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Sample records for human bone collagen

  1. Glycation Contributes to Interaction Between Human Bone Alkaline Phosphatase and Collagen Type I.

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    Halling Linder, Cecilia; Enander, Karin; Magnusson, Per

    2016-03-01

    Bone is a biological composite material comprised primarily of collagen type I and mineral crystals of calcium and phosphate in the form of hydroxyapatite (HA), which together provide its mechanical properties. Bone alkaline phosphatase (ALP), produced by osteoblasts, plays a pivotal role in the mineralization process. Affinity contacts between collagen, mainly type II, and the crown domain of various ALP isozymes were reported in a few in vitro studies in the 1980s and 1990s, but have not attracted much attention since, although such interactions may have important implications for the bone mineralization process. The objective of this study was to investigate the binding properties of human collagen type I to human bone ALP, including the two bone ALP isoforms B1 and B2. ALP from human liver, human placenta and E. coli were also studied. A surface plasmon resonance-based analysis, supported by electrophoresis and blotting, showed that bone ALP binds stronger to collagen type I in comparison with ALPs expressed in non-mineralizing tissues. Further, the B2 isoform binds significantly stronger to collagen type I in comparison with the B1 isoform. Human bone and liver ALP (with identical amino acid composition) displayed pronounced differences in binding, revealing that post-translational glycosylation properties govern these interactions to a large extent. In conclusion, this study presents the first evidence that glycosylation differences in human ALPs are of crucial importance for protein-protein interactions with collagen type I, although the presence of the ALP crown domain may also be necessary. Different binding affinities among the bone ALP isoforms may influence the mineral-collagen interface, mineralization kinetics, and degree of bone matrix mineralization, which are important factors determining the material properties of bone.

  2. Response of induced bone defects in horses to collagen matrix containing the human parathyroid hormone gene.

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    Backstrom, Kristin C; Bertone, Alicia L; Wisner, Erik R; Weisbrode, Stephen E

    2004-09-01

    To determine whether human parathyroid hormone (hPTH) gene in collagen matrix could safely promote bone formation in diaphyseal or subchondral bones of horses. 8 clinically normal adult horses. Amount, rate, and quality of bone healing for 13 weeks were determined by use of radiography, quantitative computed tomography, and histomorphometric analysis. Diaphyseal cortex and subchondral bone defects of metacarpi were filled with hPTH(1-34) gene-activated matrix (GAM) or remained untreated. Joints were assessed on the basis of circumference, synovial fluid analysis, pain on flexion, lameness, and gross and histologic examination. Bone volume index was greater for cortical defects treated with hPTH(1-34) GAM, compared with untreated defects. Bone production in cortical defects treated with hPTH(1-34) GAM positively correlated with native bone formation in untreated defects. In contrast, less bone was detected in hPTH(1-34) GAM-treated subchondral bone defects, compared with untreated defects, and histology confirmed poorer healing and residual collagen sponge. Use of hPTH(1-34) GAM induced greater total bone, specifically periosteal bone, after 13 weeks of healing in cortical defects of horses. The hPTH(1-34) GAM impeded healing of subchondral bone but was biocompatible with joint tissues. Promotion of periosteal bone formation may be beneficial for healing of cortical fractures in horses, but the delay in onset of bone formation may negate benefits. The hPTH(1-34) GAM used in this study should not be placed in articular subchondral bone defects, but contact with articular surfaces is unlikely to cause short-term adverse effects.

  3. Rapid biomimetic mineralization of collagen fibrils and combining with human umbilical cord mesenchymal stem cells for bone defects healing

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    Ye, Bihua; Luo, Xueshi; Li, Zhiwen [Department of Material Science and Engineering, Engineering Research Center of Artificial Organs and Materials, Jinan University, Guangzhou 510632 (China); Zhuang, Caiping [Department of Anesthesiology, Huizhou Central People' s Hospital, Huizhou 516001 (China); Li, Lihua, E-mail: tlihuali@jnu.edu.cn [Department of Material Science and Engineering, Engineering Research Center of Artificial Organs and Materials, Jinan University, Guangzhou 510632 (China); Lu, Lu; Ding, Shan; Tian, Jinhuan [Department of Material Science and Engineering, Engineering Research Center of Artificial Organs and Materials, Jinan University, Guangzhou 510632 (China); Zhou, Changren, E-mail: tcrz9@jnu.edu.cn [Department of Material Science and Engineering, Engineering Research Center of Artificial Organs and Materials, Jinan University, Guangzhou 510632 (China)

    2016-11-01

    Collagen biomineralization is regulated by complicated interactions between the collagen matrix and non-collagenous extracellular proteins. Here, the use of sodium tripolyphosphate to simulate the templating functional motif of the C-terminal fragment of non-collagenous proteins is reported, and a low molecular weight polyacrylic acid served as a sequestration agent to stabilize amorphous calcium phosphate into nanoprecursors. Self-assembled collagen fibrils served as a fixed template for achieving rapid biomimetic mineralization in vitro. Results demonstrated that, during the mineralization process, intrafibrillar and extrafibrillar hydroxyapatite mineral with collagen fibrils formed and did so via bottom-up nanoparticle assembly based on the non-classical crystallization approach in the presence of these dual biomimetic functional analogues. In vitro human umbilical cord mesenchymal stem cell (hUCMSC) culture found that the mineralized scaffolds have a better cytocompatibility in terms of cell viability, adhesion, proliferation, and differentiation into osteoblasts. A rabbit femoral condyle defect model was established to confirm the ability of the n-HA/collagen scaffolds to facilitate bone regeneration and repair. The images of gross anatomy, MRI, CT and histomorphology taken 6 and 12 weeks after surgery showed that the biomimetic mineralized collagen scaffolds with hUCMSCs can promote the healing speed of bone defects in vivo, and both of the scaffolds groups performing better than the bone defect control group. As new bone tissue formed, the scaffolds degraded and were gradually absorbed. All these results demonstrated that both of the scaffolds and cells have better histocompatibility. - Highlights: • A rapid and facile biomimetic mineralization approach is proposed. • Intrafibrillar and extrafibrillar mineralization of collagen fibrils was achieved. • HA/COL scaffolds promote hUCMSCs adhesion, proliferation, and differentiation. • Feasibility of h

  4. Rapid biomimetic mineralization of collagen fibrils and combining with human umbilical cord mesenchymal stem cells for bone defects healing

    International Nuclear Information System (INIS)

    Ye, Bihua; Luo, Xueshi; Li, Zhiwen; Zhuang, Caiping; Li, Lihua; Lu, Lu; Ding, Shan; Tian, Jinhuan; Zhou, Changren

    2016-01-01

    Collagen biomineralization is regulated by complicated interactions between the collagen matrix and non-collagenous extracellular proteins. Here, the use of sodium tripolyphosphate to simulate the templating functional motif of the C-terminal fragment of non-collagenous proteins is reported, and a low molecular weight polyacrylic acid served as a sequestration agent to stabilize amorphous calcium phosphate into nanoprecursors. Self-assembled collagen fibrils served as a fixed template for achieving rapid biomimetic mineralization in vitro. Results demonstrated that, during the mineralization process, intrafibrillar and extrafibrillar hydroxyapatite mineral with collagen fibrils formed and did so via bottom-up nanoparticle assembly based on the non-classical crystallization approach in the presence of these dual biomimetic functional analogues. In vitro human umbilical cord mesenchymal stem cell (hUCMSC) culture found that the mineralized scaffolds have a better cytocompatibility in terms of cell viability, adhesion, proliferation, and differentiation into osteoblasts. A rabbit femoral condyle defect model was established to confirm the ability of the n-HA/collagen scaffolds to facilitate bone regeneration and repair. The images of gross anatomy, MRI, CT and histomorphology taken 6 and 12 weeks after surgery showed that the biomimetic mineralized collagen scaffolds with hUCMSCs can promote the healing speed of bone defects in vivo, and both of the scaffolds groups performing better than the bone defect control group. As new bone tissue formed, the scaffolds degraded and were gradually absorbed. All these results demonstrated that both of the scaffolds and cells have better histocompatibility. - Highlights: • A rapid and facile biomimetic mineralization approach is proposed. • Intrafibrillar and extrafibrillar mineralization of collagen fibrils was achieved. • HA/COL scaffolds promote hUCMSCs adhesion, proliferation, and differentiation. • Feasibility of h

  5. Ectopic bone formation in nude rats using human osteoblasts seeded poly(3)hydroxybutyrate embroidery and hydroxyapatite-collagen tapes constructs.

    Science.gov (United States)

    Mai, Ronald; Hagedorn, Manolo Gunnar; Gelinsky, Michael; Werner, Carsten; Turhani, Dritan; Späth, Heike; Gedrange, Tomas; Lauer, Günter

    2006-09-01

    The aim of this study was to evaluate the ectopic bone formation using tissue engineered cell-seeded constructs with two different scaffolds and primary human maxillary osteoblasts in nude rats over an implantation period of up to 96 days. Collagen I-coated Poly(3)hydroxybutyrate (PHB) embroidery and hydroxyapatite (HAP) collagen tapes were seeded with primary human maxillary osteoblasts (hOB) and implanted into athymic rnu/run rats. A total of 72 implants were placed into the back muscles of 18 rats. 24, 48 and 96 days after implantation, histological and histomorphometric analyses were made. The osteoblastic character of the cells was confirmed by immunocytochemistry and RT-PCR for osteocalcin. Histological analysis demonstrated that all cell-seeded constructs induced ectopic bone formation after 24, 48 and 96 days of implantation. There was more mineralized tissue in PHB constructs than in HAP-collagen tapes (at day 24; p embroidery or HAP-collagen tapes can induce ectopic bone formation. However, the amount of bone formed decreased with increasing length of implantation.

  6. Increased cartilage type II collagen degradation in patients with osteogenesis imperfecta used as a human model of bone type I collagen alterations.

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    Rousseau, Jean-Charles; Chevrel, Guillaume; Schott, Anne-Marie; Garnero, Patrick

    2010-04-01

    We investigated whether cartilage degradation is altered in adult patients with mild osteogenesis imperfecta (OI) used as a human model of bone type I collagen-related osteoarthritis (OA). Sixty-four adult patients with OI (39% women, mean age+/-SD: 37+/-12 years) and 64 healthy age-matched controls (54% women, 39+/-7 years) were included. We also compared data in 87 patients with knee OA (73% women, 63+/-8 years, mean disease duration: 6 years) and 291 age-matched controls (80% women, 62+/-10 years). Urinary C-terminal cross-linked telopeptide of type II collagen (CTX-II), a marker of cartilage degradation, urinary helical peptide of type I collagen (Helix-I), a marker of bone resorption, and the urinary ratio between non-isomerised/isomerised (alpha/beta CTX-I) type I collagen C-telopeptide, a marker of type I collagen maturation, were measured. Patients with OI had CTX-II levels similar to those of subjects with knee OA (p=0.89; mean+/-SEM; 460+/-57 ng/mmol Cr for OI group and 547+/-32 ng/mmol Cr for OA group) and significantly higher than both young (144+/-7.8 ng/mmol Cr, p<0.0001) and old controls (247+/-7 ng/mmol Cr, p<0.0001). In patients with OI, increased Helix-I (p<0.0001) and alpha/beta CTX-I (p=0.0067) were independently associated with increased CTX-II and together explained 26% of its variance (p< 0.0001). In patients with knee OA, increased levels of alpha/beta CTX-I ratio were also associated with higher CTX-II levels. Adult patients with OI or knee OA are characterized by increased cartilage type II collagen degradation, which is associated with increased type I collagen degradation for OI and lower type I collagen maturation for both OI and OA. These data suggest that both quantitative and qualitative alterations of bone type I collagen metabolism are involved in increased cartilage degradation in patients with OI or knee OA. Copyright 2009 Elsevier Inc. All rights reserved.

  7. Normal Collagen and Bone Production by Gene-targeted Human Osteogenesis Imperfecta iPSCs

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    Deyle, David R; Khan, Iram F; Ren, Gaoying; Wang, Pei-Rong; Kho, Jordan; Schwarze, Ulrike; Russell, David W

    2012-01-01

    Osteogenesis imperfecta (OI) is caused by dominant mutations in the type I collagen genes. In principle, the skeletal abnormalities of OI could be treated by transplantation of patient-specific, bone-forming cells that no longer express the mutant gene. Here, we develop this approach by isolating mesenchymal cells from OI patients, inactivating their mutant collagen genes by adeno-associated virus (AAV)-mediated gene targeting, and deriving induced pluripotent stem cells (iPSCs) that were expanded and differentiated into mesenchymal stem cells (iMSCs). Gene-targeted iMSCs produced normal collagen and formed bone in vivo, but were less senescent and proliferated more than bone-derived MSCs. To generate iPSCs that would be more appropriate for clinical use, the reprogramming and selectable marker transgenes were removed by Cre recombinase. These results demonstrate that the combination of gene targeting and iPSC derivation can be used to produce potentially therapeutic cells from patients with genetic disease. PMID:22031238

  8. Age-related effect on the concentration of collagen crosslinks in human osteonal and interstitial bone tissue.

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    Nyman, Jeffry S; Roy, Anuradha; Acuna, Rae L; Gayle, Heather J; Reyes, Michael J; Tyler, Jerrod H; Dean, David D; Wang, Xiaodu

    2006-12-01

    Collagen crosslinks are important to the quality of bone and may be contributors to the age-related increase in bone fracture. This study was performed to investigate whether age and gender effects on collagen crosslinks are similar in osteonal and interstitial bone tissues. Forty human cadaveric femurs were collected and divided into two age groups: middle-aged (42-63 years of age) and elderly (69-90 years of age) with ten males and ten females in each group (n = 10). Micro-cores of bone tissue from both secondary osteons and interstitial regions in the medial quadrant of the diaphysis were extracted using a custom-modified, computer-controlled milling machine. The bone specimens were then analyzed using high performance liquid chromatography to determine the effects of age and gender on the concentration of mature, enzymatic crosslinks (hydroxylysyl-pyridinoline-HP and lysyl-pyridinoline-LP) and a non-enzymatic crosslink (pentosidine-PE) at these two microstructural sites. The results indicate that age has a significant effect on the concentration of LP and PE, while gender has a significant effect on HP and LP. In addition, the concentration of the crosslinks in the secondary osteons is significantly different from that in the interstitial bone regions. These results suggest that the amount of non-enzymatic crosslinking may increase while that of mature enzymatic crosslinking may decrease with age. Such changes could potentially reduce the inherent quality of the bone tissue in the elderly skeleton.

  9. Monomeric, porous type II collagen scaffolds promote chondrogenic differentiation of human bone marrow mesenchymal stem cells in vitro

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    Tamaddon, M.; Burrows, M.; Ferreira, S. A.; Dazzi, F.; Apperley, J. F.; Bradshaw, A.; Brand, D. D.; Czernuszka, J.; Gentleman, E.

    2017-03-01

    Osteoarthritis (OA) is a common cause of pain and disability and is often associated with the degeneration of articular cartilage. Lesions to the articular surface, which are thought to progress to OA, have the potential to be repaired using tissue engineering strategies; however, it remains challenging to instruct cell differentiation within a scaffold to produce tissue with appropriate structural, chemical and mechanical properties. We aimed to address this by driving progenitor cells to adopt a chondrogenic phenotype through the tailoring of scaffold composition and physical properties. Monomeric type-I and type-II collagen scaffolds, which avoid potential immunogenicity associated with fibrillar collagens, were fabricated with and without chondroitin sulfate (CS) and their ability to stimulate the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells was assessed. Immunohistochemical analyses showed that cells produced abundant collagen type-II on type-II scaffolds and collagen type-I on type-I scaffolds. Gene expression analyses indicated that the addition of CS - which was released from scaffolds quickly - significantly upregulated expression of type II collagen, compared to type-I and pure type-II scaffolds. We conclude that collagen type-II and CS can be used to promote a more chondrogenic phenotype in the absence of growth factors, potentially providing an eventual therapy to prevent OA.

  10. Human mandible bone defect repair by the grafting of dental pulp stem/progenitor cells and collagen sponge biocomplexes

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    R d’Aquino

    2009-11-01

    Full Text Available In this study we used a biocomplex constructed from dental pulp stem/progenitor cells (DPCs and a collagen sponge scaffold for oro-maxillo-facial (OMF bone tissue repair in patients requiring extraction of their third molars. The experiments were carried out according to our Internal Ethical Committee Guidelines and written informed consent was obtained from the patients. The patients presented with bilateral bone reabsorption of the alveolar ridge distal to the second molar secondary to impaction of the third molar on the cortical alveolar lamina, producing a defect without walls, of at least 1.5 cm in height. This clinical condition does not permit spontaneous bone repair after extraction of the third molar, and eventually leads to loss also of the adjacent second molar. Maxillary third molars were extracted first for DPC isolation and expansion. The cells were then seeded onto a collagen sponge scaffold and the obtained biocomplex was used to fill in the injury site left by extraction of the mandibular third molars. Three months after autologous DPC grafting, alveolar bone of patients had optimal vertical repair and complete restoration of periodontal tissue back to the second molars, as assessed by clinical probing and X-rays. Histological observations clearly demonstrated the complete regeneration of bone at the injury site. Optimal bone regeneration was evident one year after grafting. This clinical study demonstrates that a DPC/collagen sponge biocomplex can completely restore human mandible bone defects and indicates that this cell population could be used for the repair and/or regeneration of tissues and organs.

  11. High-strength mineralized collagen artificial bone

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    Qiu, Zhi-Ye; Tao, Chun-Sheng; Cui, Helen; Wang, Chang-Ming; Cui, Fu-Zhai

    2014-03-01

    Mineralized collagen (MC) is a biomimetic material that mimics natural bone matrix in terms of both chemical composition and microstructure. The biomimetic MC possesses good biocompatibility and osteogenic activity, and is capable of guiding bone regeneration as being used for bone defect repair. However, mechanical strength of existing MC artificial bone is too low to provide effective support at human load-bearing sites, so it can only be used for the repair at non-load-bearing sites, such as bone defect filling, bone graft augmentation, and so on. In the present study, a high strength MC artificial bone material was developed by using collagen as the template for the biomimetic mineralization of the calcium phosphate, and then followed by a cold compression molding process with a certain pressure. The appearance and density of the dense MC were similar to those of natural cortical bone, and the phase composition was in conformity with that of animal's cortical bone demonstrated by XRD. Mechanical properties were tested and results showed that the compressive strength was comparable to human cortical bone, while the compressive modulus was as low as human cancellous bone. Such high strength was able to provide effective mechanical support for bone defect repair at human load-bearing sites, and the low compressive modulus can help avoid stress shielding in the application of bone regeneration. Both in vitro cell experiments and in vivo implantation assay demonstrated good biocompatibility of the material, and in vivo stability evaluation indicated that this high-strength MC artificial bone could provide long-term effective mechanical support at human load-bearing sites.

  12. Human Bone Derived Collagen for the Development of an Artificial Corneal Endothelial Graft. In Vivo Results in a Rabbit Model.

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    Natalia Vázquez

    Full Text Available Corneal keratoplasty (penetrating or lamellar using cadaveric human tissue, is nowadays the main treatment for corneal endotelial dysfunctions. However, there is a worldwide shortage of donor corneas available for transplantation and about 53% of the world's population have no access to corneal transplantation. Generating a complete cornea by tissue engineering is still a tough goal, but an endothelial lamellar graft might be an easier task. In this study, we developed a tissue engineered corneal endothelium by culturing human corneal endothelial cells on a human purified type I collagen membrane. Human corneal endothelial cells were cultured from corneal rims after corneal penetrating keratoplasty and type I collagen was isolated from remnant cancellous bone chips. Isolated type I collagen was analyzed by western blot, liquid chromatography -mass spectrometry and quantified using the exponentially modified protein abundance index. Later on, collagen solution was casted at room temperature obtaining an optically transparent and mechanically manageable membrane that supports the growth of human and rabbit corneal endothelial cells which expressed characteristic markers of corneal endothelium: zonula ocluddens-1 and Na+/K+ ATPase. To evaluate the therapeutic efficiency of our artificial endothelial grafts, human purified type I collagen membranes cultured with rabbit corneal endothelial cells were transplanted in New Zealand white rabbits that were kept under a minimal immunosuppression regimen. Transplanted corneas maintained transparency for as long as 6 weeks without obvious edema or immune rejection and maintaining the same endothelial markers that in a healthy cornea. In conclusion, it is possible to develop an artificial human corneal endothelial graft using remnant tissues that are not employed in transplant procedures. This artificial endothelial graft can restore the integrality of corneal endothelium in an experimental model of

  13. Human bone marrow mesenchymal stem cells induce collagen production and tongue cancer invasion.

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    Sirpa Salo

    Full Text Available Tumor microenvironment (TME is an active player in carcinogenesis and changes in its composition modify cancer growth. Carcinoma-associated fibroblasts, bone marrow-derived multipotent mesenchymal stem cells (BMMSCs, and inflammatory cells can all affect the composition of TME leading to changes in proliferation, invasion and metastasis formation of carcinoma cells. In this study, we confirmed an interaction between BMMSCs and oral tongue squamous cell carcinoma (OTSCC cells by analyzing the invasion progression and gene expression pattern. In a 3-dimensional myoma organotypic invasion model the presence of BMMSCs inhibited the proliferation but increased the invasion of OTSCC cells. Furthermore, the signals originating from OTSCC cells up-regulated the expression of inflammatory chemokines by BMMSCs, whereas BMMSC products induced the expression of known invasion linked molecules by carcinoma cells. Particularly, after the cell-cell interactions, the chemokine CCL5 was abundantly secreted from BMMSCs and a function blocking antibody against CCL5 inhibited BMMSC enhanced cancer invasion area. However, CCL5 blocking antibody did not inhibit the depth of invasion. Additionally, after exposure to BMMSCs, the expression of type I collagen mRNA in OTSCC cells was markedly up-regulated. Interestingly, also high expression of type I collagen N-terminal propeptide (PINP in vivo correlated with the cancer-specific mortality of OTSCC patients, whereas there was no association between cancer tissue CCL5 levels and the clinical parameters. In conclusion, our results suggest that the interaction between BMMSC and carcinoma cells induce cytokine and matrix molecule expression, of which high level of type I collagen production correlates with the prognosis of OTSCC patients.

  14. Biomimetic soluble collagen purified from bones.

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    Ferreira, Ana Marina; Gentile, Piergiorgio; Sartori, Susanna; Pagliano, Cristina; Cabrele, Chiara; Chiono, Valeria; Ciardelli, Gianluca

    2012-11-01

    Type I collagen has been extensively exploited as a biomaterial for biomedical applications and drug delivery; however, small molecular alterations occurring during the isolation procedure and its interaction with residual bone extracellular matrix molecules or proteins might affect the overall material biocompatibility and performance. The aim of the current work is to study the potential alterations in collagen properties and organization associated with the absence of proteoglycans, which mimic pathological conditions associated with age-related diseases. A new approach for evaluating the effect of proteoglycans on the properties of isolated type I collagen from the bone matrix is described. Additional treatment with guanidine hydrochloride was introduced to remove residual proteoglycans from the collagen matrix. The properties of the isolated collagen with/without guanidine hydrochloride treatment were investigated and compared with a commercial rabbit collagen as control. We demonstrate that the absence of proteoglycans in the isolated type I collagen affects its thermal properties, the extraction into its native structure, and its ability to hydrate and self-assemble into fibers. The fine control and tuning of all these features, linked to the absence of non-collagenous proteins as proteoglycans, offer the possibility of designing new strategies and biomaterials with advanced biomimetic properties aimed at regenerating bone tissue in the case of fragility and/or defects. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Distinct characteristics of mandibular bone collagen relative to long bone collagen: relevance to clinical dentistry.

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    Matsuura, Takashi; Tokutomi, Kentaro; Sasaki, Michiko; Katafuchi, Michitsuna; Mizumachi, Emiri; Sato, Hironobu

    2014-01-01

    Bone undergoes constant remodeling throughout life. The cellular and biochemical mechanisms of bone remodeling vary in a region-specific manner. There are a number of notable differences between the mandible and long bones, including developmental origin, osteogenic potential of mesenchymal stem cells, and the rate of bone turnover. Collagen, the most abundant matrix protein in bone, is responsible for determining the relative strength of particular bones. Posttranslational modifications of collagen, such as intermolecular crosslinking and lysine hydroxylation, are the most essential determinants of bone strength, although the amount of collagen is also important. In comparison to long bones, the mandible has greater collagen content, a lower amount of mature crosslinks, and a lower extent of lysine hydroxylation. The great abundance of immature crosslinks in mandibular collagen suggests that there is a lower rate of cross-link maturation. This means that mandibular collagen is relatively immature and thus more readily undergoes degradation and turnover. The greater rate of remodeling in mandibular collagen likely renders more flexibility to the bone and leaves it more suited to constant exercise. As reviewed here, it is important in clinical dentistry to understand the distinctive features of the bones of the jaw.

  16. Distinct Characteristics of Mandibular Bone Collagen Relative to Long Bone Collagen: Relevance to Clinical Dentistry

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    Takashi Matsuura

    2014-01-01

    Full Text Available Bone undergoes constant remodeling throughout life. The cellular and biochemical mechanisms of bone remodeling vary in a region-specific manner. There are a number of notable differences between the mandible and long bones, including developmental origin, osteogenic potential of mesenchymal stem cells, and the rate of bone turnover. Collagen, the most abundant matrix protein in bone, is responsible for determining the relative strength of particular bones. Posttranslational modifications of collagen, such as intermolecular crosslinking and lysine hydroxylation, are the most essential determinants of bone strength, although the amount of collagen is also important. In comparison to long bones, the mandible has greater collagen content, a lower amount of mature crosslinks, and a lower extent of lysine hydroxylation. The great abundance of immature crosslinks in mandibular collagen suggests that there is a lower rate of cross-link maturation. This means that mandibular collagen is relatively immature and thus more readily undergoes degradation and turnover. The greater rate of remodeling in mandibular collagen likely renders more flexibility to the bone and leaves it more suited to constant exercise. As reviewed here, it is important in clinical dentistry to understand the distinctive features of the bones of the jaw.

  17. Raman spectral markers of collagen denaturation and hydration in human cortical bone tissue are affected by radiation sterilization and high cycle fatigue damage.

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    Flanagan, Christopher D; Unal, Mustafa; Akkus, Ozan; Rimnac, Clare M

    2017-11-01

    Thermal denaturation and monotonic mechanical damage alter the organic and water-related compartments of cortical bone. These changes can be detected using Raman spectroscopy. However, less is known regarding Raman sensitivity to detect the effects of cyclic fatigue damage and allograft sterilization doses of gamma radiation. To determine if Raman spectroscopic biomarkers of collagen denaturation and hydration are sensitive to the effects of (a) high cycle fatigue damage and (b) 25kGy irradiation. Unirradiated and gamma-radiation sterilized human cortical bone specimens previously tested in vitro under high-cycle (> 100,000 cycles) fatigue conditions at 15MPa, 25MPa, 35MPa, 45MPa, and 55MPa cyclic stress levels were studied. Cortical bone Raman spectral profiles from wavenumber ranges of 800-1750cm -1 and 2700-3800cm -1 were obtained and compared from: a) non-fatigue vs fatigue fracture sites and b) radiated vs. unirradiated states. Raman biomarker ratios 1670/1640 and 3220/2949, which reflect collagen denaturation and organic matrix (mainly collagen)-bound water, respectively, were assessed. One- and two-way ANOVA analyses were utilized to identify differences between groups along with interaction effects between cyclic fatigue and radiation-induced damage. Cyclic fatigue damage resulted in increases in collagen denaturation (1670/1640: 1.517 ± 0.043 vs 1.579 ± 0.021, p Raman spectroscopy can detect the effects of cyclic fatigue damage and 25kGy irradiation via increases in organic matrix (mainly collagen)-bound water. A Raman measure of collagen denaturation was sensitive to cyclic fatigue damage but not 25kGy irradiation. Collagen denaturation was correlated with organic matrix-bound water, suggesting that denaturation of collagen to gelatinous form may expose more binding sites to water by unwinding the triple alpha chains. This research may eventually be useful to help identify allograft quality and more appropriately match donors to recipients. Copyright

  18. LIQUID SOAP CHARACTERISTIC WITH THE ADDITION OF FISH BONE COLLAGEN

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    Nauli A.P.

    2018-04-01

    Full Text Available Doublewhip Threadfin Bream (Nemipterus nematophorus fish is one of the low-cost economical sea fish that can be utilized which is expected to increase its selling value. The utilization of Doublewhip threadfin bream fish can be done with the waste management of the bones in order to have a selling value. Based on the research, other than its skin, bone is also one of the collagen producers in the body of a fish that can be used to increase the amount of collagen in the human body and slowing the aging process caused by damage skin cells that exposed to free radical. Fish bones are also an alternative to the mammals bones such as cows and pigs as collagen production materials that have been damaged by certain diseases. The method used in this research was laboratory experimental using Completely Randomized Design (RAL design. This research aims to analyze the characteristics of fish bone collagen that is applied to the liquid soap, which is done by physic and high. Based on the result of the test, the stability of foam is 84.90%; viscosity 922.83; cPs; pH 10.77 and free alkali 0.031% which meets the requirements of liquid quality based on SNI 06-4085-1996 so it is safe to apply on human skin.

  19. The Mineral–Collagen Interface in Bone

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    2015-01-01

    The interface between collagen and the mineral reinforcement phase, carbonated hydroxyapatite (cAp), is essential for bone’s remarkable functionality as a biological composite material. The very small dimensions of the cAp phase and the disparate natures of the reinforcement and matrix are essential to the material’s performance but also complicate study of this interface. This article summarizes what is known about the cAp-collagen interface in bone and begins with descriptions of the matrix and reinforcement roles in composites, of the phases bounding the interface, of growth of cAp growing within the collagen matrix, and of the effect of intra- and extrafibrilar mineral on determinations of interfacial properties. Different observed interfacial interactions with cAp (collagen, water, non-collagenous proteins) are reviewed; experimental results on interface interactions during loading are reported as are their influence on macroscopic mechanical properties; conclusions of numerical modeling of interfacial interactions are also presented. The data suggest interfacial interlocking (bending of collagen molecules around cAp nanoplatelets) and water-mediated bonding between collagen and cAp are essential to load transfer. The review concludes with descriptions of areas where new research is needed to improve understanding of how the interface functions. PMID:25824581

  20. Fabrication of human hair keratin/jellyfish collagen/eggshell-derived hydroxyapatite osteoinductive biocomposite scaffolds for bone tissue engineering: From waste to regenerative medicine products.

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    Arslan, Yavuz Emre; Sezgin Arslan, Tugba; Derkus, Burak; Emregul, Emel; Emregul, Kaan C

    2017-06-01

    In the present study, we aimed at fabricating an osteoinductive biocomposite scaffold using keratin obtained from human hair, jellyfish collagen and eggshell-derived nano-sized spherical hydroxyapatite (nHA) for bone tissue engineering applications. Keratin, collagen and nHA were characterized with the modified Lowry method, free-sulfhydryl groups and hydroxyproline content analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), attenuated total reflectance-fourier transform infrared spectroscopy (ATR-FTIR) and thermal gravimetric analysis (TGA) which confirmed the success of the extraction and/or isolation processes. Human adipose mesenchymal stem cells (hAMSCs) were isolated and the cell surface markers were characterized via flow cytometry analysis in addition to multilineage differentiation capacity. The undifferentiated hAMSCs were highly positive for CD29, CD44, CD73, CD90 and CD105, but were not seen to express hematopoietic cell surface markers such as CD14, CD34 and CD45. The cells were successfully directed towards osteogenic, chondrogenic and adipogenic lineages in vitro. The microarchitecture of the scaffolds and cell attachment were evaluated using scanning electron microscopy (SEM). The cell viability on the scaffolds was assessed by the MTT assay which revealed no evidence of cytotoxicity. The osteogenic differentiation of hAMSCs on the scaffolds was determined histologically using alizarin red S, osteopontin and osteonectin stainings. Early osteogenic differentiation markers of hAMSCs were significantly expressed on the collagen-keratin-nHA scaffolds. In conclusion, it is believed that collagen-keratin-nHA osteoinductive biocomposite scaffolds have the potential of being used in bone tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Approach to the human diet of the punic population of Can Marines (Ibiza. C an N stable isotope analysis on bone collagen

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    Domingo Carlos Salazar García

    2012-09-01

    Full Text Available We report here on the results of carbon and nitrogen stable isotope analysis on bone collagen of humans from the Punic site of Can Marines (V-IVth BC from the island of Ibiza (Spain. To date, there are few isotopic studies for this period from the Mediterranean. This article reports new isotopic data from a Western Mediterranean Punic rural settlement. The results show a terrestrial based diet with no isotopic evidence of marine or freshwater protein input, and suggest the presence of C4 resources in it.

  2. Does collagen trigger the recruitment of osteoblasts into vacated bone resorption lacunae during bone remodeling?

    DEFF Research Database (Denmark)

    Abdelgawad, Mohamed Essameldin; Søe, Kent; Andersen, Thomas Levin

    2014-01-01

    matrix molecules, collagen's potency was superior and only equaled by fibronectin. Next, the majority of the newly recruited osteoblast lineage cells positioned immediately next to the osteoclasts exhibit uPARAP/Endo180, an endocytic collagen receptor reported to be involved in collagen internalization......Osteoblast recruitment during bone remodeling is obligatory to re-construct the bone resorbed by the osteoclast. This recruitment is believed to be triggered by osteoclast products and is therefore likely to start early during the remodeling cycle. Several osteoclast products with osteoblast...... recruitment potential are already known. Here we draw the attention on the osteoblast recruitment potential of the collagen that is freshly demineralized by the osteoclast. Our evidence is based on observations on adult human cancellous bone, combined with in vitro assays. First, freshly eroded surfaces where...

  3. Collagen Fiber Orientation in Primate Long Bones.

    Science.gov (United States)

    Warshaw, Johanna; Bromage, Timothy G; Terranova, Carl J; Enlow, Donald H

    2017-07-01

    Studies of variation in orientation of collagen fibers within bone have lead to the proposition that these are preferentially aligned to accommodate different kinds of load, with tension best resisted by fibers aligned longitudinally relative to the load, and compression best resisted by transversely aligned fibers. However, previous studies have often neglected to consider the effect of developmental processes, including constraints on collagen fiber orientation (CFO), particularly in primary bone. Here we use circularly polarized light microscopy to examine patterns of CFO in cross-sections from the midshaft femur, humerus, tibia, radius, and ulna in a range of living primate taxa with varied body sizes, phylogenetic relationships and positional behaviors. We find that a preponderance of longitudinally oriented collagen is characteristic of both periosteal primary and intracortically remodeled bone. Where variation does occur among groups, it is not simply understood via interpretations of mechanical loads, although prioritized adaptations to tension and/or shear are considered. While there is some suggestion that CFO may correlate with body size, this relationship is neither consistent nor easily explicable through consideration of size-related changes in mechanical adaptation. The results of our study indicate that there is no clear relationship between CFO and phylogenetic status. One of the principle factors accounting for the range of variation that does exist is primary tissue type, where slower depositing bone is more likely to comprise a larger proportion of oblique to transverse collagen fibers. Anat Rec, 300:1189-1207, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  4. Restoration of a Critical Mandibular Bone Defect Using Human Alveolar Bone-Derived Stem Cells and Porous Nano-HA/Collagen/PLA Scaffold

    Directory of Open Access Journals (Sweden)

    Xing Wang

    2016-01-01

    Full Text Available Periodontal bone defects occur in a wide variety of clinical situations. Adult stem cell- and biomaterial-based bone tissue regeneration are a promising alternative to natural bone grafts. Recent evidence has demonstrated that two populations of adult bone marrow mesenchymal stromal cells (BMSCs can be distinguished based on their embryonic origins. These BMSCs are not interchangeable, as bones preferentially heal using cells that share the same embryonic origin. However, the feasibility of tissue engineering using human craniofacial BMSCs was unclear. The goal of this study was to explore human craniofacial BMSC-based therapy for the treatment of localized mandibular defects using a standardized, minimally invasive procedure. The BMSCs’ identity was confirmed. Scanning electron microscopy, a cell proliferation assay, and supernatant detection indicated that the nHAC/PLA provided a suitable environment for aBMSCs. Real-time PCR and electrochemiluminescence immunoassays demonstrated that osteogenic markers were upregulated by osteogenic preinduction. Moreover, in a rabbit critical-size mandibular bone defect model, total bone formation in the nHAC/PLA + aBMSCs group was significantly higher than in the nHAC/PLA group but significantly lower than in the nHAC/PLA + preinduced aBMSCs. These findings demonstrate that this engineered bone is a valid alternative for the correction of mandibular bone defects.

  5. Restoration of a Critical Mandibular Bone Defect Using Human Alveolar Bone-Derived Stem Cells and Porous Nano-HA/Collagen/PLA Scaffold

    Science.gov (United States)

    Wang, Xing; Xing, Helin; Zhang, Guilan; Wu, Xia; Zou, Xuan; Feng, Lin; Wang, Dongsheng; Li, Meng; Zhao, Jing; Du, Jianwei; Lv, Yan; E, Lingling; Liu, Hongchen

    2016-01-01

    Periodontal bone defects occur in a wide variety of clinical situations. Adult stem cell- and biomaterial-based bone tissue regeneration are a promising alternative to natural bone grafts. Recent evidence has demonstrated that two populations of adult bone marrow mesenchymal stromal cells (BMSCs) can be distinguished based on their embryonic origins. These BMSCs are not interchangeable, as bones preferentially heal using cells that share the same embryonic origin. However, the feasibility of tissue engineering using human craniofacial BMSCs was unclear. The goal of this study was to explore human craniofacial BMSC-based therapy for the treatment of localized mandibular defects using a standardized, minimally invasive procedure. The BMSCs' identity was confirmed. Scanning electron microscopy, a cell proliferation assay, and supernatant detection indicated that the nHAC/PLA provided a suitable environment for aBMSCs. Real-time PCR and electrochemiluminescence immunoassays demonstrated that osteogenic markers were upregulated by osteogenic preinduction. Moreover, in a rabbit critical-size mandibular bone defect model, total bone formation in the nHAC/PLA + aBMSCs group was significantly higher than in the nHAC/PLA group but significantly lower than in the nHAC/PLA + preinduced aBMSCs. These findings demonstrate that this engineered bone is a valid alternative for the correction of mandibular bone defects. PMID:27118977

  6. Hydroxyapatite/collagen bone-like nanocomposite.

    Science.gov (United States)

    Kikuchi, Masanori

    2013-01-01

    Our group has succeeded to synthesize material with bone-like nanostructure and bone-like inorganic and organic composition via self-organization mechanism between them using simultaneous titration method under controlled pH and temperature. The hydroxyapatite/collagen (HAp/Col) bone-like nanocomposite completely incorporated into bone remodeling process to be substituted by new bone. Cells cultured on the HAp/Col revealed very interesting reactions. Osteoblast-like MG63 cells showed upregulation of alkaline phosphatase >3 times greater than MG63 cells cultured on tissue culture polystyrene (TCPS). MG63 cells 3-dimensionally cultured in a "HAp/Col sponge," a porous HAp/Col having sponge-like viscoelasticity, accumulated calcium phosphate nodules on extracellular matrices they secreted. Bone marrow cells co-cultured with osteoblasts on HAp/Col differentiated to osteoclasts without differentiation supplements. This phenomenon is not found in cells cultured on hydroxyapatite ceramics and TCPS, and rarely in cells cultured on dentin. These results suggest that HAp/Col is a good candidate for tissue engineering of bone as well as bone filler. In a clinical test as a bone filler, the HAp/Col sponge was significantly better than porous β-tricalcium phosphate. The HAp/Col sponge has been approved by the Japanese government and will be used as greatly needed bone filler in patients. In addition to the above, HAp/Col coating on titanium revealed higher osteo-conductivity than HAp-coated titanium and bare titanium and improved direct bonding between titanium and newly formed bone. The HAp/Col coating may be used for metal devices requiring osseointegration.

  7. Stable C &N isotopes in 2100 Year-B.P. human bone collagen indicate rare dietary dominance of C4 plants in NE-Italy.

    Science.gov (United States)

    Laffranchi, Zita; Huertas, Antonio Delgado; Jiménez Brobeil, Sylvia A; Torres, Arsenio Granados; Riquelme Cantal, Jose A

    2016-12-09

    C 4 plants (e.g. maize, millet), part of our current diet, are only endemic of reduced areas in South-Europe due to their need of warm climates. Since the first vestiges of agriculture in Europe remains of C 4 plants were recorded but their overall proportion in the human diet remains unknown. Therefore, isotopic (δ 13 C and δ 15 N) composition of bone collagen from the skeletal remains (human and animals) of a Celtic population, Cenomani Gauls, from Verona (3 rd to 1 st century BC) in the NE Italy provide a new perspective on this matter. The δ 13 C collagen values of 90 human skeletal individuals range between -20.2‰ and -9.7‰ (V-PDB) with a mean value of -15.3‰. As present day C 4 plants have δ 13 C values around -11‰, which is equivalent to -9.5‰ for samples of preindustrial age, the less negative δ 13 C values in these individuals indicate a diet dominated by C 4 plants. This palaeodietary study indicates that some European populations predominantly consumed cultivated C 4 plants 2100 year B.P. This is supported by the paleobotanical records and ancient Roman sources (e.g. Pliny the Elder), which indicate that millet was a staple food in South-Europe.

  8. Demineralized dentin matrix composite collagen material for bone tissue regeneration.

    Science.gov (United States)

    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.

  9. Second-harmonic generation imaging of collagen in ancient bone.

    Science.gov (United States)

    Thomas, B; McIntosh, D; Fildes, T; Smith, L; Hargrave, F; Islam, M; Thompson, T; Layfield, R; Scott, D; Shaw, B; Burrell, C L; Gonzalez, S; Taylor, S

    2017-12-01

    Second-harmonic generation imaging (SHG) captures triple helical collagen molecules near tissue surfaces. Biomedical research routinely utilizes various imaging software packages to quantify SHG signals for collagen content and distribution estimates in modern tissue samples including bone. For the first time using SHG, samples of modern, medieval, and ice age bones were imaged to test the applicability of SHG to ancient bone from a variety of ages, settings, and taxa. Four independent techniques including Raman spectroscopy, FTIR spectroscopy, radiocarbon dating protocols, and mass spectrometry-based protein sequencing, confirm the presence of protein, consistent with the hypothesis that SHG imaging detects ancient bone collagen. These results suggest that future studies have the potential to use SHG imaging to provide new insights into the composition of ancient bone, to characterize ancient bone disorders, to investigate collagen preservation within and between various taxa, and to monitor collagen decay regimes in different depositional environments.

  10. Second-harmonic generation imaging of collagen in ancient bone

    Directory of Open Access Journals (Sweden)

    B. Thomas

    2017-12-01

    Full Text Available Second-harmonic generation imaging (SHG captures triple helical collagen molecules near tissue surfaces. Biomedical research routinely utilizes various imaging software packages to quantify SHG signals for collagen content and distribution estimates in modern tissue samples including bone. For the first time using SHG, samples of modern, medieval, and ice age bones were imaged to test the applicability of SHG to ancient bone from a variety of ages, settings, and taxa. Four independent techniques including Raman spectroscopy, FTIR spectroscopy, radiocarbon dating protocols, and mass spectrometry-based protein sequencing, confirm the presence of protein, consistent with the hypothesis that SHG imaging detects ancient bone collagen. These results suggest that future studies have the potential to use SHG imaging to provide new insights into the composition of ancient bone, to characterize ancient bone disorders, to investigate collagen preservation within and between various taxa, and to monitor collagen decay regimes in different depositional environments.

  11. The collagenic architecture of human dura mater.

    Science.gov (United States)

    Protasoni, Marina; Sangiorgi, Simone; Cividini, Andrea; Culuvaris, Gloria Tiffany; Tomei, Giustino; Dell'Orbo, Carlo; Raspanti, Mario; Balbi, Sergio; Reguzzoni, Marcella

    2011-06-01

    Human dura mater is the most external meningeal sheet surrounding the CNS. It provides an efficient protection to intracranial structures and represents the most important site for CSF turnover. Its intrinsic architecture is made up of fibrous tissue including collagenic and elastic fibers that guarantee the maintenance of its biophysical features. The recent technical advances in the repair of dural defects have allowed for the creation of many synthetic and biological grafts. However, no detailed studies on the 3D microscopic disposition of collagenic fibers in dura mater are available. The authors report on the collagenic 3D architecture of normal dura mater highlighting the orientation, disposition in 3 dimensions, and shape of the collagen fibers with respect to the observed layer. Thirty-two dura mater specimens were collected during cranial decompressive surgical procedures, fixed in 2.5% Karnovsky solution, and digested in 1 N NaOH solution. After a routine procedure, the specimens were observed using a scanning electron microscope. The authors distinguished the following 5 layers in the fibrous dura mater of varying thicknesses, orientation, and structures: bone surface, external median, vascular, internal median, and arachnoid layers. The description of the ultrastructural 3D organization of the different layers of dura mater will give us more information for the creation of synthetic grafts that are as similar as possible to normal dura mater. This description will be also related to the study of the neoplastic invasion.

  12. [Polarized microscopic observation of the collagen change in bone healing during bone lengthening].

    Science.gov (United States)

    Zou, Pei; Li, Junhui; Li, Zhuyi

    2006-01-01

    To investigate the feature and regularity of the collagen change in bone healing during bone lengthening. Bone lengthening model was made in the middle segment of the rabbit tibia. Five days after the model was established, the bone was lengthened 1.5 mm per day for 14 days. The rabbits were put to death after elongation, 7, 14, 21, 30, 40, 50, 60 and 70 days after elongation. The distracted area of the bone was imbedded with paraffin. After being stained by the picric acid-sirius red staining, the slice was observed under polarized microscope. The features of the collagen change in the distracted bone were as follows: (1) In the fibrous tissue of the distracted area during lengthening period and the early stage after lengthening, there was not only collagen III but also much collagen I. (2) Collagen I , II and III were observed in the cartilage. (3) Collagen I, II and III were also observed in the pseudo-growth plate. (4) Collagen I took the dominance during lengthening period and the late stage after lengthening. New bone formation in bone lengthening is under the distracted force, so the collagen changes have different features compared with that in fracture healing. Collagen I, II and III can be identified by picric acid-sirius red staining and polarized microscope, so a new method for studying the collagen typing in bone repairing is provided.

  13. Biological effect of hydrolyzed collagen on bone metabolism.

    Science.gov (United States)

    Daneault, Audrey; Prawitt, Janne; Fabien Soulé, Véronique; Coxam, Véronique; Wittrant, Yohann

    2017-06-13

    Osteoporosis is a chronic and asymptomatic disease characterized by low bone mass and skeletal microarchitectural deterioration, increased risk of fracture, and associated comorbidities most prevalent in the elderly. Due to an increasingly aging population, osteoporosis has become a major health issue requiring innovative disease management. Proteins are important for bone by providing building blocks and by exerting specific regulatory function. This is why adequate protein intake plays a considerable role in both bone development and bone maintenance. More specifically, since an increase in the overall metabolism of collagen can lead to severe dysfunctions and a more fragile bone matrix and because orally administered collagen can be digested in the gut, cross the intestinal barrier, enter the circulation, and become available for metabolic processes in the target tissues, one may speculate that a collagen-enriched diet provides benefits for the skeleton. Collagen-derived products such as gelatin or hydrolyzed collagen (HC) are well acknowledged for their safety from a nutritional point of view; however, what is their impact on bone biology? In this manuscript, we critically review the evidence from literature for an effect of HC on bone tissues in order to determine whether HC may represent a relevant alternative in the design of future nutritional approaches to manage osteoporosis prevention.

  14. Collagen-derived markers of bone metabolism in osteogenesis imperfecta

    DEFF Research Database (Denmark)

    Lund, A M; Hansen, M; Kollerup, Gina Birgitte

    1998-01-01

    )] were measured in 78 osteogenesis imperfecta (OI) patients to investigate bone metabolism in vivo and relate marker concentrations to phenotype and in vitro collagen I defects, as shown by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). PICP and PINP were generally low....... The in vivo findings correlated with in vitro results of collagen I SDS-PAGE. Bone turnover is reduced in OI children and mildly affected OI adults, whereas bone resorption is elevated in severely affected adults. These findings may prove helpful for diagnosis and decision-making regarding therapy in OI....

  15. Odontogenic Differentiation of Human Dental Pulp Stem Cells on Hydrogel Scaffolds Derived from Decellularized Bone Extracellular Matrix and Collagen Type I.

    Science.gov (United States)

    Paduano, Francesco; Marrelli, Massimo; White, Lisa J; Shakesheff, Kevin M; Tatullo, Marco

    2016-01-01

    The aim of this study was to evaluate the level of odontogenic differentiation of dental pulp stem cells (DPSCs) on hydrogel scaffolds derived from bone extracellular matrix (bECM) in comparison to those seeded on collagen I (Col-I), one of the main components of dental pulp ECM. DPSCs isolated from human third molars were characterized for surface marker expression and odontogenic potential prior to seeding into bECM or Col-I hydrogel scaffolds. The cells were then seeded onto bECM and Col-I hydrogel scaffolds and cultured under basal conditions or with odontogenic and growth factor (GF) supplements. DPSCs cultivated on tissue culture polystyrene (TCPS) with and without supplements were used as controls. Gene expression of dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP-1) and matrix extracellular phosphoglycoprotein (MEPE) was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and mineral deposition was observed by Von Kossa staining. When DPSCs were cultured on bECM hydrogels, the mRNA expression levels of DSPP, DMP-1 and MEPE genes were significantly upregulated with respect to those cultured on Col-I scaffolds or TCPS in the absence of extra odontogenic inducers. In addition, more mineral deposition was observed on bECM hydrogel scaffolds as demonstrated by Von Kossa staining. Moreover, DSPP, DMP-1 and MEPE mRNA expressions of DPSCs cultured on bECM hydrogels were further upregulated by the addition of GFs or osteo/odontogenic medium compared to Col-I treated cells in the same culture conditions. These results demonstrate the potential of the bECM hydrogel scaffolds to stimulate odontogenic differentiation of DPSCs.

  16. A Novel HA/β-TCP-Collagen Composite Enhanced New Bone Formation for Dental Extraction Socket Preservation in Beagle Dogs

    Directory of Open Access Journals (Sweden)

    Ko-Ning Ho

    2016-03-01

    Full Text Available Past studies in humans have demonstrated horizontal and vertical bone loss after six months following tooth extraction. Many biomaterials have been developed to preserve bone volume after tooth extraction. Type I collagen serves as an excellent delivery system for growth factors and promotes angiogenesis. Calcium phosphate ceramics have also been investigated because their mineral chemistry resembles human bone. The aim of this study was to compare the performance of a novel bioresorbable purified fibrillar collagen and hydroxyapatite/β-tricalcium phosphate (HA/β-TCP ceramic composite versus collagen alone and a bovine xenograft-collagen composite in beagles. Collagen plugs, bovine graft-collagen composite and HA/β-TCP-collagen composite were implanted into the left and right first, second and third mandibular premolars, and the fourth molar was left empty for natural healing. In total, 20 male beagle dogs were used, and quantitative and histological analyses of the extraction ridge was done. The smallest width reduction was 19.09% ± 8.81% with the HA/β-TCP-collagen composite at Week 8, accompanied by new bone formation at Weeks 4 and 8. The HA/β-TCP-collagen composite performed well, as a new osteoconductive and biomimetic composite biomaterial, for socket bone preservation after tooth extraction.

  17. Vertical ridge augmentation using an equine bone and collagen block infused with recombinant human platelet-derived growth factor-BB: a randomized single-masked histologic study in non-human primates.

    Science.gov (United States)

    Nevins, Myron; Al Hezaimi, Khalid; Schupbach, Peter; Karimbux, Nadeem; Kim, David M

    2012-07-01

    This study tests the effectiveness of hydroxyapatite and collagen bone blocks of equine origin (eHAC), infused with recombinant human platelet-derived growth factor-BB (rhPDGF-BB), to augment localized posterior mandibular defects in non-human primates (Papio hamadryas). Bilateral critical-sized defects simulating severe atrophy were created at the time of the posterior teeth extraction. Test and control blocks (without growth factor) were randomly grafted into the respective sites in each non-human primate. All sites exhibited vertical ridge augmentation, with physiologic hard- and soft-tissue integration of the blocks when clinical and histologic examinations were done at 4 months after the vertical ridge augmentation procedure. There was a clear, although non-significant, tendency to increased regeneration in the test sites. As in the first two preclinical studies in this series using canines, experimental eHAC blocks infused with rhPDGF-BB proved to be a predictable and technically viable method to predictably regenerate bone and soft tissue in critical-sized defects. This investigation supplies additional evidence that eHAC blocks infused with rhPDGF-BB growth factor is a predictable and technically feasible option for vertical augmentation of severely resorbed ridges.

  18. The effects of bone marrow aspirate, bone graft, and collagen composites on fixation of titanium implants

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Sandri, Monica

    2012-01-01

    Replacement of extensive local bone loss especially in revision joint arthroplasty and spine fusion is a significant clinical challenge. Allograft and autograft have been considered as gold standards for bone replacement. However, there are several disadvantages such as donor site pain, bacterial...... contamination, and non union as well as the potential risk of disease transmission. Hydroxyapatite and collagen composites (HA/Collagen) have the potential in mimicking and replacing skeletal bones. This study attempted to determine the effects of newly developed HA/Collagen-composites with and without bone...... marrow aspirate (BMA) on enhancement of bone implant fixation. Method: Titanium alloy implants were inserted into bilateral femoral condyles of eight skeletally mature sheep, four implants per sheep. The implant had a circumferential gap of 2 mm. The gap was filled with: HA/Collagen; HA...

  19. Abnormal bone collagen morphology and decreased bone strength in growth hormone-deficient rats

    DEFF Research Database (Denmark)

    Lange, Martin; Qvortrup, Klaus; Svendsen, Ole Lander

    2004-01-01

    collagen morphology and bone mineralisation in cortical bone as well as bone strength in GHD rats to try to clarify the explanation for the increased fracture rate. The Dw-4 rat was used as a model for GHD. This strain of rats has an autosomal recessive disorder, reducing GH synthesis to approximately 10...

  20. Collagen-derived markers of bone metabolism in osteogenesis imperfecta

    DEFF Research Database (Denmark)

    Lund, A M; Hansen, M; Kollerup, Gina Birgitte

    1998-01-01

    )] were measured in 78 osteogenesis imperfecta (OI) patients to investigate bone metabolism in vivo and relate marker concentrations to phenotype and in vitro collagen I defects, as shown by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). PICP and PINP were generally low...

  1. A biocomposite of collagen nanofibers and nanohydroxyapatite for bone regeneration

    NARCIS (Netherlands)

    Ribeiro, N.; Sousa, S.R.; van Blitterswijk, Clemens; Moroni, Lorenzo; Monteiro, F.J.

    2014-01-01

    This work aims to design a synthetic construct that mimics the natural bone extracellular matrix through innovative approaches based on simultaneous type I collagen electrospinning and nanophased hydroxyapatite (nanoHA) electrospraying using non-denaturating conditions and non-toxic reagents. The

  2. Computational segmentation of collagen fibers in bone matrix indicates bone quality in ovariectomized rat spine.

    Science.gov (United States)

    Daghma, Diaa Eldin S; Malhan, Deeksha; Simon, Paul; Stötzel, Sabine; Kern, Stefanie; Hassan, Fathi; Lips, Katrin Susanne; Heiss, Christian; El Khassawna, Thaqif

    2018-05-01

    Bone loss varies according to disease and age and these variations affect bone cells and extracellular matrix. Osteoporosis rat models are widely investigated to assess mechanical and structural properties of bone; however, bone matrix proteins and their discrepant regulation of diseased and aged bone are often overlooked. The current study considered the spine matrix properties of ovariectomized rats (OVX) against control rats (Sham) at 16 months of age. Diseased bone showed less compact structure with inhomogeneous distribution of type 1 collagen (Col1) and changes in osteocyte morphology. Intriguingly, demineralization patches were noticed in the vicinity of blood vessels in the OVX spine. The organic matrix structure was investigated using computational segmentation of collagen fibril properties. In contrast to the aged bone, diseased bone showed longer fibrils and smaller orientation angles. The study shows the potential of quantifying transmission electron microscopy images to predict the mechanical properties of bone tissue.

  3. Effect of water on piezoelectricity in bone and collagen.

    Science.gov (United States)

    Netto, T G; Zimmerman, R L

    1975-01-01

    Interferometric measurements of bovine bone and tendon show that the values of the piezoelectric strain constant d14 decrease with hydration from the dry values of 0.2 X 10(-14) and 2.0 X 10(-14) m/V, respectively. The decrease of piezoelectricity in tendon is exponential with a characteristic hydration of 7% by weight from which an upper limit of the average molecular weight of the responsible electric dipole moments is deduced. The piezoelectricity in bone decreases relatively slowly with hydration indicating that the electric dipoles in bone collagen are subject to a different cancelling mechanism. PMID:1148359

  4. Endogenous collagen influences differentiation of human multipotent mesenchymal stromal cells.

    Science.gov (United States)

    Fernandes, Hugo; Mentink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    2010-05-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix mainly composed of collagen type I. Here we assessed the potential role of endogenous collagen synthesis in hMSC differentiation and stem cell maintenance. We observed a sharp reduction in proliferation rate of hMSCs in the absence of ascorbic acid, concomitant with a reduction in osteogenesis in vitro and bone formation in vivo. In line with a positive role for collagen type I in osteogenesis, gene expression profiling of hMSCs cultured in the absence of ascorbic acid demonstrated increased expression of genes involved in adipogenesis and chondrogenesis and a reduction in expression of osteogenic genes. We also observed that matrix remodeling and anti-osteoclastogenic signals were high in the presence of ascorbic acid. The presence of collagen type I during the expansion phase of hMSCs did not affect their osteogenic and adipogenic differentiation potential. In conclusion, the collagenous matrix supports both proliferation and differentiation of osteogenic hMSCs but, on the other hand, presents signals stimulating matrix remodeling and inhibiting osteoclastogenesis.

  5. Glyoxal Crosslinking of Cell-Seeded Chitosan/Collagen Hydrogels for Bone Regeneration

    Science.gov (United States)

    Wang, Limin; Stegemann, Jan P.

    2011-01-01

    Chitosan and collagen are natural biomaterials that have been used extensively in tissue engineering, both separately and as composite materials. Most methods to fabricate chitosan/collagen composites use freeze drying and chemical crosslinking to create stable porous scaffolds, which subsequently can be seeded with cells. In this study, we directly embedded human bone marrow stem cells (hBMSC) in chitosan/collagen materials by initiating gelation using β-glycerophosphate at physiological temperature and pH. We further examined the use of glyoxal, a dialdehyde with relatively low toxicity, to crosslink these materials and characterized the resulting changes in matrix and cell properties. The cytocompatibility of glyoxal and the crosslinked gels were investigated in terms of hBMSC metabolic activity, viability, proliferation, and osteogenic differentiation. These studies revealed that glyoxal was cytocompatible at concentrations below about 1 mM for periods of exposure up to 15 h, though the degree of cell spreading and proliferation were dependent on matrix composition. Glyoxal-crosslinked matrices were stiffer and compacted less than uncrosslinked controls. It was further demonstrated that hBMSC can attach and proliferate in 3D matrices composed of 50/50 chitosan/collagen, and that these materials supported osteogenic differentiation in response to stimulation. Such glyoxal-crosslinked chitosan/collagen composite materials may find utility as cell delivery vehicles for enhancing the repair of bone defects. PMID:21345389

  6. Atomic scale chemical tomography of human bone

    Science.gov (United States)

    Langelier, Brian; Wang, Xiaoyue; Grandfield, Kathryn

    2017-01-01

    Human bone is a complex hierarchical material. Understanding bone structure and its corresponding composition at the nanometer scale is critical for elucidating mechanisms of biomineralization under healthy and pathological states. However, the three-dimensional structure and chemical nature of bone remains largely unexplored at the nanometer scale due to the challenges associated with characterizing both the structural and chemical integrity of bone simultaneously. Here, we use correlative transmission electron microscopy and atom probe tomography for the first time, to our knowledge, to reveal structures in human bone at the atomic level. This approach provides an overlaying chemical map of the organic and inorganic constituents of bone on its structure. This first use of atom probe tomography on human bone reveals local gradients, trace element detection of Mg, and the co-localization of Na with the inorganic-organic interface of bone mineral and collagen fibrils, suggesting the important role of Na-rich organics in the structural connection between mineral and collagen. Our findings provide the first insights into the hierarchical organization and chemical heterogeneity in human bone in three-dimensions at its smallest length scale - the atomic level. We demonstrate that atom probe tomography shows potential for new insights in biomineralization research on bone.

  7. Greener synthesis of electrospun collagen/hydroxyapatite composite fibers with an excellent microstructure for bone tissue engineering

    Science.gov (United States)

    Zhou, Yuanyuan; Yao, Hongchang; Wang, Jianshe; Wang, Dalu; Liu, Qian; Li, Zhongjun

    2015-01-01

    In bone tissue engineering, collagen/hydroxyapatite (HAP) fibrous composite obtained via electrospinning method has been demonstrated to support the cells’ adhesion and bone regeneration. However, electrospinning of natural collagen often requires the use of cytotoxic organic solvents, and the HAP crystals were usually aggregated and randomly distributed within a fibrous matrix of collagen, limiting their clinical potential. Here, an effective and greener method for the preparation of collagen/HAP composite fibers was developed for the first time, and this green product not only had 40 times higher mechanical properties than that previously reported, but also had an excellent microstructure similar to that of natural bone. By dissolving type I collagen in environmentally friendly phosphate buffered saline/ethanol solution instead of the frequently-used cytotoxic organic solvents, followed with the key step of desalination, co-electrospinning the collagen solution with the HAP sol, generates a collagen/HAP composite with a uniform and continuous fibrous morphology. Interestingly, the nano-HAP needles were found to preferentially orient along the longitudinal direction of the collagen fibers, which mimicked the nanostructure of natural bones. Based on the characterization of the related products, the formation mechanism for this novel phenomenon was proposed. After cross-linking with 1-ethyl-3-(3-dimethyl-aminopropyl)-1-carbodiimide hydrochloride/N-hydroxysuccinimide, the obtained composite exhibited a significant enhancement in mechanical properties. In addition, the biocompatibility of the obtained composite fibers was evaluated by in vitro culture of the human myeloma cells (U2-OS). Taken together, the process outlined herein provides an effective, non-toxic approach for the fabrication of collagen/HAP composite nanofibers that could be good candidates for bone tissue engineering. PMID:25995630

  8. Investigation of the collagen-mineral-relation in bone with special respect to bone diseases with collagen defects by small-angle X-ray scattering

    International Nuclear Information System (INIS)

    Schreiber, S. A.

    1996-06-01

    Small-angle X-ray scattering (SAXS) was used to study the structure of the collagen/mineral composite of bone in the nanometer range. The most important results were: - In horse radius, the angular distribution of mineral crystals as measured by SAXS agreed well with previous measurements of collagen orientation using circularly polarized light microscopy. This shows that the crystals are parallel to the collagen fibrils. - The effect of sodium fluoride, which stimulates bone formation, and bisphosphonates, which reduce bone resorption, were analyzed. A slight increase in the average thickness of the mineral crystals as well as changes in the structure of the mineral/collagen composite were found in the case of fluoride treated animals. No differences were found between alendronate treated animals and controls. The changes with NaF correlate with bone weakening found in an earlier study with the same animals. - In cortical bone from 9 patients with Osteogenesis Imperfecta (brittle bone disease) the mean thickness of the mineral crystals was found approximately constant around 2.4 nm, while in control bones it constantly increased with age up to about 3.5 nm. In addition, the parallel alignment of the mineral crystals was less in OI-bone than in normal controls. Hence, despite the great variability of this genetic collagen defect, smaller and less well aligned mineral crystals seem to characterize the collagen/mineral composite in OI-bone. (author)

  9. The effect of bone marrow aspirate, bone graft and collagen composites on fixation of bone implants

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Overgaard, Søren

    2007-01-01

     Introduction: Replacement of extensive local bone loss especially in revision joint arthroplasties is a significant clinical challenge. Autogenous and allogenic cancellous bone grafts have been the gold standard in reconstructive orthopaedic surgery, but it is well known that there is morbidity...... associated with harvesting of autogenous bone graft and limitations in the quantity of bone available. Disadvantages of allograft include the risk of bacterial or viral contamination and non union as well as the potential risk of disease transmission. Alternative options are attractive and continue...... to be sought. Hydroxyapatite and collagen composites have the potential in mimicking and replacing skeletal bones. Aim: This study attempted to determine the effect of hydroxyapatite/collagen composites in the fixation of bone implants. The composites used in this study is produced by Institute of Science...

  10. Identification of a new hominin bone from Denisova Cave, Siberia using collagen fingerprinting and mitochondrial DNA analysis

    Science.gov (United States)

    Brown, Samantha; Higham, Thomas; Slon, Viviane; Pääbo, Svante; Meyer, Matthias; Douka, Katerina; Brock, Fiona; Comeskey, Daniel; Procopio, Noemi; Shunkov, Michael; Derevianko, Anatoly; Buckley, Michael

    2016-03-01

    DNA sequencing has revolutionised our understanding of archaic humans during the Middle and Upper Palaeolithic. Unfortunately, while many Palaeolithic sites contain large numbers of bones, the majority of these lack the diagnostic features necessary for traditional morphological identification. As a result the recovery of Pleistocene-age human remains is extremely rare. To circumvent this problem we have applied a method of collagen fingerprinting to more than 2000 fragmented bones from the site of Denisova Cave, Russia, in order to facilitate the discovery of human remains. As a result of our analysis a single hominin bone (Denisova 11) was identified, supported through in-depth peptide sequencing analysis, and found to carry mitochondrial DNA of the Neandertal type. Subsequent radiocarbon dating revealed the bone to be >50,000 years old. Here we demonstrate the huge potential collagen fingerprinting has for identifying hominin remains in highly fragmentary archaeological assemblages, improving the resources available for wider studies into human evolution.

  11. Guided Bone Regeneration in Long-Bone Defects with a Structural Hydroxyapatite Graft and Collagen Membrane

    Science.gov (United States)

    2013-01-01

    Original Articles Guided Bone Regeneration in Long-Bone Defects with a Structural Hydroxyapatite Graft and Collagen Membrane Teja Guda, PhD,1,2 John...Joint Surg Br 90-B, 1617, 2008. 6. Carlo Reis, E.C., Borges AaPB, Araujo, M.V.F., Mendes, V.C., Guan, L., and Davies, J.E. Periodontal regeneration...Regeneration of periodontal tissues: combinations of barrier membranes and grafting materials–biological foundation and preclinical evi- dence: a

  12. Immune responses to implanted human collagen graft in rats

    International Nuclear Information System (INIS)

    Quteish, D.; Dolby, A.E.

    1991-01-01

    Immunity to collagen implants may be mediated by cellular and humoral immune responses. To examine the possibility of such immunological reactivity and crossreactivity to collagen, 39 Sprague-Dawley rats (female, 10 weeks old, approximately 250 g wt) were implanted subcutaneously at thigh sites with crosslinked, freeze-dried human placental type I collagen grafts (4x4x2 mm) which had been irradiated (520 Gray) or left untreated. Blood was obtained by intracardiac sampling prior to implantation or from normal rats, and at various times afterwards when the animals were sacrificed. The sera from these animals were examined for circulating antibodies to human, bovine and rat tail (type I) collagens by enzyme-linked immunosorbent assay (ELISA). Also, the lymphoblastogenic responses of spleen lymphocytes from the irradiated collagen-implanted animals were assessed in culture by measuring thymidine uptake with autologous and normal rat sera in the presence of human bovine type I collagens. Implantation of the irradiated and non-irradiated collagen graft in rats led to a significant increase in the level of circulating antibodies to human collagen. Also antibody to bovine and rat tail collagens was detectable in the animals implanted with irradiated collagen grafts but at a lower level than the human collagen. There was a raised lymphoblastogenic response to both human and bovine collagens. The antibody level and lymphoblastogenesis to the tested collagens gradually decreased towards the end of the post-implantation period. (author)

  13. COLOSTRUM-COLLAGEN-HYDROXYAPATITE COMPOSITE, AN EXCELLENT CANDIDATE BIOMATERIAL FOR BONE REPAIR AND BONE INFECTION MANAGEMENT

    Directory of Open Access Journals (Sweden)

    Dio Nurdin Setiawan

    2014-05-01

    Full Text Available In the case ofbone fracture or defect after surgery, which is common in patients with bone cancer (osteosarcoma, it takes a long time for closure and it may cause an infection problem. The use ofcollagen-hydroxyapatite composite with a blend ofcolostrum as a scaffold is aimed to accelerate the process of osteoblast growth, inhibite the emergence of infections, and act as bone tissue repair material. The method used was the hydrogel formation process and freeze dry process to remove the solvent and to form pores. The composition of scaffold composite manufactured was 15% collagen, 75% hydroxyapatite and 10% colostrum. Combination ofscaffold collagen-hydroxyapatite-colostrum has quite reliable properties because SEM test showed that scaffold could bind to both and could bind to both and could form sufficient pores to provide enough place for bone cells (osteoblats to grow. The results of MTT assay revealed percentage of above 60%, which indicates that the material is not toxic. In conclusion, collagen-hydroxyapatite-colostrum combination is an excellent biomaterial candidate for bone repair and bone infection management.

  14. Effect of protein malnutrition on the metabolism of bone collagen in albino rats

    Energy Technology Data Exchange (ETDEWEB)

    Rao, J S; Rao, V H [Central Leather Research Inst., Madras (India)

    1981-01-01

    The effect of protein malnutrition on the metabolism of collagen in bone was studied in young female albino rats after a single injection of /sup 3/H-proline. Both specific and total radioactivities of hydroxyproline in the total collagen of the bone were found to decrease in the protein-deficient animals, indicating decreased rate of collagen synthesis. In the urine the amount of hydroxyproline excreted and total radioactivity of /sup 3/H-hydroxyproline were greatly decreased. The results of the present investigation therefore clearly indicate decreased synthesis and catabolism of collagen in bones of protein deficient animals compared to controls.

  15. Repair of rabbit radial bone defects using true bone ceramics combined with BMP-2-related peptide and type I collagen

    International Nuclear Information System (INIS)

    Li Jingfeng; Lin Zhenyu; Zheng Qixin; Guo Xiaodong; Lan Shenghui; Liu Sunan; Yang Shuhua

    2010-01-01

    An ideal bone graft material is the one characterized with good biocompatibility, biodegradation, osteoconductivity and osteoinductivity. In this study, a novel synthetic BMP-2-related peptide (designated P24) corresponding to residues of the knuckle epitope of BMP-2 was introduced into a biomimetic scaffold based on sintered bovine bone or true bone ceramics (TBC) and type I collagen (TBC/collagen I) using a simulated body fluid (SBF). Hydroxylapatite crystal mineralization with a Ca/P molar ratio of 1.63 was observed on the surface of P24/TBC/collagen I composite by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX) and X-ray diffraction (XRD) techniques. Cell adhesion rate evaluation of bone marrow stromal cells (BMSCs) seeded on materials in vitro showed that the percentage of cells attached to P24/TBC/collagen I composite was significantly higher than that of the TBC/collagen I composite. A 10 mm unilateral segmental bone defect was created in the radius of New Zealand white rabbits and randomly implanted with three groups of biomaterials (Group A: P24/TBC/collagen I composite; Group B: TBC/collagen I composite and Group C: TBC alone). Based on radiographic evaluation and histological examination, the implants of P24/TBC/collagen I composite significantly stimulated bone growth, thereby confirming the enhanced rate of bone healing compared with that of TBC/collagen I composite and TBC alone. It was concluded that BMP-2-related peptide P24 could induce nucleation of calcium phosphate crystals on the surface of TBC/collagen I composite. The TBC/collagen I composite loaded with the synthetic BMP-2-related peptide is a promising scaffold biomaterial for bone tissue engineering.

  16. Preparation of dexamethasone-loaded biphasic calcium phosphate nanoparticles/collagen porous composite scaffolds for bone tissue engineering.

    Science.gov (United States)

    Chen, Ying; Kawazoe, Naoki; Chen, Guoping

    2018-02-01

    Although bone is regenerative, its regeneration capacity is limited. For bone defects beyond a critical size, further intervention is required. As an attractive strategy, bone tissue engineering (bone TE) has been widely investigated to repair bone defects. However, the rapid and effective bone regeneration of large non-healing defects is still a great challenge. Multifunctional scaffolds having osteoinductivity and osteoconductivity are desirable to fasten functional bone tissue regeneration. In the present study, biomimetic composite scaffolds of collagen and biphasic calcium phosphate nanoparticles (BCP NPs) with a controlled release of dexamethasone (DEX) and the controlled pore structures were prepared for bone TE. DEX was introduced in the BCP NPs during preparation of the BCP NPs and hybridized with collagen scaffolds, which pore structures were controlled by using pre-prepared ice particulates as a porogen material. The composite scaffolds had well controlled and interconnected pore structures, high mechanical strength and a sustained release of DEX. The composite scaffolds showed good biocompatibility and promoted osteogenic differentiation of hMSCs when used for three-dimensional culture of human bone marrow-derived mesenchymal stem cells. Subcutaneous implantation of the composite scaffolds at the dorsa of athymic nude mice demonstrated that they facilitated the ectopic bone tissue regeneration. The results indicated the DEX-loaded BCP NPs/collagen composite scaffolds had high potential for bone TE. Scaffolds play a crucial role for regeneration of large bone defects. Biomimetic scaffolds having the same composition of natural bone and a controlled release of osteoinductive factors are desirable for promotion of bone regeneration. In this study, composite scaffolds of collagen and biphasic CaP nanoparticles (BCP NPs) with a controlled release nature of dexamethasone (DEX) were prepared and their porous structures were controlled by using ice particulates

  17. Kinetics of gene expression and bone remodelling in the clinical phase of collagen induced arthritis

    DEFF Research Database (Denmark)

    Denninger, Katja Caroline Marie; Litman, Thomas; Marstrand, Troels

    2015-01-01

    Introduction: Pathological bone changes differ considerably between inflammatory arthritic diseases and most studies have focused on bone erosion. Collagen-induced arthritis (CIA) is a model for rheumatoid arthritis, which, in addition to bone erosion, demonstrates bone formation at the time...

  18. Enhancement of tendon–bone healing via the combination of biodegradable collagen-loaded nanofibrous membranes and a three-dimensional printed bone-anchoring bolt

    Directory of Open Access Journals (Sweden)

    Chou YC

    2016-08-01

    Full Text Available Ying-Chao Chou,1,2 Wen-Lin Yeh,2 Chien-Lin Chao,1 Yung-Heng Hsu,1,2 Yi-Hsun Yu,1,2 Jan-Kan Chen,3 Shih-Jung Liu1,2 1Department of Mechanical Engineering, Chang Gung University, 2Department of Orthopedic Surgery, Chang Gung Memorial Hospital, 3Department of Physiology and Pharmacology, Chang Gung University, Taoyuan, Taiwan Abstract: A composite biodegradable polymeric model was developed to enhance tendon graft healing. This model included a biodegradable polylactide (PLA bolt as the bone anchor and a poly(D,L-lactide-co-glycolide (PLGA nanofibrous membrane embedded with collagen as a biomimic patch to promote tendon–bone interface integration. Degradation rate and compressive strength of the PLA bolt were measured after immersion in a buffer solution for 3 months. In vitro biochemical characteristics and the nanofibrous matrix were assessed using a water contact angle analyzer, pH meter, and tetrazolium reduction assay. In vivo efficacies of PLGA/collagen nanofibers and PLA bolts for tendon–bone healing were investigated on a rabbit bone tunnel model with histological and tendon pullout tests. The PLGA/collagen-blended nanofibrous membrane was a hydrophilic, stable, and biocompatible scaffold. The PLA bolt was durable for tendon–bone anchoring. Histology showed adequate biocompatibility of the PLA bolt on a medial cortex with progressive bone ingrowth and without tissue overreaction. PLGA nanofibers within the bone tunnel also decreased the tunnel enlargement phenomenon and enhanced tendon–bone integration. Composite polymers of the PLA bolt and PLGA/collagen nanofibrous membrane can effectively promote outcomes of tendon reconstruction in a rabbit model. The composite biodegradable polymeric system may be useful in humans for tendon reconstruction. Keywords: polylactide–polyglycolide nanofibers, PLGA, collagen, 3D printing, polylactide, PLA, bone-anchoring bolts, tendon healing

  19. Comparison of recombinant human bone morphogenetic protein-2-infused absorbable collagen sponge, recombinant human bone morphogenetic protein-2-coated tricalcium phosphate, and platelet-rich fibrin-mixed tricalcium phosphate for sinus augmentation in rabbits

    Directory of Open Access Journals (Sweden)

    Chul-Hun Kim

    2017-09-01

    Conclusion: Our histological evaluation demonstrates that Type I ACS can be used as a carrier of rhBMP-2, and rhBMP-2+ACS showed rapid bone formation, remodeling, and calcification at Week 2 in rabbit.

  20. Optimization of a Multi-Step Procedure for Isolation of Chicken Bone Collagen

    OpenAIRE

    Cansu, ?mran; Boran, G?khan

    2015-01-01

    Chicken bone is not adequately utilized despite its high nutritional value and protein content. Although not a common raw material, chicken bone can be used in many different ways besides manufacturing of collagen products. In this study, a multi-step procedure was optimized to isolate chicken bone collagen for higher yield and quality for manufacture of collagen products. The chemical composition of chicken bone was 2.9% nitrogen corresponding to about 15.6% protein, 9.5% fat, 14.7% mineral ...

  1. The effects of glucocorticoid on microarchitecture, collagen, mineral and mechanical properties of sheep femur cortical bone

    DEFF Research Database (Denmark)

    Ding, Ming; Danielsen, Carl C; Overgaard, Søren

    2010-01-01

    The effects of glucocorticoid on microarchitecture, collagen, mineral and mechanical properties of sheep femur cortical bone – Validation of large animal model for tissue engineering and biomaterial research Ming Ding,1* Carl Christian Danielsen,2 Søren Overgaard1 1Orthopaedic Research Laboratory......, Department of Orthopaedics and Traumatology, Odense University Hospital, Institute of Clinical Research, University of Southern Denmark, Odense C, Denmark 2Department of Connective Tissue Biology, Institute of Anatomy, University of Aarhus, Aarhus C, Denmark Osteopenia in sheep has been successfully induced...... by glucocorticoid treatment and the changes in properties of cancellous bone were comparable with those observed in humans after long-term glucocorticoid treatment. However, the influence on cortical bone has not been thoroughly elucidated. This study aimed to investigate the influence of glucocorticoid on sheep...

  2. Development of Collagen/Demineralized Bone Powder Scaffolds and Periosteum-Derived Cells for Bone Tissue Engineering Application

    Directory of Open Access Journals (Sweden)

    Wilairat Leeanansaksiri

    2013-01-01

    Full Text Available The aim of this study was to investigate physical and biological properties of collagen (COL and demineralized bone powder (DBP scaffolds for bone tissue engineering. DBP was prepared and divided into three groups, based on various particle sizes: 75–125 µm, 125–250 µm, and 250–500 µm. DBP was homogeneously mixed with type I collagen and three-dimensional scaffolds were constructed, applying chemical crosslinking and lyophilization. Upon culture with human periosteum-derived cells (PD cells, osteogenic differentiation of PD cells was investigated using alkaline phosphatase (ALP activity and calcium assay kits. The physical properties of the COL/DBP scaffolds were obviously different from COL scaffolds, irrespective of the size of DBP. In addition, PD cells cultured with COL scaffolds showed significantly higher cell adhesion and proliferation than those with COL/DBP scaffolds. In contrast, COL/DBP scaffolds exhibited greater osteoinductive potential than COL scaffolds. The PD cells with COL/DBP scaffolds possessed higher ALP activity than those with COL scaffolds. PD cells cultured with COL/DBP scaffolds with 250–500 mm particle size yielded the maximum calcium deposition. In conclusion, PD cells cultured on the scaffolds could exhibit osteoinductive potential. The composite scaffold of COL/DBP with 250–500 mm particle size could be considered a potential bone tissue engineering implant.

  3. Acceleration of bone union after structural bone grafts with a collagen-binding basic fibroblast growth factor anchored-collagen sheet for critical-size bone defects

    International Nuclear Information System (INIS)

    Ueno, Masaki; Uchida, Kentaro; Saito, Wataru; Inoue, Gen; Takahira, Naonobu; Takaso, Masashi; Matsushita, Osamu; Yogoro, Mizuki; Nishi, Nozomu; Ogura, Takayuki; Hattori, Shunji; Tanaka, Keisuke

    2014-01-01

    Bone allografts are commonly used for the repair of critical-size bone defects. However, the loss of cellular activity in processed grafts markedly reduces their healing potential compared with autografts. To overcome this obstacle, we developed a healing system for critical-size bone defects that consists of overlaying an implanted bone graft with a collagen sheet (CS) loaded with basic fibroblast growth factor (bFGF) fused to the collagen-binding domain derived from a Clostridium histolyticum collagenase (CB-bFGF). In a murine femoral defect model, defect sites treated with CS/CB-bFGF had a significantly larger callus volume than those treated with CS/native bFGF. In addition, treatment with CS/CB-bFGF resulted in the rapid formation of a hard callus bridge and a larger total callus volume at the host–graft junction than treatment with CS/bFGF. Our results suggest that the combined use of CS and CB-bFGF helps accelerate the union of allogenic bone grafts. (paper)

  4. Greener synthesis of electrospun collagen/hydroxyapatite composite fibers with an excellent microstructure for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Zhou YY

    2015-04-01

    obtained composite fibers was evaluated by in vitro culture of the human myeloma cells (U2-OS. Taken together, the process outlined herein provides an effective, non-toxic approach for the fabrication of collagen/HAP composite nanofibers that could be good candidates for bone tissue engineering.Keywords: collagen, hydroxyapatite, electrospinning, nanofibers, biomaterials

  5. Bone Collagen: New Clues to its Mineralization Mechanism From Recessive Osteogenesis Imperfecta

    Science.gov (United States)

    Eyre, David R.; Ann Weis, Mary

    2013-01-01

    Until 2006 the only mutations known to cause osteogenesis imperfecta (OI) were in the two genes coding for type I collagen chains. These dominant mutations affecting the expression or primary sequence of collagen α1(I) and α2(I) chains account for over 90% of OI cases. Since then a growing list of mutant genes causing the 5–10% of recessive cases has rapidly emerged. They include CRTAP, LEPRE1 and PPIB, which encode three proteins forming the prolyl 3-hydroxylase complex; PLOD2 and FKBP10, which encode respectively lysyl hydroxylase 2 and a foldase required for its activity in forming mature cross-links in bone collagen; SERPIN H1, which encodes the collagen chaperone HSP47; SERPIN F1, which encodes pigment epithelium-derived factor required for osteoid mineralization; and BMP1, which encodes the type I procollagen C-propeptidase. All cause fragile bone in infancy, which can include over-mineralization or under-mineralization defects as well as abnormal collagen post-translational modifications. Consistently both dominant and recessive variants lead to abnormal cross-linking chemistry in bone collagen. These recent discoveries strengthen the potential for a common pathogenic mechanism of misassembled collagen fibrils. Of the new genes identified, eight encode proteins required for collagen post-translational modification, chaperoning of newly synthesized collagen chains into native molecules or transport through the endoplasmic reticulum and Golgi for polymerization, cross-linking and mineralization. In reviewing these findings, we conclude that a common theme is emerging in the pathogenesis of brittle bone disease of mishandled collagen assembly with important insights on post-translational features of bone collagen that have evolved to optimize it as a biomineral template. PMID:23508630

  6. Effects of recombinant human collagen VI from Escherichia coli on ...

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... In this study, we reported the cloning and over expression of a gene coding for human collagen peptide. (CP6) in Escherichia coli and investigated the protective effects of CP6 on UVA-irradiated human skin fibroblasts cells. The collagen peptide (CP6) was highly soluble and the expression level was.

  7. Degradation of type IV collagen by neoplastic human skin fibroblasts

    International Nuclear Information System (INIS)

    Sheela, S.; Barrett, J.C.

    1985-01-01

    An assay for the degradation of type IV (basement membrane) collagen was developed as a biochemical marker for neoplastic cells from chemically transformed human skin fibroblasts. Type IV collagen was isolated from basement membrane of Syrian hamster lung and type I collagen was isolated from rat tails; the collagens were radioactively labelled by reductive alkylation. The abilities of normal (KD) and chemically transformed (Hut-11A) human skin fibroblasts to degrade the collagens were studied. A cell-associated assay was performed by growing either normal or transformed cells in the presence of radioactively labelled type IV collagen and measuring the released soluble peptides in the medium. This assay also demonstrated that KD cells failed to synthesize an activity capable of degrading type IV collagen whereas Hut-11A cells degraded type IV collagen in a linear manner for up to 4 h. Human serum at very low concentrations, EDTA and L-cysteine inhibited the enzyme activity, whereas protease inhibitors like phenylmethyl sulfonyl fluoride, N-ethyl maleimide or soybean trypsin inhibitor did not inhibit the enzyme from Hut-11A cells. These results suggest that the ability to degrade specifically type IV collagen may be an important marker for neoplastic human fibroblasts and supports a role for this collagenase in tumor cell invasion

  8. Collagen and mineral deposition in rabbit cortical bone during maturation and growth: effects on tissue properties.

    Science.gov (United States)

    Isaksson, Hanna; Harjula, Terhi; Koistinen, Arto; Iivarinen, Jarkko; Seppänen, Kari; Arokoski, Jari P A; Brama, Pieter A; Jurvelin, Jukka S; Helminen, Heikki J

    2010-12-01

    We characterized the composition and mechanical properties of cortical bone during maturation and growth and in adult life in the rabbit. We hypothesized that the collagen network develops earlier than the mineralized matrix. Growth was monitored, and the rabbits were euthanized at birth (newborn), and at 1, 3, 6, 9, and 18 months of age. The collagen network was assessed biochemically (collagen content, enzymatic and non-enzymatic cross-links) in specimens from the mid-diaphysis of the tibia and femur and biomechanically (tensile testing) from decalcified whole tibia specimens. The mineralized matrix was analyzed using pQCT and 3-point bend tests from intact femur specimens. The collagen content and the Young's modulus of the collagen matrix increased significantly until the rabbits were 3 months old, and thereafter remained stable. The amount of HP and LP collagen cross-links increased continuously from newborn to 18 months of age, whereas PEN cross-links increased after 6 months of age. Bone mineral density and the Young's modulus of the mineralized bone increased until the rabbits were at least 6 months old. We concluded that substantial changes take place during the normal process of development in both the biochemical and biomechanical properties of rabbit cortical bone. In cortical bone, the collagen network reaches its mature composition and mechanical strength prior to the mineralized matrix. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  9. Targeting a novel bone degradation pathway in primary bone cancer by inactivation of the collagen receptor uPARAP/Endo180

    DEFF Research Database (Denmark)

    Engelholm, Lars H; Carlsen Melander, Eva Maria; Hald, Andreas

    2016-01-01

    metastases of carcinoma origin. This opened the possibility that sarcoma cells might directly mediate bone degeneration. To examine this question, we utilized a syngeneic, osteolytic bone tumour model with transplanted NCTC-2472 sarcoma cells in mice. When analysed in vitro, these cells were capable......In osteosarcoma, a primary mesenchymal bone cancer occurring predominantly in younger patients, invasive tumour growth leads to extensive bone destruction. This process is insufficiently understood, cannot be efficiently counteracted and calls for novel means of treatment. The endocytic collagen...... receptor, uPARAP/Endo180, is expressed on various mesenchymal cell types and is involved in bone matrix turnover during normal bone growth. Human osteosarcoma specimens showed strong expression of this receptor on tumour cells, along with the collagenolytic metalloprotease, MT1-MMP. In advanced tumours...

  10. Age variations in the properties of human tibial trabecular bone

    DEFF Research Database (Denmark)

    Ding, Ming; Dalstra, M; Danielsen, CC

    1997-01-01

    We tested in compression specimens of human proximal tibial trabecular bone from 31 normal donors aged from 16 to 83 years and determined the mechanical properties, density and mineral and collagen content. Young's modulus and ultimate stress were highest between 40 and 50 years, whereas ultimate...... strain and failure energy showed maxima at younger ages. These age-related variations (except for failure energy) were non-linear. Tissue density and mineral concentration were constant throughout life, whereas apparent density (the amount of bone) varied with ultimate stress. Collagen density (the...... amount of collagen) varied with failure energy. Collagen concentration was maximal at younger ages but varied little with age. Our results suggest that the decrease in mechanical properties of trabecular bone such as Young's modulus and ultimate stress is mainly a consequence of the loss of trabecular...

  11. Collagen synthesis in human musculoskeletal tissues and skin

    DEFF Research Database (Denmark)

    Babraj, J A; Cuthbertson, D J R; Smith, K

    2005-01-01

    We have developed a direct method for the measurement of human musculoskeletal collagen synthesis on the basis of the incorporation of stable isotope-labeled proline or leucine into protein and have used it to measure the rate of synthesis of collagen in tendon, ligament, muscle, and skin....... In postabsorptive, healthy young men (28 +/- 6 yr) synthetic rates for tendon, ligament, muscle, and skin collagen were 0.046 +/- 0.005, 0.040 +/- 0.006, 0.016 +/- 0.002, and 0.037 +/- 0.003%/h, respectively (means +/- SD). In postabsorptive, healthy elderly men (70 +/- 6 yr) the rate of skeletal muscle collagen...... synthesis is greater than in the young (0.023 +/- 0.002%/h, P collagen are similar to those of mixed skeletal muscle protein in the postabsorptive state, whereas the rate for muscle collagen synthesis is much lower in both young and elderly men...

  12. Rheological, biocompatibility and osteogenesis assessment of fish collagen scaffold for bone tissue engineering.

    Science.gov (United States)

    Elango, Jeevithan; Zhang, Jingyi; Bao, Bin; Palaniyandi, Krishnamoorthy; Wang, Shujun; Wenhui, Wu; Robinson, Jeya Shakila

    2016-10-01

    In the present investigation, an attempt was made to find an alternative to mammalian collagen with better osteogenesis ability. Three types of collagen scaffolds - collagen, collagen-chitosan (CCH), and collagen-hydroxyapatite (CHA) - were prepared from the cartilage of Blue shark and investigated for their physico-functional and mechanical properties in relation to biocompatibility and osteogenesis. CCH scaffold was superior with pH 4.5-4.9 and viscosity 9.7-10.9cP. Notably, addition of chitosan and HA (hydroxyapatite) improved the stiffness (11-23MPa) and degradation rate but lowered the water binding capacity and porosity of the scaffold. Interestingly, CCH scaffolds remained for 3days before complete in-vitro biodegradation. The decreased amount of viable T-cells and higher level of FAS/APO-1 were substantiated the biocompatibility properties of prepared collagen scaffolds. Osteogenesis study revealed that the addition of CH and HA in both fish and mammalian collagen scaffolds could efficiently promote osteoblast cell formation. The ALP activity was significantly high in CHA scaffold-treated osteoblast cells, which suggests an enhanced bone-healing process. Therefore, the present study concludes that the composite scaffolds prepared from fish collagen with higher stiffness, lower biodegradation rate, better biocompatible, and osteogenesis properties were suitable biomaterial for a bone tissue engineering application as an alternative to mammalian collagen scaffolds. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Preparation and structure characterization of soluble bone collagen ...

    African Journals Online (AJOL)

    In this study, G-25 gel chromatography, X-diffraction, scanning electron microscopy (SEM), UV and Fourier transform infrared spectroscopy (FTIR) were used to analyze soluble collagen peptides chelating calcium. Collagen peptide hydrolysis can be divided into four components using G-25 gel chromatography.

  14. Microfibrous {beta}-TCP/collagen scaffolds mimic woven bone in structure and composition

    Energy Technology Data Exchange (ETDEWEB)

    Zhang Shen; Zhang Xin; Cai Qing; Yang Xiaoping [Key Laboratory of Beijing City on Preparation and Processing of Novel Polymer Materials, College of Materials Science and Engineering, Beijing University of Chemical Technology, Beijing 100029 (China); Wang Bo; Deng Xuliang, E-mail: yangxp@mail.buct.edu.c [Department of VIP Dental Service, School and Hospital of Stomatology, Peking University, Beijing 100081 (China)

    2010-12-15

    Woven bone, as the initial form of bone tissue, is always found in developing and repairing bone. It is thought of as a temporary scaffold for the deposition of osteogenic cells and the laying down of lamellar bone. Thus, we hypothesize that a matrix which resembles the architecture and components of woven bone can provide an osteoblastic microenvironment for bone cell growth and new bone formation. In this study, woven-bone-like beta-tricalcium phosphate ({beta}-TCP)/collagen scaffolds were fabricated by sol-gel electrospinning and impregnating methods. Optimization studies on sol-gel synthesis and electrospinning process were conducted respectively to prepare pure {beta}-TCP fibers with dimensions close to mineralized collagen fibrils in woven bone. The collagen-coating layer prepared by impregnation had an adhesive role that held the {beta}-TCP fibers together, and resulted in rapid degradation and matrix mineralization in in vitro tests. MG63 osteoblast-like cells seeded on the resultant scaffolds showed three-dimensional (3D) morphologies, and merged into multicellular layers after 7 days culture. Cytotoxicity test further revealed that extracts from the resultant scaffolds could promote the proliferation of MG63 cells. Therefore, the woven-bone-like matrix that we constructed favored the attachment and proliferation of MG63 cells in three dimensions. It has great potential ability to shorten the time of formation of new bone.

  15. Microfibrous β-TCP/collagen scaffolds mimic woven bone in structure and composition

    International Nuclear Information System (INIS)

    Zhang Shen; Zhang Xin; Cai Qing; Yang Xiaoping; Wang Bo; Deng Xuliang

    2010-01-01

    Woven bone, as the initial form of bone tissue, is always found in developing and repairing bone. It is thought of as a temporary scaffold for the deposition of osteogenic cells and the laying down of lamellar bone. Thus, we hypothesize that a matrix which resembles the architecture and components of woven bone can provide an osteoblastic microenvironment for bone cell growth and new bone formation. In this study, woven-bone-like beta-tricalcium phosphate (β-TCP)/collagen scaffolds were fabricated by sol-gel electrospinning and impregnating methods. Optimization studies on sol-gel synthesis and electrospinning process were conducted respectively to prepare pure β-TCP fibers with dimensions close to mineralized collagen fibrils in woven bone. The collagen-coating layer prepared by impregnation had an adhesive role that held the β-TCP fibers together, and resulted in rapid degradation and matrix mineralization in in vitro tests. MG63 osteoblast-like cells seeded on the resultant scaffolds showed three-dimensional (3D) morphologies, and merged into multicellular layers after 7 days culture. Cytotoxicity test further revealed that extracts from the resultant scaffolds could promote the proliferation of MG63 cells. Therefore, the woven-bone-like matrix that we constructed favored the attachment and proliferation of MG63 cells in three dimensions. It has great potential ability to shorten the time of formation of new bone.

  16. Distribution of Type I Collagen Morphologies in Bone: Relation to Estrogen Depletion

    Science.gov (United States)

    Wallace, Joseph M.; Erickson, Blake; Les, Clifford M.; Orr, Bradford G.; Holl, Mark M. Banaszak

    2009-01-01

    Bone is an amazing material evolved by nature to elegantly balance structural and metabolic needs in the body. Bone health is an integral part of overall health, but our lack of understanding of the ultrastructure of healthy bone precludes us from knowing how disease may impact nanoscale properties in this biological material. Here, we show that quantitative assessments of a distribution of Type I collagen fibril morphologies can be made using atomic force microscopy (AFM). We demonstrate that normal bone contains a distribution of collagen fibril morphologies and that changes in this distribution can be directly related to disease state. Specifically, by monitoring changes in the collagen fibril distribution of sham-operated and estrogen-depleted sheep, we have shown the ability to detect estrogen-deficiency-induced changes in Type I collagen in bone. This discovery provides new insight into the ultrastructure of bone as a tissue and the role of material structure in bone disease. The observation offers the possibility of a much-needed in vitro procedure to complement the current methods used to diagnose osteoporosis and other bone disease. PMID:19932773

  17. In Vitro and In Vivo Study of a Novel Porcine Collagen Membrane for Guided Bone Regeneration

    Directory of Open Access Journals (Sweden)

    Eisner Salamanca

    2016-11-01

    Full Text Available For years, in order to improve bone regeneration and prevent the need of a second stage surgery to remove non-resorbable membranes, biological absorbable membranes have gradually been developed and applied in guided tissue regeneration (GTR. The present study’s main objective was to achieve space maintenance and bone regeneration using a new freeze-dried developed porcine collagen membrane, and compare it with an already commercial collagen membrane, when both were used with a bovine xenograft in prepared alveolar ridge bone defects. Prior to surgery, the membrane’s vitality analysis showed statistically significant higher cell proliferation in the test membrane over the commercial one. In six beagle dogs, commercial bone xenograft was packed in lateral ridge bone defects prepared in the left and right side and then covered with test porcine collagen membrane or commercial collagen membrane. Alveolar height changes were measured. Histomorphometric results, in vitro and in vivo properties indicated that the new porcine collagen membrane is biocompatible, enhances bone xenograft osteoconduction, and reduces the alveolar ridge height reabsorption rate.

  18. The biochemical changes of bone collagen after high-dose irradiation

    International Nuclear Information System (INIS)

    Tajiri, Ken

    1980-01-01

    In our clinic, patients with malignant bone tumors have been treated by high-dose irradiation therapy, 10,000-20,000 rads, for primary lesions. In order to study the biochemical changes of normal bone around tumor tissue, especially bone collagen, after high-dose irradiation, the author performed the following experiments. The right knee joint of rabbits was irradiated with either 6,000, 10,000, or 15,000 rads by 60 Co-γ ray. The cortical bone of the right tibial metaphysis was used for analyses and compared with the left tibia of the same rabbit. These studies were followed for one year after the final irradiation. The calcium, phosphorous and collagen contents of irradiated bone were remarkably changed. These data indicate that collagen biosynthesis of irradiated bone was decreased and the calcification was disturbed. An increase in the amount of total soluble collagen and a decrease in the amount of hydroxylysine bound sugar were observed. The ratio of β to α chains of the collagen molecule was also changed by the irradiation. The amount of reducible cross-links per hydroxyproline residue was strikingly increased three months after the final irradiation. These changes were remarkable especially in the 10,000 and 15,000 rads irradiated group and found to be recovered approximately six months to one year after the final irradiation. These findings indicate that high-dose irradiation reduces the stability of bone collagen both with the destruction of sugar bonds of hydroxylysine residues and the replacement of matured collagen matrix to immatured one which contain mostly labile reducible cross-links. (author)

  19. Development of a nanofiltration method for bone collagen {sup 14}C AMS dating

    Energy Technology Data Exchange (ETDEWEB)

    Boudin, Mathieu, E-mail: mathieu.boudin@ugent.be [Royal Institute for Cultural Heritage, Jubelpark 1, B-1000 Brussels (Belgium); Ghent University, Faculty of Bioscience Engineering, Laboratory of Applied Physical Chemistry, Coupure Links 653, B-9000 Ghent (Belgium); Boeckx, Pascal [Ghent University, Faculty of Bioscience Engineering, Laboratory of Applied Physical Chemistry, Coupure Links 653, B-9000 Ghent (Belgium); Buekenhoudt, Anita [Flemish Institute for Technological Research, Separation and Conversion Technology, Boeretang 200, B-2400 Mol (Belgium); Vandenabeele, Peter [Ghent University, Department of Archaeology, Sint-Pietersnieuwstraat 35, B-9000 Ghent (Belgium); Van Strydonck, Mark [Royal Institute for Cultural Heritage, Jubelpark 1, B-1000 Brussels (Belgium)

    2013-01-15

    Radiocarbon dating of bones is usually performed on the collagen fraction. However, this collagen can contain exogenous molecules, including humic substances (HSs) and/or other soil components that may have a different age than the bone. Incomplete removal can result in biased {sup 14}C dates. Ultrafiltration of collagen, dissolved as gelatin (molecular weight (MW) {approx}100,000 Dalton), has received considerable attention to obtain more reliable dates. Ultrafiltration is an effective method of removal of low-molecular weight contaminants from bone collagen but it does not remove high-molecular weight contaminants, such as cross-linked humic collagen complexes. However, comparative dating studies have raised the question whether this cleaning step itself may introduce contamination with carbon from the filters used. In this study, a nanofiltration method was developed using a ceramic filter to avoid a possible extraneous carbon contamination introduced by the filter. This method should be applicable to various protein materials e.g. collagen, silk, wool, leather and should be able to remove low-molecular and high molecular weight HSs. In this study bone collagen was hot acid hydrolyzed to amino acids and nanofiltrated. A filter with a molecular weight cutoff (MWCO) of 450 Dalton was chosen in order to collect the amino acids in the permeate and the HSs in the retentate. Two pilot studies were set up. Two nanofiltration types were tested in pilot study 1: dead end and cross flow filtration. Humic substance (HS)-solutions with fossil carbon and modern hydrolyzed collagen contaminated with HSs were filtrated and analyzed with spectrofluorescence to determine the HS removal. Cross flow nanofiltration showed the most efficient HS removal. A second pilot study based upon these results was set up wherein only cross flow filtration was performed. {sup 14}C measurements of the permeates of hydrolyzed modern collagen contaminated with fossil HSs demonstrate a significant

  20. Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

    Science.gov (United States)

    Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

    2016-12-01

    Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process.

  1. Biodegradation, biocompatibility, and osteoconduction evaluation of collagen-nanohydroxyapatite cryogels for bone tissue regeneration.

    Science.gov (United States)

    Salgado, Christiane Laranjo; Grenho, Liliana; Fernandes, Maria Helena; Colaço, Bruno Jorge; Monteiro, Fernando Jorge

    2016-01-01

    Designing biomimetic biomaterials inspired by the natural complex structure of bone and other hard tissues is still a challenge nowadays. The control of the biomineralization process onto biomaterials should be evaluated before clinical application. Aiming at bone regeneration applications, this work evaluated the in vitro biodegradation and interaction between human bone marrow stromal cells (HBMSC) cultured on different collagen/nanohydroxyapatite cryogels. Cell proliferation, differentiation, morphology, and metabolic activity were assessed through different protocols. All the biocomposite materials allowed physiologic apatite deposition after incubation in simulated body fluid and the cryogel with the highest nanoHA content showed to have the highest mechanical strength (DMA). The study clearly showed that the highest concentration of nanoHA granules on the cryogels were able to support cell type's survival, proliferation, and individual functionality in a monoculture system, for 21 days. In fact, the biocomposites were also able to differentiate HBMSCs into osteoblastic phenotype. The composites behavior was also assessed in vivo through subcutaneous and bone implantation in rats to evaluate its tissue-forming ability and degradation rate. The cryogels Coll/nanoHA (30 : 70) promoted tissue regeneration and adverse reactions were not observed on subcutaneous and bone implants. The results achieved suggest that scaffolds of Coll/nanoHA (30 : 70) should be considered promising implants for bone defects that present a grotto like appearance with a relatively small access but a wider hollow inside. This material could adjust to small dimensions and when entering into the defect, it could expand inside and remain in close contact with the defect walls, thus ensuring adequate osteoconductivity. © 2015 Wiley Periodicals, Inc.

  2. Applying Knowledge on Collagen of CLRI: In Human Health Care

    Indian Academy of Sciences (India)

    Applying Knowledge on Collagen of CLRI: In Human Health Care ... Kollagen & NeuSkin are products in the market based on technologies. ... derived products of biomedical value in tissue remodeling and engineering are in advanced stage ...

  3. Probabilistic failure analysis of bone using a finite element model of mineral-collagen composites.

    Science.gov (United States)

    Dong, X Neil; Guda, Teja; Millwater, Harry R; Wang, Xiaodu

    2009-02-09

    Microdamage accumulation is a major pathway for energy dissipation during the post-yield deformation of bone. In this study, a two-dimensional probabilistic finite element model of a mineral-collagen composite was developed to investigate the influence of the tissue and ultrastructural properties of bone on the evolution of microdamage from an initial defect in tension. The probabilistic failure analyses indicated that the microdamage progression would be along the plane of the initial defect when the debonding at mineral-collagen interfaces was either absent or limited in the vicinity of the defect. In this case, the formation of a linear microcrack would be facilitated. However, the microdamage progression would be scattered away from the initial defect plane if interfacial debonding takes place at a large scale. This would suggest the possible formation of diffuse damage. In addition to interfacial debonding, the sensitivity analyses indicated that the microdamage progression was also dependent on the other material and ultrastructural properties of bone. The intensity of stress concentration accompanied with microdamage progression was more sensitive to the elastic modulus of the mineral phase and the nonlinearity of the collagen phase, whereas the scattering of failure location was largely dependent on the mineral to collagen ratio and the nonlinearity of the collagen phase. The findings of this study may help understanding the post-yield behavior of bone at the ultrastructural level and shed light on the underlying mechanism of bone fractures.

  4. Nitrogen and carbon isotopic composition of bone collagen from marine and terrestrial animals

    International Nuclear Information System (INIS)

    Schoeninger, M.J.; DeNiro, M.J.

    1984-01-01

    The stable nitrogen and carbon isotope ratios of bone collagen prepared from more than 100 animals representing 66 species of birds, fish, and mammals are presented. The delta 15 N values of bone collagen from animals that fed exclusively in the marine environment are, on average, 9 per mille more positive than those from animals that fed exclusively in the terrestrial environment: ranges for the two groups overlap by less than 1 per mille. Bone collagen delta 15 N values also serve to separate marine fish from the small number of freshwater fish we analyzed. The bone collagen delta 15 N values of birds and fish that spent part of their life cycles feeding in the marine environment and part in the freshwater environment are intermediate between those of animals that fed exclusively in one or the other system. Further, animals that fed at successive trophic levels in the marine and terrestrial environment are separated, on average, by a 3 per mille difference in the delta 15 N values of their bone collagen. Results are given and discussed. (author)

  5. Changes in tissue morphology and collagen composition during the repair of cortical bone in the adult chicken.

    Science.gov (United States)

    Glimcher, M J; Shapiro, F; Ellis, R D; Eyre, D R

    1980-09-01

    An animal model was developed to study the histology and collagen chemistry of healing cortical bone. A hole was cut through the cortex of the mid-shaft of the humerus of the adult chicken, which allowed for repair at a mechanically stable site. After one to two weeks the collagen of the repair tissue, which consisted principally of woven bone, contained almost three times as much hydroxylysine as the collagen of normal adult bone and thus resembled the collagen of embryonic long bones. By eight weeks, when lamellar one predominated, the hydroxylysine content had fallen to normal levels. Type I was the major genetic type of collagen present throughout. No type-II collagen, characteristic of cartilage, was detected; this was consistent with the histological findings. The results established that hydroxylysine-rich type-I collagen can be made by osteoblasts of adult animals as well as by those of embryos and early postnates. In order to understand the biological characteristics of fracture healing, it is vital to study not only the macroscopic organization of the repair tissue but also the chemical properties of its molecular components. The strength of healing fractured bone, and indeed of normal bone, depends largely on the properties of the structural protein collagen. To date, it is not known whether the collagen in healing fractures is the same as that in normal bone, or whether it has distinct chemical features that may suit it for bone repair.

  6. Comparative Efficacies of Collagen-Based 3D Printed PCL/PLGA/?-TCP Composite Block Bone Grafts and Biphasic Calcium Phosphate Bone Substitute for Bone Regeneration

    OpenAIRE

    Hwang, Kyoung-Sub; Choi, Jae-Won; Kim, Jae-Hun; Chung, Ho Yun; Jin, Songwan; Shim, Jin-Hyung; Yun, Won-Soo; Jeong, Chang-Mo; Huh, Jung-Bo

    2017-01-01

    The purpose of this study was to compare bone regeneration and space maintaining ability of three-dimensional (3D) printed bone grafts with conventional biphasic calcium phosphate (BCP). After mixing polycaprolactone (PCL), poly (lactic-co-glycolic acid) (PLGA), and ?-tricalcium phosphate (?-TCP) in a 4:4:2 ratio, PCL/PLGA/?-TCP particulate bone grafts were fabricated using 3D printing technology. Fabricated particulate bone grafts were mixed with atelocollagen to produce collagen-based PCL/P...

  7. Checking collagen preservation in archaeological bone by non-destructive studies (Micro-CT and IBA)

    Science.gov (United States)

    Beck, L.; Cuif, J.-P.; Pichon, L.; Vaubaillon, S.; Dambricourt Malassé, A.; Abel, R. L.

    2012-02-01

    The material to be studied is a piece of human skull discovered (1999) in Pleistocene sediments from the Orsang river (Gujarat state, India). From anatomical view point, this skull is highly composite: modern Homo sapiens characters are associated to undoubtedly more ancient features. Absolute dating by 14C is critical to understand this discovery. Prior to dating measurements, non-destructive studies have been carried out. Micro-CT reconstruction (X-ray microtomography) and Ion Beam Analysis (IBA) have been undertaken to check the structural preservation of the fossil and the collagen preservation. PIXE elemental map was used to select well-preserved bone area. RBS/EBS and NRA were used for light element quantification, in particular C, N and O contents. We also demonstrate that the PIXE-RBS/EBS combination is a effective tool for the whole characterization of archaeological and recent bones by analysing in one experiment both mineral and organic fractions. We have shown that the archaeological bone, a fragment of the potentially oldest modern Indian, is enough preserved for radiocarbon dating. We propose that Elastic Backscattering Spectrometry (EBS) using 3 MeV protons could be a good non destructive alternative to conventional CHN method using Carbon-Hydrogen-Nitrogen analyzer for measuring C and N before 14C dating.

  8. Checking collagen preservation in archaeological bone by non-destructive studies (Micro-CT and IBA)

    Energy Technology Data Exchange (ETDEWEB)

    Beck, L., E-mail: lucile.beck@cea.fr [C2RMF - UMR171 CNRS, Centre de Recherche et de Restauration des Musees de France, Palais du Louvre, Porte des Lions, 14 quai Francois Mitterrand, 75001 Paris (France); CEA, DEN, Service de Recherches de Metallurgie Physique, Laboratoire JANNUS, 91191 Gif-sur-Yvette (France); Cuif, J.-P. [UMR IDES 8148, Universite Paris XI-Orsay, 91405 Orsay cedex (France); Pichon, L. [C2RMF - UMR171 CNRS, Centre de Recherche et de Restauration des Musees de France, Palais du Louvre, Porte des Lions, 14 quai Francois Mitterrand, 75001 Paris (France); Vaubaillon, S. [CEA, INSTN, Laboratoire JANNUS, 91191 Gif-sur-Yvette (France); Dambricourt Malasse, A. [Departement de Prehistoire, Museum national d' Histoire naturelle, UMR 7194 - CNRS, Institut de Paleontologie Humaine, 1, rue Rene Panhard, 75013 Paris (France); Abel, R.L. [The Natural History Museum, London (United Kingdom)

    2012-02-15

    The material to be studied is a piece of human skull discovered (1999) in Pleistocene sediments from the Orsang river (Gujarat state, India). From anatomical view point, this skull is highly composite: modern Homo sapiens characters are associated to undoubtedly more ancient features. Absolute dating by {sup 14}C is critical to understand this discovery. Prior to dating measurements, non-destructive studies have been carried out. Micro-CT reconstruction (X-ray microtomography) and Ion Beam Analysis (IBA) have been undertaken to check the structural preservation of the fossil and the collagen preservation. PIXE elemental map was used to select well-preserved bone area. RBS/EBS and NRA were used for light element quantification, in particular C, N and O contents. We also demonstrate that the PIXE-RBS/EBS combination is a effective tool for the whole characterization of archaeological and recent bones by analysing in one experiment both mineral and organic fractions. We have shown that the archaeological bone, a fragment of the potentially oldest modern Indian, is enough preserved for radiocarbon dating. We propose that Elastic Backscattering Spectrometry (EBS) using 3 MeV protons could be a good non destructive alternative to conventional CHN method using Carbon-Hydrogen-Nitrogen analyzer for measuring C and N before {sup 14}C dating.

  9. Checking collagen preservation in archaeological bone by non-destructive studies (Micro-CT and IBA)

    International Nuclear Information System (INIS)

    Beck, L.; Cuif, J.-P.; Pichon, L.; Vaubaillon, S.; Dambricourt Malassé, A.; Abel, R.L.

    2012-01-01

    The material to be studied is a piece of human skull discovered (1999) in Pleistocene sediments from the Orsang river (Gujarat state, India). From anatomical view point, this skull is highly composite: modern Homo sapiens characters are associated to undoubtedly more ancient features. Absolute dating by 14 C is critical to understand this discovery. Prior to dating measurements, non-destructive studies have been carried out. Micro-CT reconstruction (X-ray microtomography) and Ion Beam Analysis (IBA) have been undertaken to check the structural preservation of the fossil and the collagen preservation. PIXE elemental map was used to select well-preserved bone area. RBS/EBS and NRA were used for light element quantification, in particular C, N and O contents. We also demonstrate that the PIXE-RBS/EBS combination is a effective tool for the whole characterization of archaeological and recent bones by analysing in one experiment both mineral and organic fractions. We have shown that the archaeological bone, a fragment of the potentially oldest modern Indian, is enough preserved for radiocarbon dating. We propose that Elastic Backscattering Spectrometry (EBS) using 3 MeV protons could be a good non destructive alternative to conventional CHN method using Carbon–Hydrogen–Nitrogen analyzer for measuring C and N before 14 C dating.

  10. Effects of increased collagen-matrix density on the mechanical properties and in vivo absorbability of hydroxyapatite-collagen composites as artificial bone materials

    Energy Technology Data Exchange (ETDEWEB)

    Yunoki, Shunji [Life Science Group, Tokyo Metropolitan Industrial Technology Research Institute, 2-11-1 Fukasawa, Setagaya-ku, Tokyo 158-0081 (Japan); Sugiura, Hiroaki; Kondo, Eiji; Yasuda, Kazunori [Department of Sports Medicine and Joint Surgery, Graduate School of Medicine, Hokkaido University, Kita-15 Nishi-7, Sapporo, Hokkaido 060-8638 Japan (Japan); Ikoma, Toshiyuki; Tanaka, Junzo, E-mail: yunoki.shunji@iri-tokyo.jp [Department of Metallurgy and Ceramics Science, 2-12-1-S7-1, Ookayama, Meguro-ku, Tokyo 152-8550 (Japan)

    2011-02-15

    The aim of this study was to evaluate the effects of increased collagen-matrix density on the mechanical properties and in vivo absorbability of porous hydroxyapatite (HAp)-collagen composites as artificial bone materials. Seven types of porous HAp-collagen composites were prepared from HAp nanocrystals and dense collagen fibrils. Their densities and HAp/collagen weight ratios ranged from 122 to 331 mg cm{sup -3} and from 20/80 to 80/20, respectively. The flexural modulus and strength increased with an increase in density, reaching 2.46 {+-} 0.48 and 0.651 {+-} 0.103 MPa, respectively. The porous composites with a higher collagen-matrix density exhibited much higher mechanical properties at the same densities, suggesting that increasing the collagen-matrix density is an effective way of improving the mechanical properties. It was also suggested that other structural factors in addition to collagen-matrix density are required to achieve bone-like mechanical properties. The in vivo absorbability of the composites was investigated in bone defects of rabbit femurs, demonstrating that the absorption rate decreased with increases in the composite density. An exhaustive increase in density is probably limited by decreases in absorbability as artificial bones.

  11. Effects of increased collagen-matrix density on the mechanical properties and in vivo absorbability of hydroxyapatite-collagen composites as artificial bone materials

    International Nuclear Information System (INIS)

    Yunoki, Shunji; Sugiura, Hiroaki; Kondo, Eiji; Yasuda, Kazunori; Ikoma, Toshiyuki; Tanaka, Junzo

    2011-01-01

    The aim of this study was to evaluate the effects of increased collagen-matrix density on the mechanical properties and in vivo absorbability of porous hydroxyapatite (HAp)-collagen composites as artificial bone materials. Seven types of porous HAp-collagen composites were prepared from HAp nanocrystals and dense collagen fibrils. Their densities and HAp/collagen weight ratios ranged from 122 to 331 mg cm -3 and from 20/80 to 80/20, respectively. The flexural modulus and strength increased with an increase in density, reaching 2.46 ± 0.48 and 0.651 ± 0.103 MPa, respectively. The porous composites with a higher collagen-matrix density exhibited much higher mechanical properties at the same densities, suggesting that increasing the collagen-matrix density is an effective way of improving the mechanical properties. It was also suggested that other structural factors in addition to collagen-matrix density are required to achieve bone-like mechanical properties. The in vivo absorbability of the composites was investigated in bone defects of rabbit femurs, demonstrating that the absorption rate decreased with increases in the composite density. An exhaustive increase in density is probably limited by decreases in absorbability as artificial bones.

  12. Comparative effectiveness of using resorbable membranes of polylactic acid and collagen in regeneration of bone defects in patients with periimplantitis

    Directory of Open Access Journals (Sweden)

    Gudaryan A.A.

    2014-03-01

    Full Text Available The article presents the results of comparative study of effectiveness of usage of separation membranes from polylactic acid (PLA and collagen in carrying out targeted regeneration of bone tissue in 22 patients with periimplantitis. Purpose: To conduct a comparative clinico-radiological efficiency of PLA membrane and collagen membranes in removing bone defects of the alveolar bone in patients with periimplantitis in clinic. It was found that depending on the type of membrane, bone tissue growth occurs not in the same way. Surgery in treatment of periimplantitis using osteo-inducing agent «Bio-Oss» and PLA membranes allows to reach full recovery of bone in bone defects in 90.9 % of patients versus 63.63 % of cases with collagen membranes. Thus, reconstitution of bone in bone defects in periimplantitis is more of full value in using PLA membranes than with membranes from collagen.

  13. Polycaprolactone nanofiber interspersed collagen type-I scaffold for bone regeneration: a unique injectable osteogenic scaffold

    International Nuclear Information System (INIS)

    Baylan, Nuray; Ditto, Maggie; Lawrence, Joseph G; Yildirim-Ayan, Eda; Bhat, Samerna; Lecka-Czernik, Beata

    2013-01-01

    There is an increasing demand for an injectable cell coupled three-dimensional (3D) scaffold to be used as bone fracture augmentation material. To address this demand, a novel injectable osteogenic scaffold called PN-COL was developed using cells, a natural polymer (collagen type-I), and a synthetic polymer (polycaprolactone (PCL)). The injectable nanofibrous PN-COL is created by interspersing PCL nanofibers within pre-osteoblast cell embedded collagen type-I. This simple yet novel and powerful approach provides a great benefit as an injectable bone scaffold over other non-living bone fracture stabilization polymers, such as polymethylmethacrylate and calcium content resin-based materials. The advantages of injectability and the biomimicry of collagen was coupled with the structural support of PCL nanofibers, to create cell encapsulated injectable 3D bone scaffolds with intricate porous internal architecture and high osteoconductivity. The effects of PCL nanofiber inclusion within the cell encapsulated collagen matrix has been evaluated for scaffold size retention and osteocompatibility, as well as for MC3T3-E1 cells osteogenic activity. The structural analysis of novel bioactive material proved that the material is chemically stable enough in an aqueous solution for an extended period of time without using crosslinking reagents, but it is also viscous enough to be injected through a syringe needle. Data from long-term in vitro proliferation and differentiation data suggests that novel PN-COL scaffolds promote the osteoblast proliferation, phenotype expression, and formation of mineralized matrix. This study demonstrates for the first time the feasibility of creating a structurally competent, injectable, cell embedded bone tissue scaffold. Furthermore, the results demonstrate the advantages of mimicking the hierarchical architecture of native bone with nano- and micro-size formation through introducing PCL nanofibers within macron-size collagen fibers and in

  14. Study on de novo collagen biosynthesis and degradation markers of bone

    International Nuclear Information System (INIS)

    Hanna, L.S.; Matta, T.F.; Ibrahim, I.; Meky, N.H.

    2003-01-01

    This investigation was carried out to study the performance of de novo biochemical markers of serum pro collagen type-1 amino terminal extension (PINP), as a marker of collagen biosynthesis, and urinary collagen crosslink free deoxypyridinoline (DPD) as a marker of collagen degradation. Moreover, urinary calcium C Ca) and inorganic phosphorus (P), as markers of bone demineralization, in addition to urinary creatinine (Cr), to reflect status of renal function, were also studied in order to assess the activity of bone turnover in osteoporotic (OST), postmenopausal (POST), peri menopausal(PERI), premenopausal (PRE) women and also in young adult (YON) ones. The obtained results showed that urinary creatinine levels were within the normal ranges in all women even in the elderly osteoporotic and postmenopausal women. Serum PINP did not reflect osteoblastic activity. Urinary DPD proved to be a good marker in monitoring the postmenopausal bone resorption and urinary Ca was a reliable marker for bone loss in osteoporosis and bone turnover in the postmenopausal status

  15. Differential actions of the endocytic collagen receptor uPARAP/Endo180 and the collagenase MMP-2 in bone homeostasis

    DEFF Research Database (Denmark)

    Madsen, Daniel H; Jürgensen, Henrik J; Ingvarsen, Signe

    2013-01-01

    A well-coordinated remodeling of uncalcified collagen matrices is a pre-requisite for bone development and homeostasis. Collagen turnover proceeds through different pathways, either involving extracellular reactions exclusively, or being dependent on endocytic processes. Extracellular collagen...... degradation requires the action of secreted or membrane attached collagenolytic proteases, whereas the alternative collagen degradation pathway proceeds intracellularly after receptor-mediated uptake and delivery to the lysosomes. In this study we have examined the functional interplay between...

  16. In vitro deposition of hydroxyapatite on cortical bone collagen stimulated by deformation-induced piezoelectricity.

    Science.gov (United States)

    Noris-Suárez, Karem; Lira-Olivares, Joaquin; Ferreira, Ana Marina; Feijoo, José Luis; Suárez, Nery; Hernández, Maria C; Barrios, Esteban

    2007-03-01

    In the present work, we have studied the effect of the piezoelectricity of elastically deformed cortical bone collagen on surface using a biomimetic approach. The mineralization process induced as a consequence of the piezoelectricity effect was evaluated using scanning electron microscopy (SEM), thermally stimulated depolarization current (TSDC), and differential scanning calorimetry (DSC). SEM micrographs showed that mineralization occurred predominantly over the compressed side of bone collagen, due to the effect of piezoelectricity, when the sample was immersed in the simulated body fluid (SBF) in a cell-free system. The TSDC method was used to examine the complex collagen dielectric response. The dielectric spectra of deformed and undeformed collagen samples with different hydration levels were compared and correlated with the mineralization process followed by SEM. The dielectric measurements showed that the mineralization induced significant changes in the dielectric spectra of the deformed sample. DSC and TSDC results demonstrated a reduction of the collagen glass transition as the mineralization process advanced. The combined use of SEM, TSDC, and DSC showed that, even without osteoblasts present, the piezoelectric dipoles produced by deformed collagen can produce the precipitation of hydroxyapatite by electrochemical means, without a catalytic converter as occurs in classical biomimetic deposition.

  17. Novel Vanadium-Loaded Ordered Collagen Scaffold Promotes Osteochondral Differentiation of Bone Marrow Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Ana M. Cortizo

    2016-01-01

    Full Text Available Bone and cartilage regeneration can be improved by designing a functionalized biomaterial that includes bioactive drugs in a biocompatible and biodegradable scaffold. Based on our previous studies, we designed a vanadium-loaded collagen scaffold for osteochondral tissue engineering. Collagen-vanadium loaded scaffolds were characterized by SEM, FTIR, and permeability studies. Rat bone marrow progenitor cells were plated on collagen or vanadium-loaded membranes to evaluate differences in cell attachment, growth and osteogenic or chondrocytic differentiation. The potential cytotoxicity of the scaffolds was assessed by the MTT assay and by evaluation of morphological changes in cultured RAW 264.7 macrophages. Our results show that loading of VOAsc did not alter the grooved ordered structure of the collagen membrane although it increased membrane permeability, suggesting a more open structure. The VOAsc was released to the media, suggesting diffusion-controlled drug release. Vanadium-loaded membranes proved to be a better substratum than C0 for all evaluated aspects of BMPC biocompatibility (adhesion, growth, and osteoblastic and chondrocytic differentiation. In addition, there was no detectable effect of collagen or vanadium-loaded scaffolds on macrophage viability or cytotoxicity. Based on these findings, we have developed a new ordered collagen scaffold loaded with VOAsc that shows potential for osteochondral tissue engineering.

  18. Exercise and Regulation of Bone and Collagen Tissue Biology.

    Science.gov (United States)

    Kjaer, Michael; Jørgensen, Niklas Rye; Heinemeier, Katja; Magnusson, S Peter

    2015-01-01

    The musculoskeletal system and its connective tissue include the intramuscular connective tissue, the myotendinous junction, the tendon, the joints with their cartilage and ligaments, and the bone; they all together play a crucial role in maintaining the architecture of the skeletal muscle, ensuring force transmission, storing energy, protecting joint surface and stability, and ensuring the transfer of muscular forces into resulting limb movement. The musculoskeletal connective tissue structure is relatively stable, but mechanical loading and subsequent mechanotransduction and molecular anabolic signaling can result in some adaptation of the connective tissue, its size, its strength, and its mechanical properties, whereby it can improve its capacity by 5-20% with regular physical activity. For several of the mechanically loaded connective tissues, only limited information regarding molecular and cellular signaling pathways and their adaptation to exercise is available. In contrast to tissue responses with exercise, lack of mechanical tissue loading through inactivity or immobilization of the human body will result in a dramatic loss of connective tissue content, structure, and tolerable load within weeks, to a degree (30-40%) that mimics that of contractile skeletal musculature. This illustrates the importance of regular mechanical load in order to preserve the stabilizing role of the connective tissue for the overall function of the musculoskeletal system in both daily activity and exercise. © 2015 Elsevier Inc. All rights reserved.

  19. Validating in vivo Raman spectroscopy of bone in human subjects

    Science.gov (United States)

    Esmonde-White, Francis W. L.; Morris, Michael D.

    2013-03-01

    Raman spectroscopy can non-destructively measure properties of bone related to mineral density, mineral composition, and collagen composition. Bone properties can be measured through the skin in animal and human subjects, but correlations between the transcutaneous and exposed bone measurements have only been reported for human cadavers. In this study, we examine human subjects to collect measurements transcutaneously, on surgically exposed bone, and on recovered bone fragments. This data will be used to demonstrate in vivo feasibility and to compare transcutaneous and exposed Raman spectroscopy of bone. A commercially available Raman spectrograph and optical probe operating at 785 nm excitation are used for the in vivo measurements. Requirements for applying Raman spectroscopy during a surgery are also discussed.

  20. A new procedure for extraction of collagen from modern and archaeological bones for 14C dating.

    Science.gov (United States)

    Maspero, F; Sala, S; Fedi, M E; Martini, M; Papagni, A

    2011-10-01

    Bones are potentially the best age indicators in a stratigraphic study, because they are closely related to the layer in which they are found. Collagen is the most suitable fraction and is the material normally used in radiocarbon dating. Bone contaminants can strongly alter the carbon isotopic fraction values of the samples, so chemical pretreatment for (14)C dating by accelerator mass spectrometry (AMS) is essential. The most widespread method for collagen extraction is based on the Longin procedure, which consists in HCl demineralization to dissolve the inorganic phase of the samples, followed by dissolution of collagen in a weak acid solution. In this work the possible side effects of this procedure on a modern bone are presented; the extracted collagen was analyzed by ATR-IR spectroscopy. An alternative procedure, based on use of HF instead of HCl, to minimize unwanted degradation of the organic fraction, is also given. A study by ATR-IR spectroscopic analysis of collagen collected after different demineralization times and with different acid volumes, and a study of an archaeological sample, are also presented.

  1. Influence of Nano-HA Coated Bone Collagen to Acrylic (Polymethylmethacrylate Bone Cement on Mechanical Properties and Bioactivity.

    Directory of Open Access Journals (Sweden)

    Tao Li

    Full Text Available This research investigated the mechanical properties and bioactivity of polymethylmethacrylate (PMMA bone cement after addition of the nano-hydroxyapatite(HA coated bone collagen (mineralized collagen, MC.The MC in different proportions were added to the PMMA bone cement to detect the compressive strength, compression modulus, coagulation properties and biosafety. The MC-PMMA was embedded into rabbits and co-cultured with MG 63 cells to exam bone tissue compatibility and gene expression of osteogenesis.15.0%(wt impregnated MC-PMMA significantly lowered compressive modulus while little affected compressive strength and solidification. MC-PMMA bone cement was biologically safe and indicated excellent bone tissue compatibility. The bone-cement interface crosslinking was significantly higher in MC-PMMA than control after 6 months implantation in the femur of rabbits. The genes of osteogenesis exhibited significantly higher expression level in MC-PMMA.MC-PMMA presented perfect mechanical properties, good biosafety and excellent biocompatibility with bone tissues, which has profoundly clinical values.

  2. Alpha C-telopeptide of type I collagen is associated with subchondral bone turnover and predicts progression of joint space narrowing and osteophytes in osteoarthritis.

    Science.gov (United States)

    Huebner, Janet L; Bay-Jensen, Anne C; Huffman, Kim M; He, Yi; Leeming, Diana J; McDaniel, Gary E; Karsdal, Morten A; Kraus, Virginia B

    2014-09-01

    To evaluate joint tissue remodeling using the urinary collagen biomarkers urinary α-C-telopeptide of type I collagen (α-CTX) and urinary C-telopeptide of type II collagen (CTX-II) and to determine the association of these biomarkers with osteoarthritis (OA) severity, progression, and localized knee bone turnover. Participants (n = 149) with symptomatic and radiographic knee OA underwent fixed-flexion knee radiography at baseline and 3 years, and late-phase bone scintigraphy of both knees at baseline, which were scored semiquantitatively for osteophyte and joint space narrowing (JSN) severity and uptake intensity, with scores summed across knees. Urinary concentrations of α-CTX and CTX-II were determined by enzyme-linked immunosorbent assay. Immunohistochemical analysis of human OA knees was performed to localize the joint tissue origin of the biomarker epitopes. Urinary α-CTX concentrations correlated strongly with the intensity of bone scintigraphic uptake and with JSN progression (risk ratio 13.2) and osteophyte progression (risk ratio 3). Urinary CTX-II concentrations were strongly associated with intensity of bone scintigraphic uptake, with JSN and osteophyte severity, and with OA progression based on osteophyte score. Urinary α-CTX localized primarily to high bone turnover areas in subchondral bone. CTX-II localized to the bone-cartilage interface, the tidemark, and damaged articular cartilage. Baseline urinary α-CTX, which was localized to high turnover areas of subchondral bone, was associated with dynamic bone turnover of knees, as signified by scintigraphy, and progression of both osteophytes and JSN. Urinary CTX-II correlated with JSN and osteophyte severity and progression of osteophytes. To our knowledge, this represents the first report of serologic markers reflecting subchondral bone turnover. These collagen markers may be useful for noninvasive detection and quantification of active subchondral bone turnover and joint remodeling in knee OA

  3. Reconstitution of bone-like matrix in osteogenically differentiated mesenchymal stem cell–collagen constructs: A three-dimensional in vitro model to study hematopoietic stem cell niche

    Directory of Open Access Journals (Sweden)

    WY Lai

    2013-10-01

    Full Text Available Mesenchymal stem/stromal cells (MSCs and osteoblasts are important niche cells for hematopoietic stem cells (HSCs in bone marrow osteoblastic niche. Here, we aim to partially reconstitute the bone marrow HSC niche in vitro using collagen microencapsulation for investigation of the interactions between HSCs and MSCs. Mouse MSCs (mMSCs microencapsulated in collagen were osteogenically differentiated to derive a bone-like matrix consisting of osteocalcin, osteopontin, and calcium deposits and secreted bone morphogenic protein 2 (BMP2. Decellularized bone-like matrix was seeded with fluorescence-labeled human MSCs and HSCs. Comparing with pure collagen scaffold, significantly more HSCs and HSC–MSC pairs per unit area were found in the decellularized bone-like matrix. Moreover, incubation with excess neutralizing antibody of BMP2 resulted in a significantly higher number of HSC per unit area than that without in the decellularized matrix. This work suggests that the osteogenic differentiated MSC–collagen microsphere is a valuable three-dimensional in vitro model to elucidate cell–cell and cell–matrix interactions in HSC niche.

  4. Probabilistic Failure Analysis of Bone Using a Finite Element Model of Mineral-Collagen Composites

    OpenAIRE

    Dong, X. Neil; Guda, Teja; Millwater, Harry R.; Wang, Xiaodu

    2008-01-01

    Microdamage accumulation is a major pathway for energy dissipation during the post-yield deformation of bone. In this study, a two-dimensional probabilistic finite element model of a mineral-collagen composite was developed to investigate the influence of the tissue and ultrastructural properties of bone on the evolution of microdamage from an initial defect in tension. The probabilistic failure analyses indicated that the microdamage progression would be along the plane of the initial defect...

  5. Functional grading of mineral and collagen in the attachment of tendon to bone.

    Science.gov (United States)

    Genin, Guy M; Kent, Alistair; Birman, Victor; Wopenka, Brigitte; Pasteris, Jill D; Marquez, Pablo J; Thomopoulos, Stavros

    2009-08-19

    Attachment of dissimilar materials is a major challenge because high levels of localized stress may develop at their interfaces. An effective biologic solution to this problem exists at one of nature's most extreme interfaces: the attachment of tendon (a compliant, structural "soft tissue") to bone (a stiff, structural "hard tissue"). The goal of our study was to develop biomechanical models to describe how the tendon-to-bone insertion derives its mechanical properties. We examined the tendon-to-bone insertion and found two factors that give the tendon-to-bone transition a unique grading in mechanical properties: 1), a gradation in mineral concentration, measured by Raman spectroscopy; and 2), a gradation in collagen fiber orientation, measured by polarized light microscopy. Our measurements motivate a new physiological picture of the tissue that achieves this transition, the tendon-to-bone insertion, as a continuous, functionally graded material. Our biomechanical model suggests that the experimentally observed increase in mineral accumulation within collagen fibers can provide significant stiffening of the partially mineralized fibers, but only for concentrations of mineral above a "percolation threshold" corresponding to formation of a mechanically continuous mineral network within each collagen fiber (e.g., the case of mineral connectivity extending from one end of the fiber to the other). Increasing dispersion in the orientation distribution of collagen fibers from tendon to bone is a second major determinant of tissue stiffness. The combination of these two factors may explain the nonmonotonic variation of stiffness over the length of the tendon-to-bone insertion reported previously. Our models explain how tendon-to-bone attachment is achieved through a functionally graded material composition, and provide targets for tissue engineered surgical interventions and biomimetic material interfaces.

  6. The effect of Hydroxyapatite/collagen I composites, bone marrow aspirate and bone graft on fixation of bone implants in sheep

    DEFF Research Database (Denmark)

    Babiker, Hassan

      The effect of Hydroxyapatite/collagen I composites, bone marrow aspirate and bone graft on fixation of bone implants IN SHEEP   Ph.D. Student, Hassan Babiker; Associate Professor, Ph.D. Ming Ding; Professor, dr.med., Soren Overgaard. Department of Orthopaedic Surgery, Odense University Hospital......, Odense, Denmark   Background: Hydroxyapatite and collagen composites (HA/coll) have the potential in mimicking and replacing skeletal bones. This study attempted to determine the effect of newly developed HA/coll-composites with and without bone marrow aspirate (BMA) in order to enhance the fixation...... of bone implants.   Materials and Methods: Titanium alloy implants were inserted into bilateral femoral condyles of 8 skeletally mature sheep, four in each sheep. The implant has a circumferential gap of 2 mm. The gap was filled with: HA/coll; HA/coll-BMA; autograft or allograft. Allograft was served...

  7. Collagen Fingerprinting: A New Screening Technique for Radiocarbon Dating Ancient Bone.

    Directory of Open Access Journals (Sweden)

    Virginia L Harvey

    Full Text Available Collagen is the dominant organic component of bone and is intimately locked within the hydroxyapatite structure of this ubiquitous biomaterial that dominates archaeological and palaeontological assemblages. Radiocarbon analysis of extracted collagen is one of the most common approaches to dating bone from late Pleistocene or Holocene deposits, but dating is relatively expensive compared to other biochemical techniques. Numerous analytical methods have previously been investigated for the purpose of screening out samples that are unlikely to yield reliable dates including histological analysis, UV-stimulated fluorescence and, most commonly, the measurement of percentage nitrogen (%N and ratio of carbon to nitrogen (C:N. Here we propose the use of collagen fingerprinting (also known as Zooarchaeology by Mass Spectrometry, or ZooMS, when applied to species identification as an alternative screening method for radiocarbon dating, due to its ability to provide information on collagen presence and quality, alongside species identification. The method was tested on a series of sub-fossil bone specimens from cave systems on Cayman Brac (Cayman Islands, chosen due to the observable range in diagenetic alteration, and in particular, the extent of mineralisation. Six (14C dates, of 18 initial attempts, were obtained from remains of extinct hutia, Capromys sp. (Rodentia; Capromyidae, recovered from five distinct caves on Cayman Brac, and ranging from 393 ± 25 to 1588 ± 26 radiocarbon years before present (yr BP. All of the bone samples that yielded radiocarbon dates generated excellent collagen fingerprints, and conversely those that gave poor fingerprints also failed dating. Additionally, two successfully fingerprinted bone samples were screened out from a set of 81. Both subsequently generated (14C dates, demonstrating successful utilisation of ZooMS as an alternative screening mechanism to identify bone samples that are suitable for 1(4C analysis.

  8. Collagen Fingerprinting: A New Screening Technique for Radiocarbon Dating Ancient Bone.

    Science.gov (United States)

    Harvey, Virginia L; Egerton, Victoria M; Chamberlain, Andrew T; Manning, Phillip L; Buckley, Michael

    2016-01-01

    Collagen is the dominant organic component of bone and is intimately locked within the hydroxyapatite structure of this ubiquitous biomaterial that dominates archaeological and palaeontological assemblages. Radiocarbon analysis of extracted collagen is one of the most common approaches to dating bone from late Pleistocene or Holocene deposits, but dating is relatively expensive compared to other biochemical techniques. Numerous analytical methods have previously been investigated for the purpose of screening out samples that are unlikely to yield reliable dates including histological analysis, UV-stimulated fluorescence and, most commonly, the measurement of percentage nitrogen (%N) and ratio of carbon to nitrogen (C:N). Here we propose the use of collagen fingerprinting (also known as Zooarchaeology by Mass Spectrometry, or ZooMS, when applied to species identification) as an alternative screening method for radiocarbon dating, due to its ability to provide information on collagen presence and quality, alongside species identification. The method was tested on a series of sub-fossil bone specimens from cave systems on Cayman Brac (Cayman Islands), chosen due to the observable range in diagenetic alteration, and in particular, the extent of mineralisation. Six (14)C dates, of 18 initial attempts, were obtained from remains of extinct hutia, Capromys sp. (Rodentia; Capromyidae), recovered from five distinct caves on Cayman Brac, and ranging from 393 ± 25 to 1588 ± 26 radiocarbon years before present (yr BP). All of the bone samples that yielded radiocarbon dates generated excellent collagen fingerprints, and conversely those that gave poor fingerprints also failed dating. Additionally, two successfully fingerprinted bone samples were screened out from a set of 81. Both subsequently generated (14)C dates, demonstrating successful utilisation of ZooMS as an alternative screening mechanism to identify bone samples that are suitable for 1(4)C analysis.

  9. [Bio-Oss collagen bone grafting in the treatment of endodontic-periodontic lesion].

    Science.gov (United States)

    Sun, Jiang; Liu, Qiong

    2009-09-01

    To study the effect of Bio-Oss collagen for bone grafting in the treatment of endodontic-periodontic lesion. Thirty patients (37 teeth) with endodontic-periodontic lesions received root canal therapy and periodontal treatment. Four weeks after the treatment, Bio-Oss collagen was grafted in these teeth. Clinical examinations were performed 1, 3, 6, 9, and 12 months after the surgery. The mobility of these teeth improved 1 month after the surgery. At 3, 6, 9, and 12 months after the surgery, the pocket depth reduced obviously good attachment and further improvement of the tooth mobility. Bio-Oss collagen for bone grafting can achieve good therapeutic effect for endodontic-periodontic lesions.

  10. Thermal and electron stimulated luminescence of natural bones, commercial hydroxyapatite and collagen.

    Science.gov (United States)

    Roman-Lopez, J; Correcher, V; Garcia-Guinea, J; Rivera, T; Lozano, I B

    2014-01-01

    The luminescence (cathodoluminescence and thermoluminescence) properties of natural bones (Siberian mammoth and adult elephant), commercial hydroxyapatite and collagen were analyzed. Chemical analyses of the natural bones were determined using by Electron Probe Micro-Analysis (EMPA). Structural, molecular and thermal characteristics were determined by X-ray Diffraction (XRD), Raman spectroscopy and Differential Thermal and Thermogravimetric analysis (DTA-TG). Cathodoluminescence (CL) spectra of natural bones and collagen showed similar intense broad bands at 440 and 490 nm related to luminescence of the tetrahedral anion [Formula: see text] or structural defects. A weaker luminescence exhibited at 310 nm could be attributed to small amount of rare earth elements (REEs). Four luminescent bands at 378, 424, 468 and 576 nm were observed in the commercial hydroxyapatite (HAP). Both natural bones and collagen samples exhibited natural thermoluminescence (NTL) with well-defined glow curves whereas that the induced thermoluminescence (ITL) only appears in the samples of commercial hydroxyapatite and collagen. Additional explanations for the TL anomalous fading of apatite, as a crucial difficulty performing dosimetry and dating, are also considered. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. The effects of glucocorticoid on microarchitecture, collagen, mineral and mechanical properties of sheep femur cortical bone

    DEFF Research Database (Denmark)

    Ding, Ming; Danielsen, Carl Christian; Overgaard, Søren

    2011-01-01

    of 3 months without treatment. Group 3 was left untreated and served as controls. All sheep received a restricted diet with low calcium and phosphorus. At sacrifice, cortical bone samples from the femur midshaft of each sheep were harvested, micro-CT scanned and subjected to three-point bending...... and tensile strength testing. Bone collagen and mineral were determined. Cortical porosity was significantly increased in the glucocorticoid-2 compared with the glucocorticoid-1 and control groups. Apparent density was significantly decreased in the glucocorticoid-2 compared with the glucocorticoid-1 group....... Collagen content was significantly increased in the glucocorticoid-2 compared with the glucocorticoid-1 and control groups. Bone mineral content did not differ between the groups. Neither the three-point bending mechanical properties nor the tensile mechanical properties differed significantly between...

  12. Age variations in the properties of human tibial trabecular bone and cartilage

    DEFF Research Database (Denmark)

    Ding, Ming

    2000-01-01

    , such as apparent, apparent ash and collagen densities of human tibial trabecular bone have significant relationships with age. Tissue density and mineral concentration remain constant throughout life. Trabecular bone is tougher in the younger age, i.e. fracture requires more energy. Collagen density was the single......Initiated and motivated by clinical and scientific problems such as age-related bone fracture, prosthetic loosening, bone remodeling, and degenerative bone diseases, much significant research on the properties of trabecular bone has been carried out over the last two decades. This work has mainly...... focused on the central vertebral trabecular bone, while little is known about age-related changes in the properties of human peripheral (tibial) trabecular bone. Knowledge of the properties of peripheral (tibial) trabecular bone is of major importance for the understanding of degenerative diseases...

  13. Collagen-grafted porous HDPE/PEAA scaffolds for bone reconstruction.

    Science.gov (United States)

    Kim, Chang-Shik; Jung, Kyung-Hye; Kim, Hun; Kim, Chan-Bong; Kang, Inn-Kyu

    2016-01-01

    After tumor resection, bone reconstruction such as skull base reconstruction using interconnected porous structure is absolutely necessary. In this study, porous scaffolds for bone reconstruction were prepared using heat-pressing and salt-leaching methods. High-density polyethylene (HDPE) and poly(ethylene-co-acrylic acid) (PEAA) were chosen as the polymer composites for producing a porous scaffold of high mechanical strength and having high reactivity with biomaterials such as collagen, respectively. The porous structure was observed through surface images, and its intrusion volume and porosity were measured. Owing to the carboxylic acids on PEAA, collagen was successfully grafted onto the porous HDPE/PEAA scaffold, which was confirmed by FT-IR spectroscopy and electron spectroscopy for chemical analysis. Osteoblasts were cultured on the collagen-grafted porous scaffold, and their adhesion, proliferation, and differentiation were investigated. The high viability and growth of the osteoblasts suggest that the collagen-grafted porous HDPE/PEAA is a promising scaffold material for bone generation.

  14. Some Biomaterials based on Collagen in Human Health care

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Some Biomaterials based on Collagen in Human Health care. Ophthalmology. Wound healing. Burn Dressing. Tumor Treatment. Tissue Engineered devices. for cardio-vascular functions; For managing chronic illnesses including diabetic ulcers and foot. Smart shoe.

  15. Exercise and Regulation of Bone and Collagen Tissue Biology

    DEFF Research Database (Denmark)

    Kjaer, Michael; Jørgensen, Niklas Rye; Heinemeier, Katja

    2015-01-01

    The musculoskeletal system and its connective tissue include the intramuscular connective tissue, the myotendinous junction, the tendon, the joints with their cartilage and ligaments, and the bone; they all together play a crucial role in maintaining the architecture of the skeletal muscle, ensur...

  16. In vivo bone regeneration with injectable chitosan/hydroxyapatite/collagen composites and mesenchymal stem cells

    Science.gov (United States)

    Huang, Zhi; Chen, Yan; Feng, Qing-Ling; Zhao, Wei; Yu, Bo; Tian, Jing; Li, Song-Jian; Lin, Bo-Miao

    2011-09-01

    For reconstruction of irregular bone defects, injectable biomaterials are more appropriate than the preformed biomaterials. We herein develop a biomimetic in situ-forming composite consisting of chitosan (CS) and mineralized collagen fibrils (nHAC), which has a complex hierarchical structure similar to natural bone. The CS/nHAC composites with or without mesenchymal stem cells (MSCs) are injected into cancellous bone defects at the distal end of rabbit femurs. Defects are assessed by radiographic, histological diagnosis and Raman microscopy until 12 weeks. The results show that MSCs improve the biocompatibility of CS/nHAC composites and enhance new bone formation in vivo at 12 weeks. It can be concluded that the injectable CS/nHAC composites combined with MSCs may be a novel method for reconstruction of irregular bone defects.

  17. Elastic properties of woven bone: effect of mineral content and collagen fibrils orientation.

    Science.gov (United States)

    García-Rodríguez, J; Martínez-Reina, J

    2017-02-01

    Woven bone is a type of tissue that forms mainly during fracture healing or fetal bone development. Its microstructure can be modeled as a composite with a matrix of mineral (hydroxyapatite) and inclusions of collagen fibrils with a more or less random orientation. In the present study, its elastic properties were estimated as a function of composition (degree of mineralization) and fibril orientation. A self-consistent homogenization scheme considering randomness of inclusions' orientation was used for this purpose. Lacuno-canalicular porosity in the form of periodically distributed void inclusions was also considered. Assuming collagen fibrils to be uniformly oriented in all directions led to an isotropic tissue with a Young's modulus [Formula: see text] GPa, which is of the same order of magnitude as that of woven bone in fracture calluses. By contrast, assuming fibrils to have a preferential orientation resulted in a Young's modulus in the preferential direction of 9-16 GPa depending on the mineral content of the tissue. These results are consistent with experimental evidence for woven bone in foetuses, where collagen fibrils are aligned to a certain extent.

  18. Determination of the relationship between collagen cross-links and the bone-tissue stiffness in the porcine mandibular condyle

    NARCIS (Netherlands)

    Willems, N.M.B.K.; Mulder, L.; Bank, R.A.; Grünheid, T.; Toonder, J.M.J. den; Zentner, A.; Langenbach, G.E.J.

    2011-01-01

    Although bone-tissue stiffness is closely related to the degree to which bone has been mineralized, other determinants are yet to be identified. We, therefore, examined the extent to which the mineralization degree, collagen, and its cross-links are related to bone-tissue stiffness. A total of 50

  19. Viscoelastic behavior of discrete human collagen fibrils

    DEFF Research Database (Denmark)

    Svensson, Rene; Hassenkam, Tue; P, Hansen

    2010-01-01

    Whole tendon and fibril bundles display viscoelastic behavior, but to the best of our knowledge this property has not been directly measured in single human tendon fibrils. In the present work an atomic force microscopy (AFM) approach was used for tensile testing of two human patellar tendon fibr...

  20. Changes in subchondral bone mineral density and collagen matrix organization in growing horses.

    Science.gov (United States)

    Holopainen, Jaakko T; Brama, Pieter A J; Halmesmäki, Esa; Harjula, Terhi; Tuukkanen, Juha; van Weeren, P René; Helminen, Heikki J; Hyttinen, Mika M

    2008-12-01

    The effects of growth and maturation on the mineral deposition and the collagen framework of equine subchondral bone (SCB) were studied. Osteochondral specimens (diameter 6 mm) from the left metacarpophalangeal joint of 5-(n=8), 11-(n=8) and 18-month-old (n=6) horses were investigated at two differently loaded sites (Site 1 (S1): intermittent peak loading; Site 2 (S2): habitual loading). The SCB mineral density (BMD) was measured with peripheral quantitative computer tomography (pQCT), and the data were adjusted against the volume fraction (Vv) of the bone extracellular matrix (ECM). Polarised light microscopy (PLM) was used to analyze the Vv, the collagen fibril parallelism index and the orientation angle distribution in two fractions (1 mm/fraction) beneath the osteochondral junction of the SCB. PLM analysis was made along two randomly selected perpendicularly oriented vertical sections to measure the tissue anisotropy in the x-, y-, and z-directions. The BMD of SCB at S1 and S2 increased significantly during maturation. At the same time, the Vv of the ECM increased even more. This meant that the Vv-adjusted BMD decreased. There were no significant differences between sites. The basic collagen fibril framework of SCB seems to be established already at the age of 5 months. During maturation, the extracellular matrix underwent a decrease in collagen fibril parallelism but no changes in collagen orientation. The variation was negligible in the collagen network estimates in the two section planes. Growth and maturation induce significant changes in the equine SCB. The BMD increase in SCB is primarily due to the growth of bone volume and not to any increase in mineral deposition. An increase in weight-bearing appears to greatly affect the BMD and the volume of the extracellular matrix. Growth and maturation induce a striking change in collagen fibril parallelism but not in fibril orientation. The structural anisotropy of the subchondral bone is significant along the

  1. Age-related changes in human tendo calcaneus collagen fibrils

    International Nuclear Information System (INIS)

    Sargon, Mustafa F.; Ozlu, Korhan; Oken, Fuad

    2005-01-01

    The ruptures of tendo calcaneus often occur between the age group of 30-45 years as described by several text books. It is also described that some diseases and drugs are said to be responsible in the etiology; however, there are no studies related with the detailed histological structure of collagen fibrils found in the tendon in the age groups of humans. In view there of, this study was aimed to obtain further information on the etiology and to find an answer regarding the frequency the ruptures occurring between the age of 30-45 years in human. In the study, the biopsy specimen taken from 28 patients age (1-68) years who had undergone surgery due to tendo calcaneus ruptures or acilloplasty operations were examined by transmission electron microscope. All the specimens were prepared according to routine electronic microscope tissue preparation technique. The patients were divided into 7 age groups (1-9, 10-19, 20-29, 30-39, 40-49, 50-59, >60 years) and there were 4 patients in each group. The transverse diameters of collagen fibers were measured from the ultra thin sections and statistical analysis of the results were performed. The study was carried out in the electron microscopy laboratory of the Anatomy Department of Hacettepe University, Ankara, Turkey between January 2004 and September 2004. The diameters of the collagen fibers were higher in the 20-29 year-old groups compared to other groups and it showed a statistically significant difference. In patients who were in the 30-39 year old group or older, the diameters of the collagen fibers were lesser than the 20-29 year-old group. However, an increase was observed in the collagen fibril concentration of these groups. In examination of the specimens of patients who were under 20-year old, the diameter of the collagen fibers were less than 20-29 year -old group. The electron microscopic appearance of the tissue sample of a one year-old patient had a specific organization and in this patient, both the

  2. The potential of chitosan combined with chicken shank collagen as scaffold on bone defect regeneration process in Rattus norvegicus

    Directory of Open Access Journals (Sweden)

    Fitria Rahmitasari

    2016-12-01

    Full Text Available Background: In the field of dentistry, alveolar bone damage can be caused by periodontal disease, traumatic injury due to tooth extraction, cyst enucleation, and tumor surgery. One of the ways to regenerate the bone defect is using graft scaffold. Thus, combination of chitosan and collagen can stimulate osteogenesis. Purpose: The aim of this study was to examine the potential of chitosan combined with chicken shank collagen on bone defect regeneration process. Method: Twelve Rattus norvegicus were prepared as animal models in this research. A bone defect was intentionally created at both of the right and left femoral bones of the models. Next, 24 samples were divided into four groups, namely Group 1 using chitosan – collagen scaffold (50:50, Group 2 using chitosan collagen-scaffold (80:20, Group 3 using chitosan scaffold only, and Control Group using 3% CMC-Na. On 14th day, those animals were sacrificed, and histopathological anatomy examination was conducted to observe osteoclast cells. In addition, immunohistochemistry examination was also performed to observe RANKL expressions. Result: There was a significant difference in RANKL expressions among the groups, except between Group 3 using chitosan scaffold only and control group (p value > 0.05. The highest expression of RANKL was found in Group 1 with chitosan – collagen scaffold (50:50, followed by Group 2 with chitosan-collagen scaffold (80:20. Moreover, there was also a significant difference in osteoclast generation, except between Group 1 using chitosan – collagen scaffold (50:50 and Group 2 using chitosan-collagen scaffold (80:20, p value 0.05. Less osteoclast was found in the groups using chitosan – collagen scaffold (Group 1 and Group 2. Conclusion: Combination of chitosan and chicken shank collagen scaffold can improve regeneration process of bone defect in Rattus novergicus animals through increasing of RANKL expressions, and decreasing of osteoclast.

  3. Combined oral administration of bovine collagen peptides with calcium citrate inhibits bone loss in ovariectomized rats.

    Science.gov (United States)

    Liu, JunLi; Wang, YiHu; Song, ShuJun; Wang, XiJie; Qin, YaYa; Si, ShaoYan; Guo, YanChuan

    2015-01-01

    Collagen peptides (CPs) and calcium citrate are commonly used as bone health supplements for treating osteoporosis. However, it remains unknown whether the combination of oral bovine CPs with calcium citrate is more effective than administration of either agent alone. Forty 12-week-old Sprague-Dawley rats were randomly divided into five groups (n = 8) for once-daily intragastric administration of different treatments for 3 months at 3 months after ovariectomy (OVX) as follows: sham + vehicle; OVX + vehicle; OVX + 750 mg/kg CP; OVX + CP-calcium citrate (75 mg/kg); OVX + calcium citrate (75 mg/kg). After euthanasia, the femurs were removed and analyzed by dual energy X-ray absorptiometry and micro-computed tomography, and serum samples were analyzed for bone metabolic markers. OVX rats supplemented with CPs or CP-calcium citrate showed osteoprotective effects, with reductions in the OVX-induced decreases in their femoral bone mineral density. Moreover, CP-calcium citrate prevented trabecular bone loss, improved the microarchitecture of the distal femur, and significantly inhibited bone loss with increased bone volume, connectivity density, and trabecular number compared with OVX control rats. CP or CP-calcium citrate administration significantly increased serum procollagen type I N-terminal propeptide levels and reduced serum bone-specific alkaline phosphatase, osteocalcin, and C-telopeptide of type I collagen levels. Our data indicate that combined oral administration of bovine CPs with calcium citrate inhibits bone loss in OVX rats. The present findings suggest that combined oral administration of bovine CPs with calcium citrate is a promising alternative for reducing bone loss in osteopenic postmenopausal women.

  4. Combined oral administration of bovine collagen peptides with calcium citrate inhibits bone loss in ovariectomized rats.

    Directory of Open Access Journals (Sweden)

    JunLi Liu

    Full Text Available Collagen peptides (CPs and calcium citrate are commonly used as bone health supplements for treating osteoporosis. However, it remains unknown whether the combination of oral bovine CPs with calcium citrate is more effective than administration of either agent alone.Forty 12-week-old Sprague-Dawley rats were randomly divided into five groups (n = 8 for once-daily intragastric administration of different treatments for 3 months at 3 months after ovariectomy (OVX as follows: sham + vehicle; OVX + vehicle; OVX + 750 mg/kg CP; OVX + CP-calcium citrate (75 mg/kg; OVX + calcium citrate (75 mg/kg. After euthanasia, the femurs were removed and analyzed by dual energy X-ray absorptiometry and micro-computed tomography, and serum samples were analyzed for bone metabolic markers.OVX rats supplemented with CPs or CP-calcium citrate showed osteoprotective effects, with reductions in the OVX-induced decreases in their femoral bone mineral density. Moreover, CP-calcium citrate prevented trabecular bone loss, improved the microarchitecture of the distal femur, and significantly inhibited bone loss with increased bone volume, connectivity density, and trabecular number compared with OVX control rats. CP or CP-calcium citrate administration significantly increased serum procollagen type I N-terminal propeptide levels and reduced serum bone-specific alkaline phosphatase, osteocalcin, and C-telopeptide of type I collagen levels.Our data indicate that combined oral administration of bovine CPs with calcium citrate inhibits bone loss in OVX rats. The present findings suggest that combined oral administration of bovine CPs with calcium citrate is a promising alternative for reducing bone loss in osteopenic postmenopausal women.

  5. The Structure and Function of Non-Collagenous Bone Proteins

    Science.gov (United States)

    Hook, Magnus

    1997-01-01

    The long-term goal for this program is to determine the structural and functional relationships of bone proteins and proteins that interact with bone. This information will used to design useful pharmacological compounds that will have a beneficial effect in osteoporotic patients and in the osteoporotic-like effects experienced on long duration space missions. The first phase of this program, funded under a cooperative research agreement with NASA through the Texas Medical Center, aimed to develop powerful recombinant expression systems and purification methods for production of large amounts of target proteins. Proteins expressed in sufficient'amount and purity would be characterized by a variety of structural methods, and made available for crystallization studies. In order to increase the likelihood of crystallization and subsequent high resolution solution of structures, we undertook to develop expression of normal and mutant forms of proteins by bacterial and mammalian cells. In addition to the main goals of this program, we would also be able to provide reagents for other related studies, including development of anti-fibrotic and anti-metastatic therapeutics.

  6. Uncovering nanoscale electromechanical heterogeneity in the subfibrillar structure of collagen fibrils responsible for the piezoelectricity of bone.

    Science.gov (United States)

    Minary-Jolandan, Majid; Yu, Min-Feng

    2009-07-28

    Understanding piezoelectricity, the linear electromechanical transduction, in bone and tendon and its potential role in mechanoelectric transduction leading to their growth and remodeling remains a challenging subject. With high-resolution piezoresponse force microscopy, we probed piezoelectric behavior in relevant biological samples at different scale levels: from the subfibrillar structures of single isolated collagen fibrils to bone. We revealed that, beyond the general understanding of collagen fibril being a piezoelectric material, there existed an intrinsic piezoelectric heterogeneity within a collagen fibril coinciding with the periodic variation of its gap and overlap regions. This piezoelectric heterogeneity persisted even for the collagen fibrils embedded in bone, bringing about new implications for its possible roles in structural formation and remodeling of bone.

  7. Chemical structure, biosynthesis and synthesis of free and glycosylated pyridinolines formed by cross-link of bone and synovium collagen.

    Science.gov (United States)

    Anastasia, Luigi; Rota, Paola; Anastasia, Mario; Allevi, Pietro

    2013-09-21

    This review focuses on the chemical structure, biosynthesis and synthesis of free and glycosylated pyridinolines (Pyds), fluorescent collagen cross-links, with a pyridinium salt structure. Pyds derive from the degradation of bone collagen and have attracted attention for their use as biochemical markers of bone resorption and to assess fracture risk prediction in persons suffering from osteoporosis, bone cancer and other bone or collagen diseases. We consider and critically discuss all reported syntheses of free and glycosylated Pyds evidencing an unrevised chemistry, original and of general utility, analysis of which allows us to also support a previously suggested non-enzymatic formation of Pyds in collagen better rationalizing and justifying the chemical events.

  8. Cross-correlative 3D micro-structural investigation of human bone processed into bone allografts

    International Nuclear Information System (INIS)

    Singh, Atul Kumar; Gajiwala, Astrid Lobo; Rai, Ratan Kumar; Khan, Mohd Parvez; Singh, Chandan; Barbhuyan, Tarun; Vijayalakshmi, S.; Chattopadhyay, Naibedya; Sinha, Neeraj; Kumar, Ashutosh; Bellare, Jayesh R.

    2016-01-01

    Bone allografts (BA) are a cost-effective and sustainable alternative in orthopedic practice as they provide a permanent solution for preserving skeletal architecture and function. Such BA however, must be processed to be disease free and immunologically safe as well as biologically and clinically useful. Here, we have demonstrated a processing protocol for bone allografts and investigated the micro-structural properties of bone collected from osteoporotic and normal human donor samples. In order to characterize BA at different microscopic levels, a combination of techniques such as Solid State Nuclear Magnetic Resonance (ssNMR), Scanning Electron Microscope (SEM), micro-computed tomography (μCT) and Thermal Gravimetric Analysis (TGA) were used for delineating the ultra-structural property of bone. ssNMR revealed the extent of water, collagen fine structure and crystalline order in the bone. These were greatly perturbed in the bone taken from osteoporotic bone donor. Among the processing methods analyzed, pasteurization at 60 °C and radiation treatment appeared to substantially alter the bone integrity. SEM study showed a reduction in Ca/P ratio and non-uniform distribution of elements in osteoporotic bones. μ-CT and MIMICS® (Materialize Interactive Medical Image Control System) demonstrated that pasteurization and radiation treatment affects the BA morphology and cause a shift in the HU unit. However, the combination of all these processes restored all-important parameters that are critical for BA integrity and sustainability. Cross-correlation between the various probes we used quantitatively demonstrated differences in morphological and micro-structural properties between BA taken from normal and osteoporotic human donor. Such details could also be instrumental in designing an appropriate bone scaffold. For the best restoration of bone microstructure and to be used as a biomaterial allograft, a step-wise processing method is recommended that preserves all

  9. Cross-correlative 3D micro-structural investigation of human bone processed into bone allografts

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Atul Kumar [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Gajiwala, Astrid Lobo [Tissue Bank, Tata Memorial Hospital, Parel, Mumbai 400012 (India); Rai, Ratan Kumar [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Khan, Mohd Parvez [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Singh, Chandan [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Barbhuyan, Tarun [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Vijayalakshmi, S. [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Chattopadhyay, Naibedya [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Sinha, Neeraj, E-mail: neerajcbmr@gmail.com [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Kumar, Ashutosh, E-mail: ashutoshk@iitb.ac.in [Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Mumbai 400076 (India); Bellare, Jayesh R., E-mail: jb@iitb.ac.in [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Department of Chemical Engineering, Indian Institute of Technology Bombay, Mumbai 400076 (India)

    2016-05-01

    Bone allografts (BA) are a cost-effective and sustainable alternative in orthopedic practice as they provide a permanent solution for preserving skeletal architecture and function. Such BA however, must be processed to be disease free and immunologically safe as well as biologically and clinically useful. Here, we have demonstrated a processing protocol for bone allografts and investigated the micro-structural properties of bone collected from osteoporotic and normal human donor samples. In order to characterize BA at different microscopic levels, a combination of techniques such as Solid State Nuclear Magnetic Resonance (ssNMR), Scanning Electron Microscope (SEM), micro-computed tomography (μCT) and Thermal Gravimetric Analysis (TGA) were used for delineating the ultra-structural property of bone. ssNMR revealed the extent of water, collagen fine structure and crystalline order in the bone. These were greatly perturbed in the bone taken from osteoporotic bone donor. Among the processing methods analyzed, pasteurization at 60 °C and radiation treatment appeared to substantially alter the bone integrity. SEM study showed a reduction in Ca/P ratio and non-uniform distribution of elements in osteoporotic bones. μ-CT and MIMICS® (Materialize Interactive Medical Image Control System) demonstrated that pasteurization and radiation treatment affects the BA morphology and cause a shift in the HU unit. However, the combination of all these processes restored all-important parameters that are critical for BA integrity and sustainability. Cross-correlation between the various probes we used quantitatively demonstrated differences in morphological and micro-structural properties between BA taken from normal and osteoporotic human donor. Such details could also be instrumental in designing an appropriate bone scaffold. For the best restoration of bone microstructure and to be used as a biomaterial allograft, a step-wise processing method is recommended that preserves all

  10. Coating of Biomaterial Scaffolds with the Collagen-Mimetic Peptide GFOGER for Bone Defect Repair

    OpenAIRE

    Wojtowicz, Abigail M.; Shekaran, Asha; Oest, Megan E.; Dupont, Kenneth M.; Templeman, Kellie L.; Hutmacher, Dietmar W.; Guldberg, Robert E.; García, Andrés J.

    2009-01-01

    Healing large bone defects and non-unions remains a significant clinical problem. Current treatments, consisting of auto- and allografts, are limited by donor supply and morbidity, insufficient bioactivity and risk of infection. Biotherapeutics, including cells, genes and proteins, represent promising alternative therapies, but these strategies are limited by technical roadblocks to biotherapeutic delivery, cell sourcing, high cost, and regulatory hurdles. In the present study, the collagen-m...

  11. Viscoelastic behavior of discrete human collagen fibrils

    DEFF Research Database (Denmark)

    Svensson, René; Hassenkam, Tue; Hansen, Philip

    2010-01-01

    Whole tendon and fibril bundles display viscoelastic behavior, but to the best of our knowledge this property has not been directly measured in single human tendon fibrils. In the present work an atomic force microscopy (AFM) approach was used for tensile testing of two human patellar tendon...... saline, cyclic testing was performed in the pre-yield region at different strain rates, and the elastic response was determined by a stepwise stress relaxation test. The elastic stress-strain response corresponded to a second-order polynomial fit, while the viscous response showed a linear dependence...

  12. Bone conditioned media (BCM) improves osteoblast adhesion and differentiation on collagen barrier membranes.

    Science.gov (United States)

    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

  13. Bone induction by composites of bioresorbable carriers and demineralized bone in rats: a comparative study of fibrin-collagen paste, fibrin sealant, and polyorthoester with gentamicin

    DEFF Research Database (Denmark)

    Pinholt, E M; Solheim, E; Bang, G

    1992-01-01

    fibrin-collagen paste and fibrin sealant inhibited bone induction and produced a chronic inflammation; part of the fibrin-collagen paste was still present at 4 weeks. Polyorthoester with gentamicin was almost completely absorbed, induced minimal tissue reaction, and did not inhibit osteoinduction....

  14. Collagen type I from bovine bone. Effect of animal age, bone anatomy and drying methodology on extraction yield, self-assembly, thermal behaviour and electrokinetic potential

    OpenAIRE

    Ferraro, Vincenza; Gaillard-Martinie, Brigitte; Sayd, Thierry; Chambon, Christophe; Anton, Marc; Sante-Lhoutellier, Veronique

    2017-01-01

    Natural collagen is easily available from animal tissues such as bones. Main limitations reported in the use of natural collagen are heterogeneity and loss of integrity during recovery. However, its natural complexity, functionality and bioactivity still remain to be achieved through synthetic and recombinant ways. Variability of physicochemical prope...

  15. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P < 0.05) in longitudinal bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P < 0.05). Our findings suggest that the processes regulating new collagen accretion, bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  16. Beneath the Minerals, a Layer of Round Lipid Particles Was Identified to Mediate Collagen Calcification in Compact Bone Formation

    OpenAIRE

    Xu, Shaohua; Yu, Jianqing J.

    2006-01-01

    Astronauts lose 1–2% of their bone minerals per month during space flights. A systematic search for a countermeasure relies on a good understanding of the mechanism of bone formation at the molecular level. How collagen fibers, the dominant matrix protein in bones, are mineralized remains mysterious. Atomic force microscopy was carried out, in combination with immunostaining and Western blotting, on bovine tibia to identify unrecognized building blocks involved in bone formation and for an el...

  17. Structure to function: Spider silk and human collagen

    Science.gov (United States)

    Rabotyagova, Olena S.

    , morphological features and assembly. Aside from fundamental perspectives, we anticipate that these results will provide a blueprint for the design of precise materials for a range of potential applications such as controlled release devices, functional coatings, components of tissue regeneration materials and environmentally friendly polymers in future studies. In the second part of this work, human collagen type I was studied as another representative of the family of fibrous proteins. Collagen type I is the most abundant extracellular matrix protein in the human body, providing the basis for tissue structure and directing cellular functions. Collagen has a complex structural hierarchy, organized at different length scales, including the characteristic triple helical feature. In the present study we assessed the relationship between collagen structure (native vs. denatured) and sensitivity to UV radiation with a focus on changes in the primary structure, conformation, microstructure and material properties. Free radical reactions are involved in collagen degradation and a mechanism for UV-induced collagen degradation related to structure was proposed. The results from this study demonstrated the role of collagen supramolecular organization (triple helix) in the context of the effects of electromagnetic radiation on extracellular matrices. Owing to the fact that both silks and collagens are proteins that have found widespread interest for biomaterial related needs, we anticipate that the current studies will serve as a foundation for future biomaterial designs with controlled properties. Furthermore, fundamental insight into self-assembly and environmentally-2mediated degradation, will build a foundation for fundamental understanding of the remodeling and functions of these types of fibrous proteins in vivo and in vitro. This type of insight is essential for many areas of scientific inquiry, from drug delivery, to scaffolds for tissue engineering, and to the stability of

  18. Degradability of injectable calcium sulfate/mineralized collagen-based bone repair material and its effect on bone tissue regeneration

    International Nuclear Information System (INIS)

    Chen, Zonggang; Kang, Lingzhi; Meng, Qing-Yuan; Liu, Huanye; Wang, Zhaoliang; Guo, Zhongwu; Cui, Fu-Zhai

    2014-01-01

    The nHAC/CSH composite is an injectable bone repair material with controllable injectability and self-setting properties prepared by introducing calcium sulfate hemihydrate (CSH) into mineralized collagen (nHAC). When mixed with water, the nHAC/CSH composites can be transformed into mineralized collagen/calcium sulfate dihydrate (nHAC/CSD) composites. The nHAC/CSD composites have good biocompatibility and osteogenic capability. Considering that the degradation behavior of bone repair material is another important factor for its clinical applications, the degradability of nHAC/CSD composites was studied. The results showed that the degradation ratio of the nHAC/CSD composites with lower nHAC content increased with the L/S ratio increase of injectable materials, but the variety of L/S ratio had no significant effect on the degradation ratio of the nHAC/CSD composites with higher nHAC content. Increasing nHAC content in the composites could slow down the degradation of nHAC/CSD composite. Setting accelerator had no significant effect on the degradability of nHAC/CSD composites. In vivo histological analysis suggests that the degradation rate of materials can match the growth rate of new mandibular bone tissues in the implanted site of rabbit. The regulable degradability of materials resulting from the special prescriptions of injectable nHAC/CSH composites will further improve the workability of nHAC/CSD composites. - Highlights: • The nHAC/CSH composite can be as an injectable bone repair material. • The L/S ratio and nHAC content have a significant effect on material degradability. • The degradability of bone materials can be regulated to match tissue repair. • The regulable degradability will further improve the workability of bone materials

  19. Degradability of injectable calcium sulfate/mineralized collagen-based bone repair material and its effect on bone tissue regeneration

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zonggang, E-mail: chenzg@sdu.edu.cn [National Glycoengineering Research Center, Shandong University, Jinan 250100 (China); Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Kang, Lingzhi [National Glycoengineering Research Center, Shandong University, Jinan 250100 (China); Meng, Qing-Yuan [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Liu, Huanye [Department of Prosthodontics, School of Stomatology, China Medical University, Shenyang 110001 (China); Wang, Zhaoliang [Jinan Military General Hospital of PLA, Jinan 250031 (China); Guo, Zhongwu, E-mail: zwguo@sdu.edu.cn [National Glycoengineering Research Center, Shandong University, Jinan 250100 (China); Cui, Fu-Zhai, E-mail: cuifz@mail.tsinghua.edu.cn [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China)

    2014-12-01

    The nHAC/CSH composite is an injectable bone repair material with controllable injectability and self-setting properties prepared by introducing calcium sulfate hemihydrate (CSH) into mineralized collagen (nHAC). When mixed with water, the nHAC/CSH composites can be transformed into mineralized collagen/calcium sulfate dihydrate (nHAC/CSD) composites. The nHAC/CSD composites have good biocompatibility and osteogenic capability. Considering that the degradation behavior of bone repair material is another important factor for its clinical applications, the degradability of nHAC/CSD composites was studied. The results showed that the degradation ratio of the nHAC/CSD composites with lower nHAC content increased with the L/S ratio increase of injectable materials, but the variety of L/S ratio had no significant effect on the degradation ratio of the nHAC/CSD composites with higher nHAC content. Increasing nHAC content in the composites could slow down the degradation of nHAC/CSD composite. Setting accelerator had no significant effect on the degradability of nHAC/CSD composites. In vivo histological analysis suggests that the degradation rate of materials can match the growth rate of new mandibular bone tissues in the implanted site of rabbit. The regulable degradability of materials resulting from the special prescriptions of injectable nHAC/CSH composites will further improve the workability of nHAC/CSD composites. - Highlights: • The nHAC/CSH composite can be as an injectable bone repair material. • The L/S ratio and nHAC content have a significant effect on material degradability. • The degradability of bone materials can be regulated to match tissue repair. • The regulable degradability will further improve the workability of bone materials.

  20. Automated quantification of aligned collagen for human breast carcinoma prognosis

    Directory of Open Access Journals (Sweden)

    Jeremy S Bredfeldt

    2014-01-01

    Full Text Available Background: Mortality in cancer patients is directly attributable to the ability of cancer cells to metastasize to distant sites from the primary tumor. This migration of tumor cells begins with a remodeling of the local tumor microenvironment, including changes to the extracellular matrix and the recruitment of stromal cells, both of which facilitate invasion of tumor cells into the bloodstream. In breast cancer, it has been proposed that the alignment of collagen fibers surrounding tumor epithelial cells can serve as a quantitative image-based biomarker for survival of invasive ductal carcinoma patients. Specific types of collagen alignment have been identified for their prognostic value and now these tumor associated collagen signatures (TACS are central to several clinical specimen imaging trials. Here, we implement the semi-automated acquisition and analysis of this TACS candidate biomarker and demonstrate a protocol that will allow consistent scoring to be performed throughout large patient cohorts. Methods: Using large field of view high resolution microscopy techniques, image processing and supervised learning methods, we are able to quantify and score features of collagen fiber alignment with respect to adjacent tumor-stromal boundaries. Results: Our semi-automated technique produced scores that have statistically significant correlation with scores generated by a panel of three human observers. In addition, our system generated classification scores that accurately predicted survival in a cohort of 196 breast cancer patients. Feature rank analysis reveals that TACS positive fibers are more well-aligned with each other, are of generally lower density, and terminate within or near groups of epithelial cells at larger angles of interaction. Conclusion: These results demonstrate the utility of a supervised learning protocol for streamlining the analysis of collagen alignment with respect to tumor stromal boundaries.

  1. GH receptor blocker administration and muscle-tendon collagen synthesis in humans

    DEFF Research Database (Denmark)

    Nielsen, Rie Harboe; Doessing, Simon; Goto, Kazushige

    2011-01-01

    The growth hormone (GH)/insulin-like growth factor-I (IGF-I) axis stimulates collagen synthesis in tendon and skeletal muscle, but no studies have investigated the effect of reducing IGF-I on collagen synthesis in healthy humans.......The growth hormone (GH)/insulin-like growth factor-I (IGF-I) axis stimulates collagen synthesis in tendon and skeletal muscle, but no studies have investigated the effect of reducing IGF-I on collagen synthesis in healthy humans....

  2. Scaffolds for bone regeneration made of hydroxyapatite microspheres in a collagen matrix

    Energy Technology Data Exchange (ETDEWEB)

    Cholas, Rahmatullah, E-mail: rahmat.cholas@gmail.com; Kunjalukkal Padmanabhan, Sanosh, E-mail: sanosh2001@gmail.com; Gervaso, Francesca; Udayan, Gayatri; Monaco, Graziana; Sannino, Alessandro; Licciulli, Antonio

    2016-06-01

    Biomimetic scaffolds with a structural and chemical composition similar to native bone tissue may be promising for bone tissue regeneration. In the present work hydroxyapatite mesoporous microspheres (mHA) were incorporated into collagen scaffolds containing an ordered interconnected macroporosity. The mHA were obtained by spray drying of a nano hydroxyapatite slurry prepared by the precipitation technique. X-ray diffraction (XRD) analysis revealed that the microspheres were composed only of hydroxyapatite (HA) phase, and energy-dispersive x-ray spectroscopy (EDS) analysis revealed the Ca/P ratio to be 1.69 which is near the value for pure HA. The obtained microspheres had an average diameter of 6 μm, a specific surface area of 40 m{sup 2}/g as measured by Brunauer-Emmett-Teller (BET) analysis, and Barrett-Joyner-Halenda (BJH) analysis showed a mesoporous structure with an average pore diameter of 16 nm. Collagen/HA-microsphere (Col/mHA) composite scaffolds were prepared by freeze-drying followed by dehydrothermal crosslinking. SEM observations of Col/mHA scaffolds revealed HA microspheres embedded within a porous collagen matrix with a pore size ranging from a few microns up to 200 μm, which was also confirmed by histological staining of sections of paraffin embedded scaffolds. The compressive modulus of the composite scaffold at low and high strain values was 1.7 and 2.8 times, respectively, that of pure collagen scaffolds. Cell proliferation measured by the MTT assay showed more than a 3-fold increase in cell number within the scaffolds after 15 days of culture for both pure collagen scaffolds and Col/mHA composite scaffolds. Attractive properties of this composite scaffold include the potential to load the microspheres for drug delivery and the controllability of the pore structure at various length scales. - Highlights: • Mesoporous hydroxyapatite microsphere(mHA) synthesized by spray drying method • Porous collagen/mHA composite scaffold made by freeze

  3. Calcified cartilage or bone? Collagens in the tessellated endoskeletons of cartilaginous fish (sharks and rays).

    Science.gov (United States)

    Seidel, Ronald; Blumer, Michael; Pechriggl, Elisabeth-Judith; Lyons, Kady; Hall, Brian K; Fratzl, Peter; Weaver, James C; Dean, Mason N

    2017-10-01

    The primary skeletal tissue in elasmobranchs -sharks, rays and relatives- is cartilage, forming both embryonic and adult endoskeletons. Only the skeletal surface calcifies, exhibiting mineralized tiles (tesserae) sandwiched between a cartilage core and overlying fibrous perichondrium. These two tissues are based on different collagens (Coll II and I, respectively), fueling a long-standing debate as to whether tesserae are more like calcified cartilage or bone (Coll 1-based) in their matrix composition. We demonstrate that stingray (Urobatis halleri) tesserae are bipartite, having an upper Coll I-based 'cap' that merges into a lower Coll II-based 'body' zone, although tesserae are surrounded by cartilage. We identify a 'supratesseral' unmineralized cartilage layer, between tesserae and perichondrium, distinguished from the cartilage core in containing Coll I and X (a common marker for mammalian mineralization), in addition to Coll II. Chondrocytes within tesserae appear intact and sit in lacunae filled with Coll II-based matrix, suggesting tesserae originate in cartilage, despite comprising a diversity of collagens. Intertesseral joints are also complex in their collagenous composition, being similar to supratesseral cartilage closer to the perichondrium, but containing unidentified fibrils nearer the cartilage core. Our results indicate a unique potential for tessellated cartilage in skeletal biology research, since it lacks features believed diagnostic for vertebrate cartilage mineralization (e.g. hypertrophic and apoptotic chondrocytes), while offering morphologies amenable for investigating the regulation of complex mineralized ultrastructure and tissues patterned on multiple collagens. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Onlay bone augmentation on mouse calvarial bone using a hydroxyapatite/collagen composite material with total blood or platelet-rich plasma.

    Science.gov (United States)

    Ohba, Seigo; Sumita, Yoshinori; Umebayashi, Mayumi; Yoshimura, Hitoshi; Yoshida, Hisato; Matsuda, Shinpei; Kimura, Hideki; Asahina, Izumi; Sano, Kazuo

    2016-01-01

    The aim of this study was to assess newly formed onlay bone on mouse calvarial bone using a new artificial bone material, a hydroxyapatite/collagen composite, with total blood or platelet-rich plasma. The hydroxyapatite/collagen composite material with normal saline, total blood or platelet-rich plasma was transplanted on mouse calvarial bone. The mice were sacrificed and the specimens were harvested four weeks after surgery. The newly formed bone area was measured on hematoxylin and eosin stained specimens using Image J software. The hydroxyapatite/collagen composite materials with total blood or platelet-rich plasma induced a significantly greater amount of newly formed bone than that with normal saline. Moreover, bone marrow was observed four weeks after surgery in the transplanted materials with total blood or platelet-rich plasma but not with normal saline. However, there were no significant differences in the amount of newly formed bone between materials used with total blood versus platelet-rich plasma. The hydroxyapatite/collagen composite material was valid for onlay bone augmentation and this material should be soaked in total blood or platelet-rich plasma prior to transplantation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Osteogenesis imperfecta due to mutations in non-collagenous genes: lessons in the biology of bone formation.

    Science.gov (United States)

    Marini, Joan C; Reich, Adi; Smith, Simone M

    2014-08-01

    Osteogenesis imperfecta or 'brittle bone disease' has mainly been considered a bone disorder caused by collagen mutations. Within the last decade, however, a surge of genetic discoveries has created a new paradigm for osteogenesis imperfecta as a collagen-related disorder, where most cases are due to autosomal dominant type I collagen defects, while rare, mostly recessive, forms are due to defects in genes whose protein products interact with collagen protein. This review is both timely and relevant in outlining the genesis, development, and future of this paradigm shift in the understanding of osteogenesis imperfecta. Bone-restricted interferon-induced transmembrane (IFITM)-like protein (BRIL) and pigment epithelium-derived factor (PEDF) defects cause types V and VI osteogenesis imperfecta via defective bone mineralization, while defects in cartilage-associated protein (CRTAP), prolyl 3-hydroxylase 1 (P3H1), and cyclophilin B (CYPB) cause types VII-IX osteogenesis imperfecta via defective collagen post-translational modification. Heat shock protein 47 (HSP47) and FK506-binding protein-65 (FKBP65) defects cause types X and XI osteogenesis imperfecta via aberrant collagen crosslinking, folding, and chaperoning, while defects in SP7 transcription factor, wingless-type MMTV integration site family member 1 (WNT1), trimeric intracellular cation channel type b (TRIC-B), and old astrocyte specifically induced substance (OASIS) disrupt osteoblast development. Finally, absence of the type I collagen C-propeptidase bone morphogenetic protein 1 (BMP1) causes type XII osteogenesis imperfecta due to altered collagen maturation/processing. Identification of these multiple causative defects has provided crucial information for accurate genetic counseling, inspired a recently proposed functional grouping of osteogenesis imperfecta types by shared mechanism to simplify current nosology, and has prodded investigations into common pathways in osteogenesis imperfecta. Such

  6. Effect of Simvastatin collagen graft on wound healing of defective bone

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Jun Ho; Kim, Gyu Tae [Department of Oral and Maxillofacial Radiology, School of Dentistry, Kyung Hee University, Seoul (Korea, Republic of); Choi, Yong Suk; Lee, Hyeon Woo; Hwang, Eui Hwan [Institute of Oral Biology, School of Dentistry, Kyung Hee University, Seoul (Korea, Republic of)

    2008-09-15

    To observe and evaluate the effects of Simvastatin-induced osteogenesis on the wound healing of defective bone. 64 defective bones were created in the parietal bone of 32 New Zealand White rabbits. The defects were grafted with collagen matrix carriers mixed with Simvastatin solution in the experimental group of 16 rabbits and with collagen matrix carriers mixed with water in the controlled group. The rabbits were terminated at an interval of 3, 5, 7, and 9 days, 2, 4, 6, and 8 weeks after the formation of defective bone. The wound healing was evaluated by soft X-ray radiography. The tissues within defective bones were evaluated through the analysis of flow cytometry for the manifestation of Runx2 and Osteocalcin, and observed histopathologically by using H-E stain and Masson's trichrome stain. Results : 1. In the experimental group, flow cytometry revealed more manifestation of Runx2 at 5, 7, and 9 days and Osteocalcin at 2 weeks than in the controlled groups, but there was few difference in comparison with the controlled group. 2. In the experimental group, flow cytometry revealed considerably more cells and erythrocytes at 5, 7, and 9 days in comparison with the controlled group. 3. In the experimental group, soft x-ray radiography revealed the extended formation of trabeculation at 2, 4, 6, and 8 weeks. 4. Histopathological features of the experimental group showed more fibroblasts and newly formed vessels at 5 and 7 days, and the formation of osteoid tissues at 9 days, and the newly formed trabeculations at 4 and 6 weeks. As the induced osteogenesis by Simvastatin, there was few contrast of the manifestation between Runx2 and Osteocalcin based on the differentiation of osteoblasts. But it was considered that the more formation of cells and erythrocytes depending on newly formed vessels in the experimental group obviously had an effect on the bone regeneration.

  7. Effect of Simvastatin collagen graft on wound healing of defective bone

    International Nuclear Information System (INIS)

    Kang, Jun Ho; Kim, Gyu Tae; Choi, Yong Suk; Lee, Hyeon Woo; Hwang, Eui Hwan

    2008-01-01

    To observe and evaluate the effects of Simvastatin-induced osteogenesis on the wound healing of defective bone. 64 defective bones were created in the parietal bone of 32 New Zealand White rabbits. The defects were grafted with collagen matrix carriers mixed with Simvastatin solution in the experimental group of 16 rabbits and with collagen matrix carriers mixed with water in the controlled group. The rabbits were terminated at an interval of 3, 5, 7, and 9 days, 2, 4, 6, and 8 weeks after the formation of defective bone. The wound healing was evaluated by soft X-ray radiography. The tissues within defective bones were evaluated through the analysis of flow cytometry for the manifestation of Runx2 and Osteocalcin, and observed histopathologically by using H-E stain and Masson's trichrome stain. Results : 1. In the experimental group, flow cytometry revealed more manifestation of Runx2 at 5, 7, and 9 days and Osteocalcin at 2 weeks than in the controlled groups, but there was few difference in comparison with the controlled group. 2. In the experimental group, flow cytometry revealed considerably more cells and erythrocytes at 5, 7, and 9 days in comparison with the controlled group. 3. In the experimental group, soft x-ray radiography revealed the extended formation of trabeculation at 2, 4, 6, and 8 weeks. 4. Histopathological features of the experimental group showed more fibroblasts and newly formed vessels at 5 and 7 days, and the formation of osteoid tissues at 9 days, and the newly formed trabeculations at 4 and 6 weeks. As the induced osteogenesis by Simvastatin, there was few contrast of the manifestation between Runx2 and Osteocalcin based on the differentiation of osteoblasts. But it was considered that the more formation of cells and erythrocytes depending on newly formed vessels in the experimental group obviously had an effect on the bone regeneration.

  8. Guided bone regeneration with asymmetric collagen-chitosan membranes containing aspirin-loaded chitosan nanoparticles

    Directory of Open Access Journals (Sweden)

    Zhang J

    2017-12-01

    Full Text Available Jiayu Zhang,1 Shiqing Ma,1 Zihao Liu,1 Hongjuan Geng,1 Xin Lu,1 Xi Zhang,1 Hongjie Li,1 Chenyuan Gao,2 Xu Zhang,1 Ping Gao1 1School of Dentistry, Hospital of Stomatology, Tianjin Medical University, Tianjin, 2Beijing Laboratory of Biomedical Materials, Beijing University of Chemical Technology, Beijing, People’s Republic of China Introduction: Membranes allowing the sustained release of drugs that can achieve cell adhesion are very promising for guided bone regeneration. Previous studies have suggested that aspirin has the potential to promote bone regeneration. The purpose of this study was to prepare a local drug delivery system with aspirin-loaded chitosan nanoparticles (ACS contained in an asymmetric collagen-chitosan membrane (CCM. Methods: In this study, the ACS were fabricated using different concentrations of aspirin (5 mg, 25 mg, 50 mg, and 75 mg. The drug release behavior of ACS was studied. Transmission electron microscopy (TEM and scanning electron microscopy (SEM were used to examine the micromorphology of ACS and aspirin-loaded chitosan nanoparticles contained in chitosan-collagen membranes (ACS-CCM. In vitro bone mesenchymal stem cells (BMSCs were cultured and critical-sized cranial defects on Sprague-Dawley rats were made to evaluate the effect of the ACS-CCM on bone regeneration.Results: Drug release behavior results of ACS showed that the nanoparticles fabricated in this study could successfully sustain the release of the drug. TEM showed the morphology of the nanoparticles. SEM images indicated that the asymmetric membrane comprised a loose collagen layer and a dense chitosan layer. In vitro studies showed that ACS-CCM could promote the proliferation of BMSCs, and that the degree of differentiated BMSCs seeded on CCMs containing 50 mg of ACS was higher than that of other membranes. Micro-computed tomography showed that 50 mg of ACS-CCM resulted in enhanced bone regeneration compared with the control group.Conclusion: This

  9. MT1-MMP and type II collagen specify skeletal stem cells and their bone and cartilage progeny

    DEFF Research Database (Denmark)

    Szabova, L.; Yamada, S.S.; Wimer, H.

    2009-01-01

    -expressing cells of the skeleton rescues not only diminished chondrocyte proliferation, but surprisingly, also results in amelioration of the severe skeletal dysplasia associated with MT1-MMP deficiency through enhanced bone formation. Consistent with this increased bone formation, type II collagen was identified...... from nontransgenic MT1-MMP-deficient littermates. These observations show that type II collagen is not stringently confined to the chondrocyte but is expressed in skeletal stem/progenitor cells (able to regenerate bone, cartilage, myelosupportive stroma, marrow adipocytes) and in the chondrogenic...

  10. Recombinant human bone morphogenetic protein induces bone formation

    International Nuclear Information System (INIS)

    Wang, E.A.; Rosen, V.; D'Alessandro, J.S.; Bauduy, M.; Cordes, P.; Harada, T.; Israel, D.I.; Hewick, R.M.; Kerns, K.M.; LaPan, P.; Luxenberg, D.P.; McQuaid, D.; Moutsatsos, I.K.; Nove, J.; Wozney, J.M.

    1990-01-01

    The authors have purified and characterized active recombinant human bone morphogenetic protein (BMP) 2A. Implantation of the recombinant protein in rats showed that a single BMP can induce bone formation in vivo. A dose-response and time-course study using the rat ectopic bone formation assay revealed that implantation of 0.5-115 μg of partially purified recombinant human BMP-2A resulted in cartilage by day 7 and bone formation by day 14. The time at which bone formation occurred was dependent on the amount of BMP-2A implanted; at high doses bone formation could be observed at 5 days. The cartilage- and bone-inductive activity of the recombinant BMP-2A is histologically indistinguishable from that of bone extracts. Thus, recombinant BMP-2A has therapeutic potential to promote de novo bone formation in humans

  11. Amino acid δ13C analysis of hair proteins and bone collagen using liquid chromatography/isotope ratio mass spectrometry

    DEFF Research Database (Denmark)

    Raghavan, Maanasa; McCullagh, James S. O.; Lynnerup, Niels

    2010-01-01

    We report a novel method for the chromatographic separation and measurement of stable carbon isotope ratios (delta(13)C) of individual amino acids in hair proteins and bone collagen using the LC-IsoLink system, which interfaces liquid chromatography (LC) with isotope ratio mass spectrometry (IRMS......). This paper provides baseline separation of 15 and 13 of the 18 amino acids in bone collagen and hair proteins, respectively. We also describe an approach to analysing small hair samples for compound-specific analysis of segmental hair sections. The LC/IRMS method is applied in a historical context...... by the delta(13)C analysis of hair proteins and bone collagen recovered from six individuals from Uummannaq in Greenland. The analysis of hair and bone amino acids from the same individual, compared for the first time in this study, is of importance in palaeodietary reconstruction. If hair proteins can be used...

  12. Altered distributions of bone tissue mineral and collagen properties in women with fragility fractures.

    Science.gov (United States)

    Wang, Zhen Xiang; Lloyd, Ashley A; Burket, Jayme C; Gourion-Arsiquaud, Samuel; Donnelly, Eve

    2016-03-01

    Heterogeneity of bone tissue properties is emerging as a potential indicator of altered bone quality in pathologic tissue. The objective of this study was to compare the distributions of tissue properties in women with and without histories of fragility fractures using Fourier transform infrared (FTIR) imaging. We extended a prior study that examined the relationship of the mean FTIR properties to fracture risk by analyzing in detail the widths and the tails of the distributions of FTIR properties in biopsies from fracture and non-fracture cohorts. The mineral and matrix properties of cortical and trabecular iliac crest tissue were compared in biopsies from women with a history of fragility fracture (+Fx; n=21, age: mean 54±SD 15y) and with no history of fragility fracture (-Fx; n=12, age: 57±5y). A subset of the patients included in the -Fx group were taking estrogen-plus-progestin hormone replacement therapy (HRT) (-Fx+HRT n=8, age: 58±5y) and were analyzed separately from patients with no history of HRT (-Fx-HRT n=4, age: 56±7y). When the FTIR parameter mean values were examined by treatment group, the trabecular tissue of -Fx-HRT patients had a lower mineral:matrix ratio (M:M) and collagen maturity (XLR) than that of -Fx+HRT patients (-22% M:M, -18% XLR) and +Fx patients (-17% M:M, -18% XLR). Across multiple FTIR parameters, tissue from the -Fx-HRT group had smaller low-tail (5th percentile) values than that from the -Fx+HRT or +Fx groups. In trabecular collagen maturity and crystallinity (XST), the -Fx-HRT group had smaller low-tail values than those in the -Fx+HRT group (-16% XLR, -5% XST) and the +Fx group (-17% XLR, -7% XST). The relatively low values of trabecular mineral:matrix ratio and collagen maturity and smaller low-tail values of collagen maturity and crystallinity observed in the -Fx-HRT group are characteristic of younger tissue. Taken together, our data suggest that the presence of newly formed tissue that includes small/imperfect crystals

  13. Training-induced changes in peritendinous type I collagen turnover determined by microdialysis in humans

    DEFF Research Database (Denmark)

    Langberg, Henning; Rosendal, L; Kjaer, M

    2001-01-01

    1. Acute exercise is found to increase collagen type I formation locally in peritendinous connective tissue of the Achilles' tendon in humans, as determined from changes in interstitial concentrations of collagen propeptide (PICP) and a collagen degradation product (ICTP) by the use of microdialy...

  14. Biomimetic mineralization of recombinant collagen type I derived protein to obtain hybrid matrices for bone regeneration.

    Science.gov (United States)

    Ramírez-Rodríguez, Gloria Belén; Delgado-López, José Manuel; Iafisco, Michele; Montesi, Monica; Sandri, Monica; Sprio, Simone; Tampieri, Anna

    2016-11-01

    Understanding the mineralization mechanism of synthetic protein has recently aroused great interest especially in the development of advanced materials for bone regeneration. Herein, we propose the synthesis of composite materials through the mineralization of a recombinant collagen type I derived protein (RCP) enriched with RGD sequences in the presence of magnesium ions (Mg) to closer mimic bone composition. The role of both RCP and Mg ions in controlling the precipitation of the mineral phase is in depth evaluated. TEM and X-ray powder diffraction reveal the crystallization of nanocrystalline apatite (Ap) in all the evaluated conditions. However, Raman spectra point out also the precipitation of amorphous calcium phosphate (ACP). This amorphous phase is more evident when RCP and Mg are at work, indicating the synergistic role of both in stabilizing the amorphous precursor. In addition, hybrid matrices are prepared to tentatively address their effectiveness as scaffolds for bone tissue engineering. SEM and AFM imaging show an homogeneous mineral distribution on the RCP matrix mineralized in presence of Mg, which provides a surface roughness similar to that found in bone. Preliminary in vitro tests with pre-osteoblast cell line show good cell-material interaction on the matrices prepared in the presence of Mg. To the best of our knowledge this work represents the first attempt to mineralize recombinant collagen type I derived protein proving the simultaneous effect of the organic phase (RCP) and Mg on ACP stabilization. This study opens the possibility to engineer, through biomineralization process, advanced hybrid matrices for bone regeneration. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. In vitro and in vivo evaluation of carbonate apatite-collagen scaffolds with some cytokines for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Sherman Salim

    2015-01-01

    Results and Conclusion: By histological observation and measurement of bone area ratio, CA-CS with cytokines showed higher bone formation ability (bFGF/CA-CS: 50.7 ± 7.3%, rh-BMP2/CA-CS: 54.2 ± 5.0% than other groups. From the limited results of this study, it is suggested that CA collagen scaffolds with some cytokines may become an attractive scaffold for bone regeneration.

  16. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  17. Collagen type I from bovine bone. Effect of animal age, bone anatomy and drying methodology on extraction yield, self-assembly, thermal behaviour and electrokinetic potential.

    Science.gov (United States)

    Ferraro, Vincenza; Gaillard-Martinie, Brigitte; Sayd, Thierry; Chambon, Christophe; Anton, Marc; Santé-Lhoutellier, Véronique

    2017-04-01

    Natural collagen is easily available from animal tissues such as bones. Main limitations reported in the use of natural collagen are heterogeneity and loss of integrity during recovery. However, its natural complexity, functionality and bioactivity still remain to be achieved through synthetic and recombinant ways. Variability of physicochemical properties of collagen extracted from bovine bone by acetic acid was then investigated taking into account endogenous and exogenous factors. Endogenous: bovine's bones age (4 and 7 years) and anatomy (femur and tibia); exogenous: thermal treatments (spray-drying and lyophilisation). Scanning electron microscopy, spectroscopy (EDS, FTIR, UV/Vis and CD), differential scanning calorimetry (DSC), centesimal composition, mass spectrometry, amino acids and zeta-potential analysis were used for the purpose. Age correlated negatively with yield of recovery and positively with minerals and proteoglycans content. Comparing the anatomy, higher yields were found for tibias, and higher stability of tibias collagen in solution was noticed. Whatever the age and the anatomy, collagens were able to renature and to self-assemble into tri-dimensional structures. Nonetheless thermal stability and kinetics of renaturation were different. Variability of natural collagen with bone age and anatomy, and drying methodology, may be a crucial advantage to conceive tailor-made applications in either the biological or technical sector. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Bone Healing in Extraction Sockets Covered With Collagen Membrane Alone or Associated With Porcine-Derived Bone Graft: a Comparative Histological and Histomorphometric Analysis.

    Science.gov (United States)

    Guarnieri, Renzo; Testarelli, Luca; Stefanelli, Luigi; De Angelis, Francesca; Mencio, Francesca; Pompa, Giorgio; Di Carlo, Stefano

    2017-01-01

    The present paper reports data of a randomized study aimed to analyse and compare the histologic and histomorphometric aspects of bone healing in extraction sites covered with collagen membrane alone or associated with porcine-derived bone graft. Thirty patients, with single extraction sockets without severe bone wall defects in the premolar/molar region, were included. Ten extraction sockets were grafted with porcine-derived bone and covered with collagen membrane (group 1), 10 sites were covered with collagen membrane alone (group 2), and 10 sites healed spontaneously (group 3). After 4 months of healing, 26 (8 in group 1, 9 in group 2, and 9 in group 3) bone core specimens were harvested for histologic evaluation, then dental implants were placed. Sites in the group 1 and in the group 2 showed similar histologic and histomorphometric results without significantly differences in the percentage of vital bone (57.43% [SD 4.8] vs. 60.01% [SD 3.2]), and non-mineralized connective tissue 22.99% (SD 5.3) vs. 18.53% (SD 6.2). In group 1 a 16.57% (SD 3.8) of residual material was found. Results showed that the use of collagen membrane alone or associated to porcine-derived bone improves the healing bone process compared to that of extraction sites spontaneously healed. Moreover, histomorphometric data related to bone quality, indicated that extraction sites without severe walls defects and with a vestibular bone thickness > 1.5 mm, treated with a low resorbtion rate collagen membrane alone, do not need more than 4 months for dental implant insertion.

  19. Poly (lactide-co-glycolide nanofibers coated with collagen and nano-hydroxyapatite for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Reza Tavakoli-Darestani

    2013-05-01

    Full Text Available Please cite this article as: Tavakoli-Darestani R, Kazemian GH, Emami M, Kamrani-Rad A. Poly (lactide-co-glycolide nanofibers coated with collagen and nano-hydroxyapatite for bone tissue engineering. Novel Biomed 2013;1:8-15.Background: A combination of polymeric nanofibrous scaffold and bioactive materials is potentially useful in bone regeneration applications.Materials and Methods: In the present study, Poly (lactide-co-glycolide (PLGA nanofibrous scaffolds, fabricated via electrospinning, were initially coated with Type I collagen and then with nano-hydroxyapatite. The prepared scaffolds were then characterized using SEM and their ability for bone regeneration was investigated in a rat critical size bone defect using digital mammography, multislice spiral-computed tomography (MSCT imaging, and histological analysis.Results: Electrospun scaffolds had nanofibrous structure with homogenous distribution of n-HA on collagen-grafted PLGA. After 8 weeks of implantation, no sign of inflammation or complication was observed at the site of surgery. According to digital mammography and MSCT, PLGA nanofibers coated simultaneously with collagen and HA showed the highest regeneration in rat calvarium. In addition, no significant difference was observed in bone repair in the group which received PLGA and the untreated control. This amount was lower than that observed in the group implanted with collagen-coated PLGA. Histological studies confirmed these data and showed osteointegration to the surrounding tissue.Conclusion: Taking all together, it was demonstrated that nanofibrous structures can be used as appropriate support for tissue-engineered scaffolds, and coating them with bioactive materials will provide ideal synthetic grafts. Fabricated PLGA coated with Type I collagen and HA can be used as new bone graft substitutes in orthopaedic surgery and is capable of enhancing bone regeneration via characteristics such as osteoconductivity and

  20. Osteogenesis Imperfecta due to Mutations in Non-Collagenous Genes-Lessons in the Biology of Bone Formation

    Science.gov (United States)

    Marini, Joan C.; Reich, Adi; Smith, Simone M.

    2014-01-01

    Purpose of Review Osteogenesis imperfecta (OI), or “brittle bone disease”, has mainly been considered a bone disorder caused by collagen mutations. Within the last decade, however, a surge of genetic discoveries has created a new paradigm for OI as a collagen-related disorder, where autosomal dominant type I collagen defects cause most cases, while rare, mostly recessive forms are due to defects in genes whose protein products interact with collagen protein. This review is both timely and relevant in outlining the genesis, development and future of this paradigm shift in the understanding of OI. Recent Findings BRIL and PEDF defects cause types V and VI OI via defective bone mineralization, while defects in CRTAP, P3H1 and CyPB cause types VII-IX via defective collagen post-translational modification. Hsp47 and FKBP65 defects cause types X and XI OI via aberrant collagen crosslinking, folding and chaperoning, while defects in SP7, WNT1, TRIC-B and OASIS disrupt osteoblast development. Finally, absence of the type I collagen C-propeptidase BMP1 causes type XII OI due to altered collagen maturation/processing. Summary Identification of these multiple causative defects has provided crucial information for accurate genetic counseling, inspired a recently proposed functional grouping of OI types by shared mechanism to simplify current nosology, and should prod investigations into common pathways in OI. Such investigations could yield critical information on cellular and bone tissue mechanisms and translate to new mechanistic insight into clinical therapies for patients. PMID:25007323

  1. Adherence, proliferation and collagen turnover by human fibroblasts seeded into different types of collagen sponges

    NARCIS (Netherlands)

    Middelkoop, E.; de Vries, H. J.; Ruuls, L.; Everts, V.; Wildevuur, C. H.; Westerhof, W.

    1995-01-01

    We describe an in vitro model that we have used to evaluate dermal substitutes and to obtain data on cell proliferation, the rate of degradation of the dermal equivalent, contractibility and de novo synthesis of collagen. We tested three classes of collagenous materials: (1) reconstituted

  2. ADHERENCE, PROLIFERATION AND COLLAGEN TURNOVER BY HUMAN FIBROBLASTS SEEDED INTO DIFFERENT TYPES OF COLLAGEN SPONGES

    NARCIS (Netherlands)

    MIDDELKOOP, E; DEVRIES, HJC; RUULS, L; EVERTS, [No Value; WILDEVUUR, CHR; WESTERHOF, W

    We describe an in vitro model that we have used to evaluate dermal substitutes and to obtain data on cell proliferation, the rate of degradation of the dermal equivalent, contractibility and de novo synthesis of collagen. We tested three classes of collagenous materials: (1) reconstituted

  3. In situ observation of fluoride-ion-induced hydroxyapatite-collagen detachment on bone fracture surfaces by atomic force microscopy

    International Nuclear Information System (INIS)

    Kindt, J H; Thurner, P J; Lauer, M E; Bosma, B L; Schitter, G; Fantner, G E; Izumi, M; Weaver, J C; Morse, D E; Hansma, P K

    2007-01-01

    The topography of freshly fractured bovine and human bone surfaces was determined by the use of atomic force microscopy (AFM). Fracture surfaces from both kinds of samples exhibited complex landscapes formed by hydroxyapatite mineral platelets with lateral dimensions ranging from ∼90 nm x 60 nm to ∼20 nm x 20 nm. Novel AFM techniques were used to study these fracture surfaces during various chemical treatments. Significant topographical changes were observed following exposure to aqueous solutions of ethylenediaminetetraacetic acid (EDTA) or highly concentrated sodium fluoride (NaF). Both treatments resulted in the apparent loss of the hydroxyapatite mineral platelets on a timescale of a few seconds. Collagen fibrils situated beneath the overlying mineral platelets were clearly exposed and could be resolved with high spatial resolution in the acquired AFM images. Time-dependent mass loss experiments revealed that the applied agents (NaF or EDTA) had very different resulting effects. Despite the fact that the two treatments exhibited nearly identical results following examination by AFM, bulk bone samples treated with EDTA exhibited a ∼70% mass loss after 72 h, whereas for the NaF-treated samples, the mass loss was only of the order of ∼10%. These results support those obtained from previous mechanical testing experiments, suggesting that enhanced formation of superficial fluoroapatite dramatically weakens the protein-hydroxyapatite interfaces. Additionally, we discovered that treatment with aqueous solutions of NaF resulted in the effective extraction of noncollagenous proteins from bone powder

  4. Ductile sliding between mineral crystals followed by rupture of collagen crosslinks: experimentally supported micromechanical explanation of bone strength.

    Science.gov (United States)

    Fritsch, Andreas; Hellmich, Christian; Dormieux, Luc

    2009-09-21

    There is an ongoing discussion on how bone strength could be explained from its internal structure and composition. Reviewing recent experimental and molecular dynamics studies, we here propose a new vision on bone material failure: mutual ductile sliding of hydroxyapatite mineral crystals along layered water films is followed by rupture of collagen crosslinks. In order to cast this vision into a mathematical form, a multiscale continuum micromechanics theory for upscaling of elastoplastic properties is developed, based on the concept of concentration and influence tensors for eigenstressed microheterogeneous materials. The model reflects bone's hierarchical organization, in terms of representative volume elements for cortical bone, for extravascular and extracellular bone material, for mineralized fibrils and the extrafibrillar space, and for wet collagen. In order to get access to the stress states at the interfaces between crystals, the extrafibrillar mineral is resolved into an infinite amount of cylindrical material phases oriented in all directions in space. The multiscale micromechanics model is shown to be able to satisfactorily predict the strength characteristics of different bones from different species, on the basis of their mineral/collagen content, their intercrystalline, intermolecular, lacunar, and vascular porosities, and the elastic and strength properties of hydroxyapatite and (molecular) collagen.

  5. MicroCT evaluation of bone mineral density loss in human bones

    International Nuclear Information System (INIS)

    Nogueira, Liebert P.; Braz, Delson; Lopes, Ricardo T.; Barroso, Regina C.; Oliveira, Luis F.

    2007-01-01

    Bone is a connective tissue largely composed of an organic protein, collagen and the inorganic mineral hydroxyapatite [Ca 10 (PO 4 ) 6 OH 2 ], which combine to provide a mechanical and supportive role in the body. Depending on the orientation of collagen fibers, two types of bone can be distinguished: trabecular and cortical bone. Degree of mineralization is considered an important feature of bone quality. Changes in the degree of mineralization is generally due to osteoporosis, but many recent studies have already shown that alterations in degree of mineralization can occur due to a large variety of factors. The transmission X-ray microtomography is one of the most popular methods, which provides the spatial distribution of the total absorption coefficient inside the sample. The aim of this study was to investigate the suitability of using microCT as a supplementary tool for the diagnosis of the health status of human bones. Eleven samples were constructed simulating the physiological range of bone mineral density (BMD) found in cortical human bone. The samples represent healthy mixtures of swine compact bone dried at room temperature, powdered and mixed with fat (0 - 100 % by mass). The samples were imaged by a microfocus tube (Fein-Focus) with focal size of about 60 μm (±5%), and a CCD camera (0.143 mm pixel size) coupled with an intensifier tube with fluoroscope screen at the Nuclear Instrumentation Laboratory (COPPE/UFRJ), Brazil. The images were reconstructed and treated with suitable software developed at the Nuclear Instrumentation Laboratory. The mineral content in cortical bone is defined by the volume of dry, fat-free bone per unit bulk volume of the bone. The volumes were calculated from the bone density using the relationship between volume and density. The densities of fat and bone were taken to be 0.95 g.cm -3 and 1.92 g.cm -3 respectively. The correlation of the measured absorption coefficient with the mineral content in the samples was then

  6. MicroCT evaluation of bone mineral density loss in human bones

    Energy Technology Data Exchange (ETDEWEB)

    Nogueira, Liebert P.; Braz, Delson; Lopes, Ricardo T. [Universidade Federal do Rio de Janeiro (UFRJ), RJ (Brazil). Coordenacao dos Programas de Pos-graduacao de Engenharia (COPPE). Lab. de Instrumentacao Nuclear]. E-mails: lnogueira@con.ufrj.br; Barroso, Regina C.; Oliveira, Luis F. [Universidade do Estado do Rio de Janeiro (UERJ), Rio de Janeiro, RJ (Brazil). Inst. de Fisica]. E-mail: cely@uerj.br

    2007-07-01

    Bone is a connective tissue largely composed of an organic protein, collagen and the inorganic mineral hydroxyapatite [Ca{sub 10}(PO{sub 4}){sub 6}OH{sub 2}], which combine to provide a mechanical and supportive role in the body. Depending on the orientation of collagen fibers, two types of bone can be distinguished: trabecular and cortical bone. Degree of mineralization is considered an important feature of bone quality. Changes in the degree of mineralization is generally due to osteoporosis, but many recent studies have already shown that alterations in degree of mineralization can occur due to a large variety of factors. The transmission X-ray microtomography is one of the most popular methods, which provides the spatial distribution of the total absorption coefficient inside the sample. The aim of this study was to investigate the suitability of using microCT as a supplementary tool for the diagnosis of the health status of human bones. Eleven samples were constructed simulating the physiological range of bone mineral density (BMD) found in cortical human bone. The samples represent healthy mixtures of swine compact bone dried at room temperature, powdered and mixed with fat (0 - 100 % by mass). The samples were imaged by a microfocus tube (Fein-Focus) with focal size of about 60 {mu}m ({+-}5%), and a CCD camera (0.143 mm pixel size) coupled with an intensifier tube with fluoroscope screen at the Nuclear Instrumentation Laboratory (COPPE/UFRJ), Brazil. The images were reconstructed and treated with suitable software developed at the Nuclear Instrumentation Laboratory. The mineral content in cortical bone is defined by the volume of dry, fat-free bone per unit bulk volume of the bone. The volumes were calculated from the bone density using the relationship between volume and density. The densities of fat and bone were taken to be 0.95 g.cm{sup -3} and 1.92 g.cm{sup -3} respectively. The correlation of the measured absorption coefficient with the mineral content

  7. Graphene oxide improves the biocompatibility of collagen membranes in an in vitro model of human primary gingival fibroblasts.

    Science.gov (United States)

    De Marco, Patrizia; Zara, Susi; De Colli, Marianna; Radunovic, Milena; Lazović, Vladimir; Ettorre, Valeria; Di Crescenzo, Antonello; Piattelli, Adriano; Cataldi, Amelia; Fontana, Antonella

    2017-09-13

    Commercial collagen membranes are used in oral surgical procedures as scaffolds for bone deposition in guided bone regeneration. Here, we have enriched them with graphene oxide (GO) via a simple non-covalent functionalization, exploiting the capacity of oxygenated carbon functional moieties of GO to interact through hydrogen bonding with collagen. In the present paper, the GO-coated membranes have been characterized in terms of stability, nano-roughness, biocompatibility and induction of inflammatory response in human primary gingival fibroblast cells. The obtained coated membranes are demonstrated not to leak GO in the bulk solution, and to change some features of the membrane, such as stiffness and adhesion between the membrane and the atomic force microscopy (AFM) tip. Moreover, the presence of GO increases the roughness and the total surface exposed to the cells, as demonstrated by AFM analyses. The obtained material is biocompatible, and does not induce inflammation in the tested cells.

  8. Experimental Evaluation of the Effectiveness of Demineralized Bone Matrix and Collagenated Heterologous Bone Grafts Used Alone or in Combination with Platelet-Rich Fibrin on Bone Healing in Sinus Floor Augmentation.

    Science.gov (United States)

    Peker, Elif; Karaca, Inci Rana; Yildirim, Benay

    2016-01-01

    The aim of this study was an experimental evaluation of the effectiveness of demineralized bone matrix (DBM) and collagenated heterologous bone graft (CHBG) used alone or in combination with platelet-rich fibrin on bone healing in sinus floor augmentation procedures. In this study, 36 New Zealand rabbits were used. The bilateral sinus elevation was performed, and 72 defects were obtained. The rabbit maxillary sinuses were divided into four groups according to the augmentation biomaterials obtained: demineralized bone matrix (Grafton DBM Putty, Osteotech; DBM group), DBM combined with platelet-rich fibrin (PRF; DBM + PRF group), collagenated heterologous bone graft (CHBG; Apatos Mix, OsteoBiol, Tecnoss; CHBG group), CHBG combined with PRF (CHBG + PRF group). All groups were sacrificed at 2, 4, and 8 weeks after surgery for histologic, histomorphometric, and immunohistochemical analyses. The inflammatory reaction was moderate to intense at the second week in all groups and declined from 2 to 8 weeks. New bone formation was started at the second week and increased from 2 to 8 weeks in all groups. There was no significant difference in bone formation between the experimental groups that used PRF mixed graft material and control groups that used only graft material. The percentage of new bone formation showed a significant difference in DBM groups and DBM + PRF groups compared with other groups. There were osteoclasts around all the bone graft materials used, but the percentage of residual graft particles was significantly higher in CHBG groups and CHBG + PRF groups at the eighth week. There is no beneficial effect of the application of PRF in combination with demineralized bone matrix or collagenated heterologous bone graft on bone formation in sinus floor augmentation. The results of this study showed that both collagenated heterologous bone graft and demineralized bone matrix have osteoconductive properties, but demineralized bone matrix showed more bone formation

  9. In vitro and in vivo evaluation of bone morphogenetic protein-2 (BMP-2) immobilized collagen-coated polyetheretherketone (PEEK)

    Science.gov (United States)

    Du, Ya-Wei; Zhang, Li-Nan; Ye, Xin; Nie, He-Min; Hou, Zeng-Tao; Zeng, Teng-Hui; Yan, Guo-Ping; Shang, Peng

    2015-03-01

    Polyetheretherketone (PEEK) is regarded as one of the most potential candidates of biomaterials in spinal implant applications. However, as a bioinert material, PEEK plays a limited role in osteoconduction and osseointegration. In this study, recombinant human bone morphogenetic protein-2 (rhBMP-2) was immobilized onto the surface of collagen-coated PEEK in order to prepare a multi-functional material. After adsorbed onto the PEEK surface by hydrophobic interaction, collagen was cross-linked with N-(3-dimethylaminopropyl)-N'-ethyl carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). EDC/NHS system also contributed to the immobilization of rhBMP-2. Water contact angle tests, XPS and SEM clearly demonstrated the surface changes. ELISA tests quantified the amount of rhBMP-2 immobilized and the release over a period of 30 d. In vitro evaluation proved that the osteogenesis differentiation rate was higher when cells were cultured on modified PEEK discs than on regular ones. In vivo tests were conducted and positive changes of major parameters were presented. This report demonstrates that the rhBMP-2 immobilized method for PEEK modification increase bioactivity in vitro and in vivo, suggesting its practicability in orthopedic and spinal clinical applications.

  10. Bone collagen stable carbon and nitrogen isotope variability in modern South Australian mammals: A baseline for palaeoecological inferences

    International Nuclear Information System (INIS)

    Pate, F.D.; Anson, T.J.; Noble, A.H.; Schoeninger, M.J.

    1997-01-01

    Cortical bone samples were collected from a range of modern mammals at four field sites along a 1225 km north-south transect from temperate coastal to arid interior South Australia in order to address variability in stable carbon and nitrogen isotope composition. Collection sites were located along the eastern border of the state and included Mount Gambier, Karte, Plumbago and Innamincka. Mean annual rainfall along the transect ranges from 700-800 mm at Mount Gambier to 150-200 mm at Innamincka. Bone collagen carbon and nitrogen isotope values become more positive toward the arid north in relation to increasing quantities of C-4 plants and decreasing amounts of rainfall. respectively. In addition, carnivores and herbivores can be differentiated by stable nitrogen isotope values. On average, carnivore bone collagen is approximately 6 per mil more positive than that of rabbits at Mount Gambier but only 2.6 - 3.4 per mil more positive at the three arid collection sites. In general, the large eutherian herbivores have mean bone collagen δ15N values that are 1.4 - 2.3 per mil more positive than those of the marsupial herbivores. Eutherian and marsupial bone collagen δ15N differences only disappear at the most arid collection site, Innamincka

  11. Bone collagen stable carbon and nitrogen isotope variability in modern South Australian mammals: A baseline for palaeoecological inferences.

    Energy Technology Data Exchange (ETDEWEB)

    Pate, F.D.; Anson, T.J.; Noble, A.H. [Flinders Univ. of South Australia, Bedford Park, SA (Australia). Department of Archaeology; Schoeninger, M.J. [Wisconsin Univ., Madison, WI (United States). Department of Anthropology

    1997-12-31

    Cortical bone samples were collected from a range of modern mammals at four field sites along a 1225 km north-south transect from temperate coastal to arid interior South Australia in order to address variability in stable carbon and nitrogen isotope composition. Collection sites were located along the eastern border of the state and included Mount Gambier, Karte, Plumbago and Innamincka. Mean annual rainfall along the transect ranges from 700-800 mm at Mount Gambier to 150-200 mm at Innamincka. Bone collagen carbon and nitrogen isotope values become more positive toward the arid north in relation to increasing quantities of C-4 plants and decreasing amounts of rainfall. respectively. In addition, carnivores and herbivores can be differentiated by stable nitrogen isotope values. On average, carnivore bone collagen is approximately 6 per mil more positive than that of rabbits at Mount Gambier but only 2.6 - 3.4 per mil more positive at the three arid collection sites. In general, the large eutherian herbivores have mean bone collagen {delta}15N values that are 1.4 - 2.3 per mil more positive than those of the marsupial herbivores. Eutherian and marsupial bone collagen {delta}15N differences only disappear at the most arid collection site, Innamincka.

  12. Synergistic intrafibrillar/extrafibrillar mineralization of collagen scaffolds based on a biomimetic strategy to promote the regeneration of bone defects

    Directory of Open Access Journals (Sweden)

    Wang Y

    2016-05-01

    Full Text Available Yao Wang,1 Ngo Van Manh,1,2 Haorong Wang,1 Xue Zhong,1 Xu Zhang,1 Changyi Li1 1School of Dentistry, Hospital of Stomatology, Tianjin Medical University, Tianjin, People’s Republic of China; 2Thaibinh University of Medicine and Pharmacy, Thaibinh, Vietnam Abstract: The mineralization of collagen scaffolds can improve their mechanical properties and biocompatibility, thereby providing an appropriate microenvironment for bone regeneration. The primary purpose of the present study is to fabricate a synergistically intra- and extrafibrillar mineralized collagen scaffold, which has many advantages in terms of biocompatibility, biomechanical properties, and further osteogenic potential. In this study, mineralized collagen scaffolds were fabricated using a traditional mineralization method (ie, immersed in simulated body fluid as a control group and using a biomimetic method based on the polymer-induced liquid precursor process as an experimental group. In the polymer-induced liquid precursor process, a negatively charged polymer, carboxymethyl chitosan (CMC, was used to stabilize amorphous calcium phosphate (ACP to form nanocomplexes of CMC/ACP. Collagen scaffolds mineralized based on the polymer-induced liquid precursor process were in gel form such that nanocomplexes of CMC/ACP can easily be drawn into the interstices of the collagen fibrils. Scanning electron microscopy and transmission electron microscopy were used to examine the porous micromorphology and synergistic mineralization pattern of the collagen scaffolds. Compared with simulated body fluid, nanocomplexes of CMC/ACP significantly increased the modulus of the collagen scaffolds. The results of in vitro experiments showed that the cell count and differentiated degrees in the experimental group were higher than those in the control group. Histological staining and micro-computed tomography showed that the amount of new bone regenerated in the experimental group was larger than that in the

  13. The growth of human fibroblasts and A431 epidermoid carcinoma cells on gamma-irradiated human amnion collagen substrata.

    Science.gov (United States)

    Liu, B; Harrell, R; Lamb, D J; Dresden, M H; Spira, M

    1989-10-15

    Human fibroblasts and A431 human epidermoid carcinoma cells were cultured on gamma-irradiated human amnion collagen as well as on plastic dishes and non-irradiated collagen coated dishes. The morphology, attachment, growth and short-term cytotoxicity of these culture conditions have been determined. Both irradiated and non-irradiated amnion collagen enhanced the attachment and proliferation of fibroblasts as compared to the plastic dishes. No differences in these properties were observed for A431 cells cultured on irradiated collagen when compared with culture on non-irradiated collagen substrates. Cytotoxicity assays showed that irradiated and non-irradiated collagens were not cytotoxic for either fibroblasts or A431 cells. The results demonstrated that amnion collagen irradiated at doses of 0.25-2.0 Mrads is optimal for cell growth.

  14. Nanofibrous yet injectable polycaprolactone-collagen bone tissue scaffold with osteoprogenitor cells and controlled release of bone morphogenetic protein-2

    Energy Technology Data Exchange (ETDEWEB)

    Subramanian, Gayathri; Bialorucki, Callan [Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606 (United States); Yildirim-Ayan, Eda, E-mail: eda.yildirimayan@utoledo.edu [Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606 (United States); Department of Orthopaedic Surgery, University of Toledo Medical Center, Toledo, OH 43614 (United States)

    2015-06-01

    In this work, we developed a nanofibrous, yet injectable orthobiologic tissue scaffold that is capable of hosting osteoprogenitor cells and controlling kinetic release profile of the encapsulated pro-osteogenic factor without diminishing its bioactivity over 21 days. This innovative injectable scaffold was synthesized by incorporating electrospun and subsequently O{sub 2} plasma-functionalized polycaprolactone (PCL) nanofibers within the collagen type-I solution along with MC3T3-E1 cells (pre-osteoblasts) and bone morphogenetic protein-2 (BMP2). Through changing the PCL nanofiber concentration within the injectable scaffolds, we were able to tailor the mechanical strength, protein retention capacity, bioactivity preservation, and osteoinductive potential of the scaffolds. The nanofibrous internal structure of the scaffold allowed us to use a low dose of BMP2 (200 ng/ml) to achieve osteoblastic differentiation in in vitro culture. The osteogenesis capacity of the injectable scaffolds were evaluated though measuring MC3T3-E1 cell proliferation, ALP activity, matrix mineralization, and early- and late-osteoblast specific gene expression profiles over 21 days. The results demonstrated that the nanofibrous injectable scaffold provides not only an osteoinductive environment for osteoprogenitor cells to differentiate, but also a suitable biomechanical and biochemical environment to act as a reservoir for osteogenic factors with controlled release profile. - Highlights: • Injectable nanofibrous scaffold with osteoprogenitor cells and BMP2 was synthesized. • PCL nanofiber concentration within collagen scaffold affected the BMP2 retention and bioactivity. • Optimal PCL concentration was identified for mechanical stability, injectability, and osteogenic activity. • Scaffolds exhibited long-term osteoinductive capacity for bone repair and regeneration.

  15. Sika Deer Antler Collagen Type I-Accelerated Osteogenesis in Bone Marrow Mesenchymal Stem Cells via the Smad Pathway

    Directory of Open Access Journals (Sweden)

    Na Li

    2016-01-01

    Full Text Available Deer antler preparations have been used to strengthen bones for centuries. It is particularly rich in collagen type I. This study aimed to unravel part of the purported bioremedial effect of Sika deer antler collagen type I (SDA-Col I on bone marrow mesenchymal stem cells. The results suggest that SDA-Col I might be used to promote and regulate osteoblast proliferation and differentiation. SDA-Col I might potentially provide the basis for novel therapeutic strategies in the treatment of bone injury and/or in scaffolds for bone replacement strategies. Finally, isolation of SDA-Col I from deer antler represents a renewable, green, and uncomplicated way to obtain a biomedically valuable therapeutic.

  16. PPAR-δ Agonist With Mesenchymal Stem Cells Induces Type II Collagen-Producing Chondrocytes in Human Arthritic Synovial Fluid.

    Science.gov (United States)

    Heck, Bruce E; Park, Joshua J; Makani, Vishruti; Kim, Eun-Cheol; Kim, Dong Hyun

    2017-08-01

    Osteoarthritis (OA) is an inflammatory joint disease characterized by degeneration of articular cartilage within synovial joints. An estimated 27 million Americans suffer from OA, and the population is expected to reach 67 million in the United States by 2030. Thus, it is urgent to find an effective treatment for OA. Traditional OA treatments have no disease-modifying effect, while regenerative OA therapies such as autologous chondrocyte implantation show some promise. Nonetheless, current regenerative therapies do not overcome synovial inflammation that suppresses the differentiation of mesenchymal stem cells (MSCs) to chondrocytes and the expression of type II collagen, the major constituent of functional cartilage. We discovered a synergistic combination that overcame synovial inflammation to form type II collagen-producing chondrocytes. The combination consists of peroxisome proliferator-activated receptor (PPAR) δ agonist, human bone marrow (hBM)-derived MSCs, and hyaluronic acid (HA) gel. Interestingly, those individual components showed their own strong enhancing effects on chondrogenesis. GW0742, a PPAR-δ agonist, greatly enhanced MSC chondrogenesis and the expression of type II collagen and glycosaminoglycan (GAG) in hBM-MSC-derived chondrocytes. GW0742 also increased the expression of transforming growth factor β that enhances chondrogenesis and suppresses cartilage fibrillation, ossification, and inflammation. HA gel also increased MSC chondrogenesis and GAG production. However, neither GW0742 nor HA gel could enhance the formation of type II collagen-producing chondrocytes from hBM-MSCs within human OA synovial fluid. Our data demonstrated that the combination of hBM-MSCs, PPAR-δ agonist, and HA gel significantly enhanced the formation of type II collagen-producing chondrocytes within OA synovial fluid from 3 different donors. In other words, the novel combination of PPAR-δ agonist, hBM-MSCs, and HA gel can overcome synovial inflammation to form

  17. A new procedure for extraction of collagen from modern and archaeological bones for {sup 14}C dating

    Energy Technology Data Exchange (ETDEWEB)

    Maspero, F. [CUDaM, University of Milano Bicocca, Milano (Italy); Sala, S.; Papagni, A. [University of Milano Bicocca, Materials Science Department, Milano (Italy); Fedi, M.E. [INFN sezione di Firenze, Sesto Fiorentino (Firenze) (Italy); Martini, M. [CUDaM, University of Milano Bicocca, Milano (Italy); University of Milano Bicocca, Materials Science Department, Milano (Italy); INFN sezione di Milano Bicocca, Milano (Italy)

    2011-10-15

    Bones are potentially the best age indicators in a stratigraphic study, because they are closely related to the layer in which they are found. Collagen is the most suitable fraction and is the material normally used in radiocarbon dating. Bone contaminants can strongly alter the carbon isotopic fraction values of the samples, so chemical pretreatment for {sup 14} C dating by accelerator mass spectrometry (AMS) is essential. The most widespread method for collagen extraction is based on the Longin procedure, which consists in HCl demineralization to dissolve the inorganic phase of the samples, followed by dissolution of collagen in a weak acid solution. In this work the possible side effects of this procedure on a modern bone are presented; the extracted collagen was analyzed by ATR-IR spectroscopy. An alternative procedure, based on use of HF instead of HCl, to minimize unwanted degradation of the organic fraction, is also given. A study by ATR-IR spectroscopic analysis of collagen collected after different demineralization times and with different acid volumes, and a study of an archaeological sample, are also presented. (orig.)

  18. Calcium phosphate fibers coated with collagen: In vivo evaluation of the effects on bone repair.

    Science.gov (United States)

    Ueno, Fabio Roberto; Kido, Hueliton Wilian; Granito, Renata Neves; Gabbai-Armelin, Paulo Roberto; Magri, Angela Maria Paiva; Fernandes, Kelly Rosseti; da Silva, Antonio Carlos; Braga, Francisco José Correa; Renno, Ana Claudia Muniz

    2016-08-12

    The aim of this study was to assess the characteristics of the CaP/Col composites, in powder and fiber form, via scanning electron microscopy (SEM), pH and calcium release evaluation after immersion in SBF and to evaluate the performance of these materials on the bone repair process in a tibial bone defect model. For this, four different formulations (CaP powder - CaPp, CaP powder with collagen - CaPp/Col, CaP fibers - CaPf and CaP fibers with collagen - CaPf/Col) were developed. SEM images indicated that both material forms were successfully coated with collagen and that CaPp and CaPf presented HCA precursor crystals on their surface. Although presenting different forms, FTIR analysis indicated that CaPp and CaPf maintained the characteristic peaks for this class of material. Additionally, the calcium assay study demonstrated a higher Ca uptake for CaPp compared to CaPf for up to 5 days. Furthermore, pH measurements revealed that the collagen coating prevented the acidification of the medium, leading to higher pH values for CaPp/Col and CaPf/Col. The histological analysis showed that CaPf/Col demonstrated a higher amount of newly formed bone in the region of the defect and a reduced presence of material. In summary, the results indicated that the fibrous CaP enriched with the organic part (collagen) glassy scaffold presented good degradability and bone-forming properties and also supported Runx2 and RANKL expression. These results show that the present CaP/Col fibrous composite may be used as a bone graft for inducing bone repair.

  19. Ribose mediated crosslinking of collagen-hydroxyapatite hybrid scaffolds for bone tissue regeneration using biomimetic strategies.

    Science.gov (United States)

    Krishnakumar, Gopal Shankar; Gostynska, Natalia; Campodoni, Elisabetta; Dapporto, Massimiliano; Montesi, Monica; Panseri, Silvia; Tampieri, Anna; Kon, Elizaveta; Marcacci, Maurilio; Sprio, Simone; Sandri, Monica

    2017-08-01

    This study explores for the first time the application of ribose as a highly biocompatible agent for the crosslinking of hybrid mineralized constructs, obtained by bio-inspired mineralization of self-assembling Type I collagen matrix with magnesium-doped-hydroxyapatite nanophase, towards a biomimetic mineralized 3D scaffolds (MgHA/Coll) with excellent compositional and structural mimicry of bone tissue. To this aim, two different crosslinking mechanisms in terms of pre-ribose glycation (before freeze drying) and post-ribose glycation (after freeze drying) were investigated. The obtained results explicate that with controlled freeze-drying, highly anisotropic porous structures with opportune macro-micro porosity are obtained. The physical-chemical features of the scaffolds characterized by XRD, FTIR, ICP and TGA demonstrated structural mimicry analogous to the native bone. The influence of ribose greatly assisted in decreasing solubility and increased enzymatic resistivity of the scaffolds. In addition, enhanced mechanical behaviour in response to compressive forces was achieved. Preliminary cell culture experiments reported good cytocompatibility with extensive cell adhesion, proliferation and colonization. Overall, scaffolds developed by pre-ribose glycation process are preferred, as the related crosslinking technique is more facile and robust to obtain functional scaffolds. As a proof of concept, we have demonstrated that ribose crosslinking is cost-effective, safe and functionally effective. This study also offers new insights and opportunities in developing promising scaffolds for bone tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. When size matters: differences in demineralized bone matrix particles affect collagen structure, mesenchymal stem cell behavior, and osteogenic potential.

    Science.gov (United States)

    Dozza, B; Lesci, I G; Duchi, S; Della Bella, E; Martini, L; Salamanna, F; Falconi, M; Cinotti, S; Fini, M; Lucarelli, E; Donati, D

    2017-04-01

    Demineralized bone matrix (DBM) is a natural, collagen-based, osteoinductive biomaterial. Nevertheless, there are conflicting reports on the efficacy of this product. The purpose of this study was to evaluate whether DBM collagen structure is affected by particle size and can influence DBM cytocompatibility and osteoinductivity. Sheep cortical bone was ground and particles were divided in three fractions with different sizes, defined as large (L, 1-2 mm), medium (M, 0.5-1 mm), and small (S, structure, with DBM-M being altered but not as much as DBM-S. DBM-M displayed a preferable trend in almost all biological characteristics tested, although all DBM particles revealed an optimal cytocompatibility. Subcutaneous implantation of DBM particles into immunocompromised mice resulted in bone induction only for DBM-M. When sheep MSC were seeded onto particles before implantation, all DBM particles were able to induce new bone formation with the best incidence for DBM-M and DBM-S. In conclusion, the collagen alteration in DBM-M is likely the best condition to promote bone induction in vivo. Furthermore, the choice of 0.5-1 mm particles may enable to obtain more efficient and consistent results among different research groups in bone tissue-engineering applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1019-1033, 2017. © 2017 Wiley Periodicals, Inc.

  1. Protease inhibitors enhance extracellular collagen fibril deposition in human mesenchymal stem cells.

    Science.gov (United States)

    Han, Sejin; Li, Yuk Yin; Chan, Barbara Pui

    2015-10-15

    Collagen is a widely used naturally occurring biomaterial for scaffolding, whereas mesenchymal stem cells (MSCs) represent a promising cell source in tissue engineering and regenerative medicine. It is generally known that cells are able to remodel their environment by simultaneous degradation of the scaffolds and deposition of newly synthesized extracellular matrix. Nevertheless, the interactions between MSCs and collagen biomaterials are poorly known, and the strategies enhancing the extracellular matrix deposition are yet to be defined. In this study, we aim to investigate the fate of collagen when it is in contact with MSCs and hypothesize that protease inhibition will enhance their extracellular deposition of collagen fibrils. Specifically, human MSCs (hMSCs) were exposed to fluorescence-labeled collagen with and without intracellular or extracellular protease inhibitors (or both) before tracing the collagen at both intracellular and extracellular spaces. Collagen were internalized by hMSCs and degraded intracellularly in lysosomes. In the presence of protease inhibitors, both intracellular collagen fibril growth and extracellular deposition of collagen fibrils were enhanced. Moreover, protease inhibitors work synergistically with ascorbic acid, a well-known matrix deposition-enhancing reagent, in further enhancing collagen fibril deposition at the extracellular space. These findings provide a better understanding of the interactions between hMSCs and collagen biomaterials and suggest a method to manipulate matrix remodeling and deposition of hMSCs, contributing to better scaffolding for tissue engineering and regenerative medicine.

  2. Identification and characterization of the human type II collagen gene (COL2A1).

    OpenAIRE

    Cheah, Kathryn; Stoker, N.G.; Griffin, J.R.; Grosveld, Frank; Solomon, E.

    1985-01-01

    textabstractThe gene contained in the human cosmid clone CosHcol1, previously designated an alpha 1(I) collagen-like gene, has now been identified. CosHcol1 hybridizes strongly to a single 5.9-kilobase mRNA species present only in tissue in which type II collagen is expressed. DNA sequence analysis shows that this clone is highly homologous to the chicken alpha 1(II) collagen gene. These data together suggest that CosHcol1 contains the human alpha 1(II) collagen gene COL2A1. The clone appears...

  3. Observations on morphologic changes in the aging and degenerating human disc: Secondary collagen alterations

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2002-03-01

    Full Text Available Abstract Background In the annulus, collagen fibers that make up the lamellae have a wavy, planar crimped pattern. This crimping plays a role in disc biomechanical function by allowing collagen fibers to stretch during compression. The relationship between morphologic changes in the aging/degenerating disc and collagen crimping have not been explored. Methods Ultrastructural studies were performed on annulus tissue from 29 control (normal donors (aged newborn to 79 years and surgical specimens from 49 patients (aged 16 to 77 years. Light microscopy and specialized image analysis to visualize crimping was performed on additional control and surgical specimens. Human intervertebral disc tissue from the annulus was obtained in a prospective morphologic study of the annulus. Studies were approved by the authors' Human Subjects Institutional Review Board. Results Three types of morphologic changes were found to alter the crimping morphology of collagen: 1 encircling layers of unusual matrix disrupted the lamellar collagen architecture; 2 collagen fibers were reduced in amount, and 3 collagen was absent in regions with focal matrix loss. Conclusions Although proteoglycan loss is well recognized as playing a role in the decreased shock absorber function of the aging/degenerating disc, collagen changes have received little attention. This study suggests that important stretch responses of collagen made possible by collagen crimping may be markedly altered by morphologic changes during aging/degeneration and may contribute to the early tissue changes involved in annular tears.

  4. Tendon collagen synthesis declines with immobilization in elderly humans

    DEFF Research Database (Denmark)

    Dideriksen, Kasper; Boesen, Anders P; Reitelseder, Søren

    2017-01-01

    -80 yr) were randomly assigned to NSAIDs (ibuprofen 1,200 mg/day; Ibu) or placebo (Plc). One lower limb was immobilized in a cast for 2 wk and retrained for 6 wk. Tendon collagen protein synthesis, mechanical properties, size, expression of genes related to collagen turnover and remodeling, and signal...... intensity (from magnetic resonance imaging) were investigated. Tendon collagen synthesis decreased (P ... immobilization in both groups, whereas scleraxis mRNA decreased with inactivity in the Plc group only (P collagen protein synthesis decreased after 2 wk of immobilization, whereas tendon stiffness and modulus were only marginally reduced, and NSAIDs had no influence upon this...

  5. Estrogen depletion and drug treatment alter the microstructure of type I collagen in bone

    Directory of Open Access Journals (Sweden)

    Meagan A. Cauble

    2016-12-01

    Full Text Available The impact of estrogen depletion and drug treatment on type I collagen fibril nanomorphology and collagen fibril packing (microstructure was evaluated by atomic force microscopy (AFM using an ovariectomized (OVX rabbit model of estrogen deficiency induced bone loss. Nine month-old New Zealand white female rabbits were treated as follows: sham-operated (Sham; n = 11, OVX + vehicle (OVX + Veh; n = 12, OVX + alendronate (ALN, 600 μg/kg/wk., s.c.; n = 12, and OVX + cathepsin-K inhibitor L-235 (CatKI, 10 mg/kg, daily, p.o.; n = 13 in prevention mode for 27 weeks. Samples from the cortical femur and trabecular lumbar vertebrae were polished, demineralized, and imaged using AFM. Auto-correlation of image patches was used to generate a vector field for each image that mathematically approximated the collagen fibril alignment. This vector field was used to compute an information-theoretic entropy that was employed as a quantitative fibril alignment parameter (FAP to allow image-to-image and sample-to-sample comparison. For all samples, no change was observed in the average FAP values; however significant differences in the distribution of FAP values were observed. In particular, OVX + Veh lumbar vertebrae samples contained a tail of lower FAP values representing regions of greater fibril alignment. OVX + ALN treatment resulted in a FAP distribution with a tail indicating greater alignment for cortical femur and less alignment for trabecular lumbar vertebrae. OVX + CatKI treatment gave a distribution of FAP values with a tail indicating less alignment for cortical femur and no change for trabecular lumbar vertebrae. Fibril alignment was also evaluated by considering when a fibril was part of discrete bundles or sheets (classified as parallel or not (classified as oblique. For this analysis, the percentage of parallel fibrils in cortical femur for the OVX group was 17% lower than the Sham group. OVX + ALN treatment partially

  6. Injectable calcium sulfate/mineralized collagen-based bone repair materials with regulable self-setting properties.

    Science.gov (United States)

    Chen, Zonggang; Liu, Huanye; Liu, Xi; Cui, Fu-Zhai

    2011-12-15

    An injectable and self-setting bone repair materials (nano-hydroxyapatite/collagen/calcium sulfate hemihydrate, nHAC/CSH) was developed in this study. The nano-hydroxyapatite/collagen (nHAC) composite, which is the mineralized fibril by self-assembly of nano-hydrocyapatite and collagen, has the same features as natural bone in both main hierarchical microstructure and composition. It is a bioactive osteoconductor due to its high level of biocompatibility and appropriate degradation rate. However, this material lacks handling characteristics because of its particle or solid-preformed block shape. Herein, calcium sulfate hemihydrate (CSH) was introduced into nHAC to prepare an injectable and self-setting in situ bone repair materials. The morphology of materials was observed using SEM. Most important and interesting of all, calcium sulfate dihydrate (CSD), which is not only the reactant of preparing CSH but also the final solidified product of CSH, was introduced into nHAC as setting accelerator to regulate self-setting properties of injectable nHAC/CSH composite, and thus the self-setting time of nHAC/CSH composite can be regulated from more than 100 min to about 30 min and even less than 20 min by adding various amount of setting accelerator. The compressive properties of bone graft substitute after final setting are similar to those of cancellous bone. CSD as an excellent setting accelerator has no significant effect on the mechanical property and degradability of bone repair materials. In vitro biocompatibility and in vivo histology studies demonstrated that the nHAC/CSH composite could provide more adequate stimulus for cell adhesion and proliferation, embodying favorable cell biocompatibility and a strong ability to accelerate bone formation. It can offer a satisfactory biological environment for growing new bone in the implants and for stimulating bone formation. Copyright © 2011 Wiley Periodicals, Inc.

  7. Clinical assessment of bone quality of human extraction sockets after conversion with growth factors.

    Science.gov (United States)

    Ntounis, Athanasios; Geurs, Nico; Vassilopoulos, Philip; Reddy, Michael

    2015-01-01

    The study was conducted to evaluate the effect of mineralized freeze-dried bone allograft (FDBA), alone or in combination with growth factors in extraction sockets, on subjective assessment of bone quality during implant placement. Forty-one patients whose treatment plan involved extraction of anterior or premolar teeth were randomized into four groups: Group 1, collagen plug (control); Group 2, FDBA/β-tricalcium phosphate (β-TCP)/collagen plug; Group 3, FDBA/β-TCP/platelet-rich plasma (PRP)/collagen plug; Group 4, FDBA/β-TCP/recombinant human platelet-derived growth factor BB (rhPDGF-BB)/collagen plug. After 8 weeks of healing, implants were placed. The clinicians assessed bone quality according to the Misch classification. A benchtop calibration exercise test was conducted to evaluate agreement and accuracy of operators in recognizing different bone qualities. Differences were analyzed using one-way analysis of variance (ANOVA) or chi-square tests for continuous and categorical data. Pairwise comparisons were tested using least squares means (LS means). Spearman correlation coefficients were used to evaluate the relationship of bone growth with potential confounders. P .05). Inclusion of bone grafting is associated with a shift from D4 quality to D3 quality bone. Inclusion of PRP in bone grafting eliminates the incidence of D4 bone, establishing D3 and D2 quality bone as prevalent (56% vs. 42%, respectively). Inclusion of rhPDGF-BB and β-TCP in combination with the bone grafting has the same effect, although D2 quality is less prevalent. When compared to sockets grafted with FDBA/β-TCP/collagen plug alone, the sockets with growth factors demonstrated fewer residual bone graft particles. (1) Inclusion of bone grafting enhanced bone quality as assessed during implant placement. (2) Overall inclusion of PRP and rhPDGF-BB enhanced subjective bone quality, eliminating incidence of D4 quality in human extraction sockets. (3) The use of PRP or rhPDGF-BB may

  8. The identification of proteoglycans and glycosaminoglycans in archaeological human bones and teeth.

    Science.gov (United States)

    Coulson-Thomas, Yvette M; Coulson-Thomas, Vivien J; Norton, Andrew L; Gesteira, Tarsis F; Cavalheiro, Renan P; Meneghetti, Maria Cecília Z; Martins, João R; Dixon, Ronald A; Nader, Helena B

    2015-01-01

    Bone tissue is mineralized dense connective tissue consisting mainly of a mineral component (hydroxyapatite) and an organic matrix comprised of collagens, non-collagenous proteins and proteoglycans (PGs). Extracellular matrix proteins and PGs bind tightly to hydroxyapatite which would protect these molecules from the destructive effects of temperature and chemical agents after death. DNA and proteins have been successfully extracted from archaeological skeletons from which valuable information has been obtained; however, to date neither PGs nor glycosaminoglycan (GAG) chains have been studied in archaeological skeletons. PGs and GAGs play a major role in bone morphogenesis, homeostasis and degenerative bone disease. The ability to isolate and characterize PG and GAG content from archaeological skeletons would unveil valuable paleontological information. We therefore optimized methods for the extraction of both PGs and GAGs from archaeological human skeletons. PGs and GAGs were successfully extracted from both archaeological human bones and teeth, and characterized by their electrophoretic mobility in agarose gel, degradation by specific enzymes and HPLC. The GAG populations isolated were chondroitin sulfate (CS) and hyaluronic acid (HA). In addition, a CSPG was detected. The localization of CS, HA, three small leucine rich PGs (biglycan, decorin and fibromodulin) and glypican was analyzed in archaeological human bone slices. Staining patterns were different for juvenile and adult bones, whilst adolescent bones had a similar staining pattern to adult bones. The finding that significant quantities of PGs and GAGs persist in archaeological bones and teeth opens novel venues for the field of Paleontology.

  9. The identification of proteoglycans and glycosaminoglycans in archaeological human bones and teeth.

    Directory of Open Access Journals (Sweden)

    Yvette M Coulson-Thomas

    Full Text Available Bone tissue is mineralized dense connective tissue consisting mainly of a mineral component (hydroxyapatite and an organic matrix comprised of collagens, non-collagenous proteins and proteoglycans (PGs. Extracellular matrix proteins and PGs bind tightly to hydroxyapatite which would protect these molecules from the destructive effects of temperature and chemical agents after death. DNA and proteins have been successfully extracted from archaeological skeletons from which valuable information has been obtained; however, to date neither PGs nor glycosaminoglycan (GAG chains have been studied in archaeological skeletons. PGs and GAGs play a major role in bone morphogenesis, homeostasis and degenerative bone disease. The ability to isolate and characterize PG and GAG content from archaeological skeletons would unveil valuable paleontological information. We therefore optimized methods for the extraction of both PGs and GAGs from archaeological human skeletons. PGs and GAGs were successfully extracted from both archaeological human bones and teeth, and characterized by their electrophoretic mobility in agarose gel, degradation by specific enzymes and HPLC. The GAG populations isolated were chondroitin sulfate (CS and hyaluronic acid (HA. In addition, a CSPG was detected. The localization of CS, HA, three small leucine rich PGs (biglycan, decorin and fibromodulin and glypican was analyzed in archaeological human bone slices. Staining patterns were different for juvenile and adult bones, whilst adolescent bones had a similar staining pattern to adult bones. The finding that significant quantities of PGs and GAGs persist in archaeological bones and teeth opens novel venues for the field of Paleontology.

  10. Second Harmonic Generation Imaging Analysis of Collagen Arrangement in Human Cornea.

    Science.gov (United States)

    Park, Choul Yong; Lee, Jimmy K; Chuck, Roy S

    2015-08-01

    To describe the horizontal arrangement of human corneal collagen bundles by using second harmonic generation (SHG) imaging. Human corneas were imaged with an inverted two photon excitation fluorescence microscope. The excitation laser (Ti:Sapphire) was tuned to 850 nm. Backscatter signals of SHG were collected through a 425/30-nm bandpass emission filter. Multiple, consecutive, and overlapping image stacks (z-stacks) were acquired to generate three dimensional data sets. ImageJ software was used to analyze the arrangement pattern (irregularity) of collagen bundles at each image plane. Collagen bundles in the corneal lamellae demonstrated a complex layout merging and splitting within a single lamellar plane. The patterns were significantly different in the superficial and limbal cornea when compared with deep and central regions. Collagen bundles were smaller in the superficial layer and larger in deep lamellae. By using SHG imaging, the horizontal arrangement of corneal collagen bundles was elucidated at different depths and focal regions of the human cornea.

  11. Effect of local hemostatics on bone induction in rats: a comparative study of bone wax, fibrin-collagen paste, and bioerodible polyorthoester with and without gentamicin

    DEFF Research Database (Denmark)

    Solheim, E; Pinholt, E M; Bang, G

    1992-01-01

    Local hemostatics for osseous tissue should preferably be absorbable and biocompatible and should not inhibit osteogenesis. The tissue response and effect on demineralized bone-induced heterotopic osteogenesis in the abdominal muscle of 120 male Wistar rats by different local hemostatics were...... evaluated by light microscopy and 85Sr uptake analyses. Non-absorbable bone wax of 88% beeswax and absorbable bovine fibrin-collagen paste both significantly inhibited osteoinduction, whereas a bioerodible polyorthoester drug delivery system with or without 4% gentamicin did not. Bone wax was not absorbed...

  12. Identification and characterization of the human type II collagen gene (COL2A1).

    NARCIS (Netherlands)

    K.S.E. Cheah (Kathryn); N.G. Stoker; J.R. Griffin; F.G. Grosveld (Frank); E. Solomon

    1985-01-01

    textabstractThe gene contained in the human cosmid clone CosHcol1, previously designated an alpha 1(I) collagen-like gene, has now been identified. CosHcol1 hybridizes strongly to a single 5.9-kilobase mRNA species present only in tissue in which type II collagen is expressed. DNA sequence analysis

  13. Higher iron bioavailability of a human-like collagen iron complex.

    Science.gov (United States)

    Zhu, Chenhui; Yang, Fan; Fan, Daidi; Wang, Ya; Yu, Yuanyuan

    2017-07-01

    Iron deficiency remains a public health problem around the world due to low iron intake and/or bioavailability. FeSO 4 , ferrous succinate, and ferrous glycinate chelate are rich in iron but have poor bioavailability. To solve the problem of iron deficiency, following previous research studies, a thiolated human-like collagen-ironcomplex supplement with a high iron content was prepared in an anaerobic workstation. In addition, cell viability tests were evaluated after conducting an MTT assay, and a quantitative analysis of the thiolated human-like collagen-iron digesta samples was performed using the SDS-PAGE method coupled with gel filtration chromatography. The iron bioavailability was assessed using Caco-2 cell monolayers and iron-deficiency anemia mice models. The results showed that (1) one mole of thiolated human-like collagen-iron possessed approximately 35.34 moles of iron; (2) thiolated human-like collagen-iron did not exhibit cytotoxity and (3) thiolated human-like collagen- iron digesta samples had higher bioavailability than other iron supplements, including FeSO 4 , ferrous succinate, ferrous glycine chelate and thiolated human-like collagen-Fe iron. Finally, the iron bioavailability was significantly enhanced by vitamin C. These results indicated that thiolated human-like collagen-iron is a promising iron supplement for use in the future.

  14. Reconstruction of irradiated bone segmental defects with a biomaterial associating MBCP+(R), microstructured collagen membrane and total bone marrow grafting: an experimental study in rabbits.

    Science.gov (United States)

    Jégoux, Franck; Goyenvalle, Eric; Cognet, Ronan; Malard, Olivier; Moreau, Francoise; Daculsi, Guy; Aguado, Eric

    2009-12-15

    The bone tissue engineering models used today are still a long way from any oncologic application as immediate postimplantation irradiation would decrease their osteoinductive potential. The aim of this study was to reconstruct a segmental critical size defect in a weight-bearing bone irradiated after implantation. Six white New Zealand rabbits were immediately implanted with a biomaterial associating resorbable collagen membrane EZ(R) filled and micro-macroporous biphasic calcium phosphate granules (MBCP+(R)). After a daily schedule of radiation delivery, and within 4 weeks, a total autologous bone marrow (BM) graft was injected percutaneously into the center of the implant. All the animals were sacrificed at 16 weeks. Successful osseous colonization was found to have bridged the entire length of the defects. Identical distribution of bone ingrowth and residual ceramics at the different levels of the implant suggests that the BM graft plays an osteoinductive role in the center of the defect. Periosteum-like formation was observed at the periphery, with the collagen membrane most likely playing a role. This model succeeded in bridging a large segmental defect in weight-bearing bone with immediate postimplantation fractionated radiation delivery. This has significant implications for the bone tissue engineering approach to patients with cancer-related bone defects.

  15. Ionic liquid as a potential solvent for preparation of collagen-alginate-hydroxyapatite beads as bone filler.

    Science.gov (United States)

    Iqbal, Bushra; Sarfaraz, Zenab; Muhammad, Nawshad; Ahmad, Pervaiz; Iqbal, Jibran; Khan, Zia Ul Haq; Gonfa, Girma; Iqbal, Farasat; Jamal, Arshad; Rahim, Abdur

    2018-07-01

    In this study, collagen/alginate/hydroxyapatite beads having different proportions were prepared as bone fillers for the restoration of osteological defects. Ionic liquid was used to dissolve the collagen and subsequently the solution was mixed with sodium alginate solution. Hydroxyapatite was added in different proportions, with the rationale to enhance mechanical as well as biological properties. The prepared solutions were given characteristic bead shapes by dropwise addition into calcium chloride solution. The prepared beads were characterized using FTIR, XRD, TGA and SEM analysis. Microhardness testing was used to evaluate the mechanical properties. The prepared beads were investigated for water adsorption behavior to ascertain its ability for body fluid uptake and adjusted accordingly to the bone cavity. Drug loading and subsequently the antibacterial activity was investigated for the prepared beads. The biocompatibility was assessed using the hemolysis testing and cell proliferation assay. The prepared collagen-alginate-HA beads, having biocompatibility and good mechanical properties, have showed an option of promising biologically active bone fillers for bone regeneration.

  16. Carnivore specific bone bioapatite and collagen carbon isotope fractionations: Case studies of modern and fossil grey wolf populations

    Science.gov (United States)

    Fox-Dobbs, K.; Wheatley, P. V.; Koch, P. L.

    2006-12-01

    Stable isotope analyses of modern and fossil biogenic tissues are routinely used to reconstruct present and past vertebrate foodwebs. Accurate isotopic dietary reconstructions require a consumer and tissue specific understanding of how isotopes are sorted, or fractionated, between trophic levels. In this project we address the need for carnivore specific isotope variables derived from populations that are ecologically well- characterized. Specifically, we investigate the trophic difference in carbon isotope values between mammalian carnivore (wolf) bone bioapatite and herbivore (prey) bone bioapatite. We also compare bone bioapatite and collagen carbon isotope values collected from the same individuals. We analyzed bone specimens from two modern North American grey wolf (Canis lupus) populations (Isle Royale National Park, Michigan and Yellowstone National Park, Wyoming), and the ungulate herbivores that are their primary prey (moose and elk, respectively). Because the diets of both wolf populations are essentially restricted to a single prey species, there were no confounding effects due to carnivore diet variability. We measured a trophic difference of approximately -1.3 permil between carnivore (lower value) and herbivore (higher value) bone bioapatite carbon isotope values, and an average inter-tissue difference of 5.1 permil between carnivore bone collagen (lower value) and bioapatite (higher value) carbon isotope values. Both of these isotopic differences differ from previous estimates derived from a suite of African carnivores; our carnivore-herbivore bone bioapatite carbon isotope spacing is smaller (-1.3 vs. -4.0 permil), and our carnivore collagen-bioapatite carbon difference is larger (5.1 vs. 3.0 permil). These discrepancies likely result from comparing values measured from a single hypercarnivore (wolf) to average values calculated from several carnivore species, some of which are insectivorous or partly omnivorous. The trophic and inter

  17. Three-dimensional poly (ε-caprolactone)/hydroxyapatite/collagen scaffolds incorporating bone marrow mesenchymal stem cells for the repair of bone defects

    International Nuclear Information System (INIS)

    Qi, Xin; Huang, Yinjun; Zhang, Jieyuan; Cao, Jiaqing; Jin, Xiangyun; Huang, Jinghuan; Li, Xiaolin; Wang, Ting; Han, Dan

    2016-01-01

    We previously demonstrated that three-dimensional (3D) hydroxyapatite (HAP)-collagen (COL)-coated poly(ε-caprolactone) (PCL) scaffolds (HAP-COL-PCL) possess appropriate nano-structures, surface roughness, and nutrients, providing a favorable environment for osteogenesis. However, the effect of using 3D HAP-COL-PCL scaffolds incorporating BMSCs for the repair of bone defects in rats has been not evaluated. 3D PCL scaffolds coated with HAP, collagen or HAP/COL and incorporating BMSCs were implanted into calvarial defects. At 12 weeks after surgery, the rats were sacrificed and crania were harvested to assess the bone defect repair using microcomputed tomography (micro-CT), histology, immunohistochemistry and sequential fluorescent labeling analysis. 3D micro-CT reconstructed images and quantitative analysis showed that HAP-COL-PCL groups possessed better bone-forming capacity than HAP-PCL groups or COL-PCL groups. Fluorescent labeling analysis revealed the percentage of tetracycline labeling, alizarin red labeling, and calcein labeling in HAP-COL-PCL groups were all greater than in the other two groups (P  <  0.05), and the result was confirmed by immunohistochemical staining and histological analysis of bone regeneration. This study demonstrates that 3D HAP-COL-PCL scaffolds incorporating BMSCs markedly enhance bone regeneration of bone defects in rats. (paper)

  18. Human-like collagen/nano-hydroxyapatite scaffolds for the culture of chondrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Jia, Liping; Duan, Zhiguang [Shaanxi Key Laboratory of Degradable Biomedical Materials, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Shaanxi R and D Center of Biomaterials and Fermentation Engineering, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Fan, Daidi, E-mail: fandaidi@nwu.edu.cn [Shaanxi Key Laboratory of Degradable Biomedical Materials, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Shaanxi R and D Center of Biomaterials and Fermentation Engineering, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Mi, Yu; Hui, Junfeng [Shaanxi Key Laboratory of Degradable Biomedical Materials, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Shaanxi R and D Center of Biomaterials and Fermentation Engineering, Northwest University, 229 Taibai North Road, Xi' an, Shaanxi 710069 (China); Chang, Le [School of Chemical Engineering, Northwest University, Xi' an, Shaanxi 710069 (China)

    2013-03-01

    Three dimensional (3D) biodegradable porous scaffolds play a key role in cartilage tissue repair. Freeze-drying and cross-linking techniques were used to fabricate a 3D composite scaffold that combined the excellent biological characteristics of human-like collagen (HLC) and the outstanding mechanical properties of nano-hydroxyapatite (nHA). The scaffolds were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and compression tests, using Relive Registered-Sign Artificial Bone (RAB) scaffolds as a control. HLC/nHA scaffolds displayed homogeneous interconnected macroporous structure and could withstand a compression stress of 2.67 {+-} 0.37 MPa, which was higher than that of the control group. Rabbit chondrocytes were seeded on the composite porous scaffolds and cultured for 21 days. Cell/scaffold constructs were examined using SEM, histological procedures, and biochemical assays for cell proliferation and the production of glycosaminoglycans (GAGs). The results indicated that HLC/nHA porous scaffolds were capable of encouraging cell adhesion, homogeneous distribution and abundant GAG synthesis, and maintaining natural chondrocyte morphology compared to RAB scaffolds. In conclusion, the presented data warrants the further exploration of HLC/nHA scaffolds as a potential biomimetic platform for chondrocytes in cartilage tissue engineering. - Highlights: Black-Right-Pointing-Pointer Human-like collagen was first used to prepare cartilage tissue engineering scaffold. Black-Right-Pointing-Pointer Genipin, a natural biological cross-linking agent, was introduced to treat scaffold. Black-Right-Pointing-Pointer We chose market product as a control.

  19. Toward guided tissue and bone regeneration: morphology, attachment, proliferation, and migration of cells cultured on collagen barrier membranes. A systematic review.

    NARCIS (Netherlands)

    Behring, J.; Junker, R.; Walboomers, X.F.; Chessnut, B.; Jansen, J.A.

    2008-01-01

    Collagen barrier membranes are frequently used in both guided tissue regeneration (GTR) and guided bone regeneration (GBR). Collagen used for these devices is available from different species and is often processed to alter the properties of the final product. This is necessary because unprocessed

  20. [Development of a novel absorbable nanofiber chitosan-collagen membrane by electrospinning and the preliminary study on guided bone regeneration].

    Science.gov (United States)

    Gao, B; Li, X J; Lin, M; Li, Y Y; Dong, Y

    2018-02-09

    Objective: To evaluate the application effect of nanofiber chitosan-collagen membrane (NCM) on guided bone regeneration (GBR). Methods: The mixture of collagen, chitosan, polyethylene oxide was used to make up the NCM by electrospinning, then the NCM was crosslinked by glutaraldehyde vapor. The physical property of the NCM was measured by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). MC3T3-E1 osteoblasts were cultured on NCM to characterize the biocompatibility. The effectiveness of four groups [contrast group, Bio-gide membrane (BGM), compressed chitosan-collagen menbrane (CCM), NCM/CCM] on bone regeneration were evaluated in critical-sized defects (diameter = 5 mm) in SD rats. Results: When the mixed solution consists of 4.0% collagen, 1.0% chitosan and 3.5% polyethylene oxide, the NCM could be validly fabricated by electrospinning. After cross-linking by glutaraldehyde vapor, the tensile strength and the stability of NCM in damp was enhanced. No cytotoxicity of the NCM was detected on MC3T3-E1 osteoblasts. In vivo study showed that the new bone regeneration ratio of NCM/CCM group was [(43.10±1.49)%], and this was similar to that of the group of BGM [(41.36±2.60)%] ( P> 0.05), but higher than that of the CCM group [(33.10±1.41)%] and the contrast group [(7.22±2.46)%] ( P< 0.05). Conclusions: The NCM can promote new bone regeneration effectively in GBR procedure.

  1. Paleoecological and climatic implications of stable isotope results from late Pleistocene bone collagen, Ziegeleigrube Coenen, Germany

    Science.gov (United States)

    Wißing, Christoph; Matzerath, Simon; Turner, Elaine; Bocherens, Hervé

    2015-07-01

    Climatic and ecological conditions during Marine Oxygen Isotope Stage (MIS) 3 are complex and the impact of cold spells on the ecosystems in Central Europe still needs to be investigated thoroughly. Ziegeleigrube Coenen (ZC) is a late Pleistocene MIS 3 locality in the Lower Rhine Embayment of Germany, radiocarbon-dated to > 34 14C ka BP. The site yielded a broad spectrum of mammal species. We investigated the carbon (δ13C), nitrogen (δ15N) and sulfur (δ34S) isotope signatures of bone collagen, since these are valuable tools in characterizing ecological niches, environmental conditions and aspects of climate and mobility. By comparison with pre- and post-Last Glacial Maximum (LGM) sites in Central Europe we show that ZC belongs in a cold event of MIS 3 and was climatically more similar to post-LGM sites than to pre-LGM sites. However, the trophic structure resembled that of typical pre-LGM sites in Belgium. This cold event in MIS 3 changed the bottom of the foodweb, but do not seem to have had a direct impact on the occurrence of the mammalian species and their ecological distribution. Apparently the (mega-) faunal community could adapt also to harsher environmental conditions during MIS 3.

  2. Insulin-like growth factor I enhances collagen synthesis in engineered human tendon tissue

    DEFF Research Database (Denmark)

    Herchenhan, Andreas; Bayer, Monika L.; Eliasson, Pernilla

    2015-01-01

    OBJECTIVE: Isolated human tendon cells form 3D tendon constructs that demonstrate collagen fibrillogenesis and feature structural similarities to tendon when cultured under tensile load. The exact role of circulating growth factors for collagen formation in tendon is sparsely examined. We...... investigated the influence of insulin-like growth factor I (IGF-I) on tendon construct formation in 3D cell culture. DESIGN: Tendon constructs were grown in 0.5 or 10% FBS with or without IGF-I (250 mg/ml) supplementation. Collagen content (fluorometric), mRNA levels (PCR) and fibril diameter (transmission...... electron microscopy) were determined at 7, 10, 14, 21 and 28 days. RESULTS: IGF-I revealed a stimulating effect on fibril diameter (up to day 21), mRNA for collagen (to day 28), tenomodulin (to day 28) and scleraxis (at days 10 and 14), and on overall collagen content. 10% FBS diminished the development...

  3. 3-D ultrastructure and collagen composition of healthy and overloaded human tendon

    DEFF Research Database (Denmark)

    Pingel, Jessica; Lu, Yinhui; Starborg, Tobias

    2014-01-01

    with regards to changes in the content of collagen type I and III (the major collagens in tendon), and changes in tendon fibroblast (tenocyte) shape and organization of the extracellular matrix (ECM). To gain new insights, we took biopsies from the tendinopathic region and flanking healthy region of Achilles...... block face-scanning electron microscopy were made on two individuals. In the tendinopathic regions, compared with the flanking healthy tissue, we observed: (i) an increase in the ratio of collagen III : I proteins; (ii) buckling of the collagen fascicles in the ECM; (iii) buckling of tenocytes...... and their nuclei; and (iv) an increase in the ratio of small-diameter : large-diameter collagen fibrils. In summary, load-induced non-rupture tendinopathy in humans is associated with localized biochemical changes, a shift from large- to small-diameter fibrils, buckling of the tendon ECM, and buckling of the cells...

  4. Modified silk fibroin scaffolds with collagen/decellularized pulp for bone tissue engineering in cleft palate: Morphological structures and biofunctionalities

    International Nuclear Information System (INIS)

    Sangkert, Supaporn; Meesane, Jirut; Kamonmattayakul, Suttatip; Chai, Wen Lin

    2016-01-01

    Cleft palate is a congenital malformation that generates a maxillofacial bone defect around the mouth area. The creation of performance scaffolds for bone tissue engineering in cleft palate is an issue that was proposed in this research. Because of its good biocompatibility, high stability, and non-toxicity, silk fibroin was selected as the scaffold of choice in this research. Silk fibroin scaffolds were prepared by freeze-drying before immerging in a solution of collagen, decellularized pulp, and collagen/decellularized pulp. Then, the immersed scaffolds were freeze-dried. Structural organization in solution was observed by Atomic Force Microscope (AFM). The molecular organization of the solutions and crystal structure of the scaffolds were characterized by Fourier transform infrared (FT-IR) and X-ray diffraction (XRD), respectively. The weight increase of the modified scaffolds and the pore size were determined. The morphology was observed by a scanning electron microscope (SEM). Mechanical properties were tested. Biofunctionalities were considered by seeding osteoblasts in silk fibroin scaffolds before analysis of the cell proliferation, viability, total protein assay, and histological analysis. The results demonstrated that dendrite structure of the fibrils occurred in those solutions. Molecular organization of the components in solution arranged themselves into an irregular structure. The fibrils were deposited in the pores of the modified silk fibroin scaffolds. The modified scaffolds showed a beta-sheet structure. The morphological structure affected the mechanical properties of the silk fibroin scaffolds with and without modification. Following assessment of the biofunctionalities, the modified silk fibroin scaffolds could induce cell proliferation, viability, and total protein particularly in modified silk fibroin with collagen/decellularized pulp. Furthermore, the histological analysis indicated that the cells could adhere in modified silk fibroin

  5. Early adhesive behavior of bone-marrow-derived mesenchymal stem cells on collagen electrospun fibers

    Energy Technology Data Exchange (ETDEWEB)

    Chan, Casey K; Liao, Susan; Lareu, Ricky R; Raghunath, Michael [Division of Bioengineering, National University of Singapore, 7 Engineering Drive 1, Singapore 117574 (Singapore); Li, Bojun; Ramakrishna, S [Nanoscience and Nanotechnology Initiative, National University of Singapore, 2 Engineering Drive 3, Singapore 117576 (Singapore); Larrick, James W, E-mail: doschanc@nus.edu.s [Panorama Research Institute, 2462 Wyandotte Street, Mountain View, CA 94043 (United States)

    2009-06-15

    A bioabsorbable nanofibrous scaffold was developed for early adhesion of mesenchymal stem cells (MSCs). Collagen nanofibers with diameters of 430 +- 170 nm were fabricated by electrospinning. Over 45% of the MSC population adhered to this collagen nanofiber after 30 min at room temperature. Remarkably, collagen-coated P(LLA-CL) electrospun nanofibers were almost as efficient as collagen nanofibers whereas collagen cast film did not enhance early capture when it was applied on cover slips. The adhesive efficiency could be further increased to over 20% at 20 min and over 55% at 30 min when collagen nanofibers were grafted with monoclonal antibodies recognizing CD29 or CD49a. These data demonstrate that the early adhesive behavior is highly dependent on both the surface texture and the surface chemistry of the substrate. These findings have potential applications for early capture of MSCs in an ex vivo setting under time constraints such as in a surgical setting.

  6. Early adhesive behavior of bone-marrow-derived mesenchymal stem cells on collagen electrospun fibers

    International Nuclear Information System (INIS)

    Chan, Casey K; Liao, Susan; Lareu, Ricky R; Raghunath, Michael; Li, Bojun; Ramakrishna, S; Larrick, James W

    2009-01-01

    A bioabsorbable nanofibrous scaffold was developed for early adhesion of mesenchymal stem cells (MSCs). Collagen nanofibers with diameters of 430 ± 170 nm were fabricated by electrospinning. Over 45% of the MSC population adhered to this collagen nanofiber after 30 min at room temperature. Remarkably, collagen-coated P(LLA-CL) electrospun nanofibers were almost as efficient as collagen nanofibers whereas collagen cast film did not enhance early capture when it was applied on cover slips. The adhesive efficiency could be further increased to over 20% at 20 min and over 55% at 30 min when collagen nanofibers were grafted with monoclonal antibodies recognizing CD29 or CD49a. These data demonstrate that the early adhesive behavior is highly dependent on both the surface texture and the surface chemistry of the substrate. These findings have potential applications for early capture of MSCs in an ex vivo setting under time constraints such as in a surgical setting.

  7. Differentiation of human mesenchymal stromal cells cultured on collagen sponges for cartilage repair.

    Science.gov (United States)

    Sanjurjo-Rodríguez, Clara; Martínez-Sánchez, Adela Helvia; Hermida-Gómez, Tamara; Fuentes-Boquete, Isaac; Díaz-Prado, Silvia; Blanco, Francisco J

    2016-11-01

    The aim of this study was to evaluate proliferation and chondrogenic differentiation of human bone-marrow mesenchymal stromal cells (hBMSCs) cultured on collagen biomaterials. hBMSCs were seeded on five different collagen (Col) sponges: C1C2 (types I and II Col), C1C2HS (types I and II Col plus heparan sulphate (HS)), C1C2CHS (types I and II Col plus chondroitin sulphate (CHS)), C1-OLH3 (type I Col plus low molecular weight heparin) and C1CHS (type I Col plus CHS). The resulting constructs were analyzed by histological and immunohistochemical staining, molecular biology and electron microscopy. Col released into culture media was measured by a dye-binding method Results: hBMSCs on biomaterials C1C2, C1C2HS and C1C2CHS had more capacity to attach, proliferate and synthesize Col II and proteoglycans in the extracellular matrix (ECM) than on C1-OLH3 and C1CHS. The presence of aggrecan was detected only at the gene level. Total Col liberated by the cells in the supernatants in all scaffold cultures was detected. The level of Col I in the ECM was lower in C1-OLH3 and that of Col II was highest in C1C2 and C1C2HS. Electron microscopy showed differently shaped cells, from rounded to flattened, in all constructs. Col fibers in bundles were observed in C1C2CHS by transmission electron microscopy. The results show that Col I and Col II (C1C2, C1C2HS and C1C2CHS) biomaterials allowed cell proliferation and chondrogenic-like differentiation of hBMSCs at an early stage. Constructs cultured on C1C2HS and C1C2CHS showed better cartilage-like phenotype than the other ones.

  8. Pleiotrophin commits human bone marrow mesenchymal stromal cells towards hypertrophy during chondrogenesis.

    Science.gov (United States)

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

    2014-01-01

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

  9. Pleiotrophin commits human bone marrow mesenchymal stromal cells towards hypertrophy during chondrogenesis.

    Directory of Open Access Journals (Sweden)

    Thibault Bouderlique

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

  10. Oral histories: a simple method of assigning chronological age to isotopic values from human dentine collagen.

    Science.gov (United States)

    Beaumont, Julia; Montgomery, Janet

    2015-01-01

    Stable isotope ratios of carbon (δ(13)C) and nitrogen (δ(15)N) in bone and dentine collagen have been used for over 30 years to estimate palaeodiet, subsistence strategy, breastfeeding duration and migration within burial populations. Recent developments in dentine microsampling allow improved temporal resolution for dietary patterns. A simple method is proposed which could be applied to human teeth to estimate chronological age represented by dentine microsamples in the direction of tooth growth, allowing comparison of dietary patterns between individuals and populations. The method is tested using profiles from permanent and deciduous teeth of two individuals. Using a diagrammatic representation of dentine development by approximate age for each human tooth (based on the Queen Mary University of London Atlas), this study estimated the age represented by each dentine section. Two case studies are shown: comparison of M1 and M2 from a 19th century individual from London, England, and identification of an unknown tooth from an Iron Age female adult from Scotland. The isotopic profiles demonstrate that variations in consecutively-forming teeth can be aligned using this method to extend the dietary history of an individual or identify an unknown tooth by matching the profiles.

  11. Tensile properties of human collagen fibrils and fascicles are insensitive to environmental salts

    DEFF Research Database (Denmark)

    Svensson, René B; Hassenkam, Tue; Grant, Colin A

    2010-01-01

    loading direction of tendon is along its longitudinal axis. Thus, in this study, we focus on the tensile mechanical properties of two hierarchical levels from human patellar tendon, namely: individual collagen fibrils and fascicles. Investigations on collagen fibrils and fascicles were made at pH 7...... was observed at the highest phosphate-buffered saline concentration for both the fibrils and fascicles, indicating a stabilizing effect of ionic screening, but changes were much less than reported for radial compression. Due to the small magnitude of the effects, the tensile mechanical properties of collagen...

  12. Repair of Avascular Meniscus Tears with Electrospun Collagen Scaffolds Seeded with Human Cells.

    Science.gov (United States)

    Baek, Jihye; Sovani, Sujata; Glembotski, Nicholas E; Du, Jiang; Jin, Sungho; Grogan, Shawn P; D'Lima, Darryl D

    2016-03-01

    The self-healing capacity of an injured meniscus is limited to the vascularized regions and is especially challenging in the inner avascular regions. As such, we investigated the use of human meniscus cell-seeded electrospun (ES) collagen type I scaffolds to produce meniscal tissue and explored whether these cell-seeded scaffolds can be implanted to repair defects created in meniscal avascular tissue explants. Human meniscal cells (derived from vascular and avascular meniscal tissue) were seeded on ES scaffolds and cultured. Constructs were evaluated for cell viability, gene expression, and mechanical properties. To determine potential for repair of meniscal defects, human meniscus avascular cells were seeded and cultured on aligned ES collagen scaffolds for 4 weeks before implantation. Surgical defects resembling "longitudinal tears" were created in the avascular zone of bovine meniscus and implanted with cell-seeded collagen scaffolds and cultured for 3 weeks. Tissue regeneration and integration were evaluated by histology, immunohistochemistry, mechanical testing, and magentic resonance imaging. Ex vivo implantation with cell-seeded collagen scaffolds resulted in neotissue that was significantly better integrated with the native tissue than acellular collagen scaffolds or untreated defects. Human meniscal cell-seeded ES collagen scaffolds may therefore be useful in facilitating meniscal repair of avascular meniscus tears.

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

    Science.gov (United States)

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

    2017-07-28

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

  14. The effect of acute exercise on collagen turnover in human tendons

    DEFF Research Database (Denmark)

    Mørch, Lina Steinrud; Pingel, Jessica; Boesen, Mikael

    2013-01-01

    Mechanical loading of human tendon stimulates collagen synthesis, but the relationship between acute loading responses and training status of the tendon is not clear. We tested the effect of prolonged load deprivation on the acute loading-induced collagen turnover in human tendons, by applying...... the contra-lateral leg was used habitually. Following the procedure both Achilles tendons and calf muscles were loaded with the same absolute load during a 1-h treadmill run. Tissue collagen turnover was measured by microdialysis performed post-immobilization but pre-exercise around both Achilles tendons...... and compared to values obtained by 72-h post-exercise. Power Doppler was used to monitor alterations in intratendinous blood flow velocity of the Achilles tendon and MRI used to quantitate changes in tendon cross-section area. Acute loading resulted in an increased collagen synthesis 72 h after the run in both...

  15. Parametric imaging of collagen structural changes in human osteoarthritic cartilage using optical polarization tractography

    Science.gov (United States)

    Ravanfar, Mohammadreza; Pfeiffer, Ferris M.; Bozynski, Chantelle C.; Wang, Yuanbo; Yao, Gang

    2017-12-01

    Collagen degeneration is an important pathological feature of osteoarthritis. The purpose of this study is to investigate whether the polarization-sensitive optical coherence tomography (PSOCT)-based optical polarization tractography (OPT) can be useful in imaging collagen structural changes in human osteoarthritic cartilage samples. OPT eliminated the banding artifacts in conventional PSOCT by calculating the depth-resolved local birefringence and fiber orientation. A close comparison between OPT and PSOCT showed that OPT provided improved visualization and characterization of the zonal structure in human cartilage. Experimental results obtained in this study also underlined the importance of knowing the collagen fiber orientation in conventional polarized light microscopy assessment. In addition, parametric OPT imaging was achieved by quantifying the surface roughness, birefringence, and fiber dispersion in the superficial zone of the cartilage. These quantitative parametric images provided complementary information on the structural changes in cartilage, which can be useful for a comprehensive evaluation of collagen damage in osteoarthritic cartilage.

  16. Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo [NovaCell Technology Inc., Pohang, Kyungbuk 790-784 (Korea, Republic of); Kim, So Young [Department of Dermatology, Chung-Ang University College of Medicine, Seoul 156-756 (Korea, Republic of); Department of Convergence Medicine and Pharmaceutical Biosciences, Graduate School, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Jang, Hwan-Hee [Functional Food and Nutrition Division, Department of Agrofood Resources, Rural Development Administration, Suwon 441-853 (Korea, Republic of); Ryu, Sung Ho [Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology (POSTECH), Pohang, Kyungbuk 790-784 (Korea, Republic of); Kim, Beom Joon [Department of Dermatology, Chung-Ang University College of Medicine, Seoul 156-756 (Korea, Republic of); Department of Convergence Medicine and Pharmaceutical Biosciences, Graduate School, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Lee, Taehoon G., E-mail: taehoon@novacelltech.com [NovaCell Technology Inc., Pohang, Kyungbuk 790-784 (Korea, Republic of)

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer We identify a function of the YIGSR peptide to enhance collagen synthesis in Hs27. Black-Right-Pointing-Pointer YIGSR peptide enhanced collagen type 1 synthesis both of gene and protein levels. Black-Right-Pointing-Pointer There were no changes in cell proliferation and MMP-1 level in YIGSR treatment. Black-Right-Pointing-Pointer The YIGSR effect on collagen synthesis mediated activation of FAK, pyk2 and ERK. Black-Right-Pointing-Pointer The YIGSR-induced FAK and ERK activation was modulated by FAK and MEK inhibitors. -- Abstract: The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929-933 sequence of the {beta}1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67 kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate

  17. Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts

    International Nuclear Information System (INIS)

    Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo; Kim, So Young; Jang, Hwan-Hee; Ryu, Sung Ho; Kim, Beom Joon; Lee, Taehoon G.

    2012-01-01

    Highlights: ► We identify a function of the YIGSR peptide to enhance collagen synthesis in Hs27. ► YIGSR peptide enhanced collagen type 1 synthesis both of gene and protein levels. ► There were no changes in cell proliferation and MMP-1 level in YIGSR treatment. ► The YIGSR effect on collagen synthesis mediated activation of FAK, pyk2 and ERK. ► The YIGSR-induced FAK and ERK activation was modulated by FAK and MEK inhibitors. -- Abstract: The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929–933 sequence of the β1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67 kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate peptide for the treatment of skin aging and wrinkles.

  18. Combined Effect of a Microporous Layer and Type I Collagen Coating on a Biphasic Calcium Phosphate Scaffold for Bone Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Mun-Hwan Lee

    2015-03-01

    Full Text Available In this study, type I collagen was coated onto unmodified and modified microporous biphasic calcium phosphate (BCP scaffolds. Surface characterization using a scanning electron microscope (SEM and a surface goniometer confirmed the modification of the BCP coating. The quantity of the collagen coating was investigated using Sirius Red staining, and quantitative assessment of the collagen coating showed no significant differences between the two groups. MG63 cells were used to evaluate cell proliferation and ALP activity on the modified BCP scaffolds. The modified microporous surfaces showed low contact angles and large surface areas, which enhanced cell spreading and proliferation. Coating of the BCP scaffolds with type I collagen led to enhanced cell-material interactions and improved MG63 functions, such as spreading, proliferation, and differentiation. The micropore/collagen-coated scaffold showed the highest rate of cell response. These results indicate that a combination of micropores and collagen enhances cellular function on bioengineered bone allograft tissue.

  19. Effects of Recombinant Human Bone Morphogenetic Protein-2 on Vertical Bone Augmentation in a Canine Model.

    Science.gov (United States)

    Hsu, Yung-Ting; Al-Hezaimi, Khalid; Galindo-Moreno, Pablo; O'Valle, Francisco; Al-Rasheed, Abdulaziz; Wang, Hom-Lay

    2017-09-01

    Vertical bone augmentation (VBA) remains unpredictable and challenging for most clinicians. This study aims to compare hard tissue outcomes of VBA, with and without recombinant human bone morphogenetic protein (rhBMP)-2, under space-making titanium mesh in a canine model. Eleven male beagle dogs were used in the study. Experimental ridge defects were created to form atrophic ridges. VBA was performed via guided bone regeneration using titanium mesh and allografts. In experimental hemimandibles, rhBMP-2/absorbable collagen sponge was well mixed with allografts prior to procedures, whereas a control buffer was applied within controls. Dogs were euthanized after a 4-month healing period. Clinical and radiographic examinations were performed to assess ridge dimensional changes. In addition, specimens were used for microcomputed tomography (micro-CT) assessment and histologic analysis. Membrane exposure was found on five of 11 (45.5%) rhBMP-2-treated sites, whereas it was found on nine of 11 (81.8%) non-rhBMP-2-treated sites. Within 4 months of healing, rhBMP-2-treated sites showed better radiographic bone density, greater defect fill, and significantly more bone gain in ridge height (P 0.05). Under light microscope, predominant lamellar patterns were found in the specimen obtained from rhBMP-2 sites. With inherent limitations of the canine model and the concern of such a demanding surgical technique, current findings suggest that the presence of rhBMP-2 in a composite graft allows an increase of vertical gain, with formation of ectopic bone over the titanium mesh in comparison with non-rhBMP-2 sites.

  20. Gene Expression Changes in Femoral Head Necrosis of Human Bone Tissue

    Directory of Open Access Journals (Sweden)

    Bernadett Balla

    2011-01-01

    Full Text Available Osteonecrosis of the femoral head (ONFH is the result of an interruption of the local circulation and the injury of vascular supply of bone. Multiple factors have been implicated in the development of the disease. However the mechanism of ischemia and necrosis in non-traumatic ONFH is not clear. The aim of our investigation was to identify genes that are differently expressed in ONFH vs. non-ONFH human bone and to describe the relationships between these genes using multivariate data analysis. Six bone tissue samples from ONFH male patients and 8 bone tissue samples from non-ONFH men were examined. The expression differences of selected 117 genes were analyzed by TaqMan probe-based quantitative real-time RT-PCR system. The significance test indicated marked differences in the expression of nine genes between ONFH and non-ONFH individuals. These altered genes code for collagen molecules, an extracellular matrix digesting metalloproteinase, a transcription factor, an adhesion molecule, and a growth factor. Canonical variates analysis demonstrated that ONFH and non-ONFH bone tissues can be distinguished by the multiple expression profile analysis of numerous genes controlled via canonical TGFB pathway as well as genes coding for extracellular matrix composing collagen type molecules. The markedly altered gene expression profile observed in the ONFH of human bone tissue may provide further insight into the pathogenetic process of osteonecrotic degeneration of bone.

  1. Next generation bone tissue engineering: non-viral miR-133a inhibition using collagen-nanohydroxyapatite scaffolds rapidly enhances osteogenesis

    Science.gov (United States)

    Mencía Castaño, Irene; Curtin, Caroline M.; Duffy, Garry P.; O'Brien, Fergal J.

    2016-06-01

    Bone grafts are the second most transplanted materials worldwide at a global cost to healthcare systems valued over $30 billion every year. The influence of microRNAs in the regenerative capacity of stem cells offers vast therapeutic potential towards bone grafting; however their efficient delivery to the target site remains a major challenge. This study describes how the functionalisation of porous collagen-nanohydroxyapatite (nHA) scaffolds with miR-133a inhibiting complexes, delivered using non-viral nHA particles, enhanced human mesenchymal stem cell-mediated osteogenesis through the novel focus on a key activator of osteogenesis, Runx2. This study showed enhanced Runx2 and osteocalcin expression, as well as increased alkaline phosphatase activity and calcium deposition, thus demonstrating a further enhanced therapeutic potential of a biomaterial previously optimised for bone repair applications. The promising features of this platform offer potential for a myriad of applications beyond bone repair and tissue engineering, thus presenting a new paradigm for microRNA-based therapeutics.

  2. Extraction Socket Preservation Using Porcine-Derived Collagen Membrane Alone or Associated with Porcine-Derived Bone. Clinical Results of Randomized Controlled Study

    Directory of Open Access Journals (Sweden)

    Renzo Guarnieri

    2017-03-01

    Full Text Available Objectives: The aim of present randomized controlled clinical trial was to clinically evaluate hard tissue changes after extraction socket preservation procedures compared to natural spontaneous healing. Material and Methods: Thirty patients were enrolled in the present study and underwent single-tooth extraction in the premolar/molar areas. Ten sites were grafted with porcine-derived bone covered by collagen membrane, 10 covered by porcine-derived collagen membrane alone, and 10 underwent natural spontaneous healing. Vertical and horizontal bone changes after 3-month were evaluated at implant placement. Results: The vertical and horizontal bone changes at the extraction sockets treated with collagen membrane alone (vertical: -0.55 [SD 0.11] mm, and horizontal: -1.21 [SD 0.69] mm and collagen membrane plus porcine-derived bone (vertical: -0.37 [SD 0.7] mm, and horizontal: -0.91 [SD 0.53] mm were found significantly lower (P < 0.001, when compared to non-grafted sockets (vertical: -2.09 [SD 0.19] mm, and horizontal: -3.96 [SD 0.87] mm. In type 1 extraction sockets, in premolar sites, and in presence of vestibular bone thicknesses ≥ 1.5 mm, the use of collagen membrane alone revealed similar outcomes to those with additional graft material. Conclusions: At the re-entry surgery, extraction sockets grafted with porcine-derived bone and covered by collagen membrane, and extraction sockets covered by porcine-derived collagen membrane alone, showed significantly lower vertical and horizontal bone changes, compared to extraction sockets sites underwent natural spontaneous healing. However, a complete prevention of remodelling is not achievable, irrespective of the technique used.

  3. Maxillary anterior ridge augmentation with recombinant human bone morphogenetic protein 2.

    Science.gov (United States)

    Edmunds, Ryan K; Mealey, Brian L; Mills, Michael P; Thoma, Daniel S; Schoolfield, John; Cochran, David L; Mellonig, Jim

    2014-01-01

    No human studies exist on the use of recombinant human bone morphogenetic protein 2 (rhBMP-2) on an absorbable collagen sponge (ACS) as a sole graft material for lateral ridge augmentation in large ridge defect sites. This series evaluates the treatment outcome of maxillary anterior lateral ridge augmentation with rhBMP-2/ACS. Twenty patients were treated with rhBMP-2/ACS and fixation screws for space maintenance. Cone beam volumetric tomography measurements were used to determine gain in ridge width, and a bone core biopsy was obtained. The mean horizontal ridge gain was 1.2 mm across sites, and every site gained width.

  4. Bony defect repair in rabbit using hybrid rapid prototyping polylactic co glycolic acid/β tricalciumphosphate collagen I/apatite scaffold and bone marrow mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Long Pang

    2013-01-01

    Full Text Available Background: In bone tissue engineering, extracellular matrix exerts critical influence on cellular interaction with porous biomaterial and the apatite playing an important role in the bonding process of biomaterial to bone tissue. The aim of this study was to observe the therapeutic effects of hybrid rapid prototyping (RP scaffolds comprising polylactic-co-glycolic acid (PLGA, β-tricalciumphosphate (β-TCP, collagen I and apatite (PLGA/β-TCP-collagen I/apatite on segmental bone defects in conjunction with combination with bone marrow mesenchymal stem cells (BMSCs. Materials and Methods: BMSCs were seeded into the hybrid RP scaffolds to repair 15 mm defect in the radius of rabbits. Radiograph, microcomputed tomography and histology were used to evaluate new bone formation. Results: Radiographic analysis done from 12 to 36 weeks postoperative period demonstrated that new bone formed at the radial defect site and continues to increase until the medullary cavity is recanalized and remodelling is complete. The bone defect remained unconnected in the original RP scaffolds (PLGA/β-TCP during the whole study. Histological observations conformed to the radiographic images. In hybrid RP scaffold group, woven bone united the radial defect at 12 weeks and consecutively remodeled into lamellar bone 24 weeks postoperation and finally matured into cortical bone with normal marrow cavity after another 12 weeks. No bone formation but connective tissue has been detected in RP scaffold at the same time. Conclusion: Collagen I/apatite sponge composite coating could improve new bone formation in vivo. The hybrid RP scaffold of PLGA/β-TCP skeleton with collagen I/apatite sponge composite coating is a promising candidate for bone tissue engineering.

  5. Lipo-PGE1 suppresses collagen production in human dermal fibroblasts via the ERK/Ets-1 signaling pathway.

    Directory of Open Access Journals (Sweden)

    Yoolhee Yang

    Full Text Available Dysregulation of collagen production contributes to various pathological processes, including tissue fibrosis as well as impaired wound healing. Lipo-prostaglandin E1 (Lipo-PGE1, a lipid microsphere-incorporated prostaglandin E1, is used as a vasodilator for the treatment of peripheral vascular diseases. Lipo-PGE1 was recently shown to enhance human dermal fibroblast (HDF migration and in vivo wound healing. No published study has characterized the role of Lipo-PGE1 in collagen regulation in HDFs. Here, we investigated the cellular signaling mechanism by which Lipo-PGE1 regulates collagen in HDFs. Collagen production was evaluated by the Sircol collagen assay, Western blot analysis of type I collagen and real time PCR. Unexpectedly, Lipo-PGE1 decreased mRNA expression of collagen 1A1, 1A2, and 3A1. Lipo-PGE1 markedly inhibited type I collagen and total soluble collagen production. In addition, Lipo-PGE1 inhibited transforming growth factor-β-induced collagen expression via Smad2 phosphorylation. To further investigate whether extracellular signal-regulated kinase (ERK/Ets-1 signaling, a crucial pathway in collagen regulation, is involved in Lipo-PGE1-inhibited collagen production, cells were pretreated with an ERK-specific inhibitor, PD98059, prior to the addition of Lipo-PGE1. Lipo-PGE1-inhibited collagen mRNA expression and total soluble collagen production were recovered by pretreatment with PD98059. Moreover, Lipo-PGE1 directly induced the phosphorylation of ERK. Furthermore, silencing of Ets-1 recovered Lipo-PGE1-inhibited collagen production and PD98059 blocked Lipo-PGE1-enhanced Ets-1 expression. The present study reveals an important role for Lipo-PGE1 as a negative regulator of collagen gene expression and production via ERK/Ets-1 signaling. These results suggest that Lipo-PGE1 could potentially be a therapeutic target in diseases with deregulated collagen turnover.

  6. Preparation of a biomimetic composite scaffold from gelatin/collagen and bioactive glass fibers for bone tissue engineering

    Energy Technology Data Exchange (ETDEWEB)

    Sharifi, Esmaeel; Azami, Mahmoud [Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Kajbafzadeh, Abdol-Mohammad [Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Pediatric Urology Research Center, Section of Tissue Engineering and Stem Cells Therapy, Department of Pediatric Urology, Children' s Hospital Medical Center, Tehran, Iran (IRI) (Iran, Islamic Republic of); Moztarzadeh, Fatollah [Department of Biomedical Engineering, Amirkabir University of Technology (Tehran Polytechnic), Tehran (Iran, Islamic Republic of); Faridi-Majidi, Reza [Department of Medical Nanotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Shamousi, Atefeh; Karimi, Roya [Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Ai, Jafar, E-mail: jafar_ai@tums.ac.ir [Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Brain and Spinal Injury Research Center (BASIR), Imam Khomeini Hospital, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of)

    2016-02-01

    Bone tissue is a composite material made of organic and inorganic components. Bone tissue engineering requires scaffolds that mimic bone nature in chemical and mechanical properties. This study proposes a novel method for preparing composite scaffolds that uses sub-micron bioglass fibers as the organic phase and gelatin/collagen as the inorganic phase. The scaffolds were constructed by using freeze drying and electro spinning methods and their mechanical properties were enhanced by using genipin crosslinking agent. Electron microscopy micrographs showed that the structure of composite scaffolds were porous with pore diameters of approximately 70–200 μm, this was again confirmed by mercury porosimetery. These pores are suitable for osteoblast growth. The diameters of the fibers were approximately 150–450 nm. Structural analysis confirmed the formation of desirable phases of sub-micron bioglass fibers. Cellular biocompatibility tests illustrated that scaffolds containing copper ion in the bioglass structure had more cell growth and osteoblast attachment in comparison to copper-free scaffolds. - Highlights: • Fabrication of 45S5 sub-micron bioglass fiber using electrospinning method. • Production of copper doped submicron bioglass fibers on 45S5 bioglass base by electrospinning sol gel route method. • Incorporation of bioglass/Cu-bioglass sub-micron fibers into gelatin/collagen matrix to form biomimetic composite scaffold which were non-cytotoxic according to MTT assay. • Discovering that copper can decrease the glass transition temperatures and enhance osteoblast cell adhesion and viability.

  7. Preparation of a biomimetic composite scaffold from gelatin/collagen and bioactive glass fibers for bone tissue engineering

    International Nuclear Information System (INIS)

    Sharifi, Esmaeel; Azami, Mahmoud; Kajbafzadeh, Abdol-Mohammad; Moztarzadeh, Fatollah; Faridi-Majidi, Reza; Shamousi, Atefeh; Karimi, Roya; Ai, Jafar

    2016-01-01

    Bone tissue is a composite material made of organic and inorganic components. Bone tissue engineering requires scaffolds that mimic bone nature in chemical and mechanical properties. This study proposes a novel method for preparing composite scaffolds that uses sub-micron bioglass fibers as the organic phase and gelatin/collagen as the inorganic phase. The scaffolds were constructed by using freeze drying and electro spinning methods and their mechanical properties were enhanced by using genipin crosslinking agent. Electron microscopy micrographs showed that the structure of composite scaffolds were porous with pore diameters of approximately 70–200 μm, this was again confirmed by mercury porosimetery. These pores are suitable for osteoblast growth. The diameters of the fibers were approximately 150–450 nm. Structural analysis confirmed the formation of desirable phases of sub-micron bioglass fibers. Cellular biocompatibility tests illustrated that scaffolds containing copper ion in the bioglass structure had more cell growth and osteoblast attachment in comparison to copper-free scaffolds. - Highlights: • Fabrication of 45S5 sub-micron bioglass fiber using electrospinning method. • Production of copper doped submicron bioglass fibers on 45S5 bioglass base by electrospinning sol gel route method. • Incorporation of bioglass/Cu-bioglass sub-micron fibers into gelatin/collagen matrix to form biomimetic composite scaffold which were non-cytotoxic according to MTT assay. • Discovering that copper can decrease the glass transition temperatures and enhance osteoblast cell adhesion and viability.

  8. Evaluation of human collagen biomaterials in the healing of colonic anastomoses in dogs.

    Science.gov (United States)

    Mutter, D; Aprahamian, M; Tiollier, J; Sonzini, P; Marescaux, J

    1997-04-01

    To investigate the ability of human collagen biomaterials to secure colonic anastomoses in dogs and to evaluate the biocompatibility of anastomotic protection patches (APP). Experimental open study. Experimental research centre, France. 21 mongrel dogs randomised into three groups of 7 each. Standard transverse colonic end-to-end anastomoses were secured with two-layer oxidised collagen I + III sponge covered with thin crosslinked collagen IV film (APP 1) glued around the suture (n = 7); two-layer oxidised collagen I + III sponge covered with thin non-crosslinked collagen I + III film patch (APP 2) (n = 7); or sealed by fibrin sealant (n = 7), which acted as a controls. Gross examination, radiological control (barium enemas), and microscopic examination on day 35 postoperatively. Gross clinical and radiological examinations on day 35 showed normal wound healing in all but one dog in which the anastomoses had occluded by day 16. There was significantly less stricturing with the APP 2 patch (p < 0.05 compared with the controls). Microscopic examination showed complete absorption of the APP 2 patches as well as quicker mucosal and extracellular matrix repair than controls. The APP 1 patch gave the best healing of the muscular layer but did not reduce anastomosis stricturing, and was not totally absorbed. Collagen supporting devices do not alter healing of the large bowel. Encircling patches do not increase the number of adhesions or the rate of anastomotic stricturing and a thin fibrillar collagen I + III dense layer may even improve it. The speed of absorption of the patch depends on the type of dense collagen film. These results argue for a prospective clinical evaluation in humans.

  9. Effect of indomethacin and lactoferrin on human tenocyte proliferation and collagen formation in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yaonan [Centre for Nanohealth, College of Medicine, Swansea University, Singleton Park, Swansea, UK SA2 8PP (United Kingdom); Department of Orthopaedic, Beijing Hospital of Ministry of Public Health, Beijing, China 100730 (China); Wang, Xiao; Qiu, Yiwei [Centre for Nanohealth, College of Medicine, Swansea University, Singleton Park, Swansea, UK SA2 8PP (United Kingdom); Cornish, Jillian [Department of Medicine, University of Auckland, Private Bag 92019, Auckland (New Zealand); Carr, Andrew J. [Centre for Nanohealth, College of Medicine, Swansea University, Singleton Park, Swansea, UK SA2 8PP (United Kingdom); Xia, Zhidao, E-mail: z.xia@swansea.ac.uk [Centre for Nanohealth, College of Medicine, Swansea University, Singleton Park, Swansea, UK SA2 8PP (United Kingdom)

    2014-11-14

    Highlights: • Indomethacin, a classic NSAID, inhibited human tenocyte proliferation at high concentration (100 µM). • Lactoferrin at 50-100 µg/ml promoted human tenocyte survival, proliferation and collagen synthesis. • Lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes. - Abstract: Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used in patients with injuries and inflammation of tendon and ligament, and as post-surgical analgesics. The aim of this study is to investigate the effect of indomethacin, a classic NSAID and its combinational effect with an anabolic agent of skeletal tissue, lactoferrin, on the proliferation and collagen formation of human tenocytes in vitro. A factorial experimental design was employed to study the dose-dependent effect of the combination of indomethacin and lactoferrin. The results showed that indomethacin at high concentration (100 μM) inhibited human tenocyte proliferation in culture medium with 1–10% fetal bovine serum (FBS) in vitro. Also, high dose of indomethacin inhibited the collagen formation of human tenocytes in 1% FBS culture medium. Lactoferrin at 50–100 μg/ml promoted human tenocyte survival in serum-free culture medium and enhanced proliferation and collagen synthesis of human tenocytes in 1% FBS culture medium. When 50–100 μg/ml lactoferrin was used in combination with 100–200 μM indomethacin, it partially rescued the inhibitory effects of indomethacin on human tenocyte proliferation, viability and collagen formation. To our knowledge, it is the first evidence that lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes.

  10. Effect of indomethacin and lactoferrin on human tenocyte proliferation and collagen formation in vitro

    International Nuclear Information System (INIS)

    Zhang, Yaonan; Wang, Xiao; Qiu, Yiwei; Cornish, Jillian; Carr, Andrew J.; Xia, Zhidao

    2014-01-01

    Highlights: • Indomethacin, a classic NSAID, inhibited human tenocyte proliferation at high concentration (100 µM). • Lactoferrin at 50-100 µg/ml promoted human tenocyte survival, proliferation and collagen synthesis. • Lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes. - Abstract: Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used in patients with injuries and inflammation of tendon and ligament, and as post-surgical analgesics. The aim of this study is to investigate the effect of indomethacin, a classic NSAID and its combinational effect with an anabolic agent of skeletal tissue, lactoferrin, on the proliferation and collagen formation of human tenocytes in vitro. A factorial experimental design was employed to study the dose-dependent effect of the combination of indomethacin and lactoferrin. The results showed that indomethacin at high concentration (100 μM) inhibited human tenocyte proliferation in culture medium with 1–10% fetal bovine serum (FBS) in vitro. Also, high dose of indomethacin inhibited the collagen formation of human tenocytes in 1% FBS culture medium. Lactoferrin at 50–100 μg/ml promoted human tenocyte survival in serum-free culture medium and enhanced proliferation and collagen synthesis of human tenocytes in 1% FBS culture medium. When 50–100 μg/ml lactoferrin was used in combination with 100–200 μM indomethacin, it partially rescued the inhibitory effects of indomethacin on human tenocyte proliferation, viability and collagen formation. To our knowledge, it is the first evidence that lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes

  11. Asiaticoside induces cell proliferation and collagen synthesis in human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Linda Yulianti

    2015-08-01

    Full Text Available Asiatiocoside, a saponin component isolated from Centella asiatica can improve wound healing by promoting the proliferation of human dermal fibroblasts (HDF and synthesis of collagen. The skin-renewing cells and type I and III collagen synthesis decrease with aging, resulting in the reduction of skin elasticity and delayed wound healing. Usage of natural active compounds from plants in wound healing should be evaluated and compared to retinoic acid as an active agent that regulates wound healing. The aim of this study was to compare and evaluate the effect of asiaticoside and retinoic acid to induce greater cell proliferation and type I and III collagen synthesis in human dermal fibroblast. Methods Laboratory experiments were conducted using human dermal fibroblasts (HDF isolated from human foreskin explants. Seven passages of HDF were treated with asiaticoside and retinoic acid at several doses and incubated for 24 and 48 hours. Cell viability in all groups was tested with the MTT assay to assess HDF proliferation. Type I and III collagen synthesis was examined using the respective ELISA kits. Analysis of variance was performed to compare the treatment groups. Results Asiaticoside had significantly stronger effects on HDF proliferation than retinoic acid (p<0.05. The type III collagen production was significantly greater induction with asiaticoside compared to retinoic acid (p<0.05. Conclusion Asiaticoside induces HDF proliferation and type I and III collagen synthesis in a time- and dose-dependent pattern. Asiaticoside has a similar effect as retinoic acid on type I and type III collagen synthesis.

  12. The Use of Injectable Chitosan/Nanohydroxyapatite/Collagen Composites with Bone Marrow Mesenchymal Stem Cells to Promote Ectopic Bone Formation In Vivo

    Directory of Open Access Journals (Sweden)

    Bo Yu

    2013-01-01

    Full Text Available The aim of this study was to evaluate ectopic in vivo bone formation with or without rat bone mesenchymal stem cells (rBMSCs of an injectable Chitosan/Nanohydroxyapatite/Collagen (CS/nHAC composite. The CS/nHAC composites were injected subcutaneously into the backs of Wistar rats with freshly loaded rBMSCs at a density of 10×106 cells/mL, and the CS/nHAC composites without cells were used as negative controls. New bone formation, degradation of composites, and degree of calcification were evaluated by Computed Tomography (CT and three-dimensional (3D CT reconstruction. Histological evaluations were performed to further assess bone structure and extracellular matrix by HE and Masson staining. The inflammatory reactions related to osteogenesis were also investigated in the present study. In comparison with the CS/nHAC composites, this study revealed that CS/nHAC/rBMSCs composites showed relatively higher percentage of calcification, better establishment of ECM, and less degradation rate. Meanwhile, different extents of inflammatory reactions were also observed in the CS/nHAC and CS/nHAC/rBMSCs explants at 2 and 4 weeks after implantation. Altogether, CS/nHAC/rBMSCs composites are superior to CS/nHAC composites in ectopic bone formation. In conclusion, the rBMSCs-seeded CS/nHAC composites may be beneficial to enhancing ectopic bone formation in vivo.

  13. The Great Irish Famine: Identifying Starvation in the Tissues of Victims Using Stable Isotope Analysis of Bone and Incremental Dentine Collagen.

    Directory of Open Access Journals (Sweden)

    Julia Beaumont

    Full Text Available The major components of human diet both past and present may be estimated by measuring the carbon and nitrogen isotope ratios (δ13C and δ15N of the collagenous proteins in bone and tooth dentine. However, the results from these two tissues differ substantially: bone collagen records a multi-year average whilst primary dentine records and retains time-bound isotope ratios deriving from the period of tooth development. Recent studies harnessing a sub-annual temporal sampling resolution have shed new light on the individual dietary histories of our ancestors by identifying unexpected radical short-term dietary changes, the duration of breastfeeding and migration where dietary change occurs, and by raising questions regarding factors other than diet that may impact on δ13C and δ15N values. Here we show that the dentine δ13C and δ15N profiles of workhouse inmates dating from the Great Irish Famine of the 19th century not only record the expected dietary change from C3 potatoes to C4 maize, but when used together they also document prolonged nutritional and other physiological stress resulting from insufficient sustenance. In the adults, the influence of the maize-based diet is seen in the δ13C difference between dentine (formed in childhood and rib (representing an average from the last few years of life. The demonstrated effects of stress on the δ13C and δ15N values will have an impact on the interpretations of diet in past populations even in slow-turnover tissues such as compact bone. This technique also has applicability in the investigation of modern children subject to nutritional distress where hair and nails are unavailable or do not record an adequate period of time.

  14. Bone blood flow and metabolism in humans

    DEFF Research Database (Denmark)

    Heinonen, Ilkka; Kemppainen, Jukka; Kaskinoro, Kimmo

    2012-01-01

    Human bone blood flow and metabolism during physical exercise remains poorly characterised. In the present study we measured femoral bone blood flow and glucose uptake in young healthy subjects by positron emission tomography in three separate protocols. In six women, blood flow was measured...... in femoral bone at rest and during one leg intermittent isometric exercise with increasing exercise intensities. In nine men, blood flow in femur was determined at rest and during dynamic one leg exercise, and two other physiological perturbations: moderate systemic hypoxia (14 O(2) ) at rest and during...... exercise, and during intra-femoral infusion of high-dose adenosine. Bone glucose uptake was measured at rest and during dynamic one leg exercise in five men. The results indicate that isometric exercise increased femoral bone blood flow from rest (1.8 ± 0.6 ml/100g/min) to low intensity exercise (4.1 ± 1...

  15. Uranium concentrations in human bone

    International Nuclear Information System (INIS)

    Schlenker, R.A.; Oltman, B.G.

    1981-01-01

    The uranium concentration in bone from an individual injected with 239 Pu has been determined, using the fission-track method. The data are consistent with those reported about 10 years ago by Welford and Baird for New York City area residents and by Hamilton in England. They are at variance with the more recent data of Welford et al

  16. Enhanced hyaline cartilage matrix synthesis in collagen sponge scaffolds by using siRNA to stabilize chondrocytes phenotype cultured with bone morphogenetic protein-2 under hypoxia.

    Science.gov (United States)

    Legendre, Florence; Ollitrault, David; Hervieu, Magalie; Baugé, Catherine; Maneix, Laure; Goux, Didier; Chajra, Hanane; Mallein-Gerin, Frédéric; Boumediene, Karim; Galera, Philippe; Demoor, Magali

    2013-07-01

    Cartilage healing by tissue engineering is an alternative strategy to reconstitute functional tissue after trauma or age-related degeneration. However, chondrocytes, the major player in cartilage homeostasis, do not self-regenerate efficiently and lose their phenotype during osteoarthritis. This process is called dedifferentiation and also occurs during the first expansion step of autologous chondrocyte implantation (ACI). To ensure successful ACI therapy, chondrocytes must be differentiated and capable of synthesizing hyaline cartilage matrix molecules. We therefore developed a safe procedure for redifferentiating human chondrocytes by combining appropriate physicochemical factors: hypoxic conditions, collagen scaffolds, chondrogenic factors (bone morphogenetic protein-2 [BMP-2], and insulin-like growth factor I [IGF-I]) and RNA interference targeting the COL1A1 gene. Redifferentiation of dedifferentiated chondrocytes was evaluated using gene/protein analyses to identify the chondrocyte phenotypic profile. In our conditions, under BMP-2 treatment, redifferentiated and metabolically active chondrocytes synthesized a hyaline-like cartilage matrix characterized by type IIB collagen and aggrecan molecules without any sign of hypertrophy or osteogenesis. In contrast, IGF-I increased both specific and noncharacteristic markers (collagens I and X) of chondrocytes. The specific increase in COL2A1 gene expression observed in the BMP-2 treatment was shown to involve the specific enhancer region of COL2A1 that binds the trans-activators Sox9/L-Sox5/Sox6 and Sp1, which are associated with a decrease in the trans-inhibitors of COL2A1, c-Krox, and p65 subunit of NF-kappaB. Our procedure in which BMP-2 treatment under hypoxia is associated with a COL1A1 siRNA, significantly increased the differentiation index of chondrocytes, and should offer the opportunity to develop new ACI-based therapies in humans.

  17. Inhibition of human arterial smooth muscle (HASM) cell proliferation and collagen synthesis by protamine

    International Nuclear Information System (INIS)

    Drucker, D.E.; Graham, M.F.; Diegelmann, R.F.; Greenfield, L.J.

    1986-01-01

    Atherosclerotic plaques result from vascular smooth muscle cell proliferation and collagen deposition. The authors have been studying factors which modulate HASM cell proliferation and collagen synthesis. HASM cells were isolated from the media of normal human thoracic and infrarenal aortas and grown in vitro. Cell numbers were determined by direct counting and collagen synthesis was measured by incorporation of 3 H-proline into collagenase-digestible protein. In this study, protamine (200 μg/ml) was tested and found to cause a 55% reduction of HASM cell proliferation which was reversible when the cells were returned to control medium or when heparin (100 μg/ml) was added with protamine. Protamine caused a constant 33% decrease in non-collagen protein (NCP) synthesis per cell. In contrast, collagen synthesis was inhibited in dose dependent fashion (88% reduction at 200 μg/ml). Protamine blocks HASM cell proliferation and specifically inhibits collagen production. The exact mechanism of this inhibition is unclear but may be related to a transcriptional event since protamine has a high affinity for DNA

  18. Generation of biologically active endostatin fragments from human collagen XVIII by distinct matrix metalloproteases

    International Nuclear Information System (INIS)

    Heljasvaara, Ritva; Nyberg, Pia; Luostarinen, Jani; Parikka, Mataleena; Heikkilae, Pia; Rehn, Marko; Sorsa, Timo; Salo, Tuula; Pihlajaniemi, Taina

    2005-01-01

    Endostatin, a potent inhibitor of endothelial cell proliferation, migration, angiogenesis and tumor growth, is proteolytically cleaved from the C-terminal noncollagenous NC1 domain of type XVIII collagen. We investigated the endostatin formation from human collagen XVIII by several MMPs in vitro. The generation of endostatin fragments differing in molecular size (24-30 kDa) and in N-terminal sequences was identified in the cases of MMP-3, -7, -9, -13 and -20. The cleavage sites were located in the protease-sensitive hinge region between the trimerization and endostatin domains of NC1. MMP-1, -2, -8 and -12 did not show any significant activity against the C-terminus of collagen XVIII. The anti-proliferative effect of the 20-kDa endostatin, three longer endostatin-containing fragments generated in vitro by distinct MMPs and the entire NC1 domain, on bFGF-stimulated human umbilical vein endothelial cells was established. The anti-migratory potential of some of these fragments was also studied. In addition, production of endostatin fragments between 24-30 kDa by human hepatoblastoma cells was shown to be due to MMP action on type XVIII collagen. Our results indicate that certain, especially cancer-related, MMP family members can generate biologically active endostatin-containing polypeptides from collagen XVIII and thus, by releasing endostatin fragments, may participate in the inhibition of endothelial cell proliferation, migration and angiogenesis

  19. The structural and optical properties of type III human collagen biosynthetic corneal substitutes

    Science.gov (United States)

    Hayes, Sally; Lewis, Phillip; Islam, M. Mirazul; Doutch, James; Sorensen, Thomas; White, Tomas; Griffith, May; Meek, Keith M.

    2015-01-01

    The structural and optical properties of clinically biocompatible, cell-free hydrogels comprised of synthetically cross-linked and moulded recombinant human collagen type III (RHCIII) with and without the incorporation of 2-methacryloyloxyethyl phosphorylcholine (MPC) were assessed using transmission electron microscopy (TEM), X-ray scattering, spectroscopy and refractometry. These findings were examined alongside similarly obtained data from 21 human donor corneas. TEM demonstrated the presence of loosely bundled aggregates of fine collagen filaments within both RHCIII and RHCIII-MPC implants, which X-ray scattering showed to lack D-banding and be preferentially aligned in a uniaxial orientation throughout. This arrangement differs from the predominantly biaxial alignment of collagen fibrils that exists in the human cornea. By virtue of their high water content (90%), very fine collagen filaments (2–9 nm) and lack of cells, the collagen hydrogels were found to transmit almost all incident light in the visible spectrum. They also transmitted a large proportion of UV light compared to the cornea which acts as an effective UV filter. Patients implanted with these hydrogels should be cautious about UV exposure prior to regrowth of the epithelium and in-growth of corneal cells into the implants. PMID:26159106

  20. Collagen cross-linking in sun-exposed and unexposed sites of aged human skin

    Science.gov (United States)

    Yamauchi, M.; Prisayanh, P.; Haque, Z.; Woodley, D. T.

    1991-01-01

    A recently described nonreducible, acid-heat stable compound, histidinohydroxylysinonorleucine (HHL), is a collagen cross-link isolated from mature skin tissue. Its abundance is related to chronologic aging of skin. The present communication describes the quantity of HHL from aged human skin of the same individuals in sun-exposed (wrist) and unexposed (buttock) sites. Punch biopsies were obtained from these sites from nine people of age 60 or older. HHL contents (moles/mole of collagen) at these sites were for wrist 0.13 +/- 0.07 and for buttock 0.69 +/- 0.17 (mean +/- SD, p less than 0.001). In addition, it was found that acute irradiation of the cross-linked peptides with UVA (up to 250 J/cm2) and UVB (up to 1 J/cm2) had no effect on HHL structure. The same treatment significantly degraded another nonreducible, stable collagen cross-link, pyridinoline. The results suggest that chronic sunlight exposure may be associated with an impediment to normal maturation of human dermal collagen resulting in tenuous amount of HHL. Thus, the process of photoaging in dermal collagen is different from that of chronologic aging in human skin.

  1. Eccentric rehabilitation exercise increases peritendinous type I collagen synthesis in humans with Achilles tendinosis.

    Science.gov (United States)

    Langberg, H; Ellingsgaard, H; Madsen, T; Jansson, J; Magnusson, S P; Aagaard, P; Kjaer, M

    2007-02-01

    It has been shown that 12 weeks of eccentric heavy resistance training can reduce pain in runners suffering from chronic Achilles tendinosis, but the mechanism behind the effectiveness of this treatment is unknown. The present study investigates the local effect of an eccentric training regime on elite soccer players suffering from chronic Achilles tendinosis on the turnover of the peritendinous connective tissue. Twelve elite male soccer players, of whom six suffered from unilateral tendinosis and six were healthy controls, participated in this study. All participants performed 12 weeks of heavy-resistance eccentric training apart from their regular training and soccer activity. Before and after the training period the tissue concentration of indicators of collagen turnover was measured by the use of the microdialysis technique. After training, collagen synthesis was increased in the initially injured tendon (n=6; carboxyterminal propeptide of type I collagen (PICP): pre 3.9+/-2.5 microg/L to post 19.7+/-5.4 microg/L, Ptendons in response to training (n=6; PICP: pre 8.3+/-5.2 microg/L to post 11.5+/-5.0 microg/L, P>0.05). Collagen degradation, measured as carboxyterminal telopeptide region of type I collagen (ICTP), was not affected by training neither in the injured nor in the healthy tendons. The clinical effect of the 12 weeks of eccentric training was determined by using a standardized loading procedure of the Achilles tendons showing a decrease in pain in all the chronic injured tendons (VAS before 44+/-9, after 13+/-9; Peccentric training regime. The present study demonstrates that chronically injured Achilles tendons respond to 12 weeks of eccentric training by increasing collagen synthesis rate. In contrast, the collagen metabolism in healthy control tendons seems not to be affected by eccentric training. These findings could indicate a relation between collagen metabolism and recovery from injury in human tendons.

  2. [Conservative anal fistula treatment with collagenic plug and human fibrin sealant. Preliminary results].

    Science.gov (United States)

    Gubitosi, A; Moccia, G; Malinconico, F A; Docimo, G; Ruggiero, R; Iside, G; Avenia, N; Docimo, L; Foroni, F; Gilio, F; Sparavigna, L; Agresti, M

    2009-01-01

    The authors, on the basis of a long clinical experience with human fibrin glue in general surgery, compared two different extracellular matrix (collagen), Surgisis and TissueDura, with human fibrin glue, applied during the operation, and sometimes in postoperative, to obtain the healing of perianal fistulas. The collagenic extracellular matrix provides, according to the rationale suggested, an optimal three-dimensional structure for the fibroblastic implant and neoangiogenesis, hence for the fistula "fibrotizzation" and closure. The encouraging results for transphincteric fistulas and a simple and easy technique push to researchers on samples statistically significant.

  3. Evaluation of 3D printed PCL/PLGA/β-TCP versus collagen membranes for guided bone regeneration in a beagle implant model.

    Science.gov (United States)

    Won, J-Y; Park, C-Y; Bae, J-H; Ahn, G; Kim, C; Lim, D-H; Cho, D-W; Yun, W-S; Shim, J-H; Huh, J-B

    2016-10-07

    Here, we compared 3D-printed polycaprolactone/poly(lactic-co-glycolic acid)/β-tricalcium phosphate (PCL/PLGA/β-TCP) membranes with the widely used collagen membranes for guided bone regeneration (GBR) in beagle implant models. For mechanical property comparison in dry and wet conditions and cytocompatibility determination, we analyzed the rate and pattern of cell proliferation of seeded fibroblasts and preosteoblasts using the cell counting kit-8 assay and scanning electron microscopy. Osteogenic differentiation was verified using alizarin red S staining. At 8 weeks following implantation in vivo using beagle dogs, computed tomography and histological analyses were performed after sacrifice. Cell proliferation rates in vitro indicated that early cell attachment was higher in collagen than in PCL/PLGA/β-TCP membranes; however, the difference subsided by day 7. Similar outcomes were found for osteogenic differentiation, with approximately 2.5 times greater staining in collagen than PCL/PLGA/β-TCP, but without significant difference by day 14. In vivo, bone regeneration in the defect area, represented by new bone formation and bone-to-implant contact, paralleled those associated with collagen membranes. However, tensile testing revealed that whereas the PCL/PLGA/β-TCP membrane mechanical properties were conserved in both wet and dry states, the tensile property of collagen was reduced by 99% under wet conditions. Our results demonstrate in vitro and in vivo that PCL/PLGA/β-TCP membranes have similar levels of biocompatibility and bone regeneration as collagen membranes. In particular, considering that GBR is always applied to a wet environment (e.g. blood, saliva), we demonstrated that PCL/PLGA/β-TCP membranes maintained their form more reliably than collagen membranes in a wet setting, confirming their appropriateness as a GBR membrane.

  4. Micro-CT characterization of human trabecular bone in osteogenesis imperfecta

    Science.gov (United States)

    Jameson, John; Albert, Carolyne; Smith, Peter; Molthen, Robert; Harris, Gerald

    2011-03-01

    Osteogenesis imperfecta (OI) is a genetic syndrome affecting collagen synthesis and assembly. Its symptoms vary widely but commonly include bone fragility, reduced stature, and bone deformity. Because of the small size and paucity of human specimens, there is a lack of biomechanical data for OI bone. Most literature has focused on histomorphometric analyses, which rely on assumptions to extrapolate 3-D properties. In this study, a micro-computed tomography (μCT) system was used to directly measure structural and mineral properties in pediatric OI bone collected during routine surgical procedures. Surface renderings suggested a poorly organized, plate-like orientation. Patients with a history of bone-augmenting drugs exhibited increased bone volume fraction (BV/TV), trabecular number (Tb.N), and connectivity density (Eu.Conn.D). The latter two parameters appeared to be related to OI severity. Structural results were consistently higher than those reported in a previous histomorphometric study, but these differences can be attributed to factors such as specimen collection site, drug therapy, and assumptions associated with histomorphometry. Mineral testing revealed strong correlations with several structural parameters, highlighting the importance of a dual approach in trabecular bone testing. This study reports some of the first quantitative μCT data of human OI bone, and it suggests compelling possibilities for the future of OI bone assessment.

  5. GH receptor blocker administration and muscle-tendon collagen synthesis in humans

    DEFF Research Database (Denmark)

    Nielsen, Rie Harboe; Doessing, Simon; Goto, Kazushige

    2011-01-01

    Collagen is the predominant structural protein in tendons and ligaments, and can be controlled by hormonal changes. In animals, injections of insulin-like growth factor I (IGF-I) has been shown to increase collagen synthesis in tendons and ligaments and to improve structural tissue healing......, but the effect of local IGF-I administration on tendon collagen synthesis in human has not been studied. The purpose of this study was to study whether local injections of IGF-I would have a stimulating effect on tendon collagen synthesis. Twelve healthy nonsmoking men [age 62 ± 1 years (mean ± SEM), BMI 27 ± 1......] participated. Two injections of either human recombinant IGF-I (0.1 mL Increlex©) or saline (control) into each patellar tendon were performed 24-h apart, respectively. Tendon collagen fractional synthesis rate (FSR) was measured by stable isotope technique in the hours after the second injection...

  6. Fabrication and Characterization of Collagen-Immobilized Porous PHBV/HA Nano composite Scaffolds for Bone Tissue Engineering

    International Nuclear Information System (INIS)

    Jin-Young, B.; Zhi-Cai, X.; Giseop, K.; Keun-Byoung, Y.; Soo-Young, P.; Lee, S.P.; Inn-Kyu, K.

    2012-01-01

    The porous composite scaffolds (PHBV/HA) consisting of poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) and hydroxyapatite (HA) were fabricated using a hot-press machine and salt-leaching. Collagen (type I) was then immobilized on the surface of the porous PHBV/HA composite scaffolds to improve tissue compatibility. The structure and morphology of the collagen-immobilized composite scaffolds (PHBV/HA/Col) were investigated using a scanning electron microscope (SEM), Fourier transform infrared (FTIR), and electron spectroscopy for chemical analysis (ESCA). The potential of the porous PHBV/HA/Col composite scaffolds for use as a bone scaffold was assessed by an experiment with osteoblast cells (MC3T3-E1) in terms of cell adhesion, proliferation, and differentiation. The results showed that the PHBV/HA/Col composite scaffolds possess better cell adhesion and significantly higher proliferation and differentiation than the PHBV/HA composite scaffolds and the PHBV scaffolds. These results suggest that the PHBV/HA/Col composite scaffolds have a high potential for use in the field of bone regeneration and tissue engineering.

  7. Effects of cell-attachment and extracellular matrix on bone formation in vivo in collagen-hydroxyapatite scaffolds.

    Science.gov (United States)

    Villa, Max M; Wang, Liping; Rowe, David W; Wei, Mei

    2014-01-01

    Cell-based tissue engineering can be used to replace missing or damaged bone, but the optimal methods for delivering therapeutic cells to a bony defect have not yet been established. Using transgenic reporter cells as a donor source, two different collagen-hydroxyapatite (HA) scaffolds, and a critical-size calvarial defect model, we investigated the effect of a cell-attachment period prior to implantation, with or without an extracellular matrix-based seeding suspension, on cell engraftment and osteogenesis. When quantitatively compared, the in-house scaffold implanted immediately had a higher mean radiopacity than in-house scaffolds incubated overnight. Both scaffold types implanted immediately had significantly higher area fractions of donor cells, while the in-house collagen-HA scaffolds implanted immediately had higher area fractions of the mineralization label compared with groups incubated overnight. When the cell loading was compared in vitro for each delivery method using the in-house scaffold, immediate loading led to higher numbers of delivered cells. Immediate loading may be preferable in order to ensure robust bone formation in vivo. The use of a secondary ECM carrier improved the distribution of donor cells only when a pre-attachment period was applied. These results have improved our understanding of cell delivery to bony defects in the context of in vivo outcomes.

  8. Antioxidative Peptides Derived from Enzyme Hydrolysis of Bone Collagen after Microwave Assisted Acid Pre-Treatment and Nitrogen Protection

    Directory of Open Access Journals (Sweden)

    Jin Sun

    2010-11-01

    Full Text Available This study focused on the preparation method of antioxidant peptides by enzymatic hydrolysis of bone collagen after microwave assisted acid pre-treatment and nitrogen protection. Phosphoric acid showed the highest ability of hydrolysis among the four other acids tested (hydrochloric acid, sulfuric acid and/or citric acid. The highest degree of hydrolysis (DH was 9.5% using 4 mol/L phosphoric acid with a ratio of 1:6 under a microwave intensity of 510 W for 240 s. Neutral proteinase gave higher DH among the four protease tested (Acid protease, neutral protease, Alcalase and papain, with an optimum condition of: (1 ratio of enzyme and substrate, 4760 U/g; (2 concentration of substrate, 4%; (3 reaction temperature, 55 °C and (4 pH 7.0. At 4 h, DH increased significantly (P < 0.01 under nitrogen protection compared with normal microwave assisted acid pre-treatment hydrolysis conditions. The antioxidant ability of the hydrolysate increased and reached its maximum value at 3 h; however DH decreased dramatically after 3 h. Microwave assisted acid pre-treatment and nitrogen protection could be a quick preparatory method for hydrolyzing bone collagen.

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

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Yoon Jung [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Lee, Jue Yeon [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Lee, Seung Jin [Department of Industrial Pharmacy, College of Pharmacy, Ewha Womans University, Seoul (Korea, Republic of); Chung, Chong-Pyoung, E-mail: ccpperio@snu.ac.kr [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Department of Periodontology, School of Dentistry and Dental Research Institute, Seoul National University, Seoul (Korea, Republic of); Park, Yoon Jeong, E-mail: parkyj@snu.ac.kr [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of)

    2012-03-09

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

  10. Biomimetic fabrication of a three-level hierarchical calcium phosphate/collagen/hydroxyapatite scaffold for bone tissue engineering

    International Nuclear Information System (INIS)

    Zhou, Changchun; Ye, Xingjiang; Fan, Yujiang; Tan, Yanfei; Qing, Fangzu; Zhang, Xingdong; Ma, Liang

    2014-01-01

    A three-level hierarchical calcium phosphate/collagen/hydroxyapatite (CaP/Col/HAp) scaffold for bone tissue engineering was developed using biomimetic synthesis. Porous CaP ceramics were first prepared as substrate materials to mimic the porous bone structure. A second-level Col network was then composited into porous CaP ceramics by vacuum infusion. Finally, a third-level HAp layer was achieved by biomimetic mineralization. The three-level hierarchical biomimetic scaffold was characterized using scanning electron microscopy, energy-dispersive x-ray spectra, x-ray diffraction and Fourier transform infrared spectroscopy, and the mechanical properties of the scaffold were evaluated using dynamic mechanical analysis. The results show that this scaffold exhibits a similar structure and composition to natural bone tissues. Furthermore, this three-level hierarchical biomimetic scaffold showed enhanced mechanical strength compared with pure porous CaP scaffolds. The biocompatibility and osteoinductivity of the biomimetic scaffolds were evaluated using in vitro and in vivo tests. Cell culture results indicated the good biocompatibility of this biomimetic scaffold. Faster and increased bone formation was observed in these scaffolds following a six-month implantation in the dorsal muscles of rabbits, indicating that this biomimetic scaffold exhibits better osteoinductivity than common CaP scaffolds. (papers)

  11. Age changes in human bone: an overview

    Energy Technology Data Exchange (ETDEWEB)

    Sharpe, W.D.

    1977-12-03

    The human skeleton steadily changes structure and mass during life because of a variety of internal and external factors. Extracellular substance and bone cells get old, characteristic structural remodeling occurs with age and these age-related changes are important in the discrimination between pathological and physiological changes. Perhaps 20 percent of the bone mass is lost between the fourth and the ninth decades, osteoblasts function less efficiently and gradual loss of bone substance is enhanced by delayed mineralization of an increased surface area of thin and relatively less active osteoid seams. After the fifth decade, osteoclasia and the number of Howship's lacunae increase, and with age, the number of large osteolytic osteocytes increases as the number of small osteocytes declines and empty osteocyte lacunae become more common. The result is greater liability to fracture and diminished healing or replacement of injured bone.

  12. Determination of markers for collagen type I turnover in peritendinous human tissue by microdialysis

    DEFF Research Database (Denmark)

    Olesen, J L; Langberg, Henning; Heinemeier, K M

    2006-01-01

    Previous results from our group have shown that loading of human tendon elevates tendinous type I collagen production measured by microdialysis. However, exclusion of the observed elevation as a response to trauma from inserting the microdialysis catheters or a possible influence from the collage...

  13. Mechanical properties of the collagen network in human articular cartilage as measured by osmotic stress technique

    NARCIS (Netherlands)

    Basser, P.J.; Schneiderman, R.; Bank, R.A.; Wachtel, E.; Maroudas, A.

    1998-01-01

    We have used an isotropic osmotic stress technique to assess the swelling pressures of human articular cartilage over a wide range of hydrations in order to determine from these measurements, for the first time, the tensile stress in the collagen network, P(c), as a function of hydration. Osmotic

  14. Metabolic activity and collagen turnover in human tendon in response to physical activity

    DEFF Research Database (Denmark)

    Kjaer, M; Langberg, H; Miller, B F

    2005-01-01

    Connective tissue of the human tendon plays an important role in force transmission. The extracellular matrix turnover of tendon is influenced by physical activity. Blood flow, oxygen demand, and the level of collagen synthesis and matrix metalloproteinases increase with mechanical loading. Gene...... of overuse tendon injuries occurring during sport, work or leisure-related activities....

  15. Engineered collagen hydrogels for the sustained release of biomolecules and imaging agents: promoting the growth of human gingival cells.

    Science.gov (United States)

    Choi, Jonghoon; Park, Hoyoung; Kim, Taeho; Jeong, Yoon; Oh, Myoung Hwan; Hyeon, Taeghwan; Gilad, Assaf A; Lee, Kwan Hyi

    2014-01-01

    We present here the in vitro release profiles of either fluorescently labeled biomolecules or computed tomography contrast nanoagents from engineered collagen hydrogels under physiological conditions. The collagen constructs were designed as potential biocompatible inserts into wounded human gingiva. The collagen hydrogels were fabricated under a variety of conditions in order to optimize the release profile of biomolecules and nanoparticles for the desired duration and amount. The collagen constructs containing biomolecules/nanoconstructs were incubated under physiological conditions (ie, 37°C and 5% CO2) for 24 hours, and the release profile was tuned from 20% to 70% of initially loaded materials by varying the gelation conditions of the collagen constructs. The amounts of released biomolecules and nanoparticles were quantified respectively by measuring the intensity of fluorescence and X-ray scattering. The collagen hydrogel we fabricated may serve as an efficient platform for the controlled release of biomolecules and imaging agents in human gingiva to facilitate the regeneration of oral tissues.

  16. Prdm5 Regulates Collagen Gene Transcription by Association with RNA Polymerase II in Developing Bone

    DEFF Research Database (Denmark)

    Galli, Giorgio Giacomo; Honnens de Lichtenberg, Kristian; Carrara, Matteo

    2012-01-01

    and fibrillogenesis by binding inside the Col1a1 gene body and maintaining RNA polymerase II occupancy. In vivo, Prdm5 loss results in delayed ossification involving a pronounced impairment in the assembly of fibrillar collagens. Collectively, our results define a novel role for Prdm5 in sustaining...

  17. Use of carboxymethyl cellulose and collagen carrier with equine bone lyophilisate suggests late onset bone regenerative effect in a humerus drill defect - a pilot study in six sheep

    DEFF Research Database (Denmark)

    Jensen, Jonas; Foldager, Casper Bindzus; Jakobsen, Thomas Vestergaard

    2010-01-01

    in the other. The animals were divided into three groups of two animals and observed for 8, 12 and 16 weeks. Drill holes was evaluated using quantitative computed tomography (QCT), micro computed tomography (microCT) and histomorphometry. Mean total bone mineral density (BMD) of each implantation site...... was calculated with both QCT and microCT. Bone volume to total volume (BV/TV) was analyzed using microCT and histomorphometry. Although not statistically significant, results showed increased bone BMD after 16 weeks in microCT data and an increased BV/TV after 16 weeks in both microCT and histology. Correlation...... between QCT and microCT was R(2) = 0.804. Correlation between histomorphometry and microCT BV/TV data was R(2) = 0.8935 and with an average overrepresentation of 8.2% in histomorphometry. In conclusion the CMC-Collagen + Colloss E filler seems like a viable osteogenic bone filler mid- to long term...

  18. Skeletal muscle collagen content in humans after high-force eccentric contractions

    DEFF Research Database (Denmark)

    Mackey, Abigail; Donnelly, Alan E; Turpeenniemi-Hujanen, Taina

    2004-01-01

    The purpose of this study was to investigate the effects of high-force eccentric muscle contractions on collagen remodeling and on circulating levels of matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) in humans. Nine volunteers [5 men and 4 women, mean age 23 (SD...... 4) yr] each performed a bout of 100 maximum voluntary eccentric contractions of the knee extensors. Muscle biopsies were taken before exercise and on days 4 and 22 afterward. Image analysis of stained tissue sections was used to quantify endomysial collagen staining intensity. Maximum voluntary...... contractile force declined by 39 +/- 23% (mean +/- SD) on day 2 postexercise and recovered thereafter. Serum creatine kinase activity peaked on day 4 postexercise (P Collagen type IV staining intensity increased significantly on day 22 postexercise to 126 +/- 29% (mean +/- SD) of preexercise values...

  19. Micro/Nano Multilayered Scaffolds of PLGA and Collagen by Alternately Electrospinning for Bone Tissue Engineering

    Science.gov (United States)

    Kwak, Sanghwa; Haider, Adnan; Gupta, Kailash Chandra; Kim, Sukyoung; Kang, Inn-Kyu

    2016-07-01

    The dual extrusion electrospinning technique was used to fabricate multilayered 3D scaffolds by stacking microfibrous meshes of poly(lactic acid-co-glycolic acid) (PLGA) in alternate fashion to micro/nano mixed fibrous meshes of PLGA and collagen. To fabricate the multilayered scaffold, 35 wt% solution of PLGA in THF-DMF binary solvent (3:1) and 5 wt% solution of collagen in hexafluoroisopropanol (HFIP) with and without hydroxyapatite nanorods (nHA) were used. The dual and individual electrospinning of PLGA and collagen were carried out at flow rates of 1.0 and 0.5 mL/h, respectively, at an applied voltage of 20 kV. The density of collagen fibers in multilayered scaffolds has controlled the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 cells. The homogeneous dispersion of glutamic acid-modified hydroxyapatite nanorods (nHA-GA) in collagen solution has improved the osteogenic properties of fabricated multilayered scaffolds. The fabricated multilayered scaffolds were characterized using FT-IR, X-ray photoelectron spectroscopy, and transmission electron microscopy (TEM). The scanning electron microscopy (FE-SEM) was used to evaluate the adhesion and spreads of MC3T3-E1 cells on multilayered scaffolds. The activity of MC3T3-E1 cells on the multilayered scaffolds was evaluated by applying MTT, alkaline phosphatase, Alizarin Red, von Kossa, and cytoskeleton F-actin assaying protocols. The micro/nano fibrous PLGA-Col-HA scaffolds were found to be highly bioactive in comparison to pristine microfibrous PLGA and micro/nano mixed fibrous PLGA and Col scaffolds.

  20. Regionally variant collagen alignment correlates with viscoelastic properties of the disc of the human temporomandibular joint.

    Science.gov (United States)

    Gutman, Shawn; Kim, Daniel; Tarafder, Solaiman; Velez, Sergio; Jeong, Julia; Lee, Chang H

    2018-02-01

    To determine the regionally variant quality of collagen alignment in human TMJ discs and its statistical correlation with viscoelastic properties. For quantitative analysis of the quality of collagen alignment, horizontal sections of human TMJ discs with Pricrosirius Red staining were imaged under circularly polarized microscopy. Mean angle and angular deviation of collagen fibers in each region were analyzed using a well-established automated image-processing for angular gradient. Instantaneous and relaxation moduli of each disc region were measured under stress-relaxation test both in tensile and compression. Then Spearman correlation analysis was performed between the angular deviation and the moduli. To understand the effect of glycosaminoglycans on the correlation, TMJ disc samples were treated by chondroitinase ABC (C-ABC). Our imaging processing analysis showed the region-variant direction of collagen alignment, consistently with previous findings. Interestingly, the quality of collagen alignment, not only the directions, was significantly different in between the regions. The angular deviation of fiber alignment in the anterior and intermediate regions were significantly smaller than the posterior region. Medial and lateral regions showed significantly bigger angular deviation than all the other regions. The regionally variant angular deviation values showed statistically significant correlation with the tensile instantaneous modulus and the relaxation modulus, partially dependent on C-ABC treatment. Our findings suggest the region-variant degree of collagen fiber alignment is likely attributed to the heterogeneous viscoelastic properties of TMJ disc that may have significant implications in development of regenerative therapy for TMJ disc. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Partial characterization of a low molecular weight human collagen that undergoes alternative splicing

    International Nuclear Information System (INIS)

    Pihlajaniemi, T.; Myllylea, R.; Kurkinen, M.; Prockop, D.J.

    1987-01-01

    A cDNA library prepared from RNA isolated from a cultured human tumor cell line, HT-1080, was screened with a mouse cDNA clone coding for part of the -Gly-Xaa-Yaa-domain of the α2(IV) collagen chain. Four overlapping cDNA clones were characterized that coded for a low molecular weight human collagen. The cDNA clones did not, however, code for the short-chain collagens, types IX and X. The amino acid sequences derived from the clones resembled type IV collagen in that there were short interruptions in the repeating -Gly-Xaa-Yaa-sequence. The noncollagenous, carboxyl-terminal domain was, however, much shorter and contained only 18 amino acid residues. Interestingly, one of the cDNA clones contained an additional 36 nucleotides not found in an overlapping clone. The 36 nucleotides encoded four -Gly-Xaa-Yaa-repeats without changing the reading frame. Nuclease S1 mapping using a 32 P-labelled probe demonstrated that the different between the clones was due to existence of two different mRNAs. A synthetic 24-residue peptide corresponding to the last two -Gly-Xaa-Yaa-triplets and the entire carboxyl-terminal domain was used to generate polyclonal antibodies. Electrophoretic transfer blot analysis of HT-1080 cells and normal human skin fibroblasts identified two polypeptides, M/sub r/ 67,000 and M/sub r/ 62,000, that were sensitive to bacterial collagenase

  2. Osteoinduction and proliferation of bone-marrow stromal cells in three-dimensional poly (ε-caprolactone)/ hydroxyapatite/collagen scaffolds.

    Science.gov (United States)

    Wang, Ting; Yang, Xiaoyan; Qi, Xin; Jiang, Chaoyin

    2015-05-08

    Osteoinduction and proliferation of bone-marrow stromal cells (BMSCs) in three-dimensional (3D) poly(ε-caprolactone) (PCL) scaffolds have not been studied throughly and are technically challenging. This study aimed to optimize nanocomposites of 3D PCL scaffolds to provide superior adhesion, proliferation and differentiation environment for BMSCs in this scenario. BMSCs were isolated and cultured in a novel 3D tissue culture poly(ε-caprolactone) (PCL) scaffold coated with poly-lysine, hydroxyapatite (HAp), collagen and HAp/collagen. Cell morphology was observed and BMSC biomarkers for osteogenesis, osteoblast differentiation and activation were analyzed. Scanning Electron Microscope (SEM) micrographs showed that coating materials were uniformly deposited on the surface of PCL scaffolds and BMSCs grew and aggregated to form clusters during 3D culture. Both mRNA and protein levels of the key players of osteogenesis and osteoblast differentiation and activation, including runt-related transcription factor 2 (Runx2), alkaline phosphates (ALP), osterix, osteocalcin, and RANKL, were significantly higher in BMSCs seeded in PCL scaffolds coated with HAp or HAp/collagen than those seeded in uncoated PCL scaffolds, whereas the expression levels were not significantly different in collagen or poly-lysine coated PCL scaffolds. In addition, poly-lysine, collagen, HAp/collagen, and HAp coated PCL scaffolds had significantly more viable cells than uncoated PCL scaffolds, especially scaffolds with HAp/collagen and collagen-alone coatings. That BMSCs in HAp or HAp/collagen PCL scaffolds had remarkably higher ALP activities than those in collagen-coated alone or uncoated PCL scaffolds indicating higher osteogenic differentiation levels of BMSCs in HAp or HAp/collagen PCL scaffolds. Moreover, morphological changes of BMSCs after four-week of 3D culture confirmed that BMSCs successfully differentiated into osteoblast with spread-out phenotype in HAp/collagen coated PCL scaffolds

  3. Clinical uses of radiosterilized freeze-dried human bone: its application in buccomaxillary surgery

    International Nuclear Information System (INIS)

    Wolfsohn, B.; Taramasso, F.; Godoy, J.; Wodowoz, O.; Saldias, M.; Silva, W.; Machin, D.; Sanchez, G.; Alvarez, I.

    2008-01-01

    Full text: The objective of this paper is to evaluate the uses of different human bone tissue allografts in bucomaxillary surgery between 2005 and 2007. Presentation of our experience using single freeze dried bone allografts and associated to bovine collagen membranes (commercial registered). Twenty patients were treated with cortical struts, cancellous chips, morsellized and morsellized demineralized bone. All the grafts were processed at the INDT multi tissue bank from cardiac arrest and brain death cadaveric donors. All the tissues were radiosterilized by Gamma radiation. Bone allografts were used: 1) to optimize bone support increasing maxillar or mandible bone before implant surgery. 2) in dehiscences and fissures during the implant surgery. 3) to stimulate bone regeneration in alveolar cavity, post-apicectomies, and cystectomies as well as for bone defects. The patients were periodically evaluated using standardized protocols. All the cases were successful showing clinic and radiologically osseointegration after 6 and 12 months. Results were evaluated considering surgical technique and patients bucomaxillary rehabilitation. Clinical uses of bone allografts confirm in our experience, as scientific literature outcomes shows, are useful in patients that refuses autografts ablation. (Author)

  4. Human pro. cap alpha. 1)(I) collagen: cDNA sequence for the C-propeptide domain

    Energy Technology Data Exchange (ETDEWEB)

    Maekelae, J K; Raassina, M; Virta, A; Vuorio, E

    1988-01-11

    The authors have previously constructed a cDNA clone pHCAL1, covering most of the C-terminal propeptide domain of human pro..cap alpha..1(I) collagen mRNA,by inserting a 678 bp EcoRI-XhoI fragment of cDNA into pBR322. Since the XhoI/SalI ligation prevented removal of the insert, they used the same strategy to obtain a similar clone in pUC8. RNA was isolated from fetal calvarial bones. The cDNA was digested with EcoRI and XhoI and fractionated on a 1 % agarose gel. Fragments of 650-700 bp were cloned in pUC8 at the polylinker site, which now permits easy removal of the insert. The new clone was named pHCAL1U since the RNA was isolated from another individual. The approach outlined is useful for studies on individual variation which is important to recognize when searching for disease-related mutations in type I collagen.

  5. Micromechanical properties and collagen composition of ruptured human achilles tendon

    DEFF Research Database (Denmark)

    Hansen, Philip; Kovanen, Vuokko; Hölmich, Per

    2013-01-01

    The Achilles tendon is one of the strongest tendons in the human body, and yet it frequently ruptures, which is a substantial clinical problem. However, the cause of ruptures remains elusive.......The Achilles tendon is one of the strongest tendons in the human body, and yet it frequently ruptures, which is a substantial clinical problem. However, the cause of ruptures remains elusive....

  6. A preliminary carbon and nitrogen isotopic investigation of bone collagen from skeletal remains recovered from a Pre-Columbian burial site, Matanzas Province, Cuba

    International Nuclear Information System (INIS)

    Buhay, W.M.; Chinique de Armas, Y.; Rodriguez Suárez, R.; Arredondo, C.; Smith, D.G.; Armstrong, S.D.; Roksandic, M.

    2013-01-01

    Highlights: ► Collagen isotope (carbon and nitrogen) based reconstruction of paleodiets. ► Human remains recovered from Canimar Abajo, Matanzas Province, Cuba. ► Individuals consumed marine resource diets supplemented with terrestrial plants. ► Trophic level and isotope shifts for breastfed and weaned infant/juveniles (I/J). ► I/J evidence of weaning through distinct δ 15 N enrichments and δ 13 C depletions. - Abstract: This preliminary study investigates the diet of a population of humans (n = 28) recovered from a shell-matrix site of Canimar Abajo on the Canimar River, Matanzas Province, Cuba. The site is characterized by two cemetery levels separated by a layer of occupation/ritual/midden activity that lasted 1.5 ka. Stable C (δ 13 C) and N (δ 15 N) isotope analysis of human bone collagen samples obtained from individuals (7 infant/juveniles, and 21 adults) from both cemetery levels was conducted in order to reconstruct the diet of these two populations, investigate the relative importance of marine vs. terrestrial resources, and reveal any sex- and age-related distinctions in their food sources. Initial indications suggest that individuals from both cemetery levels consumed diets that were marine resource intensive but also supplemented with varied additions of terrestrial (mostly plant) resources. This supplementation is particularly evident in the later cemetery population. Though there are no significant differences in diet according to sex, there is a trophic level and terrestrial-based shift for breastfed and weaning infant/juveniles. The infant/juveniles showed evidence of being weaned through distinct δ 15 N enrichments and δ 13 C depletions over adult females

  7. A multiscale analytical approach for bone remodeling simulations : linking scales from collagen to trabeculae

    NARCIS (Netherlands)

    Colloca, M.; Blanchard, R.; Hellmich, C.; Ito, K.; Rietbergen, van B.

    2014-01-01

    Bone is a dynamic and hierarchical porous material whose spatial and temporal mechanical properties can vary considerably due to differences in its microstructure and due to remodeling. Hence, a multiscale analytical approach, which combines bone structural information at multiple scales to the

  8. Flavonoids purified from parsley inhibit human blood platelet aggregation and adhesion to collagen under flow.

    Science.gov (United States)

    Gadi, Dounia; Bnouham, Mohamed; Aziz, Mohammed; Ziyyat, Abderrahim; Legssyer, Abdelkhaleq; Bruel, Arlette; Berrabah, Mohamed; Legrand, Chantal; Fauvel-Lafeve, Françoise; Mekhfi, Hassane

    2012-08-10

    Blood platelets are directly involved in both haemostatic and pathologic thrombotic processes, through their adhesion, secretion and aggregation. In this study, we investigated the effect of genins (aglycone flavonoids without sugar group) isolated from parsley (Petroselinum crispum) leaves in vitro on human platelet aggregation and adhesion to a collagen-coated surface under physiologic flow conditions. The aggregation and adhesion studies were monitored after pre-incubation of platelets with genins. Genins inhibited dose dependently aggregation induced by thrombin, ADP and collagen. The strongest effect was observed in collagen induced aggregation (IC50 = 0.08 ± 0.01 mg/ml). The HPLC identification of genins compounds revealed the presence of keampferol, apigenin and other not identified compounds. The aggregation tests showed that these compounds have anti-aggregating activity. In addition, adhesion of human platelets to collagen was greatly decreased (over 75 %) by genins (0.3 mg/ml). While the mechanism by which genins act is unclear, we suggest that these compounds may interfere with a multiple target step in the haemostasis process. These results show that genins isolated from parsley has a potent antiplatelet activity. It may be an important source of beneficial antiplatelet compounds that decrease thrombosis and cardiovascular diseases.

  9. Fibronectin- and collagen-mimetic ligands regulate bone marrow stromal cell chondrogenesis in three-dimensional hydrogels

    Directory of Open Access Journals (Sweden)

    JT Connelly

    2011-09-01

    Full Text Available Modification of tissue engineering scaffolds with bioactive molecules is a potential strategy for modulating cell behavior and guiding tissue regeneration. While adhesion to RGD peptides has been shown to inhibit in vitro chondrogenesis, the effects of extracellular matrix (ECM-mimetic ligands with complex secondary and tertiary structures are unknown. This study aimed to determine whether collagen- and fibronectin-mimetic ligands would retain biologic functionality in three-dimensional (3D hydrogels, whether different ECM-mimetic ligands differentially influence in vitro chondrogenesis, and if effects of ligands on differentiation depend on soluble biochemical stimuli. A linear RGD peptide, a recombinant fibronectin fragment containing the seven to ten Type III repeats (FnIII7-10 and a triple helical, collagen mimetic peptide with the GFOGER motif were covalently coupled to agarose gels using the sulfo-SANPAH crosslinker, and bone marrow stromal cells (BMSCs were cultured within the 3D hydrogels. The ligands retained biologic functionality within the agarose gels and promoted density-dependent BMSC spreading. Interactions with all adhesive ligands inhibited stimulation by chondrogenic factors of collagen Type II and aggrecan mRNA levels and deposition of sulfated glycosaminoglycans. In medium containing fetal bovine serum, interactions with the GFOGER peptide enhanced mRNA expression of the osteogenic gene osteocalcin whereas FnIII7-10 inhibited osteocalcin expression. In conclusion, modification of agarose hydrogels with ECM-mimetic ligands can influence the differentiation of BMSCs in a manner that depends strongly on the presence and nature of soluble biochemical stimuli.

  10. Collagen/hydroxyapatite scaffold enriched with polycaprolactone nanofibers, thrombocyte-rich solution and mesenchymal stem cells promotes regeneration in large bone defect in vivo

    Czech Academy of Sciences Publication Activity Database

    Prosecká, Eva; Rampichová, Michala; Litvinec, Andrej; Tonar, Z.; Králíčková, M.; Vojtová, L.; Kochová, P.; Plencner, Martin; Buzgo, Matej; Míčková, Andrea; Jančář, J.; Amler, Evžen

    2015-01-01

    Roč. 103, č. 2 (2015), s. 671-682 ISSN 1549-3296 Institutional support: RVO:68378041 Keywords : bone regeneration * mesenchymal stem cells * collagen/hydroxyapatite scaffold Subject RIV: EI - Biotechnology ; Bionics Impact factor: 3.263, year: 2015

  11. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    to initiate collagen fibrillogenesis when cultured in fixed-length fibrin gels. Fibroblasts were dissected from semitendinosus and gracilis tendons from healthy humans and cultured in 3D linear fibrin gels. The fibroblasts synthesized an extracellular matrix of parallel collagen fibrils that were aligned...

  12. Endogenous collagen influences differentiation of human multipotent mesenchymal stromal cells

    NARCIS (Netherlands)

    Fernandes, H.; Mentink, A.; Bank, R.; Stoop, R.; Blitterswijk, C. van; Boer, J. de

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  13. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, Hugo; Mentink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  14. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, H.A.M.; Mentink-Leusink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  15. Determination of composition and structure of spongy bone tissue in human head of femur by Raman spectral mapping.

    Science.gov (United States)

    Kozielski, M; Buchwald, T; Szybowicz, M; Błaszczak, Z; Piotrowski, A; Ciesielczyk, B

    2011-07-01

    Biomechanical properties of bone depend on the composition and organization of collagen fibers. In this study, Raman microspectroscopy was employed to determine the content of mineral and organic constituents and orientation of collagen fibers in spongy bone in the human head of femur at the microstructural level. Changes in composition and structure of trabecula were illustrated using Raman spectral mapping. The polarized Raman spectra permit separate analysis of local variations in orientation and composition. The ratios of ν₂PO₄³⁻/Amide III, ν₄PO₄³⁻/Amide III and ν₁CO₃²⁻/ν₂PO₄³⁻ are used to describe relative amounts of spongy bone components. The ν₁PO₄³⁻/Amide I ratio is quite susceptible to orientation effect and brings information on collagen fibers orientation. The results presented illustrate the versatility of the Raman method in the study of bone tissue. The study permits better understanding of bone physiology and evaluation of the biomechanical properties of bone.

  16. Bone Tissue Collagen Maturity and Mineral Content Increase With Sustained Hyperglycemia in the KK-Ay Murine Model of Type 2 Diabetes.

    Science.gov (United States)

    Hunt, Heather B; Pearl, Jared C; Diaz, David R; King, Karen B; Donnelly, Eve

    2018-05-01

    Type 2 diabetes mellitus (T2DM) increases fracture risk for a given bone mineral density (BMD), which suggests that T2DM changes bone tissue properties independently of bone mass. In this study, we assessed the effects of hyperglycemia on bone tissue compositional properties, enzymatic collagen crosslinks, and advanced glycation end-products (AGEs) in the KK-Ay murine model of T2DM using Fourier transform infrared (FTIR) imaging and high-performance liquid chromatography (HPLC). Compared to KK-aa littermate controls (n = 8), proximal femoral bone tissue of KK-Ay mice (n = 14) exhibited increased collagen maturity, increased mineral content, and less heterogeneous mineral properties. AGE accumulation assessed by the concentration of pentosidine, as well as the concentrations of the nonenzymatic crosslinks hydroxylysylpyridinoline (HP) and lysyl pyridinoline (LP), did not differ in the proximal femurs of KK-Ay mice compared to controls. The observed differences in tissue-level compositional properties in the KK-Ay mice are consistent with bone that is older and echo observations of reduced remodeling in T2DM. © 2017 American Society for Bone and Mineral Research. © 2017 American Society for Bone and Mineral Research.

  17. Pitfalls in comparing modern hair and fossil bone collagen C and N isotopic data to reconstruct ancient diets: a case study with cave bears (Ursus spelaeus).

    Science.gov (United States)

    Bocherens, Hervé; Grandal-d'Anglade, Aurora; Hobson, Keith A

    2014-01-01

    Stable isotope analyses provide one of the few means to evaluate diet of extinct taxa. However, interpreting isotope data from bone collagen of extinct animals based on isotopic patterns in different tissues of modern animal proxies is precarious. For example, three corrections are needed before making comparisons of recent hair and ancient bone collagen: calibration of carbon-13 variations in atmospheric CO2, different isotopic discrimination between diet-hair keratin and diet-bone collagen, and time averaging of bone collagen versus short-term record in hair keratin. Recently, Robu et al. [Isotopic evidence for dietary flexibility among European Late Pleistocene cave bears (Ursus spelaeus). Can J Zool. 2013;91:227-234] published an article comparing extant carbon (δ(13)C) and nitrogen (δ(15)N) stable isotopic data of European cave bear bone collagen with those of Yellowstone Park grizzly bear hair in order to test the prevailing assumption of a largely vegetarian diet among cave bears. The authors concluded that cave bears were carnivores. This work is unfortunately unfounded as the authors failed to consider the necessary corrections listed above. When these corrections are applied to the Romanian cave bears, these individuals can be then interpreted without involving consumption of high trophic-level food, and environmental changes are probably the reason for the unusual isotopic composition of these cave bears in comparison with other European cave bears, rather than a change of diet. We caution researchers to pay careful attention to these factors when interpreting feeding ecology of extinct fauna using stable isotope techniques.

  18. Etanercept Promotes Bone Formation via Suppression of Dickkopf-1 Expression in Rats with Collagen-Induced Arthritis

    Science.gov (United States)

    Tanida, Atsushi; Kishimoto, Yuji; Okano, Toru; Hagino, Hiroshi

    2013-01-01

    Background Various clinical reports suggest etanercept (ETN) has some efficacy in bone formation in rheumatoid arthritis (RA). To examine this effect, we investigated the gene expression of cytokines relevant to osteoblast/osteoclast differentiation, and evaluated histomorphometric findings in mature rats with collagen-induced arthritis (CIA). Methods Total RNA was extracted from knee joints with CIA after ETN or placebo administration. Subsequently, realtime-PCR was carried out to quantify the mRNAs encoding Wnt-1, Dickkopf-1 (DKK-1), receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegelin (OPG) and TNF (tumor necrosis factor)-alpha. In histomorphometric analysis, the infiltrating pannus volume and pannus surface, and the following items in contact with pannus surface were measured: osteoclast number, osteoid surface, osteoid volume and labeling surface. These were evaluated in the distal femur with CIA with or without ETN administration. Results TNF-alpha, RANKL and OPG mRNA expressions, linked to osteoclastogenesis, were not significantly different with or without ETN administration. ETN administration significantly increased Wnt-1 mRNA expression, the osteoblast promoter, and decreased DKK-1 mRNA expression, the Wnt signal inhibitor. In histomorphometric analysis, pannus volume, pannus surface and osteoclast number, parameters of bone destruction, were not significantly different among groups. Osteoid volume, osteoid surface and labeling surface, parameters of bone formation, increased significantly with ETN administration. Conclusion Our results suggest that ETN suppresses DDK-1 expression, and, as a result, Wnt expression is promoted and osteoblastogenesis becomes more active, independent of the regulation of osteoclast activity. Marked bone formation is attributed to the fact that ETN directly promotes osteoblastogenesis, not as a result of suppressing osteoclastogenesis. PMID:24031147

  19. Conservative treatment of bone tissue metabolic disorders among patients with vitamin D-dependent rickets type II with genetic abnormality of type I collagen formation

    Directory of Open Access Journals (Sweden)

    S.M. Martsyniak

    2017-08-01

    Full Text Available Background. The purpose of the article is to determine the effect of conservative therapy on genetically caused disorders of bone tissue metabolism in patients with vitamin D-dependent rickets type II and genetic abnormality of type I collagen formation (VDDR(COL1. Materials and methods. At the premises of consulting and outpatient department of SI “Institute of Traumatology and Orthopaedics of the NAMS of Ukraine”, 13 patients having VDDR type II and genetic damage of type I collagen formation were examined and treated. The medical treatment was conducted in four stages. The first stage included full examination of patients (calcium and phosphorus levels in the blood serum and their urinary excretion, as well as determination of calcidiol and calcitriol serum levels, indicators of parathyroid hormone and osteocalcin, and a marker of bone formation P1NP and osteoresorption b-CTx. At this stage, children were obligated to undergo a genetic test to detect changes (polymorphism in alleles of receptors to vitamin D and type I collagen. Besides genetic tests, examinations at the other stages were conducted in full. Results. The study has shown the following. The genetically caused abnormality of reception to vitamin D results into substantial accumulation of vitamin D active metabolite in the blood serum. When combined with gene­tic abnormality of type I collagen formation, it significantly affected bone formation and destruction processes that causes development of osteomalacia (parathormone — vitamin D — osteocalcin system. The comprehensive study of vitamin D metabolism and biochemical vitals of bone tissue in patients having VDDR (COL1 brought us to understanding of some issues related to pathogenesis and nature of osteomalacia and, in future, osteoporotic changes on different levels, ensured us to express these changes by corresponding indices in the biochemical research and, finally, to develop appropriate schemes for the treatment of

  20. Experimental model of bone response to collagenized xenografts of porcine origin (OsteoBiol® mp3): a radiological and histomorphometric study.

    Science.gov (United States)

    Calvo Guirado, Jose Luis; Ramírez Fernández, Maria Piedad; Negri, Bruno; Delgado Ruiz, Rafael Arcesio; Maté Sánchez de-Val, José Eduardo; Gómez-Moreno, Gerardo

    2013-02-01

    Adequate alveolar ridges are fundamental to successful rehabilitation with implants. There are diverse techniques for reconstructing atrophied ridges, of which bone substitute grafts is one possibility. The aim of this study was to carry out radiological and histomorphometric evaluations of bone response to collagenized porcine bone xenografts over a 4-month period following their insertion in rabbits' tibiae. Twenty New Zealand rabbits were used. Twenty collagenized porcine bone xenografts (Osteobiol® mp3, Tecnoss Dental s.r.l., Torino, Italy), in granulated form of 600 to 1,000 µm, were inserted in the proximal metaphyseal area of the animals' tibiae and 20 control areas were created. Following implantation, the animals were sacrificed in four groups of five, after 1, 2, 3, and 4 months, respectively. Radiological and histomorphometric studies were made. After 4 months, radiological images revealed bone defects with a decrease in graft volume and the complete repair of the osseous defect. No healed or residual bone alterations attributable to the presence of the implants were observed. Histomorphometric analysis at 4 months found mean values for newly formed bone, residual graft material, and non-mineralized connective tissue of 25.4 ± 1.8%, 36.37 ± 3.0%, and 38.22 ± 2.5%, respectively. There were no statistical differences in the length of cortical formation with collagenized porcine xenograft (98.9 ± 1.1%) compared with the control samples (99.1 ± 0.7%) at the end of the study period. The biomaterial used proved to be biocompatible, bioabsorbable, and osteoconductive and as such, a possible bone substitute that did not interfere with the bone's normal reparative processes. © 2011 Wiley Periodicals, Inc.

  1. Severely impaired bone material quality in Chihuahua zebrafish resembles classical dominant human osteogenesis imperfecta.

    Science.gov (United States)

    Fiedler, Imke A K; Schmidt, Felix N; Wölfel, Eva M; Plumeyer, Christine; Milovanovic, Petar; Gioia, Roberta; Tonelli, Francesca; Bale, Hrishikesh A; Jähn, Katharina; Besio, Roberta; Forlino, Antonella; Busse, Björn

    2018-04-17

    Excessive skeletal deformations and brittle fractures in the vast majority of patients suffering from osteogenesis imperfecta (OI) are a result of substantially reduced bone quality. Since the mechanical competence of bone is dependent on the tissue characteristics at small length scales, it is of crucial importance to assess how osteogenesis imperfecta manifests at the micro- and nanoscale of bone. In this context, the Chihuahua (Chi/ +) zebrafish, carrying a heterozygous glycine substitution in the α1 chain of collagen type I, has recently been proposed as suitable animal model of classical dominant OI, showing skeletal deformities, altered mineralization patterns and a smaller body size. This study assessed the bone quality properties of Chi/+ at multiple length scales using micro-computed tomography (micro-CT), histomorphometry, quantitative back-scattered electron imaging, Fourier transform infrared spectroscopy, nanoindentation and X-ray microscopy. At the skeletal level, Chi/+ display smaller body size, deformities and fracture calli in the ribs. Morphological changes at the whole bone level showed that the vertebrae in Chi/+ had a smaller size, smaller thickness and distorted shape. At the tissue level, Chi/+ displayed a higher degree of mineralization, lower collagen maturity, lower mineral maturity, altered osteoblast morphology, and lower osteocyte lacunar density compared to WT. The alterations in the cellular, compositional and structural properties of Chi/+ bones bear an explanation for the impaired local mechanical properties, which promote an increase in overall bone fragility in Chi/ +. The quantitative assessment of bone quality in Chi/+ thus further validates this mutant as an important model reflecting osseous characteristics associated with human classical dominant osteogenesis imperfecta. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  2. Osteoblast recruitment routes in human cancellous bone remodeling

    DEFF Research Database (Denmark)

    Kristensen, Helene Bjørg; Andersen, Thomas Levin; Marcussen, Niels

    2014-01-01

    It is commonly proposed that bone forming osteoblasts recruited during bone remodeling originate from bone marrow perivascular cells, bone remodeling compartment canopy cells, or bone lining cells. However, an assessment of osteoblast recruitment during adult human cancellous bone remodeling...... is lacking. We addressed this question by quantifying cell densities, cell proliferation, osteoblast differentiation markers, and capillaries in human iliac crest biopsy specimens. We found that recruitment occurs on both reversal and bone-forming surfaces, as shown by the cell density and osterix levels...

  3. Collagen Type I Improves the Differentiation of Human Embryonic Stem Cells towards Definitive Endoderm

    DEFF Research Database (Denmark)

    Rasmussen, Camilla Holzmann; Petersen, Dorthe Roenn; Møller, Jonas Bech

    2015-01-01

    Human embryonic stem cells have the ability to generate all cell types in the body and can potentially provide an unlimited source of cells for cell replacement therapy to treat degenerative diseases such as diabetes. Current differentiation protocols of human embryonic stem cells towards insulin...... and consistent differentiation of stem cells to definitive endoderm. The results shed light on the importance of extracellular matrix proteins for differentiation and also points to a cost effective and easy method to improve differentiation....... embryonic stem cells to the definitive endoderm lineage. The percentage of definitive endoderm cells after differentiation on collagen I and fibronectin was >85% and 65%, respectively. The cells on collagen I substrates displayed different morphology and gene expression during differentiation as assessed...

  4. Construction of human induced pluripotent stem cell-derived oriented bone matrix microstructure by using in vitro engineered anisotropic culture model.

    Science.gov (United States)

    Ozasa, Ryosuke; Matsugaki, Aira; Isobe, Yoshihiro; Saku, Taro; Yun, Hui-Suk; Nakano, Takayoshi

    2018-02-01

    Bone tissue has anisotropic microstructure based on collagen/biological apatite orientation, which plays essential roles in the mechanical and biological functions of bone. However, obtaining an appropriate anisotropic microstructure during the bone regeneration process remains a great challenging. A powerful strategy for the control of both differentiation and structural development of newly-formed bone is required in bone tissue engineering, in order to realize functional bone tissue regeneration. In this study, we developed a novel anisotropic culture model by combining human induced pluripotent stem cells (hiPSCs) and artificially-controlled oriented collagen scaffold. The oriented collagen scaffold allowed hiPSCs-derived osteoblast alignment and further construction of anisotropic bone matrix which mimics the bone tissue microstructure. To the best of our knowledge, this is the first report showing the construction of bone mimetic anisotropic bone matrix microstructure from hiPSCs. Moreover, we demonstrated for the first time that the hiPSCs-derived osteoblasts possess a high level of intact functionality to regulate cell alignment. © 2017 The Authors Journal of Biomedical Materials Research Part A Published by Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 360-369, 2018. © 2017 The Authors Journal of Biomedical Materials Research Part A Published by Wiley Periodicals, Inc.

  5. A New Bone Substitute Developed from 3D-Prints of Polylactide (PLA Loaded with Collagen I: An In Vitro Study

    Directory of Open Access Journals (Sweden)

    Ulrike Ritz

    2017-11-01

    Full Text Available Although a lot of research has been performed, large segmental bone defects caused by trauma, infection, bone tumors or revision surgeries still represent big challenges for trauma surgeons. New and innovative bone substitutes are needed. Three-dimensional (3D printing is a novel procedure to create 3D porous scaffolds that can be used for bone tissue engineering. In the present study, solid discs as well as porous cage-like 3D prints made of polylactide (PLA are coated or filled with collagen, respectively, and tested for biocompatibility and endotoxin contamination. Microscopic analyses as well as proliferation assays were performed using various cell types on PLA discs. Stromal-derived factor (SDF-1 release from cages filled with collagen was analyzed and the effect on endothelial cells tested. This study confirms the biocompatibility of PLA and demonstrates an endotoxin contamination clearly below the FDA (Food and Drug Administration limit. Cells of various cell types (osteoblasts, osteoblast-like cells, fibroblasts and endothelial cells grow, spread and proliferate on PLA-printed discs. PLA cages loaded with SDF-1 collagen display a steady SDF-1 release, support cell growth of endothelial cells and induce neo-vessel formation. These results demonstrate the potential for PLA scaffolds printed with an inexpensive desktop printer in medical applications, for example, in bone tissue engineering.

  6. A New Bone Substitute Developed from 3D-Prints of Polylactide (PLA) Loaded with Collagen I: An In Vitro Study.

    Science.gov (United States)

    Ritz, Ulrike; Gerke, Rebekka; Götz, Hermann; Stein, Stefan; Rommens, Pol Maria

    2017-11-29

    Although a lot of research has been performed, large segmental bone defects caused by trauma, infection, bone tumors or revision surgeries still represent big challenges for trauma surgeons. New and innovative bone substitutes are needed. Three-dimensional (3D) printing is a novel procedure to create 3D porous scaffolds that can be used for bone tissue engineering. In the present study, solid discs as well as porous cage-like 3D prints made of polylactide (PLA) are coated or filled with collagen, respectively, and tested for biocompatibility and endotoxin contamination. Microscopic analyses as well as proliferation assays were performed using various cell types on PLA discs. Stromal-derived factor (SDF-1) release from cages filled with collagen was analyzed and the effect on endothelial cells tested. This study confirms the biocompatibility of PLA and demonstrates an endotoxin contamination clearly below the FDA (Food and Drug Administration) limit. Cells of various cell types (osteoblasts, osteoblast-like cells, fibroblasts and endothelial cells) grow, spread and proliferate on PLA-printed discs. PLA cages loaded with SDF-1 collagen display a steady SDF-1 release, support cell growth of endothelial cells and induce neo-vessel formation. These results demonstrate the potential for PLA scaffolds printed with an inexpensive desktop printer in medical applications, for example, in bone tissue engineering.

  7. Bone morphogenetic protein-7 promotes chondrogenesis in human amniotic epithelial cells.

    Science.gov (United States)

    Zhou, Junjie; Yu, Guangrong; Cao, Chengfu; Pang, Jinhui; Chen, Xianqi

    2011-06-01

    Bone morphogenetic proteins (BMPs) play important roles at multiple stages of chondrogenesis. This study was undertaken to investigate the potential role of bone morphogenetic protein-7 (BMP-7) in the differentiation of chondrocytes using tissue engineering techniques. The impact of BMP-7 on human amniotic epithelial cells (hAECs) was tested. The hAECs were treated either with recombinant human BMP-7 cDNA or with transforming growth factor beta 1 (TGF-β1) as a positive control for three weeks in vitro. Cartilaginous differentiation and proliferation were assayed by quantitative RT-PCR, histology, and in situ hybridization. Our results were such that hAECs treated with either BMP-7 or TGF-β1 expressed cartilage markers (aggrecan, Sox9, CEP-68, and type II and X collagens) within three weeks. Compared with a control vector, BMP-7 induced a decrease in type I collagen expression, while the transcription of the cartilage-specific type II collagen remained stable. In induction experiments, BMP-7 transgenic hAECs exhibited the largest amount of matrix synthesis. In conclusion, these data indicate that BMP-7 plays an important role in inducing the production of cartilage by hAECs in vitro. Cartilage differentiation and matrix maturation can be promoted by BMPs in a cartilage engineering paradigm. These properties make BMPs promising tools in the engineering of cartilaginous joint bio-prostheses and as candidate biological agents or genes for cartilage stabilisation.

  8. Assessing the extent of bone degradation using glutamine deamidation in collagen.

    Science.gov (United States)

    Wilson, Julie; van Doorn, Nienke L; Collins, Matthew J

    2012-11-06

    Collagen peptides are analyzed using a low-cost, high-throughput method for assessing deamidation using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). For each chosen peptide, the theoretical distribution is calculated and the measured distribution for each sample compared with this to determine the extent of glutamine deamidation. The deamidation of glutamine (Q) to glutamic acid (E) results in a mass shift of +0.984 Da. Thus, from the resolution of our data, the second peak in the isotope distribution for a peptide containing one glutamine residue coincides with the first peak of the isotope distribution for the peptide in which the residue is deamidated. A genetic algorithm is used to determine the extent of deamidation that gives the best fit to the measured distribution. The method can be extended to peptides containing more than one glutamine residue. The extent of protein degradation assessed in this way could be used, for example, to assess the damage of collagen, and screen samples for radiocarbon dating and DNA analysis.

  9. Plastic compressed collagen as a novel carrier for expanded human corneal endothelial cells for transplantation.

    Directory of Open Access Journals (Sweden)

    Hannah J Levis

    Full Text Available Current treatments for reversible blindness caused by corneal endothelial cell failure involve replacing the failed endothelium with donor tissue using a one donor-one recipient strategy. Due to the increasing pressure of a worldwide donor cornea shortage there has been considerable interest in developing alternative strategies to treat endothelial disorders using expanded cell replacement therapy. Protocols have been developed which allow successful expansion of endothelial cells in vitro but this approach requires a supporting material that would allow easy transfer of cells to the recipient. We describe the first use of plastic compressed collagen as a highly effective, novel carrier for human corneal endothelial cells. A human corneal endothelial cell line and primary human corneal endothelial cells retained their characteristic cobblestone morphology and expression of tight junction protein ZO-1 and pump protein Na+/K+ ATPase α1 after culture on collagen constructs for up to 14 days. Additionally, ultrastructural analysis suggested a well-integrated endothelial layer with tightly opposed cells and apical microvilli. Plastic compressed collagen is a superior biomaterial in terms of its speed and ease of production and its ability to be manipulated in a clinically relevant manner without breakage. This method provides expanded endothelial cells with a substrate that could be suitable for transplantation allowing one donor cornea to potentially treat multiple patients.

  10. Analysis of bone mineral density of human bones for strength ...

    Indian Academy of Sciences (India)

    Different types of bone strength are required for various ... To statically analyse various methods to find BMD and related material ... bone study for research purpose. ..... and Dagoberto Vela Arvizo 2007 A qualitative stress analysis of a cross ...

  11. Genetics and biochemistry of collagen binding-triggered glandular differentiation in a human colon carcinoma cell line

    International Nuclear Information System (INIS)

    Pignatelli, M.; Bodmer, W.F.

    1988-01-01

    The authors have examined the interaction between collagen binding and epithelial differentiation by using a human colon carcinoma cell line (SW1222) that can differentiate structurally when grown in a three-dimensional collagen gel to form glandular structures. As much as 66% inhibition of glandular differentiation can be achieved by addition to the culture of a synthetic peptide containing the Arg-Gly-Asp-Thr (RGDT) sequence, which is a cell recognition site found in collagen. Arg-Gly-Asp-Thr also inhibited the cell attachment to collagen-coated plates. Chromosome 15 was found in all human-mouse hybrid clones that could differentiate in the collagen gel and bind collagen. Both binding to collagen and glandular differentiation of the hybrid cells were also inhibited by Arg-Gly-Asp-Thr as for the parent cell line SW1222. The ability of SW1222 cells to express the differentiated phenotype appears, therefore, to be determined by an Arg-Gly-Asp-directed collagen receptor on the cell surface that is controlled by a gene on chromosome 15

  12. Low-level laser therapy induces an upregulation of collagen gene expression during the initial process of bone healing: a microarray analysis

    Science.gov (United States)

    Tim, Carla Roberta; Bossini, Paulo Sérgio; Kido, Hueliton Wilian; Malavazi, Iran; von Zeska Kress, Marcia Regina; Carazzolle, Marcelo Falsarella; Rennó, Ana Cláudia; Parizotto, Nivaldo Antonio

    2016-08-01

    This study investigates the histological modifications produced by low level laser therapy (LLLT) on the first day of bone repair, as well as evaluates the LLLT effects on collagen expression on the site of a fracture. Twenty Wistar rats were distributed into a control group (CG) and a laser group (LG). Laser irradiation of Ga-Al-As laser 830 nm, 30 mW, 94 s, 2.8 J was performed in five sessions. Animals were euthanized on day 5 postsurgery. Histopathological analysis showed that LLLT was able to increase deposition of granulation tissue and newly formed bone at the site of the injury. In addition, picrosirius analysis showed that collagen fiber organization in the LG was enhanced compared to CG. Microarray analysis demonstrated that LLLT produced an upregulation type I collagen (COL-I). Immunohistochemical analysis revealed that the subjects that were treated presented a higher immunoexpression of COL-I. Our findings indicated that LLLT improves bone healing by producing a significant increase in the expression of collagen genes.

  13. New Altered Non-Fibrillar Collagens in Human Dilated Cardiomyopathy: Role in the Remodeling Process.

    Directory of Open Access Journals (Sweden)

    Carolina Gil-Cayuela

    Full Text Available In dilated cardiomyopathy (DCM, cardiac failure is accompanied by profound alterations of extracellular matrix associated with the progression of cardiac dilation and left ventricular (LV dysfunction. Recently, we reported alterations of non-fibrillar collagen expression in ischemic cardiomyopathy linked to fibrosis and cardiac remodeling. We suspect that expression changes in genes coding for non-fibrillar collagens may have a potential role in DCM development.This study sought to analyze changes in the expression profile of non-fibrillar collagen genes in patients with DCM and to examine relationships between cardiac remodeling parameters and the expression levels of these genes.Twenty-three human left ventricle tissue samples were obtained from DCM patients (n = 13 undergoing heart transplantation and control donors (n = 10 for RNA sequencing analysis. We found increased mRNA levels of six non-fibrillar collagen genes, such as COL4A5, COL9A1, COL21A1, and COL23A1 (P < 0.05 for all, not previously described in DCM. Protein levels of COL8A1 and COL16A1 (P < 0.05 for both, were correspondingly increased. We also identified TGF-β1 significantly upregulated and related to both COL8A1 and COL16A1. Interestingly, we found a significant relationship between LV mass index and the gene expression level of COL8A1 (r = 0.653, P < 0.05.In our research, we identified new non-fibrillar collagens with altered expression in DCM, being COL8A1 overexpression directly related to LV mass index, suggesting that they may be involved in the progression of cardiac dilation and remodeling.

  14. External fixation of femoral defects in athymic rats: Applications for human stem cell implantation and bone regeneration

    Directory of Open Access Journals (Sweden)

    Terasa Foo

    2013-01-01

    Full Text Available An appropriate animal model is critical for the research of stem/progenitor cell therapy and tissue engineering for bone regeneration in vivo. This study reports the design of an external fixator and its application to critical-sized femoral defects in athymic rats. The external fixator consists of clamps and screws that are readily available from hardware stores as well as Kirschner wires. A total of 35 rats underwent application of the external fixator with creation of a 6-mm bone defect in one femur of each animal. This model had been used in several separate studies, including implantation of collagen gel, umbilical cord blood mesenchymal stem cells, endothelial progenitor cells, or bone morphogenetic protein-2. One rat developed fracture at the proximal pin site and two rats developed deep tissue infection. Pin loosening was found in nine rats, but it only led to the failure of external fixation in two animals. In 8 to 10 weeks, various degrees of bone growth in the femoral defects were observed in different study groups, from full repair of the bone defect with bone morphogenetic protein-2 implantation to fibrous nonunion with collagen gel implantation. The external fixator used in these studies provided sufficient mechanical stability to the bone defects and had a comparable complication rate in athymic rats as in immunocompetent rats. The external fixator does not interfere with the natural environment of a bone defect. This model is particularly valuable for investigation of osteogenesis of human stem/progenitor cells in vivo.

  15. Effects of Spaceflight on Bone: The Rat as an Animal Model for Human Bone Loss

    Science.gov (United States)

    Halloran, B.; Weider, T.; Morey-Holton, E.

    1999-01-01

    The loss of weight bearing during spaceflight results in osteopenia in humans. Decrements in bone mineral reach 3-10% after as little as 75-184 days in space. Loss of bone mineral during flight decreases bone strength and increases fracture risk. The mechanisms responsible for, and the factors contributing to, the changes in bone induced by spaceflight are poorly understood. The rat has been widely used as an animal model for human bone loss during spaceflight. Despite its potential usefulness, the results of bone studies performed in the rat in space have been inconsistent. In some flights bone formation is decreased and cancellous bone volume reduced, while in others no significant changes in bone occur. In June of 1996 Drs. T. Wronski, S. Miller and myself participated in a flight experiment (STS 78) to examine the effects of glucocorticoids on bone during weightlessness. Technically the 17 day flight experiment was flawless. The results, however, were surprising. Cancellous bone volume and osteoblast surface in the proximal tibial metaphysis were the same in flight and ground-based control rats. Normal levels of cancellous bone mass and bone formation were also detected in the lumbar vertebrae and femoral neck of flight rats. Furthermore, periosteal bone formation rate was found to be identical in flight and ground-based control rats. Spaceflight had little or no effect on bone metabolism! These results prompted us to carefully review the changes in bone observed in, and the flight conditions of previous spaceflight missions.

  16. Growth hormone (GH) effects on bone and collagen turnover in healthy adults and its potential as a marker of GH abuse in sports: a double blind, placebo-controlled study. The GH-2000 Study Group.

    Science.gov (United States)

    Longobardi, S; Keay, N; Ehrnborg, C; Cittadini, A; Rosén, T; Dall, R; Boroujerdi, M A; Bassett, E E; Healy, M L; Pentecost, C; Wallace, J D; Powrie, J; Jørgensen, J O; Saccà, L

    2000-04-01

    The effects of GH on bone remodeling in healthy adults have not been systematically investigated. An analysis of these effects might provide insights into GH physiology and might yield data useful for the detection of GH doping in sports. The aim of this study was to evaluate the effects of GH administration on biochemical markers of bone and collagen turnover in healthy volunteers. Ninety-nine healthy volunteers of both sexes were enrolled in a multicenter, randomized, double blind, placebo-controlled study and assigned to receive either placebo (40 subjects) or recombinant human GH (0.1 IU/kg day in 29 subjects and 0.2 IU/kg x day in 30 subjects). The treatment duration was 28 days, followed by a 56-day wash-out period. The biochemical markers evaluated were the bone formation markers osteocalcin and C-terminal propeptide of type I procollagen, the resorption marker type I collagen telopeptide, and the soft tissue marker procollagen type III. All variables increased on days 21 and 28 in the two active treatment groups vs. levels in both the baseline (P < 0.01) and placebo (P < 0.01) groups. The increment was more pronounced in the 0.2 IU/kg-day group and remained significant on day 84 for procollagen type III (from 0.53 +/- 0.13 to 0.61 +/- 0.14 kU/L; P < 0.02) and osteocalcin (from 12.2 + 2.9 to 14.6 +/- 3.6 UG/L; P < 0.02), whereas levels of C-terminal propeptide of type I procollagen and type I collagen telopeptide declined after day 42 and were no longer significantly above baseline on day 84 (from 3.9 +/- 1.2 to 5.1 +/-1.5 microg/L and from 174 +/- 60 to 173 +/- 53 microg/L, respectively). Gender-related differences were observed in the study; females were less responsive than males to GH administration with respect to procollagen type III and type I collagen telopeptide (P < 0.001). In conclusion, exogenous GH administration affects the biochemical parameters of bone and collagen turnover in a dose- and gender-dependent manner. As GH-induced modifications

  17. Mechanical and mineral properties of osteogenesis imperfecta human bones at the tissue level.

    Science.gov (United States)

    Imbert, Laurianne; Aurégan, Jean-Charles; Pernelle, Kélig; Hoc, Thierry

    2014-08-01

    Osteogenesis imperfecta (OI) is a genetic disorder characterized by an increase in bone fragility on the macroscopic scale, but few data are available to describe the mechanisms involved on the tissue scale and the possible correlations between these scales. To better understand the effects of OI on the properties of human bone, we studied the mechanical and chemical properties of eight bone samples from children suffering from OI and compared them to the properties of three controls. High-resolution computed tomography, nanoindentation and Raman microspectroscopy were used to assess those properties. A higher tissue mineral density was found for OI bone (1.131 gHA/cm3 vs. 1.032 gHA/cm3, p=0.032), along with a lower Young's modulus (17.6 GPa vs. 20.5 GPa, p=0.024). Obviously, the mutation-induced collagen defects alter the collagen matrix, thereby affecting the mineralization. Raman spectroscopy showed that the mineral-to-matrix ratio was higher in the OI samples, while the crystallinity was lower, suggesting that the mineral crystals were smaller but more abundant in the case of OI. This change in crystal size, distribution and composition contributes to the observed decrease in mechanical strength. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. Localized ridge defect augmentation using human pericardium membrane and demineralized bone matrix.

    Science.gov (United States)

    Vidyadharan, Arun Kumar; Ravindran, Anjana

    2014-01-01

    Patient wanted to restore her lost teeth with implants in the lower left first molar and second premolar region. Cone beam computerized tomography (CBCT) revealed inadequate bone width and height around future implant sites. The extraction socket of second premolar area revealed inadequate socket healing with sparse bone fill after 4 months of extraction. To evaluate the clinical feasibility of using a collagen physical resorbable barrier made of human pericardium (HP) to augment localized alveolar ridge defects for the subsequent placement of dental implants. Ridge augmentation was done in the compromised area using Puros® demineralized bone matrix (DBM) Putty with chips and an HP allograft membrane. Horizontal (width) and vertical hard tissue measurements with CBCT were recorded on the day of ridge augmentation surgery, 4 month and 7 months follow-up. Intra oral periapical taken 1 year after implant installation showed minimal crestal bone loss. Bone volume achieved through guided bone regeneration was a gain of 4.8 mm horizontally (width) and 6.8 mm vertically in the deficient ridge within a period of 7 months following the procedure. The results suggested that HP Allograft membrane may be a suitable component for augmentation of localized alveolar ridge defects in conjunction with DBM with bone chips.

  19. Nicotine promotes proliferation and collagen synthesis of chondrocytes isolated from normal human and osteoarthritis patients.

    Science.gov (United States)

    Ying, Xiaozhou; Cheng, Shaowen; Shen, Yue; Cheng, Xiaojie; An Rompis, Ferdinand; Wang, Wei; Lin, Zhongqin; Chen, Qingyu; Zhang, Wei; Kou, Dongquan; Peng, Lei; Tian, Xin Qiao; Lu, Chuan Zhu

    2012-01-01

    The aims of the study were to show the direct effect of nicotine with different concentrations (0, 25, 50, and 100 ng/ml) on chondrocytes isolated from normal human and osteoarthritis patients, respectively. Microscopic observation was performed during the culture with an inverted microscope. Methyl thiazolyl tetrazolium (MTT) assay method was adopted to observe the influence of nicotine on the proliferation of chondrocytes, and real-time PCR and ELISA were used to assay the mRNA and protein expression of type II collagen and aggrecan, respectively. We discovered that the OA chondrocytes were similar to fibroblasts in shape and grow slower than normal chondrocytes. The proliferation of the two kinds of chondrocytes was increased in a concentration-dependent manner and in a time-dependent manner (P<0.05). Also, we found that the mRNA level of type II collagen were upregulated under 25-100 ng/ml nicotine doses both in the two kinds of chondrocytes compared with control. The expression of protein levels of type II collagen were synthesized in line with the increase in mRNA. No effect was observed on aggrecan synthesis with any nicotine dose. We concluded that nicotine has the same effect on both chondrocytes, obtained either from osteoarthritis patients or from normal human, and the positive effect of smoking in OA may relate to the alteration in metabolism of chondrocytes.

  20. Normal human bone marrow and its variations in MRI

    International Nuclear Information System (INIS)

    Vahlensieck, M.; Schmidt, H.M.

    2000-01-01

    Physiology and age dependant changes of human bone marrow are described. The resulting normal distribution patterns of active and inactive bone marrow including the various contrasts on different MR-sequences are discussed. (orig.) [de

  1. Stable carbon isotope variability of bone collagen and hair within a modern population of red kangaroos (Macropus rufus) in south western Queensland: some implications for palaeoecological research

    International Nuclear Information System (INIS)

    Witt, G.B.

    1997-01-01

    Full text: Before any palaeo-reconstruction work can be attempted using stable isotope analysis of macropod remains it will be necessary to determine the nature of natural variability within contemporary populations. This research indicates that δ 13 C of bone collagen is strongly related to age. Furthermore, bone collagen δ 13 C not at equilibrium with dietary δ 13 C, as indicated by analysis of hair, until animals are several years old. These preliminary data suggest that in younger macropods most carbon in bone collagen has been derived via the mother's milk which may have undergone fractionation. These findings have significant implications for any palaeoecological research using bone or tooth. Teeth of macropods erupt from the rear of the jaw and move forward in molar progression. Since the rate of eruption is variable, and many of the forward molars are well formed while the joey is still at the pouch, teeth formed early in the life of a macropod may be isotopically distinct from those that develop later. This hypothesis is currently under investigation

  2. Two-layer membranes of calcium phosphate/collagen/PLGA nanofibres: in vitro biomineralisation and osteogenic differentiation of human mesenchymal stem cells

    Science.gov (United States)

    Hild, Nora; Schneider, Oliver D.; Mohn, Dirk; Luechinger, Norman A.; Koehler, Fabian M.; Hofmann, Sandra; Vetsch, Jolanda R.; Thimm, Benjamin W.; Müller, Ralph; Stark, Wendelin J.

    2011-02-01

    The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a-CaP/Col/PLGA compositions were synthesised by electrospinning to mimic the actual composition of bone tissue. Immersion in simulated body fluid and in cell culture medium resulted in the deposition of a hydroxyapatite layer. Incubation of hMSC for 4 weeks allowed for assessment of the proliferation and osteogenic differentiation of the cells on both sides of the double membrane. Confocal laser scanning microscopy was used to observe the proper adhesion of the cells. Calcium and collagen content was proven by Alizarin red S and Sirius red assays. Acute cytotoxic effects of the nanoparticles or the chemicals used in the scaffold preparation could be excluded based on viability assays (alamarBlue and alkaline phosphatase activity). The findings suggest possible application of such double membranes is in treatment of bone defects with complex geometries as wound dressing material.The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a

  3. Articular cartilage repair with recombinant human type II collagen/polylactide scaffold in a preliminary porcine study.

    Science.gov (United States)

    Muhonen, Virpi; Salonius, Eve; Haaparanta, Anne-Marie; Järvinen, Elina; Paatela, Teemu; Meller, Anna; Hannula, Markus; Björkman, Mimmi; Pyhältö, Tuomo; Ellä, Ville; Vasara, Anna; Töyräs, Juha; Kellomäki, Minna; Kiviranta, Ilkka

    2016-05-01

    The purpose of this study was to investigate the potential of a novel recombinant human type II collagen/polylactide scaffold (rhCo-PLA) in the repair of full-thickness cartilage lesions with autologous chondrocyte implantation technique (ACI). The forming repair tissue was compared to spontaneous healing (spontaneous) and repair with a commercial porcine type I/III collagen membrane (pCo). Domestic pigs (4-month-old, n = 20) were randomized into three study groups and a circular full-thickness chondral lesion with a diameter of 8 mm was created in the right medial femoral condyle. After 3 weeks, the chondral lesions were repaired with either rhCo-PLA or pCo together with autologous chondrocytes, or the lesion was only debrided and left untreated for spontaneous repair. The repair tissue was evaluated 4 months after the second operation. Hyaline cartilage formed most frequently in the rhCo-PLA treatment group. Biomechanically, there was a trend that both treatment groups resulted in better repair tissue than spontaneous healing. Adverse subchondral bone reactions developed less frequently in the spontaneous group (40%) and the rhCo-PLA treated group (50%) than in the pCo control group (100%). However, no statistically significant differences were found between the groups. The novel rhCo-PLA biomaterial showed promising results in this proof-of-concept study, but further studies will be needed in order to determine its effectiveness in articular cartilage repair. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:745-753, 2016. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  4. Aging, human immunodeficiency virus, and bone health

    Directory of Open Access Journals (Sweden)

    Kim C Mansky

    2010-09-01

    Full Text Available Kim C ManskyDivision of Orthodontics, Department of Developmental and Surgical Sciences, School of Dentistry, University of Minnesota, Minneapolis, MN, USAAbstract: Highly active antiretroviral therapy (HAART has had a profound impact on improving the long-term prognosis for individuals infected with human immunodeficiency virus (HIV. HAART has been available for close to two decades, and now a significant number of patients with access to HAART are over the age of 50 years. Many clinical studies have indicated that HIV infection, as well as components of HAART, can increase the risk in these individuals to a variety of noninfectious complications, including a risk to bone health. There is a significant need for detailed mechanistic analysis of the aging, HIV-infected population regarding the risk of HIV infection and therapy in order to maintain bone health. Insights from basic mechanistic studies will help to shed light on the role of HIV infection and the components of HAART that impact bone health, and will help in identifying preventative countermeasures, particularly for individuals 50 years of age and older.Keywords: osteopenia, osteomalacia, osteoporosis, bisphosphonates, tenofovir, osteoimmunology

  5. Three-dimensional structure of human lamellar bone: the presence of two different materials and new insights into the hierarchical organization.

    Science.gov (United States)

    Reznikov, Natalie; Shahar, Ron; Weiner, Steve

    2014-02-01

    Lamellar bone is the most common bone type in humans. The predominant components of individual lamellae are plywood-like arrays of mineralized collagen fibrils aligned in different directions. Using a dual-beam electron microscope and the Serial Surface View (SSV) method we previously identified a small, but significantly different layer in rat lamellar bone, namely a disordered layer with collagen fibrils showing little or no preferred orientation. Here we present a 3D structural analysis of 12 SSV volumes (25 complete lamellae) from femora of 3 differently aged human individuals. We identify the ordered and disordered motifs in human bone as in the rat, with several significant differences. The ordered motif shows two major preferred orientations, perpendicular to the long axis of the bone, and aligned within 10-20° of the long axis, as well as fanning arrays. At a higher organizational level, arrays of ordered collagen fibrils are organized into 'rods' around 2 to 3μm in diameter, and the long axes of these 'rods' are parallel to the lamellar boundaries. Human bone also contains a disordered component that envelopes the rods and fills in the spaces between them. The disordered motif is especially well-defined between adjacent layers of rods. The disordered motif and its interfibrillar substance stain heavily with osmium tetroxide and Alcian blue indicating the presence of another organic component in addition to collagen. The canalicular network is confined to the disordered material, along with voids and individual collagen fibrils, some of which are also aligned more or less perpendicular to the lamellar boundaries. The organization of the ordered fibril arrays into rods enveloped in the continuous disordered structure was not observed in rat lamellar bone. We thus conclude that human lamellar bone is comprised of two distinct materials, an ordered material and a disordered material, and contains an additional hierarchical level of organization composed of

  6. Collagen Orientation and Crystallite Size in Human Dentin: A Small Angle X-ray Scattering Study

    Energy Technology Data Exchange (ETDEWEB)

    Pople, John A

    2001-03-29

    The mechanical properties of dentin are largely determined by the intertubular dentin matrix, which is a complex composite of type I collagen fibers and a carbonate-rich apatite mineral phase. The authors perform a small angle x-ray scattering (SAXS) study on fully mineralized human dentin to quantify this fiber/mineral composite architecture from the nanoscopic through continuum length scales. The SAXS results were consistent with nucleation and growth of the apatite phase within periodic gaps in the collagen fibers. These mineralized fibers were perpendicular to the dentinal tubules and parallel with the mineralization growth front. Within the plane of the mineralization front, the mineralized collagen fibers were isotropic near the pulp, but became mildly anisotropic in the mid-dentin. Analysis of the data also indicated that near the pulp the mineral crystallites were approximately needle-like, and progressed to a more plate-like shape near the dentino-enamel junction. The thickness of these crystallites, {approx} 5 nm, did not vary significantly with position in the tooth. These results were considered within the context of dentinogenesis and maturation.

  7. Long-term Culture of Human iPS Cell-derived Telencephalic Neuron Aggregates on Collagen Gel.

    Science.gov (United States)

    Oyama, Hiroshi; Takahashi, Koji; Tanaka, Yoshikazu; Takemoto, Hiroshi; Haga, Hisashi

    2018-01-01

    It takes several months to form the 3-dimensional morphology of the human embryonic brain. Therefore, establishing a long-term culture method for neuronal tissues derived from human induced pluripotent stem (iPS) cells is very important for studying human brain development. However, it is difficult to keep primary neurons alive for more than 3 weeks in culture. Moreover, long-term adherent culture to maintain the morphology of telencephalic neuron aggregates induced from human iPS cells is also difficult. Although collagen gel has been widely used to support long-term culture of cells, it is not clear whether human iPS cell-derived neuron aggregates can be cultured for long periods on this substrate. In the present study, we differentiated human iPS cells to telencephalic neuron aggregates and examined long-term culture of these aggregates on collagen gel. The results indicated that these aggregates could be cultured for over 3 months by adhering tightly onto collagen gel. Furthermore, telencephalic neuronal precursors within these aggregates matured over time and formed layered structures. Thus, long-term culture of telencephalic neuron aggregates derived from human iPS cells on collagen gel would be useful for studying human cerebral cortex development.Key words: Induced pluripotent stem cell, forebrain neuron, collagen gel, long-term culture.

  8. Comparison of Animal Discs Used in Disc Research to Human Lumbar Disc: Torsion Mechanics and Collagen Content

    Science.gov (United States)

    Showalter, Brent L.; Beckstein, Jesse C.; Martin, John T.; Beattie, Elizabeth E.; Orías, Alejandro A. Espinoza; Schaer, Thomas P.; Vresilovic, Edward J.; Elliott, Dawn M.

    2012-01-01

    Study Design Experimental measurement and normalization of in vitro disc torsion mechanics and collagen content for several animal species used in intervertebral disc research and comparing these to the human disc. Objective To aid in the selection of appropriate animal models for disc research by measuring torsional mechanical properties and collagen content. Summary of Background Data There is lack of data and variability in testing protocols for comparing animal and human disc torsion mechanics and collagen content. Methods Intervertebral disc torsion mechanics were measured and normalized by disc height and polar moment of inertia for 11 disc types in 8 mammalian species: the calf, pig, baboon, goat, sheep, rabbit, rat, and mouse lumbar, and cow, rat, and mouse caudal. Collagen content was measured and normalized by dry weight for the same discs except the rat and mouse. Collagen fiber stretch in torsion was calculated using an analytical model. Results Measured torsion parameters varied by several orders of magnitude across the different species. After geometric normalization, only the sheep and pig discs were statistically different from human. Fiber stretch was found to be highly dependent on the assumed initial fiber angle. The collagen content of the discs was similar, especially in the outer annulus where only the calf and goat discs were statistically different from human. Disc collagen content did not correlate with torsion mechanics. Conclusion Disc torsion mechanics are comparable to human lumbar discs in 9 of 11 disc types after normalization by geometry. The normalized torsion mechanics and collagen content of the multiple animal discs presented is useful for selecting and interpreting results for animal models of the disc. Structural composition of the disc, such as initial fiber angle, may explain the differences that were noted between species after geometric normalization. PMID:22333953

  9. Characterization of the effects of x-ray irradiation on the hierarchical structure and mechanical properties of human cortical bone

    Energy Technology Data Exchange (ETDEWEB)

    Barth, Holly; Zimmermann, Elizabeth; Schaible, Eric; Tang, Simon; Alliston, Tamara; Ritchie, Robert

    2011-08-19

    Bone comprises a complex structure of primarily collagen, hydroxyapatite and water, where each hierarchical structural level contributes to its strength, ductility and toughness. These properties, however, are degraded by irradiation, arising from medical therapy or bone-allograft sterilization. We provide here a mechanistic framework for how irradiation affects the nature and properties of human cortical bone over a range of characteristic (nano to macro) length-scales, following x-­ray exposures up to 630 kGy. Macroscopically, bone strength, ductility and fracture resistance are seen to be progressively degraded with increasing irradiation levels. At the micron-­scale, fracture properties, evaluated using in-situ scanning electron microscopy and synchrotron x-ray computed micro-tomography, provide mechanistic information on how cracks interact with the bone-matrix structure. At sub-micron scales, strength properties are evaluated with in-situ tensile tests in the synchrotron using small-/wide-angle x-ray scattering/diffraction, where strains are simultaneously measured in the macroscopic tissue, collagen fibrils and mineral. Compared to healthy bone, results show that the fibrillar strain is decreased by ~40% following 70 kGy exposures, consistent with significant stiffening and degradation of the collagen. We attribute the irradiation-­induced deterioration in mechanical properties to mechanisms at multiple length-scales, including changes in crack paths at micron-­scales, loss of plasticity from suppressed fibrillar sliding at sub-­micron scales, and the loss and damage of collagen at the nano-­scales, the latter being assessed using Raman and Fourier-Transform-Infrared spectroscopy and a fluorometric assay.

  10. Analysis of bone mineral density of human bones for strength ...

    Indian Academy of Sciences (India)

    The bone density (BMD) is a medical term normally referring to the amount of mineral matter per square centimetre of bones. Twenty-five patients (18 female and 7 male patients with a mean age of 71.3 years) undergoing both lumbar spine DXA scans and computed tomography imaging were evaluated to determine if HU ...

  11. Offsets of atmospheric radiocarbon concentration from IntCal09 in East Asia appeared in human bones excavated at Imdangdong tombs, Jingshan, Korea

    International Nuclear Information System (INIS)

    Sakamoto, Minoru; Takada, Kanta; Lee, Chang-hee; Fujio, Shin'ichiro; Woo, Byoung-cheul; Park, Seung-kyu

    2013-01-01

    Radiocarbon dating of human bones excavated from Imdangdong toms, Korea, was carried out. The preservation of bone collagen was poor, and even though the measurement was successful, their calibrated ages were not in good agreement with archaeological observations as well. However, in comparing to the radiocarbon offsets appeared in Japanese tree rings, the calibration age shifted to the proper age. This might show that the regional effects on calibration curve were coincident between Japanese archipelagos and Korean peninsula. (author)

  12. Synthesis and characterization of hyaluronic acid/human-like collagen hydrogels

    International Nuclear Information System (INIS)

    Zhang, Jingjing; Ma, Xiaoxuan; Fan, Daidi; Zhu, Chenhui; Deng, Jianjun; Hui, Junfeng; Ma, Pei

    2014-01-01

    Injectable hydrogel plays an important role in soft tissue filling and repair. We report an injectable hydrogel based on hyaluronic acid (HA) and human-like collagen (HLC), both with favorable biocompatibility and biodegradability. These two types of biomacromolecules were crosslinked with 1,4-butanediol diglycidyl ether to form a three-dimensional network. The redundant crosslinker was removed by dialysis and distillation. An HA-based hydrogel prepared by the same method was used as a control. The cytocompatibility was studied with a Cell Counting Kit-8 (CCK-8) test. Carbazole colorimetry was used to analyze the in vitro degradation rate. The histocompatibility was evaluated by hematoxylin and eosin (H and E) staining analysis and immunohistochemical analysis. The CCK-8 assay demonstrated that the HA/HLC hydrogel was less cytotoxic than the HA-based hydrogel and could promote baby hamster kidney cell (BHK) proliferation. The cell adhesion indicated that BHK could grow well on the surface of the materials and maintain good cell viability. The in vitro degradation test showed that the HA/HLC hydrogel had a longer degradation time and an excellent antienzyme ability. In vivo injection showed that there was little inflammatory response to HA/HLC after 1, 2, and 4 weeks. Therefore, the HA/HLC hydrogel is a promising biomaterial for soft tissue filling and repair. - Highlights: • Human-like collagen was used with hyaluronic acid to prepare soft tissue filling meterials. • 1,4-Butanediol diglycidyl ether (BDDE) was introduced to treat the hydrogels. • The addition of human-like collagen could improve the biological properties of hydrogels

  13. Synthesis and characterization of hyaluronic acid/human-like collagen hydrogels

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Jingjing; Ma, Xiaoxuan, E-mail: xiaoxuanma@163.com; Fan, Daidi, E-mail: fandaidi@nwu.edu.cn; Zhu, Chenhui; Deng, Jianjun; Hui, Junfeng; Ma, Pei

    2014-10-01

    Injectable hydrogel plays an important role in soft tissue filling and repair. We report an injectable hydrogel based on hyaluronic acid (HA) and human-like collagen (HLC), both with favorable biocompatibility and biodegradability. These two types of biomacromolecules were crosslinked with 1,4-butanediol diglycidyl ether to form a three-dimensional network. The redundant crosslinker was removed by dialysis and distillation. An HA-based hydrogel prepared by the same method was used as a control. The cytocompatibility was studied with a Cell Counting Kit-8 (CCK-8) test. Carbazole colorimetry was used to analyze the in vitro degradation rate. The histocompatibility was evaluated by hematoxylin and eosin (H and E) staining analysis and immunohistochemical analysis. The CCK-8 assay demonstrated that the HA/HLC hydrogel was less cytotoxic than the HA-based hydrogel and could promote baby hamster kidney cell (BHK) proliferation. The cell adhesion indicated that BHK could grow well on the surface of the materials and maintain good cell viability. The in vitro degradation test showed that the HA/HLC hydrogel had a longer degradation time and an excellent antienzyme ability. In vivo injection showed that there was little inflammatory response to HA/HLC after 1, 2, and 4 weeks. Therefore, the HA/HLC hydrogel is a promising biomaterial for soft tissue filling and repair. - Highlights: • Human-like collagen was used with hyaluronic acid to prepare soft tissue filling meterials. • 1,4-Butanediol diglycidyl ether (BDDE) was introduced to treat the hydrogels. • The addition of human-like collagen could improve the biological properties of hydrogels.

  14. L-arginine mediated renaturation enhances yield of human, α6 Type IV collagen non-collagenous domain from bacterial inclusion bodies.

    Science.gov (United States)

    Gunda, Venugopal; Boosani, Chandra Shekhar; Verma, Raj Kumar; Guda, Chittibabu; Sudhakar, Yakkanti Akul

    2012-10-01

    The anti-angiogenic, carboxy terminal non-collagenous domain (NC1) derived from human Collagen type IV alpha 6 chain, [α6(IV)NC1] or hexastatin, was earlier obtained using different recombinant methods of expression in bacterial systems. However, the effect of L-arginine mediated renaturation in enhancing the relative yields of this protein from bacterial inclusion bodies has not been evaluated. In the present study, direct stirring and on-column renaturation methods using L-arginine and different size exclusion chromatography matrices were applied for enhancing the solubility in purifying the recombinant α6(IV)NC1 from bacterial inclusion bodies. This methodology enabled purification of higher quantities of soluble protein from inclusion bodies, which inhibited endothelial cell proliferation, migration and tube formation. Thus, the scope for L-arginine mediated renaturation in obtaining higher yields of soluble, biologically active NC1 domain from bacterial inclusion bodies was evaluated.

  15. A murine model of human myeloma bone disease

    NARCIS (Netherlands)

    Garrett, I.R.; Dallas, S.; Radl, J.; Mundy, G.R.

    1997-01-01

    Myeloma causes a devastating and unique form of osteolytic bone disease. Although osteoclast activation is responsible for bone destruction, the precise mechanisms by which myeloma cells increase osteoclast activity have not been defined. An animal model of human myeloma bone disease mould help in

  16. Type I collagen gene suppresses tumor growth and invasion of malignant human glioma cells

    Directory of Open Access Journals (Sweden)

    Miyata Teruo

    2007-06-01

    Full Text Available Abstract Background Invasion is a hallmark of a malignant tumor, such as a glioma, and the progression is followed by the interaction of tumor cells with an extracellular matrix (ECM. This study examined the role of type I collagen in the invasion of the malignant human glioma cell line T98G by the introduction of the human collagen type I α1 (HCOL1A1 gene. Results The cells overexpressing HCOL1A1 were in a cluster, whereas the control cells were scattered. Overexpression of HCOL1A1 significantly suppressed the motility and invasion of the tumor cells. The glioma cell growth was markedly inhibited in vitro and in vivo by the overexpression of HCOL1A1; in particular, tumorigenicity completely regressed in nude mice. Furthermore, the HCOL1A1 gene induced apoptosis in glioma cells. Conclusion These results indicate that HCOL1A1 have a suppressive biological function in glioma progression and that the introduction of HCOL1A1 provides the basis of a novel therapeutic approach for the treatment of malignant human glioma.

  17. Introduction of the human proα1(I) collagen gene into proα1(I)-deficient Mov-13 mouse cells leads to formation of functional mouse-human hybrid type I collagen

    International Nuclear Information System (INIS)

    Schnieke, A.; Dziadek, M.; Bateman, J.; Mascara, T.; Harbers, K.; Gelinas, R.; Jaenisch, R.

    1987-01-01

    The Mov-13 mouse strain carries a retroviral insertion in the proα1(I) collagen gene that prevents transcription of the gene. Cell lines derived from homozygous embryos do not express type I collagen although normal amounts of proα2 mRNA are synthesized. The authors have introduced genomic clones of either the human or mouse proα1(I) collagen gene into homozygous cell lines to assess whether the human or mouse proα1(I) chains can associate with the endogenous mouse proα2(I) chain to form stable type I collagen. The human gene under control of the simian virus 40 promoter was efficiently transcribed in the transfected cells. Protein analyses revealed that stable heterotrimers consisting of two human α1 chains and one mouse α2 chain were formed and that type I collagen was secreted by the transfected cells at normal rates. However, the electrophoretic migration of both α1(I) and α2(I) chains in the human-mouse hybrid molecules were retarded, compared to the α(I) chains in control mouse cells. Inhibition of the posttranslational hydroxylation of lysine and proline resulted in comigration of human and mouse α1 and α2 chains, suggesting that increased posttranslational modification caused the altered electrophoretic migration in the human-mouse hybrid molecules. Amino acid sequence differences between the mouse and human α chains may interfere with the normal rate of helix formation and increase the degree of posttranslational modifications similar to those observed in patients with lethal perinatal osteogenesis imperfecta. The Mov-13 mouse system should allow the authors to study the effect specific mutations introduced in transfected proα1(I) genes have on the synthesis, assembly, and function of collagen I

  18. Engineering bone tissue from human embryonic stem cells

    OpenAIRE

    Marolt, Darja; Campos, Iván Marcos; Bhumiratana, Sarindr; Koren, Ana; Petridis, Petros; Zhang, Geping; Spitalnik, Patrice F.; Grayson, Warren L.; Vunjak-Novakovic, Gordana

    2012-01-01

    In extensive bone defects, tissue damage and hypoxia lead to cell death, resulting in slow and incomplete healing. Human embryonic stem cells (hESC) can give rise to all specialized lineages found in healthy bone and are therefore uniquely suited to aid regeneration of damaged bone. We show that the cultivation of hESC-derived mesenchymal progenitors on 3D osteoconductive scaffolds in bioreactors with medium perfusion leads to the formation of large and compact bone constructs. Notably, the i...

  19. Nicotine-induced chondrogenic differentiation of human bone marrow stromal cells in vitro.

    Science.gov (United States)

    Ying, Xiaozhou; Zhang, Wei; Cheng, Shaowen; Nie, Pengfei; Cheng, Xiaojie; Shen, Yue; Wang, Wei; Xue, Enxing; Chen, Qingyu; Kou, Dongquan; Peng, Lei; Zhang, Yu; Lu, Chuanzhu

    2012-11-01

    Nicotine has been reported that it has a dose-dependent effect on matrix mineralization by human bone marrow cells. However, there is no relevant research concerning on chondrogenic differentiation potential of bone marrow stromal stem cells (BMSCs) treated with nicotine in vitro. The aims of the study were to examine the effects of nicotine (0, 10(-7), 10(-6) and 10(-5) M) on the proliferation and chondrogenic differentiation of BMSCs from three healthy donors in vitro. BMSCs proliferation was analyzed by CCK8 assay and real-time polymerase chain reaction was used to assay the expression of type II collagen, aggrecan, type I collagen and type X collagen. The proteoglycan content was stained by Alcian blue, and the sulfated glycosaminoglycan (sGAG) content of BMSCs was quantified spectrofluorometrically using dimethylmethylene blue. The cell viability was not significantly impaired until up to a concentration of 10(-5) M nicotine. Nicotine promoted the proliferation and enhanced the expression of type II collagen at the level up to 10(-6) M (P < 0.05). The expression of aggrecan was reduced at the concentration of 10(-5) M nicotine at day 14 (P < 0.05), and there was no significant difference in aggrecan gene expression at 10(-7) and 10(-6) M nicotine levels compared to control group (n.s.). Also the fibroblastic and hypertrophic gene expressions were down-regulated in the chondrogenic medium with 10(-7)-10(-5) M nicotine (P < 0.05). It was implied that local application of nicotine at an appropriate concentration may be a promising approach for enhancing chondrogenic differentiation capacity of BMSCs in cell-based cartilage tissue engineering. Also these results indicate that nicotine maybe a potentially useful drug for the treatment of Osteoarthritis.

  20. MCF-7 human mammary adenocarcinoma cells exhibit augmented responses to human insulin on a collagen IV surface

    DEFF Research Database (Denmark)

    Listov-Saabye, Nicolai; Jensen, Marianne Blirup; Kiehr, Benedicte

    2009-01-01

    Human mammary cell lines are extensively used for preclinical safety assessment of insulin analogs. However, it is essentially unknown how mitogenic responses can be optimized in mammary cell-based systems. We developed an insulin mitogenicity assay in MCF-7 human mammary adenocarcinoma cells......, under low serum (0.1% FCS) and phenol red-free conditions, with 3H thymidine incorporation as endpoint. Based on EC50 values determined from 10-fold dilution series, beta-estradiol was the most potent mitogen, followed by human IGF-1, human AspB10 insulin and native human insulin. AspB10 insulin...... was significantly more mitogenic than native insulin, validating the ability of the assay to identify hypermitogenic human insulin analogs. With MCF-7 cells on a collagen IV surface, the ranking of mitogens was maintained, but fold mitogenic responses and dynamic range and steepness of dose-response curves were...

  1. Extracellular matrix collagen alters cell proliferation and cell cycle progression of human uterine leiomyoma smooth muscle cells.

    Science.gov (United States)

    Koohestani, Faezeh; Braundmeier, Andrea G; Mahdian, Arash; Seo, Jane; Bi, JiaJia; Nowak, Romana A

    2013-01-01

    Uterine leiomyomas (ULs) are benign tumors occurring in the majority of reproductive aged women. Despite the high prevalence of these tumors, little is known about their etiology. A hallmark of ULs is the excessive deposition of extracellular matrix (ECM), primarily collagens. Collagens are known to modulate cell behavior and function singularly or through interactions with integrins and growth factor-mediated mitogenic pathways. To better understand the pathogenesis of ULs and the role of ECM collagens in their growth, we investigated the interaction of leiomyoma smooth muscle cells (LSMCs) with two different forms of collagen, non-polymerized collagen (monomeric) and polymerized collagen (fibrillar), in the absence or presence of platelet-derived growth factor (PDGF), an abundant growth factor in ULs. Primary cultures of human LSMCS from symptomatic patients were grown on these two different collagen matrices and their morphology, cytoskeletal organization, cellular proliferation, and signaling pathways were evaluated. Our results showed that LSMCs had distinct morphologies on the different collagen matrices and their basal as well as PDGF-stimulated proliferation varied on these matrices. These differences in proliferation were accompanied by changes in cell cycle progression and p21, an inhibitory cell cycle protein. In addition we found alterations in the phosphorylation of focal adhesion kinase, cytoskeletal reorganization, and activation of the mitogen activated protein kinase (MAPK) signaling pathway. In conclusion, our results demonstrate a direct effect of ECM on the proliferation of LSMCs through interplay between the collagen matrix and the PDGF-stimulated MAPK pathway. In addition, these findings will pave the way for identifying novel therapeutic approaches for ULs that target ECM proteins and their signaling pathways in ULs.

  2. Microstructural and Photoacoustic Infrared Spectroscopic Studies of Human Cortical Bone with Osteogenesis Imperfecta

    Science.gov (United States)

    Gu, Chunju; Katti, Dinesh R.; Katti, Kalpana S.

    2016-04-01

    The molecular basis of bone disease osteogenesis imperfecta (OI) and the mineralization of hydroxyapatite in OI bone have been of significant research interest. To further investigate the mechanism of OI disease and bone mineralization, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy, and x-ray diffraction (XRD) are used in the present study to describe the structural and compositional differences between OI and healthy bone. OI bone exhibits more porous, fibrous features, abnormal collagen fibrils, and abnormal mineral deposits. Likewise, photoacoustic-FTIR experiments indicate an aberrant collagen structure and an altered mineral structure in OI. In contrast, there is neither significant difference in the non-collagenous proteins (NCPs) composition observed nor apparent change in the crystal structure between OI and healthy bone minerals as shown in XRD and energy-dispersive x-ray spectroscopy (EDS) results. This observation indicates that the biomineralization process is more controlled by the bone cells and non-collagenous phosphorylated proteins. The present study also confirms that there is an orientational influence on the stoichiometry of the mineral in OI bone. Also, a larger volume of the hydrated layer in the transverse plane than the longitudinal plane of the mineral crystal structure is proposed. The appearance of a new C-S band in the FTIR spectra in OI bone suggests the substitution of glycine by cysteine in collagen molecules or/and an increased amount of cysteine-rich osteonectin that relates to mineral nucleation and mineral crystal formation.

  3. Hypoxia Is a Critical Parameter for Chondrogenic Differentiation of Human Umbilical Cord Blood Mesenchymal Stem Cells in Type I/III Collagen Sponges

    Directory of Open Access Journals (Sweden)

    Tangni Gómez-Leduc

    2017-09-01

    Full Text Available Umbilical cord blood (UCB is an attractive alternative to bone marrow for isolation of mesenchymal stem cells (MSCs to treat articular cartilage defects. Here, we set out to determine the growth factors (bone morphogenetic protein 2 (BMP-2 and transforming growth factor-β (TGF-β1 and oxygen tension effects during chondrogenesis of human UCB-MSCs for cartilage engineering. Chondrogenic differentiation was induced using 3D cultures in type I/III collagen sponges with chondrogenic factors in normoxia (21% O2 or hypoxia (<5% O2 for 7, 14 and 21 days. Our results show that UCB-MSCs can be committed to chondrogenesis in the presence of BMP-2+TGF-β1. Normoxia induced the highest levels of chondrocyte-specific markers. However, hypoxia exerted more benefit by decreasing collagen X and matrix metalloproteinase-13 (MMP13 expression, two chondrocyte hypertrophy markers. However, a better chondrogenesis was obtained by switching oxygen conditions, with seven days in normoxia followed by 14 days in hypoxia, since these conditions avoid hypertrophy of hUCB-MSC-derived chondrocytes while maintaining the expression of chondrocyte-specific markers observed in normoxia. Our study demonstrates that oxygen tension is a key factor for chondrogenesis and suggests that UBC-MSCs 3D-culture should begin in normoxia to obtain a more efficient chondrocyte differentiation before placing them in hypoxia for chondrocyte phenotype stabilization. UCB-MSCs are therefore a reliable source for cartilage engineering.

  4. Growth hormone stimulates the collagen synthesis in human tendon and skeletal muscle without affecting myofibrillar protein synthesis

    DEFF Research Database (Denmark)

    Doessing, Simon; Heinemeier, Katja M; Holm, Lars

    2010-01-01

    young individuals. rhGH administration caused an increase in serum GH, serum IGF-I, and IGF-I mRNA expression in tendon and muscle. Tendon collagen I mRNA expression and tendon collagen protein synthesis increased by 3.9-fold and 1.3-fold, respectively (P ...RNA expression and muscle collagen protein synthesis increased by 2.3-fold and 5.8-fold, respectively (P protein synthesis was unaffected by elevation of GH and IGF-I. Moderate exercise did not enhance the effects of GH manipulation. Thus, increased GH availability stimulates...... matrix collagen synthesis in skeletal muscle and tendon, but without any effect upon myofibrillar protein synthesis. The results suggest that GH is more important in strengthening the matrix tissue than for muscle cell hypertrophy in adult human musculotendinous tissue....

  5. Regulation of bone blood flow in humans

    DEFF Research Database (Denmark)

    Heinonen, Ilkka; Boushel, Robert; Hellsten, Ylva

    2018-01-01

    of cyclooxygenase (COX) enzyme, thus prostaglandin (PG) synthesis on femoral bone marrow blood flow by positron emission tomography in healthy young men at rest and during one leg dynamic exercise. In an additional group of healthy men, the role of adenosine (ADO) in the regulation of BBF during exercise......The mechanisms that regulate bone blood flow (BBF) in humans are largely unknown. Animal studies suggest that nitric oxide (NO) could be involved and in the present study we investigated the effects of inhibition of nitric oxide synthase (NOS) alone and in combination with inhibition.......036), but did not affect BBF significantly during exercise (5.5±1.4 ml/100g/min, p=0.25). On the other hand, while combined NOS and COX inhibition did not cause any further reduction of blood flow at rest (0.6±0.2 ml/100g/min), the combined blockade reduced BBF during exercise by ~21%, to 5.0±1.8 ml/100g/min (p...

  6. Confocal Raman mapping of collagen cross-link and crystallinity of human dentin-enamel junction

    Science.gov (United States)

    Slimani, Amel; Nouioua, Fares; Desoutter, Alban; Levallois, Bernard; Cuisinier, Frédéric J. G.; Tassery, Hervé; Terrer, Elodie; Salehi, Hamideh

    2017-08-01

    The separation zone between enamel and dentin [dentin-enamel junction (DEJ)] with different properties in biomechanical composition has an important role in preventing crack propagation from enamel to dentin. The understanding of the chemical structure (inorganic and organic components), physical properties, and chemical composition of the human DEJ could benefit biomimetic materials in dentistry. Spatial distribution of calcium phosphate crystallinity and the collagen crosslinks near DEJ were studied using confocal Raman microscopy and calculated by different methods. To obtain collagen crosslinking, the ratio of two peaks 1660 cm-1 over 1690 cm-1 (amide I bands) is calculated. For crystallinity, the inverse full-width at half maximum of phosphate peak at 960 cm-1, and the ratio of two Raman peaks of phosphate at 960/950 cm-1 is provided. In conclusion, the study of chemical and physical properties of DEJ provides many benefits in the biomaterial field to improve the synthesis of dental materials in respect to the natural properties of human teeth. Confocal Raman microscopy as a powerful tool provides the molecular structure to identify the changes along DEJ and can be expanded for other mineralized tissues.

  7. STAT6-Dependent Collagen Synthesis in Human Fibroblasts Is Induced by Bovine Milk.

    Directory of Open Access Journals (Sweden)

    Stefan Kippenberger

    Full Text Available Since the domestication of the urus, 10.000 years ago, mankind utilizes bovine milk for different purposes. Besides usage as a nutrient also the external application of milk on skin has a long tradition going back to at least the ancient Aegypt with Cleopatra VII as a great exponent. In order to test whether milk has impact on skin physiology, cultures of human skin fibroblasts were exposed to commercial bovine milk. Our data show significant induction of proliferation by milk (max. 2,3-fold, EC50: 2,5% milk without toxic effects. Surprisingly, bovine milk was identified as strong inducer of collagen 1A1 synthesis at both, the protein (4-fold, EC50: 0,09% milk and promoter level. Regarding the underlying molecular pathways, we show functional activation of STAT6 in a p44/42 and p38-dependent manner. More upstream, we identified IGF-1 and insulin as key factors responsible for milk-induced collagen synthesis. These findings show that bovine milk contains bioactive molecules that act on human skin cells. Therefore, it is tempting to test the herein introduced concept in treatment of atrophic skin conditions induced e.g. by UV light or corticosteroids.

  8. Pathogenesis of age-related bone loss in humans.

    Science.gov (United States)

    Khosla, Sundeep

    2013-10-01

    Although data from rodent systems are extremely useful in providing insights into possible mechanisms of age-related bone loss, concepts evolving from animal models need to ultimately be tested in humans. This review provides an update on mechanisms of age-related bone loss in humans based on the author's knowledge of the field and focused literature reviews. Novel imaging, experimental models, biomarkers, and analytic techniques applied directly to human studies are providing new insights into the patterns of bone mass acquisition and loss as well as the role of sex steroids, in particular estrogen, on bone metabolism and bone loss with aging in women and men. These studies have identified the onset of trabecular bone loss at multiple sites that begins in young adulthood and remains unexplained, at least based on current paradigms of the mechanisms of bone loss. In addition, estrogen appears to be a major regulator of bone metabolism not only in women but also in men. Studies assessing mechanisms of estrogen action on bone in humans have identified effects of estrogen on RANKL expression by several different cell types in the bone microenvironment, a role for TNF-α and IL-1β in mediating effects of estrogen deficiency on bone, and possible regulation of the Wnt inhibitor, sclerostin, by estrogen. There have been considerable advances in our understanding of age-related bone loss in humans. However, there are also significant gaps in knowledge, particularly in defining cell autonomous changes in bone in human studies to test or validate concepts emerging from studies in rodents. Decision Editor: Luigi Ferrucci, MD, PhD.

  9. Asiaticoside induces cell proliferation and collagen synthesis in human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Linda Yuliati

    2015-12-01

    Asiaticoside induces HDF proliferation and type I and III collagen synthesis in a time- and dose-dependent pattern. Asiaticoside has a similar effect as retinoic acid on type I and type III collagen synthesis.

  10. Basic Fibroblast Growth Factor Fused with Tandem Collagen-Binding Domains from Clostridium histolyticum Collagenase ColG Increases Bone Formation

    Directory of Open Access Journals (Sweden)

    Hiroyuki Sekiguchi

    2018-01-01

    Full Text Available Basic fibroblast growth factor 2 (bFGF accelerates bone formation during fracture healing. Because the efficacy of bFGF decreases rapidly following its diffusion from fracture sites, however, repeated dosing is required to ensure a sustained therapeutic effect. We previously developed a fusion protein comprising bFGF, a polycystic kidney disease domain (PKD; s2b, and collagen-binding domain (CBD; s3 sourced from the Clostridium histolyticum class II collagenase, ColH, and reported that the combination of this fusion protein with a collagen-like peptide, poly(Pro-Hyp-Gly10, induced mesenchymal cell proliferation and callus formation at fracture sites. In addition, C. histolyticum produces class I collagenase (ColG with tandem CBDs (s3a and s3b at the C-terminus. We therefore hypothesized that a bFGF fusion protein containing ColG-derived tandem CBDs (s3a and s3b would show enhanced collagen-binding activity, leading to improved bone formation. Here, we examined the binding affinity of four collagen anchors derived from the two clostridial collagenases to H-Gly-Pro-Arg-Gly-(Pro-Hyp-Gly12-NH2, a collagenous peptide, by surface plasmon resonance and found that tandem CBDs (s3a-s3b have the highest affinity for the collagenous peptide. We also constructed four fusion proteins consisting of bFGF and s3 (bFGF-s3, s2b-s3b (bFGF-s2b-s3, s3b (bFGF-s3b, and s3a-s3b (bFGF-s3a-s3b and compared their biological activities to those of a previous fusion construct (bFGF-s2b-s3 using a cell proliferation assay in vitro and a mouse femoral fracture model in vivo. Among these CB-bFGFs, bFGF-s3a-s3b showed the highest capacity to induce mesenchymal cell proliferation and callus formation in the mice fracture model. The poly(Pro-Hyp-Gly10/bFGF-s3a-s3b construct may therefore have the potential to promote bone formation in clinical settings.

  11. Effect of Tris-acetate buffer on endotoxin removal from human-like collagen used biomaterials.

    Science.gov (United States)

    Zhang, Huizhi; Fan, Daidi; Deng, Jianjun; Zhu, Chenghui; Hui, Junfeng; Ma, Xiaoxuan

    2014-09-01

    Protein preparation, which has active ingredients designated for the use of biomaterials and therapeutical protein, is obtained by genetic engineering, but products of genetic engineering are often contaminated by endotoxins. Because endotoxin is a ubiquitous and potent proinflammatory agent, endotoxin removal or depletion from protein is essential for researching any biomaterials. In this study, we have used Tris-acetate (TA) buffer of neutral pH value to evaluate endotoxins absorbed on the Pierce high-capacity endotoxin removal resin. The effects of TA buffer on pH, ionic strength, incubation time as well as human-like collagen (HLC) concentration on eliminating endotoxins are investigated. In the present experiments, we design an optimal method for TA buffer to remove endotoxin from recombinant collagen and use a chromogenic tachypleus amebocyte lysate (TAL) test kit to measure the endotoxin level of HLC. The present results show that, the endotoxins of HLC is dropped to 8.3EU/ml at 25 mM TA buffer (pH7.8) with 150 mM NaCl when setting incubation time at 6h, and HLC recovery is about 96%. Under this experimental condition, it is proved to exhibit high efficiencies of both endotoxin removal and collagen recovery. The structure of treated HLC was explored by Transmission Electron Microscopy (TEM), demonstrating that the property and structure of HLC treated by TA buffer are maintained. Compared to the most widely used endotoxin removal method, Triton X-114 extraction, using TA buffer can obtain the non-toxic HLC without extra treatment for removing the toxic substances in Triton X-114. In addition, the present study aims at establishing a foundation for further work in laboratory animal science and providing a foundation for medical grade biomaterials. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Exercise-dependent IGF-I, IGFBPs, and type I collagen changes in human peritendinous connective tissue determined by microdialysis

    DEFF Research Database (Denmark)

    Olesen, Jens L; Heinemeier, Katja M; Gemmer, Carsten

    2007-01-01

    Microdialysis studies indicate that mechanical loading of human tendon during exercise elevates type I collagen production in tendon. However, the possibility that the insertion of microdialysis fibers per se may increase the local collagen production due to trauma has not been explored. Insulin......-terminal propeptide (PICP) and COOH-terminal telopeptide of type I collagen] were measured by microdialysis in peritendinous tissue of the human Achilles tendon in an exercise group (performing a 36-km run, n = 6) and a control group (no intervention, n = 6). An increase in local PICP concentration was seen in both...... and exercise groups after 48 h (P human peritendinous tissue in response to prolonged mechanical loading with part of the increase due to trauma from the sampling...

  13. Collagen Type I alpha1 (COL1A1 Gene Polymorphism and Bone Mineral Density in Postmenopausal Kazakh Women

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    Akbota Aitkulova

    2014-12-01

    Full Text Available Introduction: Single nucleotide polymorphism (SNP at the collagen type I alpha 1 gene (COL1A1 rs1800012 has been widely studied and has shown an association with bone mineral density (BMD and fractures. A minor allele TT of this SNP was found to be greatly overrepresented in individuals with fractures compared to controls, thus becoming a good predictor of  increased fracture risk. The aim of this investigation was to evaluate potential association between COL1A1 gene polymorphism and osteoporosis in Kazakh postmenopausal women.Methods: The study population included 103 postmenopausal women recruited from Pavlodar and Almaty clinics. BMD was measured using DEXA. Genomic DNA was extracted from peripheral venous blood of study participants with Wizard® Genomic DNA Purification Kit (Promega, USA. Detection of COL1A1 +1245G/T (Sp1 polymorphism was done by the TaqMan® SNP Genotyping Assay of real-time PCR.Results: Densitometry results revealed 36 osteoporotic, 42 osteopenic, and 25 normal postmenopausal women. Data analysis of 1245G>T polymorphism in COL1A1 gene in the group of women with osteopenia and osteoporosis revealed deviation from Hardy-Weinberg equilibrium. The mutant TT genotype was prevalent compared to the heterozygous genotype GT in both groups. Distributions were 83% GG, 3% GT, and 14% TT in the group with osteopenia and 80% GG, 6% GT, and 14% TT in the group with osteoporosis. The distribution of genotypes frequency in the group of normal postmenopausal women was 76% GG, 16% GT, and 8% TT.Conclusion: These results suggest that TT genotype of COL1A1 +1245G/T (Sp1 polymorphism is associated with risk of postmenopausal osteoporosis in Kazakh women. Further studies involving a larger number of women are needed to clarify the relationship of this polymorphism with risk of osteoporosis. 

  14. The ratio 1660/1690 cm(-1) measured by infrared microspectroscopy is not specific of enzymatic collagen cross-links in bone tissue.

    Science.gov (United States)

    Farlay, Delphine; Duclos, Marie-Eve; Gineyts, Evelyne; Bertholon, Cindy; Viguet-Carrin, Stéphanie; Nallala, Jayakrupakar; Sockalingum, Ganesh D; Bertrand, Dominique; Roger, Thierry; Hartmann, Daniel J; Chapurlat, Roland; Boivin, Georges

    2011-01-01

    In postmenopausal osteoporosis, an impairment in enzymatic cross-links (ECL) occurs, leading in part to a decline in bone biomechanical properties. Biochemical methods by high performance liquid chromatography (HPLC) are currently used to measure ECL. Another method has been proposed, by Fourier Transform InfraRed Imaging (FTIRI), to measure a mature PYD/immature DHLNL cross-links ratio, using the 1660/1690 cm(-1) area ratio in the amide I band. However, in bone, the amide I band composition is complex (collagens, non-collagenous proteins, water vibrations) and the 1660/1690 cm(-1) by FTIRI has never been directly correlated with the PYD/DHLNL by HPLC. A study design using lathyritic rats, characterized by a decrease in the formation of ECL due to the inhibition of lysyl oxidase, was used in order to determine the evolution of 1660/1690 cm(-1) by FTIR Microspectroscopy in bone tissue and compare to the ECL quantified by HPLC. The actual amount of ECL was quantified by HPLC on cortical bone from control and lathyritic rats. The lathyritic group exhibited a decrease of 78% of pyridinoline content compared to the control group. The 1660/1690 cm(-1) area ratio was increased within center bone compared to inner bone, and this was also correlated with an increase in both mineral maturity and mineralization index. However, no difference in the 1660/1690 cm(-1) ratio was found between control and lathyritic rats. Those results were confirmed by principal component analysis performed on multispectral infrared images. In bovine bone, in which PYD was physically destructed by UV-photolysis, the PYD/DHLNL (measured by HPLC) was strongly decreased, whereas the 1660/1690 cm(-1) was unmodified. In conclusion, the 1660/1690 cm(-1) is not related to the PYD/DHLNL ratio, but increased with age of bone mineral, suggesting that a modification of this ratio could be mainly due to a modification of the collagen secondary structure related to the mineralization process.

  15. The Ratio 1660/1690 cm−1 Measured by Infrared Microspectroscopy Is Not Specific of Enzymatic Collagen Cross-Links in Bone Tissue

    Science.gov (United States)

    Farlay, Delphine; Duclos, Marie-Eve; Gineyts, Evelyne; Bertholon, Cindy; Viguet-Carrin, Stéphanie; Nallala, Jayakrupakar; Sockalingum, Ganesh D.; Bertrand, Dominique; Roger, Thierry; Hartmann, Daniel J.; Chapurlat, Roland; Boivin, Georges

    2011-01-01

    In postmenopausal osteoporosis, an impairment in enzymatic cross-links (ECL) occurs, leading in part to a decline in bone biomechanical properties. Biochemical methods by high performance liquid chromatography (HPLC) are currently used to measure ECL. Another method has been proposed, by Fourier Transform InfraRed Imaging (FTIRI), to measure a mature PYD/immature DHLNL cross-links ratio, using the 1660/1690 cm−1 area ratio in the amide I band. However, in bone, the amide I band composition is complex (collagens, non-collagenous proteins, water vibrations) and the 1660/1690 cm−1 by FTIRI has never been directly correlated with the PYD/DHLNL by HPLC. A study design using lathyritic rats, characterized by a decrease in the formation of ECL due to the inhibition of lysyl oxidase, was used in order to determine the evolution of 1660/1690 cm−1 by FTIR Microspectroscopy in bone tissue and compare to the ECL quantified by HPLC. The actual amount of ECL was quantified by HPLC on cortical bone from control and lathyritic rats. The lathyritic group exhibited a decrease of 78% of pyridinoline content compared to the control group. The 1660/1690 cm−1 area ratio was increased within center bone compared to inner bone, and this was also correlated with an increase in both mineral maturity and mineralization index. However, no difference in the 1660/1690 cm−1 ratio was found between control and lathyritic rats. Those results were confirmed by principal component analysis performed on multispectral infrared images. In bovine bone, in which PYD was physically destructed by UV-photolysis, the PYD/DHLNL (measured by HPLC) was strongly decreased, whereas the 1660/1690 cm−1 was unmodified. In conclusion, the 1660/1690 cm−1 is not related to the PYD/DHLNL ratio, but increased with age of bone mineral, suggesting that a modification of this ratio could be mainly due to a modification of the collagen secondary structure related to the mineralization process. PMID:22194900

  16. Colonic complications following human bone marrow transplantation

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    Paulino Martínez Hernández-Magro

    2015-01-01

    Full Text Available Background: Human bone marrow transplantation (BMT becomes an accepted treatment of leukemia, aplastic anemia, immunodeficiency syndromes, and hematologic malignancies. Colorectal surgeons must know how to determine and manage the main colonic complications. Objective: To review the clinical features, clinical and pathological staging of graft vs host disease (GVHD, and treatment of patients suffering with colonic complications of human bone marrow transplantation. Patients and methods: We have reviewed the records of all patients that received an allogeneic bone marrow transplant and were evaluated at our Colon and Rectal Surgery department due to gastrointestinal symptoms, between January 2007 and January 2012. The study was carried out in patients who developed colonic complications, all of them with clinical, histopathological or laboratory diagnosis. Results: The study group was constituted by 77 patients, 43 male and 34 female patients. We identified colonic complications in 30 patients (38.9%; five patients developed intestinal toxicity due to pretransplant chemotherapy (6.4%; graft vs. host disease was present in 16 patients (20%; 13 patients (16.8% developed acute colonic GVHD, and 3 (3.8% chronic GVHD. Infection was identified in 9 patients (11.6%. Conclusions: The three principal colonic complications are the chemotherapy toxicity, GVHD, and superinfection; the onset of symptoms could help to suspect the type of complication (0–20 day chemotherapy toxicity, 20 and more GVHD, and infection could appear in any time of transplantation. Resumo: Experiência: O transplante de medula óssea humana (MOH passou a ser um tratamento adotado para leucemia, anemia aplástica, síndromes de imunodeficiência e neoplasias hematológicas. Cirurgiões colorretais devem saber como determinar e tratar as principais complicações do cólon. Objetivo: Revisar as características clínicas, estadiamentos clínico e patológico da doença do enxerto

  17. Effects of the Nd:YAG laser on DNA synthesis and collagen production in human skin fibroblast cultures

    Energy Technology Data Exchange (ETDEWEB)

    Castro, D.J.; Abergel, R.P.; Meeker, C.; Dwyer, R.M.; Lesavoy, M.A.; Uitto, J.

    1983-09-01

    Human skin fibroblasts were subjected to treatment with a Neodymium:YAG laser at 1060 nm with varying levels of energy determined by a reproducible method of dosimetry. DNA synthesis in the cells was measured by the incorporation of (3H)thymidine, and collagen production was monitored by the synthesis of nondialyzable (3H)hydroxyproline after incubation of cells with (3H)proline. Using energy levels equal to 1.7 X 10(3) J/cm2, a significant reduction in DNA synthesis was noted, while the cells remained viable as tested by the trypan blue exclusion test. With energy levels higher or equal to 2.3 X 10(3) J/cm2, the suppression of DNA synthesis was accompanied by cell nonviability. The collagen production, when measured immediately following the treatment with 1.7 X 10(3) J/cm2, was markedly reduced, and similar effects were observed with higher energy levels. However, when the cells were tested for collagen production at 20 hours following laser treatment, there was a significant decrease in collagen production at energy levels as low as 1.1 X 10(3) J/cm2, a dose that did not affect DNA synthesis or cell viability. Thus, the results indicate that the Nd:YAG laser can selectively suppress collagen production without affecting cell proliferation. These observations suggest that laser treatment could potentially be used to reduce collagen deposition in conditions such as keloids and hypertrophic scars.

  18. Human platelet lysate improves human cord blood derived ECFC survival and vasculogenesis in three dimensional (3D) collagen matrices.

    Science.gov (United States)

    Kim, Hyojin; Prasain, Nutan; Vemula, Sasidhar; Ferkowicz, Michael J; Yoshimoto, Momoko; Voytik-Harbin, Sherry L; Yoder, Mervin C

    2015-09-01

    Human cord blood (CB) is enriched in circulating endothelial colony forming cells (ECFCs) that display high proliferative potential and in vivo vessel forming ability. Since diminished ECFC survival is known to dampen the vasculogenic response in vivo, we tested how long implanted ECFC survive and generate vessels in three-dimensional (3D) type I collagen matrices in vitro and in vivo. We hypothesized that human platelet lysate (HPL) would promote cell survival and enhance vasculogenesis in the 3D collagen matrices. We report that the percentage of ECFC co-cultured with HPL that were alive was significantly enhanced on days 1 and 3 post-matrix formation, compared to ECFC alone containing matrices. Also, co-culture of ECFC with HPL displayed significantly more vasculogenic activity compared to ECFC alone and expressed significantly more pro-survival molecules (pAkt, p-Bad and Bcl-xL) in the 3D collagen matrices in vitro. Treatment with Akt1 inhibitor (A-674563), Akt2 inhibitor (CCT128930) and Bcl-xL inhibitor (ABT-263/Navitoclax) significantly decreased the cell survival and vasculogenesis of ECFC co-cultured with or without HPL and implicated activation of the Akt1 pathway as the critical mediator of the HPL effect on ECFC in vitro. A significantly greater average vessel number and total vascular area of human CD31(+) vessels were present in implants containing ECFC and HPL, compared to the ECFC alone implants in vivo. We conclude that implantation of ECFC with HPL in vivo promotes vasculogenesis and augments blood vessel formation via diminishing apoptosis of the implanted ECFC. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Improvement of the Chondrocyte-Specific Phenotype upon Equine Bone Marrow Mesenchymal Stem Cell Differentiation: Influence of Culture Time, Transforming Growth Factors and Type I Collagen siRNAs on the Differentiation Index

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    Thomas Branly

    2018-02-01

    Full Text Available Articular cartilage is a tissue characterized by its poor intrinsic capacity for self-repair. This tissue is frequently altered upon trauma or in osteoarthritis (OA, a degenerative disease that is currently incurable. Similar musculoskeletal disorders also affect horses and OA incurs considerable economic loss for the equine sector. In the view to develop new therapies for humans and horses, significant progress in tissue engineering has led to the emergence of new generations of cartilage therapy. Matrix-associated autologous chondrocyte implantation is an advanced 3D cell-based therapy that holds promise for cartilage repair. This study aims to improve the autologous chondrocyte implantation technique by using equine mesenchymal stem cells (MSCs from bone marrow differentiated into chondrocytes that can be implanted in the chondral lesion. The optimized protocol relies on culture under hypoxia within type I/III collagen sponges. Here, we explored three parameters that influence MSC differentiation: culture times, growth factors and RNA interference strategies. Our results suggest first that an increase in culture time from 14 to 28 or 42 days lead to a sharp increase in the expression of chondrocyte markers, notably type II collagen (especially the IIB isoform, along with a concomitant decrease in HtrA1 expression. Nevertheless, the expression of type I collagen also increased with longer culture times. Second, regarding the growth factor cocktail, TGF-β3 alone showed promising result but the previously tested association of BMP-2 and TGF-β1 better limits the expression of type I collagen. Third, RNA interference targeting Col1a2 as well as Col1a1 mRNA led to a more significant knockdown, compared with a conventional strategy targeting Col1a1 alone. This chondrogenic differentiation strategy showed a strong increase in the Col2a1:Col1a1 mRNA ratio in the chondrocytes derived from equine bone marrow MSCs, this ratio being considered as an

  20. Improvement of the Chondrocyte-Specific Phenotype upon Equine Bone Marrow Mesenchymal Stem Cell Differentiation: Influence of Culture Time, Transforming Growth Factors and Type I Collagen siRNAs on the Differentiation Index.

    Science.gov (United States)

    Branly, Thomas; Contentin, Romain; Desancé, Mélanie; Jacquel, Thibaud; Bertoni, Lélia; Jacquet, Sandrine; Mallein-Gerin, Frédéric; Denoix, Jean-Marie; Audigié, Fabrice; Demoor, Magali; Galéra, Philippe

    2018-02-01

    Articular cartilage is a tissue characterized by its poor intrinsic capacity for self-repair. This tissue is frequently altered upon trauma or in osteoarthritis (OA), a degenerative disease that is currently incurable. Similar musculoskeletal disorders also affect horses and OA incurs considerable economic loss for the equine sector. In the view to develop new therapies for humans and horses, significant progress in tissue engineering has led to the emergence of new generations of cartilage therapy. Matrix-associated autologous chondrocyte implantation is an advanced 3D cell-based therapy that holds promise for cartilage repair. This study aims to improve the autologous chondrocyte implantation technique by using equine mesenchymal stem cells (MSCs) from bone marrow differentiated into chondrocytes that can be implanted in the chondral lesion. The optimized protocol relies on culture under hypoxia within type I/III collagen sponges. Here, we explored three parameters that influence MSC differentiation: culture times, growth factors and RNA interference strategies. Our results suggest first that an increase in culture time from 14 to 28 or 42 days lead to a sharp increase in the expression of chondrocyte markers, notably type II collagen (especially the IIB isoform), along with a concomitant decrease in HtrA1 expression. Nevertheless, the expression of type I collagen also increased with longer culture times. Second, regarding the growth factor cocktail, TGF-β3 alone showed promising result but the previously tested association of BMP-2 and TGF-β1 better limits the expression of type I collagen. Third, RNA interference targeting Col1a2 as well as Col1a1 mRNA led to a more significant knockdown, compared with a conventional strategy targeting Col1a1 alone. This chondrogenic differentiation strategy showed a strong increase in the Col2a1 : Col1a1 mRNA ratio in the chondrocytes derived from equine bone marrow MSCs, this ratio being considered as an index of the

  1. Hypoxia-Induced Collagen Synthesis of Human Lung Fibroblasts by Activating the Angiotensin System

    Directory of Open Access Journals (Sweden)

    Shan-Shan Liu

    2013-12-01

    Full Text Available The exact molecular mechanism that mediates hypoxia-induced pulmonary fibrosis needs to be further clarified. The aim of this study was to explore the effect and underlying mechanism of angiotensin II (Ang II on collagen synthesis in hypoxic human lung fibroblast (HLF cells. The HLF-1 cell line was used for in vitro studies. Angiotensinogen (AGT, angiotensin converting enzyme (ACE, angiotensin II type 1 receptor (AT1R and angiotensin II type 2 receptor (AT2R expression levels in human lung fibroblasts were analysed using real-time polymerase chain reaction (RT-PCR after hypoxic treatment. Additionally, the collagen type I (Col-I, AT1R and nuclear factor κappaB (NF-κB protein expression levels were detected using Western blot analysis, and NF-κB nuclear translocation was measured using immunofluorescence localization analysis. Ang II levels in HLF-1 cells were measured with an enzyme-linked immunosorbent assay (ELISA. We found that hypoxia increased Col-I mRNA and protein expression in HLF-1 cells, and this effect could be inhibited by an AT1R or AT2R inhibitor. The levels of NF-κB, RAS components and Ang II production in HLF-1 cells were significantly increased after the hypoxia exposure. Hypoxia or Ang II increased NF-κB-p50 protein expression in HLF-1 cells, and the special effect could be inhibited by telmisartan (TST, an AT1R inhibitor, and partially inhibited by PD123319, an AT2R inhibitor. Importantly, hypoxia-induced NF-κB nuclear translocation could be nearly completely inhibited by an AT1R or AT2R inhibitor. Furthermore pyrrolidine dithiocarbamate (PDTC, a NF-κB blocker, abolished the expression of hypoxia-induced AT1R and Col-I in HLF-1 cells. Our results indicate that Ang II-mediated NF-κB signalling via ATR is involved in hypoxia-induced collagen synthesis in human lung fibroblasts.

  2. Human bone ingrowth into a porous tantalum acetabular cup

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    Gregory N. Haidemenopoulos

    2017-11-01

    Full Text Available Porous Tantalum is increasingly used as a structural scaffold in orthopaedic applications. Information on the mechanisms of human bone ingrowth into trabecular metal implants is rather limited. In this work we have studied, qualitatively, human bone ingrowth into a retrieved porous tantalum monoblock acetabular cup using optical microscopy, scanning electron microscopy and energy dispersive X-ray analysis. According to the results and taking into account the short operational life (4 years of the implant, bone ingrowth on the acetabular cup took place in the first two-rows of porous tantalum cells to an estimated depth of 1.5 to 2 mm. The bone material, grown inside the first raw of cells, had almost identical composition with the attached bone on the cup surface, as verified by the same Ca:P ratio. Bone ingrowth has been a gradual process starting with Ca deposition on the tantalum struts, followed by bone formation into the tantalum cells, with gradual densification of the bone tissue into hydroxyapatite. A critical step in this process has been the attachment of bone material to the tantalum struts following the topology of the porous tantalum scaffold. These results provide insight to the human bone ingrowth process into porous tantalum implants.

  3. Histologic and Radiographic Analysis of Nonhealing Extraction Sockets Treated with Bio-Oss Collagen After a 4-Month Healing Period: A Prospective Descriptive Study in Humans.

    Science.gov (United States)

    Tirone, Federico; Salzano, Stefano; Pagano, Marco

    2018-03-07

    Healing of extraction sockets may sometimes result in formation of fibrous tissue instead of bone, even after 4 months, an occurrence that may hinder implant placement. The aim of this preliminary observational study was to histologically evaluate quality and amount of bone regeneration after treating nonhealing sockets with a bovine-derived xenograft enriched with porcine collagen (Bio-Oss Collagen, Geistlich) without barrier membranes. Biopsy specimens were collected during implant placement, 4 months after grafting. A total of 10 cases were treated and evaluated. In all cases, correct implant placement was possible and no implant failure occurred up to 6 months after loading. The histologic analysis demonstrated new bone formation in all specimens. The percentage of newly formed bone was 29.1% (SD 20.71%; range 5% to 48%). Xenograft particles in direct contact with newly formed bone were visible, and mature lamellar bone was observed in 8 cases.

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

    Science.gov (United States)

    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.

  5. Crystal structure of the second fibronectin type III (FN3) domain from human collagen α1 type XX.

    Science.gov (United States)

    Zhao, Jingfeng; Ren, Jixia; Wang, Nan; Cheng, Zhong; Yang, Runmei; Lin, Gen; Guo, Yi; Cai, Dayong; Xie, Yong; Zhao, Xiaohong

    2017-12-01

    Collagen α1 type XX, which contains fibronectin type III (FN3) repeats involving six FN3 domains (referred to as the FN#1-FN#6 domains), is an unusual member of the fibril-associated collagens with interrupted triple helices (FACIT) subfamily of collagens. The results of standard protein BLAST suggest that the FN3 repeats might contribute to collagen α1 type XX acting as a cytokine receptor. To date, solution NMR structures of the FN#3, FN#4 and FN#6 domains have been determined. To obtain further structural evidence to understand the relationship between the structure and function of the FN3 repeats from collagen α1 type XX, the crystal structure of the FN#2 domain from human collagen α1 type XX (residues Pro386-Pro466; referred to as FN2-HCXX) was solved at 2.5 Å resolution. The crystal structure of FN2-HCXX shows an immunoglobulin-like fold containing a β-sandwich structure, which is formed by a three-stranded β-sheet (β1, β2 and β5) packed onto a four-stranded β-sheet (β3, β4, β6 and β7). Two consensus domains, tencon and fibcon, are structural analogues of FN2-HCXX. Fn8, an FN3 domain from human oncofoetal fibronectin, is the closest structural analogue of FN2-HCXX derived from a naturally occurring sequence. Based solely on the structural similarity of FN2-HCXX to other FN3 domains, the detailed functions of FN2-HCXX and the FN3 repeats in collagen α1 type XX cannot be identified.

  6. EFEK KOLAGEN DARI BERBAGAI JENIS TULANG IKAN TERHADAP KUALITAS MIOFIBRIL PROTEIN IKAN SELAMA PROSES DEHIDRASI [Effect of Various Fish Bone Collagens on the Quality of Myofibril Fish Protein During Dehydration Process

    Directory of Open Access Journals (Sweden)

    Yudhomenggolo Sastro Darmanto*

    2012-06-01

    Full Text Available Increase in fish fillet export in Indonesia has caused an increase in its waste such as bones, spines, skin and entrails of fish. Fish bones can be processed by demineralization to produce collagen, an important food additive. The effect of addition of 5% of collagen obtained from fresh water, brackish water and sea water fish bone on the fish protein miofibril of grouper was investigated in this research. Water sorption isotherm, Ca-ATPase activity, gel strength, water holding capacity, folding test and viscosity during dehydration process were evaluated. The results showed that collagens made from various fish bones have different Ca-ATPase activity. The reduction rate of Ca-ATPase activity were in accordance with the reduction of water sorbtion isotherm, gel forming ability, water holding capacity, viscosity and folding test during dehydration process.

  7. Bone dosimetry using synthetic images to represent trabecular bones of five regions of the human body

    Energy Technology Data Exchange (ETDEWEB)

    Lima Filho, Jose de M. [Instituto Federal de Educacao, Ciencia e Tecnologia de Pernambuco (IFPE), Recife, PE (Brazil); Vieira, Jose W. [Escola Politecnica de Pernambuco (POLI). Universidade de Pernambuco (UPE), Recife, PE (Brazil); Lima, Vanildo J. de M., E-mail: vjr@ufpe.br [Departamento de Anatomia. Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil); Lima, Lindeval F., E-mail: lindeval@dmat.ufrr.br [Departamento de Matematica (DMAT). Universidade Federal de Roraima (UFRR), Boa Vista, RR (Brazil); Lima, Fernando R.A., E-mail: falima@cnen.gov.br [Centro Regional de Ciencias Nucleares (CRCN/NE-CNEN-PE), Recife, PE (Brazil); Vasconcelos, Wagner E. de [Departamento de Energia Nuclear (DEN). Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil)

    2011-07-01

    One of the greatest challenges in numerical dosimetry of ionizing radiation is to estimate the absorbed dose by bone tissue in the human body. The bone tissues of greater radiosensitivity are the red bone marrow (RBM), that consist of the hematopoietic cells, located within the trabecular bones, and the bone surface cells (BSC), called osteogenic cells. The report 70 of the ICRP lists five spongiosa regions with their respective volume percent of trabecular bone: ribs (also contemplating the clavicles and sternum), spine, long bones, pelvis and skull (also contemplating mandible). The Grupo de Pesquisa em Dosimetria Numerica (GDN/CNPq) has been built exposure computational models (ECMs) based on voxel phantoms and EGSnrc Monte Carlo code. To estimate the energy deposited in the RBM and in the BSC of a phantom, the GDN/CNPq has used a method based on micro-CT images of the five trabecular regions mentioned above. These images were provided by other research institutes and were obtained from scan of bone samples of adult. Here is the greatest difficulty in reproducing this method: besides the need for bone images of real people with micrometer resolution, the distribution of bone marrow in the human body, according to ICRP 70, varies with age. This article presents some proposals of the GDN/CNPQ for replacing in the ECMs the micro-CT images by images synthesized by the computer, based on Monte Carlo sampling. (author)

  8. Bone marrow blood vessel ossification and "microvascular dead space" in rat and human long bone.

    Science.gov (United States)

    Prisby, Rhonda D

    2014-07-01

    Severe calcification of the bone microvascular network was observed in rats, whereby the bone marrow blood vessels appeared ossified. This study sought to characterize the magnitude of ossification in relation to patent blood vessels and adipocyte content in femoral diaphyses. Additionally, this study confirmed the presence of ossified vessels in patients with arteriosclerotic vascular disease and peripheral vascular disease and cellulitis. Young (4-6 month; n=8) and old (22-24 month; n=8) male Fischer-344 rats were perfused with barium sulfate to visualize patent bone marrow blood vessels. Femoral shafts were processed for bone histomorphometry to quantify ossified (Goldner's Trichrome) and calcified (Alizarin Red) vessels. Adipocyte content was also determined. Additional femora (n=5/age group) were scanned via μCT to quantify microvascular ossification. Bone marrow blood vessels from the rats and the human patients were also isolated and examined via microscopy. Ossified vessels (rats and humans) had osteocyte lacunae on the vessel surfaces and "normal" vessels were transitioning into bone. The volume of ossified vessels was 4800% higher (pnecrosis. Progression of bone microvascular ossification may provide the common link associated with age-related changes in bone and bone marrow. The clinical implications may be evident in the difficulties treating bone disease in the elderly. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Bone dosimetry using synthetic images to represent trabecular bones of five regions of the human body

    International Nuclear Information System (INIS)

    Lima Filho, Jose de M.; Vieira, Jose W.; Lima, Vanildo J. de M.; Lima, Lindeval F.; Lima, Fernando R.A.; Vasconcelos, Wagner E. de

    2011-01-01

    One of the greatest challenges in numerical dosimetry of ionizing radiation is to estimate the absorbed dose by bone tissue in the human body. The bone tissues of greater radiosensitivity are the red bone marrow (RBM), that consist of the hematopoietic cells, located within the trabecular bones, and the bone surface cells (BSC), called osteogenic cells. The report 70 of the ICRP lists five spongiosa regions with their respective volume percent of trabecular bone: ribs (also contemplating the clavicles and sternum), spine, long bones, pelvis and skull (also contemplating mandible). The Grupo de Pesquisa em Dosimetria Numerica (GDN/CNPq) has been built exposure computational models (ECMs) based on voxel phantoms and EGSnrc Monte Carlo code. To estimate the energy deposited in the RBM and in the BSC of a phantom, the GDN/CNPq has used a method based on micro-CT images of the five trabecular regions mentioned above. These images were provided by other research institutes and were obtained from scan of bone samples of adult. Here is the greatest difficulty in reproducing this method: besides the need for bone images of real people with micrometer resolution, the distribution of bone marrow in the human body, according to ICRP 70, varies with age. This article presents some proposals of the GDN/CNPQ for replacing in the ECMs the micro-CT images by images synthesized by the computer, based on Monte Carlo sampling. (author)

  10. The effect of radiation sterilization on human transplantable bone

    International Nuclear Information System (INIS)

    Triantafyllou, N.; Karatzas, P.

    1974-11-01

    In order to study the effect of radiation sterilization on human transplantable bones, work was carried out on human and bovine bone tissue samples. Factors causing possible alterations in the mechanical structures of the preserved bone allografts were considered to be deep freezing (-35degC), lyophylization, irradiation, or a combination of lyophylization and irradiation. The latter could be shown to lower the mechanical strength of the bone. Crystal lattice of the bone did not show any alterations in x-ray diffraction pattern, following freeze drying and/or irradiation with doses up to 10 Mrad of gamma radiation. Deterioration in mechanical properties is probably due to damage to the organic phase of the bone matrix

  11. Otosclerosis: Temporal Bone Pathology.

    Science.gov (United States)

    Quesnel, Alicia M; Ishai, Reuven; McKenna, Michael J

    2018-04-01

    Otosclerosis is pathologically characterized by abnormal bony remodeling, which includes bone resorption, new bone deposition, and vascular proliferation in the temporal bone. Sensorineural hearing loss in otosclerosis is associated with extension of otosclerosis to the cochlear endosteum and deposition of collagen throughout the spiral ligament. Persistent or recurrent conductive hearing loss after stapedectomy has been associated with incomplete footplate fenestration, poor incus-prosthesis connection, and incus resorption in temporal bone specimens. Human temporal bone pathology has helped to define the role of computed tomography imaging for otosclerosis, confirming that computed tomography is highly sensitive for diagnosis, yet limited in assessing cochlear endosteal involvement. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Elastin and collagen fibre microstructure of the human aorta in ageing and disease: a review

    Science.gov (United States)

    Tsamis, Alkiviadis; Krawiec, Jeffrey T.; Vorp, David A.

    2013-01-01

    Aortic disease is a significant cause of death in developed countries. The most common forms of aortic disease are aneurysm, dissection, atherosclerotic occlusion and ageing-induced stiffening. The microstructure of the aortic tissue has been studied with great interest, because alteration of the quantity and/or architecture of the connective fibres (elastin and collagen) within the aortic wall, which directly imparts elasticity and strength, can lead to the mechanical and functional changes associated with these conditions. This review article summarizes the state of the art with respect to characterization of connective fibre microstructure in the wall of the human aorta in ageing and disease, with emphasis on the ascending thoracic aorta and abdominal aorta where the most common forms of aortic disease tend to occur. PMID:23536538

  13. Proportion of collagen type II in the extracellular matrix promotes the differentiation of human adipose-derived mesenchymal stem cells into nucleus pulposus cells.

    Science.gov (United States)

    Tao, Yiqing; Zhou, Xiaopeng; Liu, Dongyu; Li, Hao; Liang, Chengzhen; Li, Fangcai; Chen, Qixin

    2016-01-01

    During degeneration process, the catabolism of collagen type II and anabolism of collagen type I in nucleus pulposus (NP) may influence the bioactivity of transplanted cells. Human adipose-derived mesenchymal stem cells (hADMSCs) were cultured as a micromass or in a series of gradual proportion hydrogels of a mix of collagen types I and II. Cell proliferation and cytotoxicity were detected using CCK-8 and LDH assays respectively. The expression of differentiation-related genes and proteins, including SOX9, aggrecan, collagen type I, and collagen type II, was examined using RT-qPCR and Western blotting. Novel phenotypic genes were also detected by RT-qPCR and western blotting. Alcian blue and dimethylmethylene blue assays were used to investigate sulfate proteoglycan expression, and PI3K/AKT, MAPK/ERK, and Smad signaling pathways were examined by Western blotting. The results showed collagen hydrogels have good biocompatibility, and cell proliferation increased after collagen type II treatment. Expressions of SOX9, aggrecan, and collagen type II were increased in a collagen type II dependent manner. Sulfate proteoglycan synthesis increased in proportion to collagen type II concentration. Only hADMSCs highly expressed NP cell marker KRT19 in collagen type II culture. Additionally, phosphorylated Smad3, which is associated with phosphorylated ERK, was increased after collagen type II-stimulation. The concentration and type of collagen affect hADMSC differentiation into NP cells. Collagen type II significantly ameliorates hADMSC differentiation into NP cells and promotes extracellular matrix synthesis. Therefore, anabolism of collagen type I and catabolism of type II may attenuate the differentiation and biosynthesis of transplanted stem cells. © 2016 International Union of Biochemistry and Molecular Biology.

  14. Analyses of stable isotopes in camelids collagen bones from Tulan Ravine, Atacama Puna, early formative period (CA 3,1000-2,400BP)

    International Nuclear Information System (INIS)

    Lopez, Patricio; Cartajena, Isabel; Nunez, Lautaro

    2013-01-01

    This paper presents the results of isotope analysis (δ 13 C y δ 15 N) conducted on bone collagen found in Lama guanicoe and Lama glama remains from Tulan-85 and Tulan-54 archaeological sites. Both sites have been dated to the Early Formative Period (ca. 3,100-2,400 ap) and are located southeast of the Atacama Puna basin. Faunal samples were selected using anatomical and morphometric criteria. The results indicate divergences in the diets of both species, reflecting vegetation variation in the Tulan Quebrada caused by altitude differences and linked to hunting and herding areas [es

  15. Comparative cortical bone thickness between the long bones of humans and five common non-human mammal taxa.

    Science.gov (United States)

    Croker, Sarah L; Reed, Warren; Donlon, Denise

    2016-03-01

    The task of identifying fragments of long bone shafts as human or non-human is difficult but necessary, for both forensic and archaeological cases, and a fast simple method is particularly useful. Previous literature suggests there may be differences in the thickness of the cortical bone between these two groups, but this has not been tested thoroughly. The aim of this study was not only to test this suggestion, but also to provide data that could be of practical assistance for future comparisons. The major limb bones (humerus, radius, femur and tibia) of 50 Caucasoid adult skeletons of known age and sex were radiographed, along with corresponding skeletal elements from sheep, pigs, cattle, large dogs and kangaroos. Measurements were taken from the radiographs at five points along the bone shaft, of shaft diameter, cortical bone thickness, and a cortical thickness index (sum of cortices divided by shaft diameter) in both anteroposterior and mediolateral orientations. Each variable for actual cortical bone thickness as well as cortical thickness indices were compared between the human group (split by sex) and each of the non-human groups in turn, using Student's t-tests. Results showed that while significant differences did exist between the human groups and many of the non-human groups, these were not all in the same direction. That is, some variables in the human groups were significantly greater than, and others were significantly less than, the corresponding variable in the non-human groups, depending on the particular non-human group, sex of the human group, or variable under comparison. This was the case for measurements of both actual cortical bone thickness and cortical thickness index. Therefore, for bone shaft fragments for which the skeletal element is unknown, the overlap in cortical bone thickness between different areas of different bones is too great to allow identification using this method alone. However, by providing extensive cortical bone

  16. Lower strength of the human posterior patellar tendon seems unrelated to mature collagen cross-linking and fibril morphology

    DEFF Research Database (Denmark)

    Hansen, Philip; Haraldsson, Bjarki Thor; Aagaard, Per

    2010-01-01

    The human patellar tendon is frequently affected by tendinopathy, but the etiology of the condition is not established, although differential loading of the anterior and posterior tendon may be associated with the condition. We hypothesized that changes in fibril morphology and collagen cross-lin...

  17. In vitro tendon tissue development from human fibroblasts demonstrates collagen fibril diameter growth associated with a rise in mechanical strength

    DEFF Research Database (Denmark)

    Herchenhan, Andreas; Bayer, Monika L; Svensson, René B

    2013-01-01

    Collagen-rich tendons and ligaments are important for joint stability and force transmission, but the capacity to form new tendon is poorly understood. In the present study, we investigated mechanical strength, fibril size, and structure during development of tendon-like tissue from adult human...

  18. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    Tendon fibroblasts synthesize collagen and form fibrils during embryonic development, but to what extent mature fibroblasts are able to recapitulate embryonic development and develop normal tendon structure is unknown. The present study examined the capability of mature human tendon fibroblasts t...

  19. Cytocompatibility and biologic characteristics of synthetic scaffold materials of rabbit acellular vascular matrix combining with human-like collagen I.

    Science.gov (United States)

    Liu, Xuqian; Wang, Jie; Dong, Fusheng; Song, Peng; Tian, Songbo; Li, Hexiang; Hou, Yali

    2017-10-01

    Scaffold material provides a three-dimensional growing environment for seed cells in the research field of tissue engineering. In the present study, rabbit arterial blood vessel cells were chemically removed with trypsin and Triton X-100 to prepare rabbit acellular vascular matrix scaffold material. Observation by He&Masson staining revealed that no cellular components or nuclei existed in the vascular intima and media after decellularization. Human-like collagen I was combined with acellular vascular matrix by freeze-drying to prepare an acellular vascular matrix-0.25% human-like collagen I scaffold to compensate for the extracellular matrix loss during the decellularization process. We next performed a series of experiments to test the water absorbing quality, biomechanics, pressure resistance, cytotoxicity, and ultra-micro structure of the acellular vascular matrix composite material and natural rabbit artery and found that the acellular vascular matrix-0.25% human-like collagen I material behaved similarly to natural rabbit artery. In conclusion, the acellular vascular matrix-0.25% human-like collagen I composite material provides a new approach and lays the foundation for novel scaffold material research into tissue engineering of blood vessels.

  20. Age-related liquefaction of the human vitreous body : LM and TEM evaluation of the role of proteoglycans and collagen

    NARCIS (Netherlands)

    Los, Leonoor I.; van der Worp, Roelofje J; van Luyn, Marja J. A.; Hooymans, Johanna M. M.

    PURPOSE. To evaluate morphologic aspects of age-related liquefaction of the human vitreous body by fight and electron microscopy to provide a basis from which future studies directed at the pathogenesis of this phenomenon can be undertaken. The study focuses on changes in fibrillar Collagen and

  1. Collagen turnover in normal and degenerate human intervertebral discs as determined by the racemization of aspartic acid

    NARCIS (Netherlands)

    Sivan, S.-S.; Wachtel, E.; Tsitron, E.; Sakkee, N.; Ham, F. van der; Groot, J.de; Roberts, S.; Maroudas, A.

    2008-01-01

    Knowledge of rates of protein turnover is important for a quantitative understanding of tissue synthesis and catabolism. In this work, we have used the racemization of aspartic acid as a marker for the turnover of collagen obtained from healthy and pathological human intervertebral disc matrices. We

  2. Neurotrophin 3 upregulates proliferation and collagen production in human aortic valve interstitial cells: a potential role in aortic valve sclerosis.

    Science.gov (United States)

    Yao, Qingzhou; Song, Rui; Ao, Lihua; Cleveland, Joseph C; Fullerton, David A; Meng, Xianzhong

    2017-06-01

    Calcific aortic valve disease (CAVD) is a leading cardiovascular disorder in the elderly. Diseased aortic valves are characterized by sclerosis (fibrosis) and nodular calcification. Sclerosis, an early pathological change, is caused by aortic valve interstitial cell (AVIC) proliferation and overproduction of extracellular matrix (ECM) proteins. However, the mechanism of aortic valve sclerosis remains unclear. Recently, we observed that diseased human aortic valves overexpress growth factor neurotrophin 3 (NT3). In the present study, we tested the hypothesis that NT3 is a profibrogenic factor to human AVICs. AVICs isolated from normal human aortic valves were cultured in M199 growth medium and treated with recombinant human NT3 (0.10 µg/ml). An exposure to NT3 induced AVIC proliferation, upregulated the production of collagen and matrix metalloproteinase (MMP), and augmented collagen deposition. These changes were abolished by inhibition of the Trk receptors. NT3 induced Akt phosphorylation and increased cyclin D1 protein levels in a Trk receptor-dependent fashion. Inhibition of Akt abrogated the effect of NT3 on cyclin D1 production. Furthermore, inhibition of either Akt or cyclin D1 suppressed NT3-induced cellular proliferation and MMP-9 and collagen production, as well as collagen deposition. Thus, NT3 upregulates cellular proliferation, ECM protein production, and collagen deposition in human AVICs. It exerts these effects through the Trk-Akt-cyclin D1 cascade. NT3 is a profibrogenic mediator in human aortic valve, and overproduction of NT3 by aortic valve tissue may contribute to the mechanism of valvular sclerosis. Copyright © 2017 the American Physiological Society.

  3. Registration-based Bone Morphometry for Shape Analysis of the Bones of the Human Wrist

    Science.gov (United States)

    Joshi, Anand A.; Leahy, Richard M.; Badawi, Ramsey D.; Chaudhari, Abhijit J.

    2015-01-01

    We present a method that quantifies point-wise changes in surface morphology of the bones of the human wrist. The proposed method, referred to as Registration-based Bone Morphometry (RBM), consists of two steps: an atlas selection step and an atlas warping step. The atlas for individual wrist bones was selected based on the shortest l2 distance to the ensemble of wrist bones from a database of a healthy population of subjects. The selected atlas was then warped to the corresponding bones of individuals in the population using a non-linear registration method based on regularized l2 distance minimization. The displacement field thus calculated showed local differences in bone shape that then were used for the analysis of group differences. Our results indicate that RBM has potential to provide a standardized approach to shape analysis of bones of the human wrist. We demonstrate the performance of RBM for examining group differences in wrist bone shapes based on sex and between those of the right and left wrists in healthy individuals. We also present data to show the application of RBM for tracking bone erosion status in rheumatoid arthritis. PMID:26353369

  4. Type I Collagen and Strontium-Containing Mesoporous Glass Particles as Hybrid Material for 3D Printing of Bone-Like Materials.

    Science.gov (United States)

    Montalbano, Giorgia; Fiorilli, Sonia; Caneschi, Andrea; Vitale-Brovarone, Chiara

    2018-04-28

    Bone tissue engineering offers an alternative promising solution to treat a large number of bone injuries with special focus on pathological conditions, such as osteoporosis. In this scenario, the bone tissue regeneration may be promoted using bioactive and biomimetic materials able to direct cell response, while the desired scaffold architecture can be tailored by means of 3D printing technologies. In this context, our study aimed to develop a hybrid bioactive material suitable for 3D printing of scaffolds mimicking the natural composition and structure of healthy bone. Type I collagen and strontium-containing mesoporous bioactive glasses were combined to obtain suspensions able to perform a sol-gel transition under physiological conditions. Field emission scanning electron microscopy (FESEM) analyses confirmed the formation of fibrous nanostructures homogeneously embedding inorganic particles, whereas bioactivity studies demonstrated the large calcium phosphate deposition. The high-water content promoted the strontium ion release from the embedded glass particles, potentially enhancing the osteogenic behaviour of the composite. Furthermore, the suspension printability was assessed by means of rheological studies and preliminary extrusion tests, showing shear thinning and fast material recovery upon deposition. In conclusion, the reported results suggest that promising hybrid systems suitable for 3D printing of bioactive scaffolds for bone tissue engineering have been developed.

  5. Type II collagen C2C epitope in human synovial fluid and serum after knee injury

    DEFF Research Database (Denmark)

    Kumahashi, N; Swärd, P; Larsson, S

    2015-01-01

    PURPOSE: Investigate in a cross-sectional study time-dependent changes of synovial fluid type II collagen epitope C2C concentrations after knee injury and correlate to other joint injury biomarkers. METHODS: Synovial fluid samples were aspirated between 0 days and 7 years after injury (n = 235...... = 0.403, P type II collagen (r = 0.444, P = 0.003), ARGS-aggrecan (r = 0.337, P ... with an immediate and sustained local degradation of type II collagen....

  6. Sandwich ELISA for quantitative detection of human collagen prolyl 4-hydroxylase

    Directory of Open Access Journals (Sweden)

    Myllyharju Johanna

    2010-06-01

    Full Text Available Abstract Background We describe a method for specific, quantitative and quick detection of human collagen prolyl 4-hydroxylase (C-P4H, the key enzyme for collagen prolyl-4 hydroxylation, in crude samples based on a sandwich ELISA principle. The method is relevant to active C-P4H level monitoring during recombinant C-P4H and collagen production in different expression systems. The assay proves to be specific for the active C-P4H α2β2 tetramer due to the use of antibodies against its both subunits. Thus in keeping with the method C-P4H is captured by coupled to an anti-α subunit antibody magnetic beads and an anti-β subunit antibody binds to the PDI/β subunit of the protein. Then the following holoenzyme detection is accomplished by a goat anti-rabbit IgG labeled with alkaline phosphatase which AP catalyzes the reaction of a substrate transformation with fluorescent signal generation. Results We applied an experimental design approach for the optimization of the antibody concentrations used in the sandwich ELISA. The assay sensitivity was 0.1 ng of C-P4H. The method was utilized for the analysis of C-P4H accumulation in crude cell extracts of E. coli overexpressing C-P4H. The sandwich ELISA signals obtained demonstrated a very good correlation with the detected protein activity levels measured with the standard radioactive assay. The developed assay was applied to optimize C-P4H production in E. coli Origami in a system where the C-P4H subunits expression acted under control by different promoters. The experiments performed in a shake flask fed-batch system (EnBase® verified earlier observations that cell density and oxygen supply are critical factors for the use of the inducer anhydrotetracycline and thus for the soluble C-P4H yield. Conclusions Here we show an example of sandwich ELISA usage for quantifying multimeric proteins. The method was developed for monitoring the amount of recombinant C-P4H tetramer in crude E. coli extracts. Due

  7. Effect of Tris-acetate buffer on endotoxin removal from human-like collagen used biomaterials

    International Nuclear Information System (INIS)

    Zhang, Huizhi; Fan, Daidi; Deng, Jianjun; Zhu, Chenghui; Hui, Junfeng; Ma, Xiaoxuan

    2014-01-01

    Protein preparation, which has active ingredients designated for the use of biomaterials and therapeutical protein, is obtained by genetic engineering, but products of genetic engineering are often contaminated by endotoxins. Because endotoxin is a ubiquitous and potent proinflammatory agent, endotoxin removal or depletion from protein is essential for researching any biomaterials. In this study, we have used Tris-acetate (TA) buffer of neutral pH value to evaluate endotoxins absorbed on the Pierce high-capacity endotoxin removal resin. The effects of TA buffer on pH, ionic strength, incubation time as well as human-like collagen (HLC) concentration on eliminating endotoxins are investigated. In the present experiments, we design an optimal method for TA buffer to remove endotoxin from recombinant collagen and use a chromogenic tachypleus amebocyte lysate (TAL) test kit to measure the endotoxin level of HLC. The present results show that, the endotoxins of HLC is dropped to 8.3 EU/ml at 25 mM TA buffer (pH 7.8) with 150 mM NaCl when setting incubation time at 6 h, and HLC recovery is about 96%. Under this experimental condition, it is proved to exhibit high efficiencies of both endotoxin removal and collagen recovery. The structure of treated HLC was explored by Transmission Electron Microscopy (TEM), demonstrating that the property and structure of HLC treated by TA buffer are maintained. Compared to the most widely used endotoxin removal method, Triton X-114 extraction, using TA buffer can obtain the non-toxic HLC without extra treatment for removing the toxic substances in Triton X-114. In addition, the present study aims at establishing a foundation for further work in laboratory animal science and providing a foundation for medical grade biomaterials. - Graphical abstract: The processes of endotoxins adsorbed from HLC. - Highlights: • TA buffer is a mild buffer system for endotoxins removal of HLC. • TA buffer may facilitate endotoxins adsorbed on the

  8. Correlation of collagen synthesis with polarization-sensitive optical coherence tomography imaging of in vitro human atherosclerosis

    Science.gov (United States)

    Kuo, Wen-Chuan; Shyu, Jeou-Jong; Chou, Nai-Kuan; Lai, Chih-Ming; Tien, En-Kuang; Huang, Huan-Jang; Chou, Chien; Jan, Gwo-Jen

    2005-04-01

    Atherosclerosis is unquestionably the leading cause of morbidity and mortality in developed countries. In the mean time, the worldwide importance of acute vascular syndromes is increasing. Because collagen fiber is a critical component of atherosclerotic lesions; it constitutes up to 60% of the total atherosclerotic plaque protein. The uncontrolled collagen accumulation leads to arterial stenosis, whereas excessive collagen breakdown weakens plaques thereby making them prone to rupture finally. Thus, in this study, we present the first application, to our knowledge, of using polarization-sensitive optical coherence tomography (PS-OCT) in human atherosclerosis. We demonstrate this technique for imaging of intensity, birefringence, and fast-axis orientation simultaneously in atherosclerotic plaques. This in vitro study suggests that the birefringence change in plaque is due to the prominent deposition of collagen according to the correlation of PS-OCT images with histological counterpart. Moreover, we can acquire quantitative criteria based on the change of polarization of incident beam to estimate whether the collagen synthesized is "too much" or "not enough". Thus by combining of high resolution intensity imaging and birefringence detection makes PS-OCT could be a potentially powerful tool for early assessment of atherosclerosis appearance and the prediction of plaque rupture in clinic.

  9. Investigation of the effect of hydration on dermal collagen in ex vivo human skin tissue using second harmonic generation microscopy

    Science.gov (United States)

    Samatham, Ravikant; Wang, Nicholas K.; Jacques, Steven L.

    2016-02-01

    Effect of hydration on the dermal collagen structure in human skin was investigated using second harmonic generation microscopy. Dog ears from the Mohs micrographic surgery department were procured for the study. Skin samples with subject aged between 58-90 years old were used in the study. Three dimensional Multiphoton (Two-photon and backward SHG) control data was acquired from the skin samples. After the control measurement, the skin tissue was either soaked in deionized water for 2 hours (Hydration) or kept at room temperature for 2 hours (Desiccation), and SHG data was acquired. The data was normalized for changes in laser power and detector gain. The collagen signal per unit volume from the dermis was calculated. The desiccated skin tissue gave higher backward SHG compared to respective control tissue, while hydration sample gave a lower backward SHG. The collagen signal decreased with increase in hydration of the dermal collagen. Hydration affected the packing of the collagen fibrils causing a change in the backward SHG signal. In this study, the use of multiphoton microscopy to study the effect of hydration on dermal structure was demonstrated in ex vivo tissue.

  10. Evaluation of autogenous PRGF+β-TCP with or without a collagen membrane on bone formation and implant osseointegration in large size bone defects. A preclinical in vivo study.

    Science.gov (United States)

    Batas, Leonidas; Stavropoulos, Andreas; Papadimitriou, Serafim; Nyengaard, Jens R; Konstantinidis, Antonios

    2016-08-01

    The aim of this study was to evaluate whether the adjunctive use of a collagen membrane enhances bone formation and implant osseointegration in non-contained defects grafted with chair-side prepared autologous platelet-rich growth factor (PRGF) adsorbed on a β-TCP particulate carrier. Large box-type defects (10 × 6 mm; W × D) were prepared in the edentulated and completely healed mandibles of six Beagles dogs. An implant with moderately rough surface was placed in the center of each defect leaving the coronal 6 mm of the implant not covered with bone. The remaining defect space was then filled out with chair-side prepared autologous PRGF adsorbed on β-TCP particles and either covered with a collagen membrane (PRGF/β-TCP+CM) (6 defects) or left without a membrane (PRGF/β-TCP) (5 defects). Histology 4 months post-op showed new lamellar and woven bone formation encompassing almost entirely the defect and limited residual β-TCP particles. Extent of osseointegration of the previously exposed portion of the implants varied, but in general was limited. Within the defect, new mineralized bone (%) averaged 43.2 ± 9.86 vs. 39.9 ± 13.7 in the PRGF/β-TCP+CM and PRGF/β-TCP group (P = 0.22) and relative mineralized bone-to-implant contact (%) averaged 26.2 ± 16.45 vs. 35.91 ± 24.45, respectively (P = 0.5). First, bone-to-implant contact from the implant top was 4.1 ± 1.5 and 3.2 ± 2.3 (P = 0.9), in the PRGF/β-TCP+CM and PRGF/β-TCP group, respectively. Implantation of chair-side prepared autologous PRGF adsorbed on a β-TCP carrier in non-contained peri-implant defects resulted in large amounts of bone regeneration, but osseointegration was limited. Provisions for GBR with a collagen membrane did not significantly enhance bone regeneration or implant osseointegration. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. The carboxy-terminal pyridinoline cross-linked telopeptide of type I collagen in serum as a marker of bone resorption

    DEFF Research Database (Denmark)

    Hassager, C; Jensen, L T; Pødenphant, J

    1994-01-01

    Carboxy-terminal pyridinoline cross-linked telopeptide of type I collagen (ICTP) in serum has recently been proposed as a new biochemical marker of bone resorption. In the present study we compared serum ICTP with radiopharmaceutical and histomorphometric measurements of bone turnover...... in postmenopausal women with mild osteoporosis, and assessed the effect of hormone replacement therapy (HRT) (2 mg 17 beta-estradiol plus 1 mg norethisterone daily) and anabolic steroid therapy (50 mg nandrolone decanoate (ND) i.m. every 3 weeks) on serum ICTP in two double-blind placebo-controlled studies with 55...... to 75-year-old women. Serum ICTP measured by radioimmunoassay (RIA) correlated significantly with the 24-hour whole body retention of 99m-technetium diphosphonate (Rho = 0.47, P

  12. Age-related changes in the plasticity and toughness of human cortical bone at multiple length-scales

    Energy Technology Data Exchange (ETDEWEB)

    Zimmermann, Elizabeth A.; Schaible, Eric; Bale, Hrishikesh; Barth, Holly D.; Tang, Simon Y.; Reichert, Peter; Busse, Bjoern; Alliston, Tamara; Ager III, Joel W.; Ritchie, Robert O.

    2011-08-10

    The structure of human cortical bone evolves over multiple length-scales from its basic constituents of collagen and hydroxyapatite at the nanoscale to osteonal structures at nearmillimeter dimensions, which all provide the basis for its mechanical properties. To resist fracture, bone’s toughness is derived intrinsically through plasticity (e.g., fibrillar sliding) at structural-scales typically below a micron and extrinsically (i.e., during crack growth) through mechanisms (e.g., crack deflection/bridging) generated at larger structural-scales. Biological factors such as aging lead to a markedly increased fracture risk, which is often associated with an age-related loss in bone mass (bone quantity). However, we find that age-related structural changes can significantly degrade the fracture resistance (bone quality) over multiple lengthscales. Using in situ small-/wide-angle x-ray scattering/diffraction to characterize sub-micron structural changes and synchrotron x-ray computed tomography and in situ fracture-toughness measurements in the scanning electron microscope to characterize effects at micron-scales, we show how these age-related structural changes at differing size-scales degrade both the intrinsic and extrinsic toughness of bone. Specifically, we attribute the loss in toughness to increased non-enzymatic collagen cross-linking which suppresses plasticity at nanoscale dimensions and to an increased osteonal density which limits the potency of crack-bridging mechanisms at micron-scales. The link between these processes is that the increased stiffness of the cross-linked collagen requires energy to be absorbed by “plastic” deformation at higher structural levels, which occurs by the process of microcracking.

  13. The effects of collagen-rich extracellular matrix on the intracellular delivery of glycol chitosan nanoparticles in human lung fibroblasts.

    Science.gov (United States)

    Yhee, Ji Young; Yoon, Hong Yeol; Kim, Hyunjoon; Jeon, Sangmin; Hergert, Polla; Im, Jintaek; Panyam, Jayanth; Kim, Kwangmeyung; Nho, Richard Seonghun

    2017-01-01

    Recent progress in nanomedicine has shown a strong possibility of targeted therapy for obstinate chronic lung diseases including idiopathic pulmonary fibrosis (IPF). IPF is a fatal lung disease characterized by persistent fibrotic fibroblasts in response to type I collagen-rich extracellular matrix. As a pathological microenvironment is important in understanding the biological behavior of nanoparticles, in vitro cellular uptake of glycol chitosan nanoparticles (CNPs) in human lung fibroblasts was comparatively studied in the presence or absence of type I collagen matrix. Primary human lung fibroblasts from non-IPF and IPF patients (n=6/group) showed significantly increased cellular uptake of CNPs (>33.6-78.1 times) when they were cultured on collagen matrix. To elucidate the underlying mechanism of enhanced cellular delivery of CNPs in lung fibroblasts on collagen, cells were pretreated with chlorpromazine, genistein, and amiloride to inhibit clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis, respectively. Amiloride pretreatment remarkably reduced the cellular uptake of CNPs, suggesting that lung fibroblasts mainly utilize the macropinocytosis-dependent mechanism when interacted with collagen. In addition, the internalization of CNPs was predominantly suppressed by a phosphoinositide 3-kinase (PI3K) inhibitor in IPF fibroblasts, indicating that enhanced PI3K activity associated with late-stage macropinocytosis can be particularly important for the enhanced cellular delivery of CNPs in IPF fibroblasts. Our study strongly supports the concept that a pathological microenvironment which surrounds lung fibroblasts has a significant impact on the intracellular delivery of nanoparticles. Based on the property of enhanced intracellular delivery of CNPs when fibroblasts are made to interact with a collagen-rich matrix, we suggest that CNPs may have great potential as a drug-carrier system for targeting fibrotic lung fibroblasts.

  14. Early effects of zoledronic acid and teriparatide on bone microarchitecture, remodeling and collagen crosslinks: comparison between iliac crest and lumbar vertebra in ewes.

    Science.gov (United States)

    Portero-Muzy, N R; Chavassieux, P M; Bouxsein, M L; Gineyts, E; Garnero, P; Chapurlat, R D

    2012-10-01

    Iliac crest bone biopsies are used to assess the mechanism of action of drug treatments, yet there are little data comparing this site to sites prone to fracture. The purpose of this study was to compare the delay and the amplitude of responses to treatment in two different bone sites. The short-term effects of zoledronic acid and teriparatide on microarchitecture, collagen crosslinks and bone remodeling were evaluated in iliac crest and lumbar vertebrae. Aged ewes (n=8/gr) received either vehicle (CTRL) or a single injection of zoledronic acid (ZOL, 10mg) or daily injections of teriparatide (TPTD, 20 μg/d) for 3 months. Blood samples were collected monthly for assessing bone turnover markers. At the end of the study, a transiliac bone biopsy (IC) and L1 lumbar vertebrae (LV1) were collected to assess bone microarchitecture; pyridinoline (PYD), deoxypyridinoline (DPD), pentosidine (PEN) content, static and dynamic parameters of bone remodeling. In CTRL, Tb-BV/TV was significantly higher in LV1 than IC (psALP (p<0.001) and sCTX (p<0.001) were observed in the ZOL-group whereas in TPTD-group, after transient increases, they returned to baseline values. When compared to their respective CTRL, ZOL induced significant increases in Tb.BV/TV, Conn.D, Tb.N and Tb.Sp, in IC but not in LV1. Regardless of the site, ZOL markedly depressed the bone turnover: The static parameters of bone formation significantly decreased and the diminution of MS/BS, BFR/BS and Ac.f varied from -94 to -98% vs CTRL (p<0.01 to 0.001). It was associated with a diminution of the DPD content and the PYD/DPD ratio mainly in IC cortices. In contrast, after 3 months, TPTD did not modify the 3D structure and microarchitecture in IC and LV1, except a trend of higher Conn.D in IC, compared to IC-CTRL. TPTD treatment induced a significant increase in cortical porosity in LV1 (p<0.05) when compared to LV1-CTRL. Static parameters of bone formation and resorption were augmented in both sites, significantly

  15. Collagen VII deficient mice show morphologic and histologic corneal changes that phenotypically mimic human dystrophic epidermolysis bullosa of the eye.

    Science.gov (United States)

    Chen, Vicki M; Shelke, Rajani; Nyström, Alexander; Laver, Nora; Sampson, James F; Zhiyi, Cao; Bhat, Najma; Panjwani, Noorjahan

    2018-06-16

    Absence of collagen VII causes blistering of the skin, eyes and many other tissues. This disease is termed dystrophic epidermolysis bullosa (DEB). Corneal fibrosis occurs in up to 41% and vision loss in up to 64% of patients. Standard treatments are supportive and there is no cure. The immune-histologic and morphologic changes in the corneas of the mouse model for this disease have not been described in the literature. Our purpose is to characterize the eyes of these mice to determine if this is an appropriate model for study of human therapeutics. Western blot analysis (WB) and immunohistochemistry (IHC) were performed to assess the relative collagen VII protein levels and its location within the cornea. Additional IHC for inflammatory and fibrotic biomarkers alpha-smooth muscle actin (α-SMA), transforming growth factor-beta (TGF-β), connective tissue growth factor (CTGF), proteinase 3, tenascin C and collagen III were performed. Clinical photographs documenting opacification of the corneas of animals of differing ages were assessed and scored independently by 2 examiners. Histology was then used to investigate morphologic changes. IHC and WB confirmed that these mice are deficient in collagen VII production at the level of the basement membrane when compared with wild-types. IHC showed anomalous deposition of collagen III throughout the stroma. Of the 5 biomarkers tested, TGF-β showed the strongest and most consistently staining. Photographs documented corneal opacities only in mice older than 10 weeks, opacities were not seen in younger animals. Histology showed multiple abnormalities, including epithelial hyperplasia, ulceration, fibrosis, edema, dysplasia, neovascularization and bullae formation. The collagen VII hypomorphic mouse shows reduced collagen VII production at the level of the corneal basement membrane. Corneal changes are similar to pathology seen in humans with this disease. The presence of anomalous stromal collagen III and TGF-β appear to be

  16. Telomerase reverse transcriptase mediated immortalization of human bone marrow stromal cells

    Directory of Open Access Journals (Sweden)

    Yong Teng

    2014-02-01

    Full Text Available Primary human bone marrow stromal cells (hMSCs were transfected with human telomerase reverse transcriptase (hTERT gene with lipofection method. The hTERT transfected hMSCs of passage 100 underwent chondrogenesis induction with dexamethasone, transforming the growth factor β and vitamin C, osteogenesis induction with dexamethasone, β glycerophosphoric acid and vitamin C, and cardiomyocyte induction with 5-azacytidine. After 7, 14, 21 and 28 days of induction, immunocytochemistry was performed to detect the expressions of type I and II collagen and osteocalcin, and alizarin red staining was performed to detect the bone nodule formation in osteogenesis induction. Immunocytochemistry was carried out to detect the striated muscle actin expression in cardiomyocytes. The hMSCs undergoing successful transfection were positive for the hTERT. The hTERT transfected cells were grown in vitro successfully and passaged for 136 generations. Results showed that these cells could be induced to differentiate into chondrocytes, bone and myocardial cells. Introduction of exogenous hTERT into hMSCs could achieve immortalized hMSCs with the potential of multi-directional differentiation. Thus, these cells could be applied as seed cells in tissue engineering.

  17. Details of the Collagen and Elastin Architecture in the Human Limbal Conjunctiva, Tenon's Capsule and Sclera Revealed by Two-Photon Excited Fluorescence Microscopy.

    Science.gov (United States)

    Park, Choul Yong; Marando, Catherine M; Liao, Jason A; Lee, Jimmy K; Kwon, Jiwon; Chuck, Roy S

    2016-10-01

    To investigate the architecture and distribution of collagen and elastin in human limbal conjunctiva, Tenon's capsule, and sclera. The limbal conjunctiva, Tenon's capsule, and sclera of human donor corneal buttons were imaged with an inverted two-photon excited fluorescence microscope. No fixation process was necessary. The laser (Ti:sapphire) was tuned at 850 nm for two-photon excitation. Backscatter signals of second harmonic generation (SHG) and autofluorescence (AF) were collected through a 425/30-nm and a 525/45-nm emission filter, respectively. Multiple, consecutive, and overlapping (z-stack) images were acquired. Collagen signals were collected with SHG, whereas elastin signals were collected with AF. The size and density of collagen bundles varied widely depending on depth: increasing from conjunctiva to sclera. In superficial image planes, collagen bundles were image planes (episclera and superficial sclera), collagen bundles were thicker (near 100 μm in width) and densely packed. Comparatively, elastin fibers were thinner and sparse. The orientation of elastin fibers was independent of collagen fibers in superficial layers; but in deep sclera, elastin fibers wove through collagen interbundle gaps. At the limbus, both collagen and elastin fibers were relatively compact and were distributed perpendicular to the limbal annulus. Two-photon excited fluorescence microscopy has enabled us to understand in greater detail the collagen and elastin architecture of the human limbal conjunctiva, Tenon's capsule, and sclera.

  18. A clinical stydy on the effectiveness of slow - resorbing collagen membrane barrier therapy to guide regeneration in mandibular class II furcations in human

    Directory of Open Access Journals (Sweden)

    Abolfazli N

    1998-09-01

    Full Text Available The present clinical trial was designed to evaluate the regenerative potential of periodontal tissues in degree II"nfurcation defects at mandibular molars of human using a slow-resorbing collagen membrane and a surgical treatment"ntechnique based on the principles of guided tissue regeneration."nThe patient sampleinclude 8 subjects who had periodontal lessions in right and left mandibular molars regions, including moderate to advance periodonal destruction within the radicular area. Following a baseline examination including recording the clinical measurements (PD, Al, HC, F.G.M , the furcation- involved molars were randomly assigned in each patient to either a test or a control treatment procedure. Included the evevation of mucoperiosteal flaps, recording measurement from the cemento enamel junction (C.E.J directly coronal to the furcation area to the alveolar crest and to the base of the defect-Horizontal furcation measurements were also made using a William's probe, finally a collagen membrrane placed on the involved area to cover the entrance of the furcation and adjucent root surfaces as well as a portion of the alveolar bone apical to the crest. The flaps were repositioned and secured with interdental sutures. A procedure identical to the one used at the test teeth was Performed at the control teeth region with the exception of the placement of the collagen membrance. Following surgery all patients were placed on a plaque control regimen. All Patients received normal postsurgical care and at 6 month post-surgery were scheduled for re-entry surgery. Before re-entry surgery all clinical parameters recorded again. The re-entry mucoperiosteal flaps were designed to expose the furcation area for measurements, as describedabove. There was clinical improvement in all measurements made in both the test and control patients (especially in test group over the 6 month period. The horizontal and vertical furcation measurements did yield a

  19. Promotion of minTBP-1-PRGDN on the attachment, proliferation and collagen I synthesis of human keratocyte on titanium

    Directory of Open Access Journals (Sweden)

    Xin-Yu Li

    2014-02-01

    Full Text Available AIM:To investigate the influence of minTBP-1-PRGDN on the attachment, proliferation and collagen I synthesis of human keratocyte on titanium (Ti surface.METHODS:The chimeric peptide RKLPDAPRGDN (minTBP-1-PRGDN was synthesized by connecting RKLPDA (minTBP-1 to the N-terminal of PRGDN , the influence of minTBP-1-PRGDN on the attachment, proliferation and collagen I synthesis of human keratocyte on Ti surface were tested using PRGDN and minTBP-1as controls. The keratocytes attached to the surface of Ti were either stained with FITC-labeled phalloidin and viewed with fluorescence microscope or quantified with alamar Blue method. The proliferation of keratocytes on Ti were quantified with 3-(4,5-dim- ethylthiazol-2-yl-2, 5-diphenyltetrazolium bromide up-taking methods. The secretion of type I collagen were determined using an ELISA kit.RESULTS:The results showed that minTBP-1-PRGDN at a concentration of 100ng/mL was the most potent peptide to enhance the attachment of human keratocytes to the surface of Ti (1.40±0.03 folds, P=0.003, to promote the proliferation (1.26±0.05 folds, P=0.014 and the synthesis of type I collagen (1.530±0.128, P=0.008. MinTBP-1 at the same concentration could only promote the attachment (1.13±0.04 folds, P=0.020 and proliferation(1.15±0.06 folds, P=0.021, while PRGDN had no significant influence (P>0.05.CONCLUSION:Our data shows that the novel chimeric peptide minTBP-1-PRGDN could promote the attachment, proliferation and type I collagen synthesis of human keratocytes on the surface of Ti.

  20. In vivo observation of age-related structural changes of dermal collagen in human facial skin using collagen-sensitive second harmonic generation microscope equipped with 1250-nm mode-locked Cr:Forsterite laser

    Science.gov (United States)

    Yasui, Takeshi; Yonetsu, Makoto; Tanaka, Ryosuke; Tanaka, Yuji; Fukushima, Shu-ichiro; Yamashita, Toyonobu; Ogura, Yuki; Hirao, Tetsuji; Murota, Hiroyuki; Araki, Tsutomu

    2013-03-01

    In vivo visualization of human skin aging is demonstrated using a Cr:Forsterite (Cr:F) laser-based, collagen-sensitive second harmonic generation (SHG) microscope. The deep penetration into human skin, as well as the specific sensitivity to collagen molecules, achieved by this microscope enables us to clearly visualize age-related structural changes of collagen fiber in the reticular dermis. Here we investigated intrinsic aging and/or photoaging in the male facial skin. Young subjects show dense distributions of thin collagen fibers, whereas elderly subjects show coarse distributions of thick collagen fibers. Furthermore, a comparison of SHG images between young and elderly subjects with and without a recent life history of excessive sun exposure show that a combination of photoaging with intrinsic aging significantly accelerates skin aging. We also perform image analysis based on two-dimensional Fourier transformation of the SHG images and extracted an aging parameter for human skin. The in vivo collagen-sensitive SHG microscope will be a powerful tool in fields such as cosmeceutical sciences and anti-aging dermatology.

  1. From mechanical loading to collagen synthesis, structural changes and function in human tendon

    DEFF Research Database (Denmark)

    Kjaer, M; Langberg, H; Heinemeier, K

    2009-01-01

    The adaptive response of connective tissue to loading requires increased synthesis and turnover of matrix proteins, with special emphasis on collagen. Collagen formation and degradation in the tendon increases with both acute and chronic loading, and data suggest that a gender difference exists...

  2. Clinical comparison of guided tissue regeneration, with collagen membrane and bone graft, versus connective tissue graft in the treatment of gingival recessions

    Directory of Open Access Journals (Sweden)

    Haghighati F

    2006-06-01

    Full Text Available Background and Aim: Increasing patient demands for esthetic, put the root coverage procedures in particular attention. Periodontal regeneration with GTR based root coverage methods is the most common treatment used. The purpose of this study was to compare guided tissue regeneration (GTR with collagen membrane and a bone graft, with sub-epithelial connective tissue graft (SCTG, in treatment of gingival recession. Materials and Methods: In this randomized clinical trial study, eleven healthy patients with no systemic diseases who had miller’s class I or II recession defects (gingival recession  2mm were treated with SCTG or GTR using a collagen membrane and a bone graft. Clinical measurements were obtained at baseline and 6 months after surgery. These clinical measurements included recession depth (RD, recession width (RW, probing depth (PD, and clinical attachment level (CAL. Data were analyzed using independent t test with p<0.05 as the limit of significance. Results: Both treatment methods resulted in a statistically significant reduction of recession depth (SCTG=2.3mm, GTR=2.1mm; P<0.0001. CAL gain after 6 months was also improved in both groups (SCG= 2.5mm, GTR=2.1mm, compared to baseline (P<0.0001. No statistical differences were observed in RD, RW, CAL between test and control groups. Root coverage was similar in both methods (SCTG= 74.2%, GTR= 62.6%, P=0.87. Conclusion: Based on the results of this study, the two techniques are clinically comparable. Therefore the use of collagen membrane and a bovine derived xenograft may alleviate the need for connective tissue graft.

  3. Coordinated collagen and muscle protein synthesis in human patella tendon and quadriceps muscle after exercise

    DEFF Research Database (Denmark)

    Miller, Benjamin F; Olesen, Jens L; Hansen, Mette

    2005-01-01

    We hypothesized that an acute bout of strenuous, non-damaging exercise would increase rates of protein synthesis of collagen in tendon and skeletal muscle but these would be less than those of muscle myofibrillar and sarcoplasmic proteins. Two groups (n = 8 and 6) of healthy young men were studied...... collagen (0.077% h(-1)), muscle collagen (0.054% h(-1)), myofibrillar protein (0.121% h(-1)), and sarcoplasmic protein (0.134% h(-1))). The rates decreased toward basal values by 72 h although rates of tendon collagen and myofibrillar protein synthesis remained elevated. There was no tissue damage...... of muscle visible on histological evaluation. Neither tissue microdialysate nor serum concentrations of IGF-I and IGF binding proteins (IGFBP-3 and IGFBP-4) or procollagen type I N-terminal propeptide changed from resting values. Thus, there is a rapid increase in collagen synthesis after strenuous exercise...

  4. An Evaluation of Collagen Metabolism in Non Human Primates Associated with the Bion 11 Space Program-Markers of Urinary Collagen Turnover and Muscle Connective Tissue

    Science.gov (United States)

    Vailas, Arthur C.; Martinez, Daniel A.

    1999-01-01

    Patients exhibiting changes in connective tissue and bone metabolism also show changes in urinary by-products of tissue metabolism. Furthermore, the changes in urinary connective tissue and bone metabolites precede alterations at the tissue macromolecular level. Astronauts and Cosmonauts have also shown suggestive increases in urinary by-products of mineralized and non-mineralized tissue degradation. Thus, the idea of assessing connective tissue and bone response in spaceflight monkeys by measurement of biomarkers in urine has merit. Other investigations of bone and connective histology, cytology and chemistry in the Bion 11 monkeys will allow for further validation of the relationship of urinary biomarkers and tissue response. In future flights the non-invasive procedure of urinary analysis may be useful in early detection of changes in these tissues. The purpose of this grant investigation was to evaluate mineralized and non-mineralized connective tissue responses of non-human primates to microgravity by the non-invasive analysis of urinary biomarkers. Secondly, we also wanted to assess muscle connective tissue adaptive changes in three weight-bearing skeletal muscles: the soleus, media] gastrocnemius and tibialis anterior by obtaining pre-flight and post-flight small biopsy specimens in collaboration with Dr. V. Reggie Edgerton's laboratory at the University of California at Los Angeles.

  5. Quantitative analysis of the synthesis and secretion of type VII collagen in cultured human dermal fibroblasts with a sensitive sandwich enzyme-linked immunoassay.

    Science.gov (United States)

    Amano, Satoshi; Ogura, Yuki; Akutsu, Nobuko; Nishiyama, Toshio

    2007-02-01

    Type VII collagen is the major component of anchoring fibrils in the epidermal basement membrane. Its expression has been analyzed by immunostaining or Northern blotting, but rarely at the protein level. In this study, we have quantitatively examined the effects of ascorbic acid and various cytokines/growth factors on the protein synthesis and secretion of type VII collagen by human dermal fibroblasts in culture, using a developed, highly sensitive sandwich enzyme-linked immunoassay with two kinds of specific monoclonal antibodies against the non-collagenous domain-1. Ascorbic acid and its derivative induced a twofold increase in type VII collagen synthesis, and markedly increased the secretion of type VII collagen into the medium when compared with the control culture. This effect was not influenced by the presence of transforming growth factor-beta1 (TGF-beta1). The synthesis of type VII collagen was elevated by TGF-beta1, platelet-derived growth factor, tumor necrosis factor-alpha, and interleukin-1beta, but not by TGF-alpha. Thus, our data indicate that the synthesis and secretion of type VII collagen in human dermal fibroblasts are regulated by ascorbate and the enhancement of type VII collagen gene expression by cytokines/growth factors is accompanied with elevated production of type VII collagen at the protein level.

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

    Directory of Open Access Journals (Sweden)

    Christopher A Smith

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

  7. Is fatty acid composition of human bone marrow significant to bone health?

    Science.gov (United States)

    Pino, Ana María; Rodríguez, J Pablo

    2017-12-16

    The bone marrow adipose tissue (BMAT) is a conserved component of the marrow microenvironment, providing storage and release of energy and stabilizing the marrow extent. Also, it is recognized both the amount and quality of BMAT are relevant to preserve the functional relationships between BMAT, bone, and blood cell production. In this article we ponder the information supporting the tenet that the quality of BMAT is relevant to bone health. In the human adult the distribution of BMAT is heterogeneous over the entire skeleton, and both BMAT accumulation and bone loss come about with aging in healthy populations. But some pathological conditions which increase BMAT formation lead to bone impairment and fragility. Analysis in vivo of the relative content of saturated and unsaturated fatty acids (FA) in BMAT indicates site-related bone marrow fat composition and an association between increased unsaturation index (UI) and bone health. With aging some impairment ensues in the regulation of bone marrow cells and systemic signals leading to local chronic inflammation. Most of the bone loss diseases which evolve altered BMAT composition have as common factors aging and/or chronic inflammation. Both saturated and unsaturated FAs originate lipid species which are active mediators in the inflammation process. Increased free saturated FAs may lead to lipotoxicity of bone marrow cells. The pro-inflammatory, anti-inflammatory or resolving actions of compounds derived from long chain poly unsaturated FAs (PUFA) on bone cells is varied, and depending on the metabolism of the parent n:3 or n:6 PUFAs series. Taking together the evidence substantiate that marrow adipocyte function is fundamental for an efficient link between systemic and marrow fatty acids to accomplish specific energy or regulatory needs of skeletal and marrow cells. Further, they reveal marrow requirements of PUFAs. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Effect of boron on osteogenic differentiation of human bone marrow stromal cells.

    Science.gov (United States)

    Ying, Xiaozhou; Cheng, Shaowen; Wang, Wei; Lin, Zhongqin; Chen, Qingyu; Zhang, Wei; Kou, Dongquan; Shen, Yue; Cheng, Xiaojie; Rompis, Ferdinand An; Peng, Lei; Zhu Lu, Chuan

    2011-12-01

    Bone marrow stromal cells (BMSCs) have been well established as an ideal source of cell-based therapy for bone tissue engineering applications. Boron (B) is a notable trace element in humans; so far, the effects of boron on the osteogenic differentiation of BMSCs have not been reported. The aim of this study was to evaluate the effects of boron (0, 1, 10,100, and 1,000 ng/ml) on osteogenic differentiation of human BMSCs. In this study, BMSCs proliferation was analyzed by cell counting kit-8 (CCK8) assay, and cell osteogenic differentiation was evaluated by alkaline phosphatase (ALP) activity assay, Von Kossa staining, and real-time PCR. The results indicated that the proliferation of BMSCs was no different from the control group when added with B at the concentration of 1, 10, and 100 ng/ml respectively (P > 0.05); in contrast, 1,000 ng/ml B inhibited the proliferation of BMSCs at days 4, 7, and 14 (P < 0.05). By ALP staining, we discovered that BMSCs treated with 10 and 100 ng/ml B presented a higher ALP activity compared with control (P < 0.05). By real-time PCR, we detected the messenger RNA expression of ALP, osteocalcin, collagen type I, and bone morphogenetic proteins 7 were also increased in 10 and 100 ng/ml B treatment groups (P < 0.05). The calcium depositions were increased in 1 and 10 ng/ml B treatment groups (P < 0.05). Taken all together, it was the first time to report that B could increase osteogenic effect by stimulating osteogenic differentiation-related marker gene synthesis during the proliferation and differentiation phase in human BMSCs and could be a promising approach for enhancing osteogenic capacity of cell-based construction in bone tissue engineering.

  9. Collagen degradation as a possibility to determine the post-mortem interval (PMI) of animal bones: a validation study referring to an original study of Boaks et al. (2014).

    Science.gov (United States)

    Jellinghaus, Katharina; Hachmann, Carolin; Hoeland, Katharina; Bohnert, Michael; Wittwer-Backofen, Ursula

    2018-05-01

    Estimation of the post-mortem interval (PMI) of unknown skeletal remains is a common forensic task. Boaks and colleagues demonstrated a new method for PMI estimation in showing a reduction of the collagen to non-collagen content (Co/NCo ratio) in porcine bones after a PMI of 12 months using the Sirius Red/Fast Green Collagen Staining Kit from Chondrex in 2014 (Boaks et al. Forensic Sci Int 240: 104-110, 2014). The aim of our study was to reproduce this method and to investigate if the method could be used for forensic issues. Sixteen fresh porcine bones were placed in prepared boxes where they were treated regularly with distilled water or with water from hay infusions. For determining the Co/NCo ratio, we used the Sirius Red/Fast Green Collagen Staining Kit from Chondrex, which stains collagenous (Co) proteins red and non-collagenous (NCo) proteins green Chondrex Inc. (2008). After a PMI of 1-3 months, an analysis of porcine bone thin sections was performed on the one hand with spectrophotometry, on the other hand with stereomicroscopy. Using spectrophotometry, we go low and partially negative Co/NCo ratios which were up to 100-fold lower than the results we expected to get. The data we got by stereomicroscopy and calculating the Co/NCo ratio from extracting the red and green content with the software MATLAB and so calculating the Co/NCo ratio showed a correlation between PMI and the Co/NCo ratio in the porcine bone samples. Regular addition of distilled water or water from a hay infusion did not produce any significant differences so that an increased presence of microorganisms had obviously no influence on collagen degradation.

  10. Parameters in three-dimensional osteospheroids of telomerized human mesenchymal (stromal) stem cells grown on osteoconductive scaffolds that predict in vivo bone-forming potential

    DEFF Research Database (Denmark)

    Burns, Jorge S; Hansen, Pernille Lund; Larsen, Kenneth H

    2010-01-01

    Osteoblastic differentiation of human mesenchymal stem cells (hMSC) in monolayer culture is artefactual, lacking an organized bone-like matrix. We present a highly reproducible microwell protocol generating three-dimensional ex vivo multicellular aggregates of telomerized hMSC (hMSC-telomerase re......Osteoblastic differentiation of human mesenchymal stem cells (hMSC) in monolayer culture is artefactual, lacking an organized bone-like matrix. We present a highly reproducible microwell protocol generating three-dimensional ex vivo multicellular aggregates of telomerized hMSC (h......, was deposited in the scaffold concavities. Here, mature osteoblasts stained positively for differentiated osteoblast markers TAZ, biglycan, osteocalcin, and phospho-AKT. Quantification of collagen birefringence and relatively high expression of genes for matrix proteins, including type I collagen, biglycan...

  11. Mutations in collagen 18A1 (COL18A1 and their relevance to the human phenotype

    Directory of Open Access Journals (Sweden)

    Passos-Bueno Maria Rita

    2006-01-01

    Full Text Available Collagen XVIII, a proteoglycan, is a component of basement membranes (BMs. There are three distinct isoforms that differ only by their N-terminal, but with a specific pattern of tissue and developmental expression. Cleavage of its C-terminal produces endostatin, an inhibitor of angiogenesis. In its N-terminal, there is a frizzled motif which seems to be involved in Wnt signaling. Mutations in this gene cause Knobloch syndrome KS, an autosomal recessive disorder characterized by vitreoretinal and macular degeneration and occipital encephalocele. This review discusses the effect of both rare and polymorphic alleles in the human phenotype, showing that deficiency of one of the collagen XVIII isoforms is sufficient to cause KS and that null alleles causing deficiency of all collagen XVIII isoforms are associated with a more severe ocular defect. This review besides illustrating the functional importance of collagen XVIII in eye development and its structure maintenance throughout life, it also shows its role in other tissues and organs, such as nervous system and kidney.

  12. Sericin Enhances the Bioperformance of Collagen-Based Matrices Preseeded with Human-Adipose Derived Stem Cells (hADSCs

    Directory of Open Access Journals (Sweden)

    Marieta Costache

    2013-01-01

    Full Text Available Current clinical strategies for adipose tissue engineering (ATE, including autologous fat implants or the use of synthetic surrogates, not only are failing in the long term, but also can’t face the latest requirements regarding the aesthetic restoration of the resulted imperfections. In this context, modern strategies in current ATE applications are based on the implantation of 3D cell-scaffold bioconstructs, designed for prospective achievement of in situ functional de novo tissue. Thus, in this paper, we reported for the first time the evaluation of a spongious 60% collagen and 40% sericin scaffold preseeded with human adipose-derived stem cells (hADSCs in terms of biocompatibility and adipogenic potential in vitro. We showed that the addition of the sticky protein sericin in the composition of a classical collagen sponge enhanced the adhesion and also the proliferation rate of the seeded cells, thus improving the biocompatibility of the novel scaffold. In addition, sericin stimulated PPARγ2 overexpression, triggering a subsequent upregulated expression profile of FAS, aP2 and perilipin adipogenic markers. These features, together with the already known sericin stimulatory potential on cellular collagen production, promote collagen-sericin biomatrix as a good candidate for soft tissue reconstruction and wound healing applications.

  13. Early reversal cells in adult human bone remodeling

    DEFF Research Database (Denmark)

    Abdelgawad, Mohamed Essameldin; Delaissé, Jean-Marie; Hinge, Maja

    2016-01-01

    The mechanism coupling bone resorption and formation is a burning question that remains incompletely answered through the current investigations on osteoclasts and osteoblasts. An attractive hypothesis is that the reversal cells are likely mediators of this coupling. Their nature is a big matter...... of debate. The present study performed on human cancellous bone is the first one combining in situ hybridization and immunohistochemistry to demonstrate their osteoblastic nature. It shows that the Runx2 and CD56 immunoreactive reversal cells appear to take up TRAcP released by neighboring osteoclasts....... Earlier preclinical studies indicate that reversal cells degrade the organic matrix left behind by the osteoclasts and that this degradation is crucial for the initiation of the subsequent bone formation. To our knowledge, this study is the first addressing these catabolic activities in adult human bone...

  14. Endocytic collagen degradation

    DEFF Research Database (Denmark)

    Madsen, Daniel H.; Jürgensen, Henrik J.; Ingvarsen, Signe Ziir

    2012-01-01

    it crucially important to understand both the collagen synthesis and turnover mechanisms in this condition. Here we show that the endocytic collagen receptor, uPARAP/Endo180, is a major determinant in governing the balance between collagen deposition and degradation. Cirrhotic human livers displayed a marked...... up-regulation of uPARAP/Endo180 in activated fibroblasts and hepatic stellate cells located close to the collagen deposits. In a hepatic stellate cell line, uPARAP/Endo180 was shown to be active in, and required for, the uptake and intracellular degradation of collagen. To evaluate the functional...... groups of mice clearly revealed a fibrosis protective role of uPARAP/Endo180. This effect appeared to directly reflect the activity of the collagen receptor, since no compensatory events were noted when comparing the mRNA expression profiles of the two groups of mice in an array system focused on matrix-degrading...

  15. Assessing Collagen and Elastin Pressure-dependent Microarchitectures in Live, Human Resistance Arteries by Label-free Fluorescence Microscopy

    DEFF Research Database (Denmark)

    Bloksgaard, Maria; Thorsted, Bjarne; Brewer, Jonathan R

    2018-01-01

    The pathogenic contribution of resistance artery remodeling is documented in essential hypertension, diabetes and the metabolic syndrome. Investigations and development of microstructurally motivated mathematical models for understanding the mechanical properties of human resistance arteries...... in health and disease have the potential to aid understanding how disease and medical treatments affect the human microcirculation. To develop these mathematical models, it is essential to decipher the relationship between the mechanical and microarchitectural properties of the microvascular wall...... of interest. Image analyses are described for quantifying i) pressure-induced changes in internal elastic lamina branching angles and adventitial collagen straightness using Fiji and ii) collagen and elastin volume densities determined using Ilastik software. Preferably all mechanical and imaging measurements...

  16. Protease inhibitors enhance extracellular collagen fibril deposition in human mesenchymal stem cells

    OpenAIRE

    Han, Sejin; Li, Yuk Yin; Chan, Barbara Pui

    2015-01-01

    Introduction Collagen is a widely used naturally occurring biomaterial for scaffolding, whereas mesenchymal stem cells (MSCs) represent a promising cell source in tissue engineering and regenerative medicine. It is generally known that cells are able to remodel their environment by simultaneous degradation of the scaffolds and deposition of newly synthesized extracellular matrix. Nevertheless, the interactions between MSCs and collagen biomaterials are poorly known, and the strategies enhanci...

  17. DNA and bone structure preservation in medieval human skeletons.

    Science.gov (United States)

    Coulson-Thomas, Yvette M; Norton, Andrew L; Coulson-Thomas, Vivien J; Florencio-Silva, Rinaldo; Ali, Nadir; Elmrghni, Samir; Gil, Cristiane D; Sasso, Gisela R S; Dixon, Ronald A; Nader, Helena B

    2015-06-01

    Morphological and ultrastructural data from archaeological human bones are scarce, particularly data that have been correlated with information on the preservation of molecules such as DNA. Here we examine the bone structure of macroscopically well-preserved medieval human skeletons by transmission electron microscopy and immunohistochemistry, and the quantity and quality of DNA extracted from these skeletons. DNA technology has been increasingly used for analyzing physical evidence in archaeological forensics; however, the isolation of ancient DNA is difficult since it is highly degraded, extraction yields are low and the co-extraction of PCR inhibitors is a problem. We adapted and optimised a method that is frequently used for isolating DNA from modern samples, Chelex(®) 100 (Bio-Rad) extraction, for isolating DNA from archaeological human bones and teeth. The isolated DNA was analysed by real-time PCR using primers targeting the sex determining region on the Y chromosome (SRY) and STR typing using the AmpFlSTR(®) Identifiler PCR Amplification kit. Our results clearly show the preservation of bone matrix in medieval bones and the presence of intact osteocytes with well preserved encapsulated nuclei. In addition, we show how effective Chelex(®) 100 is for isolating ancient DNA from archaeological bones and teeth. This optimised method is suitable for STR typing using kits aimed specifically at degraded and difficult DNA templates since amplicons of up to 250bp were successfully amplified. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  18. Dating of cremated bones

    NARCIS (Netherlands)

    Lanting, JN; Aerts-Bijma, AT; van der Plicht, J; Boaretto, E.; Carmi, I.

    2001-01-01

    When dating unburnt bone, bone collagen, the organic fraction of the bone, is used. Collagen does not survive the heat of the cremation pyre, so dating of cremated bone has been considered impossible. Structural carbonate in the mineral fraction of the bone, however, survives the cremation process.

  19. Histologic and histomorphometric evaluation of bone regeneration using nanocrystalline hydroxyapatite and human freeze-dried bone graft : An experimental study in rabbit.

    Science.gov (United States)

    Sadeghi, Rokhsareh; Najafi, Mohammad; Semyari, Hassan; Mashhadiabbas, Fatemeh

    2017-03-01

    Bone regeneration is an important concern in periodontal treatment and implant dentistry. Different biomaterials and surgical techniques have been used for this purpose. The aim of the present study was to compare the effect of nanocrystalline hydroxyapatite and human freeze-dried bone graft (FDBG) in regeneration of rabbit calvarium bony defects by histologic and histomorphometric evaluation. In this experimental study, three similar defects, measuring 8 mm in diameter, were created in the calvaria of 16 white New Zealand rabbits. Two defects were filled with FDBG and nanocrystalline hydroxyapatite silica gel, while the other one remained unfilled to be considered as control. All the defects were covered with collagen membranes. During the healing period, two animals perished; so 14 rabbits were divided into two groups: half of them were euthanized after 6 weeks of healing and the other half after 12 weeks. The specimens were subjected to histologic and histomorphometric examinations for assessment of the following variables: percentage of bone formation and residual graft material, inflammation scores, patterns of bone formation and type of newly formed bone. The percentages of new bone formation after 6 weeks were 14.22 ± 7.85, 21.57 ± 6.91, and 20.54 ± 10.07% in FDBG, NanoBone, and control defects. These values were 27.54 ± 20.19, 23.86 ± 6.27, and 26.48 ± 14.18% in 12-week specimens, respectively. No significant differences were found in the amount of bone formation between the groups. With regard to inflammation, the control and NanoBone groups showed significantly less inflammation compared to FDBG at the 6-week healing phase (P = 0.04); this difference was not significant in the 12-week specimens. Based on the results of this experimental study, both NanoBone and FDBG exhibited a similar effect on bone formation.

  20. Radiation dosimetry in human bone using electron paramagnetic resonance

    International Nuclear Information System (INIS)

    Breen, S.L.

    1995-01-01

    Accurate measurements of dose in bone are required in order to improve the dosimetry of systemic radiotherapy for osseous metastases. Bone is an integrating dosimeter which records the radiation history of the skeleton. During irradiation, electrons become trapped in the crystalline component of bone mineral (hydroxyapatite). The traps are very stable; at room temperature, emptying of the traps occurs with a half-life of many years. The population of trapped unpaired electrons is proportional to the radiation dose administered to the bone and can be measured in excised bone samples using electron paramagnetic resonance (EPR). EPR spectra of synthetic hydroxyapatite, irradiated with Co-60, were obtained at room temperature and at 77 K. At room temperature, the radiation-induced signal, with a g-value of 2.001 ± 0.001 increased linearly with absorbed dose above a lower threshold of 3 Gy, up to doses of 200 Gy. In contrast with pure hydroxyapatite, EPR spectra of excised human bone showed a broad 'native' signal, due to the organic component of bone, which masks the dosimetrically important signal. This native signal is highly variable from sample to sample and precludes the use of EPR as an absolute dosimetry technique. However, after subtraction of the background signal, irradiated human bone showed a linear response with a lower limit of measurement similar to that of synthetic hydroxyapatite. Bone is an in vivo linear dosimeter which can be exploited to develop accurate estimates of the radiation dose delivered during systemic radiotherapy and teletherapy. However, improved sensitivity of the EPR dosimetry technique is necessary before it can be applied reliably in clinical situations. (author)

  1. SU-E-J-107: Supervised Learning Model of Aligned Collagen for Human Breast Carcinoma Prognosis

    International Nuclear Information System (INIS)

    Bredfeldt, J; Liu, Y; Conklin, M; Keely, P; Eliceiri, K; Mackie, T

    2014-01-01

    Purpose: Our goal is to develop and apply a set of optical and computational tools to enable large-scale investigations of the interaction between collagen and tumor cells. Methods: We have built a novel imaging system for automating the capture of whole-slide second harmonic generation (SHG) images of collagen in registry with bright field (BF) images of hematoxylin and eosin stained tissue. To analyze our images, we have integrated a suite of supervised learning tools that semi-automatically model and score collagen interactions with tumor cells via a variety of metrics, a method we call Electronic Tumor Associated Collagen Signatures (eTACS). This group of tools first segments regions of epithelial cells and collagen fibers from BF and SHG images respectively. We then associate fibers with groups of epithelial cells and finally compute features based on the angle of interaction and density of the collagen surrounding the epithelial cell clusters. These features are then processed with a support vector machine to separate cancer patients into high and low risk groups. Results: We validated our model by showing that eTACS produces classifications that have statistically significant correlation with manual classifications. In addition, our system generated classification scores that accurately predicted breast cancer patient survival in a cohort of 196 patients. Feature rank analysis revealed that TACS positive fibers are more well aligned with each other, generally lower density, and terminate within or near groups of epithelial cells. Conclusion: We are working to apply our model to predict survival in larger cohorts of breast cancer patients with a diversity of breast cancer types, predict response to treatments such as COX2 inhibitors, and to study collagen architecture changes in other cancer types. In the future, our system may be used to provide metastatic potential information to cancer patients to augment existing clinical assays

  2. SU-E-J-107: Supervised Learning Model of Aligned Collagen for Human Breast Carcinoma Prognosis

    Energy Technology Data Exchange (ETDEWEB)

    Bredfeldt, J; Liu, Y; Conklin, M; Keely, P; Eliceiri, K; Mackie, T [University of Wisconsin, Madison, WI (United States)

    2014-06-01

    Purpose: Our goal is to develop and apply a set of optical and computational tools to enable large-scale investigations of the interaction between collagen and tumor cells. Methods: We have built a novel imaging system for automating the capture of whole-slide second harmonic generation (SHG) images of collagen in registry with bright field (BF) images of hematoxylin and eosin stained tissue. To analyze our images, we have integrated a suite of supervised learning tools that semi-automatically model and score collagen interactions with tumor cells via a variety of metrics, a method we call Electronic Tumor Associated Collagen Signatures (eTACS). This group of tools first segments regions of epithelial cells and collagen fibers from BF and SHG images respectively. We then associate fibers with groups of epithelial cells and finally compute features based on the angle of interaction and density of the collagen surrounding the epithelial cell clusters. These features are then processed with a support vector machine to separate cancer patients into high and low risk groups. Results: We validated our model by showing that eTACS produces classifications that have statistically significant correlation with manual classifications. In addition, our system generated classification scores that accurately predicted breast cancer patient survival in a cohort of 196 patients. Feature rank analysis revealed that TACS positive fibers are more well aligned with each other, generally lower density, and terminate within or near groups of epithelial cells. Conclusion: We are working to apply our model to predict survival in larger cohorts of breast cancer patients with a diversity of breast cancer types, predict response to treatments such as COX2 inhibitors, and to study collagen architecture changes in other cancer types. In the future, our system may be used to provide metastatic potential information to cancer patients to augment existing clinical assays.

  3. ECM microenvironment unlocks brown adipogenic potential of adult human bone marrow-derived MSCs.

    Science.gov (United States)

    Lee, Michelle H; Goralczyk, Anna G; Kriszt, Rókus; Ang, Xiu Min; Badowski, Cedric; Li, Ying; Summers, Scott A; Toh, Sue-Anne; Yassin, M Shabeer; Shabbir, Asim; Sheppard, Allan; Raghunath, Michael

    2016-02-17

    Key to realizing the diagnostic and therapeutic potential of human brown/brite adipocytes is the identification of a renewable, easily accessible and safe tissue source of progenitor cells, and an efficacious in vitro differentiation protocol. We show that macromolecular crowding (MMC) facilitates brown adipocyte differentiation in adult human bone marrow mesenchymal stem cells (bmMSCs), as evidenced by substantially upregulating uncoupling protein 1 (UCP1) and uncoupled respiration. Moreover, MMC also induced 'browning' in bmMSC-derived white adipocytes. Mechanistically, MMC creates a 3D extracellular matrix architecture enshrouding maturing adipocytes in a collagen IV cocoon that is engaged by paxillin-positive focal adhesions also at the apical side of cells, without contact to the stiff support structure. This leads to an enhanced matrix-cell signaling, reflected by increased phosphorylation of ATF2, a key transcription factor in UCP1 regulation. Thus, tuning the dimensionality of the microenvironment in vitro can unlock a strong brown potential dormant in bone marrow.

  4. Recombinant human IGF-1 produced by transgenic plant cell suspension culture enhances new bone formation in calvarial defects.

    Science.gov (United States)

    Poudel, Sher Bahadur; Bhattarai, Govinda; Kook, Sung-Ho; Shin, Yun-Ji; Kwon, Tae-Ho; Lee, Seung-Youp; Lee, Jeong-Chae

    2017-10-01

    Transgenic plant cell suspension culture systems have been utilized extensively as convenient and efficient expression systems for the production of recombinant human growth factors. We produced insulin-like growth factor-1 using a plant suspension culture system (p-IGF-1) and explored its effect on new bone formation in calvarial defects. We also compared the bone regenerating potential of p-IGF-1 with commercial IGF-1 derived from Escherichia coli (e-IGF-1). Male C57BL/6 mice underwent calvarial defect surgery, and the defects were loaded with absorbable collagen sponge (ACS) only (ACS group) or ACS impregnated with 13μg of p-IGF-1 (p-IGF-1 group) or e-IGF-1 (e-IGF-1 group). The sham group did not receive any treatment with ACS or IGFs after surgery. Live μCT and histological analyses showed critical-sized bone defects in the sham group, whereas greater bone formation was observed in the p-IGF-1 and e-IGF-1 groups than the ACS group both 5 and 10weeks after surgery. Bone mineral density, bone volume, and bone surface values were also higher in the IGF groups than in the ACS group. Local delivery of p-IGF-1 or e-IGF-1 more greatly enhanced the expression of osteoblast-specific markers, but inhibited osteoclast formation, in newly formed bone compared with ACS control group. Specifically, p-IGF-1 treatment induced higher expression of alkaline phosphatase, osteocalcin, and osteopontin in the defect site than did e-IGF-1. Furthermore, treatment with p-IGF-1, but not e-IGF-1, increased mineralization of MC3T3-E1 cells, with the attendant upregulation of osteogenic marker genes. Collectively, our findings suggest the potential of p-IGF-1 in promoting the processes required for bone regeneration. Copyright © 2017. Published by Elsevier Ltd.

  5. Overexpression of Human Bone Alkaline Phosphatase in Pichia Pastoris

    Science.gov (United States)

    Karr, Laurel; Malone, Christine, C.; Rose, M. Franklin (Technical Monitor)

    2000-01-01

    The Pichiapastoris expression system was utilized to produce functionally active human bone alkaline phosphatase in gram quantities. Bone alkaline phosphatase is a key enzyme in bone formation and biomineralization, yet important questions about its structural chemistry and interactions with other cellular enzymes in mineralizing tissues remain unanswered. A soluble form of human bone alkaline phosphatase was constructed by deletion of the 25 amino acid hydrophobic C-terminal region of the encoding cDNA and inserted into the X-33 Pichiapastoris strain. An overexpression system was developed in shake flasks and converted to large-scale fermentation. Alkaline phosphatase was secreted into the medium to a level of 32mgAL when cultured in shake flasks. Enzyme activity was 12U/mg measured by a spectrophotometric assay. Fermentation yielded 880mgAL with enzymatic activity of 968U/mg. Gel electrophoresis analysis indicates that greater than 50% of the total protein in the fermentation is alkaline phosphatase. A purification scheme has been developed using ammonium sulfate precipitation followed by hydrophobic interaction chromatography. We are currently screening crystallization conditions of the purified recombinant protein for subsequent X-ray diffraction analyses. Structural data should provide additional information on the role of alkaline phosphatase in normal bone mineralization and in certain bone mineralization anomalies.

  6. Cardiopulmonary dysfunction in the Osteogenesis imperfecta mouse model Aga2 and human patients are caused by bone-independent mechanisms.

    Science.gov (United States)

    Thiele, Frank; Cohrs, Christian M; Flor, Armando; Lisse, Thomas S; Przemeck, Gerhard K H; Horsch, Marion; Schrewe, Anja; Gailus-Durner, Valerie; Ivandic, Boris; Katus, Hugo A; Wurst, Wolfgang; Reisenberg, Catherine; Chaney, Hollis; Fuchs, Helmut; Hans, Wolfgang; Beckers, Johannes; Marini, Joan C; Hrabé de Angelis, Martin

    2012-08-15

    Osteogenesis imperfecta (OI) is an inherited connective tissue disorder with skeletal dysplasia of varying severity, predominantly caused by mutations in the collagen I genes (COL1A1/COL1A2). Extraskeletal findings such as cardiac and pulmonary complications are generally considered to be significant secondary features. Aga2, a murine model for human OI, was systemically analyzed in the German Mouse Clinic by means of in vivo and in vitro examinations of the cardiopulmonary system, to identify novel mechanisms accounting for perinatal lethality. Pulmonary and, especially, cardiac fibroblast of perinatal lethal Aga2/+ animals display a strong down-regulation of Col1a1 transcripts in vivo and in vitro, resulting in a loss of extracellular matrix integrity. In addition, dysregulated gene expression of Nppa, different types of collagen and Agt in heart and lung tissue support a bone-independent vicious cycle of heart dysfunction, including hypertrophy, loss of myocardial matrix integrity, pulmonary hypertension, pneumonia and hypoxia leading to death in Aga2. These murine findings are corroborated by a pediatric OI cohort study, displaying significant progressive decline in pulmonary function and restrictive pulmonary disease independent of scoliosis. Most participants show mild cardiac valvular regurgitation, independent of pulmonary and skeletal findings. Data obtained from human OI patients and the mouse model Aga2 provide novel evidence for primary effects of type I collagen mutations on the heart and lung. The findings will have potential benefits of anticipatory clinical exams and early intervention in OI patients.

  7. Connective matrix organization in human pulmonary fibrosis. Collagen polymorphism analysis in fibrotic deposits by immunohistological methods.

    Science.gov (United States)

    Takiya, C; Peyrol, S; Cordier, J F; Grimaud, J A

    1983-01-01

    In the interstitium of the alveolar septa in the peripheral parts of the lung, four molecular types of collagen (I, III, IV and V) each with different morphological appearances, can be identified. The structural integrity of collagens accounts for the physiological efficiency of the lung. Fibrous thickening of alveolar septa is an invariable result of various diseases affecting the interstitium of the lung. The light and electron microscopic findings, and the immunological typing of collagens in six cases of fibrotic alveolar disease, are described. In the alveolar septa, two different compartments (the alveolo-capillary junction and the supportive axis) were affected by fibrosis: the alveolo-capillary junction was widened by the addition of interstitial collagens to basement membranes. In the axis, the increase of interstitial (types I and III) collagen gave rise to different patterns of connective matrix organization, graded as Loose or Dense depending on quantitative alterations of the type I/III ratio. The mode of organization of the fibrotic lung connective matrix, which depends on the quality of deposits in the matrix, may be correlated with the evolution of interstitial pulmonary fibrosis, in terms of its stability, remodelling ability and reversibility.

  8. Regenerative surgical therapy for peri-implantitis using deproteinized bovine bone mineral with 10% collagen, enamel matrix derivative and Doxycycline-A prospective 3-year cohort study.

    Science.gov (United States)

    Mercado, Faustino; Hamlet, Stephen; Ivanovski, Saso

    2018-05-16

    There is limited evidence regarding the long-term efficacy of regenerative treatment for peri-implantitis. The aim of this study was to evaluate a combination therapy of deproteinized bovine bone mineral with 10% collagen (DBBMC), enamel matrix derivative (EMD) and Doxycycline in the regeneration of bone defects associated with peri-implantitis. Thirty patients diagnosed with peri-implantitis (BoP/suppuration, probing depth greater than 4 mm, minimum radiographic bone loss of 20%, at least 2 years in function) were enrolled in the study. Clinical measurements included probing depths, recession, radiographic bone fill, gingival inflammation and bleeding on probing/suppuration. Following surgical access and debridement, the implant surfaces were decontaminated with 24% EDTA for 2 min, and the bone defects were filled with a combined mixture of DBBMC, EMD and Doxycycline powder. The defects were covered with connective tissue grafts where necessary. Clinical measurements were recorded after 12, 24 and 36 months. The mean probing depth and bone loss at the initial visit was 8.9 mm (±1.9) and 6.92 mm (±1.26), respectively. Both mean probing depth and bone loss reduced significantly from baseline to 3.55 mm (±0.50) and 2.85 mm (±0.73) at 12 months, 3.50 (±0.50) and 2.62 mm (±0.80) at 24 months and 3.50 mm (±0.50) and 2.60 mm (±0.73) at 36 months. 56.6% of the implants were considered successfully treated (according to Successful Treatment Outcome Criterion: PD 10%, no BoP/suppuration, no recession >0.5 mm for anterior implants and >1.5 mm for posterior implants) after 36 months. Regenerative treatment of peri-implantitis using a combined mixture of DBBMC, EMD and Doxycycline achieved promising results. The benefits of this protocol incorporating EMD should be tested in randomized clinical trials. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Osteogenic medium is superior to growth factors in differentiation of human adipose stem cells towards bone-forming cells in 3D culture

    Directory of Open Access Journals (Sweden)

    L Tirkkonen

    2013-01-01

    Full Text Available Human adipose stem cells (hASCs have been recently used to treat bone defects in clinical practice. Yet there is a need for more optimal scaffolds and cost-effective approaches to induce osteogenic differentiation of hASCs. Therefore, we compared the efficiency of bone morphogenetic proteins (BMP-2 and BMP-7, vascular endothelial growth factor (VEGF, and osteogenic medium (OM for the osteo-induction of hASCs in 3D culture. In addition, growth factors were tested in combination with OM. Commercially available bioactive glass scaffolds (BioRestore and biphasic calcium phosphate granules (BoneCeramic were evaluated as prospective carriers for hASCs. Both biomaterials supported hASC-viability, but BioRestore resulted in higher cell number than BoneCeramic, whereas BoneCeramic supported more significant collagen production. The most efficient osteo-induction was achieved with plain OM, promoting higher alkaline phosphatase activity and collagen production than growth factors. In fact, treatment with BMP-2 or VEGF did not increase osteogenic differentiation or cell number significantly more than maintenance medium with either biomaterial. Moreover, BMP-7 treatment consistently inhibited proliferation and osteogenic differentiation of hASCs. Interestingly, there was no benefit from growth factors added to OM. This is the first study to demonstrate that OM enhances hASC-differentiation towards bone-forming cells significantly more than growth factors in 3D culture.

  10. Probing the interaction forces of prostate cancer cells with collagen I and bone marrow derived stem cells on the single cell level.

    Directory of Open Access Journals (Sweden)

    Ediz Sariisik

    Full Text Available Adhesion of metastasizing prostate carcinoma cells was quantified for two carcinoma model cell lines LNCaP (lymph node-specific and PC3 (bone marrow-specific. By time-lapse microscopy and force spectroscopy we found PC3 cells to preferentially adhere to bone marrow-derived mesenchymal stem cells (SCP1 cell line. Using atomic force microscopy (AFM based force spectroscopy, the mechanical pattern of the adhesion to SCP1 cells was characterized for both prostate cancer cell lines and compared to a substrate consisting of pure collagen type I. PC3 cells dissipated more energy (27.6 aJ during the forced de-adhesion AFM experiments and showed significantly more adhesive and stronger bonds compared to LNCaP cells (20.1 aJ. The characteristic signatures of the detachment force traces revealed that, in contrast to the LNCaP cells, PC3 cells seem to utilize their filopodia in addition to establish adhesive bonds. Taken together, our study clearly demonstrates that PC3 cells have a superior adhesive affinity to bone marrow mesenchymal stem cells, compared to LNCaP. Semi-quantitative PCR on both prostate carcinoma cell lines revealed the expression of two Col-I binding integrin receptors, α1β1 and α2β1 in PC3 cells, suggesting their possible involvement in the specific interaction to the substrates. Further understanding of the exact mechanisms behind this phenomenon might lead to optimized therapeutic applications targeting the metastatic behavior of certain prostate cancer cells towards bone tissue.

  11. Imaging and modeling of acute pressure-induced changes of collagen and elastin microarchitectures in pig and human resistance arteries.

    Science.gov (United States)

    Bloksgaard, Maria; Leurgans, Thomas M; Spronck, Bart; Heusinkveld, Maarten H G; Thorsted, Bjarne; Rosenstand, Kristoffer; Nissen, Inger; Hansen, Ulla M; Brewer, Jonathan R; Bagatolli, Luis A; Rasmussen, Lars M; Irmukhamedov, Akhmadjon; Reesink, Koen D; De Mey, Jo G R

    2017-07-01

    The impact of disease-related changes in the extracellular matrix (ECM) on the mechanical properties of human resistance arteries largely remains to be established. Resistance arteries from both pig and human parietal pericardium (PRA) display a different ECM microarchitecture compared with frequently used rodent mesenteric arteries. We hypothesized that the biaxial mechanics of PRA mirror pressure-induced changes in the ECM microarchitecture. This was tested using isolated pig PRA as a model system, integrating vital imaging, pressure myography, and mathematical modeling. Collagenase and elastase digestions were applied to evaluate the load-bearing roles of collagen and elastin, respectively. The incremental elastic modulus linearly related to the straightness of adventitial collagen fibers circumferentially and longitudinally (both R 2 ≥ 0.99), whereas there was a nonlinear relationship to the internal elastic lamina elastin fiber branching angles. Mathematical modeling suggested a collagen recruitment strain (means ± SE) of 1.1 ± 0.2 circumferentially and 0.20 ± 0.01 longitudinally, corresponding to a pressure of ~40 mmHg, a finding supported by the vital imaging. The integrated method was tested on human PRA to confirm its validity. These showed limited circumferential distensibility and elongation and a collagen recruitment strain of 0.8 ± 0.1 circumferentially and 0.06 ± 0.02 longitudinally, reached at a distending pressure below 20 mmHg. This was confirmed by vital imaging showing negligible microarchitectural changes of elastin and collagen upon pressurization. In conclusion, we show here, for the first time in resistance arteries, a quantitative relationship between pressure-induced changes in the extracellular matrix and the arterial wall mechanics. The strength of the integrated methods invites for future detailed studies of microvascular pathologies. NEW & NOTEWORTHY This is the first study to quantitatively relate pressure

  12. Collagen-coated polylactic-glycolic acid (PLGA) seeded with neural-differentiated human mesenchymal stem cells as a potential nerve conduit.

    Science.gov (United States)

    Sulong, Ahmad Fadzli; Hassan, Nur Hidayah; Hwei, Ng Min; Lokanathan, Yogeswaran; Naicker, Amaramalar Selvi; Abdullah, Shalimar; Yusof, Mohd Reusmaazran; Htwe, Ohnmar; Idrus, Ruszymah Bt Hj; Haflah, Nor Hazla Mohamed

    2014-01-01

    Autologous nerve grafts to bridge nerve gaps pose various drawbacks. Nerve tissue engineering to promote nerve regeneration using artificial neural conduits has emerged as a promising alternative. To develop an artificial nerve conduit using collagen-coated polylactic-glycolic acid (PLGA) and to analyse the survivability and propagating ability of the neuro-differentiated human mesenchymal stem cells in this conduit. The PLGA conduit was constructed by dip-molding method and coated with collagen by immersing the conduit in collagen bath. The ultra structure of the conduits were examined before they were seeded with neural-differentiated human mesenchymal stem cells (nMSC) and implanted sub-muscularly on nude mice thighs. The non-collagen-coated PLGA conduit seeded with nMSC and non-seeded non-collagen-coated PLGA conduit were also implanted for comparison purposes. The survivability and propagation ability of nMSC was studied by histological and immunohistochemical analysis. The collagen-coated conduits had a smooth inner wall and a highly porous outer wall. Conduits coated with collagen and seeded with nMSCs produced the most number of cells after 3 weeks. The best conduit based on the number of cells contained within it after 3 weeks was the collagen-coated PLGA conduit seeded with neuro-transdifferentiated cells. The collagen-coated PLGA conduit found to be suitable for attachment, survival and proliferation of the nMSC. Minimal cell infiltration was found in the implanted conduits where nearly all of the cells found in the cell seeded conduits are non-mouse origin and have neural cell markers, which exhibit the biocompatibility of the conduits. The collagen-coated PLGA conduit is biocompatible, non-cytotoxic and suitable for use as artificial nerve conduits.

  13. Organotypic culture of human bone marrow adipose tissue.

    Science.gov (United States)

    Uchihashi, Kazuyoshi; Aoki, Shigehisa; Shigematsu, Masamori; Kamochi, Noriyuki; Sonoda, Emiko; Soejima, Hidenobu; Fukudome, Kenji; Sugihara, Hajime; Hotokebuchi, Takao; Toda, Shuji

    2010-04-01

    The precise role of bone marrow adipose tissue (BMAT) in the marrow remains unknown. The purpose of the present study was therefore to describe a novel method for studying BMAT using 3-D collagen gel culture of BMAT fragments, immunohistochemistry, ELISA and real-time reverse transcription-polymerase chain reaction. Mature adipocytes and CD45+ leukocytes were retained for >3 weeks. Bone marrow stromal cells (BMSC) including a small number of lipid-laden preadipocytes and CD44+/CD105+ mesenchymal stem cell (MSC)-like cells, developed from BMAT. Dexamethasone (10 micromol/L), but not insulin (20 mU/mL), significantly increased the number of preadipocytes. Dexamethasone and insulin also promoted leptin production and gene expression in BMAT. Adiponectin production by BMAT was BMAT, in which adiponectin protein secretion is normally very low, and that BMAT may exhibit a different phenotype from that of the visceral and subcutaneous adipose tissues. BMAT-osteoblast interactions were also examined, and it was found that osteoblasts inhibited the development of BMSC and reduced leptin production, while BMAT inhibited the growth and differentiation of osteoblasts. The present novel method proved to be useful for the study of BMAT biology.

  14. [Comparation on Haversian system between human and animal bones by imaging analysis].

    Science.gov (United States)

    Lu, Hui-Ling; Zheng, Jing; Yao, Ya-Nan; Chen, Sen; Wang, Hui-Pin; Chen, Li-Xian; Guo, Jing-Yuan

    2006-04-01

    To explore the differences in Haversian system between human and animal bones through imaging analysis and morphology description. Thirty-five slices grinding from human being as well as dog, pig, cow and sheep bones were observed to compare their structure, then were analysed with the researchful microscope. Plexiform bone or oeston band was not found in human bones; There were significant differences in the shape, size, location, density of Haversian system, between human and animal bones. The amount of Haversian lamella and diameter of central canal in human were the biggest; Significant differences in the central canal diameter and total area percentage between human and animal bones were shown by imaging analysis. (1) Plexiform bone and osteon band could be the exclusive index in human bone; (2) There were significant differences in the structure of Haversian system between human and animal bones; (3) The percentage of central canals total area was valuable in species identification through imaging analysis.

  15. Collagen: A review on its sources and potential cosmetic applications.

    Science.gov (United States)

    Avila Rodríguez, María Isabela; Rodríguez Barroso, Laura G; Sánchez, Mirna Lorena

    2018-02-01

    Collagen is a fibrillar protein that conforms the conjunctive and connective tissues in the human body, essentially skin, joints, and bones. This molecule is one of the most abundant in many of the living organisms due to its connective role in biological structures. Due to its abundance, strength and its directly proportional relation with skin aging, collagen has gained great interest in the cosmetic industry. It has been established that the collagen fibers are damaged with the pass of time, losing thickness and strength which has been strongly related with skin aging phenomena [Colágeno para todo. 60 y más. 2016. http://www.revista60ymas.es/InterPresent1/groups/revistas/documents/binario/ses330informe.pdf.]. As a solution, the cosmetic industry incorporated collagen as an ingredient of different treatments to enhance the user youth and well-being, and some common presentations are creams, nutritional supplement for bone and cartilage regeneration, vascular and cardiac reconstruction, skin replacement, and augmentation of soft skin among others [J App Pharm Sci. 2015;5:123-127]. Nowadays, the biomolecule can be obtained by extraction from natural sources such as plants and animals or by recombinant protein production systems including yeast, bacteria, mammalian cells, insects or plants, or artificial fibrils that mimic collagen characteristics like the artificial polymer commercially named as KOD. Because of its increased use, its market size is valued over USD 6.63 billion by 2025 [Collagen Market By Source (Bovine, Porcine, Poultry, Marine), Product (Gelatin, Hydrolyzed Collagen), Application (Food & Beverages, Healthcare, Cosmetics), By Region, And Segment Forecasts, 2014 - 2025. Grand View Research. http://www.grandviewresearch.com/industry-analysis/collagen-market. Published 2017.]. Nevertheless, there has been little effort on identifying which collagen types are the most suitable for cosmetic purposes, for which the present review will try to enlighten

  16. Complete amino acid sequence of the human alpha 5 (IV) collagen chain and identification of a single-base mutation in exon 23 converting glycine 521 in the collagenous domain to cysteine in an Alport syndrome patient

    DEFF Research Database (Denmark)

    Zhou, J; Hertz, Jens Michael; Leinonen, A

    1992-01-01

    We have generated and characterized cDNA clones providing the complete amino acid sequence of the human type IV collagen chain whose gene has been shown to be mutated in X chromosome-linked Alport syndrome. The entire translation product has 1,685 amino acid residues. There is a 26-residue signal...

  17. H-ras oncogene-transformed human bronchial epithelial cells (TBE-1) secrete a single metalloprotease capable of degrading basement membrane collagen

    International Nuclear Information System (INIS)

    Collier, I.E.; Wilhelm, S.M.; Eisen, A.Z.

    1988-01-01

    H-ras transformed human bronchial epithelial cells (TBE-1) secrete a single major extracellular matrix metalloprotease which is not found in the normal parental cells. The enzyme is secreted in a latent form which can be activated to catalyze the cleavage of the basement membrane macromolecule type IV collagen. The substrates in their order of preference are: gelatin, type IV collagen, type V collagen, fibronectin, and type VII collagen; but the enzyme does not cleave the interstitial collagens or laminin. This protease is identical to gelatinase isolated from normal human skin explants, normal human skin fibroblasts, and SV40-transformed human lung fibroblasts. Based on this ability to initiate the degradation of type IV collagen in a pepsin-resistant portion of the molecule, it will be referred to as type IV collagenase. This enzyme is most likely the human analog of type IV collagenase detected in several rodent tumors. Type IV collagenase consists of three domains. Type IV collagenase represents the third member of a newly recognized gene family coding for secreted extracellular matrix metalloproteases, which includes interstitial fibroblast collagenase and stromelysin

  18. Sphingosine 1-phosphate (S1P) suppresses the collagen-induced activation of human platelets via S1P4 receptor.

    Science.gov (United States)

    Onuma, Takashi; Tanabe, Kumiko; Kito, Yuko; Tsujimoto, Masanori; Uematsu, Kodai; Enomoto, Yukiko; Matsushima-Nishiwaki, Rie; Doi, Tomoaki; Nagase, Kiyoshi; Akamatsu, Shigeru; Tokuda, Haruhiko; Ogura, Shinji; Iwama, Toru; Kozawa, Osamu; Iida, Hiroki

    2017-08-01

    Sphingosine 1-phosphate (S1P) is as an extracellular factor that acts as a potent lipid mediator by binding to specific receptors, S1P receptors (S1PRs). However, the precise role of S1P in human platelets that express S1PRs has not yet been fully clarified. We previously reported that heat shock protein 27 (HSP27) is released from human platelets accompanied by its phosphorylation stimulated by collagen. In the present study, we investigated the effect of S1P on the collagen-induced platelet activation. S1P pretreatment markedly attenuated the collagen-induced aggregation. Co-stimulation with S1P and collagen suppressed collagen-induced platelet activation, but the effect was weaker than that of S1P-pretreatment. The collagen-stimulated secretion of platelet-derived growth factor (PDGF)-AB and the soluble CD40 ligand (sCD40L) release were significantly reduced by S1P. In addition, S1P suppressed the collagen-induced release of HSP27 as well as the phosphorylation of HSP27. S1P significantly suppressed the collagen-induced phosphorylation of p38 mitogen-activated protein kinase. S1P increased the levels of GTP-bound Gαi and GTP-bound Gα13 coupled to S1PPR1 and/or S1PR4. CYM50260, a selective S1PR4 agonist, but not SEW2871, a selective S1PR1 agonist, suppressed the collagen-stimulated platelet aggregation, PDGF-AB secretion and sCD40L release. In addition, CYM50260 reduced the release of phosphorylated-HSP27 by collagen as well as the phosphorylation of HSP27. The selective S1PR4 antagonist CYM50358, which failed to affect collagen-induced HSP27 phosphorylation, reversed the S1P-induced attenuation of HSP27 phosphorylation by collagen. These results strongly suggest that S1P inhibits the collagen-induced human platelet activation through S1PR4 but not S1PR1. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Differentiating human versus non-human bone by exploring the nutrient foramen: implications for forensic anthropology.

    Science.gov (United States)

    Johnson, Vail; Beckett, Sophie; Márquez-Grant, Nicholas

    2017-11-01

    One of the roles of a forensic anthropologist is to assist medico-legal investigations in the identification of human skeletal remains. In some instances, only small fragments of bone may be present. In this study, a non-destructive novel technique is presented to distinguish between human and non-human long bones. This technique is based on the macroscopic and computed tomography (CT) analysis of nutrient foramina. The nutrient foramen of long bone diaphyses transmits the nutrient artery which provides much of the oxygen and nutrients to the bone. The nutrient foramen and its canal were analysed in six femora and humeri of human, sheep (Ovies aries) and pig (Sus scrofa) species. The location, position and direction of the nutrient foramina were measured macroscopically. The length of the canal, angle of the canal, circumference and area of the entrance of the foramen were measured from CT images. Macroscopic analysis revealed the femora nutrient foramina are more proximal, whereas humeri foramina are more distal. The human bones and sheep humerus conform to the perceived directionality, but the pig bones and sheep femur do not. Amongst the parameters measured in the CT analysis, the angle of the canal had a discriminatory power. This study shows the potential of this technique to be used independently or complementary to other methods in distinguishing between human and non-human bone in forensic anthropology.

  20. Tissue engineering for lateral ridge augmentation with recombinant human bone morphogenetic protein 2 combination therapy: a case report.

    Science.gov (United States)

    Mandelaris, George A; Spagnoli, Daniel B; Rosenfeld, Alan L; McKee, James; Lu, Mei

    2015-01-01

    This case report describes a tissue-engineered reconstruction with recombinant human bone morphogenetic protein 2/acellular collagen sponge (rhBMP-2/ ACS) + cancellous allograft and space maintenance via Medpor Contain mesh in the treatment of a patient requiring maxillary and mandibular horizontal ridge augmentation to enable implant placement. The patient underwent a previously unsuccessful corticocancellous bone graft at these sites. Multiple and contiguous sites in the maxilla and in the mandibular anterior, demonstrating advanced lateral ridge deficiencies, were managed using a tissue engineering approach as an alternative to autogenous bone harvesting. Four maxillary and three mandibular implants were placed 9 and 10 months, respectively, after tissue engineering reconstruction, and all were functioning successfully after 24 months of follow-up. Histomorphometric analysis of a bone core obtained at the time of the maxillary implant placement demonstrated a mean of 76.1% new vital bone formation, 22.2% marrow/cells, and 1.7% residual graft tissue. Tissue engineering for lateral ridge augmentation with combination therapy requires further research to determine predictability and limitations.

  1. Distribution of radium and plutonium in human bone

    International Nuclear Information System (INIS)

    Schlenker, R.A.

    1985-01-01

    This review covers studies of the microdistribution of radium and plutonium in human bone, conducted at Argonne with emphasis on the alpha-spectrometric method of measurement. Alpha spectrometry offers high spatial resolution and is well suited to the measurement of radionuclide concentrations near bone surfaces. With these techniques surface deposit thicknesses have been measured to be about 1 μm thick for isotopes of lead, radium and the actinides, and volume deposits of 226 Ra have been found to be quite nonuniform near bone surfaces, leading to endosteal tissue dose rates that are higher than expected under the assumption of uniform volume concentration normally used in radiation protection calculations. With autoradiography, the bony septa of the mastoid air cell system have been found to be depleted in radium relative to the bone tissue surrounding them; this is expected to have a significant influence on the dosimetry of the mastoid epithelia. A combination of autoradiographic and morphometric measurements indicates that specific activities in the axial skeleton are higher than in the appendicular skeleton, primarily because the former has higher bone surface-to-volume ratios and higher bone surface concentrations of plutonium. 19 references, 14 figures, 6 tables

  2. Crestal Sinus Augmentation with Recombinant Human Bone Morphogenetic Protein 2: Clinical and Radiographic Outcomes of 2-Year Pilot Trial.

    Science.gov (United States)

    Kuchler, Ulrike; Rudelstorfer, Claudia M; Barth, Barbara; Tepper, Gabor; Lidinsky, Dominika; Heimel, Patrick; Watzek, Georg; Gruber, Reinhard

    Recombinant human bone morphogenetic protein 2 (rhBMP-2) together with an absorbable collagen carrier (ACS) was approved for augmentation of the maxillary sinus prior to implant placement. The original registration trial was based on a lateral window approach. Clinical outcomes of crestal sinus augmentation with rhBMP-2 have not been reported so far. An uncontrolled pilot trial in which seven patients with a residual maxillary height below 5 mm were enrolled to receive crestal sinus augmentation with rhBMP-2/ACS was conducted. Elevation of the sinus mucosa was performed by gel pressure. Primary endpoints were the gain in augmentation height and volume measured by computed tomography after 6 months. Evaluation of bone quality at the time of implant placement was based on histology. Secondary endpoints were the clinical and radiologic evaluation of the implants and patient satisfaction by visual analog scale (VAS) at the 2-year follow-up. Median gain in augmentation height was 7.2 mm (range 0.0 to 17.5 mm). Five patients gained at least 5 mm of bone height. Two patients with a perforation of the sinus mucosa failed to respond to rhBMP-2/ACS and underwent lateral window augmentation. The median gain in augmentation volume of the five patients was 781.3 mm³ (range 426.9 to 1,242.8 mm³). Biopsy specimens showed a cancellous network consisting of primary plexiform bone with little secondary lamellar bone. After 2 years, implants were in function with no signs of inflammation or peri-implant bone loss. Patients were satisfied with the esthetic outcomes and chewing function. This pilot clinical trial supports the original concept that rhBMP-2/ACS supports bone formation, also in crestal sinus augmentation, and emphasizes the relevance of the integrity of the sinus mucosa to predict the bone gain.

  3. Strontium-90 content of human bone collected in 1967

    International Nuclear Information System (INIS)

    Jeanmaire, L.; Patti, F.

    1969-01-01

    This report follows report CEA-R-3381 and presents the strontium 90 content of human bones collected in 1967 in the Paris area. The main trend is much the same as during 1966; contamination levels are falling down in infants up to 5 year old. Beyond this age, the values are the same or experience a slight increase. (authors) [fr

  4. Ancient pathogen DNA in human teeth and petrous bones

    DEFF Research Database (Denmark)

    Margaryan, Ashot; Hansen, Henrik B.; Rasmussen, Simon

    2018-01-01

    Recent ancient DNA (aDNA) studies of human pathogens have provided invaluable insights into their evolutionary history and prevalence in space and time. Most of these studies were based on DNA extracted from teeth or postcranial bones. In contrast, no pathogen DNA has been reported from the petro...

  5. Specific depletion of mature T lymphocytes from human bone marrow

    DEFF Research Database (Denmark)

    Geisler, C; Møller, J; Plesner, T

    1989-01-01

    An effective method for specific depletion of mature T lymphocytes from human bone marrow mononuclear cells (BMMC) with preservation of prethymic T cells and natural killer (NK) cells is presented. The BMMC were incubated with F101.01, a monoclonal antibody recognizing an epitope of the T...

  6. Serum Bone Resorption Markers after Parathyroidectomy for Renal Hyperparathyroidism: Correlation Analyses for the Cross-Linked N-telopeptide of Collagen I and Tartrate-Resistant Acid Phosphatase

    Directory of Open Access Journals (Sweden)

    Kuo-Chin Hung

    2012-01-01

    Full Text Available Patients on long-term dialysis may develop secondary hyperparathyroidism (SHPT with increased serum concentrations of bone resorption markers such as the cross-linked N-telopeptide of type I collagen (NTX and type-5b tartrate-resistant acid phosphatase (TRAP. When SHPT proves refractory to treatment, parathyroidectomy (PTX may be needed. Renal patients on maintenance HD who received PTX for refractory SHPT (n=23 or who did not develop refractory SHPT (control subjects; n=25 were followed prospectively for 4 weeks. Serum intact parathyroid hormone (iPTH, NTX, TRAP, and bone alkaline phosphatase (BAP concentrations were measured serially and correlation analyses were performed. iPTH values decreased rapidly and dramatically. BAP values increased progressively with peak increases observed at 2 weeks after surgery. NTX and TRAP values decreased concurrently and progressively through 4 weeks following PTX. A significant correlation between TRAP and NTX values was observed before PTX but not at 4 weeks after PTX. Additionally, the fractional changes in serum TRAP were larger than those in serum NTX at all times examined after PTX. Serum iPTH, TRAP, and NTX values declined rapidly following PTX for SHPT. Serum TRAP values declined to greater degrees than serum NTX values throughout the 4-week period following PTX.

  7. The effect of growth factors on both collagen synthesis and tensile strength of engineered human ligaments.

    Science.gov (United States)

    Hagerty, Paul; Lee, Ann; Calve, Sarah; Lee, Cassandra A; Vidal, Martin; Baar, Keith

    2012-09-01

    Growth factors play a central role in the development and remodelling of musculoskeletal tissues. To determine which growth factors optimized in vitro ligament formation and mechanics, a Box-Behnken designed array of varying concentrations of growth factors and ascorbic acid were applied to engineered ligaments and the collagen content and mechanics of the grafts were determined. Increasing the amount of transforming growth factor (TGF) β1 and insulin-like growth factor (IGF)-1 led to an additive effect on ligament collagen and maximal tensile load (MTL). In contrast, epidermal growth factor (EGF) had a negative effect on both collagen content and MTL. The predicted optimal growth media (50 μg/ml TGFβ, IGF-1, and GDF-7 and 200 μM ascorbic acid) was then validated in two separate trials: showing a 5.7-fold greater MTL and 5.2-fold more collagen than a minimal media. Notably, the effect of the maximized growth media was scalable such that larger constructs developed the same material properties, but larger MTL. These results show that optimizing the interactions between growth factors and engineered ligament volume results in an engineered ligament of clinically relevant function. Copyright © 2012 Elsevier Ltd. All rights reserved.

  8. Secondary cytotoxicity of (crosslinked) dermal sheep collagen during repeated exposure to human fibroblasts

    NARCIS (Netherlands)

    van Luyn, M.J.A.; van Wachem, P.B.; Olde damink, L.H.H.; Olde Damink, L.H.H.; Dijkstra, Pieter J.; Feijen, Jan; Nieuwenhuis, P.

    1992-01-01

    We investigated commercially available dermal sheep collagen either cross-linked with hexamethylenediisocyanate, or cross-linked with glutaraldehyde. In previous in vitro studies we could discriminate primary, i.e. extractable, and secondary cytotoxicity, due to cell-biomaterial interactions, i.e.

  9. Effect of Collagen Type I or Type II on Chondrogenesis by Cultured Human Articular Chondrocytes

    NARCIS (Netherlands)

    Rutgers, M.; Saris, Daniël B.F.; Vonk, L.A.; van Rijen, M.H.P.; Akrum, V.; Langeveld, D.; van Boxtel, A.; Dhert, W.J.A.; Creemers, L.B.

    2013-01-01

    Introduction: Current cartilage repair procedures using autologous chondrocytes rely on a variety of carriers for implantation. Collagen types I and II are frequently used and valuable properties of both were shown earlier in vitro, although a preference for either was not demonstrated. Recently,

  10. SECONDARY CYTOTOXICITY OF CROSS-LINKED DERMAL SHEEP COLLAGENS DURING REPEATED EXPOSURE TO HUMAN FIBROBLASTS

    NARCIS (Netherlands)

    VANLUYN, MJA; VANWACHEM, PB; DAMINK, LHHO; DIJKSTRA, PJ; FEIJEN, J; NIEUWENHUIS, P

    1992-01-01

    We investigated commercially available dermal sheep collagen either cross-linked with hexamethylenedlisocyanate, or cross-linked with glutaraldehyde. In previous in vitro studies we could discriminate primary, i.e. extractable, and secondary cytotoxicity, due to cell-biomaterial interactions, i.e.

  11. Effect of albumin-bound DHA on phosphoinositide phosphorylation in collagen stimulated human platelets

    International Nuclear Information System (INIS)

    Gaudette, D.C.; Holub, B.J.

    1990-01-01

    The effect of exogenous albumin-bound docosahexaenoic acid (22:6n-3) (DHA), arachidonic acid (20:4n-6) (AA), and eicosapendaenoic acid (20:5n-3) (EPA) on phosphoinositide metabolism following collagen stimulation was studied using [3H]inositol prelabelled platelets. Collagen stimulation (3 min, 1.8 micrograms/ml) increased the labelling of both phosphatidylinositol 4-monophosphate (PIP), and phosphatidylinositol 4,5-biphosphate (PIP2). Of the fatty acids tested, only pre-incubation (2 min) with DHA (20 microM) significantly attenuated the collagen-induced increased PIP and PIP2 labelling; EPA was without effect, while AA enhanced PIP labelling. Forty microM DHA was less effective at attenuating the increased PIP and PIP2 labelling even though this concentration of DHA resulted in greater inhibition of platelet aggregation. Neither concentration of DHA attenuated the increased polyphosphoinositide labelling resulting from stimulation by the endoperoxide analogue U46619, or the phorbol ester, PMA. These data suggest that the effect of DHA on attenuating the increased PIP and PIP2 labelling following collagen stimulation likely occurs before thromboxane receptor occupancy, may not occur at the level of protein kinase C activation, and could be mediated in part via a lessened synthesis of thromboxane A2

  12. Supramolecular Organization of Collagen Fibrils in Healthy and Osteoarthritic Human Knee and Hip Joint Cartilage.

    Directory of Open Access Journals (Sweden)

    Riccardo Gottardi

    Full Text Available Cartilage matrix is a composite of discrete, but interacting suprastructures, i.e. cartilage fibers with microfibrillar or network-like aggregates and penetrating extrafibrillar proteoglycan matrix. The biomechanical function of the proteoglycan matrix and the collagen fibers are to absorb compressive and tensional loads, respectively. Here, we are focusing on the suprastructural organization of collagen fibrils and the degradation process of their hierarchical organized fiber architecture studied at high resolution at the authentic location within cartilage. We present electron micrographs of the collagenous cores of such fibers obtained by an improved protocol for scanning electron microscopy (SEM. Articular cartilages are permeated by small prototypic fibrils with a homogeneous diameter of 18 ± 5 nm that can align in their D-periodic pattern and merge into larger fibers by lateral association. Interestingly, these fibers have tissue-specific organizations in cartilage. They are twisted ropes in superficial regions of knee joints or assemble into parallel aligned cable-like structures in deeper regions of knee joint- or throughout hip joints articular cartilage. These novel observations contribute to an improved understanding of collagen fiber biogenesis, function, and homeostasis in hyaline cartilage.

  13. Indirect MR-arthrography in osteochondral autograft and crushed bone graft with a collagen membrane-Correlation with histology

    Energy Technology Data Exchange (ETDEWEB)

    Streitparth, F. [Klinik fuer Strahlenheilkunde, Charite, Campus Virchow-Klinikum, Humboldt-University, Berlin (Germany)], E-mail: florian.streitparth@charite.de; Schoettle, P.; Schell, H. [Center for Musculoskeletal Surgery, Charite, Campus Virchow-Klinikum, Humboldt-University, Berlin (Germany); Lehmkuhl, L.; Madej, T.; Wieners, G. [Klinik fuer Strahlenheilkunde, Charite, Campus Virchow-Klinikum, Humboldt-University, Berlin (Germany); Duda, G.N. [Center for Musculoskeletal Surgery, Charite, Campus Virchow-Klinikum, Humboldt-University, Berlin (Germany); Schroeder, R.J. [Klinik fuer Strahlenheilkunde, Charite, Campus Virchow-Klinikum, Humboldt-University, Berlin (Germany)

    2009-04-15

    Objective: To analyze the spectrum of findings in indirect MR-arthrography following osteochondral autograft transfer system (OATS) and crushed bone graft using a magnetic resonance imaging (MRI) scoring and grading system in relation to histology as the standard of reference. Materials and methods: Iatrogenic lesions were set at ovine condylar facets (n = 6/group), treated with OATS or crushed bone graft. 1.5 T MRI was performed 6 months after surgery using PD-weighted (w fat saturated (fs) fast spin echo (FSE), T1-w 2D, and 3D fs gradient echo (GE) sequences 30 min. after i.v. Gd-DTPA administration and passive joint exercise. The repair tissue was evaluated by two independent radiologists. The MR findings were compared to histology. Results: In all cases, MRI and histologic grading correlated well and showed significant superior repair in OATS at 6 months (p < 0.05), reproducing the original articular contour and a good subchondral restoration. FsT1-w3DGE proved most appropriate identifying characteristic post-operative findings: the OATS group demonstrated bone marrow edema at the donor site and the graft/host interface showed significant enhancement in indirect MR-arthrography, indicating fibrocartilage. After crushed bone graft, we found an irregular structure and significant contrast uptake, consistent with remnants of bone grafts surrounded by inflammatory tissue. Conclusion: Indirect MR-arthrography is an accurate, non-invasive monitoring tool following OATS and crushed bone graft as the MRI scoring and grading system allows a reliable evaluation of normal and pathological osteochondral repair with a high histologic correlation.

  14. Functional collagen conduits combined with human mesenchymal stem cells promote regeneration after sciatic nerve transection in dogs.

    Science.gov (United States)

    Cui, Yi; Yao, Yao; Zhao, Yannan; Xiao, Zhifeng; Cao, Zongfu; Han, Sufang; Li, Xing; Huan, Yong; Pan, Juli; Dai, Jianwu

    2018-05-01

    Numerous studies have focused on the development of novel and innovative approaches for the treatment of peripheral nerve injury using artificial nerve guide conduits. In this study, we attempted to bridge 3.5-cm defects of the sciatic nerve with a longitudinally oriented collagen conduit (LOCC) loaded with human umbilical cord mesenchymal stem cells (hUC-MSCs). The LOCC contains a bundle of longitudinally aligned collagenous fibres enclosed in a hollow collagen tube. Our previous studies showed that an LOCC combined with neurotrophic factors enhances peripheral nerve regeneration. However, it remained unknown whether an LOCC seeded with hUC-MSCs could also promote regeneration. In this study, using various histological and electrophysiological analyses, we found that an LOCC provides mechanical support to newly growing nerves and functions as a structural scaffold for cells, thereby stimulating sciatic nerve regeneration. The LOCC and hUC-MSCs synergistically promoted regeneration and improved the functional recovery in a dog model of sciatic nerve injury. Therefore, the combined use of an LOCC and hUC-MSCs might have therapeutic potential for the treatment of peripheral nerve injury. Copyright © 2018 John Wiley & Sons, Ltd.

  15. Original Article. Toxic effect of sodium fluoride on hydroxyproline level and expression of collagen-1 gene in rat bone and its amelioration by Tamrindus indica L. fruit pulp extract

    Directory of Open Access Journals (Sweden)

    Gupta Amit Raj

    2016-03-01

    Full Text Available Excessive fluoride intoxication plays an important role in the development of dental, skeletal and non-skeletal fluorosis. The aim of this study was to ascertain the toxic effect of excessive fluoride ingestion on the level of hydroxyproline and expression of type 1 collagen gene in rat bone and its amelioration by supplementation with Tamarindus indica fruit pulp extract. Forty albino rats were randomly assigned to four groups. The first group served as control and received only tap water. The second group received sodium fluoride (200 ppm through drinking water. The third group received T. indica fruit pulp extract (200 mg/kg body weight alone and the fourth group received the T. indica fruit pulp extract (200 mg/kg body weight along with fluorinated drinking water (200 ppm daily by gavage for a period of 90 days. The level of hydroxyproline and expression of type 1 collagen gene using quantitative real time PCR in the tibia bone decreased significantly with continuous exposure to sodium fluoride. Co-administration of T. indica fruit pulp extract during exposure to fluoride through drinking water restored the level of calcium, phosphorus and alkaline phosphatase in serum and the concentration of hydroxyproline in urine. It increased the level of hydroxyproline and expression of type 1 collagen gene in the tibia as compared to untreated fluoride-exposed rats. It is concluded that T. indica fruit pulp extract has an ameliorative potential to protect the bone from fluoride induced collagen damage.

  16. Glaucoma-related Changes in the Mechanical Properties and Collagen Micro-architecture of the Human Sclera

    Science.gov (United States)

    Coudrillier, Baptiste; Pijanka, Jacek K.; Jefferys, Joan L.; Goel, Adhiraj; Quigley, Harry A.; Boote, Craig; Nguyen, Thao D.

    2015-01-01

    Objective The biomechanical behavior of the sclera determines the level of mechanical insult from intraocular pressure to the axons and tissues of the optic nerve head, as is of interest in glaucoma. In this study, we measure the collagen fiber structure and the strain response, and estimate the material properties of glaucomatous and normal human donor scleras. Methods Twenty-two posterior scleras from normal and diagnosed glaucoma donors were obtained from an eyebank. Optic nerve cross-sections were graded to determine the presence of axon loss. The specimens were subjected to pressure-controlled inflation testing. Full-field displacement maps were measured by digital image correlation (DIC) and spatially differentiated to compute surface strains. Maps of the collagen fiber structure across the posterior sclera of each inflated specimen were obtained using synchrotron wide-angle X-ray scattering (WAXS). Finite element (FE) models of the posterior scleras, incorporating a specimen-specific representation of the collagen structure, were constructed from the DIC-measured geometry. An inverse finite element analysis was developed to estimate the stiffness of the collagen fiber and inter-fiber matrix. Results The differences between glaucoma and non-glaucoma eyes were small in magnitude. Sectorial variations of degree of fiber alignment and peripapillary scleral strain significantly differed between normal and diagnosed glaucoma specimens. Meridional strains were on average larger in diagnosed glaucoma eyes compared with normal specimens. Non-glaucoma specimens had on average the lowest matrix and fiber stiffness, followed by undamaged glaucoma eyes, and damaged glaucoma eyes but the differences in stiffness were not significant. Conclusion The observed biomechanical and microstructural changes could be the result of tissue remodeling occuring in glaucoma and are likely to alter the mechanical environment of the optic nerve head and contribute to axonal damage. PMID

  17. Type VII collagen is enriched in the enamel organic matrix associated with the dentin-enamel junction of mature human teeth.

    Science.gov (United States)

    McGuire, Jacob D; Walker, Mary P; Mousa, Ahmad; Wang, Yong; Gorski, Jeff P

    2014-06-01

    The inner enamel region of erupted teeth is known to exhibit higher fracture toughness and crack growth resistance than bulk phase enamel. However, an explanation for this behavior has been hampered by the lack of compositional information for the residual enamel organic matrix. Since enamel-forming ameloblasts are known to express type VII collagen and type VII collagen null mice display abnormal amelogenesis, the aim of this study was to determine whether type VII collagen is a component of the enamel organic matrix at the dentin-enamel junction (DEJ) of mature human teeth. Immunofluorescent confocal microscopy of demineralized tooth sections localized type VII collagen to the organic matrix surrounding individual enamel rods near the DEJ. Morphologically, immunoreactive type VII collagen helical-bundles resembled the gnarled-pattern of enamel rods detected by Coomassie Blue staining. Western blotting of whole crown or enamel matrix extracts also identified characteristic Mr=280 and 230 kDa type VII dimeric forms, which resolved into 75 and 25 kDa bands upon reduction. As expected, the collagenous domain of type VII collagen was resistant to pepsin digestion, but was susceptible to purified bacterial collagenase. These results demonstrate the inner enamel organic matrix in mature teeth contains macromolecular type VII collagen. Based on its physical association with the DEJ and its well-appreciated capacity to complex with other collagens, we hypothesize that enamel embedded type VII collagen fibrils may contribute not only to the structural resilience of enamel, but may also play a role in bonding enamel to dentin. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. Rotating three-dimensional dynamic culture of adult human bone marrow-derived cells for tissue engineering of hyaline cartilage.

    Science.gov (United States)

    Sakai, Shinsuke; Mishima, Hajime; Ishii, Tomoo; Akaogi, Hiroshi; Yoshioka, Tomokazu; Ohyabu, Yoshimi; Chang, Fei; Ochiai, Naoyuki; Uemura, Toshimasa

    2009-04-01

    The method of constructing cartilage tissue from bone marrow-derived cells in vitro is considered a valuable technique for hyaline cartilage regenerative medicine. Using a rotating wall vessel (RWV) bioreactor developed in a NASA space experiment, we attempted to efficiently construct hyaline cartilage tissue from human bone marrow-derived cells without using a scaffold. Bone marrow aspirates were obtained from the iliac crest of nine patients during orthopedic operation. After their proliferation in monolayer culture, the adherent cells were cultured in the RWV bioreactor with chondrogenic medium for 2 weeks. Cells from the same source were cultured in pellet culture as controls. Histological and immunohistological evaluations (collagen type I and II) and quantification of glycosaminoglycan were performed on formed tissues and compared. The engineered constructs obtained using the RWV bioreactor showed strong features of hyaline cartilage in terms of their morphology as determined by histological and immunohistological evaluations. The glycosaminoglycan contents per microg DNA of the tissues were 10.01 +/- 3.49 microg/microg DNA in the case of the RWV bioreactor and 6.27 +/- 3.41 microg/microg DNA in the case of the pellet culture, and their difference was significant. The RWV bioreactor could provide an excellent environment for three-dimensional cartilage tissue architecture that can promote the chondrogenic differentiation of adult human bone marrow-derived cells.

  19. Three-dimensional, bioactive, biodegradable, polymer-bioactive glass composite scaffolds with improved mechanical properties support collagen synthesis and mineralization of human osteoblast-like cells in vitro.

    Science.gov (United States)

    Lu, Helen H; El-Amin, Saadiq F; Scott, Kimberli D; Laurencin, Cato T

    2003-03-01

    In the past decade, tissue engineering-based bone grafting has emerged as a viable alternative to biological and synthetic grafts. The biomaterial component is a critical determinant of the ultimate success of the tissue-engineered graft. Because no single existing material possesses all the necessary properties required in an ideal bone graft, our approach has been to develop a three dimensional (3-D), porous composite of polylactide-co-glycolide (PLAGA) and 45S5 bioactive glass (BG) that is biodegradable, bioactive, and suitable as a scaffold for bone tissue engineering (PLAGA-BG composite). The objectives of this study were to examine the mechanical properties of a PLAGA-BG matrix, to evaluate the response of human osteoblast-like cells to the PLAGA-BG composite, and to evaluate the ability of the composite to form a surface calcium phosphate layer in vitro. Structural and mechanical properties of PLAGA-BG were measured, and the formation of a surface calcium phosphate layer was evaluated by surface analysis methods. The growth and differentiation of human osteoblast-like cells on PLAGA-BG were also examined. A hypothesis was that the combination of PLAGA with BG would result in a biocompatible and bioactive composite, capable of supporting osteoblast adhesion, growth and differentiation, with mechanical properties superior to PLAGA alone. The addition of bioactive glass granules to the PLAGA matrix resulted in a structure with higher compressive modulus than PLAGA alone. Moreover, the PLAGA-BA composite was found to be a bioactive material, as it formed surface calcium phosphate deposits in a simulated body fluid (SBF), and in the presence of cells and serum proteins. The composite supported osteoblast-like morphology, stained positively for alkaline phosphatase, and supported higher levels of Type I collagen synthesis than tissue culture polystyrene controls. We have successfully developed a degradable, porous, polymer bioactive glass composite possessing

  20. Survival of human mesenchymal stromal cells from bone marrow and adipose tissue after xenogenic transplantation in immunocompetent mice

    DEFF Research Database (Denmark)

    Niemeyer, P; Vohrer, J; Schmal, H

    2008-01-01

    of the current paper was to evaluate the survival of undifferentiated and osteogenically induced human MSC from different origins after transplantation in immunocompetent mice. METHODS: Human MSC were isolated from bone marrow (BMSC) and adipose tissue (ASC). After cultivation on mineralized collagen, MSC were......INTRODUCTION: Mesenchymal stromal cells (MSC) represent an attractive cell population for tissue engineering purposes. As MSC are described as immunoprivileged, non-autologous applications seem possible. A basic requirement is the survival of MSC after transplantation in the host. The purpose...... transplanted subcutaneously into immunocompetent mice (n=12). Undifferentiated MSC (group A) were compared with osteogenic-induced MSC (group B). Human-specific in situ hybridization and anti-vimentin staining was used to follow MSC after transplantation. Quantitative evaluation of lymphocytes and macrophages...