBM) and Mg-incorporated (Mg and RBM/Mg) surfaces than those on Ma surfaces. Mg incorporation further increased the mRNA expressions of key osteoblast genes and integrins (α1, α2, α5, and β1) in cells grown on both the smooth and the micro-rough surfaces. These results indicate that an Mg-incorporated nanoporous Ti oxide surface produced by hydrothermal treatment may improve implant bone healing by enhancing the attachment and differentiation of osteoblastic cells. © 2010 John Wiley & Sons A/S.

Zoledronate promotes bone formation by blocking osteocyte-osteoblast communication during bone defect healing.

PubMed

Cui, Pingping; Liu, Hongrui; Sun, Jing; Amizuka, Norio; Sun, Qinfeng; Li, Minqi

2018-01-01

Nitrogen-containing bisphosphonates (N-BPs) are potent antiresorptive drugs and their actions on osteoclasts have been studied extensively. Recent studies have suggested that N-BPs also target bone-forming cells. However, the precise mechanism of N-BPs in osteoblasts is paradoxical, and the specific role of osteocytes is worthy of in-depth study. Here, we investigated the cellular mechanisms of N-BPs regulating bone defect healing by zoledronate (ZA). Bone histomorphometry confirmed an increase in new bone formation by systemic ZA administration. ZA induced more alkaline phosphatase-positive osteoblasts and tartrate-resistant acid phosphatase-positive osteoclasts residing on the bone surface. Inexplicably, ZA increased SOST expression in osteocytes embedded in the bone matrix, which was not compatible with the intense osteoblast activity on the bone surface. ZA induced heterogeneous osteocytes and disturbed the distribution of the osteocytic-canalicular system (OLCS). Furthermore, according to the degree of OLCS regularity, dentin matrix protein 1 reactivity had accumulated around osteocytes in the ZA group, but it was distributed evenly in the OLCS of the control group. The control group showed a dense array of the gap junction protein connexin 43. However, connexin 43 was extremely sparse after ZA administration. In summary, ZA treatment reduces gap junction connections and blocks cellular communication between osteocytes and osteoblasts. Retaining SOST expression in osteocytes leads to activation of the Wnt signaling pathway and subsequent bone formation.

Cell therapy with bone marrow mononuclear cells in elastase-induced pulmonary emphysema.

PubMed

Longhini-Dos-Santos, Nathalia; Barbosa-de-Oliveira, Valter Abraão; Kozma, Rodrigo Heras; Faria, Carolina Arruda de; Stessuk, Talita; Frei, Fernando; Ribeiro-Paes, João Tadeu

2013-04-01

Emphysema is characterized by destruction of alveolar walls with loss of gas exchange surface and consequent progressive dyspnea. This study aimed to evaluate the efficiency of cell therapy with bone marrow mononuclear cells (BMMC) in an animal model of elastase-induced pulmonary emphysema. Emphysema was induced in C57Bl/J6 female mice by intranasal instillation of elastase. After 21 days, the mice received bone marrow mononuclear cells from EGFP male mice with C57Bl/J6 background. The groups were assessed by comparison and statistically significant differences (p < 0.05) were observed among the groups treated with BMMC and evaluated after 7, 14 and 21 days. Analysis of the mean linear intercept (Lm) values for the different groups allowed to observe that the group treated with BMMC and evaluated after 21 days showed the most significant result. The group that received no treatment showed a statistically significant difference when compared to other groups, except the group treated and evaluated after 21 days, evidencing the efficacy of cell therapy with BMMC in pulmonary emphysema.

Effects of low level laser therapy (LLLT) on pressured human osteoblasts: A histomorphologic and quantitative study

NASA Astrophysics Data System (ADS)

Pyo, S. J.; Song, W. W.; Kim, I. R.; Park, B. S.; Kim, C. H.; Kim, S. S.; Chung, I. K.; Kim, Y. D.

2012-03-01

Previous research has investigated the effects of LLLT during titanium implantation, tooth movement and bone graft using deproteinized bovine bone and recognized that these circumstances were nothing more than intentional controlled overpressure against static cells since this controlled trauma could affect cell function/malfunction, or cell recovery/apoptosis. The present preliminary study was conducted to prove if LLL would influence cell viability and cell function after excessive damage, which is enough to diminish cell numbers and distort the features of cells. Our aim is to evaluate whether low level laser irradiation (LLLi) could be helpful in the recovery of traumatized osteoblasts (pressure damaged cells) by observing the morphology and the survival rate of those cells. This model used bone cell cultures which were traumatized by a pressure with 250 G of centripetal force and observed their response to such trauma and low level laser irradiation. In this experiment, a Ga-Al-As diode LLL (IMPRA-ORT, NDLux, Seoul, KOREA) was used with a wavelength of 808 nm, a focus of 14 × 24 mm, which was wide enough to cover the whole dish surface or well within at least 2 times radiation, and an output of 100 mW. Statistical analysis showed a higher recovery rate of damaged osteoblasts in the radiation group than the non-radiation group ( p < 0.05). The nonradiation group had a very poor proliferation rate in comparison to the control group ( p < 0.05) in every time period. In the control group, actin filaments showed a random orientation and cell process branched variously around each cell. In contrast, compressed cells, these patterns were turned into thicker and shorter cytoskeletons. As time progressed, every living cell recovered from the severe stress and recovered both form and function. In summary, the present study showed the capacity of LLLT to aid the recovery of the cell skeleton and affect cell viability on overpressured osteoblasts. These results may contribute toward a better understanding of the effect of LLLT on the recovery of cells after trauma. In addition, our results demonstrated that LLLT could be used in the field of bone tissue engineering to traumatized bone conditions and repair large bone defects such as bone graft and implant installation.

Impaired CXCR4 Expression and Cell Engraftment of Bone Marrow-derived Cells from Aged Atherogenic Mice

PubMed Central

Xu, Qiyuan; Wang, Jian’An; He, Jinlin; Zhou, Mingsheng; Adi, Jennipher; Webster, Keith A; Yu, Hong

2011-01-01

Objectives Reduced numbers and activity of circulating progenitor cells are associated with aging and have been linked with coronary artery disease. To determine the impact of aging and atherosclerotic disease on the chemotaxic activity of bone marrow derived cells (BMCs), we examined CXCR4 surface expression on BMCs from aged and atherosclerotic mice. Methods CXCR4 expression and cellular mobility were compared between BMCs of young (6-week old) ApoE null mice (ApoE−/−) and aged ApoE−/− mice that had been fed with a high-fat, high-cholesterol diet for 6-months. Results Age and atherosclerosis correlated with significantly lower surface expression of CXCR4 that was less inducible by calcium. The impaired calcium response was associated with defective calcium influx and was partially recovered by treatment with the calcium ionophore ionomycin. ApoE−/− mice fed high fat diet for 6-months had defective CXCR4 expression and SDF-1 regulation that is equivalent to that of 24-month old wild type mice. BMCs from aged, atherogenic ApoE−/− mice also displayed defective homing to SDF-1, and the animals had lower serum and bone marrow levels of SDF-1. Conclusion Evolution of atherosclerosis in ApoE−/− mice is paralleled by progressive loss of mobility of BMCs with reductions of CXCR4 expression, and reduced levels of SDF-1 in both serum and bone marrow. These changes mute the homing capability of BMCs and may contribute to the progression of atherosclerosis in this model. PMID:21855069

Peripheral mineralization of a 3D biodegradable tubular construct as a way to enhance guidance stabilization in spinal cord injury regeneration.

PubMed

Oliveira, A L; Sousa, E C; Silva, N A; Sousa, N; Salgado, A J; Reis, R L

2012-11-01

Spinal cord injuries (SCI) present a major challenge to therapeutic development due to its complexity. Combinatorial approaches using biodegradable polymers that can simultaneously provide a tissue scaffold, a cell vehicle, and a reservoir for sustained drug delivery have shown very promising results. In our previous studies we have developed a novel hybrid system consisting of starch/poly-e-caprolactone (SPCL) semi-rigid tubular porous structure, based on a rapid prototyping technology, filled by a gellan gum hydrogel concentric core for the regeneration within spinal-cord injury sites. In the present work we intend to promote enhanced osteointegration on these systems by pre-mineralizing specifically the external surfaces of the SPCL tubular structures, though a biomimetic strategy, using a sodium silicate gel as nucleating agent. The idea is to create two different cell environments to promote axonal regeneration in the interior of the constructs while inducing osteogenic activity on its external surface. By using a Teflon cylinder to isolate the interior of the scaffold, it was possible to observe the formation of a bone-like poorly crystalline carbonated apatite layer continuously formed only in the external side of the tubular structure. This biomimetic layer was able to support the adhesion of Bone Marrow Mesenchymal Stem Cells, which have gone under cytoskeleton reorganization in the first hours of culture when compared to cells cultured on uncoated scaffolds. This strategy can be a useful route for locally stimulate bone tissue regeneration and facilitating early bone ingrowth.

Effect of functional end groups of silane self assembled monolayer surfaces on apatite formation, fibronectin adsorption and osteoblast cell function

PubMed Central

Toworfe, G.K.; Bhattacharyya, S.; Composto, R.J.; Adams, C.S.; Shapiro, I.M.; Ducheyne, P.

2008-01-01

Bioactive glass (BG) can directly bond to living bone without fibrous tissue encapsulation. Key mechanistic steps of BG’s activity are attributed to calcium phosphate formation, surface hydroxylation and fibronectin (FN) adsorption. In the present study, self-assembled monolayers (SAMs) of alkanesilanes with different surface chemistry (OH, NH2, and COOH) were used as a model system to mimic BG’s surface activity. Calcium phosphate (Ca-P) was formed on SAMs by immersion in a solution which simulates the electrolyte content of physiological fluids. FN adsorption kinetics and monolayer coverage was determined on SAMs with or without Ca-P coating. The surface roughness was also examined on these substrates before and after FN adsorption. The effects of FN-adsorbed, Ca-P coated SAMs on the function of MC3T3-E1 were evaluated by cell growth, expression of alkaline phosphatase activity, and actin cytoskeleton formation. We demonstrate that, although the FN monolayer coverage and the rms roughness are similar on −OH and −COOH terminated SAMs with or without Ca-P coating, higher levels of ALP activity, more actin cytoskeleton formation and more cell growth are obtained on −OH and −COOH terminated SAMs with Ca-P coating. In addition, although the FN monolayer coverage is higher on Ca-P coated −NH2 terminated SAMs and SiOx surfaces, higher levels of ALP activity and more cell growth are obtained on Ca-P coated −OH and −COOH terminated SAMs. Thus with same Ca-P coatings, different surface functional groups have different effects on the function of osteoblastic cells. These findings represent new insights into the mechanism of bioactivity of BG and, thereby, may lead to designing superior constructs for bone grafting. PMID:19012271

The biological response to laser-aided direct metal-coated Titanium alloy (Ti6Al4V)

PubMed Central

Shin, T.; Lim, D.; Kim, Y. S.; Kim, S. C.; Jo, W. L.

2018-01-01

Objectives Laser-engineered net shaping (LENS) of coated surfaces can overcome the limitations of conventional coating technologies. We compared the in vitro biological response with a titanium plasma spray (TPS)-coated titanium alloy (Ti6Al4V) surface with that of a Ti6Al4V surface coated with titanium using direct metal fabrication (DMF) with 3D printing technologies. Methods The in vitro ability of human osteoblasts to adhere to TPS-coated Ti6Al4V was compared with DMF-coating. Scanning electron microscopy (SEM) was used to assess the structure and morphology of the surfaces. Biological and morphological responses to human osteoblast cell lines were then examined by measuring cell proliferation, alkaline phosphatase activity, actin filaments, and RUNX2 gene expression. Results Morphological assessment of the cells after six hours of incubation using SEM showed that the TPS- and DMF-coated surfaces were largely covered with lamellipodia from the osteoblasts. Cell adhesion appeared similar in both groups. The differences in the rates of cell proliferation and alkaline phosphatase activities were not statistically significant. Conclusions The DMF coating applied using metal 3D printing is similar to the TPS coating, which is the most common coating process used for bone ingrowth. The DMF method provided an acceptable surface structure and a viable biological surface. Moreover, this method is automatable and less complex than plasma spraying. Cite this article: T. Shin, D. Lim, Y. S. Kim, S. C. Kim, W. L. Jo, Y. W. Lim. The biological response to laser-aided direct metal-coated Titanium alloy (Ti6Al4V). Bone Joint Res 2018;7:357–361. DOI: 10.1302/2046-3758.75.BJR-2017-0222.R1. PMID:29922456

Niche matters: The comparison between bone marrow stem cells and endometrial stem cells and stromal fibroblasts reveal distinct migration and cytokine profiles in response to inflammatory stimulus

PubMed Central

Sorjamaa, Anna; Kangasniemi, Marika; Sutinen, Meeri; Salo, Tuula; Liakka, Annikki; Lehenkari, Petri; Tapanainen, Juha S.; Vuolteenaho, Olli; Chen, Joseph C.; Lehtonen, Siri; Piltonen, Terhi T.

2017-01-01

Objective Intrinsic inflammatory characteristics play a pivotal role in stem cell recruitment and homing through migration where the subsequent change in niche has been shown to alter these characteristics. The bone marrow mesenchymal stem cells (bmMSCs) have been demonstrated to migrate to the endometrium contributing to the stem cell reservoir and regeneration of endometrial tissue. Thus, the aim of the present study was to compare the inflammation-driven migration and cytokine secretion profile of human bmMSCs to endometrial mesenchymal stem cells (eMSCs) and endometrial fibroblasts (eSFs). Materials and methods The bmMSCs were isolated from bone marrow aspirates through culturing, whereas eMSCs and eSFs were FACS-isolated. All cell types were tested for their surface marker, proliferation profiles and migration properties towards serum and inflammatory attractants. The cytokine/chemokine secretion profile of 35 targets was analysed in each cell type at basal level along with lipopolysaccharide (LPS)-induced state. Results Both stem cell types, bmMSCs and eMSCs, presented with similar stem cell surface marker profiles as well as possessed high proliferation and migration potential compared to eSFs. In multiplex assays, the secretion of 16 cytokine targets was detected and LPS stimulation expanded the cytokine secretion pattern by triggering the secretion of several targets. The bmMSCs exhibited higher cytokine secretion of vascular endothelial growth factor (VEGF)-A, stromal cell-derived factor-1 alpha (SDF)-1α, interleukin-1 receptor antagonist (IL-1RA), IL-6, interferon-gamma inducible protein (IP)-10, monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)1α and RANTES compared to eMSCs and/or eSFs after stimulation with LPS. The basal IL-8 secretion was higher in both endometrial cell types compared to bmMSCs. Conclusion Our results highlight that similar to bmMSCs, the eMSCs possess high migration activity while the differentiation process towards stromal fibroblasts seemed to result in loss of stem cell surface markers, minimal migration activity and a subtler cytokine profile likely contributing to normal endometrial function. PMID:28419140

Cell Adhesion and in Vivo Osseointegration of Sandblasted/Acid Etched/Anodized Dental Implants

PubMed Central

Kim, Mu-Hyon; Park, Kyeongsoon; Choi, Kyung-Hee; Kim, Soo-Hong; Kim, Se Eun; Jeong, Chang-Mo; Huh, Jung-Bo

2015-01-01

The authors describe a new type of titanium (Ti) implant as a Modi-anodized (ANO) Ti implant, the surface of which was treated by sandblasting, acid etching (SLA), and anodized techniques. The aim of the present study was to evaluate the adhesion of MG-63 cells to Modi-ANO surface treated Ti in vitro and to investigate its osseointegration characteristics in vivo. Four different types of Ti implants were examined, that is, machined Ti (control), SLA, anodized, and Modi-ANO Ti. In the cell adhesion study, Modi-ANO Ti showed higher initial MG-63 cell adhesion and induced greater filopodia growth than other groups. In vivo study in a beagle model revealed the bone-to-implant contact (BIC) of Modi-ANO Ti (74.20% ± 10.89%) was much greater than those of machined (33.58% ± 8.63%), SLA (58.47% ± 12.89), or ANO Ti (59.62% ± 18.30%). In conclusion, this study demonstrates that Modi-ANO Ti implants produced by sandblasting, acid etching, and anodizing improve cell adhesion and bone ongrowth as compared with machined, SLA, or ANO Ti implants. These findings suggest that the application of Modi-ANO surface treatment could improve the osseointegration of dental implant. PMID:25955650

Improved Mobilization of Exogenous Mesenchymal Stem Cells to Bone for Fracture Healing and Sex Difference

PubMed Central

Yao, Wei; Evan Lay, Yu-An; Kot, Alexander; Liu, Ruiwu; Zhang, Hongliang; Chen, Haiyan; Lam, Kit; Lane, Nancy E.

2017-01-01

Mesenchymal stem cell (MSC) transplantation has been tested in animal and clinical fracture studies. We have developed a bone-seeking compound, LLP2A-Alendronate (LLP2A-Ale) that augments MSC homing to bone. The purpose of this study was to determine whether treatment with LLP2A-Ale or a combination of LLP2A-Ale and MSCs would accelerate bone healing in a mouse closed fracture model and if the effects are sex dependent. A right mid-femur fracture was induced in two-month-old osterix-mCherry (Osx-mCherry) male and female reporter mice. The mice were subsequently treated with placebo, LLP2A-Ale (500 µg/kg, IV), MSCs derived from wild-type female Osx-mCherry adipose tissue (ADSC, 3 × 105, IV) or ADSC + LLP2A-Ale. In phosphate buffered saline-treated mice, females had higher systemic and surface-based bone formation than males. However, male mice formed a larger callus and had higher volumetric bone mineral density and bone strength than females. LLP2A-Ale treatment increased exogenous MSC homing to the fracture gaps, enhanced incorporation of these cells into callus formation, and stimulated endochondral bone formation. Additionally, higher engraftment of exogenous MSCs in fracture gaps seemed to contribute to overall fracture healing and improved bone strength. These effects were sex-independent. There was a sex-difference in the rate of fracture healing. ADSC and LLP2A-Ale combination treatment was superior to on callus formation, which was independent of sex. Increased mobilization of exogenous MSCs to fracture sites accelerated endochondral bone formation and enhanced bone tissue regeneration. PMID:27334693

Hematopoietic stem cells found in lineage-positive subsets in the bone marrow of 5-fluorouracil-treated mice.

PubMed

Nishi, N; Osawa, M; Ishikawa, R; Nishikawa, M; Tsumura, H; Inoue, H; Sudo, T

1995-09-01

It is known that treatment of mice with 5-fluorouracil (5-FU, 150 mg/kg) confers radioprotection. To investigate this effect, we performed bone marrow transplantation (BMT) using C57BL/6-Ly5 congenic mice treated with 5-FU five days prior to experiments. The mononuclear cells (MNC) in 5-FU-treated bone marrow (BM) were 10 times more radioprotective than those in untreated BM. Moreover, the number of BM MNC expressing c-kit on their surface from 5-FU-treated mice was markedly decreased relative to those from untreated controls. These results showed that the surface characteristics of cells that contributed to this radio-protective effect differ from those of stem cells as reported recently. BM MNC of mice treated with 5-FU were separated on the basis of expression of the lineage-specific antigens (Lin), c-kit, and Ly6A/E. When injected into lethally irradiated mice, 1,000 Lin+ and Lin-c-kit+Ly6A/E+ cells showed radioprotective effects such that 100% and 60% survived, respectively. Flow cytometric analysis 165 days after BMT showed that 88.8% and 65.1% of peripheral blood (PB) in mice transplanted with Lin+ and Lin-c-kit+Ly6A/E+ was derived from donor mice, respectively. After six months, donor-derived Lin-c-kit+Ly6A/E+ cells which showed radioprotective effects on a secondary irradiated host were detected from mice transplanted with Lin+ cells from 5-FU-treated mice. Taken together, these findings demonstrated that stem cells expressing Lin+ present in the BM of mice treated with 5-FU other than Lin-c-kit+Ly6A/E+ cells and these Lin+ cells play an important role in the recovery of myeloablative mice.

Fabrication of Biocompatible Potassium Sodium Niobate Piezoelectric Ceramic as an Electroactive Implant

PubMed Central

Chen, Wei; Yu, Zunxiong; Pang, Jinshan; Yu, Peng; Tan, Guoxin; Ning, Chengyun

2017-01-01

The discovery of piezoelectricity in natural bone has attracted extensive research in emulating biological electricity for various tissue regeneration. Here, we carried out experiments to build biocompatible potassium sodium niobate (KNN) ceramics. Then, influence substrate surface charges on bovine serum albumin (BSA) protein adsorption and cell proliferation on KNN ceramics surfaces was investigated. KNN ceramics with piezoelectric constant of ~93 pC/N and relative density of ~93% were fabricated. The adsorption of protein on the positive surfaces (Ps) and negative surfaces (Ns) of KNN ceramics with piezoelectric constant of ~93 pC/N showed greater protein adsorption capacity than that on non-polarized surfaces (NPs). Biocompatibility of KNN ceramics was verified through cell culturing and live/dead cell staining of MC3T3. The cells experiment showed enhanced cell growth on the positive surfaces (Ps) and negative surfaces (Ns) compared to non-polarized surfaces (NPs). These results revealed that KNN ceramics had great potential to be used to understand the effect of surface potential on cells processes and would benefit future research in designing piezoelectric materials for tissue regeneration. PMID:28772704

Fabrication of Biocompatible Potassium Sodium Niobate Piezoelectric Ceramic as an Electroactive Implant.

PubMed

Chen, Wei; Yu, Zunxiong; Pang, Jinshan; Yu, Peng; Tan, Guoxin; Ning, Chengyun

2017-03-26

The discovery of piezoelectricity in natural bone has attracted extensive research in emulating biological electricity for various tissue regeneration. Here, we carried out experiments to build biocompatible potassium sodium niobate (KNN) ceramics. Then, influence substrate surface charges on bovine serum albumin (BSA) protein adsorption and cell proliferation on KNN ceramics surfaces was investigated. KNN ceramics with piezoelectric constant of ~93 pC/N and relative density of ~93% were fabricated. The adsorption of protein on the positive surfaces (Ps) and negative surfaces (Ns) of KNN ceramics with piezoelectric constant of ~93 pC/N showed greater protein adsorption capacity than that on non-polarized surfaces (NPs). Biocompatibility of KNN ceramics was verified through cell culturing and live/dead cell staining of MC3T3. The cells experiment showed enhanced cell growth on the positive surfaces (Ps) and negative surfaces (Ns) compared to non-polarized surfaces (NPs). These results revealed that KNN ceramics had great potential to be used to understand the effect of surface potential on cells processes and would benefit future research in designing piezoelectric materials for tissue regeneration.

Low-magnitude, high-frequency vibration promotes the adhesion and the osteogenic differentiation of bone marrow-derived mesenchymal stem cells cultured on a hydroxyapatite-coated surface: The direct role of Wnt/β-catenin signaling pathway activation.

PubMed

Chen, Bailing; Lin, Tao; Yang, Xiaoxi; Li, Yiqiang; Xie, Denghui; Zheng, Wenhui; Cui, Haowen; Deng, Weimin; Tan, Xin

2016-11-01

The positive effect of low-magnitude, high‑frequency (LMHF) vibration on implant osseointegration has been demonstrated; however, the underlying cellular and molecular mechanisms remain unknown. The aim of this study was to explore the effect of LMHF vibration on the adhesion and the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) cultured on hydroxyapatite (HA)-coated surfaces in an in vitro model as well as to elucidate the molecular mechanism responsible for the effects of LMHF vibration on osteogenesis. LMHF vibration resulted in the increased expression of fibronectin, which was measured by immunostaining and RT-qPCR. Stimulation of BMSCs by LMHF vibration resulted in the rearrangement of the actin cytoskeleton with more prominent F-actin. Moreover, the expression of β1 integrin, vinculin and paxillin was notably increased following LMHF stimulation. Scanning electron microscope observations revealed that there were higher cell numbers and more extracellular matrix attached to the HA-coated surface in the LMHF group. Alkaline phosphatase activity as well as the expression of osteogenic-specific genes, namely Runx2, osterix, collagen I and osteocalcin, were significantly elevated in the LMHF group. In addition, the protein expression of Wnt10B, β-catenin, Runx2 and osterix was increased following exposure to LMHF vibration. Taken together, the findings of this study indicate that LMHF vibration promotes the adhesion and the osteogenic differentiation of BMSCs on HA-coated surfaces in vitro, and LMHF vibration may directly induce osteogenesis by activating the Wnt/β‑catenin signaling pathway. These data suggest that LMHF vibration enhances the osseointegration of bone to a HA-coated implant, and provide a scientific foundation for improving bone-implant osseointegration through the application of LMHF vibration.

High hydrostatic pressure-induced cell death in human chondrocytes and chondrosarcoma cells.

PubMed

Naal, Florian-Dominique; Mengele, Karin; Schauwecker, Johannes; Gollwitzer, Hans; Gerdesmeyer, Ludger; Reuning, Ute; Mittelmeier, Wolfram; Gradinger, Reiner; Schmitt, Manfred; Diehl, Peter

2005-01-01

In orthopedic surgery, sterilization of bone used for reconstruction of osteoarticular defects caused by malignant tumors is carried out in different ways. At present, to devitalize tumor-bearing osteochondral segments, extracorporal irradiation or autoclaving is mainly used, although both methods have substantial disadvantages, e.g. loss of biomechanical and/or biological integrity of the bone and destabilization of the articular surface. In this regard, high hydrostatic pressure (HHP) treatment of bone is a new, advancing technology, now being used in preclinical testing to inactivate tumor cells. To find out if this technique is also suited for extracorporal inactivation of chondrocytes and chondral tumor cells, the effect of HHP on cell viability and morphology of human chondrocytes / chondrosarcoma cells was investigated in the present study. SW1353 chondrosarcoma cells and chondrocytes were subjected to HHP in the range of 50 to 350 MPa (10 min, 37 degrees C) and, subsequently, cell viability and cell morphology assessed. After exposure at 350 MPa, all HHP-treated chondral cells showed explicit morphological changes, evident by membrane ruffling and bleb formation; chondrosarcoma cells treated this way were irreversibly damaged and not alive. We anticipate that, in orthopedic surgery, HHP eventually can serve as a novel, promising technical approach for cell inactivation (including tumor cells) and allow subsequent reimplantation of the osteoarticular autograft.

Multiscale mechanobiology of de novo bone generation, remodeling and adaptation of autograft in a common ovine femur model.

PubMed

Knothe Tate, Melissa L; Dolejs, Scott; McBride, Sarah H; Matthew Miller, R; Knothe, Ulf R

2011-08-01

The link between mechanics and biology in the generation and the adaptation of bone has been studied for more than a century in the context of skeletal development and fracture healing. However, the interplay between mechanics and biology in de novo generation of bone in postnatal defects as well as healing of morcellized bone graft or massive cortical bone autografts is less well understood. To address this, here we integrate insights from our previously published studies describing the mechanobiology on both de novo bone generation and graft healing in a common ovine femoral defect model. Studying these effects in a common experimental model provides a unique opportunity to elucidate factors conducive to harnessing the regenerative power of the periosteum, and ultimately, to provide mechanistic insights into the multiscale mechanobiology of bone generation, remodeling and adaptation. Taken together, the studies indicate that, as long as adequate, directional transport of cells and molecules can be insured (e.g. with periosteum in situ or a delivery device), biological factors intrinsic to the periosteum suffice to bridge critical sized bone defects, even in the absence of a patent blood supply. Furthermore, mechanical stimuli are crucial for the success of periosteal bone generation and bone graft healing. Interestingly, areas of highest periosteal strain around defects correlate with greatest amounts albeit not greatest mineralization of newly generated bone. This may indicate a role for convection enhanced transport of cells and molecules in modulation of tissue generation by pluripotent cells that ingress into the defect center, away from the periosteum and toward the surface of the intramedullary nail that fills the medullary cavity. These insights bring us much closer to understanding the mechanobiological environment and stimuli that stimulate the proliferation and differentiation of periosteum-derived progenitor cells and ultimately drive the generation of new bone tissue. Furthermore, these insights provide a foundation to create virtual predictive computational models of bone mechanophysiology, to develop cell seeding protocols for scale up and manufacture of engineered tissues, to optimize surgical procedures, and to develop post-surgical therapies with the ultimate goal of achieving the best possible healing outcomes for treatment and/or reconstruction of postnatal bone defects. Copyright © 2011 Elsevier Ltd. All rights reserved.

Fabrication and characterization of novel nano-biocomposite scaffold of chitosan-gelatin-alginate-hydroxyapatite for bone tissue engineering.

PubMed

Sharma, Chhavi; Dinda, Amit Kumar; Potdar, Pravin D; Chou, Chia-Fu; Mishra, Narayan Chandra

2016-07-01

A novel nano-biocomposite scaffold was fabricated in bead form by applying simple foaming method, using a combination of natural polymers-chitosan, gelatin, alginate and a bioceramic-nano-hydroxyapatite (nHAp). This approach of combining nHAp with natural polymers to fabricate the composite scaffold, can provide good mechanical strength and biological property mimicking natural bone. Environmental scanning electron microscopy (ESEM) images of the nano-biocomposite scaffold revealed the presence of interconnected pores, mostly spread over the whole surface of the scaffold. The nHAp particulates have covered the surface of the composite matrix and made the surface of the scaffold rougher. The scaffold has a porosity of 82% with a mean pore size of 112±19.0μm. Swelling and degradation studies of the scaffold showed that the scaffold possesses excellent properties of hydrophilicity and biodegradability. Short term mechanical testing of the scaffold does not reveal any rupturing after agitation under physiological conditions, which is an indicative of good mechanical stability of the scaffold. In vitro cell culture studies by seeding osteoblast cells over the composite scaffold showed good cell viability, proliferation rate, adhesion and maintenance of osteoblastic phenotype as indicated by MTT assay, ESEM of cell-scaffold construct, histological staining and gene expression studies, respectively. Thus, it could be stated that the nano-biocomposite scaffold of chitosan-gelatin-alginate-nHAp has the paramount importance for applications in bone tissue-engineering in future regenerative therapies. Copyright © 2016 Elsevier B.V. All rights reserved.

STATs and macrophage fusion.

PubMed

Miyamoto, Takeshi

2013-07-01

Macrophages play a pivotal role in host defense against multiple foreign materials such as bacteria, parasites and artificial devices. Some macrophage lineage cells, namely osteoclasts and foreign body giant cells (FBGCs), form multi-nuclear giant cells by the cell-cell fusion of mono-nuclear cells. Osteoclasts are bone-resorbing cells, and are formed in the presence of RANKL on the surface of bones, while FBGCs are formed in the presence of IL-4 or IL-13 on foreign materials such as artificial joints, catheters and parasites. Recently, fusiogenic mechanisms and the molecules required for the cell-cell fusion of these macrophage lineage cells were, at least in part, clarified. Dendritic cell specific transmembrane protein (DC-STAMP) and osteoclast stimulatory transmembrane protein (OC-STAMP), both of which comprise seven transmembrane domains, are required for both osteoclast and FBGC cell-cell fusion. STAT6 was demonstrated to be required for the cell-cell fusion of FBGCs but not osteoclasts. In this review, advances in macrophage cell-cell fusion are discussed.

[Minimal residual disease in breast carcinoma].

PubMed

Janků, F; Novotný, J; Vedralová, J; Kleibl, Z; Matous, B; Petruzelka, L

2002-08-30

The incidence of breast cancer continuously increases in developed countries. The introduction of screening methods such as mammography or ultrasound lead to higher proportion of early diagnosed tumors. However, even in early stage tumors occult neoplastic cells can spread to the organism. Such tumor cells are very likely precursors of distant metastases. Using several monoclonal antibodies against epithelial mucins or cytokeratins on the cell surface could be detected one tumor cell among 10(5) or 10(6) of normal bone marrow cells. These cells are not detectable by routine histopathologic exam. More sensitive but also more costly and technically demanding are PCR assays. The sensitivity might reach almost 1:10(7). Prospective clinical trials using immunocytochemistry have shown that the presence of stained cells in bone marrow is clearly associated with shorter disease free survival and overall survival. In the near future we may use the bone marrow examination for the presence of occult tumor cells in order to improve current staging system or as a surrogate marker in the decision-making in regard to adjuvant systemic therapy or in the assessment of efficacy of adjuvant treatment. The review summarizes contemporary knowledge assembled in preclinical and clinical studies.

In vitro evaluation of three-dimensional single-walled carbon nanotube composites for bone tissue engineering.

PubMed

Gupta, Ashim; Main, Benjamin J; Taylor, Brittany L; Gupta, Manu; Whitworth, Craig A; Cady, Craig; Freeman, Joseph W; El-Amin, Saadiq F

2014-11-01

The purpose of this study was to develop three-dimensional single-walled carbon nanotube composites (SWCNT/PLAGA) using 10-mg single-walled carbon nanotubes (SWCNT) for bone regeneration and to determine the mechanical strength of the composites, and to evaluate the interaction of MC3T3-E1 cells via cell adhesion, growth, survival, proliferation, and gene expression. PLAGA (polylactic-co-glycolic acid) and SWCNT/PLAGA microspheres and composites were fabricated, characterized, and mechanical testing was performed. MC3T3-E1 cells were seeded and cell adhesion/morphology, growth/survival, proliferation, and gene expression analysis were performed to evaluate biocompatibility. Imaging studies demonstrated microspheres with uniform shape and smooth surfaces, and uniform incorporation of SWCNT into PLAGA matrix. The microspheres bonded in a random packing manner while maintaining spacing, thus resembling trabeculae of cancellous bone. Addition of SWCNT led to greater compressive modulus and ultimate compressive strength. Imaging studies revealed that MC3T3-E1 cells adhered, grew/survived, and exhibited normal, nonstressed morphology on the composites. SWCNT/PLAGA composites exhibited higher cell proliferation rate and gene expression compared with PLAGA. These results demonstrate the potential of SWCNT/PLAGA composites for musculoskeletal regeneration, for bone tissue engineering, and are promising for orthopedic applications as they possess the combined effect of increased mechanical strength, cell proliferation, and gene expression. © 2014 Wiley Periodicals, Inc.

Behavior and biocompatibility of rabbit bone marrow mesenchymal stem cells with bacterial cellulose membrane

PubMed Central

Leite, Yulla Klinger de Carvalho; de Carvalho, Camila Ernanda Sousa; Feitosa, Matheus Levi Tajra; Alves, Michel Muálem de Moraes; Carvalho, Fernando Aécio de Amorim; Neto, Bartolomeu Cruz Viana; Miglino, Maria Angélica

2018-01-01

Background Tissue engineering has been shown to exhibit great potential for the creation of biomaterials capable of developing into functional tissues. Cellular expansion and integration depends on the quality and surface-determinant factors of the scaffold, which are required for successful biological implants. The objective of this research was to characterize and evaluate the in vitro characteristics of rabbit bone marrow mesenchymal stem cells (BM-MSCs) associated with a bacterial cellulose membrane (BCM). We assessed the adhesion, expansion, and integration of the biomaterial as well as its ability to induce macrophage activation. Finally, we evaluated the cytotoxicity and toxicity of the BCM. Methods Samples of rabbit bone marrow were collected. Mesenchymal stem cells were isolated from medullary aspirates to establish fibroblast colony-forming unit assay. Osteogenic, chondrogenic, and adipogenic differentiation was performed. Integration with the BCM was assessed by scanning electron microscopy at 1, 7, and 14 days. Cytotoxicity was assessed via the production of nitric oxide, and BCM toxicity was assessed with the MTT assay; phagocytic activity was also determined. Results The fibroblastoid colony-forming unit (CFU-F) assay showed cells with a fibroblastoid morphology organized into colonies, and distributed across the culture area surface. In the growth curve, two distinct phases, lag and log phase, were observed at 15 days. Multipotentiality of the cells was evident after induction of osteogenic, chondrogenic, and adipogenic lineages. Regarding the BM-MSCs’ bioelectrical integration with the BCM, BM-MSCs were anchored in the BCM in the first 24 h. On day 7 of culture, the cytoplasm was scattered, and on day 14, the cells were fully integrated with the biomaterial. We also observed significant macrophage activation; analysis of the MTT assay and the concentration of nitric oxide revealed no cytotoxicity of the biomaterial. Conclusion The BCM allowed the expansion and biointegration of bone marrow progenitor cells with a stable cytotoxic profile, thus presenting itself as a biomaterial with potential for tissue engineering. PMID:29736332

Development of a new method to harvest chondroprogenitor cells from underneath cartilage defects in the knees.

PubMed

Elvenes, Jan; Knutsen, Gunnar; Johansen, Oddmund; Moe, Bjørn T; Martinez, Inigo

2009-07-01

Mesenchymal progenitor cells from bone marrow hold great potential as a cell source for cartilage repair. Aspiration from the iliac crest is the most widely used method to harvest bone marrow cells for cartilage repair. The objective of our study was to establish a new method to isolate mesenchymal progenitor cells by direct aspiration of bone marrow from the subchondral spongious bone underneath cartilage defects during microfracture treatment and to confirm the chondrogenic potential of the resulting cell cultures. Bone marrow was aspirated arthroscopically from patients treated for isolated cartilage defects. Adherent stromal cells were isolated, expanded in monolayer cultures, and characterized by flow cytometry. Chondrogenic induction of cells was achieved by combination of spheroid cultures in hanging drops and the concomitant use of transforming growth factor-beta (TGFbeta). Articular chondrocytes established in three-dimensional (3D) cultures were used as positive cartilage-forming units, and skin fibroblasts were used as negative controls. Three-dimensional constructs were stained for immunohistochemical and histological examination, and a real-time polymerase chain reaction (PCR) was performed to quantify the expression of aggrecan, collagen types 1 and 2, and Sox9. Mesenchymal stem cell-like progenitor cells (MSCs) displaying chondrogenic differentiation capacity were harvested arthroscopically from underneath cartilage lesions on distal femurs using the one-hole technique. Stem cell-related surface antigens analyzed by flow cytometry confirmed the nature of the isolated adherent cells. MSC spheroids stained positive for glycosaminoglycans and collagen type 2. Realtime PCR showed that MSCs in 3D spheroids significantly increased gene expression of collagen type 2, aggrecan, and Sox 9 and down-regulated expression of collagen type 1 when compared to the mRNA levels measured in MSCs monolayers. We describe a new technique that may be applied for harvesting bone marrow cells from cartilage defects during arthroscopic intervention of the knee. Cells harvested in this way hold full chondrogenic differentiation potential. Our data imply that MSC storage may be established by using marrow from this approach, bypassing the need for cell aspiration from the iliac crest.

Accelerated Bone Repair After Plasma Laser Corticotomies

PubMed Central

Leucht, Philipp; Lam, Kentson; Kim, Jae-Beom; Mackanos, Mark A.; Simanovskii, Dmitrii M.; Longaker, Michael T.; Contag, Christopher H.; Schwettman, H Alan; Helms, Jill A.

2007-01-01

Objective: To reveal, on a cellular and molecular level, how skeletal regeneration of a corticotomy is enhanced when using laser-plasma mediated ablation compared with conventional mechanical tissue removal. Summary Background Data: Osteotomies are well-known for their most detrimental side effect: thermal damage. This thermal and mechanical trauma to adjacent bone tissue can result in the untoward consequences of cell death and eventually in a delay in healing. Methods: Murine tibial corticotomies were performed using a conventional saw and a Ti:Sapphire plasma-generated laser that removes tissue with minimal thermal damage. Our analyses began 24 hours after injury and proceeded to postsurgical day 6. We investigated aspects of wound repair ranging from vascularization, inflammation, cell proliferation, differentiation, and bone remodeling. Results: Histology of mouse corticotomy sites uncovered a significant difference in the onset of bone healing; whereas laser corticotomies showed abundant bone matrix deposition at postsurgical day 6, saw corticotomies only exhibited undifferentiated tissue. Our analyses uncovered that cutting bone with a saw caused denaturation of the collagen matrix due to thermal effects. This denatured collagen represented an unfavorable scaffold for subsequent osteoblast attachment, which in turn impeded deposition of a new bony matrix. The matrix degradation induced a prolonged inflammatory reaction at the cut edge to create a surface favorable for osteochondroprogenitor cell attachment. Laser corticotomies were absent of collagen denaturation, therefore osteochondroprogenitor cell attachment was enabled shortly after surgery. Conclusion: In summary, these data demonstrate that corticotomies performed with Ti:Sapphire lasers are associated with a reduced initial inflammatory response at the injury site leading to accelerated osteochondroprogenitor cell migration, attachment, differentiation, and eventually matrix deposition. PMID:17592303

Effects of graphene modification on the bioactivation of polyethylene-terephthalate-based artificial ligaments.

PubMed

Wang, Chun-Hui; Guo, Zhong-Shang; Pang, Fei; Zhang, Li-Yuan; Yan, Ming; Yan, Jin-Hong; Li, Ke-Wen; Li, Xiao-Jie; Li, Yong; Bi, Long; Han, Yi-Sheng

2015-07-22

The objective of this study was to investigate whether surface coating with graphene could enhance the surface bioactivation of PET-based artificial ligaments to accelerate graft-to-bone healing after anterior cruciate ligament reconstruction. In an in vitro study, the proliferation of MC3T3-E1 cells and their differentiation on the scaffolds were quantified via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and real-time polymerase chain reaction assays. The significantly higher optical-density values and transcription levels of osteoblast-specific genes indicated that graphene modification could promote the proliferation of MC3T3-E1 cells and accelerate their specific differentiation into osteogenic lineages on scaffolds. In an in vivo test, rabbits were used to establish an extra-articular graft-to-bone healing model. At 4, 8, and 12 weeks after surgery, biomechanical tests, microcomputed tomography analysis, and histological observations were performed. The final results demonstrated that the microstructural parameters, the average mineral apposition rate of the bone, and the biomechanical properties of the graphene-coated polyethylene terephthalate (PET)-based artificial ligament (G-PET-AL) group were significantly higher than those of the PET-AL graft group (P < 0.05). The results of Van Gieson staining indicated that in the G-PET-AL group, there was more newly formed bone than there was in the group in which nongraphene-coated PET-ALs were used. In conclusion, graphene exhibits considerable potential for enhancing the surface bioactivation of materials.

Numerical simulation of electrically stimulated osteogenesis in dental implants.

PubMed

Vanegas-Acosta, J C; Garzón-Alvarado, D A; Lancellotti, V

2014-04-01

Cell behavior and tissue formation are influenced by a static electric field (EF). Several protocols for EF exposure are aimed at increasing the rate of tissue recovery and reducing the healing times in wounds. However, the underlying mechanisms of the EF action on cells and tissues are still a matter of research. In this work we introduce a mathematical model for electrically stimulated osteogenesis at the bone-dental implant interface. The model describes the influence of the EF in the most critical biological processes leading to bone formation at the bone-dental implant interface. The numerical solution is able to reproduce the distribution of spatial-temporal patterns describing the influence of EF during blood clotting, osteogenic cell migration, granulation tissue formation, displacements of the fibrillar matrix, and formation of new bone. In addition, the model describes the EF-mediated cell behavior and tissue formation which lead to an increased osteogenesis in both smooth and rough implant surfaces. Since numerical results compare favorably with experimental evidence, the model can be used to predict the outcome of using electrostimulation in other types of wounds and tissues. Copyright © 2013 Elsevier B.V. All rights reserved.

Gametocytes of the Malaria Parasite Plasmodium falciparum Interact With and Stimulate Bone Marrow Mesenchymal Cells to Secrete Angiogenetic Factors

PubMed Central

Messina, Valeria; Valtieri, Mauro; Rubio, Mercedes; Falchi, Mario; Mancini, Francesca; Mayor, Alfredo; Alano, Pietro; Silvestrini, Francesco

2018-01-01

The gametocytes of Plasmodium falciparum, responsible for the transmission of this malaria parasite from humans to mosquitoes, accumulate and mature preferentially in the human bone marrow. In the 10 day long sexual development of P. falciparum, the immature gametocytes reach and localize in the extravascular compartment of this organ, in contact with several bone marrow stroma cell types, prior to traversing the endothelial lining and re-entering in circulation at maturity. To investigate the host parasite interplay underlying this still obscure process, we developed an in vitro tridimensional co-culture system in a Matrigel scaffold with P. falciparum gametocytes and self-assembling spheroids of human bone marrow mesenchymal cells (hBM-MSCs). Here we show that this co-culture system sustains the full maturation of the gametocytes and that the immature, but not the mature, gametocytes adhere to hBM-MSCs via trypsin-sensitive parasite ligands exposed on the erythrocyte surface. Analysis of a time course of gametocytogenesis in the co-culture system revealed that gametocyte maturation is accompanied by the parasite induced stimulation of hBM-MSCs to secrete a panel of 14 cytokines and growth factors, 13 of which have been described to play a role in angiogenesis. Functional in vitro assays on human bone marrow endothelial cells showed that supernatants from the gametocyte mesenchymal cell co-culture system enhance ability of endothelial cells to form vascular tubes. These results altogether suggest that the interplay between immature gametocytes and hBM-MSCs may induce functional and structural alterations in the endothelial lining of the human bone marrow hosting the P. falciparum transmission stages. PMID:29546035

Potential for thermal damage to articular cartilage by PMMA reconstruction of a bone cavity following tumor excision: A finite element study.

PubMed

Radev, Boyko R; Kase, Jonathan A; Askew, Michael J; Weiner, Scott D

2009-05-29

Benign, giant cell tumors are often treated by intralesional excision and reconstruction with polymethylmethacrylate (PMMA) bone cement. The exothermic reaction of the in-situ polymerizing PMMA is believed to beneficially kill remaining tumor cells. However, at issue is the extent of this necrotic effect into the surrounding normal bone and the adjacent articular cartilage. Finite element analysis (ABAQUS 6.4-1) was used to determine the extent of possible thermal necrosis around prismatically shaped, PMMA implants (8-24cc in volume), placed into a peripheral, sagittally symmetric, metaphyseal defect in the proximal tibia. Temperature/exposure time conditions indicating necrotic potential during the exotherm of the polymerizing bone cement were found in regions of the cancellous bone within 3mm of the superior surface of the PMMA implant. If less than 3mm of cancellous bone existed between the PMMA implant and the subchondral bone layer, regions of the subchondral bone were also exposed to thermally necrotic conditions. However, as long as there were at least 2mm of uniform subchondral bone above the PMMA implant, the necrotic regions did not extend into the overlying articular cartilage. This was the case even when the PMMA was in direct contact with the subchondral bone. If the subchondral bone is not of sufficient thickness, or is not continuous, then care should be taken to protect the articular cartilage from thermal damage as a result of the reconstruction of the tumor cavity with PMMA bone cement.

Variation in primary and culture-expanded cells derived from connective tissue progenitors in human bone marrow space, bone trabecular surface and adipose tissue.

PubMed

Qadan, Maha A; Piuzzi, Nicolas S; Boehm, Cynthia; Bova, Wesley; Moos, Malcolm; Midura, Ronald J; Hascall, Vincent C; Malcuit, Christopher; Muschler, George F

2018-03-01

Connective tissue progenitors (CTPs) embody the heterogeneous stem and progenitor cell populations present in native tissue. CTPs are essential to the formation and remodeling of connective tissue and represent key targets for tissue-engineering and cell-based therapies. To better understand and characterize CTPs, we aimed to compare the (i) concentration and prevalence, (ii) early in vitro biological behavior and (iii) expression of surface-markers and transcription factors among cells derived from marrow space (MS), trabecular surface (TS), and adipose tissues (AT). Cancellous-bone and subcutaneous-adipose tissues were collected from 8 patients. Cells were isolated and cultured. Colony formation was assayed using Colonyze software based on ASTM standards. Cell concentration ([Cell]), CTP concentration ([CTP]) and CTP prevalence (P CTP ) were determined. Attributes of culture-expanded cells were compared based on (i) effective proliferation rate and (ii) expression of surface-markers CD73, CD90, CD105, SSEA-4, SSEA-3, SSEA-1/CD15, Cripto-1, E-Cadherin/CD324, Ep-CAM/CD326, CD146, hyaluronan and transcription factors Oct3/4, Sox-2 and Nanog using flow cytometry. Mean [Cell], [CTP] and P CTP were significantly different between MS and TS samples (P = 0.03, P = 0.008 and P= 0.0003), respectively. AT-derived cells generated the highest mean total cell yield at day 6 of culture-4-fold greater than TS and more than 40-fold greater than MS per million cells plated. TS colonies grew with higher mean density than MS colonies (290 ± 11 versus 150 ± 11 cell per mm 2 ; P = 0.0002). Expression of classical-mesenchymal stromal cell (MSC) markers was consistently recorded (>95%) from all tissue sources, whereas all the other markers were highly variable. The prevalence and biological potential of CTPs are different between patients and tissue sources and lack variation in classical MSC markers. Other markers are more likely to discriminate differences between cell populations in biological performance. Understanding the underlying reasons for variation in the concentration, prevalence, marker expression and biological potential of CTPs between patients and source tissues and determining the means of managing this variation will contribute to the rational development of cell-based clinical diagnostics and targeted cell-based therapies. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

CD147 deficiency blocks IL-8 secretion and inhibits lung cancer-induced osteoclastogenesis

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

Wang, Hongkai; Zhuo, Yunyun; Hu, Xu

2015-03-06

Bone is a frequent target of lung cancer metastasis, which is associated with significant morbidity and poor prognosis; however, the molecular basis of this process is still unknown. This study investigated the role of extracellular matrix metalloproteinase inducer (also known as cluster of differentiation (CD)147) in osteoclastogenesis resulting from bone metastasis, based on the enrichment of this glycoprotein on the surface of many malignant bone tumors. RNA interference was used to silence CD147 expression in A549 human lung cancer cells. Compared with conditioned medium (CM) from control cells (A549-CM), CM from CD147-deficient cells (A549-si-CM) suppressed receptor activator of nuclear factormore » κB ligand-stimulated osteoclastogenesis in RAW 264.7 cells and bone marrow-derived macrophages. The mRNA levels of osteoclast-specific genes such as tartrate-resistant acid phosphatase, calcitonin receptor, and cathepsin K were also reduced in the presence of A549-si-CM. CD147 knockdown in A549 cells decreased interleukin (IL)-8mRNA and protein expression. IL-8 is present in large amounts in A549-CM and mimicked its inductive effect on osteoclastogenesis; this was reversed by depletion of IL-8 from the medium. Taken together, these results indicate that CD147 promotes lung cancer-induced osteoclastogenesis by modulating IL-8 secretion, and suggest that CD147 is a potential therapeutic target for cancer-associated bone resorption in lung cancer patients. - Highlights: • Bone loss frequently results from lung cancer metastasis. • Cluster of differentiation (CD)147 was depleted in A549 lung adenocarcinoma cells. • RAW 264.7 cell osteoclastogenesis was blocked by medium from CD147-deficient cells. • Interleukin (IL)-8 level was reduced in the conditioned medium. • Osteoclastogenesis induced by lung tumor cells requires CD147-mediated IL-8 release.« less

Bone-like apatite coating on functionalized poly(etheretherketone) surface via tailored silanization layers technique.

PubMed

Zheng, Yanyan; Xiong, Chengdong; Zhang, Shenglan; Li, Xiaoyu; Zhang, Lifang

2015-10-01

Poly(etheretherketone) (PEEK) is a rigid semi-crystalline polymer with outstanding mechanical properties, bone-like stiffness and suitable biocompatibility that has attracted much interest as a biomaterial for orthopedic and dental implants. However, the bio-inert surface of PEEK limits its biomedical applications when direct osteointegration between the implants and the host tissue is desired. In this work, -PO4H2, -COOH and -OH groups were introduced on the PEEK surface by further chemical treatments of the vinyl-terminated silanization layers formed on the hydroxylation-pretreated PEEK surface. Both the surface-functionalized and pristine specimens were characterized by X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and water contact angle measurements. When placed in 1.5 strength simulated body fluid (SBF) solution, apatite was observed to form uniformly on the functionalized PEEK surface and firmly attach to the substrate. The characterized results demonstrated that the coating was constituted by poorly crystallized bone-like apatite and the effect of surface functional groups on coating formation was also discussed in detail. In addition, in vitro biocompatibility of PEEK, in terms of pre-osteoblast cell (MC3T3-E1) attachment, spreading and proliferation, was remarkably enhanced by the bone-like apatite coating. Thus, this study provides a method to enhance the bioactivity of PEEK and expand its applications in orthopedic and dental implants. Copyright © 2015 Elsevier B.V. All rights reserved.

Bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber composite: biomechanical properties and biocompatibility

PubMed Central

Qiao, Bo; Li, Jidong; Zhu, Qingmao; Guo, Shuquan; Qi, Xiaotong; Li, Weichao; Wu, Jun; Liu, Yang; Jiang, Dianming

2014-01-01

An ideal bone plate for internal fixation of bone fractures should have good biomechanical properties and biocompatibility. In this study, we prepared a new nondegradable bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber (n-HA/PA66/GF) composite. A breakage area on the n-HA/PA66/GF plate surface was characterized by scanning electron microscopy. Its mechanical properties were investigated using bone-plate constructs and biocompatibility was evaluated in vitro using bone marrow-derived mesenchymal stem cells. The results confirmed that adhesion between the n-HA/PA66 matrix and the glass fibers was strong, with only a few fibers pulled out at the site of breakage. Fractures fixed by the n-HA/PA66/GF plate showed lower stiffness and had satisfactory strength compared with rigid fixation using a titanium plate. Moreover, the results with regard to mesenchymal stem cell morphology, MTT assay, Alizarin Red S staining, enzyme-linked immunosorbent assay, and reverse transcription polymerase chain reaction for alkaline phosphatase and osteocalcin showed that the n-HA/PA66/GF composite was suitable for attachment and proliferation of mesenchymal stem cells, and did not have a negative influence on matrix mineralization or osteogenic differentiation of mesenchymal stem cells. These observations indicate that the n-HA/PA66/GF plate has good biomechanical properties and biocompatibility, and may be considered a new option for internal fixation in orthopedic surgery. PMID:24669191

Bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber composite: biomechanical properties and biocompatibility.

PubMed

Qiao, Bo; Li, Jidong; Zhu, Qingmao; Guo, Shuquan; Qi, Xiaotong; Li, Weichao; Wu, Jun; Liu, Yang; Jiang, Dianming

2014-01-01

An ideal bone plate for internal fixation of bone fractures should have good biomechanical properties and biocompatibility. In this study, we prepared a new nondegradable bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber (n-HA/PA66/GF) composite. A breakage area on the n-HA/PA66/GF plate surface was characterized by scanning electron microscopy. Its mechanical properties were investigated using bone-plate constructs and biocompatibility was evaluated in vitro using bone marrow-derived mesenchymal stem cells. The results confirmed that adhesion between the n-HA/PA66 matrix and the glass fibers was strong, with only a few fibers pulled out at the site of breakage. Fractures fixed by the n-HA/PA66/GF plate showed lower stiffness and had satisfactory strength compared with rigid fixation using a titanium plate. Moreover, the results with regard to mesenchymal stem cell morphology, MTT assay, Alizarin Red S staining, enzyme-linked immunosorbent assay, and reverse transcription polymerase chain reaction for alkaline phosphatase and osteocalcin showed that the n-HA/PA66/GF composite was suitable for attachment and proliferation of mesenchymal stem cells, and did not have a negative influence on matrix mineralization or osteogenic differentiation of mesenchymal stem cells. These observations indicate that the n-HA/PA66/GF plate has good biomechanical properties and biocompatibility, and may be considered a new option for internal fixation in orthopedic surgery.

Periosteum tissue engineering-a review.

PubMed

Li, Nanying; Song, Juqing; Zhu, Guanglin; Li, Xiaoyu; Liu, Lei; Shi, Xuetao; Wang, Yingjun

2016-10-18

As always, the clinical therapy of critical size bone defects caused by trauma, tumor removal surgery or congenital malformation is facing great challenges. Currently, various approaches including autograft, allograft and cell-biomaterial composite based tissue-engineering strategies have been implemented to reconstruct injured bone. However, due to damage during the transplantation processes or design negligence of the bionic scaffolds, these methods expose vulnerabilities without the assistance of periosteum, a bilayer membrane on the outer surface of the bone. Periosteum plays a significant role in bone formation and regeneration as a store for progenitor cells, a source of local growth factors and a scaffold to recruit cells and growth factors, and more and more researchers have recognized its great value in tissue engineering application. Besides direct transplantation, periosteum-derived cells can be cultured on various scaffolds for osteogenesis or chondrogenesis application due to their availability. Research studies also provide a biomimetic methodology to synthesize artificial periosteum which mimic native periosteum in structure or function. According to the studies, these tissue-engineered periostea did obviously enhance the therapeutic effects of bone graft and scaffold engineering while they could be directly used as substitutes of native periosteum. Periosteum tissue engineering, whose related research studies have provided new opportunities for the development of bone tissue engineering and therapy, has gradually become a hot spot and there are still lots to consummate. In this review, tissue-engineered periostea were classified into four kinds and discussed, which might help subsequent researchers get a more systematic view of pseudo-periosteum.

UV-activated 7-dehydrocholesterol-coated titanium implants promote differentiation of human umbilical cord mesenchymal stem cells into osteoblasts.

PubMed

Satué, María; Ramis, Joana M; Monjo, Marta

2016-01-01

Vitamin D metabolites are essential for bone regeneration and mineral homeostasis. The vitamin D precursor 7-dehydrocholesterol can be used after UV irradiation to locally produce active vitamin D by osteoblastic cells. Furthermore, UV-irradiated 7-dehydrocholesterol is a biocompatible coating for titanium implants with positive effects on osteoblast differentiation. In this study, we examined the impact of titanium implants surfaces coated with UV-irradiated 7-dehydrocholesterol on the osteogenic differentiation of human umbilical cord mesenchymal stem cells. First, the synthesis of cholecalciferol (D3) was achieved through the incubation of the UV-activated 7-dehydrocholesterol coating for 48 h at 23℃. Further, we investigated in vitro the biocompatibility of this coating in human umbilical cord mesenchymal stem cells and its potential to enhance their differentiation towards the osteogenic lineage. Human umbilical cord mesenchymal stem cells cultured onto UV-irradiated 7-dehydrocholesterol-coated titanium implants surfaces, combined with osteogenic supplements, upregulated the gene expression of several osteogenic markers and showed higher alkaline phosphatase activity and calcein blue staining, suggesting increased mineralization. Thus, our results show that the use of UV irradiation on 7-dehydrocholesterol -treated titanium implants surfaces generates a bioactive coating that promotes the osteogenic differentiation of human umbilical cord mesenchymal stem cells, with regenerative potential for improving osseointegration in titanium-based bone anchored implants. © The Author(s) 2015.

Zero-order controlled release of BMP2-derived peptide P24 from the chitosan scaffold by chemical grafting modification technique for promotion of osteogenesis in vitro and enhancement of bone repair in vivo.

PubMed

Chen, Yan; Liu, Xujie; Liu, Rui; Gong, Yong; Wang, Mingbo; Huang, Qianli; Feng, Qingling; Yu, Bo

2017-01-01

Combination of tissue-engineered bone scaffolds with cell-adhesive, osteoconductive, or osteoinductive biomolecules is a critical strategy to improve their properties that significantly influence cellular behaviors, such as adhesion, proliferation, and differentiation, which is beneficial for critical-sized bone defects repairing. However, the traditional surface modification techniques, such as physical adsorption, coating, and plasma treatment, et al, have great limitations for immobilization of bioactive molecules due to undesirable controlled delivery performance or overly complex multistep procedures. In this study, we functionalized the chitosan/hydroxyapatite (CS/HA) biomimetic composite scaffold for controlled delivery of BMP2-derived peptide (P24) by the chemical grafting modification technique: firstly, P24 was conjugated with a thiolated chitosan, chitosan-4-thiobutylamidine (CS-TBA); secondly, the resultant CS-P24 was then combined with HA to prepare CS-P24/HA scaffolds. The effect of CS-P24/HA scaffolds on bone regeneration was evaluated, along with the underlying biological mechanisms responsible in vitro and in vivo . In vitro , the controlled and sustained release of bioactive P24 could last up to 90 days, furthermore, the release profiles of CS-5%P24/HA and CS-10%P24/HA were linear and could be fitted according to zero-order kinetic model (R 2 =0.9929; R 2 =0.9757); P24 on the scaffold significantly promoted cell adhesion, proliferation, osteodifferentiation, and mineralization with synergistic effects. Bone marrow stromal cells (BMSCs) revealed spindle-shaped surface morphology, indicating the CS-P24/HA scaffolds supported cell adhesion and possessed a high proliferation rate that varied according to the P24 concentration levels. Furthermore, mRNA levels for OCN, Runx2, and collagen I were significantly up-regulated on CS-P24/HA scaffolds compared with cells grown on CS/HA scaffolds in vitro ( p < 0.05). Similarly, the BMSCs exhibited a higher ALP expression and calcium deposition level on CS-P24/HA scaffolds compared with CS/HA scaffolds ( p < 0.05). In vivo , osteoinductive studies revealed a significantly higher ectopic osteogenesis level of CS-10%P24/HA scaffolds in rat dorsal muscle pockets compared with that of CS/HA scaffolds. Finally, CS-P24/HA scaffolds showed superior performance in the reconstruction of rat calvarial bone defects. This novel CS-P24/HA scaffold is deemed a strong potential candidate for the repair of bone defects in human bone tissue engineering.

Zero-order controlled release of BMP2-derived peptide P24 from the chitosan scaffold by chemical grafting modification technique for promotion of osteogenesis in vitro and enhancement of bone repair in vivo

PubMed Central

Chen, Yan; Liu, Xujie; Liu, Rui; Gong, Yong; Wang, Mingbo; Huang, Qianli; Feng, Qingling; Yu, Bo

2017-01-01

Combination of tissue-engineered bone scaffolds with cell-adhesive, osteoconductive, or osteoinductive biomolecules is a critical strategy to improve their properties that significantly influence cellular behaviors, such as adhesion, proliferation, and differentiation, which is beneficial for critical-sized bone defects repairing. However, the traditional surface modification techniques, such as physical adsorption, coating, and plasma treatment, et al, have great limitations for immobilization of bioactive molecules due to undesirable controlled delivery performance or overly complex multistep procedures. In this study, we functionalized the chitosan/hydroxyapatite (CS/HA) biomimetic composite scaffold for controlled delivery of BMP2-derived peptide (P24) by the chemical grafting modification technique: firstly, P24 was conjugated with a thiolated chitosan, chitosan-4-thiobutylamidine (CS-TBA); secondly, the resultant CS-P24 was then combined with HA to prepare CS-P24/HA scaffolds. The effect of CS-P24/HA scaffolds on bone regeneration was evaluated, along with the underlying biological mechanisms responsible in vitro and in vivo. In vitro, the controlled and sustained release of bioactive P24 could last up to 90 days, furthermore, the release profiles of CS-5%P24/HA and CS-10%P24/HA were linear and could be fitted according to zero-order kinetic model (R2=0.9929; R2=0.9757); P24 on the scaffold significantly promoted cell adhesion, proliferation, osteodifferentiation, and mineralization with synergistic effects. Bone marrow stromal cells (BMSCs) revealed spindle-shaped surface morphology, indicating the CS-P24/HA scaffolds supported cell adhesion and possessed a high proliferation rate that varied according to the P24 concentration levels. Furthermore, mRNA levels for OCN, Runx2, and collagen I were significantly up-regulated on CS-P24/HA scaffolds compared with cells grown on CS/HA scaffolds in vitro (p < 0.05). Similarly, the BMSCs exhibited a higher ALP expression and calcium deposition level on CS-P24/HA scaffolds compared with CS/HA scaffolds (p < 0.05). In vivo, osteoinductive studies revealed a significantly higher ectopic osteogenesis level of CS-10%P24/HA scaffolds in rat dorsal muscle pockets compared with that of CS/HA scaffolds. Finally, CS-P24/HA scaffolds showed superior performance in the reconstruction of rat calvarial bone defects. This novel CS-P24/HA scaffold is deemed a strong potential candidate for the repair of bone defects in human bone tissue engineering. PMID:28435449

Diminished potential for B-lymphoid differentiation after murine leukemia virus infection in vivo and in EML hematopoietic progenitor cells.

PubMed

Finstad, Samantha L; Rosenberg, Naomi; Levy, Laura S

2007-07-01

Infection with a recombinant murine-feline gammaretrovirus, MoFe2, or with the parent virus, Moloney murine leukemia virus, caused significant reduction in B-lymphoid differentiation of bone marrow at 2 to 8 weeks postinfection. The suppression was selective, in that myeloid potential was significantly increased by infection. Analysis of cell surface markers and immunoglobulin H gene rearrangements in an in vitro model demonstrated normal B-lymphoid differentiation after infection but significantly reduced viability of differentiating cells. This reduction in viability may confer a selective advantage on undifferentiated lymphoid progenitors in the bone marrow of gammaretrovirus-infected animals and thereby contribute to the establishment of a premalignant state.

Human alveolar bone cell proliferation, expression of osteoblastic phenotype, and matrix mineralization on porous titanium produced by powder metallurgy.

PubMed

Rosa, Adalberto Luiz; Crippa, Grasiele Edilaine; de Oliveira, Paulo Tambasco; Taba, Mario; Lefebvre, Louis-Philippe; Beloti, Marcio Mateus

2009-05-01

This study aimed at investigating the influence of the porous titanium (Ti) structure on the osteogenic cell behaviour. Porous Ti discs were fabricated by the powder metallurgy process with the pore size typically between 50 and 400 microm and a porosity of 60%. Osteogenic cells obtained from human alveolar bone were cultured until subconfluence and subcultured on dense Ti (control) and porous Ti for periods of up to 17 days. Cultures grown on porous Ti exhibited increased cell proliferation and total protein content, and lower levels of alkaline phosphatase (ALP) activity than on dense Ti. In general, gene expression of osteoblastic markers-runt-related transcription factor 2, collagen type I, alkaline phosphatase, bone morphogenetic protein-7, and osteocalcin was lower at day 7 and higher at day 17 in cultures grown on porous Ti compared with dense Ti, a finding consistent with the enhanced growth rate for such cultures. The amount of mineralized matrix was greater on porous Ti compared with the dense one. These results indicate that the porous Ti is an appropriate substrate for osteogenic cell adhesion, proliferation, and production of a mineralized matrix. Because of the three-dimensional environment it provides, porous Ti should be considered an advantageous substrate for promoting desirable implant surface-bone interactions.

Integrins in bone metastasis formation and potential therapeutic implications.

PubMed

Clëzardin, P

2009-11-01

Integrins constitute a family of cell surface receptors that are heterodimers composed of noncovalently associated alpha and beta subunits. Integrins bind to extracellular matrix proteins and immunogobulin superfamily molecules. They exert a stringent control on cell migration, survival and proliferation. However, their expression and functions are often deregulated in cancer, and many lines of evidence implicate them as key regulators during progression from primary tumor growth to metastasis. Here, we review the role of integrins in bone metastasis formation and present evidence that the use of integrin-targeted therapeutic agents may be an efficient strategy to block tumor metastasis.

Bisphosphonate-coated BSA nanoparticles lack bone targeting after systemic administration.

PubMed

Wang, Guilin; Kucharski, Cezary; Lin, Xiaoyue; Uludağ, Hasan

2010-09-01

A polymeric conjugate of polyethyleneimine-graft-poly(ethylene glycol) and 2-(3-mercaptopropylsulfanyl)-ethyl-1,1-bisphosphonic acid (PEI-PEG-thiolBP) was prepared and used for surface coating of bovine serum albumin (BSA) nanoparticles (NPs) designed for bone-specific delivery of bone morphogenetic protein-2 (BMP-2). The NP coating was achieved with a dialysis and an evaporation method, and the obtained NPs were characterized by particle size, zeta-potential, morphology, and cytotoxicity in vitro. The particle size and surface charge of the NPs could be effectively tuned by the PEG and thiolBP substitution ratios of the conjugate, the coating method, and the polymer concentration used for coating. The PEG modification on PEI reduced the toxicity of PEI and the coated NPs, based on in vitro assessment with human C2C12 cells and rat bone marrow stromal cells. On the basis of an alkaline phosphatase (ALP) induction assay, the NP-encapsulated BMP-2 displayed full retention of its bioactivity, except for BMP-2 in PEI-coated NPs. By encapsulating (125)I-labeled BMP-2, the polymer-coated NPs were assessed for hydroxyapatite (HA) affinity; all NP-encapsulated BMP-2 showed significant affinity to HA as compared with free BMP-2 in vitro, and the PEI-PEG-thiolBP coated NPs improved the in vivo retention of BMP-2 compared with uncoated NPs. However, the biodistribution of NPs after intravenous injection in a rat model indicated no beneficial effects of thiolBP-coated NPs for bone targeting. Our results suggested that the BP-conjugated NPs are useful for localized delivery of BMP-2 in bone repair and regeneration, but they are not effective for bone targeting after intravenous administration.

Enhancement of healing in osteochondral defects by collagen sponge implants.

PubMed

Speer, D P; Chvapil, M; Volz, R G; Holmes, M D

1979-10-01

Implants of porous, highly cross-linked collagen sponge (CS) were tested for their capacity to enhance the healing of osteochondral defects in rabbits. Comparison was made to the healing of similar defects with polyvinyl alcohol sponge (PVAS) implants and with no implants (CONT). Evaluation was carried out up to 44 weeks following implantation and included observation of host cellular response, biodegradability of implant, gross appearance of restored joint surface, collagenous architecture of repair tissue, and properties of the junctions of implants and host articular cartilage, subchondral bone, and medullary bone. Collagen sponge proved most effective in promoting healing of osteochondral defects with fibrous and fibrocartilaginous tissue over restored subchondral bone. Collagen sponge showed many desirable properties as a potential material for biologic resurfacing of damaged joints. These properties included porosity, biodegradability, biocompatability, ability to mechanically protect cells and matrix while directing cell ingrowth, and an available chemical technology for modifying its biomechanical and biological properties. Comparative analysis of results of healing of CS, PVAS, and CONT osteochondral defects suggest rational design criteria for implant materials to improve their effectiveness in restoration of articular surfaces.

DC-STAMP Is an Osteoclast Fusogen Engaged in Periodontal Bone Resorption

PubMed Central

Wisitrasameewong, W.; Kajiya, M.; Movila, A.; Rittling, S.; Ishii, T.; Suzuki, M.; Matsuda, S.; Mazda, Y.; Torruella, M.R.; Azuma, M.M.; Egashira, K.; Freire, M.O.; Sasaki, H.; Wang, C.Y.; Han, X.; Taubman, M.A.; Kawai, T.

2017-01-01

Dendritic cell-specific transmembrane protein (DC-STAMP) plays a key role in the induction of osteoclast (OC) cell fusion, as well as DC-mediated immune regulation. While DC-STAMP gene expression is upregulated in the gingival tissue with periodontitis, its pathophysiological roles in periodontitis remain unclear. To evaluate the effects of DC-STAMP in periodontitis, anti-DC-STAMP–monoclonal antibody (mAb) was tested in a mouse model of ligature-induced periodontitis (n = 6–7/group) where Pasteurella pneumotropica (Pp)-reactive immune response activated T cells to produce receptor activator of nuclear factor kappa-B ligand (RANKL), which, in turn, promotes the periodontal bone loss via upregulation of osteoclastogenesis. DC-STAMP was expressed on the cell surface of mature multinuclear OCs, as well as immature mononuclear OCs, in primary cultures of RANKL-stimulated bone marrow cells. Anti-DC-STAMP-mAb suppressed the emergence of large, but not small, multinuclear OCs, suggesting that DC-STAMP is engaged in the late stage of cell fusion. Anti-DC-STAMP-mAb also inhibited pit formation caused by RANKL-stimulated bone marrow cells. Attachment of ligature to a second maxillary molar induced DC-STAMP messenger RNA and protein, along with elevated tartrate-resistant acid phosphatase–positive (TRAP+) OCs and alveolar bone loss. As we expected, systemic administration of anti-DC-STAMP-mAb downregulated the ligature-induced alveolar bone loss. Importantly, local injection of anti-DC-STAMP-mAb also suppressed alveolar bone loss and reduced the total number of multinucleated TRAP+ cells in mice that received ligature attachment. Attachment of ligature induced significantly elevated tumor necrosis factor–α, interleukin-1β, and RANKL in the gingival tissue compared with the control site without ligature (P < 0.05), which was unaffected by local injection with either anti-DC-STAMP-mAb or control-mAb. Neither in vivo anti-Pp IgG antibody nor in vitro anti-Pp T-cell response and resultant production of RANKL was affected by anti-DC-STAMP-mAb. This study illustrated the roles of DC-STAMP in promoting local OC cell fusion without affecting adaptive immune responses to oral bacteria. Therefore, it is plausible that a novel therapeutic regimen targeting DC-STAMP could suppress periodontal bone loss. PMID:28199142

A New In Vitro Model of Breast Cancer Metastasis to Bone

DTIC Science & Technology

2010-04-01

excessive cell vacuolization, ruffled cell margins , and increasing rates of cell-surface detachment. Thus, conventional tissue-culture strategies...determined. 3050 DHURJATI ET AL. apoptotic bodies, cytoplasmic vesiculation, and chromatin margination (Fig. 3) and a distinct loss of insoluble ECM...respond abnormally to various hor- mones and cytokines compared to normal, differen- tiated human osteoblasts. In effort to overcome these limitations

Biology and Clinical Relevance of Acute Myeloid Leukemia Stem Cells.

PubMed

Reinisch, Andreas; Chan, Steven M; Thomas, Daniel; Majeti, Ravindra

2015-07-01

Evidence for the cancer stem cell model was first demonstrated in xenotransplanted blood and bone marrow samples from patients with acute myeloid leukemia (AML) almost two decades ago, supporting the concept that a rare clonal and mutated leukemic stem cell (LSC) population is sufficient to drive leukemic growth. The inability to eliminate LSCs with conventional therapies is thought to be the primary cause of disease relapse in AML patients, and as such, novel therapies with the ability to target this population are required to improve patient outcomes. An important step towards this goal is the identification of common immunophenotypic surface markers and biological properties that distinguish LSCs from normal hematopoietic stem and progenitor cells (HSPCs) across AML patients. This work has resulted in the development of a large number of potential LSC-selective therapies that target cell surface molecules, intracellular signaling pathways, and the bone marrow microenvironment. Here, we will review the basic biology, immunophenotypic detection, and clinical relevance of LSCs, as well as emerging biological and small-molecule strategies that either directly target LSCs or indirectly target these cells through modulation of their microenvironment. Copyright © 2015 Elsevier Inc. All rights reserved.

Composite bone cements loaded with a bioactive and ferrimagnetic glass-ceramic: Leaching, bioactivity and cytocompatibility.

PubMed

Verné, Enrica; Bruno, Matteo; Miola, Marta; Maina, Giovanni; Bianco, Carlotta; Cochis, Andrea; Rimondini, Lia

2015-08-01

In this work, composite bone cements, based on a commercial polymethylmethacrylate matrix (Palamed®) loaded with ferrimagnetic bioactive glass-ceramic particles (SC45), were produced and characterized in vitro. The ferrimagnetic bioactive glass-ceramic belongs to the system SiO2-Na2O-CaO-P2O5-FeO-Fe2O3 and contains magnetite (Fe3O4) crystals into a residual amorphous bioactive phase. Three different formulations (containing 10, 15 and 20 wt.% of glass-ceramic particles respectively) have been investigated. These materials are intended to be applied as bone fillers for the hyperthermic treatment of bone tumors. The morphological, compositional, calorimetric and mechanical properties of each formulation have been already discussed in a previous paper. The in vitro properties of the composite bone cements described in the present paper are related to iron ion leaching test (by graphite furnace atomic absorption spectrometer), bioactivity (i.e. the ability to stimulate the formation of a hydroxyapatite - HAp - layer on their surface after soaking in simulated body fluid SBF) and cytocompatibility toward human osteosarcoma cells (ATCC CRL-1427, Mg63). Morphological and chemical characterizations by scanning electron microscopy and energy dispersion spectrometry have been performed on the composite samples after each test. The iron release was negligible and all the tested samples showed the growth of HAp on their surface after 28 days of immersion in a simulated body fluid (SBF). Cells showed good viability, morphology, adhesion, density and the ability to develop bridge-like structures on all investigated samples. A synergistic effect between bioactivity and cell mineralization was also evidenced. Copyright © 2015 Elsevier B.V. All rights reserved.

Chlorhexidine-loaded hydroxyapatite microspheres as an antimicrobial delivery system and its effect on in vivo osteo-conductive properties.

PubMed

Soriano-Souza, Carlos Alberto; Rossi, Andre L; Mavropoulos, Elena; Hausen, Moema A; Tanaka, Marcelo N; Calasans-Maia, Mônica D; Granjeiro, Jose M; Rocha-Leão, Maria Helena M; Rossi, Alexandre M

2015-04-01

Hydroxyapatite (HA) has been investigated as a delivery system for antimicrobial and antibacterial agents to simultaneously stimulate bone regeneration and prevent infection. Despite evidence supporting the bactericidal efficiency of these HA carriers, few studies have focused on the effect of this association on bone regeneration. In this work, we evaluated the physico-chemical properties of hydroxyapatite microspheres loaded with chlorhexidine (CHX) at two different concentrations, 0.9 and 9.1 μgCHX/cm2 HA, and characterized their effects on in vitro osteoblast viability and bone regeneration. Ultraviolet-visible spectroscopy, scanning and transmission electron microscopy associated with energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy were used to characterize the association of CHX and HA nanoparticles. The high CHX loading dose induced formation of organic CHX plate-like aggregates on the HA surface, whereas a Langmuir film was formed at the low CHX surface concentration. Quantitative evaluation of murine osteoblast viability parameters, including adhesion, mitochondrial activity and membrane integrity of cells exposed to HA/CHX extracts, revealed a cytotoxic effect for both loading concentrations. Histomorphological analysis upon implantation into the dorsal connective tissues and calvaria of rats for 7 and 42 days showed that the high CHX concentration induced the infiltration of inflammatory cells, resulting in retarded bone growth. Despite a strong decrease in in vitro cell viability, the low CHX loading dose did not impair the biocompatibility and osteoconductivity of HA during bone repair. These results indicate that high antimicrobial doses may activate a strong local inflammatory response and disrupt the long-term osteoconductive properties of CHX-HA delivery systems.

Biocompatible polymer coating of titania nanotube arrays for improved drug elution and osteoblast adhesion.

PubMed

Gulati, Karan; Ramakrishnan, Saminathan; Aw, Moom Sinn; Atkins, Gerald J; Findlay, David M; Losic, Dusan

2012-01-01

Bacterial infection, extensive inflammation and poor osseointegration have been identified as the major reasons for [early] orthopaedic implant failures based on titanium. Creating implants with drug-eluting properties to locally deliver drugs is an appealing way to address some of these problems. To improve properties of titanium for orthopaedic applications, this study explored the modification of titanium surfaces with titaniananotube (TNT) arrays, and approach that combines drug delivery into bone and potentially improved bone integration. A titania layer with an array of nanotube structures (∼120 nm in diameter and 50 μm in length) was synthesized on titanium surfaces by electrochemical anodization and loaded with the water-insoluble anti-inflammatory drug indomethacin. A simple dip-coating process of polymer modification formed thin biocompatible polymer films over the drug-loaded TNTs to create TNTs with predictable drug release characteristics. Two biodegradable and antibacterial polymers, chitosan and poly(lactic-co-glycolic acid), were tested for their ability to extend the drug release time of TNTs and produce favourable bone cell adhesion properties. Dependent on polymer thickness, a significant improvement in the drug release characteristics was demonstrated, with reduced burst release (from 77% to >20%) and extended overall release from 4 days to more than 30 days. Excellent osteoblast adhesion and cell proliferation on polymer-coated TNTs compared with uncoated TNTs were also observed. These results suggest that polymer-modified implants with a TNT layer are capable of delivering a drug to a bone site over an extended period and with predictable kinetics. In addition, favourable bone cell adhesion suggests that such an implant would have good biocompatibility. The described approach is broadly applicable to a wide range of drugs and implants currently used in orthopaedic practice. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

Macrophage cell lines derived from major histocompatibility complex II-negative mice

NASA Technical Reports Server (NTRS)

Beharka, A. A.; Armstrong, J. W.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

1998-01-01

Two bone-marrow-derived macrophage cell lines, C2D and C2Dt, were isolated from major histocompatibility class II negative knock-out mice. The C2D cell line was stabilized by continuous culture in colony-stimulating factor-1 and the C2Dt cell line was transformed with SV40 virus large T antigen. These cells exhibited phenotypic properties of macrophages including morphology and expression of Mac 1 and Mac 2 cell surface molecules. These cells also had comparable growth to the bone-marrow-derived macrophage cell line B6MP102. These new cell lines were not spontaneously cytotoxic and were only capable of modest killing of F5b tumor cells when stimulated with LPS and interferon-gamma, but not when stimulated with LPS alone or with staphylococcal exotoxin. C2D and C2Dt cells phagocytosed labeled Staphylococcus aureus similarly to B6MP102 cells but less well than C2D peritoneal macrophages. These cell lines secreted interleukin-6, but not tumor necrosis factor or nitric oxide in response to LPS or staphlococcal enterotoxins A or B C2D(t) cells were tumorigenic in C2D and C57BL/6J mice but C2D cells were not. These data suggest that macrophage cell lines can be established from bone marrow cells of major histocompatibility complex II-negative mice.

Defective CXCR4 expression in aged bone marrow cells impairs vascular regeneration

PubMed Central

Shao, Hongwei; Xu, Qiyuan; Wu, Qiuling; Ma, Qi; Salgueiro, Luis; Wang, Jian’An; Eton, Darwin; Webster, Keith A; Yu, Hong

2011-01-01

The chemokine stromal cell-derived factor-1 (SDF-1) plays a critical role in mobilizing precursor cells in the bone marrow and is essential for efficient vascular regeneration and repair. We recently reported that calcium augments the expression of chemokine receptor CXCR4 and enhances the angiogenic potential of bone marrow derived cells (BMCs). Neovascularization is impaired by aging therefore we suggested that aging may cause defects of CXCR4 expression and cellular responses to calcium. Indeed we found that both the basal and calcium-induced surface expression of CXCR4 on BMCs was significantly reduced in 25-month-old mice compared with 2-month-old mice. Reduced Ca-induced CXCR4 expression in BMC from aged mice was associated with defective calcium influx. Diminished CXCR4 surface expression in BMC from aged mice correlated with diminished neovascularization in an ischemic hindlimb model with less accumulation of CD34+ progenitor cells in the ischemic muscle with or without local overexpression of SDF-1. Intravenous injection of BMCs from old mice homed less efficiently to ischemic muscle and stimulated significantly less neovascularization compared with the BMCs from young mice. Transplantation of old BMCs into young mice did not reconstitute CXCR4 functions suggesting that the defects were not reversible by changing the environment. We conclude that defects of basal and calcium-regulated functions of the CXCR4/SDF-1 axis in BMCs contribute significantly to the age-related loss of vasculogenic responses. PMID:21143386

In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading

PubMed Central

Jing, Da; Baik, Andrew D.; Lu, X. Lucas; Zhou, Bin; Lai, Xiaohan; Wang, Liyun; Luo, Erping; Guo, X. Edward

2014-01-01

Osteocytes have been hypothesized to be the major mechanosensors in bone. How in situ osteocytes respond to mechanical stimuli is still unclear because of technical difficulties. In vitro studies have shown that osteocytes exhibited unique calcium (Ca2+) oscillations to fluid shear. However, whether this mechanotransduction phenomenon holds for in situ osteocytes embedded within a mineralized bone matrix under dynamic loading remains unknown. Using a novel synchronized loading/imaging technique, we successfully visualized in real time and quantified Ca2+ responses in osteocytes and bone surface cells in situ under controlled dynamic loading on intact mouse tibia. The resultant fluid-induced shear stress on the osteocyte in the lacunocanalicular system (LCS) was also quantified. Osteocytes, but not surface cells, displayed repetitive Ca2+ spikes in response to dynamic loading, with spike frequency and magnitude dependent on load magnitude, tissue strain, and shear stress in the LCS. The Ca2+ oscillations were significantly reduced by endoplasmic reticulum (ER) depletion and P2 purinergic receptor (P2R)/phospholipase C (PLC) inhibition. This study provides direct evidence that osteocytes respond to in situ mechanical loading by Ca2+ oscillations, which are dependent on the P2R/PLC/inositol trisphosphate/ER pathway. This study develops a novel approach in skeletal mechanobiology and also advances our fundamental knowledge of bone mechanotransduction.—Jing, D., Baik, A. D., Lu, X. L., Zhou, B., Lai, X., Wang, L., Luo, E., Guo, X. E. In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading. PMID:24347610

Multiphasic Scaffolds for Periodontal Tissue Engineering

PubMed Central

Ivanovski, S.; Vaquette, C.; Gronthos, S.; Hutmacher, D.W.; Bartold, P.M.

2014-01-01

For a successful clinical outcome, periodontal regeneration requires the coordinated response of multiple soft and hard tissues (periodontal ligament, gingiva, cementum, and bone) during the wound-healing process. Tissue-engineered constructs for regeneration of the periodontium must be of a complex 3-dimensional shape and adequate size and demonstrate biomechanical stability over time. A critical requirement is the ability to promote the formation of functional periodontal attachment between regenerated alveolar bone, and newly formed cementum on the root surface. This review outlines the current advances in multiphasic scaffold fabrication and how these scaffolds can be combined with cell- and growth factor–based approaches to form tissue-engineered constructs capable of recapitulating the complex temporal and spatial wound-healing events that will lead to predictable periodontal regeneration. This can be achieved through a variety of approaches, with promising strategies characterized by the use of scaffolds that can deliver and stabilize cells capable of cementogenesis onto the root surface, provide biomechanical cues that encourage perpendicular alignment of periodontal fibers to the root surface, and provide osteogenic cues and appropriate space to facilitate bone regeneration. Progress on the development of multiphasic constructs for periodontal tissue engineering is in the early stages of development, and these constructs need to be tested in large animal models and, ultimately, human clinical trials. PMID:25139362

Multiphasic scaffolds for periodontal tissue engineering.

PubMed

Ivanovski, S; Vaquette, C; Gronthos, S; Hutmacher, D W; Bartold, P M

2014-12-01

For a successful clinical outcome, periodontal regeneration requires the coordinated response of multiple soft and hard tissues (periodontal ligament, gingiva, cementum, and bone) during the wound-healing process. Tissue-engineered constructs for regeneration of the periodontium must be of a complex 3-dimensional shape and adequate size and demonstrate biomechanical stability over time. A critical requirement is the ability to promote the formation of functional periodontal attachment between regenerated alveolar bone, and newly formed cementum on the root surface. This review outlines the current advances in multiphasic scaffold fabrication and how these scaffolds can be combined with cell- and growth factor-based approaches to form tissue-engineered constructs capable of recapitulating the complex temporal and spatial wound-healing events that will lead to predictable periodontal regeneration. This can be achieved through a variety of approaches, with promising strategies characterized by the use of scaffolds that can deliver and stabilize cells capable of cementogenesis onto the root surface, provide biomechanical cues that encourage perpendicular alignment of periodontal fibers to the root surface, and provide osteogenic cues and appropriate space to facilitate bone regeneration. Progress on the development of multiphasic constructs for periodontal tissue engineering is in the early stages of development, and these constructs need to be tested in large animal models and, ultimately, human clinical trials. © International & American Associations for Dental Research.

Structural and surface property characterization of titanium dioxide nanotubes for orthopedic implants

NASA Astrophysics Data System (ADS)

Shokuhfar, Tolou

This research focused on the to modification of the surface structure of titanium implants with nanostructured morphology of TiO2 nanotubes and studied the interaction of nanotubes with osteoblast cells to understand the parameters that affect the cell growth. The electrical, mechanical, and structural properties of TiO2 nanotubes were characterized to establish a better understanding on the properties of such nanoscale morphological structures. To achieve the objectives of this research work I transformed the titanium and its alloys, either in bulk sheet form, bulk machined form, or thin film deposited on another substrate into a surface of titania nanotubes using a low cost and environmentally friendly process. The process requires only a simple electrolyte, low cost electrode, and a DC power supply. With this simple approach of scalable nanofabrication, a typical result is nanotubes that are each approximately 100nm in diameter and have a wall thickness of about 20nm. By changing the fabrication parameters, independent nanotubes can be fabricated with open volume between them. Titanium in this form is termed onedimensional since electron transport is narrowly confined along the length of the nanotube. My Ph.D. accomplishments have successfully shown that osteoblast cells, the cells that are the precursors to bone, have a strong tendency to attach to the inside and outside of the titanium nanotubes onto which they are grown using their filopodia -- cell's foot used for locomotion -- anchored to titanium nanotubes. In fact it was shown that the cell prefers to find many anchoring sites. These sites are critical for cell locomotion during the first several weeks of maturity and upon calcification as a strongly anchored bone cell. In addition I have shown that such a surface has a greater cell density than a smooth titanium surface. My work also developed a process that uses a focused and controllably rastered ion beam as a nano-scalpel to cut away sections of the osteoblast cells to probe the attachment beneath the main cell body. Ultimately the more rapid growth of osteoblasts, coupled with a stronger cell-surface interface, could provide cost reduction, shorter rehabilitation, and fewer follow-on surgeries due to implant loosening.

Vesicular delivery of crystalline calcium minerals to ECM in biomineralized nanoclay composites

NASA Astrophysics Data System (ADS)

Katti, Kalpana S.; Ambre, Avinash H.; Payne, Scott; Katti, Dinesh R.

2015-04-01

The mechanisms of mineralization and new bone formation were explored in newly formed extracellular matrix in a nanoclay based composite. Nanoclay films were prepared by intercalating the clays with amino acids and using the amino acids for mineralization of hydroxyapatite. The biomineralized hydroxyapatite (HAP) inside nanoclay galleries or in situ HAP/clay was further used to make films (substrates) using polycaprolactone (PCL) that were seeded with mesenchymal stem cells in a two-stage seeding process. SEM imaging experiments performed on PCL/in situ HAPclay composite films seeded with human MSCs indicated formation of matrix vesicles. The vesicles appear to emerge from the cells that are adhered to the nanoclay HAP films and also deposited in the extracellular space. Vesicles are also observed to be embedded in the cells or under the surface of cells. Crystalline structures with Ca and P were found inside vesicles. The Ca/P ratios obtained using energy dispersive spectroscopy indicate values ranging from below 0.7 to the stoichiometric HAP value of 1.67. The Ca/P ratios were obtained to be closer to the stoichiometric value for single seeding experiments as compared to the double seeding experiments indicating more new bone formation in double seeding experiments. New bone formation with bone mimetic mineralization is thus observed on the in situ HAP nanoclay PCL samples. Hence the PCL/in situ HAPclay composites besides being osteoinductive are also capable of providing a favorable micro-environment for cell dependent processes involved in bone mineral formation.