Science.gov

Sample records for collagen fibril surface

  1. Surface-Sensitive Raman Spectroscopy of Collagen I Fibrils

    PubMed Central

    Gullekson, Corinne; Lucas, Leanne; Hewitt, Kevin; Kreplak, Laurent

    2011-01-01

    Collagen fibrils are the main constituent of the extracellular matrix surrounding eukaryotic cells. Although the assembly and structure of collagen fibrils is well characterized, very little appears to be known about one of the key determinants of their biological function—namely, the physico-chemical properties of their surface. One way to obtain surface-sensitive structural and chemical data is to take advantage of the near-field nature of surface- and tip-enhanced Raman spectroscopy. Using Ag and Au nanoparticles bound to Collagen type-I fibrils, as well as tips coated with a thin layer of Ag, we obtained Raman spectra characteristic to the first layer of collagen molecules at the surface of the fibrils. The most frequent Raman peaks were attributed to aromatic residues such as phenylalanine and tyrosine. In several instances, we also observed Amide I bands with a full width at half-maximum of 10–30 cm−1. The assignment of these Amide I band positions suggests the presence of 310-helices as well as α- and β-sheets at the fibril's surface. PMID:21463598

  2. Collagen fibril surface displays a constellation of sites capable of promoting fibril assembly, stability, and hemostasis

    SciTech Connect

    Orgel, J.P.; Antipova, O.; Sagi, I.; Bitler, A.; Qiu, D.; Wang, R.; Xu, Y.; San Antonio, J.D.

    2011-12-14

    Fibrillar collagens form the structural basis of organs and tissues including the vasculature, bone, and tendon. They are also dynamic, organizational scaffolds that present binding and recognition sites for ligands, cells, and platelets. We interpret recently published X-ray diffraction findings and use atomic force microscopy data to illustrate the significance of new insights into the functional organization of the collagen fibril. These data indicate that collagen's most crucial functional domains localize primarily to the overlap region, comprising a constellation of sites we call the 'master control region.' Moreover, the collagen's most exposed aspect contains its most stable part - the C-terminal region that controls collagen assembly, cross-linking, and blood clotting. Hidden beneath the fibril surface exists a constellation of 'cryptic' sequences poised to promote hemostasis and cell - collagen interactions in tissue injury and regeneration. These findings begin to address several important, and previously unresolved, questions: How functional domains are organized in the fibril, which domains are accessible, and which require proteolysis or structural trauma to become exposed? Here we speculate as to how collagen fibrillar organization impacts molecular processes relating to tissue growth, development, and repair.

  3. Decorin Core Protein (Decoron) Shape Complements Collagen Fibril Surface Structure and Mediates Its Binding

    SciTech Connect

    Orgel, Joseph P.R.O.; Eid, Aya; Antipova, Olga; Bella, Jordi; Scott, John E.

    2010-02-11

    Decorin is the archetypal small leucine rich repeat proteoglycan of the vertebrate extracellular matrix (ECM). With its glycosaminoglycuronan chain, it is responsible for stabilizing inter-fibrillar organization. Type I collagen is the predominant member of the fibrillar collagen family, fulfilling both organizational and structural roles in animal ECMs. In this study, interactions between decoron (the decorin core protein) and binding sites in the d and e1 bands of the type I collagen fibril were investigated through molecular modeling of their respective X-ray diffraction structures. Previously, it was proposed that a model-based, highly curved concave decoron interacts with a single collagen molecule, which would form extensive van der Waals contacts and give rise to strong non-specific binding. However, the large well-ordered aggregate that is the collagen fibril places significant restraints on modes of ligand binding and necessitates multi-collagen molecular contacts. We present here a relatively high-resolution model of the decoron-fibril collagen complex. We find that the respective crystal structures complement each other well, although it is the monomeric form of decoron that shows the most appropriate shape complementarity with the fibril surface and favorable calculated energies of interaction. One molecule of decoron interacts with four to six collagen molecules, and the binding specificity relies on a large number of hydrogen bonds and electrostatic interactions, primarily with the collagen motifs KXGDRGE and AKGDRGE (d and e{sub 1} bands). This work helps us to understand collagen-decorin interactions and the molecular architecture of the fibrillar ECM in health and disease.

  4. Electrostatic effects in collagen fibrillization

    NASA Astrophysics Data System (ADS)

    Morozova, Svetlana; Muthukumar, Murugappan

    2014-03-01

    Using light scattering and AFM techniques, we have measured the kinetics of fibrillization of collagen (pertinent to the vitreous of human eye) as a function of pH and ionic strength. At higher and lower pH, collagen triple-peptides remain stable in solution without fibrillization. At neutral pH, the fibrillization occurs and its growth kinetics is slowed upon either an increase in ionic strength or a decrease in temperature. We present a model, based on polymer crystallization theory, to describe the observed electrostatic nature of collagen assembly.

  5. Collagen fibril formation during development

    SciTech Connect

    Fleischmajer, R.; Perlish, J.S.; Timpl, R.; Olsen, B.R.

    1987-05-01

    Studies with embryonic skin and bone suggested that the aminopropeptide (AP) and carboxylpropeptide (CP) of type I pro-callagen (pro-col) play a role in fibril formation. Chick leg metatarsal tendons were studied by electron microscopy. AP and CP of type I pro-col were purified from chick leg tendons; antibodies developed in rabbits and purity tested by radioimmunoassays. Antibodies were used for immunofluorescence microscopy (IFM) and immunoblotting (IB). The peritendineum, consisting of thin 20-30 nm fibrils, revealed the AP of type I and type III procol. In the tendon area, collagen fibrils were arranged within small compartments and were of uniform diameter at 10d, 14d and 18d. However, beyond 21d, there was confluency of the compartments and a wide range of fibril diameters. IFM revealed fine streaks of collagen, staining with the AP of type I throughout the tendon. The CP was mainly intracellular with only a small amount present in the extracellular space. IB revealed procollagen, pN-collagen (AP+collagen) and pC-collagen, (CP+collagen) at all stages of development. Ratios of pN/pC collagen, determined by spectrophotometric scanning of autoradiographs, correlated well with the distribution of fibril diameter. This study suggests the hypothesis that AP initiates fibrillogenesis while CP may regulate additional fibril growth.

  6. Thermal Memory in Self-Assembled Collagen Fibril Networks

    PubMed Central

    de Wild, Martijn; Pomp, Wim; Koenderink, Gijsje H.

    2013-01-01

    Collagen fibrils form extracellular networks that regulate cell functions and provide mechanical strength to tissues. Collagen fibrillogenesis is an entropy-driven process promoted by warming and reversed by cooling. Here, we investigate the influence of noncovalent interactions mediated by the collagen triple helix on fibril stability. We measure the kinetics of cold-induced disassembly of fibrils formed from purified collagen I using turbimetry, probe the fibril morphology by atomic force microscopy, and measure the network connectivity by confocal microscopy and rheometry. We demonstrate that collagen fibrils disassemble by subunit release from their sides as well as their ends, with complex kinetics involving an initial fast release followed by a slow release. Surprisingly, the fibrils are gradually stabilized over time, leading to thermal memory. This dynamic stabilization may reflect structural plasticity of the collagen fibrils arising from their complex structure. In addition, we propose that the polymeric nature of collagen monomers may lead to slow kinetics of subunit desorption from the fibril surface. Dynamic stabilization of fibrils may be relevant in the initial stages of collagen assembly during embryogenesis, fibrosis, and wound healing. Moreover, our results are relevant for tissue repair and drug delivery applications, where it is crucial to control fibril stability. PMID:23823240

  7. Type V collagen controls the initiation of collagen fibril assembly.

    PubMed

    Wenstrup, Richard J; Florer, Jane B; Brunskill, Eric W; Bell, Sheila M; Chervoneva, Inna; Birk, David E

    2004-12-17

    Vertebrate collagen fibrils are heterotypically composed of a quantitatively major and minor fibril collagen. In non-cartilaginous tissues, type I collagen accounts for the majority of the collagen mass, and collagen type V, the functions of which are poorly understood, is a minor component. Type V collagen has been implicated in the regulation of fibril diameter, and we reported recently preliminary evidence that type V collagen is required for collagen fibril nucleation (Wenstrup, R. J., Florer, J. B., Cole, W. G., Willing, M. C., and Birk, D. E. (2004) J. Cell. Biochem. 92, 113-124). The purpose of this study was to define the roles of type V collagen in the regulation of collagen fibrillogenesis and matrix assembly. Mouse embryos completely deficient in pro-alpha1(V) chains were created by homologous recombination. The col5a1-/- animals die in early embryogenesis, at approximately embryonic day 10. The type V collagen-deficient mice demonstrate a virtual lack of collagen fibril formation. In contrast, the col5a1+/- animals are viable. The reduced type V collagen content is associated with a 50% reduction in fibril number and dermal collagen content. In addition, relatively normal, cylindrical fibrils are assembled with a second population of large, structurally abnormal collagen fibrils. The structural properties of the abnormal matrix are decreased relative to the wild type control animals. These data indicate a central role for the evolutionary, ancient type V collagen in the regulation of fibrillogenesis. The complete dependence of fibril formation on type V collagen is indicative of the critical role of the latter in early fibril initiation. In addition, this fibril collagen is important in the determination of fibril structure and matrix organization. PMID:15383546

  8. The collagen fibril organization in human articular cartilage.

    PubMed Central

    Minns, R J; Steven, F S

    1977-01-01

    In this scanning electron microscopic study blocks of collagen fibrils were prepared from human articular cartilage, using two techinques which selectively removed either the proteoglycans alone, or both the proteoglycans and the collagen fibrils, of the non-calcified cartilage layer. Amino acid analysis of the fibrils confirmed the purity of the collagen after proteoglycan extraction. The cartilage was scanned in four different ways: (1) normal to the articular surface, (2) in superficial sections, (3) on surfaces of blocks which had been broken in planes parallel to artificial splits make by the insertion of a pin, and (4) on fracture surfaces which traversed the calcified cartilage and the subchondral bone. Five features of the organization of the collagen fibrils were specially noted: (1) Individual fibrils within the trabeculae joined to form small fibre bundles which became grouped into larger bundles at the calcified/uncalcified interface. (2) Fibrils in the deep and middle zones which, exhibiting the characteristic surface periodicity of collagen, were generally oriented towars the articular surface in large bundles approximately 55 micronm across. (3) In the superficial zone, fibrils ran parallel to the surface. (4) The surface fibrils had random orientation, even at the bases of empty lacunae vacated by chondrocytes during specimen preparation. (5) The collagen fibrils of the lacunar walls appeared to be thinner and more closely packed than thos between the lacunae. The fine collagen fibrils associated with the lacunar walls were frequently observed to pass through a large lacunar space, resulting in the formation of two or more compartments, each of which was presumably filled with a chondrocyte in the living cartilage. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 Fig. 13 Fig. 14 PMID:870478

  9. Nanointerfacial strength between non-collagenous protein and collagen fibrils in antler bone

    PubMed Central

    Hang, Fei; Gupta, Himadri S.; Barber, Asa H.

    2014-01-01

    Antler bone displays considerable toughness through the use of a complex nanofibrous structure of mineralized collagen fibrils (MCFs) bound together by non-collagenous proteins (NCPs). While the NCP regions represent a small volume fraction relative to the MCFs, significant surface area is evolved upon failure of the nanointerfaces formed at NCP–collagen fibril boundaries. The mechanical properties of nanointerfaces between the MCFs are investigated directly in this work using an in situ atomic force microscopy technique to pull out individual fibrils from the NCP. Results show that the NCP–fibril interfaces in antler bone are weak, which highlights the propensity for interface failure at the nanoscale in antler bone and extensive fibril pullout observed at antler fracture surfaces. The adhesion between fibrils and NCP is additionally suggested as being rate dependent, with increasing interfacial strength and fracture energy observed when pullout velocity decreases. PMID:24352676

  10. Tension tests on mammalian collagen fibrils.

    PubMed

    Liu, Yehe; Ballarini, Roberto; Eppell, Steven J

    2016-02-01

    A brief overview of isolated collagen fibril mechanics testing is followed by presentation of the first results testing fibrils isolated from load-bearing mammalian tendons using a microelectromechanical systems platform. The in vitro modulus (326 ± 112 MPa) and fracture stress (71 ± 23 MPa) are shown to be lower than previously measured on fibrils extracted from sea cucumber dermis and tested with the same technique. Scanning electron microscope images show the fibrils can fail with a mechanism that involves circumferential rupture, whereas the core of the fibril stays at least partially intact. PMID:26855757

  11. Fibril-forming collagens in lamprey.

    PubMed

    Kelly, J; Tanaka, S; Hardt, T; Eikenberry, E F; Brodsky, B

    1988-01-15

    Five types of collagen with triple-helical regions approximately 300 nm in length were found in lamprey tissues which show characteristic D-periodic collagen fibrils. These collagens are members of the fibril forming family of this primitive vertebrate. Lamprey collagens were characterized with respect to solubility, mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, carboxylmethyl-cellulose chromatography, peptide digestion patterns, composition, susceptibility to vertebrate collagenase, thermal stability, and segment long spacing-banding pattern. Comparison with fibril-forming collagens in higher vertebrates (types I, II, III, V, and XI) identified three lamprey collagens as types II, V, and XI. Both lamprey dermis and major body wall collagens had properties similar to type I but not the typical heterotrimer composition. Dermis molecules had only alpha 1(I)-like chains, while body wall molecules had alpha 2(I)-like chains combined with chains resembling lamprey type II. Neither collagen exhibited the interchain disulfide linkages or solubility properties of type III. The conservation of fibril organization in type II/type XI tissues in contrast to the major developments in type I and type III tissues after the divergence of lamprey and higher vertebrates is consistent with these results. The presence of type II and type I-like molecules as major collagens and types V and XI as minor collagens in the lamprey, and the differential susceptibility of these molecules to vertebrate collagenase is analogous to the findings in higher vertebrates. PMID:3335531

  12. Collagen fibrils as skeletal frame in monkey trabecular meshwork.

    PubMed

    Nishida, S; Mizutani, S

    1994-01-01

    In an attempt to identify the characteristic skeletal frame of the trabecular meshwork, the three-dimensional architecture of collagen fibrillar bundles (collagen bundles) was studied by applying the NaOH cell-maceration method to the anterior segment of cynomolgus monkey eyes. Collagen bundles in the trabecular meshwork were found to be continuous with thinner collagen lamellae in the peripheral cornea and with the collagen fibril plexus in the ciliary body. The collagen core in the uveal cord was columnar in shape and was arranged parallel to the long axis. Collagen bundles were arranged as the layered network forming the fundamental framework of the trabecular meshwork. Most collagen bundles of the corneoscleral sheet were arranged circularly, parallel to the circumference of the limbus, and numerous slender bundles were observed side by side, forming the flat and membranous configuration of the sheet. The endothelial meshwork consisted of a network of fine and sparse collagen fibrils forming extracellular spaces and intercommunicating openings. The inner wall of the canal of Schlemm was seen as a netlike surface of sparse collagen fibrils with variously sized circular openings. The openings suggested the possibility that development of giant vacuoles corresponds to the circular openings under the inner endothelial cell lining. PMID:7933694

  13. Study of Native Type I Collagen Fibrils

    NASA Astrophysics Data System (ADS)

    Heim, August

    2006-03-01

    Presented in this work is direct imaging and force microscopy of native, intact type I collagen fibrils extracted from the sea cucumber Cucumaria frondosa dermis with affiliated proteoglycan molecules. The prototypical collagen fibril structure is well conserved through higher mammalian species and presents a model for study of the mechanical properties of the primary individual components of the dermis and skeletal ligature. Common practice is to use reconstituted fibrils which lack the precise conformal structure and affiliated proteoglycans. We have performed force microscopy to probe the mechanical properties of native fibrils and extract the elastic modulus under natural conditions. This knowledge is combined transmission and atomic force imaging, in conjunction with applied computation models, to demonstrate an inherent semitubular structure of these fibrils.

  14. Nature designs tough collagen: Explaining the nanostructure of collagen fibrils

    PubMed Central

    Buehler, Markus J.

    2006-01-01

    Collagen is a protein material with superior mechanical properties. It consists of collagen fibrils composed of a staggered array of ultra-long tropocollagen (TC) molecules. Theoretical and molecular modeling suggests that this natural design of collagen fibrils maximizes the strength and provides large energy dissipation during deformation, thus creating a tough and robust material. We find that the mechanics of collagen fibrils can be understood quantitatively in terms of two critical molecular length scales χS and χR that characterize when (i) deformation changes from homogeneous intermolecular shear to propagation of slip pulses and when (ii) covalent bonds within TC molecules begin to fracture, leading to brittle-like failure. The ratio χS/χR indicates which mechanism dominates deformation. Our modeling rigorously links the chemical properties of individual TC molecules to the macroscopic mechanical response of fibrils. The results help to explain why collagen fibers found in nature consist of TC molecules with lengths in the proximity of 300 nm and advance the understanding how collagen diseases that change intermolecular adhesion properties influence mechanical properties. PMID:16895989

  15. Mineralization of collagen may occur on fibril surfaces: evidence from conventional and high-voltage electron microscopy and three-dimensional imaging

    NASA Technical Reports Server (NTRS)

    Landis, W. J.; Hodgens, K. J.; Song, M. J.; Arena, J.; Kiyonaga, S.; Marko, M.; Owen, C.; McEwen, B. F.

    1996-01-01

    The interaction between collagen and mineral crystals in the normally calcifying leg tendons from the domestic turkey, Meleagris gallopavo, has been investigated at an ultrastructural level with conventional and high-voltage electron microscopy, computed tomography, and three-dimensional image reconstruction methods. Specimens treated by either aqueous or anhydrous techniques and resin-embedded were appropriately sectioned and regions of early tendon mineralization were photographed. On the basis of individual photomicrographs, stereoscopic pairs of images, and tomographic three-dimensional image reconstructions, platelet-shaped crystals may be demonstrated for the first time in association with the surface of collagen fibrils. Mineral is also observed in closely parallel arrays within collagen hole and overlap zones. The mineral deposition at these spatially distinct locations in the tendon provides insight into possible means by which calcification is mediated by collagen as a fundamental event in skeletal and dental formation among vertebrates.

  16. Viscoelastic Properties of Isolated Collagen Fibrils

    PubMed Central

    Shen, Zhilei Liu; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J.

    2011-01-01

    Understanding the viscoelastic behavior of collagenous tissues with complex hierarchical structures requires knowledge of the properties at each structural level. Whole tissues have been studied extensively, but less is known about the mechanical behavior at the submicron, fibrillar level. Using a microelectromechanical systems platform, in vitro coupled creep and stress relaxation tests were performed on collagen fibrils isolated from the sea cucumber dermis. Stress-strain-time data indicate that isolated fibrils exhibit viscoelastic behavior that could be fitted using the Maxwell-Weichert model. The fibrils showed an elastic modulus of 123 ± 46 MPa. The time-dependent behavior was well fit using the two-time-constant Maxwell-Weichert model with a fast time response of 7 ± 2 s and a slow time response of 102 ± 5 s. The fibrillar relaxation time was smaller than literature values for tissue-level relaxation time, suggesting that tissue relaxation is dominated by noncollagenous components (e.g., proteoglycans). Each specimen was tested three times, and the only statistically significant difference found was that the elastic modulus is larger in the first test than in the subsequent two tests, indicating that viscous properties of collagen fibrils are not sensitive to the history of previous tests. PMID:21689535

  17. Elastic model for crimped collagen fibrils

    NASA Technical Reports Server (NTRS)

    Freed, Alan D.; Doehring, Todd C.

    2005-01-01

    A physiologic constitutive expression is presented in algorithmic format for the nonlinear elastic response of wavy collagen fibrils found in soft connective tissues. The model is based on the observation that crimped fibrils in a fascicle have a three-dimensional structure at the micron scale that we approximate as a helical spring. The symmetry of this wave form allows the force/displacement relationship derived from Castigliano's theorem to be solved in closed form: all integrals become analytic. Model predictions are in good agreement with experimental observations for mitral-valve chordae tendinece.

  18. Elastic Response of Crimped Collagen Fibrils

    NASA Technical Reports Server (NTRS)

    Freed, Alan D.; Doehring, Todd C.

    2005-01-01

    A physiologic constitutive expression is presented in algorithmic format for the elastic response of wavy collagen fibrils found in soft connective tissues. The model is based on the observation that crimped fibrils have a three-dimensional structure at the micrometer scale that we approximate as a helical spring. The symmetry of this waveform allows the force/displacement relationship derived from Castigliano's theorem to be solved in closed form. Model predictions are in good agreement with experimental observations for mitral-valve chordae tendineae

  19. Collagen fibril biosynthesis in tendon: a review and recent insights.

    PubMed

    Canty, E G; Kadler, K E

    2002-12-01

    The development and evolution of multicellular animals relies on the ability of certain cell types to synthesise an extracellular matrix (ECM) comprising very long collagen fibrils that are arranged in very ordered 3-dimensional scaffolds. Tendon is a good example of a highly ordered ECM, in which tens of millions of collagen fibrils, each hundreds of microns long, are synthesised parallel to the tendon long axis. This review highlights recent discoveries showing that the assembly of collagen fibrils in tendon is hierarchical, and involves the formation of fairly short "collagen early fibrils" that are the fusion precursors of the very long fibrils that occur in mature tendon. PMID:12485687

  20. Structural investigations on native collagen type I fibrils using AFM

    SciTech Connect

    Strasser, Stefan; Zink, Albert; Janko, Marek; Heckl, Wolfgang M.; Thalhammer, Stefan . E-mail: stefan.thalhammer@gsf.de

    2007-03-02

    This study was carried out to determine the elastic properties of single collagen type I fibrils with the use of atomic force microscopy (AFM). Native collagen fibrils were formed by self-assembly in vitro characterized with the AFM. To confirm the inner assembly of the collagen fibrils, the AFM was used as a microdissection tool. Native collagen type I fibrils were dissected and the inner core uncovered. To determine the elastic properties of collagen fibrils the tip of the AFM was used as a nanoindentor by recording force-displacement curves. Measurements were done on the outer shell and in the core of the fibril. The structural investigations revealed the banding of the shell also in the core of native collagen fibrils. Nanoindentation experiments showed the same Young's modulus on the shell as well as in the core of the investigated native collagen fibrils. In addition, the measurements indicate a higher adhesion in the core of the collagen fibrils compared to the shell.

  1. Enzyme-etching technique to fabricate micropatterns of aligned collagen fibrils

    PubMed Central

    Liu, Honghai; Chen, Ruikai; Yang, Huaxiao; Qin, Wan; Borg, Thomas K.; Dean, Delphine; Xu, Meifeng; Gao, Bruce Z.

    2014-01-01

    A technique to tailor-make pre-coated, pre-aligned bovine collagen fibrils, derived from neonatal cardiomyocytes, on the surface of a glass slide into a designated pattern is reported. The unwanted collagen-coated area was erased by a collagenase solution and the tailored area was retained by attaching a microfabricated polydimethylsiloxane stamp directly to the collagen-coated surface. Using this technique, collagen patterns with designated orientations and with clear pattern boundaries and defined shapes were fabricated. PMID:24562408

  2. An equilibrium double-twist model for the radial structure of collagen fibrils.

    PubMed

    Brown, Aidan I; Kreplak, Laurent; Rutenberg, Andrew D

    2014-11-14

    Mammalian tissues contain networks and ordered arrays of collagen fibrils originating from the periodic self-assembly of helical 300 nm long tropocollagen complexes. The fibril radius is typically between 25 to 250 nm, and tropocollagen at the surface appears to exhibit a characteristic twist-angle with respect to the fibril axis. Similar fibril radii and twist-angles at the surface are observed in vitro, suggesting that these features are controlled by a similar self-assembly process. In this work, we propose a physical mechanism of equilibrium radius control for collagen fibrils based on a radially varying double-twist alignment of tropocollagen within a collagen fibril. The free-energy of alignment is similar to that of liquid crystalline blue phases, and we employ an analytic Euler-Lagrange and numerical free energy minimization to determine the twist-angle between the molecular axis and the fibril axis along the radial direction. Competition between the different elastic energy components, together with a surface energy, determines the equilibrium radius and twist-angle at the fibril surface. A simplified model with a twist-angle that is linear with radius is a reasonable approximation in some parameter regimes, and explains a power-law dependence of radius and twist-angle at the surface as parameters are varied. Fibril radius and twist-angle at the surface corresponding to an equilibrium free-energy minimum are consistent with existing experimental measurements of collagen fibrils. Remarkably, in the experimental regime, all of our model parameters are important for controlling equilibrium structural parameters of collagen fibrils. PMID:25238208

  3. Stress-strain experiments on individual collagen fibrils.

    PubMed

    Shen, Zhilei L; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J

    2008-10-01

    Collagen, a molecule consisting of three braided protein helices, is the primary building block of many biological tissues including bone, tendon, cartilage, and skin. Staggered arrays of collagen molecules form fibrils, which arrange into higher-ordered structures such as fibers and fascicles. Because collagen plays a crucial role in determining the mechanical properties of these tissues, significant theoretical research is directed toward developing models of the stiffness, strength, and toughness of collagen molecules and fibrils. Experimental data to guide the development of these models, however, are sparse and limited to small strain response. Using a microelectromechanical systems platform to test partially hydrated collagen fibrils under uniaxial tension, we obtained quantitative, reproducible mechanical measurements of the stress-strain curve of type I collagen fibrils, with diameters ranging from 150-470 nm. The fibrils showed a small strain (epsilon < 0.09) modulus of 0.86 +/- 0.45 GPa. Fibrils tested to strains as high as 100% demonstrated strain softening (sigma(yield) = 0.22 +/- 0.14 GPa; epsilon(yield) = 0.21 +/- 0.13) and strain hardening, time-dependent recoverable residual strain, dehydration-induced embrittlement, and susceptibility to cyclic fatigue. The results suggest that the stress-strain behavior of collagen fibrils is dictated by global characteristic dimensions as well as internal structure. PMID:18641067

  4. Stress-Strain Experiments on Individual Collagen Fibrils

    PubMed Central

    Shen, Zhilei L.; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J.

    2008-01-01

    Collagen, a molecule consisting of three braided protein helices, is the primary building block of many biological tissues including bone, tendon, cartilage, and skin. Staggered arrays of collagen molecules form fibrils, which arrange into higher-ordered structures such as fibers and fascicles. Because collagen plays a crucial role in determining the mechanical properties of these tissues, significant theoretical research is directed toward developing models of the stiffness, strength, and toughness of collagen molecules and fibrils. Experimental data to guide the development of these models, however, are sparse and limited to small strain response. Using a microelectromechanical systems platform to test partially hydrated collagen fibrils under uniaxial tension, we obtained quantitative, reproducible mechanical measurements of the stress-strain curve of type I collagen fibrils, with diameters ranging from 150–470 nm. The fibrils showed a small strain (ɛ < 0.09) modulus of 0.86 ± 0.45 GPa. Fibrils tested to strains as high as 100% demonstrated strain softening (σyield = 0.22 ± 0.14 GPa; ɛyield = 0.21 ± 0.13) and strain hardening, time-dependent recoverable residual strain, dehydration-induced embrittlement, and susceptibility to cyclic fatigue. The results suggest that the stress-strain behavior of collagen fibrils is dictated by global characteristic dimensions as well as internal structure. PMID:18641067

  5. Micromechanical analysis of native and cross-linked collagen type I fibrils supports the existence of microfibrils.

    PubMed

    Yang, L; van der Werf, K O; Dijkstra, P J; Feijen, J; Bennink, M L

    2012-02-01

    The mechanical properties of individual collagen fibrils of approximately 200 nm in diameter were determined using a slightly adapted AFM system. Single collagen fibrils immersed in PBS buffer were attached between an AFM cantilever and a glass surface to perform tensile tests at different strain rates and stress relaxation measurements. The stress-strain behavior of collagen fibrils immersed in PBS buffer comprises a toe region up to a stress of 5 MPa, followed by the heel and linear region at higher stresses. Hysteresis and strain-rate dependent stress-strain behavior of collagen fibrils were observed, which suggest that single collagen fibrils have viscoelastic properties. The stress relaxation process of individual collagen fibrils could be best fitted using a two-term Prony series. Furthermore, the influence of different cross-linking agents on the mechanical properties of single collagen fibrils was investigated. Based on these results, we propose that sliding of microfibrils with respect to each other plays a role in the viscoelastic behavior of collagen fibrils in addition to the sliding of collagen molecules with respect to each other. Our finding provides a better insight into the relationship between the structure and mechanical properties of collagen and the micro-mechanical behavior of tissues. PMID:22301184

  6. Molecular packing in bone collagen fibrils prior to mineralization

    NASA Astrophysics Data System (ADS)

    Hsiao, Benjamin; Zhou, Hong-Wen; Burger, Christian; Chu, Benjamin; Glimcher, Melvin J.

    2012-02-01

    The three-dimensional packing of collagen molecules in bone collagen fibrils has been largely unknown because even in moderately mineralized bone tissues, the organic matrix structure is severely perturbed by the deposition of mineral crystals. During the past decades, the structure of tendon collagen (e.g. rat tail) --- a tissue that cannot mineralize in vivo, has been assumed to be representative for bone collagen fibrils. Small-angle X-ray diffraction analysis of the native, uncalcified intramuscular fish bone has revealed a new molecular packing scheme, significantly different from the quasi-hexagonal arrangement often found in tendons. The deduced structure in bone collagen fibrils indicates the presence of spatially discrete microfibrils, and an arrangement of intrafibrillar space to form ``channels'', which could accommodate crystals with dimensions typically found in bone apatite.

  7. Electron microscopic stereological study of collagen fibrils in bovine articular cartilage: volume and surface densities are best obtained indirectly (from length densities and diameters) using isotropic uniform random sampling

    PubMed Central

    LÅNGSJÖ, TEEMU K.; HYTTINEN, MIKA; PELTTARI, ALPO; KIRALY, KARI; AROKOSKI, JARI; HELMINEN, HEIKKI J.

    1999-01-01

    Results obtained by the indirect zonal isotropic uniform random (IUR) estimation were compared with those obtained by the direct point and interception counting methods on vertical (VS) or IUR sections in a stereological study of bovine articular cartilage collagen fibrils at the ultrastructural level. Besides comparisons between the direct and indirect estimations (direct IUR vs indirect IUR estimations) and between different sampling methods (VS vs IUR sampling), simultaneous comparison of the 2 issues took place (direct VS vs indirect IUR estimation). Using the direct VS method, articular cartilage superficial zone collagen volume fraction (Vv 41%) was 67% and fibril surface density (Sv 0.030 nm2/nm3) 15% higher (P<0.05) than values obtained by the indirect IUR method (Vv 25% and Sv 0.026 nm2/nm3). The same was observed when the direct IUR method was used: collagen volume fraction (Vv 40%) was 63% and fibril surface density (Sv 0.032 nm2/nm3) 21% higher (P<0.05) than those obtained by the indirect IUR technique. Similarly, in the deep zone of articular cartilage direct VS and direct IUR methods gave 50 and 55% higher (P<0.05) collagen fibril volume fractions (Vv 43 and 44% vs 29%) and the direct IUR method 25% higher (P<0.05) fibril surface density values (Sv 0.025 vs 0.020 nm2/nm3) than the indirect IUR estimation. On theoretical grounds, scrutiny calculations, as well as earlier reports, it is concluded that the direct VS and direct IUR methods systematically overestimated the Vv and Sv of collagen fibrils. This bias was due to the overprojection which derives from the high section thickness in relation to collagen fibril diameter. On the other hand, factors that during estimation tend to underestimate Vv and Sv, such as profile overlapping and truncation (‘fuzzy’ profiles), seemed to cause less bias. As length density (Lv) and collagen fibril diameter are minimally biased by the high relative section thickness, the indirect IUR method, based on

  8. Stabilization and Anomalous Hydration of Collagen Fibril under Heating

    PubMed Central

    Gevorkian, Sasun G.; Allahverdyan, Armen E.; Gevorgyan, David S.; Simonian, Aleksandr L.; Hu, Chin-Kun

    2013-01-01

    Background Type I collagen is the most common protein among higher vertebrates. It forms the basis of fibrous connective tissues (tendon, chord, skin, bones) and ensures mechanical stability and strength of these tissues. It is known, however, that separate triple-helical collagen macromolecules are unstable at physiological temperatures. We want to understand the mechanism of collagen stability at the intermolecular level. To this end, we study the collagen fibril, an intermediate level in the collagen hierarchy between triple-helical macromolecule and tendon. Methodology/Principal Finding When heating a native fibril sample, its Young’s modulus decreases in temperature range 20–58°C due to partial denaturation of triple-helices, but it is approximately constant at 58–75°C, because of stabilization by inter-molecular interactions. The stabilization temperature range 58–75°C has two further important features: here the fibril absorbs water under heating and the internal friction displays a peak. We relate these experimental findings to restructuring of collagen triple-helices in fibril. A theoretical description of the experimental results is provided via a generalization of the standard Zimm-Bragg model for the helix-coil transition. It takes into account intermolecular interactions of collagen triple-helices in fibril and describes water adsorption via the Langmuir mechanism. Conclusion/Significance We uncovered an inter-molecular mechanism that stabilizes the fibril made of unstable collagen macromolecules. This mechanism can be relevant for explaining stability of collagen. PMID:24244320

  9. Extracellular compartments in matrix morphogenesis: collagen fibril, bundle, and lamellar formation by corneal fibroblasts.

    PubMed

    Birk, D E; Trelstad, R L

    1984-12-01

    The regulation of collagen fibril, bundle, and lamella formation by the corneal fibroblasts, as well as the organization of these elements into an orthogonal stroma, was studied by transmission electron microscopy and high voltage electron microscopy. Transmission and high voltage electron microscopy of chick embryo corneas each demonstrated a series of unique extracellular compartments. Collagen fibrillogenesis occurred within small surface recesses. These small recesses usually contained between 5 and 12 collagen fibrils with typically mature diameters and constant intrafibrillar spacing. The lateral fusion of the recesses resulted in larger recesses and consequent formation of prominent cell surface foldings. Within these surface foldings, bundles that contained 50-100 collagen fibrils were formed. The surface foldings continued to fuse and the cell surface retracted, forming large surface-associated compartments in which bundles coalesced to form lamellae. High voltage electron microscopy of 0.5 micron sections cut parallel to the corneal surface revealed that the corneal fibroblasts and their processes had two major axes at approximately right angles to one another. The surface compartments involved in the production of the corneal stroma were aligned along the fibroblast axes and the orthogonality of the cell was in register with that of the extracellular matrix. In this manner, corneal fibroblasts formed collagen fibrils, bundles, and lamellae within a controlled environment and thereby determined the architecture of the corneal stroma by the configuration of the cell and its associated compartments. PMID:6542105

  10. Measurement of the Mechanical Properties of Intact Collagen Fibrils

    NASA Astrophysics Data System (ADS)

    Mercedes, H.; Heim, A.; Matthews, W. G.; Koob, T.

    2006-03-01

    Motivated by the genetic disorder Ehlers-Danlos syndrome (EDS), in which proper collagen synthesis is interrupted, we are investigating the structural and mechanical properties of collagen fibrils. The fibrous glycoprotein collagen is the most abundant protein found in the human body and plays a key role in the extracellular matrix of the connective tissue, the properties of which are altered in EDS. We have selected as our model system the collagen fibrils of the sea cucumber dermis, a naturally mutable tissue. This system allows us to work with native fibrils which have their proteoglycan complement intact, something that is not possible with reconstituted mammalian collagen fibrils. Using atomic force microscopy, we measure, as a function of the concentration of divalent cations, the fibril diameter, its response to force loading, and the changes in its rigidity. Through these experiments, we will shed light on the mechanisms which control the properties of the sea cucumber dermis and hope to help explain the altered connective tissue extracellular matrix properties associated with EDS.

  11. Collagen XII: Protecting bone and muscle integrity by organizing collagen fibrils.

    PubMed

    Chiquet, Matthias; Birk, David E; Bönnemann, Carsten G; Koch, Manuel

    2014-08-01

    Collagen XII, largest member of the fibril-associated collagens with interrupted triple helix (FACIT) family, assembles from three identical α-chains encoded by the COL12A1 gene. The molecule consists of three threadlike N-terminal noncollagenous NC3 domains, joined by disulfide bonds and a short interrupted collagen triple helix toward the C-terminus. Splice variants differ considerably in size and properties: "small" collagen XIIB (220 kDa subunit) is similar to collagen XIV, whereas collagen XIIA (350 kDa) has a much larger NC3 domain carrying glycosaminoglycan chains. Collagen XII binds to collagen I-containing fibrils via its collagenous domain, whereas its large noncollagenous arms interact with other matrix proteins such as tenascin-X. In dense connective tissues and bone, collagen XII is thought to regulate organization and mechanical properties of collagen fibril bundles. Accordingly, recent findings show that collagen XII mutations cause Ehlers-Danlos/myopathy overlap syndrome associated with skeletal abnormalities and muscle weakness in mice and humans. PMID:24801612

  12. Molecules in Focus: Collagen XII: Protecting bone and muscle integrity by organizing collagen fibrils

    PubMed Central

    Chiquet, Matthias; Birk, David E.; Bönnemann, Carsten G.; Koch, Manuel

    2014-01-01

    Collagen XII, largest member of the fibril-associated collagens with interrupted triple helix (FACIT) family, assembles from three identical α-chains encoded by the COL12A1 gene. The molecule consists of three threadlike N-terminal noncollagenous NC3 domains, joined by disulfide bonds and a short interrupted collagen triple helix towards the C-terminus. Splice variants differ considerably in size and properties: "small" collagen XIIB (220 kDa subunit) is similar to collagen XIV, whereas collagen XIIA (350 kDa) has a much larger NC3 domain carrying glycosaminoglycan chains. Collagen XII binds to collagen I-containing fibrils via its collagenous domain, whereas its large noncollagenous arms interact with other matrix proteins such as tenascin-X. In dense connective tissues and bone, collagen XII is thought to regulate organization and mechanical properties of collagen fibril bundles. Accordingly, recent findings show that collagen XII mutations cause Ehlers-Danlos/myopathy overlap syndrome associated with skeletal abnormalities and muscle weakness in mice and humans. PMID:24801612

  13. Interpreting Second-Harmonic Generation Images of Collagen I Fibrils

    PubMed Central

    Williams, Rebecca M.; Zipfel, Warren R.; Webb, Watt W.

    2005-01-01

    Fibrillar collagen, being highly noncentrosymmetric, possesses a tremendous nonlinear susceptibility. As a result, second-harmonic generation (SHG) microscopy of collagen produces extremely bright and robust signals, providing an invaluable tool for imaging tissue structure with submicron resolution. Here we discuss fundamental principles governing SHG phase matching with the tightly focusing optics used in microscopy. Their application to collagen imaging yields several biophysical features characteristic of native collagen structure: SHG radiates from the shell of a collagen fibril, rather than from its bulk. This SHG shell may correspond to the supporting element of the fibril. Physiologically relevant changes in solution ionic strength alter the ratio of forward-to-backward propagating SHG, implying a resulting change in the SHG shell thickness. Fibrillogenesis can be resolved in immature tissue by directly imaging backward-propagating SHG. Such findings are crucial to the design and development of forthcoming diagnostic and research tools. PMID:15533922

  14. Characteristics and Young's Modulus of Collagen Fibrils from Expanded Skin Using Anisotropic Controlled Rate Self-Inflating Tissue Expander.

    PubMed

    Manssor, Nur Aini S; Radzi, Zamri; Yahya, Noor Azlin; Mohamad Yusof, Loqman; Hariri, Firdaus; Khairuddin, Nurul Hayah; Abu Kasim, Noor Hayaty; Czernuszka, Jan T

    2016-01-01

    Mechanical properties of expanded skin tissue are different from normal skin, which is dependent mainly on the structural and functional integrity of dermal collagen fibrils. In the present study, mechanical properties and surface topography of both expanded and nonexpanded skin collagen fibrils were evaluated. Anisotropic controlled rate self-inflating tissue expanders were placed beneath the skin of sheep's forelimbs. The tissue expanders gradually increased in height and reached equilibrium in 2 weeks. They were left in situ for another 2 weeks before explantation. Expanded and normal skin samples were surgically harvested from the sheep (n = 5). Young's modulus and surface topography of collagen fibrils were measured using an atomic force microscope. A surface topographic scan showed organized hierarchical structural levels: collagen molecules, fibrils and fibers. No significant difference was detected for the D-banding pattern: 63.5 ± 2.6 nm (normal skin) and 63.7 ± 2.7 nm (expanded skin). Fibrils from expanded tissues consisted of loosely packed collagen fibrils and the width of the fibrils was significantly narrower compared to those from normal skin: 153.9 ± 25.3 and 106.7 ± 28.5 nm, respectively. Young's modulus of the collagen fibrils in the expanded and normal skin was not statistically significant: 46.5 ± 19.4 and 35.2 ± 27.0 MPa, respectively. In conclusion, the anisotropic controlled rate self-inflating tissue expander produced a loosely packed collagen network and the fibrils exhibited similar D-banding characteristics as the control group in a sheep model. However, the fibrils from the expanded skin were significantly narrower. The stiffness of the fibrils from the expanded skin was higher but it was not statistically different. PMID:26836267

  15. Rapid oriented fibril formation of fish scale collagen facilitates early osteoblastic differentiation of human mesenchymal stem cells.

    PubMed

    Matsumoto, Rena; Uemura, Toshimasa; Xu, Zhefeng; Yamaguchi, Isamu; Ikoma, Toshiyuki; Tanaka, Junzo

    2015-08-01

    We studied the effect of fibril formation of fish scale collagen on the osteoblastic differentiation of human mesenchymal stem cells (hMSCs). We found that hMSCs adhered easily to tilapia scale collagen, which remarkably accelerated the early stage of osteoblastic differentiation in hMSCs during in vitro cell culture. Osteoblastic markers such as ALP activity, osteopontin, and bone morphogenetic protein 2 were markedly upregulated when the hMSCs were cultured on a tilapia collagen surface, especially in the early osteoblastic differentiation stage. We hypothesized that this phenomenon occurs due to specific fibril formation of tilapia collagen. Thus, we examined the time course of collagen fibril formation using high-speed atomic force microscopy. Moreover, to elucidate the effect of the orientation of fibril formation on the differentiation of hMSCs, we measured ALP activity of hMSCs cultured on two types of tilapia scale collagen membranes with different degrees of fibril formation. The ALP activity in hMSCs cultured on a fibrous collagen membrane was significantly higher than on a non-fibrous collagen membrane even before adding osteoblastic differentiation medium. These results showed that the degree of the fibril formation of tilapia collagen was essential for the osteoblastic differentiation of hMSCs. PMID:25546439

  16. Supramolecular assembly of collagen fibrils into collagen fiber in fish scales of red seabream, Pagrus major.

    PubMed

    Youn, Hwa Shik; Shin, Tae Joo

    2009-11-01

    Supramolecular assembly of collagen fibrils into collagen fiber and its distribution in fish scales of red seabream, Pagrus major, were investigated. By virtue of Zernike phase-contrast hard X-ray microscopy, it has been firstly observed that collagen fiber consists of helical substructures of collagen fibrils wrapped with incrustation. As it close to the scalar focus (that is, with aging), loosened- and deteriorated-helical assemblies started to be observed with loosing wrapping incrustation, indicative of the distortion of the basic helical assembly. Various distributions and packing arrangements of collagen fibers were observed dependent on subdivisions of fish scale. Freshly growing edge region of fish scale, embedded into fish skin, showed rarely patched and one directionally arranged collagen fibers, in which specifically triple helical assemblies of collagen fibrils were found. On the contrary, relatively aged region of the rostral field close to the scalar focus displayed randomly directed and densely packed collagen fibers, in which loosened- and deteriorated-helical assemblies of collagen fibrils were mostly found. Our results have demonstrated that hard X-ray microscope can be a powerful tool to study in situ internal structure of biological specimens in an atmospheric pressure. PMID:19666125

  17. Viscoelastic behavior of discrete human collagen fibrils.

    PubMed

    Svensson, René B; Hassenkam, Tue; Hansen, Philip; Peter Magnusson, S

    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 fibrils. Fibrils were obtained from intact human fascicles, without any pre-treatment besides frozen storage. In the dry state a single isolated fibril was anchored to a substrate using epoxy glue, and the end of the fibril was glued on to an AFM cantilever for tensile testing. In phosphate buffered 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 on the strain. The slope of the viscous response showed a strain rate dependence corresponding to a power function of powers 0.242 and 0.168 for the two patellar tendon fibrils, respectively. In conclusion, the present work provides direct evidence of viscoelastic behavior at the single fibril level, which has not been previously measured. PMID:19878908

  18. Mechanical Properties of Mineralized Collagen Fibrils As Influenced By Demineralization

    SciTech Connect

    Balooch, M.; Habelitz, S.; Kinney, J.H.; Marshall, S.J.; Marshall, G.W.

    2009-05-11

    Dentin and bone derive their mechanical properties from a complex arrangement of collagen type-I fibrils reinforced with nanocrystalline apatite mineral in extra- and intrafibrillar compartments. While mechanical properties have been determined for the bulk of the mineralized tissue, information on the mechanics of the individual fibril is limited. Here, atomic force microscopy was used on individual collagen fibrils to study structural and mechanical changes during acid etching. The characteristic 67 nm periodicity of gap zones was not observed on the mineralized fibril, but became apparent and increasingly pronounced with continuous demineralization. AFM-nanoindentation showed a decrease in modulus from 1.5 GPa to 50 MPa during acid etching of individual collagen fibrils and revealed that the modulus profile followed the axial periodicity. The nanomechanical data, Raman spectroscopy and SAXS support the hypothesis that intrafibrillar mineral etches at a substantially slower rate than the extrafibrillar mineral. These findings are relevant for understanding the biomechanics and design principles of calcified tissues derived from collagen matrices.

  19. Nanostructural and nanomechanical responses of collagen fibrils in the collagenase-induced Achilles tendinitis rat model.

    PubMed

    Lee, Gi-Ja; Choi, Samjin; Chon, Jinmann; Yoo, Seung Don; Kim, Hee-Sang; Park, Hun-Kuk; Chung, Joo Ho

    2013-11-01

    Achilles tendons are vulnerable to acute or chronic injuries that lead to inflammation. We investigated nanostructural and nanomechanical changes in collagen fibrils from rat Achilles tendons over a period of 9 weeks after injury using atomic force microscopy (AFM). To evaluate the nanostructural changes in Achilles tendons, we measured the diameter and D-banding of collagen fibrils by AFM. And the adhesion forces, which were related to cross-linking of collagen, were calculated from the retraction process of a force-distance curve. We successfully observed the time course of changes in collagen fibrils during healing using AFM. The diameters and D-banding in healed tendons were similar to those of uninjured tendons at 9 weeks after injury, but the adhesion forces remained different from those of uninjured tendons. Our AFM results depicted the minute changes in Achilles tendon surface by natural healing quite well, even drawbacks to naturally healed tendon. Understanding changes in collagen cross-linking and structure while healing will lead to better understanding of healing mechanisms and subsequent improvements in treatment. And AFM can be applied as powerful tool to evaluate structural and property changes in collagen fibrils before and after injury and/or treatment in Achilles tendon. PMID:24245243

  20. Exploring a Role in Tanning for the Gap Region of the Collagen Fibril: Catechin-Collagen Interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Electron micrographs of stained collagen fibrils display a pattern of alternating light and dark bands perpendicular to the axis of the collagen fibril. Light bands correspond to regions of more dense lateral packing where adjacent collagen monomers overlap, and dark bands correspond to 'gap' regio...

  1. Techniques to assess bone ultrastructure organization: orientation and arrangement of mineralized collagen fibrils.

    PubMed

    Georgiadis, Marios; Müller, Ralph; Schneider, Philipp

    2016-06-01

    Bone's remarkable mechanical properties are a result of its hierarchical structure. The mineralized collagen fibrils, made up of collagen fibrils and crystal platelets, are bone's building blocks at an ultrastructural level. The organization of bone's ultrastructure with respect to the orientation and arrangement of mineralized collagen fibrils has been the matter of numerous studies based on a variety of imaging techniques in the past decades. These techniques either exploit physical principles, such as polarization, diffraction or scattering to examine bone ultrastructure orientation and arrangement, or directly image the fibrils at the sub-micrometre scale. They make use of diverse probes such as visible light, X-rays and electrons at different scales, from centimetres down to nanometres. They allow imaging of bone sections or surfaces in two dimensions or investigating bone tissue truly in three dimensions, in vivo or ex vivo, and sometimes in combination with in situ mechanical experiments. The purpose of this review is to summarize and discuss this broad range of imaging techniques and the different modalities of their use, in order to discuss their advantages and limitations for the assessment of bone ultrastructure organization with respect to the orientation and arrangement of mineralized collagen fibrils. PMID:27335222

  2. Techniques to assess bone ultrastructure organization: orientation and arrangement of mineralized collagen fibrils

    PubMed Central

    Georgiadis, Marios; Müller, Ralph; Schneider, Philipp

    2016-01-01

    Bone's remarkable mechanical properties are a result of its hierarchical structure. The mineralized collagen fibrils, made up of collagen fibrils and crystal platelets, are bone's building blocks at an ultrastructural level. The organization of bone's ultrastructure with respect to the orientation and arrangement of mineralized collagen fibrils has been the matter of numerous studies based on a variety of imaging techniques in the past decades. These techniques either exploit physical principles, such as polarization, diffraction or scattering to examine bone ultrastructure orientation and arrangement, or directly image the fibrils at the sub-micrometre scale. They make use of diverse probes such as visible light, X-rays and electrons at different scales, from centimetres down to nanometres. They allow imaging of bone sections or surfaces in two dimensions or investigating bone tissue truly in three dimensions, in vivo or ex vivo, and sometimes in combination with in situ mechanical experiments. The purpose of this review is to summarize and discuss this broad range of imaging techniques and the different modalities of their use, in order to discuss their advantages and limitations for the assessment of bone ultrastructure organization with respect to the orientation and arrangement of mineralized collagen fibrils. PMID:27335222

  3. In Vitro Fracture Testing of Submicron Diameter Collagen Fibril Specimens

    PubMed Central

    Shen, Zhilei Liu; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J.

    2010-01-01

    Mechanical testing of collagenous tissues at different length scales will provide improved understanding of the mechanical behavior of structures such as skin, tendon, and bone, and also guide the development of multiscale mechanical models. Using a microelectromechanical-systems (MEMS) platform, stress-strain response curves up to failure of type I collagen fibril specimens isolated from the dermis of sea cucumbers were obtained in vitro. A majority of the fibril specimens showed brittle fracture. Some displayed linear behavior up to failure, while others displayed some nonlinearity. The fibril specimens showed an elastic modulus of 470 ± 410 MPa, a fracture strength of 230 ± 160 MPa, and a fracture strain of 80% ± 44%. The fibril specimens displayed significantly lower elastic modulus in vitro than previously measured in air. Fracture strength/strain obtained in vitro and in air are both significantly larger than those obtained in vacuo, indicating that the difference arises from the lack of intrafibrillar water molecules produced by vacuum drying. Furthermore, fracture strength/strain of fibril specimens were different from those reported for collagenous tissues of higher hierarchical levels, indicating the importance of obtaining these properties at the fibrillar level for multiscale modeling. PMID:20858445

  4. In vitro fracture testing of submicron diameter collagen fibril specimens.

    PubMed

    Shen, Zhilei Liu; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J

    2010-09-22

    Mechanical testing of collagenous tissues at different length scales will provide improved understanding of the mechanical behavior of structures such as skin, tendon, and bone, and also guide the development of multiscale mechanical models. Using a microelectromechanical-systems (MEMS) platform, stress-strain response curves up to failure of type I collagen fibril specimens isolated from the dermis of sea cucumbers were obtained in vitro. A majority of the fibril specimens showed brittle fracture. Some displayed linear behavior up to failure, while others displayed some nonlinearity. The fibril specimens showed an elastic modulus of 470 ± 410 MPa, a fracture strength of 230 ± 160 MPa, and a fracture strain of 80% ± 44%. The fibril specimens displayed significantly lower elastic modulus in vitro than previously measured in air. Fracture strength/strain obtained in vitro and in air are both significantly larger than those obtained in vacuo, indicating that the difference arises from the lack of intrafibrillar water molecules produced by vacuum drying. Furthermore, fracture strength/strain of fibril specimens were different from those reported for collagenous tissues of higher hierarchical levels, indicating the importance of obtaining these properties at the fibrillar level for multiscale modeling. PMID:20858445

  5. Constitutive modeling of crimped collagen fibrils in soft tissues.

    PubMed

    Grytz, Rafael; Meschke, Günther

    2009-10-01

    A microstructurally oriented constitutive formulation for the hyperelastic response of crimped collagen fibrils existing in soft connective tissues is proposed. The model is based on observations that collagen fibrils embedded in a soft matrix crimp into a smooth three-dimensional pattern when unloaded. Following ideas presented by Beskos and Jenkins [Beskos, D., Jenkins, J., 1975. A mechanical model for mammalian tendon. ASME Journal of Applied Mechanics 42, 755-758] and Freed and Doehring [Freed, A., Doehring, T., 2005. Elastic model for crimped collagen fibrils. Journal of Biomechanical Engineering 127, 587-593] the collagen fibril crimp is approximated by a cylindrical helix to represent the constitutive behavior of the hierarchical organized substructure of biological tissues at the fibrillar level. The model is derived from the nonlinear axial force-stretch relationship of an extensible helical spring, including the full extension of the spring as a limit case. The geometrically nonlinear solution of the extensible helical spring is carried out by an iterative procedure. The model only requires one material parameter and two geometrical parameters to be determined from experiments. The ability of the proposed model to reproduce the biomechanical response of fibrous tissues is demonstrated for fascicles from rat tail tendons, for porcine cornea strips, and for bovine Achilles tendons. PMID:19627859

  6. Structure and formation of the twisted plywood pattern of collagen fibrils in rat lamellar bone.

    PubMed

    Yamamoto, Tsuneyuki; Hasegawa, Tomoka; Sasaki, Muneteru; Hongo, Hiromi; Tabata, Chihiro; Liu, Zhusheng; Li, Minqi; Amizuka, Norio

    2012-04-01

    This study was designed to elucidate details of the structure and formation process of the alternate lamellar pattern known to exist in lamellar bone. For this purpose, we examined basic internal lamellae in femurs of young rats by transmission and scanning electron microscopy, the latter employing two different macerations with NaOH at concentrations of 10 and 24%. Observations after the maceration with 10% NaOH showed that the regular and periodic rotation of collagen fibrils caused an alternation between two types of lamellae: one consisting of transversely and nearly transversely cut fibrils, and the other consisting of longitudinally and nearly longitudinally cut fibrils. This finding confirms the consistency of the twisted plywood model. The maceration method with 24% NaOH removed bone components other than cells, thus allowing for three-dimensional observations of osteoblast morphology. Osteoblasts extended finger-like processes paralleling the inner bone surface, and grouped in such a way that, within a group, the processes arranged in a similar direction. Transmission electron microscopy showed that newly deposited fibrils were arranged alongside these processes. For the formation of the alternating pattern, our findings suggest that: (1) osteoblasts control the collagen fibril arrangement through their finger-like process position; (2) osteoblasts behave similarly within a group; (3) osteoblasts move their processes synchronously and periodically to promote alternating different fibril orientation; and (4) this dynamic sequential deposition of fibrils results in the alternate lamellar (or twisted plywood) pattern. PMID:22362877

  7. Exploring a role in tanning for the gap region of the collagen fibril

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Three-dimensional structures of fibrillar collagens have been the subject of numerous studies for more than 60 years. Electron micrographs of stained collagen fibrils display a pattern of alternating light and dark bands perpendicular to the axis of the collagen fibril. Light bands correspond to r...

  8. Exploring a role in tanning for the gap region of the collagen fibril

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Three-dimensional structures of fibrillar collagens have been the subject of numerous studies for more than 60 years. Electron micrographs of stained collagen fibrils display a pattern of alternating light and dark bands perpendicular to the axis of the collagen fibril. Light bands correspond to reg...

  9. In vitro formation and thermal transition of novel hybrid fibrils from type I fish scale collagen and type I porcine collagen

    NASA Astrophysics Data System (ADS)

    Chen, Song; Ikoma, Toshiyuki; Ogawa, Nobuhiro; Migita, Satoshi; Kobayashi, Hisatoshi; Hanagata, Nobutaka

    2010-06-01

    Novel type I collagen hybrid fibrils were fabricated by neutralizing a mixture of type I fish scale collagen solution and type I porcine collagen solution with a phosphate buffer saline at 28 °C. Their structure was discussed in terms of the volume ratio of fish/porcine collagen solution. Scanning electron and atomic force micrographs showed that the diameter of collagen fibrils derived from the collagen mixture was larger than those derived from each collagen, and all resultant fibrils exhibited a typical D-periodic unit of ~67 nm, irrespective of volume ratio of both collagens. Differential scanning calorimetry revealed only one endothermic peak for the fibrils derived from collagen mixture or from each collagen solution, indicating that the resultant collagen fibrils were hybrids of type I fish scale collagen and type I porcine collagen.

  10. Large Deformation Mechanisms, Plasticity, and Failure of an Individual Collagen Fibril With Different Mineral Content.

    PubMed

    Depalle, Baptiste; Qin, Zhao; Shefelbine, Sandra J; Buehler, Markus J

    2016-02-01

    Mineralized collagen fibrils are composed of tropocollagen molecules and mineral crystals derived from hydroxyapatite to form a composite material that combines optimal properties of both constituents and exhibits incredible strength and toughness. Their complex hierarchical structure allows collagen fibrils to sustain large deformation without breaking. In this study, we report a mesoscale model of a single mineralized collagen fibril using a bottom-up approach. By conserving the three-dimensional structure and the entanglement of the molecules, we were able to construct finite-size fibril models that allowed us to explore the deformation mechanisms which govern their mechanical behavior under large deformation. We investigated the tensile behavior of a single collagen fibril with various intrafibrillar mineral content and found that a mineralized collagen fibril can present up to five different deformation mechanisms to dissipate energy. These mechanisms include molecular uncoiling, molecular stretching, mineral/collagen sliding, molecular slippage, and crystal dissociation. By multiplying its sources of energy dissipation and deformation mechanisms, a collagen fibril can reach impressive strength and toughness. Adding mineral into the collagen fibril can increase its strength up to 10 times and its toughness up to 35 times. Combining crosslinks with mineral makes the fibril stiffer but more brittle. We also found that a mineralized fibril reaches its maximum toughness to density and strength to density ratios for a mineral density of around 30%. This result, in good agreement with experimental observations, attests that bone tissue is optimized mechanically to remain lightweight but maintain strength and toughness. PMID:26866939

  11. Epitaxially guided assembly of collagen layers on mica surfaces.

    PubMed

    Leow, Wee Wen; Hwang, Wonmuk

    2011-09-01

    Ordered assembly of collagen molecules on flat substrates has potential for various applications and serves as a model system for studying the assembly process. While previous studies demonstrated self-assembly of collagen on muscovite mica into highly ordered layers, the mechanism by which different conditions affect the resulting morphology remains to be elucidated. Using atomic force microscopy, we follow the assembly of collagen on muscovite mica at a concentration lower than the critical fibrillogenesis concentration in bulk. Initially, individual collagen molecules adsorb to mica and subsequently nucleate into fibrils possessing the 67 nm D-periodic bands. Emergence of fibrils aligned in parallel despite large interfibril distances agrees with an alignment mechanism guided by the underlying mica. The epitaxial growth was further confirmed by the formation of novel triangular networks of collagen fibrils on phlogopite mica, whose surface lattice is known to have a hexagonal symmetry, whereas the more widely used muscovite does not. Comparing collagen assembly on the two types of mica at different potassium concentrations revealed that potassium binds to the negatively charged mica surface and neutralizes it, thereby reducing the binding affinity of collagen and enhancing surface diffusion. These results suggest that collagen assembly on mica follows the surface adsorption, diffusion, nucleation, and growth pathway, where the growth direction is determined at the nucleation step. Comparison with other molecules that assemble similarly on mica supports generality of the proposed assembly mechanism, the knowledge of which will be useful for controlling the resulting surface morphologies. PMID:21740026

  12. Candidate Cell and Matrix Interaction Domains on the Collagen Fibril, the Predominant Protein of Vertebrates

    SciTech Connect

    Sweeney, Shawn M.; Orgel, Joseph P.; Fertala, Andrzej; McAuliffe, Jon D.; Turner, Kevin R.; Di Lullo, Gloria A.; Chen, Steven; Antipova, Olga; Perumal, Shiamalee; Ala-Kokko, Leena; Forlinoi, Antonella; Cabral, Wayne A.; Barnes, Aileen M.; Marini, Joan C.; San Antonio, James D.

    2008-07-18

    Type I collagen, the predominant protein of vertebrates, polymerizes with type III and V collagens and non-collagenous molecules into large cable-like fibrils, yet how the fibril interacts with cells and other binding partners remains poorly understood. To help reveal insights into the collagen structure-function relationship, a data base was assembled including hundreds of type I collagen ligand binding sites and mutations on a two-dimensional model of the fibril. Visual examination of the distribution of functional sites, and statistical analysis of mutation distributions on the fibril suggest it is organized into two domains. The 'cell interaction domain' is proposed to regulate dynamic aspects of collagen biology, including integrin-mediated cell interactions and fibril remodeling. The 'matrix interaction domain' may assume a structural role, mediating collagen cross-linking, proteoglycan interactions, and tissue mineralization. Molecular modeling was used to superimpose the positions of functional sites and mutations from the two-dimensional fibril map onto a three-dimensional x-ray diffraction structure of the collagen microfibril in situ, indicating the existence of domains in the native fibril. Sequence searches revealed that major fibril domain elements are conserved in type I collagens through evolution and in the type II/XI collagen fibril predominant in cartilage. Moreover, the fibril domain model provides potential insights into the genotype-phenotype relationship for several classes of human connective tissue diseases, mechanisms of integrin clustering by fibrils, the polarity of fibril assembly, heterotypic fibril function, and connective tissue pathology in diabetes and aging.

  13. Structure-mechanics relationships of collagen fibrils in the osteogenesis imperfecta mouse model.

    PubMed

    Andriotis, O G; Chang, S W; Vanleene, M; Howarth, P H; Davies, D E; Shefelbine, S J; Buehler, M J; Thurner, P J

    2015-10-01

    The collagen molecule, which is the building block of collagen fibrils, is a triple helix of two α1(I) chains and one α2(I) chain. However, in the severe mouse model of osteogenesis imperfecta (OIM), deletion of the COL1A2 gene results in the substitution of the α2(I) chain by one α1(I) chain. As this substitution severely impairs the structure and mechanics of collagen-rich tissues at the tissue and organ level, the main aim of this study was to investigate how the structure and mechanics are altered in OIM collagen fibrils. Comparing results from atomic force microscopy imaging and cantilever-based nanoindentation on collagen fibrils from OIM and wild-type (WT) animals, we found a 33% lower indentation modulus in OIM when air-dried (bound water present) and an almost fivefold higher indentation modulus in OIM collagen fibrils when fully hydrated (bound and unbound water present) in phosphate-buffered saline solution (PBS) compared with WT collagen fibrils. These mechanical changes were accompanied by an impaired swelling upon hydration within PBS. Our experimental and atomistic simulation results show how the structure and mechanics are altered at the individual collagen fibril level as a result of collagen gene mutation in OIM. We envisage that the combination of experimental and modelling approaches could allow mechanical phenotyping at the collagen fibril level of virtually any alteration of collagen structure or chemistry. PMID:26468064

  14. Structure–mechanics relationships of collagen fibrils in the osteogenesis imperfecta mouse model

    PubMed Central

    Andriotis, O. G.; Chang, S. W.; Vanleene, M.; Howarth, P. H.; Davies, D. E.; Shefelbine, S. J.; Buehler, M. J.; Thurner, P. J.

    2015-01-01

    The collagen molecule, which is the building block of collagen fibrils, is a triple helix of two α1(I) chains and one α2(I) chain. However, in the severe mouse model of osteogenesis imperfecta (OIM), deletion of the COL1A2 gene results in the substitution of the α2(I) chain by one α1(I) chain. As this substitution severely impairs the structure and mechanics of collagen-rich tissues at the tissue and organ level, the main aim of this study was to investigate how the structure and mechanics are altered in OIM collagen fibrils. Comparing results from atomic force microscopy imaging and cantilever-based nanoindentation on collagen fibrils from OIM and wild-type (WT) animals, we found a 33% lower indentation modulus in OIM when air-dried (bound water present) and an almost fivefold higher indentation modulus in OIM collagen fibrils when fully hydrated (bound and unbound water present) in phosphate-buffered saline solution (PBS) compared with WT collagen fibrils. These mechanical changes were accompanied by an impaired swelling upon hydration within PBS. Our experimental and atomistic simulation results show how the structure and mechanics are altered at the individual collagen fibril level as a result of collagen gene mutation in OIM. We envisage that the combination of experimental and modelling approaches could allow mechanical phenotyping at the collagen fibril level of virtually any alteration of collagen structure or chemistry. PMID:26468064

  15. A micro-mechanical model to determine changes of collagen fibrils under cyclic loading

    NASA Astrophysics Data System (ADS)

    Chen, Michelle L.; Susilo, Monica E.; Ruberti, Jeffrey A.; Nguyen, Thao D.

    Dynamic mechanical loading induces growth and remodeling in biological tissues. It can alter the degradation rate and intrinsic mechanical properties of collagen through cellular activity. Experiments showed that repeated cyclic loading of a dense collagen fibril substrate increased collagen stiffness and strength, lengthened the substrate, but did not significantly change the fibril areal fraction or fibril anisotropy (Susilo, et al. ``Collagen Network Hardening Following Cyclic Tensile Loading'', Interface Focus, submitted). We developed a model for the collagen fibril substrate (Tonge, et al. ``A micromechanical modeling study of the mechanical stabilization of enzymatic degradation of collagen tissues'', Biophys J, in press.) to probe whether changes in the fibril morphology and mechanical properties can explain the tissue-level properties observed during cyclic loading. The fibrils were modeled as a continuous distribution of wavy elastica, based on experimental measurements of fibril density and collagen anisotropy, and can experience damage after a critical stress threshold. Other mechanical properties in the model were fit to the stress response measured before and after the extended cyclic loading to determine changes in the strength and stiffness of collagen fibrils.

  16. Changes in collagen fibril pattern and adhesion force with collagenase-induced injury in rat Achilles tendon observed via AFM.

    PubMed

    Lee, Gi-Ja; Choi, Samjin; Chon, Jinmann; Yoo, Seungdon; Cho, Ilsung; Park, Hun-Kuk

    2011-01-01

    The Achilles tendon consists mainly of type I collagen fibers that contain collagen fibrils. When the Achilles tendon is injured, it is inflamed. The collagenase-induced model has been widely used to study tendinitis. The major advantages of atomic force microscopy (AFM) over conventional optical and electron microscopy for bio-imaging include its non-requirement of a special coating and vacuum, and its capability to perform imaging in all environments. AFM force-distance measurements have become a fundamental tool in the fields of surface chemistry, biochemistry and materials science. Therefore, the changes in the ultrastructure and adhesion force of the collagen fibrils on the Achilles tendons of rats with Achilles tendinitis were observed using AFM. The changes in the structure of the Achilles tendons were evaluated based on the diameter and D-banding of the collagen fibrils. Collagenase-induced Achilles tendinitis was induced with the injection of 30 microl crude collagenase into 7-week-old male Sprague-Dawley rats. The animals were each sacrificed on the first, second, third, fifth and seventh day after the collagenase injection. The normal and injured Achilles tendons were fixed in 4% buffered formalin and dehydrated with increasing concentrations of ethanol. AFM was performed using the non-contact mode at the resolution of 512 x 512 pixels, with a scan speed of 0.8 line/sec. The adhesion force was measured via the force-distance curve that resulted from the interactions between the AFM tip and the collagen fibril sample using the contact mode. The diameter of the collagen fibrils in the Achilles tendons significantly decreased (p < 0.05) after the collagenase injection, and the pattern of the D-banding of the collagen fibrils was similar to that of the diameter changes. The adhesion force decreased until the fifth day after the collagenase injection, but increased on the seventh day after the collagenase injection (p < 0.0001). PMID:21446543

  17. Rapid Patterning of 1-D Collagenous Topography as an ECM Protein Fibril Platform for Image Cytometry

    PubMed Central

    Xue, Niannan; Li, Xia; Bertulli, Cristina; Li, Zhaoying; Patharagulpong, Atipat; Sadok, Amine; Huang, Yan Yan Shery

    2014-01-01

    Cellular behavior is strongly influenced by the architecture and pattern of its interfacing extracellular matrix (ECM). For an artificial culture system which could eventually benefit the translation of scientific findings into therapeutic development, the system should capture the key characteristics of a physiological microenvironment. At the same time, it should also enable standardized, high throughput data acquisition. Since an ECM is composed of different fibrous proteins, studying cellular interaction with individual fibrils will be of physiological relevance. In this study, we employ near-field electrospinning to create ordered patterns of collagenous fibrils of gelatin, based on an acetic acid and ethyl acetate aqueous co-solvent system. Tunable conformations of micro-fibrils were directly deposited onto soft polymeric substrates in a single step. We observe that global topographical features of straight lines, beads-on-strings, and curls are dictated by solution conductivity; whereas the finer details such as the fiber cross-sectional profile are tuned by solution viscosity. Using these fibril constructs as cellular assays, we study EA.hy926 endothelial cells' response to ROCK inhibition, because of ROCK's key role in the regulation of cell shape. The fibril array was shown to modulate the cellular morphology towards a pre-capillary cord-like phenotype, which was otherwise not observed on a flat 2-D substrate. Further facilitated by quantitative analysis of morphological parameters, the fibril platform also provides better dissection in the cells' response to a H1152 ROCK inhibitor. In conclusion, the near-field electrospun fibril constructs provide a more physiologically-relevant platform compared to a featureless 2-D surface, and simultaneously permit statistical single-cell image cytometry using conventional microscopy systems. The patterning approach described here is also expected to form the basics for depositing other protein fibrils, seen among

  18. Modeling of bovine type-I collagen fibrils: interaction with pickling and retanning agents.

    PubMed

    Bulo, Rosa E; Siggel, Lorenz; Molnar, Ferenc; Weiss, Horst

    2007-02-12

    Bovine Type I collagen was investigated, building on a large scale computer model of a collagen fibril in water, and focusing on two stages of the leather manufacturing process. The effects of different salts (NaCl, CaCl(2), and Na(2)SO(4)) on the swelling behavior of collagen at low pH (the pickling process) were studied. The salts suppress the swelling of the fibrils at low pH and we find specific stabilizing influences for CaCl(2) and Na(2)SO(4), due to weak Ca(2+)/Cl(-) and strong SO(4) (2-)/lysine/arginine interactions, respectively. Using state-of-the-art sampling techniques, such as the metadynamics algorithm, to allow an efficient exploration of configuration space, we were able to investigate the effect of polyacrylate and poly(methyl acrylate) - two polymeric retanning agents - on the fibril. Both polymers interact with the ammonium groups on the surface, but polyacrylate shows significantly stronger interactions. We suggest that it is this stronger interaction that contributes to the reduced suitability of PAA as a tanning agent. PMID:17295396

  19. Collagen fibril architecture, domain organization, and triple-helical conformation govern its proteolysis

    SciTech Connect

    Perumal, Shiamalee; Antipova, Olga; Orgel, Joseph P.R.O.

    2008-06-24

    We describe the molecular structure of the collagen fibril and how it affects collagen proteolysis or 'collagenolysis.' The fibril-forming collagens are major components of all mammalian connective tissues, providing the structural and organizational framework for skin, blood vessels, bone, tendon, and other tissues. The triple helix of the collagen molecule is resistant to most proteinases, and the matrix metalloproteinases that do proteolyze collagen are affected by the architecture of collagen fibrils, which are notably more resistant to collagenolysis than lone collagen monomers. Until now, there has been no molecular explanation for this. Full or limited proteolysis of the collagen fibril is known to be a key process in normal growth, development, repair, and cell differentiation, and in cancerous tumor progression and heart disease. Peptide fragments generated by collagenolysis, and the conformation of exposed sites on the fibril as a result of limited proteolysis, regulate these processes and that of cellular attachment, but it is not known how or why. Using computational and molecular visualization methods, we found that the arrangement of collagen monomers in the fibril (its architecture) protects areas vulnerable to collagenolysis and strictly governs the process. This in turn affects the accessibility of a cell interaction site located near the cleavage region. Our observations suggest that the C-terminal telopeptide must be proteolyzed before collagenase can gain access to the cleavage site. Collagenase then binds to the substrate's 'interaction domain,' which facilitates the triple-helix unwinding/dissociation function of the enzyme before collagenolysis.

  20. Tyrosine-rich acidic matrix protein (TRAMP) accelerates collagen fibril formation in vitro.

    PubMed

    MacBeath, J R; Shackleton, D R; Hulmes, D J

    1993-09-15

    Tyrosine-rich acidic matrix protein (TRAMP) is a recently discovered protein that co-purifies with porcine skin lysyl oxidase and is equivalent to the M(r) 22,000 extracellular matrix protein from bovine skin that co-purifies with dermatan sulfate proteoglycans (Cronshaw, A. D., MacBeath, J. R. E., Shackleton, D. R., Collins, J. F., Fothergill-Gilmore, L. A., and Hulmes, D. J. S. (1993) Matrix 13, 255-266; Neame, P. J., Choi, H. U., and Rosenberg, L. C. (1989) J. Biol. Chem. 264, 5474-5479). The effect of TRAMP on collagen fibril formation was studied in vitro by reconstitution of fibrils from lathyritic rat skin collagen I. Fibril formation was initiated by the warm start procedure, in which acidic collagen solutions and double strength neutral buffer, both preincubated separately at 34 degrees C, were mixed. When monitored by turbidimetry, TRAMP was found to accelerate collagen fibril formation. Acceleration occurred at sub-stoichiometric molar ratios of TRAMP collagen, and the presence of TRAMP stabilized the fibrils against low temperature dissociation. It was confirmed by centrifugation that the amount of fibrillar collagen formed in the presence of TRAMP was greater than in its absence. By SDS-polyacrylamide gel electrophoresis and scanning densitometry, binding of TRAMP to collagen was detected that approached saturation with a molar ratio of TRAMP to collagen of approximately 1:2. Fibrils formed in the presence of TRAMP were normal when observed by electron microscopy, although fibril diameters were smaller than the controls. TRAMP was found to partially reverse the inhibitory effects of urea and increased ionic strength on the kinetics of fibril formation, although inhibition by glucose was unaffected. TRAMP also accelerated the assembly of pepsin-treated collagen, where the non-helical, telopeptide regions were partially removed. Acceleration of collagen fibril formation by TRAMP is discussed in the light of the known effects of other extracellular matrix

  1. Coalignment of microtubules, cytokeratin intermediate filaments, and collagen fibrils in a collagen-secreting cell system.

    PubMed

    McBeath, E; Fujiwara, K

    1989-12-01

    The distribution of microtubules and intermediate filaments in the collagen-secreting scleroblasts of the goldfish scale was investigated by immunofluorescence and electron microscopy. Many of the microtubules and cytokeratin type intermediate filaments formed bundles that were aligned with the underlying, parallel collagen fibrils. The intermediate filament bundles were evenly spaced and located adjacent to the basal plasma membrane. The microtubules, on the other hand, were located further away from the membrane, although many were found very close to the intermediate filament bundles. No detectable change was observed in scleroblast microtubules when cells on scales were treated with colchicine or cooled (greater than or equal to 0 degrees C) for up to 1 h. Cells had to be cooled overnight before the microtubules were affected. The final number and length of the microtubules in the cell depended only on the final steady-state temperature and not the temperature history of the scale cell, and steady state was reached more slowly at colder temperatures. The microtubules but not the intermediate filaments rapidly (within 5 min) and reversibly depolymerized when cells were chilled to -2 approximately -4 degrees C. When chilled cells were warmed, the microtubules polymerized back, within 15 min at room temperature, to the same pattern of parallel coalignment with the underlying collagen. They appeared to repolymerize via two different pathways: (1) a radial growth outwards from the microtubule organizing center followed by a progressive realignment with the underlying collagen and (2) a gradual and simultaneous polymerization along cold-stable, antitubulin staining fibers. These fibers were also aligned with the collagen fibrils and may be related to the aligned intermediate filaments. PMID:2483378

  2. Large Deformation Mechanisms, Plasticity, and Failure of an Individual Collagen Fibril With Different Mineral Content

    PubMed Central

    Depalle, Baptiste; Qin, Zhao; Shefelbine, Sandra J

    2016-01-01

    ABSTRACT Mineralized collagen fibrils are composed of tropocollagen molecules and mineral crystals derived from hydroxyapatite to form a composite material that combines optimal properties of both constituents and exhibits incredible strength and toughness. Their complex hierarchical structure allows collagen fibrils to sustain large deformation without breaking. In this study, we report a mesoscale model of a single mineralized collagen fibril using a bottom‐up approach. By conserving the three‐dimensional structure and the entanglement of the molecules, we were able to construct finite‐size fibril models that allowed us to explore the deformation mechanisms which govern their mechanical behavior under large deformation. We investigated the tensile behavior of a single collagen fibril with various intrafibrillar mineral content and found that a mineralized collagen fibril can present up to five different deformation mechanisms to dissipate energy. These mechanisms include molecular uncoiling, molecular stretching, mineral/collagen sliding, molecular slippage, and crystal dissociation. By multiplying its sources of energy dissipation and deformation mechanisms, a collagen fibril can reach impressive strength and toughness. Adding mineral into the collagen fibril can increase its strength up to 10 times and its toughness up to 35 times. Combining crosslinks with mineral makes the fibril stiffer but more brittle. We also found that a mineralized fibril reaches its maximum toughness to density and strength to density ratios for a mineral density of around 30%. This result, in good agreement with experimental observations, attests that bone tissue is optimized mechanically to remain lightweight but maintain strength and toughness. © 2015 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research (ASBMR). PMID:26866939

  3. Structural changes in collagen fibrils across a mineralized interface revealed by cryo-TEM.

    PubMed

    Quan, Bryan D; Sone, Eli D

    2015-08-01

    The structure of the mineralized collagen fibril, which is the basic building block of mineralized connective tissues, is critical to its function. We use cryo-TEM to study collagen structure at a well-defined hard-soft tissue interface, across which collagen fibrils are continuous, in order to evaluate changes to collagen upon mineralization. To establish a basis for the analysis of collagen banding, we compared cryo-TEM images of rat-tail tendon collagen to a model based on the X-ray structure. While there is close correspondence of periodicity, differences in band intensity indicate fibril regions with high density but lacking order, providing new insight into collagen fibrillar structure. Across a mineralized interface, we show that mineralization results in an axial contraction of the fibril, concomitant with lateral expansion, and that this contraction occurs only in the more flexible gap region of the fibril. Nevertheless, the major features of the banding pattern are not significantly changed, indicating that the axial arrangement of molecules remains largely intact. These results suggest a mechanism by which collagen fibrils are able to accommodate large amounts of mineral without significant disruption of their molecular packing, leading to synergy of mechanical properties. PMID:25892483

  4. Three-dimensional modeling and computational analysis of the human cornea considering distributed collagen fibril orientations.

    PubMed

    Pandolfi, Anna; Holzapfel, Gerhard A

    2008-12-01

    Experimental tests on human corneas reveal distinguished reinforcing collagen lamellar structures that may be well described by a structural constitutive model considering distributed collagen fibril orientations along the superior-inferior and the nasal-temporal meridians. A proper interplay between the material structure and the geometry guarantees the refractive function and defines the refractive properties of the cornea. We propose a three-dimensional computational model for the human cornea that is able to provide the refractive power by analyzing the structural mechanical response with the nonlinear regime and the effect the intraocular pressure has. For an assigned unloaded geometry we show how the distribution of the von Mises stress at the top surface of the cornea and through the corneal thickness and the refractive power depend on the material properties and the fibril dispersion. We conclude that a model for the human cornea must not disregard the peculiar collagen fibrillar structure, which equips the cornea with the unique biophysical, mechanical, and optical properties. PMID:19045535

  5. The Collagen Fibril Structure in the Superficial Zone of Articular Cartilage by μMRI

    PubMed Central

    Zheng, ShaoKuan; Xia, Yang

    2009-01-01

    Objective To investigate the fibril architecture of the collage matrix in the superficial zone of articular cartilage non-destructively by microscopic MRI (μMRI) T2 anisotropy. Method Six specimens of canine humeral cartilage were rotated in such a way that the normal axis of the articular surface of the cartilage specimen remained stationary and perpendicular to the static magnetic field, over a range of 180° and at a step of 15°. At each rotation angle, a quantitative T2 image was constructed at 13μm pixel resolution. Results A set of complex and depth-dependent patterns was found in the μMRI T2 anisotropy along the depth of the tissue. In the superficial zone, the T2 anisotropy is clearly periodic, which demonstrates that the distribution of the collagen fibrils in the superficial zone is not random. In the transitional zone, the periodicity of the T2 anisotropy approximately doubles with respect to that in the superficial zone. In the initial part of the radial zone, the T2 anisotropy is also periodic but inverse to that in the superficial zone. In the deep part of the radial zone, the T2 anisotropy becomes increasingly weaker and eventually disappears. Conclusion There exists a certain degree of collagen anisotropy in all zones of articular cartilage. The anisotropic imaging data can be interpreted with the aid of a collagen architecture model. PMID:19527808

  6. Minerals and aligned collagen fibrils in tilapia fish scales: structural analysis using dark-field and energy-filtered transmission electron microscopy and electron tomography.

    PubMed

    Okuda, Mitsuhiro; Ogawa, Nobuhiro; Takeguchi, Masaki; Hashimoto, Ayako; Tagaya, Motohiro; Chen, Song; Hanagata, Nobutaka; Ikoma, Toshiyuki

    2011-10-01

    The mineralized structure of aligned collagen fibrils in a tilapia fish scale was investigated using transmission electron microscopy (TEM) techniques after a thin sample was prepared using aqueous techniques. Electron diffraction and electron energy loss spectroscopy data indicated that a mineralized internal layer consisting of aligned collagen fibrils contains hydroxyapatite crystals. Bright-field imaging, dark-field imaging, and energy-filtered TEM showed that the hydroxyapatite was mainly distributed in the hole zones of the aligned collagen fibrils structure, while needle-like materials composed of calcium compounds including hydroxyapatite existed in the mineralized internal layer. Dark-field imaging and three-dimensional observation using electron tomography revealed that hydroxyapatite and needle-like materials were mainly found in the matrix between the collagen fibrils. It was observed that hydroxyapatite and needle-like materials were preferentially distributed on the surface of the hole zones in the aligned collagen fibrils structure and in the matrix between the collagen fibrils in the mineralized internal layer of the scale. PMID:21899811

  7. Collagen Self-Assembly on Orthopedic Magnesium Biomaterials Surface and Subsequent Bone Cell Attachment

    PubMed Central

    Zhao, Nan; Zhu, Donghui

    2014-01-01

    Magnesium (Mg) biomaterials are a new generation of biodegradable materials and have promising potential for orthopedic applications. After implantation in bone tissues, these materials will directly interact with extracellular matrix (ECM) biomolecules and bone cells. Type I collagen, the major component of bone ECM, forms the architecture scaffold that provides physical support for bone cell attachment. However, it is still unknown how Mg substrate affects collagen assembly on top of it as well as subsequent cell attachment and growth. Here, we studied the effects of collagen monomer concentration, pH, assembly time, and surface roughness of two Mg materials (pure Mg and AZ31) on collagen fibril formation. Results showed that formation of fibrils would not initiate until the monomer concentration reached a certain level depending on the type of Mg material. The thickness of collagen fibril increased with the increase of assembly time. The structures of collagen fibrils formed on semi-rough surfaces of Mg materials have a high similarity to that of native bone collagen. Next, cell attachment and growth after collagen assembly were examined. Materials with rough surface showed higher collagen adsorption but compromised bone cell attachment. Interestingly, surface roughness and collagen structure did not affect cell growth on AZ31 for up to a week. Findings from this work provide some insightful information on Mg-tissue interaction at the interface and guidance for future surface modifications of Mg biomaterials. PMID:25303459

  8. Determination of the elastic modulus of native collagen fibrils via radial indentation

    NASA Astrophysics Data System (ADS)

    Heim, August J.; Matthews, William G.; Koob, Thomas J.

    2006-10-01

    The authors studied the elastic response of single, native collagen fibrils extracted from tissues of the inner dermis of the sea cucumber, Cucumaria frondosa, via local nanoscale indentation with an atomic force microscope (AFM). AFM imaging of fibrils under ambient conditions are presented, demonstrating a peak-to-peak periodicity, the d band, of dehydrated, unfixed fibrils to be ˜64.5nm. Radial indentation experiments were performed, and the measured value for the reduced modulus is 1-2GPa.

  9. Chondroitin sulfate perlecan enhances collagen fibril formation. Implications for perlecan chondrodysplasias.

    PubMed

    Kvist, Alexander J; Johnson, Anna E; Mörgelin, Matthias; Gustafsson, Erika; Bengtsson, Eva; Lindblom, Karin; Aszódi, Attila; Fässler, Reinhard; Sasaki, Takako; Timpl, Rupert; Aspberg, Anders

    2006-11-01

    Inactivation of the perlecan gene leads to perinatal lethal chondrodysplasia. The similarity to the phenotypes of the Col2A1 knock-out and the disproportionate micromelia mutation suggests perlecan involvement in cartilage collagen matrix assembly. We now present a mechanism for the defect in collagen type II fibril assembly by perlecan-null chondrocytes. Cartilage perlecan is a heparin sulfate or a mixed heparan sulfate/chondroitin sulfate proteoglycan. The latter form binds collagen and accelerates fibril formation in vitro, with more defined fibril morphology and increased fibril diameters produced in the presence of perlecan. Interestingly, the enhancement of collagen fibril formation is independent on the core protein and is mimicked by chondroitin sulfate E but neither by chondroitin sulfate D nor dextran sulfate. Furthermore, perlecan chondroitin sulfate contains the 4,6-disulfated disaccharides typical for chondroitin sulfate E. Indeed, purified glycosaminoglycans from perlecan-enriched fractions of cartilage extracts contain elevated levels of 4,6-disulfated chondroitin sulfate disaccharides and enhance collagen fibril formation. The effect on collagen assembly is proportional to the content of the 4,6-disulfated disaccharide in the different cartilage extracts, with growth plate cartilage glycosaminoglycan being the most efficient enhancer. These findings demonstrate a role for perlecan chondroitin sulfate side chains in cartilage extracellular matrix assembly and provide an explanation for the perlecan-null chondrodysplasia. PMID:16956876

  10. Thermal denaturation behavior of collagen fibrils in wet and dry environment.

    PubMed

    Suwa, Yosuke; Nam, Kwangwoo; Ozeki, Kazuhide; Kimura, Tsuyoshi; Kishida, Akio; Masuzawa, Toru

    2016-04-01

    We have developed a new minimally invasive technique--integrated low-level energy adhesion technique (ILEAT)--which uses heat, pressure, and low-frequency vibrations for binding living tissues. Because the adhesion mechanism of the living tissues is not fully understood, we investigated the effect of thermal energy on the collagen structure in living tissues using ILEAT. To study the effect of thermal energy and heating time on the structure of the collagen fibril, samples were divided in two categories-wet and dry. Further, atomic force microscopy was used to analyze the collagen fibril structure before and after heating. Results showed that collagen fibrils in water denatured after 1 minute at temperatures higher than 80 °C, while partial denaturation was observed at temperatures of 80 °C and a heating time of 1 min. Furthermore, complete denaturation was achieved after 90 min, suggesting that the denaturation rate is temperature and time dependent. Moreover, the collagen fibrils in dry condition maintained their native structure even after being heated to 120 °C for 90 min in the absence of water, which specifically suppressed denaturation. However, partial denaturation of collagen fibrils could not be prevented, because this determines the adhesion between the collagen molecules, and stabilizes tissue bonding. PMID:25952296

  11. Second harmonic generation quantitative measurements on collagen fibrils through correlation to electron microscopy

    NASA Astrophysics Data System (ADS)

    Bancelin, S.; Aimé, C.; Gusachenko, I.; Kowalczuk, L.; Latour, G.; Coradin, T.; Schanne-Klein, M.-C.

    2015-03-01

    Type I collagen is a major structural protein in mammals that shows highly structured macromolecular organizations specific to each tissue. This biopolymer is synthesized as triple helices, which self-assemble into fibrils (Ø =10-300 nm) and further form various 3D organization. In recent years, Second Harmonic Generation (SHG) microscopy has emerged as a powerful technique to probe in situ the fibrillar collagenous network within tissues. However, this optical technique cannot resolve most of the fibrils and is a coherent process, which has impeded quantitative measurements of the fibril diameter so far. In this study, we correlated SHG microscopy with Transmission Electron Microscopy to determine the sensitivity of SHG microscopy and to calibrate SHG signals as a function of the fibril diameter in reconstructed collagen gels. To that end, we synthetized isolated fibrils with various diameters and successfully imaged the very same fibrils with both techniques, down to 30 nm diameter. We observed that SHG signals scaled as the fourth power of the fibril diameter, as expected from analytical and numerical calculations. This calibration was then applied to diabetic rat cornea in which we successfully recovered the diameter of hyperglycemia-induced fibrils in the Descemet's membrane without having to resolve them. Finally we derived the first hyperpolarizability from a single collagen triple helix which validates the bottom-up approach used to calculate the non-linear response at the fibrillar scale and denotes a parallel alignment of triple helices within the fibrils. These results represent a major step towards quantitative SHG imaging of nm-sized collagen fibrils.

  12. Nanoscale characterization of the biomechanical properties of collagen fibrils in the sclera

    SciTech Connect

    Papi, M.; Paoletti, P.; Geraghty, B.; Akhtar, R.

    2014-03-10

    We apply the PeakForce Quantitative Nanomechanical Property Mapping (PFQNM) atomic force microscopy mode for the investigation of regional variations in the nanomechanical properties of porcine sclera. We examine variations in the collagen fibril diameter, adhesion, elastic modulus and dissipation in the posterior, equatorial and anterior regions of the sclera. The mean fibril diameter, elastic modulus and dissipation increased from the posterior to the anterior region. Collagen fibril diameter correlated linearly with elastic modulus. Our data matches the known macroscopic mechanical behavior of the sclera. We propose that PFQNM has significant potential in ocular biomechanics and biophysics research.

  13. Fracture Mechanics of Collagen Fibrils: Influence of Natural Cross-Links

    PubMed Central

    Svensson, Rene B.; Mulder, Hindrik; Kovanen, Vuokko; Magnusson, S. Peter

    2013-01-01

    Tendons are important load-bearing structures, which are frequently injured in both sports and work. Type I collagen fibrils are the primary components of tendons and carry most of the mechanical loads experienced by the tissue, however, knowledge of how load is transmitted between and within fibrils is limited. The presence of covalent enzymatic cross-links between collagen molecules is an important factor that has been shown to influence mechanical behavior of the tendons. To improve our understanding of how molecular bonds translate into tendon mechanics, we used an atomic force microscopy technique to measure the mechanical behavior of individual collagen fibrils loaded to failure. Fibrils from human patellar tendons, rat-tail tendons (RTTs), NaBH4 reduced RTTs, and tail tendons of Zucker diabetic fat rats were tested. We found a characteristic three-phase stress-strain behavior in the human collagen fibrils. There was an initial rise in modulus followed by a plateau with reduced modulus, which was finally followed by an even greater increase in stress and modulus before failure. The RTTs also displayed the initial increase and plateau phase, but the third region was virtually absent and the plateau continued until failure. The importance of cross-link lability was investigated by NaBH4 reduction of the rat-tail fibrils, which did not alter their behavior. These findings shed light on the function of cross-links at the fibril level, but further studies will be required to establish the underlying mechanisms. PMID:23746520

  14. The organisation of collagen fibrils in the superficial zones of articular cartilage.

    PubMed Central

    Clark, J M

    1990-01-01

    The origin and structure of collagen fibres in the surface of articular cartilage were studied using SEM. Cryofracture was used to create orthogonal fracture surfaces in three planes. Fibres which originated in the radial zone could be traced into the surface where they flattened and overlapped in a common direction. Thick fibres from the periosteum ran into the surface as well, but apparently ended there and did not enter the radial zone. The tangential fibres were covered by a dense, separate layer of small fibrils. The fundamental aspects of the model proposed by Benninghoff are supported by these findings. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 PMID:2081698

  15. Two-Dimensional Nanoscale Structural and Functional Imaging in Individual Collagen Type I Fibrils

    PubMed Central

    Harnagea, Catalin; Vallières, Martin; Pfeffer, Christian P.; Wu, Dong; Olsen, Bjorn R.; Pignolet, Alain; Légaré, François; Gruverman, Alexei

    2010-01-01

    Abstract The piezoelectric properties of single collagen type I fibrils in fascia were imaged with sub-20 nm spatial resolution using piezoresponse force microscopy. A detailed analysis of the piezoresponse force microscopy signal in controlled tip-fibril geometry revealed shear piezoelectricity parallel to the fibril axis. The direction of the displacement is preserved along the whole fiber length and is independent of the fiber conformation. It is shown that individual fibrils within bundles in skeletal muscle fascia can have opposite polar orientations and are organized into domains, i.e., groups of several fibers having the same polar orientation. We were also able to detect piezoelectric activity of collagen fibrils in the high-frequency range up to 200 kHz, suggesting that the mechanical response time of biomolecules to electrical stimuli can be ∼5 μs. PMID:20550920

  16. Characterization of type I collagen fibril formation using thioflavin T fluorescent dye.

    PubMed

    Morimoto, Koichi; Kawabata, Kazuya; Kunii, Saori; Hamano, Kaori; Saito, Takuya; Tonomura, Ben'ichiro

    2009-05-01

    Collagen is composed of fibrils that are formed by self-assembly of smaller units, monomers which are triple-helical polypeptide. However, the mechanism of fibril formation at the level of individual molecules has remained to be clarified. We found that the fluorescence of thioflavin T, which has been widely used as a specific dye for amyloid fibrils, also increased by binding with fibrils of atelocollagen prepared from yellowfin tuna skin. There was a linear correlation between the fluorescence increase and the amount of atelocollagen within a collagen concentration range of 0-0.15 mg/ml at pH 6.5 with 50 microM thioflavin T. In contrast, neither actinidain-processed collagen that keeps monomeric nature nor heat-denatured collagen could cause the fluorescence increase of thioflavin T at all. The relationship between the fluorescence increase and thioflavin T concentration was fit to a theoretical binary binding curve. An apparent dissociation constant, K(d), and a maximal fluorescence increase, DeltaF(max), were calculated at various pHs. The values of K(d) and DeltaF(max) were dependent on pH (K(d) was 9.4 microM at pH 6.5). The present finding demonstrates that thioflavin T specifically binds to collagen fibrils and may be used as a sensitive tool for the study of collagen structure. PMID:19204013

  17. Effects of hydration on the mechanical response of individual collagen fibrils

    NASA Astrophysics Data System (ADS)

    Grant, Colin A.; Brockwell, David J.; Radford, Sheena E.; Thomson, Neil H.

    2008-06-01

    Collagen fibrils prepared from bovine Achilles tendon have been mechanically tested through nanoindentation by an atomic force microscope using force volume analysis. In ambient conditions where the fibrils are expected to be dehydrated, the elastic modulus was determined to be 1.9±0.5GPa, while under aqueous fluid, it decreased by three orders of magnitude to 1.2±0.1MPa. In air, fibril fracture occurred along the axis and the crack lengths were quantized to the D-banding periodicity. The apparent hardness of the fibrils was estimated to be in the range of 0.35-0.6GPa.

  18. Effects of photochemical riboflavin-mediated crosslinks on the physical properties of collagen constructs and fibrils.

    PubMed

    Rich, Harvey; Odlyha, Marianne; Cheema, Umber; Mudera, Vivek; Bozec, Laurent

    2014-01-01

    The use of collagen scaffold in tissue engineering is on the rise, as modifications to mechanical properties are becoming more effective in strengthening constructs whilst preserving the natural biocompatibility. The combined technique of plastic compression and cross-linking is known to increase the mechanical strength of the collagen construct. Here, a modified protocol for engineering these collagen constructs is used to bring together a plastic compression method, combined with controlled photochemical crosslinking using riboflavin as a photoinitiator. In order to ascertain the effects of the photochemical crosslinking approach and the impact of the crosslinks created upon the properties of the engineered collagen constructs, the constructs were characterized both at the macroscale and at the fibrillar level. The resulting constructs were found to have a 2.5 fold increase in their Young's modulus, reaching a value of 650 ± 73 kPa when compared to non-crosslinked control collagen constructs. This value is not yet comparable to that of native tendon, but it proves that combining a crosslinking methodology to collagen tissue engineering may offer a new approach to create stronger, biomimetic constructs. A notable outcome of crosslinking collagen with riboflavin is the collagen's greater affinity for water; it was demonstrated that riboflavin crosslinked collagen retains water for a longer period of time compared to non-cross-linked control samples. The affinity of the cross-linked collagen to water also resulted in an increase of individual collagen fibrils' cross-sectional area as function of the crosslinking. These changes in water affinity and fibril morphology induced by the process of crosslinking could indicate that the crosslinked chains created during the photochemical crosslinking process may act as intermolecular hydrophilic nanosprings. These intermolecular nanosprings would be responsible for a change in the fibril morphology to accommodate variable

  19. Bowstring Stretching and Quantitative Imaging of Single Collagen Fibrils via Atomic Force Microscopy.

    PubMed

    Quigley, Andrew S; Veres, Samuel P; Kreplak, Laurent

    2016-01-01

    Collagen is the primary structural protein in animals. Serving as nanoscale biological ropes, collagen fibrils are responsible for providing strength to a variety of connective tissues such as tendon, skin, and bone. Understanding structure-function relationships in collagenous tissues requires the ability to conduct a variety of mechanical experiments on single collagen fibrils. Though significant advances have been made, certain tests are not possible using the techniques currently available. In this report we present a new atomic force microscopy (AFM) based method for tensile manipulation and subsequent nanoscale structural assessment of single collagen fibrils. While the method documented here cannot currently capture force data during loading, it offers the great advantage of allowing structural assessment after subrupture loading. To demonstrate the utility of this technique, we describe the results of 23 tensile experiments in which collagen fibrils were loaded to varying levels of strain and subsequently imaged in both the hydrated and dehydrated states. We show that following a dehydration-rehydration cycle (necessary for sample preparation), fibrils experience an increase in height and decrease in radial modulus in response to one loading-unloading cycle to strain <5%. This change is not altered by a second cycle to strain >5%. In fibril segments that ruptured during their second loading cycle, we show that the fibril structure is affected away from the rupture site in the form of discrete permanent deformations. By comparing the severity of select damage sites in both hydrated and dehydrated conditions, we demonstrate that dehydration masks damage features, leading to an underestimate of the degree of structural disruption. Overall, the method shows promise as a powerful tool for the investigation of structure-function relationships in nanoscale fibrous materials. PMID:27598334

  20. Metastatic Bladder Cancer Cells Distinctively Sense and Respond to Physical Cues of Collagen Fibril-Mimetic Nanotopography

    PubMed Central

    Iuliano, James N.; Kutscha, Paul D.; Biderman, Norbert J.; Subbaram, Sita; J; Groves, Timothy R.; Tenenbaum, Scott A.; Hempel, Nadine

    2015-01-01

    Tumor metastasis is characterized by enhanced invasiveness and migration of tumor cells through the extracellular matrix (ECM), resulting in extravasation into the blood and lymph and colonization at secondary sites. The ECM provides a physical scaffold consisting of components such as collagen fibrils, which have distinct dimensions at the nano-scale. In addition to the interaction of peptide moieties with tumor cell integrin clusters, the ECM provides a physical guide for tumor cell migration. Using nanolithography we set out to mimic the physical dimensions of collagen fibrils using lined nanotopographical silicon surfaces and to explore whether metastatic tumor cells are uniquely able to respond to these physical dimensions. Etched silicon surfaces containing nanoscale lined-patterns with varying trench and ridge sizes (65-500 nm) were evaluated for their ability to distinguish between a non-metastatic (253J) and a highly metastatic (253-J-BV) derivative bladder cancer cell line. Enhanced alignment was distinctively observed for the metastatic cell lines on feature sizes that mimic the dimensions of collagen fibrils (65-100 nm lines, 1:1-1:1.5 pitch). Further, these sub-100 nm lines acted as guides for migration of metastatic cancer cells. Interestingly, even at this sub-cellular scale, metastatic cell migration was abrogated when cells were forced to move perpendicular to these lines. Compared to flat surfaces, 65 nm lines enhanced the formation of actin stress fibers and filopodia of metastatic cells. This was accompanied by increased formation of focal contacts, visualized by immunofluorescent staining of phospho-focal adhesion Kinase (FAK) along the protruding lamellopodia. Simple lined nanotopography appears to be an informative platform for studying the physical cues of the ECM in a pseudo-3D format and likely mimics physical aspects of collagen fibrils. Metastatic cancer cells appear distinctively well-adapted to sense these features using filopodia

  1. Dissolution of type I collagen fibrils by gingival fibroblasts isolated from patients of various periodontitis categories.

    PubMed

    Havemose-Poulsen, A; Holmstrup, P; Stoltze, K; Birkedal-Hansen, H

    1998-07-01

    The classification of periodontitis in various disease categories, including juvenile periodontitis, rapidly progressive adult periodontitis and slowly progressive adult periodontitis is based mainly on differences in disease progression and age group susceptibility. Because dissolution of collagen fibers is an integral part of periodontal attachment loss, we investigated whether the clinical differences among these periodontitis/control groups are reflected in the collagen-degrading activity of gingival fibroblasts isolated from affected tissues. All fibroblast strains isolated from the 4 groups (n = 48) displayed cell-associated collagenolytic activity when seeded in contact with a reconstituted film of type I collagen fibrils. Cells from the control group (n = 14) dissolved the collagen fibril film twice as fast as those from each of the 3 disease groups (juvenile periodontitis (n = 13), rapidly progressive adult periodontitis (n = 7), and slowly progressive adult periodontitis (n = 14)). Both interleukin-1 beta and phorbolester accelerated the rate of dissolution 2-4-fold, but even after cytokine or phorbolester stimulation control cells were still considerably more effective in dissolving the collagen fibrils than cells from the disease groups. The observation made in this study, that dissolution of collagen fibrils by gingival fibroblasts from periodontally diseased individuals is significantly slower than by cells from healthy control subjects, challenges disease paradigms based on a direct relationship between collagenolytic potential and disease activity. PMID:9777595

  2. 3D Raman mapping of the collagen fibril orientation in human osteonal lamellae.

    PubMed

    Schrof, Susanne; Varga, Peter; Galvis, Leonardo; Raum, Kay; Masic, Admir

    2014-09-01

    Chemical composition and fibrillar organization are the major determinants of osteonal bone mechanics. However, prominent methodologies commonly applied to investigate mechanical properties of bone on the micro scale are usually not able to concurrently describe both factors. In this study, we used polarized Raman spectroscopy (PRS) to simultaneously analyze structural and chemical information of collagen fibrils in human osteonal bone in a single experiment. Specifically, the three-dimensional arrangement of collagen fibrils in osteonal lamellae was assessed. By analyzing the anisotropic intensity of the amide I Raman band of collagen as a function of the orientation of the incident laser polarization, different parameters related to the orientation of the collagen fibrils and the degree of alignment of the fibrils were derived. Based on the analysis of several osteons, two major fibrillar organization patterns were identified, one with a monotonic and another with a periodically changing twist direction. These results confirm earlier reported twisted and oscillating plywood arrangements, respectively. Furthermore, indicators of the degree of alignment suggested the presence of disordered collagen within the lamellar organization of the osteon. The results show the versatility of the analytical PRS approach and demonstrate its capability in providing not only compositional, but also 3D structural information in a complex hierarchically structured biological material. The concurrent assessment of chemical and structural features may contribute to a comprehensive characterization of the microstructure of bone and other collagen-based tissues. PMID:25025981

  3. Intracellular collagen fibrils: evidence of an intracellular source from experiments with tendon fibroblasts and fibroblastic tumour cells.

    PubMed Central

    Michna, H

    1988-01-01

    This study was designed to substantiate one or both of the two hypotheses for the explanation of intracellular collagen fibrils in collagen-producing cells. The more obvious is the phagocytosis of extracellular collagen fibrils by the cell and the other is a form of autophagocytosis of newly synthesised collagenous products. Information was collected on fibroblasts from murine tendons after exercise and simultaneously stimulating collagen synthesis by treatment with an anabolic steroid hormone. Moreover, in vivo and in vitro fibroblastic tumour cells which demonstrate enhanced protein synthesis were also treated with the anabolic steroid. The findings of intracellular collagen fibrils in tendon fibroblasts and the sarcoma cells after experimentally stimulating collagen synthesis are discussed in the light of the hypothesis that the findings may represent steps of autophagocytosis of newly synthesised collagenous products in the absence of a control mechanism to remove collagenous products which cannot be secreted. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:3225213

  4. Nanoscale characterization of isolated individual type I collagen fibrils: polarization and piezoelectricity

    NASA Astrophysics Data System (ADS)

    Minary-Jolandan, Majid; Yu, Min-Feng

    2009-02-01

    Piezoresponse force microscopy was applied to directly study individual type I collagen fibrils with diameters of ~100 nm isolated from bovine Achilles tendon. It was revealed that single collagen fibrils behave predominantly as shear piezoelectric materials with a piezoelectric coefficient on the order of 1 pm V-1, and have unipolar axial polarization throughout their entire length. It was estimated that, under reasonable shear load conditions, the fibrils were capable of generating an electric potential up to tens of millivolts. The result substantiates the nanoscale origin of piezoelectricity in bone and tendons, and implies also the potential importance of the shear load-transfer mechanism, which has been the principle basis of the nanoscale mechanics model of collagen, in mechanoelectric transduction in bone.

  5. The 3D structure of the collagen fibril network in human trabecular bone: relation to trabecular organization.

    PubMed

    Reznikov, Natalie; Chase, Hila; Brumfeld, Vlad; Shahar, Ron; Weiner, Steve

    2015-02-01

    Trabecular bone is morphologically and functionally different from compact bone at the tissue level, but both are composed of lamellae at the micrometer-scale level. We present a three-dimensional study of the collagenous network of human trabecular lamellar bone from the proximal femur using the FIB-SEM serial surface view method. The results are compared to human compact lamellar bone of the femoral shaft, studied by the same method. Both demineralized trabecular and compact lamellar bone display the same overall structural organization, namely the presence of ordered and disordered materials and the confinement of the canalicular network to the disordered material. However, in trabecular bone lamellae a significant proportion of the ordered collagen fibril arrays is aligned with the long axis of the trabecula and, unlike in compact bone, is not related to the anatomical axis of the whole femur. The remaining ordered collagen fibrils are offset from the axis of a trabecula either by about 30° or 70°. Interestingly, at the tissue scale of millimeters, the most abundant angles between any two connected trabeculae - the inter-trabecular angles - center around 30° and 70°. This implies that within a framework of interconnected trabeculae the same lamellar structure will always have a significant component of the fibrils aligned with the long axes of connected trabeculae. This structural complementarity at different hierarchical levels presumably reflects an adaptation of trabecular bone to function. PMID:25445457

  6. Nano-mechanical properties of individual mineralized collagen fibrils from bone tissue.

    PubMed

    Hang, Fei; Barber, Asa H

    2011-04-01

    Mineralized collagen fibrils (MCFs) are distinct building blocks for bone material and perform an important mechanical function. A novel experimental technique using combined atomic force microscopy and scanning electron microscopy is used to manipulate and measure the mechanical properties of individual MCFs from antler, which is a representative bone tissue. The recorded stress-strain response of individual MCFs under tension shows an initial linear deformation region for all fibrils, followed by inhomogeneous deformation above a critical strain. This inhomogeneous deformation is indicative of fibrils exhibiting either yield or strain hardening and suggests possible mineral compositional changes within each fibril. A phenomenological model is used to describe the fibril nano-mechanical behaviour. PMID:20961895

  7. Second harmonic generation imaging of the collagen in myocardium for atrial fibrillation diagnosis

    NASA Astrophysics Data System (ADS)

    Tsai, Ming-Rung; Chiou, Yu-We; Sun, Chi-Kuang

    2009-02-01

    Myocardial fibrosis, a common sequela of cardiac hypertrophy, has been shown to be associated with arrhythmias in experimental models. Some research has indicated that myocardial fibrosis plays an important role in predisposing patients to atrial fibrillation. Second harmonic generation (SHG) is an optically nonlinear coherent process to image the collagen network. In this presentation, we observe the SHG images of the collagen matrix in atrial myocardium and we analyzed of collagen fibers arrangement by using Fourier-transform analysis. Moreover, comparing the SHG images of the collagen fibers in atrial myocardium between normal sinus rhythm (NSR) and atrial fibrillation (AF), our result indicated that it is possible to realize the relation between myocardial fibrosis and AF.

  8. Effect of γ-PGA on the formation of collagen fibrils in vitro.

    PubMed

    Ding, Cuicui; Zheng, Zhigong; Liu, Xinzhong; Li, Hengda; Zhang, Min

    2016-07-01

    The effect of γ-poly(glutamic acid) (γ-PGA) on the self-assembly of collagen was studied. Under physiological conditions, the kinetic curves for fibril formation showed that the turbidity of collagen/γ-PGA blends at 313 nm was increased with the addition of γ-PGA. Furthermore, it was shown using both field-emission scanning electron microscopy (FESEM) and atomic force microscopy (AFM) that fibrils with a larger diameter were obtained following the addition of γ-PGA, probably due to the electrostatic and hydrogen bond interactions between collagen and γ-PGA, which promoted the lateral association of collagen molecules. In addition, both the thermal stability and viscoelastic properties of the hybrid hydrogels, which were evaluated by differential scanning calorimetry and rheological measurements, respectively, were improved by the addition of γ-PGA. PMID:26940941

  9. Poisson's ratio of collagen fibrils measured by small angle X-ray scattering of strained bovine pericardium

    SciTech Connect

    Wells, Hannah C.; Sizeland, Katie H.; Kayed, Hanan R.; Haverkamp, Richard G.; Kirby, Nigel; Hawley, Adrian; Mudie, Stephen T.

    2015-01-28

    Type I collagen is the main structural component of skin, tendons, and skin products, such as leather. Understanding the mechanical performance of collagen fibrils is important for understanding the mechanical performance of the tissues that they make up, while the mechanical properties of bulk tissue are well characterized, less is known about the mechanical behavior of individual collagen fibrils. In this study, bovine pericardium is subjected to strain while small angle X-ray scattering (SAXS) patterns are recorded using synchrotron radiation. The change in d-spacing, which is a measure of fibril extension, and the change in fibril diameter are determined from SAXS. The tissue is strained 0.25 (25%) with a corresponding strain in the collagen fibrils of 0.045 observed. The ratio of collagen fibril width contraction to length extension, or the Poisson's ratio, is 2.1 ± 0.7 for a tissue strain from 0 to 0.25. This Poisson's ratio indicates that the volume of individual collagen fibrils decreases with increasing strain, which is quite unlike most engineering materials. This high Poisson's ratio of individual fibrils may contribute to high Poisson's ratio observed for tissues, contributing to some of the remarkable properties of collagen-based materials.

  10. Poisson's ratio of collagen fibrils measured by small angle X-ray scattering of strained bovine pericardium

    NASA Astrophysics Data System (ADS)

    Wells, Hannah C.; Sizeland, Katie H.; Kayed, Hanan R.; Kirby, Nigel; Hawley, Adrian; Mudie, Stephen T.; Haverkamp, Richard G.

    2015-01-01

    Type I collagen is the main structural component of skin, tendons, and skin products, such as leather. Understanding the mechanical performance of collagen fibrils is important for understanding the mechanical performance of the tissues that they make up, while the mechanical properties of bulk tissue are well characterized, less is known about the mechanical behavior of individual collagen fibrils. In this study, bovine pericardium is subjected to strain while small angle X-ray scattering (SAXS) patterns are recorded using synchrotron radiation. The change in d-spacing, which is a measure of fibril extension, and the change in fibril diameter are determined from SAXS. The tissue is strained 0.25 (25%) with a corresponding strain in the collagen fibrils of 0.045 observed. The ratio of collagen fibril width contraction to length extension, or the Poisson's ratio, is 2.1 ± 0.7 for a tissue strain from 0 to 0.25. This Poisson's ratio indicates that the volume of individual collagen fibrils decreases with increasing strain, which is quite unlike most engineering materials. This high Poisson's ratio of individual fibrils may contribute to high Poisson's ratio observed for tissues, contributing to some of the remarkable properties of collagen-based materials.

  11. Second-harmonic generation imaging of collagen fibers in myocardium for atrial fibrillation diagnosis

    NASA Astrophysics Data System (ADS)

    Tsai, Ming-Rung; Chiu, Yu-Wei; Lo, Men Tzung; Sun, Chi-Kuang

    2010-03-01

    Atrial fibrillation (AF) is the most common irregular heart rhythm and the mortality rate for patients with AF is approximately twice the mortality rate for patients with normal sinus rhythm (NSR). Some research has indicated that myocardial fibrosis plays an important role in predisposing patients to AF. Therefore, realizing the relationship between myocardial collagen fibrosis and AF is significant. Second-harmonic generation (SHG) is an optically nonlinear coherent process to image the collagen network. We perform SHG microscopic imaging of the collagen fibers in the human atrial myocardium. Utilizing the SHG images, we can identify the differences in morphology and the arrangement of collagen fibers between NSR and AF tissues. We also quantify the arrangement of the collagen fibers using Fourier transform images and calculating the values of angle entropy. We indicate that SHG imaging, a nondestructive and reproducible method to analyze the arrangement of collagen fibers, can provide explicit information about the relationship between myocardial fibrosis and AF.

  12. Three-dimensional imaging of collagen fibril organization in rat circumferential lamellar bone using a dual beam electron microscope reveals ordered and disordered sub-lamellar structures.

    PubMed

    Reznikov, Natalie; Almany-Magal, Rotem; Shahar, Ron; Weiner, Steve

    2013-02-01

    Lamellar bone is a major component of most mammalian skeletons. A prominent component of individual lamellae are parallel arrays of mineralized type I collagen fibrils, organized in a plywood like motif. Here we use a dual beam microscope and the serial surface view (SSV) method to investigate the three dimensional collagen organization of circumferential lamellar bone from rat tibiae after demineralization and osmium staining. Fast Fourier transform analysis is used to quantitatively identify the mean collagen array orientations and local collagen fibril dispersion. Based on collagen fibril array orientations and variations in fibril dispersion, we identify 3 distinct sub-lamellar structural motifs: a plywood-like fanning sub-lamella, a unidirectional sub-lamella and a disordered sub-lamella. We also show that the disordered sub-lamella is less mineralized than the other sub-lamellae. The hubs and junctions of the canalicular network, which connect radially oriented canaliculi, are intimately associated with the disordered sub-lamella. We also note considerable variations in the proportions of these 3 sub-lamellar structural elements among different lamellae. This new application of Serial Surface View opens the way to quantitatively compare lamellar bone from different sources, and to clarify the 3-dimensional structures of other bone types, as well as other biological structural materials. PMID:23153959

  13. A new model to simulate the elastic properties of mineralized collagen fibril.

    SciTech Connect

    Yuan, F.; Stock, S.R.; Haeffner, D.R.; Almer, J.D.; Dunand , D.C.; Brinson, K.

    2011-01-01

    Bone, because of its hierarchical composite structure, exhibits an excellent combination of stiffness and toughness, which is due substantially to the structural order and deformation at the smaller length scales. Here, we focus on the mineralized collagen fibril, consisting of hydroxyapatite plates with nanometric dimensions aligned within a protein matrix, and emphasize the relationship between the structure and elastic properties of a mineralized collagen fibril. We create two- and three-dimensional representative volume elements to represent the structure of the fibril and evaluate the importance of the parameters defining its structure and properties of the constituent mineral and collagen phase. Elastic stiffnesses are calculated by the finite element method and compared with experimental data obtained by synchrotron X-ray diffraction. The computational results match the experimental data well, and provide insight into the role of the phases and morphology on the elastic deformation characteristics. Also, the effects of water, imperfections in the mineral phase and mineral content outside the mineralized collagen fibril upon its elastic properties are discussed.

  14. A new model to simulate the elastic properties of mineralized collagen fibril

    SciTech Connect

    Yuan, F.; Stock, S.R.; Haeffner, D.R.; Almer, J.D.; Dunand, D.C.; Brinson, L.C.

    2012-05-02

    Bone, because of its hierarchical composite structure, exhibits an excellent combination of stiffness and toughness, which is due substantially to the structural order and deformation at the smaller length scales. Here, we focus on the mineralized collagen fibril, consisting of hydroxyapatite plates with nanometric dimensions aligned within a protein matrix, and emphasize the relationship between the structure and elastic properties of a mineralized collagen fibril. We create two- and three-dimensional representative volume elements to represent the structure of the fibril and evaluate the importance of the parameters defining its structure and properties of the constituent mineral and collagen phase. Elastic stiffnesses are calculated by the finite element method and compared with experimental data obtained by synchrotron X-ray diffraction. The computational results match the experimental data well, and provide insight into the role of the phases and morphology on the elastic deformation characteristics. Also, the effects of water, imperfections in the mineral phase and mineral content outside the mineralized collagen fibril upon its elastic properties are discussed.

  15. High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils.

    PubMed

    Watanabe-Nakayama, Takahiro; Itami, Masahiro; Kodera, Noriyuki; Ando, Toshio; Konno, Hiroki

    2016-01-01

    Bacterial collagenases involved in donor infection are widely applied in many fields due to their high activity and specificity; however, little is known regarding the mechanisms by which bacterial collagenases degrade insoluble collagen in host tissues. Using high-speed atomic force microscopy, we simultaneously visualized the hierarchical structure of collagen fibrils and the movement of a representative bacterial collagenase, Clostridium histolyticum type I collagenase (ColG), to determine the relationship between collagen structure and collagenase movement. Notably, ColG moved ~14.5 nm toward the collagen N terminus in ~3.8 s in a manner dependent on a catalytic zinc ion. While ColG was engaged, collagen molecules were not only degraded but also occasionally rearranged to thicken neighboring collagen fibrils. Importantly, we found a similarity of relationship between the enzyme-substrate interface structure and enzyme migration in collagen-collagenase and DNA-nuclease systems, which share a helical substrate structure, suggesting a common strategy in enzyme evolution. PMID:27373458

  16. High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils

    PubMed Central

    Watanabe-Nakayama, Takahiro; Itami, Masahiro; Kodera, Noriyuki; Ando, Toshio; Konno, Hiroki

    2016-01-01

    Bacterial collagenases involved in donor infection are widely applied in many fields due to their high activity and specificity; however, little is known regarding the mechanisms by which bacterial collagenases degrade insoluble collagen in host tissues. Using high-speed atomic force microscopy, we simultaneously visualized the hierarchical structure of collagen fibrils and the movement of a representative bacterial collagenase, Clostridium histolyticum type I collagenase (ColG), to determine the relationship between collagen structure and collagenase movement. Notably, ColG moved ~14.5 nm toward the collagen N terminus in ~3.8 s in a manner dependent on a catalytic zinc ion. While ColG was engaged, collagen molecules were not only degraded but also occasionally rearranged to thicken neighboring collagen fibrils. Importantly, we found a similarity of relationship between the enzyme-substrate interface structure and enzyme migration in collagen-collagenase and DNA-nuclease systems, which share a helical substrate structure, suggesting a common strategy in enzyme evolution. PMID:27373458

  17. Cross-linking connectivity in bone collagen fibrils: the COOH-terminal locus of free aldehyde

    NASA Technical Reports Server (NTRS)

    Otsubo, K.; Katz, E. P.; Mechanic, G. L.; Yamauchi, M.

    1992-01-01

    Quantitative analyses of the chemical state of the 16c residue of the alpha 1 chain of bone collagen were performed on samples from fetal (4-6-month embryo) and mature (2-3 year old) bovine animals. All of this residue could be accounted for in terms of three chemical states, in relative amounts which depended upon the age of the animal. Most of the residue was incorporated into either bifunctional or trifunctional cross-links. Some of it, however, was present as free aldehyde, and the content increased with maturation. This was established by isolating and characterizing the aldehyde-containing peptides generated by tryptic digestion of NaB3H4-reduced mature bone collagen. We have concluded that the connectivity of COOH-terminal cross-linking in bone collagen fibrils changes with maturation in the following way: at first, each 16c residue in each of the two alpha 1 chains of the collagen molecule is incorporated into a sheet-like pattern of intermolecular iminium cross-links, which stabilizes the young, nonmineralized fibril as a whole. In time, some of these labile cross-links maturate into pyridinoline while others dissociate back to their precursor form. The latter is likely due to changes in the molecular packing brought about by the mineralization of the collagen fibrils. The resultant reduction in cross-linking connectivity may provide a mechanism for enhancing certain mechanical characteristics of the skeleton of a mature animal.

  18. Acellular and cellular high-density, collagen-fibril constructs with suprafibrillar organization.

    PubMed

    Blum, Kevin M; Novak, Tyler; Watkins, Lauren; Neu, Corey P; Wallace, Joseph M; Bart, Zachary R; Voytik-Harbin, Sherry L

    2016-04-22

    Collagen is used extensively for tissue engineering due to its prevalence in connective tissues and its role in defining tissue biophysical and biological signalling properties. However, traditional collagen-based materials fashioned from atelocollagen and telocollagen have lacked collagen densities, multi-scale organization, mechanical integrity, and proteolytic resistance found within tissues in vivo. Here, highly interconnected low-density matrices of D-banded fibrils were created from collagen oligomers, which exhibit fibrillar as well as suprafibrillar assembly. Confined compression then was applied to controllably reduce the interstitial fluid while maintaining fibril integrity. More specifically, low-density (3.5 mg mL(-1)) oligomer matrices were densified to create collagen-fibril constructs with average concentrations of 12.25 mg mL(-1) and 24.5 mg mL(-1). Control and densified constructs exhibited nearly linear increases in ultimate stress, Young's modulus, and compressive modulus over the ranges of 65 to 213 kPa, 400 to 1.26 MPa, and 20 to 150 kPa, respectively. Densification also increased construct resistance to collagenase degradability. Finally, this process was amenable to creating high-density cellularized tissues; all constructs maintained high cell viability (at least 97%) immediately following compression as well as after 1 day and 7 days of culture. This method, which integrates the suprafibrillar assembly capacity of oligomers and controlled fluid reduction by confined compression, supports the rational and scalable design of a broad range of collagen-fibril materials and cell-encapsulated tissue constructs for tissue engineering applications. PMID:26902645

  19. Mechanisms of collagen fibril alignment in tendon injury: from tendon regeneration to artificial tendon.

    PubMed

    Torigoe, Kojun; Tanaka, Hirohito F; Yonenaga, Kazumichi; Ohkochi, Hiroki; Miyasaka, Muneo; Sato, Ryota; Kuzumaki, Toru; Yoshida, Kazuharu; Yoshida, Toshiko

    2011-12-01

    The process by which collagen fibrils are aligned following tendon injury remains unknown. Therefore, we analyzed the process of tendon regeneration by transmission electron microscopy, using a film model method. In mice, the Achilles tendon of medial head was transected. On day 3, after only the proximal end of the transected tendon was placed on film and kept in vivo, a translucent substance containing granules, called tendon gel, was secreted. On day 5, the granules assembled in a loose (L) layer, and coalesced tightly in a dense (D) layer, forming an L-D-L layered pattern. On day 10, granules showed high electron density in H layers, which developed into D-H-D layers on day 13. The distal end was placed on film to face the proximal end. On day 10, the tendon gel showed a D-H-D layer pattern. On day 11, mechanical stress from muscular constriction changed the tendon gel to aligned collagen fibrils (6 ± 2 nm in diameter). Thereafter, the diameter of the fibrils increased. Tendon gel harvested on day 5 or day 10 was pulled manually or by hanging weights (about 0.6 MPa). Aligned collagen fibrils (32 ± 7 nm in diameter) were created by traction using tendon gel harvested on day 10. PMID:21618275

  20. Investigation of the three-dimensional orientation of mineralized collagen fibrils in human lamellar bone using synchrotron X-ray phase nano-tomography.

    PubMed

    Varga, Peter; Pacureanu, Alexandra; Langer, Max; Suhonen, Heikki; Hesse, Bernhard; Grimal, Quentin; Cloetens, Peter; Raum, Kay; Peyrin, Françoise

    2013-09-01

    We investigate the three-dimensional (3-D) organization of mineralized collagen fibrils in human cortical bone based on synchrotron X-ray phase nano-tomography images. In lamellar bone the collagen fibrils are assumed to have a plywood-like arrangement, but due to experimental limitations the 3-D fibril structure has only been deduced from section surfaces so far and the findings have been controversial. Breakthroughs in synchrotron tomographic imaging have given access to direct 3-D information on the bone structure at the nanoscale level. Using an autocorrelation-based orientation measure we confirm that the fibrils are unidirectional in quasi-planes of sub-lamellae and find two specific dominant patterns, oscillating and twisted plywoods coexisting in a single osteon. Both patterns exhibit smooth orientation changes between adjacent quasi-planes. Moreover, we find that the periodic changes in collagen fibril orientation are independent of fluctuations in local mass density. These data improve our understanding of the lamellar arrangement in bone and allow more detailed investigations of structure-function relationships at this scale, providing templates for bio-inspired materials. The presented methodology can be applied to non-destructive 3-D characterization of the sub-micron scale structure of other natural and artificial mineralized biomaterials. PMID:23707503

  1. Influence of the mineral staggering on the elastic properties of the mineralized collagen fibril in lamellar bone.

    PubMed

    Vercher-Martínez, Ana; Giner, Eugenio; Arango, Camila; Fuenmayor, F Javier

    2015-02-01

    In this work, a three-dimensional finite element model of the staggered distribution of the mineral within the mineralized collagen fibril has been developed to characterize the lamellar bone elastic behavior at the sub-micro length scale. Minerals have been assumed to be embedded in a collagen matrix, and different degrees of mineralization have been considered allowing the growth of platelet-shaped minerals both in the axial and the transverse directions of the fibril, through the variation of the lateral space between platelets. We provide numerical values and trends for all the elastic constants of the mineralized collagen fibril as a function of the volume fraction of mineral. In our results, we verify the high influence of the mineral overlapping on the mechanical response of the fibril and we highlight that the lateral distance between crystals is relevant to the mechanical behavior of the fibril and not only the mineral overlapping in the axial direction. PMID:25498297

  2. The Relation Between Collagen Fibril Kinematics and Mechanical Properties in the Mitral Valve Anterior Leaflet

    SciTech Connect

    Liao,J.; Yang, L.; Grashow, J.; Sacks, M.

    2007-01-01

    We have recently demonstrated that the mitral valve anterior leaflet (MVAL) exhibited minimal hysteresis, no strain rate sensitivity, stress relaxation but not creep (Grashow et al., 2006, Ann Biomed Eng., 34(2), pp. 315-325; Grashow et al., 2006, Ann Biomed. Eng., 34(10), pp. 1509-1518). However, the underlying structural basis for this unique quasi-elastic mechanical behavior is presently unknown. As collagen is the major structural component of the MVAL, we investigated the relation between collagen fibril kinematics (rotation and stretch) and tissue-level mechanical properties in the MVAL under biaxial loading using small angle X-ray scattering. A novel device was developed and utilized to perform simultaneous measurements of tissue level forces and strain under a planar biaxial loading state. Collagen fibril D-period strain ({epsilon}{sub D}) and the fibrillar angular distribution were measured under equibiaxial tension, creep, and stress relaxation to a peak tension of 90 N/m. Results indicated that, under equibiaxial tension, collagen fibril straining did not initiate until the end of the nonlinear region of the tissue-level stress-strain curve. At higher tissue tension levels, {epsilon}{sub D} increased linearly with increasing tension. Changes in the angular distribution of the collagen fibrils mainly occurred in the tissue toe region. Using {epsilon}{sub D}, the tangent modulus of collagen fibrils was estimated to be 95.5{+-}25.5 MPa, which was {approx}27 times higher than the tissue tensile tangent modulus of 3.58{+-}1.83 MPa. In creep tests performed at 90 N/m equibiaxial tension for 60 min, both tissue strain and D remained constant with no observable changes over the test length. In contrast, in stress relaxation tests performed for 90 min {epsilon}{sub D} was found to rapidly decrease in the first 10 min followed by a slower decay rate for the remainder of the test. Using a single exponential model, the time constant for the reduction in collagen

  3. Influence of telopeptides, fibrils and crosslinking on physicochemical properties of type I collagen films.

    PubMed

    Walton, Robin S; Brand, David D; Czernuszka, Jan T

    2010-02-01

    Type I collagen is widely used in various different forms for research and commercial applications. Different forms of collagen may be classified according to their source, extraction method, crosslinking and resultant ultrastructure. In this study, afibrillar and reconstituted fibrillar films, derived from acid soluble and pepsin digested Type I collagen, were analysed using Lateral Force Microscopy (LFM), Fourier Transform Infra-Red Spectroscopy (FTIR), Differential Scanning Calorimetry (DSC) and enzymatic stability assays to asses the influence of telopeptides, fibrils and crosslinking. LFM proved to be a useful technique to confirm an afibrillar/fibrillar ultrastructure and to elucidate fibril diameters. FTIR has proved insensitive to ultrastructural differences involving telopeptides and fibrils. DSC results showed a significant increase in T(d) for crosslinked samples (+22-28 degrees C), and demonstrated that the thermal behaviour of hydrated, afibrillar films is more akin to reconstituted fibrillar films than monomeric solutions. The enzymatic stability assay has provided new evidence to show that afibrillar films of Type I collagen can be significantly more resistant to collagenase (by up to 3.5 times), than reconstituted fibrillar films, as a direct consequence of the different spatial arrangement of collagen molecules. A novel mechanism for this phenomenon is proposed and discussed. Additionally, the presence of telopeptide regions in afibrillar tropocollagen samples has been shown to increase resistance to collagenase by greater than 3.5 times compared to counterpart afibrillar atelocollagen samples. One-factor ANOVA analysis, with Fisher's LSD post-hoc test, confirms these key findings to be of statistical significance (P < 0.05). The profound physicochemical effects of collagen ultrastructure demonstrated in this study reiterates the need for comprehensive materials disclosure and classification when using these biomaterials. PMID:19851839

  4. Modification by UV radiation of the surface of thin films based on collagen extracted from fish scales.

    PubMed

    Sionkowska, Alina; Kozłowska, Justyna; Lazare, Sylvain

    2014-06-01

    Collagen was extracted from fish scales (Esox lucius) through demineralization process. Thin films by solvent evaporation from collagen extracted from fish scales were prepared. The surface of thin films made of fish scales collagen was modified by ultraviolet (UV)-irradiation with the wavelength λ = 254 nm. The amino acid composition of the Esox lucius scale collagen was analyzed before and after UV-irradiation by means of high-pressure liquid chromatography. The surface properties of films were investigated using the technique of atomic force microscopy (AFM) and by means of contact angle measurements allowing the calculation of surface free energy. Measurements of the contact angle for diiodomethane (D) and glycerol (G) on the surface of fish collagen films were made and surface free energy was calculated. The structure of collagen before and after UV-irradiation was studied using Fourier-transform infrared spectroscopy. It was found that after UV-irradiation the amount of all amino acids present in collagen molecule decreased. It was found also that the contact angle and the surface free energy were altered by UV-irradiation of collagen film. AFM showed that the surface roughness of collagen films was also altered by UV-irradiation. UV-irradiation caused the decrease of surface roughness due to photochemical processes, which occurred in the top layer of collagen film. The formation of collagen fibrils after solvent evaporation was observed using AFM. The diameter of collagen fibrils was bigger for irradiated collagen film than the diameter of collagen fibrils before UV-irradiation. PMID:24985207

  5. Interfibrillar shear stress is the loading mechanism of collagen fibrils in tendon

    PubMed Central

    Szczesny, Spencer E.; Elliott, Dawn M.

    2014-01-01

    Despite the critical role tendons play in transmitting loads throughout the musculoskeletal system, little is known about the microstructural mechanisms underlying their mechanical function. Of particular interest is whether collagen fibrils in tendon fascicles bear load independently or if load is transferred between fibrils through interfibrillar shear forces. We conducted multiscale experimental testing and developed a microstructural shear lag model to explicitly test whether interfibrillar shear load transfer is indeed the fibrillar loading mechanism in tendon. Experimental correlations between fascicle macroscale mechanics and microscale interfibrillar sliding suggest that fibrils are discontinuous and share load. Moreover, for the first time, we demonstrate that a shear lag model can replicate the fascicle macroscale mechanics as well as predict the microscale fibrillar deformations. Since interfibrillar shear stress is the fundamental loading mechanism assumed in the model, this result provides strong evidence that load is transferred between fibrils in tendon and possibly other aligned collagenous tissues. Conclusively establishing this fibrillar loading mechanism and identifying the involved structural components should help develop repair strategies for tissue degeneration and guide the design of tissue engineered replacements. PMID:24530560

  6. Homogenized stiffness matrices for mineralized collagen fibrils and lamellar bone using unit cell finite element models.

    PubMed

    Vercher, Ana; Giner, Eugenio; Arango, Camila; Tarancón, José E; Fuenmayor, F Javier

    2014-04-01

    Mineralized collagen fibrils have been usually analyzed like a two-phase composite material where crystals are considered as platelets that constitute the reinforcement phase. Different models have been used to describe the elastic behavior of the material. In this work, it is shown that when Halpin-Tsai equations are applied to estimate elastic constants from typical constituent properties, not all crystal dimensions yield a model that satisfy thermodynamic restrictions. We provide the ranges of platelet dimensions that lead to positive definite stiffness matrices. On the other hand, a finite element model of a mineralized collagen fibril unit cell under periodic boundary conditions is analyzed. By applying six canonical load cases, homogenized stiffness matrices are numerically calculated. Results show a monoclinic behavior of the mineralized collagen fibril. In addition, a 5-layer lamellar structure is also considered where crystals rotate in adjacent layers of a lamella. The stiffness matrix of each layer is calculated applying Lekhnitskii transformations, and a new finite element model under periodic boundary conditions is analyzed to calculate the homogenized 3D anisotropic stiffness matrix of a unit cell of lamellar bone. Results are compared with the rule-of-mixtures showing in general good agreement. PMID:23793930

  7. Epitaxially Grown Collagen Fibrils Reveal Diversity in Contact Guidance Behavior among Cancer Cells

    PubMed Central

    2015-01-01

    Invasion of cancer cells into the surrounding tissue is an important step during cancer progression and is driven by cell migration. Cell migration can be random, but often it is directed by various cues such as aligned fibers composed of extracellular matrix (ECM), a process called contact guidance. During contact guidance, aligned fibers bias migration along the long axis of the fibers. These aligned fibers of ECM are commonly composed of type I collagen, an abundant structural protein around tumors. In this paper, we epitaxially grew several different patterns of organized type I collagen on mica and compared the morphology and contact guidance behavior of two invasive breast cancer cell lines (MDA-MB-231 and MTLn3 cells). Others have shown that these cells randomly migrate in qualitatively different ways. MDA-MB-231 cells exert large traction forces, tightly adhere to the ECM, and migrate with spindle-shaped morphology and thus adopt a mesenchymal mode of migration. MTLn3 cells exert small traction forces, loosely adhere to the ECM, and migrate with a more rounded morphology and thus adopt an amoeboid mode of migration. As the degree of alignment of type I collagen fibrils increases, cells become more elongated and engage in more directed contact guidance. MDA-MB-231 cells perceive the directional signal of highly aligned type I collagen fibrils with high fidelity, elongating to large extents and migrating directionally. Interestingly, behavior in MTLn3 cells differs. While highly aligned type I collagen fibril patterns facilitate spreading and random migration of MTLn3 cells, they do not support elongation or directed migration. Thus, different contact guidance cues bias cell migration differently and the fidelity of contact guidance is cell type dependent, suggesting that ECM alignment is a permissive cue for contact guidance, but requires a cell to have certain properties to interpret that cue. PMID:25531276

  8. Biochemical characterisation and assessment of fibril-forming ability of collagens extracted from Bester sturgeon Huso huso × Acipenser ruthenus.

    PubMed

    Zhang, Xi; Ookawa, Mika; Tan, Yongkai; Ura, Kazuhiro; Adachi, Shinji; Takagi, Yasuaki

    2014-10-01

    Collagens purified from Bester sturgeon organs were characterised biochemically, and their fibril-forming abilities and fibril morphologies formed in vitro clarified. Yields of collagens were 2.1%, 11.9%, 0.4%, 18.1%, 0.4%, 0.8% and 0.03% (collagen dry weight/tissue wet weight) from scales, skin, muscle, swim bladder, digestive tract, notochord and snout cartilage, respectively. Using SDS-PAGE and amino acid composition analyses, collagens from scales, skin, muscle, the swim bladder and digestive tract were characterised as type I, and collagens from the notochord and snout cartilage as type II. Denaturation temperatures of the collagens, measured using circular dichroism, were 29.6, 26.8, 29.0, 32.9, 31.6 and 36.3 °C in scales, skin, muscle, swim bladder, digestive tract, and notochord, respectively. For fibril formation, swim bladder and skin collagen showed a more rapid rate of increase in turbidity, a shorter time to attain the maximum turbidity, and formed thicker fibrils compared with porcine tendon type I collagen. PMID:24799243

  9. Tectorins crosslink type II collagen fibrils and connect the tectorial membrane to the spiral limbus.

    PubMed

    Andrade, Leonardo R; Salles, Felipe T; Grati, M'hamed; Manor, Uri; Kachar, Bechara

    2016-05-01

    All inner ear organs possess extracellular matrix appendices over the sensory epithelia that are crucial for their proper function. The tectorial membrane (TM) is a gelatinous acellular membrane located above the hearing sensory epithelium and is composed mostly of type II collagen, and α and β tectorins. TM molecules self-assemble in the endolymph fluid environment, interacting medially with the spiral limbus and distally with the outer hair cell stereocilia. Here, we used immunogold labeling in freeze-substituted mouse cochleae to assess the fine localization of both tectorins in distinct TM regions. We observed that the TM adheres to the spiral limbus through a dense thin matrix enriched in α- and β-tectorin, both likely bound to the membranes of interdental cells. Freeze-etching images revealed that type II collagen fibrils were crosslinked by short thin filaments (4±1.5nm, width), resembling another collagen type protein, or chains of globular elements (15±3.2nm, diameter). Gold-particles for both tectorins also localized adjacent to the type II collagen fibrils, suggesting that these globules might be composed essentially of α- and β-tectorins. Finally, the presence of gold-particles at the TM lower side suggests that the outer hair cell stereocilia membrane has a molecular partner to tectorins, probably stereocilin, allowing the physical connection between the TM and the organ of Corti. PMID:26806019

  10. Small-Angle X-ray Scattering Study of Intramuscular Fish Bone: Collagen Fibril Superstructure Determined from Equidistant Meridional Reflections

    SciTech Connect

    Burger,C.; Zhou, H.; Sics, I.; Hsiao, B.; Chu, B.; Graham, L.; Glimcher, M.

    2008-01-01

    New insights into the bone collagen fibril superstructure have been obtained by novel small-angle X-ray scattering analysis. The analysis was carried out on the small-angle equidistant meridional reflections resulting from the periodic structure of collagen fibrils in their axial direction. Conventional two-dimensional analysis is difficult because of the large discrepancy of longitudinal and lateral length scales for individual fibrils, as well as their preferred orientation. The new approach represents an unapproximated analysis of the equidistant meridional reflections, which takes the exact separation of preferred orientation and fibril size effects into account. The analytical results (e.g. axial period, fibril diameter etc.) agree well with the parameters obtained from transmission electron microscopy.

  11. Modeling the collagen fibril network of biological tissues as a nonlinearly elastic material using a continuous volume fraction distribution function

    PubMed Central

    Shirazi, Reza; Vena, Pasquale; Sah, Robert L.; Klisch, Stephen M.

    2012-01-01

    Despite distinct mechanical functions, biological soft tissues have a common microstructure in which a ground matrix is reinforced by a collagen fibril network. The microstructural properties of the collagen network contribute to continuum mechanical tissue properties that are strongly anisotropic with tensile-compressive asymmetry. In this study, a novel approach based on a continuous distribution of collagen fibril volume fractions is developed to model fibril reinforced soft tissues as a nonlinearly elastic and anisotropic material. Compared with other approaches that use a normalized number of fibrils for the definition of the distribution function, this representation is based on a distribution parameter (i.e. volume fraction) that is commonly measured experimentally while also incorporating pre-stress of the collagen fibril network in a tissue natural configuration. After motivating the form of the collagen strain energy function, examples are provided for two volume fraction distribution functions. Consequently, collagen second-Piola Kirchhoff stress and elasticity tensors are derived, first in general form and then specifically for a model that may be used for immature bovine articular cartilage. It is shown that the proposed strain energy is a convex function of the deformation gradient tensor and, thus, is suitable for the formation of a polyconvex tissue strain energy function. PMID:23390357

  12. Influence of cross-link structure, density and mechanical properties in the mesoscale deformation mechanisms of collagen fibrils

    PubMed Central

    Depalle, Baptiste; Qin, Zhao; Shefelbine, Sandra J.; Buehler, Markus J.

    2015-01-01

    Collagen is a ubiquitous protein with remarkable mechanical properties. It is highly elastic, shows large fracture strength and enables substantial energy dissipation during deformation. Most of the connective tissue in humans consists of collagen fibrils composed of a staggered array of tropocollagen molecules, which are connected by intermolecular cross-links. In this study, we report a three-dimensional coarse-grained model of collagen and analyze the influence of enzymatic cross-links on the mechanics of collagen fibrils. Two representatives immature and mature cross-links are implemented in the mesoscale model using a bottom-up approach. By varying the number, type and mechanical properties of cross-links in the fibrils and performing tensile test on the models, we systematically investigate the deformation mechanisms of cross-linked collagen fibrils. We find that cross-linked fibrils exhibit a three phase behavior, which agrees closer with experimental results than what was obtained using previous models. The fibril mechanical response is characterized by: (i) an initial elastic deformation corresponding to the collagen molecule uncoiling, (ii) a linear regime dominated by molecule sliding and (iii) the second stiffer elastic regime related to the stretching of the backbone of the tropocollagen molecules until the fibril ruptures. Our results suggest that both cross-link density and type dictate the stiffness of large deformation regime by increasing the number of interconnected molecules while cross-links mechanical properties determine the failure strain and strength of the fibril. These findings reveal that cross-links play an essential role in creating an interconnected fibrillar material of tunable toughness and strength. PMID:25153614

  13. Effect of cyclic loading on the nanoscale deformation of hydroxyapatite and collagen fibrils in bovine bone.

    PubMed

    Singhal, Anjali; Stock, Stuart R; Almer, Jonathan D; Dunand, David C

    2014-06-01

    Cyclic compressive loading tests were carried out on bovine femoral bones at body temperature (37 °C), with varying mean stresses (-55 to -80 MPa) and loading frequencies (0.5-5 Hz). At various times, the cyclic loading was interrupted to carry out high-energy X-ray scattering measurements of the internal strains developing in the hydroxyapatite (HAP) platelets and the collagen fibrils. The residual strains upon unloading were always tensile in the HAP and compressive in the fibrils, and each increases in magnitude with loading cycles, which can be explained from damage at the HAP–collagen interface and accumulation of plastic deformation within the collagen phase. The samples tested at a higher mean stress and stress amplitude, and at lower loading frequencies exhibit greater plastic deformation and damage accumulation, which is attributed to greater contribution of creep. Synchrotron microcomputed tomography of some of the specimens showed that cracks are produced during cyclic loading and that they mostly occur concentric with Haversian canals. PMID:23958833

  14. Modelling the mechanics of partially mineralized collagen fibrils, fibres and tissue

    PubMed Central

    Liu, Yanxin; Thomopoulos, Stavros; Chen, Changqing; Birman, Victor; Buehler, Markus J.; Genin, Guy M.

    2014-01-01

    Progressive stiffening of collagen tissue by bioapatite mineral is important physiologically, but the details of this stiffening are uncertain. Unresolved questions about the details of the accommodation of bioapatite within and upon collagen's hierarchical structure have posed a central hurdle, but recent microscopy data resolve several major questions. These data suggest how collagen accommodates bioapatite at the lowest relevant hierarchical level (collagen fibrils), and suggest several possibilities for the progressive accommodation of bioapatite at higher hierarchical length scales (fibres and tissue). We developed approximations for the stiffening of collagen across spatial hierarchies based upon these data, and connected models across hierarchies levels to estimate mineralization-dependent tissue-level mechanics. In the five possible sequences of mineralization studied, percolation of the bioapatite phase proved to be an important determinant of the degree of stiffening by bioapatite. The models were applied to study one important instance of partially mineralized tissue, which occurs at the attachment of tendon to bone. All sequences of mineralization considered reproduced experimental observations of a region of tissue between tendon and bone that is more compliant than either tendon or bone, but the size and nature of this region depended strongly upon the sequence of mineralization. These models and observations have implications for engineered tissue scaffolds at the attachment of tendon to bone, bone development and graded biomimetic attachment of dissimilar hierarchical materials in general. PMID:24352669

  15. Synchrotron radiation x-ray scattering in the early stages of in vitro collagen fibril formation

    SciTech Connect

    Suarez, G.; Oronsky, A.L.; Bordas, J.; Koch, M.H.

    1985-07-01

    The time course of in vitro collagen fibril formation was monitored by synchrotron radiation x-ray scattering. Collagen polymerization was induced by a temperature jump from 4 degrees C to 32 degrees C and the solution scattering pattern was recorded continuously with a time resolution of a few seconds. The scattered intensity increased as soon as the final temperature was attained, without discernible lag phase, when the collagen concentration was about 0.56 mg/ml, whereas turbidimetric measurements revealed a lag phase of ca. 2.5 min at 1.05 mg/ml. A direct correlation was found between the temperature and both the rate of formation and the total amount of early aggregates. The formation of these aggregates was only partially reversed by lowering the temperature, except when the collagen had been prevented from forming Schiff base-mediated intermolecular crosslinks by sodium borohydride reduction. In this case, formation of aggregates was completely reversible. The authors conclude that the aggregates that appear to correspond to the subfibrils proposed on the basis of independent methods are formed and simultaneously crosslinked in the early phases of in vitro collagen self-assembly.

  16. Agent-based modeling traction force mediated compaction of cell-populated collagen gels using physically realistic fibril mechanics.

    PubMed

    Reinhardt, James W; Gooch, Keith J

    2014-02-01

    Agent-based modeling was used to model collagen fibrils, composed of a string of nodes serially connected by links that act as Hookean springs. Bending mechanics are implemented as torsional springs that act upon each set of three serially connected nodes as a linear function of angular deflection about the central node. These fibrils were evaluated under conditions that simulated axial extension, simple three-point bending and an end-loaded cantilever. The deformation of fibrils under axial loading varied <0.001% from the analytical solution for linearly elastic fibrils. For fibrils between 100 μm and 200 μm in length experiencing small deflections, differences between simulated deflections and their analytical solutions were <1% for fibrils experiencing three-point bending and <7% for fibrils experiencing cantilever bending. When these new rules for fibril mechanics were introduced into a model that allowed for cross-linking of fibrils to form a network and the application of cell traction force, the fibrous network underwent macroscopic compaction and aligned between cells. Further, fibril density increased between cells to a greater extent than that observed macroscopically and appeared similar to matrical tracks that have been observed experimentally in cell-populated collagen gels. This behavior is consistent with observations in previous versions of the model that did not allow for the physically realistic simulation of fibril mechanics. The significance of the torsional spring constant value was then explored to determine its impact on remodeling of the simulated fibrous network. Although a stronger torsional spring constant reduced the degree of quantitative remodeling that occurred, the inclusion of torsional springs in the model was not necessary for the model to reproduce key qualitative aspects of remodeling, indicating that the presence of Hookean springs is essential for this behavior. These results suggest that traction force mediated matrix

  17. Characterization of the viscoelastic behavior of a simplified collagen micro-fibril based on molecular dynamics simulations.

    PubMed

    Ghodsi, Hossein; Darvish, Kurosh

    2016-10-01

    Collagen fibril is a major component of connective tissues such as bone, tendon, blood vessels, and skin. The mechanical properties of this highly hierarchical structure are greatly influenced by the presence of covalent cross-links between individual collagen molecules. This study investigates the viscoelastic behavior of a collagen lysine-lysine cross-link based on creep simulations with applied forces in the range or 10 to 2000pN using steered molecular dynamics (SMD). The viscoelastic model of the cross-link was combined with a system composed by two segments of adjacent collagen molecules hence representing a reduced viscoelastic model for a simplified micro-fibril. It was found that the collagen micro-fibril assembly had a steady-state Young׳s modulus ranging from 2.24 to 3.27GPa, which is in agreement with reported experimental measurements. The propagation of longitudinal force wave along the molecule was implemented by adding a delay element to the model. The force wave speed was found to be correlated with the speed of one-dimensional elastic waves in rods. The presented reduced model with three degrees of freedom can serve as a building block for developing models of the next level of hierarchy, i.e., a collagen fibril. PMID:27341288

  18. Molecular and intermolecular effects in collagen fibril mechanics: a multiscale analytical model compared with atomistic and experimental studies.

    PubMed

    Marino, Michele

    2016-02-01

    Both atomistic and experimental studies reveal the dependence of collagen fibril mechanics on biochemical and biophysical features such as, for instance, cross-link density, water content and protein sequence. In order to move toward a multiscale structural description of biological tissues, a novel analytical model for collagen fibril mechanics is herein presented. The model is based on a multiscale approach that incorporates and couples: thermal fluctuations in collagen molecules; the uncoiling of collagen triple helix; the stretching of molecular backbone; the straightening of the telopeptide in which covalent cross-links form; slip-pulse mechanisms due to the rupture of intermolecular weak bonds; molecular interstrand delamination due to the rupture of intramolecular weak bonds; the rupture of covalent bonds within molecular strands. The effectiveness of the proposed approach is verified by comparison with available atomistic results and experimental data, highlighting the importance of cross-link density in tuning collagen fibril mechanics. The typical three-region shape and hysteresis behavior of fibril constitutive response, as well as the transition from a yielding-like to a brittle-like behavior, are recovered with a special insight on the underlying nanoscale mechanisms. The model is based on parameters with a clear biophysical and biochemical meaning, resulting in a promising tool for analyzing the effect of pathological or pharmacological-induced histochemical alterations on the functional mechanical response of collagenous tissues. PMID:26220454

  19. Elemental distribution analysis of type I collagen fibrils in tilapia fish scale with energy-filtered transmission electron microscope.

    PubMed

    Okuda, Mitsuhiro; Takeguchi, Masaki; Tagaya, Motohiro; Tonegawa, Toru; Hashimoto, Ayako; Hanagata, Nobutaka; Ikoma, Toshiyuki

    2009-01-01

    Elemental distribution of calcium, phosphorus, oxygen, and carbon in a single collagen fibril obtained from tilapia fish scales was identified with an electron energy-loss spectroscopy and an energy-filtered transmission electron microscopy, for the first time. The carbon intensity profile of the single collagen fibril showed the specific D-periodic pattern at 67 nm of type I collagen fibrils. The calcium L(2,3)-edge and oxygen K-edge peak positions were detected at 347/350 eV and 137 eV, respectively, and these positions were identical to those of hydroxyapatite. Calcium, phosphorus, and oxygen were present in the hole zones as the amorphous phase, while carbon was present in the overlap zone. Our results indicated that the hole zones preferentially attract calcium and phosphate ions and thus serve as possible nucleation sites for mineralization. PMID:19419879

  20. Observations on the different substrate behavior of tropocollagen molecules in solution and intermolecularly cross-linked tropocollagen within insoluble polymeric collagen fibrils.

    PubMed Central

    Steven, F S

    1976-01-01

    Bacterial collagenase was used to compare the extent of digestion of tropocollagen monomers in solution and in reconstituted fibrils with that of tropocollagen molecules intermolecularly cross-linked within insoluble polymeric collagen fibrils obtained from mature tendons at given time-intervals. The extent of digestion of tropocollagen monomers in solution was directly proportional to the enzyme concentration (a range of enzyme substrate molar ratios 1:200 to 1:10 was used). The extent of digestion of polymeric collagen was followed by measuring the solubilization of fluorescent peptides from fluorescent-labelled insoluble polymeric collagen fibrils. The extent of digestion of tropocollagen within polymeric collagen was linear over a very small range of enzyme concentrations, when the enzyme/substrate ratio in the reaction mixture was less than 1:400 on a molecular basis. The behavior of tropocollagen in the form of reconstituted collagen fibrils, which had been matured at 37 degrees C for 8 weeks, was intermediate between the behaviour of solutions of tropocollagen and insoluble polymeric collagen fibrils. The significance of the results is discussed in terms of the structure of polymeric collagen fibrils and the protection against enzymic attack provided by tropocollagen molecules on the circumference of the fibril. The results suggest that assays of collagenase activities based on tropocollagen as substrate cannot be directly related to the ability of these enzymes to degrade mature insoluble collagen fibrils. PMID:180984

  1. Corneal Opacity in Lumican-Null Mice: Defects in Collagen Fibril Structure and Packing in the Posterior Stroma

    PubMed Central

    Chakravarti, Shukti; Petroll, W. Matthew; Hassell, John R.; Jester, James V.; Lass, Jonathan H.; Paul, Jennifer; Birk, David E.

    2015-01-01

    Purpose Gene targeted lumican-null mutants (lumtm1sc/lumtm1sc) have cloudy corneas with abnormally thick collagen fibrils. The purpose of the present study was to analyze the loss of transparency quantitatively and to define the associated corneal collagen fibril and stromal defects. Methods Backscattering of light, a function of corneal haze and opacification, was determined regionally using in vivo confocal microscopy in lumican-deficient and wild-type control mice. Fibril organization and structure were analyzed using transmission electron microscopy. Biochemical approaches were used to quantify glycosaminoglycan contents. Lumican distribution in the cornea was elucidated immunohistochemically. Results Compared with control stromas, lumican-deficient stromas displayed a threefold increase in backscattered light with maximal increase confined to the posterior stroma. Confocal microscopy through-focusing (CMTF) measurement profiles also indicated a 40% reduction in stromal thickness in the lumican-null mice. Transmission electron microscopy indicated significant collagen fibril abnormalities in the posterior stroma, with the anterior stroma remaining relatively unremarkable. The lumican-deficient posterior stroma displayed a pronounced increase in fibril diameter, large fibril aggregates, altered fibril packing, and poor lamellar organization. Immunostaining of wild-type corneas demonstrated high concentrations of lumican in the posterior stroma. Biochemical assessment of keratan sulfate (KS) content of whole eyes revealed a 25% reduction in KS content in the lumican-deficient mice. Conclusions The structural defects and maximum backscattering of light clearly localized to the posterior stroma of lumican-deficient mice. In normal mice, an enrichment of lumican was observed in the posterior stroma compared with that in the anterior stroma. Taken together, these observations indicate a key role for lumican in the posterior stroma in maintaining normal fibril

  2. A possible role of collagen fibrils in the process of calcification observed in the capsule of the pineal gland in aging rats.

    PubMed

    Humbert, W; Cuisinier, F; Voegel, J C; Pévet, P

    1997-06-01

    The relationship between collagen fibrils and calcified concretions exclusively appearing in the pineal gland of adult/aging rats has been investigated. Deposits of lanthanum, which replace calcium ions are distributed along collagen fibrils with a repeating period of about 70 nm. Calcium has been detected histochemically between collagen bundles surrounding extracellular concretions by means of the pyroantimonate method and by X-ray microanalysis. It is associated with phosphorus. The data presented here suggest that collagen fibrils are involved in the genesis and growth of extracellular concretions located in the connective tissue surrounding the pineal gland of aging rats. PMID:9134857

  3. Pseudo-hyperelastic model of tendon hysteresis from adaptive recruitment of collagen type I fibrils.

    PubMed

    Ciarletta, Pasquale; Dario, Paolo; Micera, Silvestro

    2008-02-01

    Understanding the functional relationship between the viscoelasticity and the morphology of soft collagenous tissues is fundamental for many applications in bioengineering science. This work presents a pseudo-hyperelastic constitutive theory aiming at describing the time-dependant hysteretic response of tendons subjected to uniaxial tensile loads. A macroscopic tendon is modeled as a composite homogeneous tissue with the anisotropic reinforcement of collagen type I fibrils. The tissue microstructure is considered as an adaptive network of fibrillar units connected in temporary junctions. The processes of breakage and reformation of active fibrils are thermally activated, and are occurring at random times. An internal softening variable and a dissipation energy function account for the adaptive arrangement of the fibrillar network in the pseudo-hyperelastic model. Cyclic uniaxial tensile tests have been performed in vitro on porcine flexor digital tendons. The theoretical predictions fit accurately the experimental stress-strain data both for the loading and the unloading processes. The hysteresis behavior reflects the improvement in the efficiency and performance of the motion of the muscle-tendon unit at high strain rates. The results of the model demonstrate the microstructural importance of proteoglycans in determining the functional viscoelastic adaptability of the macroscopic tendon. PMID:17997481

  4. The Impact of Collagen Fibril Polarity on Second Harmonic Generation Microscopy.

    PubMed

    Couture, Charles-André; Bancelin, Stéphane; Van der Kolk, Jarno; Popov, Konstantin; Rivard, Maxime; Légaré, Katherine; Martel, Gabrielle; Richard, Hélène; Brown, Cameron; Laverty, Sheila; Ramunno, Lora; Légaré, François

    2015-12-15

    In this work, we report the implementation of interferometric second harmonic generation (SHG) microscopy with femtosecond pulses. As a proof of concept, we imaged the phase distribution of SHG signal from the complex collagen architecture of juvenile equine growth cartilage. The results are analyzed in respect to numerical simulations to extract the relative orientation of collagen fibrils within the tissue. Our results reveal large domains of constant phase together with regions of quasi-random phase, which are correlated to respectively high- and low-intensity regions in the standard SHG images. A comparison with polarization-resolved SHG highlights the crucial role of relative fibril polarity in determining the SHG signal intensity. Indeed, it appears that even a well-organized noncentrosymmetric structure emits low SHG signal intensity if it has no predominant local polarity. This work illustrates how the complex architecture of noncentrosymmetric scatterers at the nanoscale governs the coherent building of SHG signal within the focal volume and is a key advance toward a complete understanding of the structural origin of SHG signals from tissues. PMID:26682809

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

    PubMed

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

    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 12weeks 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. PMID:27523994

  6. Purification, characterization and cloning of tensilin, the collagen-fibril binding and tissue-stiffening factor from Cucumaria frondosa dermis.

    PubMed

    Tipper, Jennifer P; Lyons-Levy, Gillian; Atkinson, Mark A L; Trotter, John A

    2002-12-01

    The inner dermis of the sea cucumber, Cucumaria frondosa, is a mutable collagenous tissue characterized by rapid and reversible changes in its mechanical properties regulated by one or more protein effectors that are released from neurosecretory cells. One such effector, tensilin, is a collagen-fibril binding protein, named for its ability to induce dermis stiffening. Tensilin was purified using an affinity column constructed from C. frondosa collagen-fibrils. The protein migrates as a single band on SDS-PAGE (Mr approximately 33 kDa) and has an isoelectric point of 5.8. Equilibrium sedimentation experiments suggest a molecular mass of approximately 28.5-29.4 kDa. Carbohydrate analysis of tensilin revealed no measurable sugar content. The molar amount of tensilin was determined to be 0.38% that of collagen and 47% that of stiparin, a constitutive matrix glycoprotein. A full-length cDNA clone for tensilin was obtained from a C. frondosa inner dermis cDNA expression library. Predicted properties derived from the deduced peptide sequence were in agreement with those of the native protein. A noted feature of tensilin's deduced peptide sequence, particularly in its N-terminal domain, is its homology to tissue inhibitor of metalloproteinases. Tensilin's C-terminal tail has no known homology to other proteins but contains a putative collagen-fibril binding site. PMID:12524049

  7. Surface Chemistry of Nanoscale Mineralized Collagen Regulates Periodontal Ligament Stem Cell Fate.

    PubMed

    Fu, Yu; Liu, Shuai; Cui, Sheng-Jie; Kou, Xiao-Xing; Wang, Xue-Dong; Liu, Xiao-Mo; Sun, Yue; Wang, Gao-Nan; Liu, Yan; Zhou, Yan-Heng

    2016-06-29

    The interplay between stem cells and their extracellular microenvironment is of critical importance to the stem cell-based therapeutics in regenerative medicine. Mineralized collagen is the main component of bone extracellular matrix, but the effect of interfacial properties of mineralized collagen on subsequent cellular behaviors is unclear. This study examined the role of surface chemistry of nanoscale mineralized collagen on human periodontal ligament stem cell (hPDLSC) fate decisions. The intrafibrillarly mineralized collagen (IMC), fabricated by a biomimetic bottom-up approach, showed a bonelike hierarchy with nanohydroxyapatites (HAs) periodically embedded within fibrils. The infrared spectrum of the IMC showed the presence of phosphate, carbonate, amide I and II bands; and infrared mapping displayed uniform and higher spatial distribution of mineralization in the IMC. However, the distribution of the phosphate group differed far from that of the amide I group in the extrafibrillarly mineralized collagen (EMC), in which flowerlike HA clusters randomly depositing around the surface of the fibrils. Moreover, a large quantity of extrafibrillar HAs covered up the C═O stretch and N-H in-plane bend, resulting in substantial reduction of amide I and II bands. Cell experiments demonstrated that the hPDLSCs seeded on the IMC exhibited a highly branched, osteoblast-like polygonal shape with extended pseudopodia and thick stress fiber formation; while cells on the EMC displayed a spindle shape with less branch points and thin actin fibril formation. Furthermore, the biocompatibility of EMC was much lower than that of IMC. Interestingly, even without osteogenic induction, mRNA levels of major osteogenic differentiation genes were highly expressed in the IMC during cultivation time. These data suggest that the IMC with a similar nanotopography and surface chemistry to natural mineralized collagen directs hPDLSCs toward osteoblast differentiation, providing a promising

  8. A Novel 3D Fibril Force Assay Implicates Src in Tumor Cell Force Generation in Collagen Networks

    PubMed Central

    Polackwich, Robert J.; Koch, Daniel; Arevalo, Richard; Miermont, Anne M.; Jee, Kathleen J.; Lazar, John; Urbach, Jeffrey; Mueller, Susette C.; McAllister, Ryan G.

    2013-01-01

    New insight into the biomechanics of cancer cell motility in 3D extracellular matrix (ECM) environments would significantly enhance our understanding of aggressive cancers and help identify new targets for intervention. While several methods for measuring the forces involved in cell-matrix interactions have been developed, previous to this study none have been able to measure forces in a fibrillar environment. We have developed a novel assay for simultaneously measuring cell mechanotransduction and motility in 3D fibrillar environments. The assay consists of a controlled-density fibrillar collagen gel atop a controlled-stiffness polyacrylamide (PAA) surface. Forces generated by living cells and their migration in the 3D collagen gel were measured with the 3D motion of tracer beads within the PAA layer. Here, this 3D fibril force assay is used to study the role of the invasion-associated protein kinase Src in mechanotransduction and motility. Src expression and activation are linked with proliferation, invasion, and metastasis, and have been shown to be required in 2D for invadopodia membranes to direct and mediate invasion. Breast cancer cell line MDA-MD-231 was stably transfected with GFP-tagged constitutively active Src or wild-type Src. In 3D fibrillar collagen matrices we found that, relative to wild-type Src, constitutively active Src: 1) increased the strength of cell-induced forces on the ECM, 2) did not significantly change migration speed, and 3) increased both the duration and the length, but not the number, of long membrane protrusions. Taken together, these results support the hypothesis that Src controls invasion by controlling the ability of the cell to form long lasting cellular protrusions to enable penetration through tissue barriers, in addition to its role in promoting invadopodia matrix-degrading activity. PMID:23536784

  9. Interleukin-4 Receptor α Signaling in Myeloid Cells Controls Collagen Fibril Assembly in Skin Repair.

    PubMed

    Knipper, Johanna A; Willenborg, Sebastian; Brinckmann, Jürgen; Bloch, Wilhelm; Maaß, Tobias; Wagener, Raimund; Krieg, Thomas; Sutherland, Tara; Munitz, Ariel; Rothenberg, Marc E; Niehoff, Anja; Richardson, Rebecca; Hammerschmidt, Matthias; Allen, Judith E; Eming, Sabine A

    2015-10-20

    Activation of the immune response during injury is a critical early event that determines whether the outcome of tissue restoration is regeneration or replacement of the damaged tissue with a scar. The mechanisms by which immune signals control these fundamentally different regenerative pathways are largely unknown. We have demonstrated that, during skin repair in mice, interleukin-4 receptor α (IL-4Rα)-dependent macrophage activation controlled collagen fibril assembly and that this process was important for effective repair while having adverse pro-fibrotic effects. We identified Relm-α as one important player in the pathway from IL-4Rα signaling in macrophages to the induction of lysyl hydroxylase 2 (LH2), an enzyme that directs persistent pro-fibrotic collagen cross-links, in fibroblasts. Notably, Relm-β induced LH2 in human fibroblasts, and expression of both factors was increased in lipodermatosclerosis, a condition of excessive human skin fibrosis. Collectively, our findings provide mechanistic insights into the link between type 2 immunity and initiation of pro-fibrotic pathways. PMID:26474656

  10. Contribution of Long Fibrils and Peptides to Surface and Foaming Behavior of Soy Protein Fibril System.

    PubMed

    Wan, Zhili; Yang, Xiaoquan; Sagis, Leonard M C

    2016-08-16

    When soy glycinin (11S) is heated for a prolonged time at pH 2 (20 h at 85 °C), a mixture is formed consisting of long semiflexible 11S fibrils and small peptides. The surface and foaming properties of this mixture were investigated at different pHs, and compared to the behavior of pure fibrils and pure peptides, to determine the individual contributions of these two factions to the behavior of the mixture. The adsorption of these three systems at air-water interfaces and the resulting surface rheological properties were studied by combining drop shape analysis tensiometry, ellipsometry, and surface large amplitude oscillatory dilatational (LAOD) rheology. Lissajous plots of surface pressure versus deformation were used to analyze the surface rheological response in terms of interfacial microstructure. Our results show that the adsorption kinetics, dilatational rheological properties, and the foaming behavior of the mixture were mainly dominated by the small peptides in the fibril system. Compared to pH 2, the fibril mixture at pH 5 and 7 provides much better foam stability and appears to be a very promising protein material to make stable foams, even at low protein concentration (0.1 wt %). The presence of fibril clusters and peptide aggregates at pH 5 and 7 contributed to foam stability of the mixture. In contrast, pure fibril formed an interface with a highly pH-responsive adsorption and rheological behavior, and the foamability and foam stability of the pure fibrils were very poor. PMID:27452662

  11. Surfaces Self-Assembly and Rapid Growth of Amyloid Fibrils

    NASA Astrophysics Data System (ADS)

    Lin, Yichih; Petersson, E. James; Fakhraai, Zahra

    2014-03-01

    The mechanism of surface-mediated fibrillization has been considered as a key issue in understanding the origins of the neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases. In vitro, amyloid proteins fold through nucleation-elongation process. There is a critical concentration for early nucleating stage. However, some studies indicate that surfaces can modulate the fibril's formation under physiological conditions, even when the concentration is much lower than the critical concentration. Here, we use a label-free procedure to monitor the growth of fibrils across many length scales. We show that near a surface, the fibrillization process appears to bypass the nucleation step and fibrils grow through a self-assembly mechanism instead. We control and measure the pre-fibrillar morphology at different stages of this process on various surfaces. The interplay between the surface concentration and diffusion constant can help identify the detailed mechanisms of surface-mediated fibril growth, which remains largely unexplored. Our works provide a new insight in designing new probes and therapies. Supported by the National Institute On Aging of the National Institutes of Health under Award Number P30AG010124.

  12. Thrombogenic collagen-mimetic peptides: Self-assembly of triple helix-based fibrils driven by hydrophobic interactions

    PubMed Central

    Cejas, Mabel A.; Kinney, William A.; Chen, Cailin; Vinter, Jeremy G.; Almond, Harold R.; Balss, Karin M.; Maryanoff, Cynthia A.; Schmidt, Ute; Breslav, Michael; Mahan, Andrew; Lacy, Eilyn; Maryanoff, Bruce E.

    2008-01-01

    Collagens are integral structural proteins in animal tissues and play key functional roles in cellular modulation. We sought to discover collagen model peptides (CMPs) that would form triple helices and self-assemble into supramolecular fibrils exhibiting collagen-like biological activity without preorganizing the peptide chains by covalent linkages. This challenging objective was accomplished by placing aromatic groups on the ends of a representative 30-mer CMP, (GPO)10, as with l-phenylalanine and l-pentafluorophenylalanine in 32-mer 1a. Computational studies on homologous 29-mers 1a′–d′ (one less GPO), as pairs of triple helices interacting head-to-tail, yielded stabilization energies in the order 1a′ > 1b′ > 1c′ > 1d′, supporting the hypothesis that hydrophobic aromatic groups can drive CMP self-assembly. Peptides 1a–d were studied comparatively relative to structural properties and ability to stimulate human platelets. Although each 32-mer formed stable triple helices (CD) spectroscopy, only 1a and 1b self-assembled into micrometer-scale fibrils. Light microscopy images for 1a depicted long collagen-like fibrils, whereas images for 1d did not. Atomic force microscopy topographical images indicated that 1a and 1b self-organize into microfibrillar species, whereas 1c and 1d do not. Peptides 1a and 1b induced the aggregation of human blood platelets with a potency similar to type I collagen, whereas 1c was much less effective, and 1d was inactive (EC50 potency: 1a/1b ≫ 1c > 1d). Thus, 1a and 1b spontaneously self-assemble into thrombogenic collagen-mimetic materials because of hydrophobic aromatic interactions provided by the special end-groups. These findings have important implications for the design of biofunctional CMPs. PMID:18559857

  13. New method for Raman investigation of the orientation of collagen fibrils and crystallites in the Haversian system of bone.

    PubMed

    Falgayrac, Guillaume; Facq, Sébastien; Leroy, Gérard; Cortet, Bernard; Penel, Guillaume

    2010-07-01

    Knowledge of the organization of the components of bone is of primary importance in understanding how this tissue responds to stresses and provides a starting point for the design and development of biomaterials. Bone structure has been the subject of numerous studies. The mineralized fiber arrangement in cortical bone is either a twisted or orthogonal plywood structure. Both mineral models coexist in compact bone. Raman polarized spectroscopy offers definite advantages in the study of biological samples, enabling the simultaneous analysis of mineral and organic components and the determination of molecular orientation through the polarization properties of the Raman scattering. In this study, we used the Raman polarization approach to simultaneously investigate the orientation of collagen fibrils and apatite crystals in human cortical bone. Raman bands ratios were monitored as a function of sample orientation. Specific ratios were chosen--such as nu(3) PO(4)/nu(1) PO(4), amide III (1271 cm(-1))/amide III (1243 cm(-1)), and amide I/amide III (1243 cm(-1))--due to their sensitivity to apatite-crystal and collagen-fibril orientation. Based on this original approach, spatial changes were monitored as a function of distance from the Haversian canal. The results revealed simultaneous tilting in intra-lamellar collagen-fibril and mineral crystal orientations. These results are consistent with a twisted plywood organization in the Haversian bone structure at the lamellar level. But at molecular level, the co-alignment of the collagen fibrils and the apatite crystal is observed in the innermost lamellae and becomes gradually less ordered as the distance from the Haversian canal increases. This work highlights the interest of Raman spectroscopy for the multiscale investigation of bone structure. PMID:20615291

  14. Nano measurements with micro-devices: mechanical properties of hydrated collagen fibrils

    PubMed Central

    Eppell, S.J; Smith, B.N; Kahn, H; Ballarini, R

    2005-01-01

    The mechanical response of a biological material to applied forces reflects deformation mechanisms occurring within a hierarchical architecture extending over several distinct length scales. Characterizing and in turn predicting the behaviour of such a material requires an understanding of the mechanical properties of the substructures within the hierarchy, the interaction between the substructures, and the relative influence of each substructure on the overall behaviour. While significant progress has been made in mechanical testing of micrometre to millimetre sized biological specimens, quantitative reproducible experimental techniques for making mechanical measurements on specimens with characteristic dimensions in the smaller range of 10–1000 nm are lacking. Filling this void in experimentation is a necessary step towards the development of realistic multiscale computational models useful to predict and mitigate the risk of bone fracture, design improved synthetic replacements for bones, tendons and ligaments, and engineer bioinspired efficient and environmentally friendly structures. Here, we describe a microelectromechanical systems device for directly measuring the tensile strength, stiffness and fatigue behaviour of nanoscale fibres. We used the device to obtain the first stress–strain curve of an isolated collagen fibril producing the modulus and some fatigue properties of this soft nanofibril. PMID:16849223

  15. Age- and diabetes-related nonenzymatic crosslinks in collagen fibrils: candidate amino acids involved in Advanced Glycation End-products.

    PubMed

    Gautieri, Alfonso; Redaelli, Alberto; Buehler, Markus J; Vesentini, Simone

    2014-02-01

    Ageing and diabetes share a common deleterious phenomenon, the formation of Advanced Glycation Endproducts (AGEs), which accumulate predominantly in collagen due to its low turnover. Though the general picture of glycation has been identified, the detailed knowledge of which collagen amino acids are involved in AGEs is still missing. In this work we use an atomistic model of a collagen fibril to pinpoint, for the first time, the precise location of amino acids involved in the most relevant AGE, glucosepane. The results show that there are 14 specific lysine-arginine pairs that, due to their relative position and configuration, are likely to form glucosepane. We find that several residues involved in AGE crosslinks are within key collagen domains, such as binding sites for integrins, proteoglycans and collagenase, hence providing molecular-level explanations of previous experimental results showing decreased collagen affinity for key molecules. Altogether, these findings reveal the molecular mechanism by which glycation affects the biological properties of collagen tissues, which in turn contribute to age- and diabetes-related pathological states. PMID:24060753

  16. Hierarchical ordering of amyloid fibrils on the mica surface

    NASA Astrophysics Data System (ADS)

    Zhou, Xingfei; Zhang, Yingying; Zhang, Feng; Pillai, Saju; Liu, Jianhua; Li, Rong; Dai, Bin; Li, Bin; Zhang, Yi

    2013-05-01

    The aggregation of amyloid peptides into ordered fibrils is closely associated with many neurodegenerative diseases. The surfaces of cell membranes and biomolecules are believed to play important roles in modulation of peptide aggregation under physiological conditions. Experimental studies of fibrillogenesis at the molecular level in vivo, however, are inherently challenging, and the molecular mechanisms of how surface affects the structure and ordering of amyloid fibrils still remain elusive. Herein we have investigated the aggregation behavior of insulin peptides within water films adsorbed on the mica surface. AFM measurements revealed that the structure and orientation of fibrils were significantly affected by the mica lattice and the peptide concentration. At low peptide concentration (~0.05 mg mL-1), there appeared a single layer of short and well oriented fibrils with a mean height of 1.6 nm. With an increase of concentration to a range of 0.2-2.0 mg mL-1, a different type of fibrils with a mean height of 3.8 nm was present. Interestingly, when the concentration was above 2.0 mg mL-1, the thicker fibrils exhibited two-dimensional liquid-crystal-like ordering probably caused by the combination of entropic and electrostatic forces. These results could help us gain better insight into the effects of the substrate on amyloid fibrillation.The aggregation of amyloid peptides into ordered fibrils is closely associated with many neurodegenerative diseases. The surfaces of cell membranes and biomolecules are believed to play important roles in modulation of peptide aggregation under physiological conditions. Experimental studies of fibrillogenesis at the molecular level in vivo, however, are inherently challenging, and the molecular mechanisms of how surface affects the structure and ordering of amyloid fibrils still remain elusive. Herein we have investigated the aggregation behavior of insulin peptides within water films adsorbed on the mica surface. AFM

  17. Modification of Ti6Al4V surfaces using collagen I, III, and fibronectin. I. Biochemical and morphological characteristics of the adsorbed matrix.

    PubMed

    Bierbaum, Susanne; Beutner, René; Hanke, Thomas; Scharnweber, Dieter; Hempel, Ute; Worch, Hartmut

    2003-11-01

    Studies in developmental and cell biology have established the fact that responses of cells are influenced to a large degree by morphology and composition of the extracellular matrix. Goal of this work is to use this basic principle to improve the biological acceptance of implants by modifying the surfaces with components of the extracellular matrix (ECM). Aiming at load-bearing applications in bone contact, in this study the modification of titanium surfaces with the collagen types I and III in combination with fibronectin was undertaken; fibrillogenesis, fibril morphology and adsorption of type I, III and I/III-cofibrils onto titanium were assessed. Increasing the collagen type III amount resulted in a decrease of fibril diameter, while no significant changes in adsorption could be detected. The amount of fibronectin bound to the heterotypic fibrils depended on fibrillogenesis parameters such as ionic strength or concentration of phosphate, and varied with the percentage of integrated type III collagen. PMID:14566782

  18. Characterization via atomic force microscopy of discrete plasticity in collagen fibrils from mechanically overloaded tendons: Nano-scale structural changes mimic rope failure.

    PubMed

    Baldwin, Samuel J; Kreplak, Laurent; Lee, J Michael

    2016-07-01

    Tendons exposed to tensile overload show a structural alteration at the fibril scale termed discrete plasticity. Serial kinks appear along individual collagen fibrils that are susceptible to enzymatic digestion and are thermally unstable. Using atomic force microscopy we mapped the topography and mechanical properties in dehydrated and hydrated states of 25 control fibrils and 25 fibrils displaying periodic kinks, extracted from overloaded bovine tail tendons. Using the measured modulus of the hydrated fibrils as a probe of molecular density, we observed a non-linear negative correlation between molecular density and kink density of individual fibrils. This is accompanied by an increase in water uptake with kink density and a doubling of the coefficient of variation of the modulus between kinked, and control fibrils. The mechanical property maps of kinked collagen fibrils show radial heterogeneity that can be modeled as a high-density core surrounded by a low-density shell. The core of the fibril contains the kink structures characteristic of discrete plasticity; separated by inter-kink regions, which often retain the D-banding structure. We propose that the shell and kink structures mimic characteristic damage motifs observed in laid rope strands. PMID:26925699

  19. Contact activation of blood coagulation on a defined kaolin/collagen surface in a microfluidic assay

    PubMed Central

    Zhu, Shu; Diamond, Scott L.

    2014-01-01

    Generation of active Factor XII (FXIIa) triggers blood clotting on artificial surfaces and may also enhance intravascular thrombosis. We developed a patterned kaolin (0 to 0.3 pg/μm2)/type 1 collagen fibril surface for controlled microfluidic clotting assays. Perfusion of whole blood (treated only with a low level of 4 μg/mL of the XIIa inhibitor, corn trypsin inhibitor) drove platelet deposition followed by fibrin formation. At venous wall shear rate (100 s−1), kaolin accelerated onset of fibrin formation by ~100 sec when compared to collagen alone (250 sec vs. 350 sec), with little effect on platelet deposition. Even with kaolin present, arterial wall shear rate (1000 s−1) delayed and suppressed fibrin formation compared to venous wall shear rate. A comparison of surfaces for extrinsic activation (tissue factor TF/collagen) versus contact activation (kaolin/collagen) that each generated equal platelet deposition at 100 s−1 revealed: (1) TF surfaces promoted much faster fibrin onset (at 100 sec) and more endpoint fibrin at 600 sec at either 100 s−1 or 1000 s−1, and (2) kaolin and TF surfaces had a similar sensitivity for reduced fibrin deposition at 1000 s−1 (compared to fibrin formed at 100 s−1) despite differing coagulation triggers. Anti-platelet drugs inhibiting P2Y1, P2Y12, cyclooxygenase-1 or activating IP-receptor or guanylate cyclase reduced platelet and fibrin deposition on kaolin/collagen. Since FXIIa or FXIa inhibition may offer safe antithrombotic therapy, especially for biomaterial thrombosis, these defined collagen/kaolin surfaces may prove useful in drug screening tests or in clinical diagnostic assays of blood under flow conditions. PMID:25303860

  20. COLLAGEN PROCESSING

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Collagen dispersions, produced from fibrils recovered from milled bovine collagen, have shown promise in environmental remediation in applications as settling aids, filtration aids, fractionation media, oil drop stabilizers, and water purification aids. Macroporous structures, processed by controll...

  1. Softenin, a Novel Protein That Softens the Connective Tissue of Sea Cucumbers through Inhibiting Interaction between Collagen Fibrils

    PubMed Central

    Takehana, Yasuhiro; Yamada, Akira; Tamori, Masaki; Motokawa, Tatsuo

    2014-01-01

    The dermis in the holothurian body wall is a typical catch connective tissue or mutable collagenous tissue that shows rapid changes in stiffness. Some chemical factors that change the stiffness of the tissue were found in previous studies, but the molecular mechanisms of the changes are not yet fully understood. Detection of factors that change the stiffness by working directly on the extracellular matrix was vital to clarify the mechanisms of the change. We isolated from the body wall of the sea cucumber Stichopus chloronotus a novel protein, softenin, that softened the body-wall dermis. The apparent molecular mass was 20 kDa. The N-terminal sequence of 17 amino acids had low homology to that of known proteins. We performed sequential chemical and physical dissections of the dermis and tested the effects of softenin on each dissection stage by dynamic mechanical tests. Softenin softened Triton-treated dermis whose cells had been disrupted by detergent. The Triton-treated dermis was subjected to repetitive freeze-and-thawing to make Triton-Freeze-Thaw (TFT) dermis that was softer than the Triton-treated dermis, implying that some force-bearing structure had been disrupted by this treatment. TFT dermis was stiffened by tensilin, a stiffening protein of sea cucumbers. Softenin softened the tensilin-stiffened TFT dermis while it had no effect on the TFT dermis without tensilin treatment. We isolated collagen from the dermis. When tensilin was applied to the suspending solution of collagen fibrils, they made a large compact aggregate that was dissolved by the application of softenin or by repetitive freeze-and-thawing. These results strongly suggested that softenin decreased dermal stiffness through inhibiting cross-bridge formation between collagen fibrils; the formation was augmented by tensilin and the bridges were broken by the freeze-thaw treatment. Softenin is thus the first softener of catch connective tissue shown to work on the cross-bridges between

  2. Temperature-Responsive Gelation of Type I Collagen Solutions Involving Fibril Formation and Genipin Crosslinking as a Potential Injectable Hydrogel

    PubMed Central

    Yunoki, Shunji

    2013-01-01

    We investigated the temperature-responsive gelation of collagen/genipin solutions using pepsin-solubilized collagen (PSC) and acid-solubilized collagen (ASC) as substrates. Gelation occurred in the PSC/genipin solutions at genipin concentrations 0–2 mM under moderate change in temperature from 25 to 37°C. The PSC/genipin solutions exhibited fluidity at room temperature for at least 30 min, whereas the ASC/genipin solutions rapidly reached gel points. In specific cases PSC would be preferred over ASC as an injectable gel system. The temperature-responsive gelation of PSC/genipin solutions was due to temperature responses to genipin crosslinking and collagen fibril formation. The elastic modulus of the 0.5% PSC/genipin gel system could be adjusted in a range of 2.5 to 50 kPa by the PSC and genipin concentrations, suggesting that a PSC/genipin solution is a potential injectable gel system for drug and cell carriers, with mechanical properties matching those of living tissues. PMID:24222766

  3. Attachment and conformational changes of collagen on bioactive glass surface.

    PubMed

    Magyari, K; Vanea, E; Baia, L; Simon, V

    2016-05-12

    The proteins adsorption on biomaterials surface leads to changes in their structural conformation that may further influence the adhesion, migration and growth of cells. The aim of this study was to examine the attachment of collagen (calf skin type I) on bioactive glass powders and the conformational changes of the protein. Scanning electron microscopy analysis and X-ray photoelectron spectroscopy measurements indicate that the collagen cover the glass surface in a nanometric thin layer. The infrared amide I absorption signal shows pronounced changes in the secondary structure of the adsorbed collagen. PMID:27175468

  4. Collagen nanofilm immobilized on at surfaces by electrodeposition method.

    PubMed

    Yang, Xiudong; Jiang, Bo; Huang, Yi; Tian, Yunfei; Chen, Hong; Chen, Jiyong; Yang, Bangcheng

    2009-08-01

    A simple electrodeposition method is presented for the preparing of collagen nanofilms (EAT) on anodic oxidized titanium surfaces (AT). The nanofilms were observed by scanning electron microscopy and atomic force microscopy. Functional TiOx layers with anionic groups of --PO(4), --SO(4) and --OH were investigated on the AT surface by X-ray photoelectron spectroscopy; X-ray diffraction results indicated that the AT surface was composed mainly of anatase and rutile. The bioactive electrodeposited TiOx layers on the AT surface showed lower water contact angles and higher surface energy than pure titanium surfaces (CT) and displayed higher collagen molecule immobilization. PMID:19165768

  5. Structural constraints on the evolution of the collagen fibril: convergence on a 1014-residue COL domain

    PubMed Central

    Slatter, David Anthony; Farndale, Richard William

    2015-01-01

    Type I collagen is the fundamental component of the extracellular matrix. Its α1 gene is the direct descendant of ancestral fibrillar collagen and contains 57 exons encoding the rod-like triple-helical COL domain. We trace the evolution of the COL domain from a primordial collagen 18 residues in length to its present 1014 residues, the limit of its possible length. In order to maintain and improve the essential structural features of collagen during evolution, exons can be added or extended only in permitted, non-random increments that preserve the position of spatially sensitive cross-linkage sites. Such sites cannot be maintained unless the twist of the triple helix is close to 30 amino acids per turn. Inspection of the gene structure of other long structural proteins, fibronectin and titin, suggests that their evolution might have been subject to similar constraints. PMID:25994354

  6. Investigation of ethanol infiltration into demineralized dentin collagen fibrils using molecular dynamics simulations.

    PubMed

    Jee, Sang Eun; Zhou, Jienfeng; Tan, Jianquo; Breschi, Lorenzo; Tay, Franklin R; Grégoire, Geneviève; Pashley, David H; Jang, Seung Soon

    2016-05-01

    The purpose of this study is to investigate the interaction of neat ethanol with bound and non-bound water in completely demineralized dentin that is fully hydrated, using molecular dynamics (MD) simulation method. The key to creating ideal resin-dentin bonds is the removal of residual free water layers and its replacement by ethanol solvent in which resin monomers are soluble, using the ethanol wet-bonding technique. The test null hypotheses were that ethanol cannot remove any collagen-bound water, and that ethanol cannot infiltrate into the spacing between collagen triple helix due to narrow interlayer spacing. Collagen fibrillar structures of overlap and gap regions were constructed by aligning the collagen triple helix of infinite length in hexagonal packing. Three layers of the water molecules were specified as the layers of 0.15-0.22nm, 0.22-0.43nm and 0.43-0.63nm from collagen atoms by investigating the water distribution surrounding collagen molecules. Our simulation results show that ethanol molecules infiltrated into the intermolecular spacing in the gap region, which increased due to the lateral shrinkage of the collagen structures in contact with ethanol solution, while there was no ethanol infiltration observed in the overlap region. Infiltrated ethanol molecules in the gap region removed residual water molecules via modifying mostly the third water layer (50% decrease), which would be considered as a loosely-bound water layer. The first and second hydration layers, which would be considered as tightly bound water layers, were not removed by the ethanol molecules, thus maintaining the helical structures of the collagen molecules. PMID:26969524

  7. Age-associated reduction of cellular spreading/mechanical force up-regulates matrix metalloproteinase-1 expression and collagen fibril fragmentation via c-Jun/AP-1 in human dermal fibroblasts

    PubMed Central

    Qin, Zhaoping; Voorhees, John J; Fisher, Gary J; Quan, Taihao

    2014-01-01

    The dermal compartment of human skin is largely composed of dense collagen-rich fibrils, which provide structural and mechanical support. Skin dermal fibroblasts, the major collagen-producing cells, are interact with collagen fibrils to maintain cell spreading and mechanical force for function. A characteristic feature of aged human skin is fragmentation of collagen fibrils, which is initiated by matrix metalloproteinase 1 (MMP-1). Fragmentation impairs fibroblast attachment and thereby reduces spreading. Here, we investigated the relationship among fibroblast spreading, mechanical force, MMP-1 expression, and collagen fibril fragmentation. Reduced fibroblast spreading due to cytoskeletal disruption was associated with reduced cellular mechanical force, as determined by atomic force microscopy. These reductions substantially induced MMP-1 expression, which led to collagen fibril fragmentation and disorganization in three-dimensional collagen lattices. Constraining fibroblast size by culturing on slides coated with collagen micropatterns also significantly induced MMP-1 expression. Reduced spreading/mechanical force induced transcription factor c-Jun and its binding to a canonical AP-1 binding site in the MMP-1 proximal promoter. Blocking c-Jun function with dominant negative mutant c-Jun significantly reduced induction of MMP-1 expression in response to reduced spreading/mechanical force. Furthermore, restoration of fibroblast spreading/mechanical force led to decline of c-Jun and MMP-1 levels and eliminated collagen fibril fragmentation and disorganization. These data reveal a novel mechanism by which alteration of fibroblast shape/mechanical force regulates c-Jun/AP-1-dependent expression of MMP-1 and consequent collagen fibril fragmentation. This mechanism provides a foundation for understanding the cellular and molecular basis of age-related collagen fragmentation in human skin. PMID:25201474

  8. Plasmodium falciparum Merozoite Surface Protein 2 is Unstructured and Forms Amyloid-Like Fibrils

    PubMed Central

    Adda, Christopher G.; Murphy, Vince J.; Sunde, Margaret; Waddington, Lynne J.; Jesse, Schloegel; Talbo, Gert H.; Vingas, Kleo; Kienzle, Vivian; Masciantonio, Rosella; Howlett, Geoffrey J.; Hodder, Anthony N.; Foley, Michael; Anders, Robin F.

    2009-01-01

    Several merozoite surface proteins are being assessed as potential components of a vaccine against Plasmodium falciparum, the cause of the most serious form of human malaria. One of these proteins, merozoite surface protein 2 (MSP2), is unusually hydrophilic and contains tandem sequence repeats, characteristics of intrinsically unstructured proteins. A range of physicochemical studies have confirmed that recombinant forms of MSP2 are largely unstructured. Both dimorphic types of MSP2 (3D7 and FC27) are equivalently extended in solution and form amyloid-like fibrils although with different kinetics and structural characteristics. These fibrils have a regular underlying β-sheet structure and both fibril types stain with Congo Red, but only the FC27 fibrils stain with Thioflavin T. 3D7 MSP2 fibrils seeded the growth of fibrils from 3D7 or FC27 MSP2 monomer indicating the involvement of a conserved region of MSP2 in fibril formation. Consistent with this, digestion of fibrils with proteinase K generated resistant peptides, which included the N-terminal conserved region of MSP2. A monoclonal antibody that reacted preferentially with monomeric recombinant MSP2 did not react with the antigen in situ on the merozoite surface. Glutaraldehyde cross-linking of infected erythrocytes generated MSP2 oligomers similar to those formed by polymeric recombinant MSP2. We conclude that MSP2 oligomers containing intermolecular β-strand interactions similar to those in amyloid fibrils may be a component of the fibrillar surface coat on P. falciparum merozoites. PMID:19450733

  9. Plasmodium falciparum merozoite surface protein 2 is unstructured and forms amyloid-like fibrils.

    PubMed

    Adda, Christopher G; Murphy, Vince J; Sunde, Margaret; Waddington, Lynne J; Schloegel, Jesse; Talbo, Gert H; Vingas, Kleo; Kienzle, Vivian; Masciantonio, Rosella; Howlett, Geoffrey J; Hodder, Anthony N; Foley, Michael; Anders, Robin F

    2009-08-01

    Several merozoite surface proteins are being assessed as potential components of a vaccine against Plasmodium falciparum, the cause of the most serious form of human malaria. One of these proteins, merozoite surface protein 2 (MSP2), is unusually hydrophilic and contains tandem sequence repeats, characteristics of intrinsically unstructured proteins. A range of physicochemical studies has confirmed that recombinant forms of MSP2 are largely unstructured. Both dimorphic types of MSP2 (3D7 and FC27) are equivalently extended in solution and form amyloid-like fibrils although with different kinetics and structural characteristics. These fibrils have a regular underlying beta-sheet structure and both fibril types stain with Congo Red, but only the FC27 fibrils stain with Thioflavin T. 3D7 MSP2 fibrils seeded the growth of fibrils from 3D7 or FC27 MSP2 monomer indicating the involvement of a conserved region of MSP2 in fibril formation. Consistent with this, digestion of fibrils with proteinase K generated resistant peptides, which included the N-terminal conserved region of MSP2. A monoclonal antibody that reacted preferentially with monomeric recombinant MSP2 did not react with the antigen in situ on the merozoite surface. Glutaraldehyde cross-linking of infected erythrocytes generated MSP2 oligomers similar to those formed by polymeric recombinant MSP2. We conclude that MSP2 oligomers containing intermolecular beta-strand interactions similar to those in amyloid fibrils may be a component of the fibrillar surface coat on P. falciparum merozoites. PMID:19450733

  10. Streptococcus salivarius strains carry either fibrils or fimbriae on the cell surface.

    PubMed Central

    Handley, P S; Carter, P L; Fielding, J

    1984-01-01

    Strains of Streptococcus salivarius were screened by negative staining for the presence of surface structures. Two structural subgroups were found, carrying either fibrils or fimbriae, projecting from the cell surface. Eight strains carried a very dense peritrichous array of fibrils of two distinct lengths. Long fibrils had an average length of 175 nm, and short fibrils had an average length of 95 nm. Two strains carried only long fibrils, one strain carried only short fibrils, and another strain carried a lateral tuft of very prominent fibrils of two lengths, with a fibrillar fuzz covering the remainder of the cell surface. In all the strains in which they were present, the long fibrils were unaffected by protease or trypsin treatment. In contrast, the short fibrils were completely digested by protease and partially removed by trypsin. Neither long nor short fibrils were affected structurally by mild pepsin digestion or by lipase. The Lancefield extraction procedure removed both long and short fibrils. These twelve fibrillar strains were therefore divisible into four structural subgroups. Extracts of all the fibrillar strains reacted with group K antiserum. The second main structural subgroup consisted of nine strains of S. salivarius, all of which carried morphologically identical, flexible fimbriae arranged peritrichously over the cell surface. The fimbriae were structurally distinct from fibrils and measured 0.5 to 1.0 micron long and 3 to 4 nm wide, with an irregular outline and no obvious substructure. There was no obvious reduction in the number of fimbriae after protease or trypsin treatment. Extracts of the fimbriated strains did not react with the group K antiserum. The two serological and structural subgroups could also be distinguished by colony morphology. Images PMID:6197404

  11. Genetic linkage of type VII collagen (COL7A1) to dominant dystrophic epidermolysis bullosa in families with abnormal anchoring fibrils.

    PubMed Central

    Ryynänen, M; Ryynänen, J; Sollberg, S; Iozzo, R V; Knowlton, R G; Uitto, J

    1992-01-01

    Epidermolysis bullosa (EB) in a group of genodermatoses characterized by the fragility of skin. Previous studies on the dystrophic (scarring) forms of EB have suggested abnormalities in anchoring fibrils, morphologically recognizable attachment structures that provide stability to the association of the cutaneous basement membrane to the underlying dermis. Since type VII collagen is the major component of the anchoring fibrils, we examined the genetic linkage of dominant dystrophic EB (EBDD) and the type VII collagen gene (COL7A1) locus, which we have recently mapped to chromosome 3p, in three large kindreds with abnormal anchoring fibrils. Strong genetic linkage of EBDD and COL7A1 loci was demonstrated with the maximum logarithm of odds (LOD) score of 8.77 at theta = 0. This linkage was further confirmed with two additional markers in this region of the short arm of chromosome 3, and these analyses allowed further refinement of the map locus of COL7A1. Since there were no recombinants between the COL7A1 and EBDD loci, our findings suggest that type VII collagen is the candidate gene that may harbor the mutations responsible for the EB phenotype in these three families. Images PMID:1347297

  12. On the computation of stress in affine versus nonaffine fibril kinematics within planar collagen network models.

    PubMed

    Pence, Thomas J; Monroe, Ryan J; Wright, Neil T

    2008-08-01

    Some recent analyses modeled the response of collagenous tissues, such as epicardium, using a hypothetical network consisting of interconnected springlike fibers. The fibers in the network were organized such that internal nodes served as the connection point between three such collagen springs. The results for assumed affine and nonaffine deformations are contrasted after a homogeneous deformation at the boundary. Affine deformation provides a stiffer mechanical response than nonaffine deformation. In contrast to nonaffine deformation, affine deformation determines the displacement of internal nodes without imposing detailed force balance, thereby complicating the simplest intuitive notion of stress, one based on free body cuts, at the single node scale. The standard notion of stress may then be recovered via average field theory computations based on large micromesh realizations. An alternative and by all indications complementary viewpoint for the determination of stress in these collagen fiber networks is discussed here, one in which stress is defined using elastic energy storage, a notion which is intuitive at the single node scale. It replaces the average field theory computations by an averaging technique over randomly oriented isolated simple elements. The analytical operations do not require large micromesh realizations, but the tedious nature of the mathematical manipulation is clearly aided by symbolic algebra calculation. For the example case of linear elastic deformation, this results in material stiffnesses that relate the infinitesimal strain and stress. The result that the affine case is stiffer than the nonaffine case is recovered, as would be expected. The energy framework also lends itself to the natural inclusion of changes in mechanical response due to the chemical, electrical, or thermal environment. PMID:18601451

  13. Body Surface Mapping to Guide Atrial Fibrillation Ablation

    PubMed Central

    Yamashita, Seigo; Shah, Ashok J; Mahida, Saagar; Sellal, Jean-Marc; Berte, Benjamin; Hooks, Darren; Frontera, Antonio; Jefairi, Nora Al; Wielandts, Jean-Yves; Lim, Han S; Amraoui, Sana; Denis, Arnaud; Derval, Nicolas; Sacher, Frédéric; Cochet, Hubert; Hocini, Mélèze; Jaïs, Pierre; Haïssaguerre, Michel

    2015-01-01

    Atrial fibrillation (AF) is the most common rhythm disorder, and is strongly associated with thromboembolic events and heart failure. Over the past decade, catheter ablation of AF has advanced considerably with progressive improvement in success rates. However, interventional treatment is still challenging, especially for persistent and long-standing persistent AF. Recently, AF analysis using a non-invasive body surface mapping technique has been shown to identify localised reentrant and focal sources, which play an important role in driving and perpetuating AF. Non-invasive mapping-guided ablation has also been reported to be effective for persistent AF. In this review, we describe new clinical insights obtained from non-invasive mapping of persistent AF to guide catheter ablation. PMID:26835121

  14. Body Surface Mapping to Guide Atrial Fibrillation Ablation.

    PubMed

    Yamashita, Seigo; Shah, Ashok J; Mahida, Saagar; Sellal, Jean-Marc; Berte, Benjamin; Hooks, Darren; Frontera, Antonio; Jefairi, Nora Al; Wielandts, Jean-Yves; Lim, Han S; Amraoui, Sana; Denis, Arnaud; Derval, Nicolas; Sacher, Frédéric; Cochet, Hubert; Hocini, Mélèze; Jaïs, Pierre; Haïssaguerre, Michel

    2015-12-01

    Atrial fibrillation (AF) is the most common rhythm disorder, and is strongly associated with thromboembolic events and heart failure. Over the past decade, catheter ablation of AF has advanced considerably with progressive improvement in success rates. However, interventional treatment is still challenging, especially for persistent and long-standing persistent AF. Recently, AF analysis using a non-invasive body surface mapping technique has been shown to identify localised reentrant and focal sources, which play an important role in driving and perpetuating AF. Non-invasive mapping-guided ablation has also been reported to be effective for persistent AF. In this review, we describe new clinical insights obtained from non-invasive mapping of persistent AF to guide catheter ablation. PMID:26835121

  15. Two-way regulation between cells and aligned collagen fibrils: local 3D matrix formation and accelerated neural differentiation of human decidua parietalis placental stem cells.

    PubMed

    Li, Wen; Zhu, Bofan; Strakova, Zuzana; Wang, Rong

    2014-08-01

    It has been well established that an aligned matrix provides structural and signaling cues to guide cell polarization and cell fate decision. However, the modulation role of cells in matrix remodeling and the feedforward effect on stem cell differentiation have not been studied extensively. In this study, we report on the concerted changes of human decidua parietalis placental stem cells (hdpPSCs) and the highly ordered collagen fibril matrix in response to cell-matrix interaction. With high-resolution imaging, we found the hdpPSCs interacted with the matrix by deforming the cell shape, harvesting the nearby collagen fibrils, and reorganizing the fibrils around the cell body to transform a 2D matrix to a localized 3D matrix. Such a unique 3D matrix prompted high expression of β-1 integrin around the cell body that mediates and facilitates the stem cell differentiation toward neural cells. The study offers insights into the coordinated, dynamic changes at the cell-matrix interface and elucidates cell modulation of its matrix to establish structural and biochemical cues for effective cell growth and differentiation. PMID:25003322

  16. Lipidation Effect on Surface Adsorption and Associated Fibrillation of the Model Protein Insulin.

    PubMed

    Hedegaard, Sofie Fogh; Cárdenas, Marité; Barker, Robert; Jorgensen, Lene; van de Weert, Marco

    2016-07-19

    Lipidation of proteins is used in the pharmaceutical field to increase the therapeutic efficacy of proteins. In this study, we investigate the effect of a 14-carbon fatty acid modification on the adsorption behavior of human insulin to a hydrophobic solid surface and the subsequent fibrillation development under highly acidic conditions and elevated temperature by comparing to the fibrillation of human insulin. At these stressed conditions, the lipid modification accelerates the rate of fibrillation in bulk solution. With the use of several complementary surface-sensitive techniques, including quartz crystal microbalance with dissipation monitoring (QCM-D), atomic force microscopy (AFM), and neutron reflectivity (NR), we show that there are two levels of structurally different protein organization at a hydrophobic surface for both human insulin and the lipidated analogue: a dense protein layer formed within minutes on the surface and a diffuse outer layer of fibrillar structures which took hours to form. The two layers may only be weakly connected, and proteins from both layers are able to desorb from the surface. The lipid modification increases the protein surface coverage and the thickness of both layer organizations. Upon lipidation not only the fibrillation extent but also the morphology of the fibrillar structures changes from fibril clusters on the surface to a more homogeneous network of fibrils covering the entire hydrophobic surface. PMID:27348237

  17. Liquid crystalline collagen: a self-assembled morphology for the orientation of mammalian cells.

    PubMed

    Kirkwood, John E; Fuller, Gerald G

    2009-03-01

    We report the creation of collagen films having a cholesteric banding structure with an orientation that can be systematically controlled. The action of hydrodynamic flow and rapid desiccation was used to influence the orientation of collagen fibrils, producing a film with a twisted plywood architecture. Adult human fibroblasts cultured on these substrates orient in the direction of the flow deposition, and filopodia are extended onto individual bands. Atomic force microscopy reveals the assembly of 30 nm collagen fibrils into the uniform cholesteric collagen films with a periodic surface relief. The generation of collagen with a reticular, "basket-weave" morphology when using lower concentrations is also discussed. PMID:19437784

  18. Collagen coated tantalum substrate for cell proliferation.

    PubMed

    Li, Yinli; Zhang, Shuai; Guo, Lijun; Dong, Mingdong; Liu, Bo; Mamdouh, Wael

    2012-06-15

    The extracellular matrix (ECM) plays a key role in cell culture in various physiological and pathological processes in the field of tissue engineering. Recently, the type I collagen ECM has been widely utilized in vitro model systems for the attachment of many different cell lines since it has multi-functions in human tissues. For example it accounts for 6% of the weight of strong, tendinous muscles. In this paper, we reported a new material by coating tantalum (Ta), one highly biocompatible metal, with type I collagen fibrils. The morphology of the new material was studied by high resolution atomic force microscope. It was shown that the adhesion force between type I collagen fibrils network and Ta was strong enough to overcome surface defects. A possible way to explain the phenomenon is that the longitudinal periodicity of collagen fibrils matches the grain size of the Ta domains, which results in increase of the physical adsorption contact area, thereby inducing the dramatic adhesion enhancement between collagen fibrils and Ta. The obtained material was then employed as a template for cell proliferation. Although the surface of this template is more hydrophobic by comparison with the bare Ta surface, the cells on this material were successfully incubated, indicating that the collagen coated Ta might be used as the buffer layer for proliferating cells in hydrophobic biomaterials. PMID:22494669

  19. Dependencies of multi-component T 2 and T 1 ρ relaxation on the anisotropy of collagen fibrils in bovine nasal cartilage

    NASA Astrophysics Data System (ADS)

    Wang, Nian; Xia, Yang

    2011-09-01

    Both NMR spectroscopy and MRI were used to investigate the dependencies of multi-component T2 and T1ρ relaxation on the anisotropy of bovine nasal cartilage (BNC). The non-negative least square (NNLS) method and the multi-exponential fitting method were used to analyze all experimental data. When the collagen fibrils in nasal cartilage were oriented at the magic angle (55°) to the magnetic field B 0, both T2 and T1ρ were single component, regardless of the spin-lock field strength or the echo spacing time in the pulse sequences. When the collagen fibrils in nasal cartilage were oriented at 0° to B 0, both T2 and T1ρ at a spin-lock field of 500 Hz had two components. When the spin-lock field was increased to 1000 Hz or higher, T1ρ relaxation in nasal cartilage became a single component, even when the specimen orientation was 0°. These results demonstrate that the specimen orientation must be considered for any multi-component analysis, even for nasal cartilage that is commonly considered homogenously structured. Since the rapidly and slowly relaxing components can be attributed to different portions of the water population in tissue, the ability to resolve different relaxation components could be used to quantitatively examine individual molecular components in connective tissues.

  20. Lateral growth limitation of corneal fibrils and their lamellar stacking depend on covalent collagen cross-linking by transglutaminase-2 and lysyl oxidases, respectively.

    PubMed

    Wang, Lei; Uhlig, Philipp C; Eikenberry, Eric F; Robenek, Horst; Bruckner, Peter; Hansen, Uwe

    2014-01-10

    Corneal stroma contains an extracellular matrix of orthogonal lamellae formed by parallel and equidistant fibrils with a homogeneous diameter of ~35 nm. This is indispensable for corneal transparency and mechanical functions. However, the mechanisms controlling corneal fibrillogenesis are incompletely understood and the conditions required for lamellar stacking are essentially unknown. Under appropriate conditions, chick embryo corneal fibroblasts can produce an extracellular matrix in vitro resembling primary corneal stroma during embryonic development. Among other requirements, cross-links between fibrillar collagens, introduced by tissue transglutaminase-2, are necessary for the self-assembly of uniform, small diameter fibrils but not their lamellar stacking. By contrast, the subsequent lamellar organization into plywood-like stacks depends on lysyl aldehyde-derived cross-links introduced by lysyl oxidase activity, which, in turn, only weakly influences fibril diameters. These cross-links are introduced at early stages of fibrillogenesis. The enzymes are likely to be important for a correct matrix deposition also during repair of the cornea. PMID:24265319

  1. Cytoskeletal organization and collagen orientation in the fish scales.

    PubMed

    Zylberberg, L; Bereiter-Hahn, J; Sire, J Y

    1988-09-01

    Immunofluorescence and electron microscopy were used to analyze the relationships between the organization of collagen fibrils in elasmoid scales, and the orientation of microtubules and actin microfilaments in the scleroblasts producing this collagenous stroma. Attention was focused on the basal plate of the scales because of the highly ordered three-dimensional arrangement of the collagen fibrils in superimposed plies forming an acellular plywood-like structure. The collagen fibrils are synthesized by the scleroblasts forming a monolayered pseudo-epithelium, the hyposquama, at the lowest surface of the scale. Fully developed scales with a low collagen deposition rate were compared with regenerating scales active in fibrillogenesis. When an ordered array of the collagen fibrils is found, the innermost collagen fibrils are coaligned with microtubules and actin microfilaments. Thus, because of this coalignment, microtubules and actin microfilaments of the hyposquamal scleroblasts are subjected to consecutive alterations during the formation of the plies of the basal plate. The sequence of events when the collagen fibrils change their direction from one ply to the other in the basal plate is deduced from immunofluorescence and phase-contrast-microscopic observations. During the formation of the orthogonal plywood-like structure in the regenerating scales, first microtubules may change their curse with a rotating angle of about 90 degrees; then, actin microfilaments are disorganized and reorganized by interacting mechanically with the microtubules with which they are coaligned. Collagen fibrils are synthesized in a direction that is roughly perpendicular to that of the preceding ply. The unknown signals inducing the change in direction of the cytoskeleton may be transmitted throughout the hyposquama via gap junctions. PMID:3052849

  2. Binding Modes of Thioflavin T on the Surface of Amyloid Fibrils Studied by NMR.

    PubMed

    Ivancic, Valerie A; Ekanayake, Oshini; Lazo, Noel D

    2016-08-18

    The mechanism for the interaction of thioflavin T (ThT) with amyloid fibrils at the molecular level is not known. Here, we used (1) H NMR spectroscopy to determine the binding mode of ThT on the surface of fibrils from lysozyme and insulin. Relayed rotating-frame Overhauser enhancements in ThT were observed, indicating that the orientation of ThT is orthogonal to the fibril surface. Importantly, the assembly state of ThT on both surfaces is different. On the surface of insulin fibrils, ThT is oligomeric, as indicated by rapid (1) H spin-lattice relaxation rate in the rotating frame (R1ρ ), presumably due to intermolecular dipole-dipole interactions between ThT molecules. In contrast, ThT on the surface of lysozyme fibrils is a monomer, as indicated by slower (1) H R1ρ . These results shed new light into the mechanism for the enhancement of ThT fluorescence and may lead to more efficient detectors of amyloid assemblies, which have escaped detection by ThT in monomer form. PMID:27165642

  3. Gold-Induced Fibril Growth: The Mechanism of Surface-Facilitated Amyloid Aggregation.

    PubMed

    Gladytz, Anika; Abel, Bernd; Risselada, Herre Jelger

    2016-09-01

    The question of how amyloid fibril formation is influenced by surfaces is crucial for a detailed understanding of the process in vivo. We applied a combination of kinetic experiments and molecular dynamics simulations to elucidate how (model) surfaces influence fibril formation of the amyloid-forming sequences of prion protein SUP35 and human islet amyloid polypeptide. The kinetic data suggest that structural reorganization of the initial peptide corona around colloidal gold nanoparticles is the rate-limiting step. The molecular dynamics simulations reveal that partial physisorption to the surface results in the formation of aligned monolayers, which stimulate the formation of parallel, critical oligomers. The general mechanism implies that the competition between the underlying peptide-peptide and peptide-surface interactions must strike a balance to accelerate fibril formation. PMID:27513605

  4. Interaction of Lubricin with Collagen II Surfaces: Adsorption, Friction, and Normal Forces

    PubMed Central

    Chang, Debby P.; Guilak, Farshid; Jay, Gregory; Zauscher, Stefan

    2014-01-01

    One of the major constituents of the synovial fluid that is thought to be responsible for chondroprotection and boundary lubrication is the glycoprotein lubricin (PRG4); however, the molecular mechanisms by which lubricin carries out its critical functions still remain largely unknown. We hypothesized that the interaction of lubricin with type II collagen, the main component of the cartilage extracellular matrix, results in enhanced tribological and wear properties. In this study, we examined: i) the molecular details by which lubricin interacts with type II collagen and how binding is related to boundary lubrication and adhesive interactions; and, ii) whether collagen structure can affect lubricin adsorption and its chondroprotective properties. We found that lubricin adsorbs strongly onto denatured, amorphous, and fibrillar collagen surfaces. Furthermore, we found large repulsive interactions between the collagen surfaces in presence of lubricin, which increased with increasing lubricin concentration. Lubricin attenuated the large friction and also the long-range adhesion between fibrillar collagen surfaces. Interestingly, lubricin adsorbed onto and mediated the frictional response between the denatured and native amorphous collagen surfaces equally and showed no preference on the supramolecular architecture of collagen. However, the coefficient of friction was lowest on fibrillar collagen in the presence of lubricin. We speculate that an important role of lubricin in mediating interactions at the cartilage surface is to attach to the cartilage surface and provide a protective coating that maintains the contacting surfaces in a sterically repulsive state. PMID:24406099

  5. Endothelial Cell Growth and Differentiation on Collagen-Immobilized Polycaprolactone Nanowire Surfaces.

    PubMed

    Leszczak, Victoria; Baskett, Dominique A; Popat, Ketul C

    2015-06-01

    The success of cardiovascular implants is associated with the development of an endothelium on material surface, critical to the prevention of intimal hyperplasia, calcification and thrombosis. A thorough understanding of the interaction between vascular endothelial cells and the biomaterial involved is essential in order to have a successful application which promotes healing and regeneration through integration with native tissue. In this study, we have developed collagen immobilized nanostructured surfaces with controlled arrays of high aspect ratio nanowires for the growth and maintenance of human microvascular endothelial cells (HMVECs). The nanowire surfaces were fabricated from polycaprolactone using a novel nanotemplating technique, and were immobilized with collagen utilizing an aminolysis method. The collagen immobilized nanowire surfaces were characterized using contact angle measurements, scanning electron microscopy and X-ray photoelectron spectroscopy. Human microvascular endothelial cells were used to evaluate the efficacy of the collagen immobilized nanowire surfaces to promote cell adhesion, proliferation, viability and differentiation. The results presented here indicate significantly higher cellular adhesion, proliferation and viability on nanowire and collagen immobilized surfaces as compared to the control surface. Further, HMVECs have a more elongated body and low shape factor on nanostructured surfaces. The differentiation potential of collagen immobilized nanowire surfaces was also evaluated by immunostaining and western blotting for key endothelial cell markers that are expressed when human microvascular endothelial cells are differentiated. Results indicate that expression of VE-cadherin is increased on collagen immobilized surfaces while the expression of von Willebrand factor is statistically similar on all surfaces. PMID:26353596

  6. In Situ Quantification of Surface Chemistry in Porous Collagen Biomaterials.

    PubMed

    Tzeranis, Dimitrios S; Soller, Eric C; Buydash, Melissa C; So, Peter T C; Yannas, Ioannis V

    2016-03-01

    Cells inside a 3D matrix (such as tissue extracellular matrix or biomaterials) sense their insoluble environment through specific binding interactions between their adhesion receptors and ligands present on the matrix surface. Despite the critical role of the insoluble matrix in cell regulation, there exist no widely-applicable methods for quantifying the chemical stimuli provided by a matrix to cells. Here, we describe a general-purpose technique for quantifying in situ the density of ligands for specific cell adhesion receptors of interest on the surface of a 3D matrix. This paper improves significantly the accuracy of the procedure introduced in a previous publication by detailed marker characterization, optimized staining, and improved data interpretation. The optimized methodology is utilized to quantify the ligands of integrins α 1 β 1, α 2 β 1 on two kinds of matched porous collagen scaffolds, which are shown to possess significantly different ligand density, and significantly different ability to induce peripheral nerve regeneration in vivo. Data support the hypothesis that cell adhesion regulates contractile cell phenotypes, recently shown to be inversely related to organ regeneration. The technique provides a standardized way to quantify the surface chemistry of 3D matrices, and a means for introducing matrix effects in quantitative biological models. PMID:26369635

  7. Identification of novel short peptides derived from the {alpha}4, {alpha}5, and {alpha}6 fibrils of type IV collagen with anti-angiogenic properties

    SciTech Connect

    Karagiannis, Emmanouil D. . E-mail: ekaragi1@jhmi.edu; Popel, Aleksander S.

    2007-03-09

    Angiogenesis, or neovascularization, is tightly controlled by positive and negative regulators, many of which reside in the extracellular matrix. We have now identified eight novel 19- to 20-residue peptides derived from the {alpha}4, {alpha}5, and {alpha}6 fibrils of type IV collagen, which we have designated tetrastatins, pentastatins, and hexastatins, respectively. We have shown that these endogenous peptides suppress the proliferation and migration of HUVECs in vitro. By performing clustering analyses of the sequences using sequence similarity criteria and of the experimental results using a hierarchical algorithm, we report that the clusters identified by the experimental results coincide with the sequence-based clusters, indicating a tight relationship between peptide sequence and anti-angiogenic potency. These peptides may have potential as anti-angiogenic therapeutic agents.

  8. Nanomechanics of Type I Collagen.

    PubMed

    Varma, Sameer; Orgel, Joseph P R O; Schieber, Jay D

    2016-07-12

    Type I collagen is the predominant collagen in mature tendons and ligaments, where it gives them their load-bearing mechanical properties. Fibrils of type I collagen are formed by the packing of polypeptide triple helices. Higher-order structures like fibril bundles and fibers are assembled from fibrils in the presence of other collagenous molecules and noncollagenous molecules. Curiously, however, experiments show that fibrils/fibril bundles are less resistant to axial stress compared to their constituent triple helices-the Young's moduli of fibrils/fibril bundles are an order-of-magnitude smaller than the Young's moduli of triple helices. Given the sensitivity of the Young's moduli of triple helices to solvation environment, a plausible explanation is that the packing of triple helices into fibrils perhaps reduces the Young's modulus of an individual triple helix, which results in fibrils having smaller Young's moduli. We find, however, from molecular dynamics and accelerated conformational sampling simulations that the Young's modulus of the buried core of the fibril is of the same order as that of a triple helix in aqueous phase. These simulations, therefore, suggest that the lower Young's moduli of fibrils/fibril bundles cannot be attributed to the specific packing of triple helices in the fibril core. It is not the fibril core that yields initially to axial stress. Rather, it must be the portion of the fibril exposed to the solvent and/or the fibril-fibril interface that bears the initial strain. Overall, this work provides estimates of Young's moduli and persistence lengths at two levels of collagen's structural assembly, which are necessary to quantitatively investigate the response of various biological factors on collagen mechanics, including congenital mutations, posttranslational modifications and ligand binding, and also engineer new collagen-based materials. PMID:27410733

  9. Three-dimensional collagen architecture in bovine articular cartilage.

    PubMed

    Jeffery, A K; Blunn, G W; Archer, C W; Bentley, G

    1991-09-01

    The three-dimensional architecture of bovine articular cartilage collagen and its relationship to split lines has been studied with scanning electron microscopy. In the middle and superficial zones, collagen was organised in a layered or leaf-like manner. The orientation was vertical in the intermediate zone, curving to become horizontal and parallel to the articular surface in the superficial zone. Each leaf consisted of a fine network of collagen fibrils. Adjacent leaves merged or were closely linked by bridging fibrils and were arranged according to the split-line pattern. The surface layer (lamina splendens) was morphologically distinct. Although ordered, the overall collagen structure was different in each plane (anisotropic) a property described in previous morphological and biophysical studies. As all components of the articular cartilage matrix interact closely, the three-dimensional organisation of collagen is important when considering cartilage function and the processes of cartilage growth, injury and repair. PMID:1894669

  10. COLLAGEN STRUCTURE AND STABILITY

    PubMed Central

    Shoulders, Matthew D.; Raines, Ronald T.

    2010-01-01

    Collagen is the most abundant protein in animals. This fibrous, structural protein comprises a right-handed bundle of three parallel, left-handed polyproline II-type helices. Much progress has been made in elucidating the structure of collagen triple helices and the physicochemical basis for their stability. New evidence demonstrates that stereoelectronic effects and preorganization play a key role in that stability. The fibrillar structure of type I collagen–the prototypical collagen fibril–has been revealed in detail. Artificial collagen fibrils that display some properties of natural collagen fibrils are now accessible using chemical synthesis and self-assembly. A rapidly emerging understanding of the mechanical and structural properties of native collagen fibrils will guide further development of artificial collagenous materials for biomedicine and nanotechnology. PMID:19344236

  11. Type I Collagen and Collagen Mimetics as Angiogenesis Promoting Superpolymers

    SciTech Connect

    Twardowski, T.; Fertala, A.; Orgel, J.P.R.O.; San Antonio, J.D.

    2008-07-18

    Angiogenesis, the development of blood vessels from the pre-existing vasculature, is a key component of embryogenesis and tissue regeneration. Angiogenesis also drives pathologies such as tumor growth and metastasis, and hemangioma development in newborns. On the other hand, promotion of angiogenesis is needed in tissues with vascular insufficiencies, and in bioengineering, to endow tissue substitutes with appropriate microvasculatures. Therefore, much research has focused on defining mechanisms of angiogenesis, and identifying pro- and anti-angiogenic molecules. Type I collagen, the most abundant protein in humans, potently stimulates angiogenesis in vitro and in vivo. Crucial to its angiogenic activity appears to be ligation and possibly clustering of endothelial cell (EC) surface {alpha}1{beta}1/{alpha}2{beta}1 integrin receptors by the GFPGER502-507 sequence of the collagen fibril. However, additional aspects of collagen structure and function that may modulate its angiogenic properties are discussed. Moreover, type I collagen and fibrin, another angiogenic polymer, share several structural features. These observations suggest strategies for creating 'angiogenic superpolymers', including: modifying type I collagen to influence its biological half-life, immunogenicity, and integrin binding capacity; genetically engineering fibrillar collagens to include additional integrin binding sites or angiogenic determinants, and remove unnecessary or deleterious sequences without compromising fibril integrity; and exploring the suitability of poly(ortho ester), PEG-lysine copolymer, tubulin, and cholesteric cuticle as collagen mimetics, and suggesting means of modifying them to display ideal angiogenic properties. The collagenous and collagen mimetic angiogenic superpolymers described here may someday prove useful for many applications in tissue engineering and human medicine.

  12. Reaction efficiency and retention of poly(styrene-co-maleimide) nanoparticles deposited on fibrillated cellulose surfaces.

    PubMed

    Rastogi, Vibhore Kumar; Stanssens, Dirk; Samyn, Pieter

    2016-05-01

    Surface modification of micro- and nanofibrillated cellulose (MFC and NFC) under aqueous environment was performed by deposition of poly(styrene-co-maleimide) nanoparticles synthesized by imidization of poly(styrene-co-maleic anhydride) in presence of wax and ammonium hydroxide in variable amounts. Specifically, the influences of fiber fibrillation on nanoparticle formation (i.e., reaction efficiency) and permanent nanoparticle deposition on the fiber surface (i.e., retention) were investigated. The surface modification was mainly governed by the fiber diameter, surface charges and amount of wax. As such, the MFC affected the imidization reaction to a smaller extent (i.e., high reaction efficiency) and was more densely deposited by nanoparticles than NFC (i.e., high retention). Moreover, wax protected the fibers against fibrillation and peeling-off at high temperature and favored nanoparticle deposition. As a result, water contact angles of 142° were obtained for modified MFC in parallel with a surface coverage of 92%. PMID:26877019

  13. Dynamic shear-influenced collagen self-assembly.

    PubMed

    Saeidi, Nima; Sander, Edward A; Ruberti, Jeffrey W

    2009-12-01

    The ability to influence the direction of polymerization of a self-assembling biomolecular system has the potential to generate materials with extremely high anisotropy. In biological systems where highly-oriented cellular populations give rise to aligned and often load-bearing tissue such organized molecular scaffolds could aid in the contact guidance of cells for engineered tissue constructs (e.g. cornea and tendon). In this investigation we examine the detailed dynamics of pepsin-extracted type I bovine collagen assembly on a glass surface under the influence of flow between two plates. Differential Interference Contrast (DIC) imaging (60x-1.4NA) with focal plane stabilization was used to resolve and track the growth of collagen aggregates on borosilicate glass for 4 different shear rates (500, 80, 20, and 9s(-1)). The detailed morphology of the collagen fibrils/aggregates was examined using Quick Freeze Deep Etch (QFDE) electron microscopy. Nucleation of fibrils on the glass was observed to occur rapidly (approximately 2 min) followed by continued growth of the fibrils. The growth rates were dependent on flow in a complex manner with the highest rate of axial growth (0.1 micro/s) occurring at a shear rate of 9s(-1). The lowest growth rate occurred at the highest shear. Fibrils were observed to both branch and join during the experiments. The best alignment of fibrils was observed at intermediate shear rates of 20 and 80s(-1). However, the investigation revealed that fibril directional growth was not stable. At high shear rates, fibrils would often turn downstream forming what we term "hooks" which are likely the combined result of monomer interaction with the initial collagen layer or "mat" and the high shear rate. Further, QFDE examination of fibril morphology demonstrated that the assembled fibrillar structure did not possess native D-periodicity. Instead, fibrils comprised a collection of generally aligned, monomers which were self-assembled to form a fibril

  14. The initial attachment of cemental fibrils to the root dentin surface in acellular and cellular cementogenesis in rat molars.

    PubMed

    Yamamoto, T; Domon, T; Takahashi, S; Islam, M N; Suzuki, R

    2001-03-01

    To elucidate the initial attachment mechanism of cemental fibrils to the root dentin surface in acellular and cellular cementogenesis, developing rat molars were observed by light microscopy and scanning electron microscopy combined with NaOH maceration. The NaOH maceration was used to observe details of the positional association of cemental and dentinal fibrils during cementogenesis. An initial hematoxylin stained, cementum layer began to form on the root dentin surface with the first dentin mineralization in both acellular and cellular cementogenesis. The initial attachment of cemental fibrils to the dentin surface also began at this point. At the initial attachment the intermingling of cemental and dentinal fibrils occurred only in places. With advanced cementogenesis the initial cementum layer became the fibril-poor cemento-dentinal junction. This suggests that cemental fibrils attach on the initial cementum layer, and not directly on dentinal fibrils, so that the layer results in the fibril-poor cemento-dentinal junction. The present study suggests that an intervening adhesive is necessary for the cemento-dentinal attachment at any stage of cementogenesis in rat molars. PMID:11325058

  15. Collagen IX is required for the integrity of collagen II fibrils and the regulation of vascular plexus formation in zebrafish caudal fins.

    PubMed

    Huang, Cheng-chen; Wang, Tai-Chuan; Lin, Bo-Hung; Wang, Yi-Wen; Johnson, Stephen L; Yu, John

    2009-08-15

    Capillary plexuses form during both vasculogenesis and angiogenesis and are remodeled into mature vessel types and patterns which are delicately orchestrated with the sizes and shapes of other tissues and organs. We isolated a zebrafish mutation named prp (for persistent plexus) that causes persistent formation of vascular plexuses in the caudal fins and consequent mispatterning of bony fin rays and the fin shape. Detailed analyses revealed that the prp mutation causes a significant reduction in the size and dramatic structural defects in collagen II-rich extracellular matrices called actinotrichia of both embryonic finfolds and adult fins. prp was mapped to chromosome 19 and found to encode the zebrafish collagen9alpha1 (col9alpha1) gene which is abundantly expressed in developing finfolds. A point mutation resulting in a leucine-to-histidine change was detected in the thrombospondin domain of the col9alpha1 gene in prp. Morpholino-mediated knockdown of col9alpha1 phenocopied the prp small-finfold phenotype in wild-type embryos, and an injection of plasmids containing the col9alpha1 cDNA into prp embryos locally restored the finfold size. Furthermore, we found that osteoblasts in prp mutants were mispatterned apparently following the abnormal vascular plexus pattern, demonstrating that blood vessels play an important role in the patterning of bony rays in zebrafish caudal fins. PMID:19501583

  16. Collagen interactions: Drug design and delivery.

    PubMed

    An, Bo; Lin, Yu-Shan; Brodsky, Barbara

    2016-02-01

    Collagen is a major component in a wide range of drug delivery systems and biomaterial applications. Its basic physical and structural properties, together with its low immunogenicity and natural turnover, are keys to its biocompatibility and effectiveness. In addition to its material properties, the collagen triple-helix interacts with a large number of molecules that trigger biological events. Collagen interactions with cell surface receptors regulate many cellular processes, while interactions with other ECM components are critical for matrix structure and remodeling. Collagen also interacts with enzymes involved in its biosynthesis and degradation, including matrix metalloproteinases. Over the past decade, much information has been gained about the nature and specificity of collagen interactions with its partners. These studies have defined collagen sequences responsible for binding and the high-resolution structures of triple-helical peptides bound to its natural binding partners. Strategies to target collagen interactions are already being developed, including the use of monoclonal antibodies to interfere with collagen fibril formation and the use of triple-helical peptides to direct liposomes to melanoma cells. The molecular information about collagen interactions will further serve as a foundation for computational studies to design small molecules that can interfere with specific interactions or target tumor cells. Intelligent control of collagen biological interactions within a material context will expand the effectiveness of collagen-based drug delivery. PMID:26631222

  17. Surface adsorption considerations when working with amyloid fibrils in multiwell plates and Eppendorf tubes.

    PubMed

    Murray, Amber N; Palhano, Fernando L; Bieschke, Jan; Kelly, Jeffery W

    2013-11-01

    The accumulation of cross-β-sheet amyloid fibrils is the hallmark of amyloid diseases. Recently, we reported the discovery of amyloid disaggregase activities in extracts from mammalian cells and Caenorhabditis elegans. However, we have discovered a problem with the interpretation of our previous results as Aβ disaggregation in vitro. Here, we show that Aβ fibrils adsorb to the plastic surface of multiwell plates and Eppendorf tubes. This adsorption is markedly increased in the presence of complex biological mixtures subjected to a denaturing air-water interface. The time-dependent loss of thioflavin T fluorescence that we interpreted previously as disaggregation is due to increased adsorption of Aβ amyloid to the surfaces of multiwell plates and Eppendorf tubes in the presence of biological extracts. As the proteins in biological extracts denature over time at the air-water interface due to agitation/shaking, their adsorption increases, in turn promoting adsorption of amyloid fibrils. We delineate important control experiments that quantify the extent of amyloid adsorption to the surface of plastic and quartz containers. Based on the results described in this article, we conclude that our interpretation of the kinetic fibril disaggregation assay data previously reported in Bieschke et al., Protein Sci 2009;18:2231-2241 and Murray et al., Protein Sci 2010;19:836-846 is invalid when used as evidence for a disaggregase activity. Thus, we correct the two prior publications reporting that worm or mammalian cell extracts disaggregate Aβ amyloid fibrils in vitro at 37°C (see Corrigenda in this issue of Protein Science). We apologize for misinterpreting our previous data and for any confounding experimental efforts this may have caused. PMID:23963844

  18. Expression of type I and type V collagen mRNAs in the elasmoid scales of a teleost fish as revealed by in situ hybridization.

    PubMed

    Le Guellec, D; Zylberberg, L

    1998-01-01

    The ability of scale-forming cells to produce both type I and type V collagens was investigated by in situ hybridization at the light and electron microscope levels. Biochemical analyses reported that type I collagen, the predominant component, was associated with the minor type V collagen in the collagenous matrix of the teleost scales where, thin and thick collagen fibrils formed distinct layers. Thin collagen fibrils of the external layer were produced by the episquamal scleroblasts scattered on the outer scale surface, while thick collagen fibrils forming the compact basal plate were produced by the hyposquamal scleroblasts lining the inner surface of the scale. We demonstrated that episquamal and hyposquamal scleroblasts contained mRNAs for alpha1(I) and alpha1(V) collagens. Quantification by image analysis of the relative amount of alpha1(I) and alpha1(V) mRNAs in episquamal and hyposquamal scleroblasts suggests that the gene expression of type V collagen was proportionally higher in episquamal scleroblasts. These results support our hypothesis that the diameter of the thin fibrils of the external layer is regulated by the significant amount of type V collagen that interacts with type I collagen. PMID:11063006

  19. Surface characterization of collagen/elastin based biomaterials for tissue regeneration

    NASA Astrophysics Data System (ADS)

    Skopinska-Wisniewska, J.; Sionkowska, A.; Kaminska, A.; Kaznica, A.; Jachimiak, R.; Drewa, T.

    2009-07-01

    Collagen and elastin are the main proteins of extracellular matrix. Collagen plays a crucial role in tensile strength of tissues, whereas elastin provides resilience to many organs. Both biopolymers are readily available and biocompatible. These properties point out that collagen and elastin are good components of materials for many potential medical applications. The surface properties of biomaterials play an important role in biomedicine as the majority of biological reactions occur on the surface of implanted materials. One of the methods of surface modification is UV-irradiation. The exposition of the biomaterial on ultraviolet light can alterate surface properties of the materials, their chemical stability, swelling properties and mechanical properties as well. The aim of our work was to study the surface properties and biocompatibility of new collagen/elastin based biomaterials and consideration of the influence of ultraviolet light on these properties. The surface properties of collagen/elastin based biomaterials modified by UV-irradiation were studied using the technique of atomic force microscopy (AFM) and contact angle measurements. On the basis of the results the surface free energy and its polar component was calculated using Owens-Wendt method. To assess the biological performance of films based on collagen, elastin and their blends, the response of 3T3 cell was investigated. It was found that the surface of collagen/elastin film is enriched in less polar component - collagen. Exposition on UV light increases polarity of collagen/elastin based films, due to photooxidation process. The AFM images have shown that topography and roughness of the materials had been also affected by UV-irradiation. The changes in surface properties influence on interaction between the material's surface and cells. The investigation of 3T3 cells grown on films based on collagen, elastin and their blends, leads to the conclusion that higher content of elastin in biomaterial

  20. Fibulin-4 E57K Knock-in Mice Recapitulate Cutaneous, Vascular and Skeletal Defects of Recessive Cutis Laxa 1B with both Elastic Fiber and Collagen Fibril Abnormalities.

    PubMed

    Igoucheva, Olga; Alexeev, Vitali; Halabi, Carmen M; Adams, Sheila M; Stoilov, Ivan; Sasaki, Takako; Arita, Machiko; Donahue, Adele; Mecham, Robert P; Birk, David E; Chu, Mon-Li

    2015-08-28

    Fibulin-4 is an extracellular matrix protein essential for elastic fiber formation. Frameshift and missense mutations in the fibulin-4 gene (EFEMP2/FBLN4) cause autosomal recessive cutis laxa (ARCL) 1B, characterized by loose skin, aortic aneurysm, arterial tortuosity, lung emphysema, and skeletal abnormalities. Homozygous missense mutations in FBLN4 are a prevalent cause of ARCL 1B. Here we generated a knock-in mouse strain bearing a recurrent fibulin-4 E57K homozygous missense mutation. The mutant mice survived into adulthood and displayed abnormalities in multiple organ systems, including loose skin, bent forelimb, aortic aneurysm, tortuous artery, and pulmonary emphysema. Biochemical studies of dermal fibroblasts showed that fibulin-4 E57K mutant protein was produced but was prone to dimer formation and inefficiently secreted, thereby triggering an endoplasmic reticulum stress response. Immunohistochemistry detected a low level of fibulin-4 E57K protein in the knock-in skin along with altered expression of selected elastic fiber components. Processing of a precursor to mature lysyl oxidase, an enzyme involved in cross-linking of elastin and collagen, was compromised. The knock-in skin had a reduced level of desmosine, an elastin-specific cross-link compound, and ultrastructurally abnormal elastic fibers. Surprisingly, structurally aberrant collagen fibrils and altered organization into fibers were characteristics of the knock-in dermis and forelimb tendons. Type I collagen extracted from the knock-in skin had decreased amounts of covalent intermolecular cross-links, which could contribute to the collagen fibril abnormalities. Our studies provide the first evidence that fibulin-4 plays a role in regulating collagen fibril assembly and offer a preclinical platform for developing treatments for ARCL 1B. PMID:26178373

  1. Self-healing Characteristics of Collagen Coatings with Respect to Surface Abrasion

    PubMed Central

    Kim, Chang-Lae; Kim, Dae-Eun

    2016-01-01

    A coating based on collagen with self-healing properties was developed for applications in mechanical components that are prone to abrasion due to contact with a counter surface. The inherent swelling behavior of collagen in water was exploited as the fundamental mechanism behind self-healing of a wear scar formed on the surface. The effects of freeze-drying process and water treatment of the collagen coatings on their mechanical and self-healing properties were analyzed. Water was also used as the medium to trigger the self-healing effect of the collagen coatings after the wear test. It was found that collagen coatings without freeze-drying did not demonstrate any self-healing effect whereas the coatings treated by freeze-drying process showed remarkable self-healing effect. Overall, collagen coatings that were freeze-dried and water treated showed the best friction and self-healing properties. Repeated self-healing ability of these coatings with respect to wear scar was also demonstrated. It was also confirmed that the self-healing property of the collagen coating was effective over a relatively wide range of temperature. PMID:27010967

  2. Self-healing Characteristics of Collagen Coatings with Respect to Surface Abrasion

    NASA Astrophysics Data System (ADS)

    Kim, Chang-Lae; Kim, Dae-Eun

    2016-03-01

    A coating based on collagen with self-healing properties was developed for applications in mechanical components that are prone to abrasion due to contact with a counter surface. The inherent swelling behavior of collagen in water was exploited as the fundamental mechanism behind self-healing of a wear scar formed on the surface. The effects of freeze-drying process and water treatment of the collagen coatings on their mechanical and self-healing properties were analyzed. Water was also used as the medium to trigger the self-healing effect of the collagen coatings after the wear test. It was found that collagen coatings without freeze-drying did not demonstrate any self-healing effect whereas the coatings treated by freeze-drying process showed remarkable self-healing effect. Overall, collagen coatings that were freeze-dried and water treated showed the best friction and self-healing properties. Repeated self-healing ability of these coatings with respect to wear scar was also demonstrated. It was also confirmed that the self-healing property of the collagen coating was effective over a relatively wide range of temperature.

  3. Revealing molecular-level surface structure of amyloid fibrils in liquid by means of frequency modulation atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Fukuma, Takeshi; Mostaert, Anika S.; Serpell, Louise C.; Jarvis, Suzanne P.

    2008-09-01

    We have investigated the surface structure of islet amyloid polypeptide (IAPP) fibrils and α-synuclein protofibrils in liquid by means of frequency modulation atomic force microscopy (FM-AFM). Ångström-resolution FM-AFM imaging of isolated macromolecules in liquid is demonstrated for the first time. Individual β-strands aligned perpendicular to the fibril axis with a spacing of 0.5 nm are resolved in FM-AFM images, which confirms cross-β structure of IAPP fibrils in real space. FM-AFM images also reveal the existence of 4 nm periodic domains along the axis of IAPP fibrils. Stripe features with 0.5 nm spacing are also found in images of α-synuclein protofibrils. However, in contrast to the case for IAPP fibrils, the stripes are oriented 30° from the axis, suggesting the possibility of β-strand alignment in protofibrils different from that in mature fibrils or the regular arrangement of thioflavin T molecules present during the fibril preparation aligned at the surface of the protofibrils.

  4. Surface engineering of stainless steel materials by covalent collagen immobilization to improve implant biocompatibility.

    PubMed

    Müller, Rainer; Abke, Jochen; Schnell, Edith; Macionczyk, Frank; Gbureck, Uwe; Mehrl, Robert; Ruszczak, Zbigniev; Kujat, Richard; Englert, Carsten; Nerlich, Michael; Angele, Peter

    2005-12-01

    It was shown recently that the deposition of thin films of tantalum and tantalum oxide enhanced the long-term biocompatibility of stainless steel biomaterials due to an increase in their corrosion resistance. In this study, we used this tantalum oxide coating as a basis for covalent immobilization of a collagen layer, which should result in a further improvement of implant tissue integration. Because of the high degradation rate of natural collagen in vivo, covalent immobilization as well as carbodiimide induced cross-linking of the protein was performed. It was found that the combination of the silane-coupling agent aminopropyl triethoxysilane and the linker molecule N,N'-disulphosuccinimidyl suberate was a very effective system for collagen immobilizing. Mechanical and enzymatic stability testing revealed a higher stability of covalent bound collagen layers compared to physically adsorbed collagen layers. The biological response induced by the surface modifications was evaluated by in vitro cell culture with human mesenchymal stem cells as well as by in vivo subcutaneous implantation into nude mice. The presence of collagen clearly improved the cytocompatibility of the stainless steel implants which, nevertheless, significantly depended on the cross-linking degree of the collagen layer. PMID:15967497

  5. Surface engineering of PHBV by covalent collagen immobilization to improve cell compatibility.

    PubMed

    Wang, Yingjun; Ke, Yu; Ren, Li; Wu, Gang; Chen, Xiaofeng; Zhao, Qichun

    2009-03-01

    Covalent immobilization of collagen onto poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) film was achieved to improve its cell compatibility. Amide groups photografted on PHBV films were initially converted into amine groups through Hofmann degradation and collagen was then chemically bonded to amine groups, consequently forming the amide, amine, and collagen-modified PHBV. The structures of these modified PHBV films were confirmed by ATR-FTIR, XPS, and SEM analyses. Compared with that of PHBV film, surface wettability of the modified PHBV films enhanced remarkably. In particular, water contact angle of the collagen-modified PHBV film decreased from 65.0 degrees to 2.1 degrees within 130 s. Sheep chondrocytes cultured on PHBV and modified PHBV films were evaluated by cell adhesion test, MTT assay, and morphological observation under SEM. Results showed that the collagen-modified PHBV film had better cell adhesion and proliferation than other modified PHBV films and PHBV film. Chondrocytes on the collagen-modified PHBV film adhered through filopodia, spread by cytoplasmic webbing, and formed cells layer earlier than other modified ones, indicating that the collagen-modified PHBV is a promising biomaterial for cartilage tissue engineering. PMID:18314894

  6. Surface-Driven Collagen Self-Assembly Affects Early Osteogenic Stem Cell Signaling.

    PubMed

    Razafiarison, Tojo; Silván, Unai; Meier, Daniela; Snedeker, Jess G

    2016-06-01

    This study reports how extracellular matrix (ECM) ligand self-assembly on biomaterial surfaces and the resulting nanoscale architecture can drive stem cell behavior. To isolate the biological effects of surface wettability on protein deposition, folding, and ligand activity, a polydimethylsiloxane (PDMS)-based platform was developed and characterized with the ability to tune wettability of elastomeric substrates with otherwise equivalent topology, ligand loading, and mechanical properties. Using this platform, markedly different assembly of covalently bound type I collagen monomers was observed depending on wettability, with hydrophobic substrates yielding a relatively rough layer of collagen aggregates compared to a smooth collagen layer on more hydrophilic substrates. Cellular and molecular investigations with human bone marrow stromal cells revealed higher osteogenic differentiation and upregulation of focal adhesion-related components on the resulting smooth collagen layer coated substrates. The initial collagen assembly driven by the PDMS surface directly affected α1β1 integrin/discoidin domain receptor 1 signaling, activation of the extracellular signal-regulated kinase/mitogen activated protein kinase pathway, and ultimately markers of osteogenic stem cell differentiation. We demonstrate for the first time that surface-driven ligand assembly on material surfaces, even on materials with otherwise identical starting topographies and mechanical properties, can dominate the biomaterial surface-driven cell response. PMID:27125602

  7. Molecular crowding of collagen: a pathway to produce highly-organized collagenous structures.

    PubMed

    Saeidi, Nima; Karmelek, Kathryn P; Paten, Jeffrey A; Zareian, Ramin; DiMasi, Elaine; Ruberti, Jeffrey W

    2012-10-01

    Collagen in vertebrate animals is often arranged in alternating lamellae or in bundles of aligned fibrils which are designed to withstand in vivo mechanical loads. The formation of these organized structures is thought to result from a complex, large-area integration of individual cell motion and locally-controlled synthesis of fibrillar arrays via cell-surface fibripositors (direct matrix printing). The difficulty of reproducing such a process in vitro has prevented tissue engineers from constructing clinically useful load-bearing connective tissue directly from collagen. However, we and others have taken the view that long-range organizational information is potentially encoded into the structure of the collagen molecule itself, allowing the control of fibril organization to extend far from cell (or bounding) surfaces. We here demonstrate a simple, fast, cell-free method capable of producing highly-organized, anistropic collagen fibrillar lamellae de novo which persist over relatively long-distances (tens to hundreds of microns). Our approach to nanoscale organizational control takes advantage of the intrinsic physiochemical properties of collagen molecules by inducing collagen association through molecular crowding and geometric confinement. To mimic biological tissues which comprise planar, aligned collagen lamellae (e.g. cornea, lamellar bone or annulus fibrosus), type I collagen was confined to a thin, planar geometry, concentrated through molecular crowding and polymerized. The resulting fibrillar lamellae show a striking resemblance to native load-bearing lamellae in that the fibrils are small, generally aligned in the plane of the confining space and change direction en masse throughout the thickness of the construct. The process of organizational control is consistent with embryonic development where the bounded planar cell sheets produced by fibroblasts suggest a similar confinement/concentration strategy. Such a simple approach to nanoscale

  8. Configurational effects of collagen/ALP coatings on enzyme immobilization and surface mineralization

    NASA Astrophysics Data System (ADS)

    Bosco, R.; Leeuwenburgh, S. C. G.; Jansen, J. A.; van den Beucken, J. J. J. P.

    2014-08-01

    The ultimate goal for surface modifications in bone implants is to achieve biologically active surface able to control and trigger specific tissue response. In this study was evaluated the effects of organic compound, derived from extracellular matrix, involved in tissue mineralization. Alkaline phosphatase (ALP) plays a fundamental role in bone mineralization concurrently with collagen, the main organic components of bones. Electrospray deposition (ESD) was used to coat titanium disks with ALP and collagen at room temperature. To verify the synergistic role of ALP and collagen different conformations of coatings (mixed and layered) were obtained and their mineralization capacity was tested in vitro. The mineralization tests indicated the fundamental role of collagen to increase ALP coating retention. Analyses indicated that the coating conformation has a role; in fact the mixed group showed improved ALP retention, enzymatic activity and unique mineralized surface morphology. ESD demonstrated to be a successful method to deposit organic molecules preserving their properties as indicated by the in vitro results. These findings proved the synergistic effect of ALP and collagen in inducing mineralization offering an intriguing coating constituent for medical device that aim to trigger surface mineralization such as bone implants.

  9. Thermoresponsive poly(N-isopropyl acrylamide)-grafted polycaprolactone films with surface immobilization of collagen.

    PubMed

    Xu, F J; Zheng, Y Q; Zhen, W J; Yang, W T

    2011-06-15

    Thermoresponsive poly(N-isopropylacrylamide) (P(NIPAAm))-grafted polycaprolactone (PCL) films with a suitable amount of immobilized cell-adhesive collagen were prepared to improve cell adhesion and proliferation above the lower critical solution temperature (LCST, 32°C) of P(NIPAAm) without destroying cell detachment properties at lower temperatures. Covalently tethered P(NIPAAm) brushes on PCL film surfaces were first prepared via surface-initiated atom transfer radical polymerization (ATRP). The alkyl bromide end groups of the grafted P(NIPAAm) brushes were used in nucleophilic substitution reactions for the direct coupling of collagen to produce the collagen-immobilized thermoresponsive PCL surface. At 37°C, the cell attachments on the collagen-immobilized thermoresponsive PCL surface were enhanced substantially. The attached cells could be recovered simply by lowering culture temperature. The P(NIPAAm)-grafted PCL films with immobilized collagen are potentially useful as adhesion modifiers for advanced cell culture and tissue engineering applications. PMID:20980132

  10. Northern pike (Esox lucius) collagen: Extraction, characterization and potential application.

    PubMed

    Kozlowska, J; Sionkowska, A; Skopinska-Wisniewska, J; Piechowicz, K

    2015-11-01

    Acid soluble collagen (ASC) and pepsin soluble collagen (PSC) from the scales of northern pike (Esox lucius) were extracted and characterized. It was the first time that this species was used as sources of collagen. FT-IR and amino acid analysis results revealed the presence of collagen. Glycine accounts for one-third of its amino acid residues and specific for collagen amino acid - hydroxyproline - is present in isolated protein. The content of imino acid: proline and hydroxyproline in ASC and PSC was similar (12.5% Pro and 6.5% Hyp). Both ASC and PSC were type I collagen. The denaturation temperature of ASC and PSC were 28.5 and 27°C, respectively. Thin collagen films were obtained by casting of collagen solution onto glass plates. The surface properties of ASC and PSC films were different - the surface of ASC collagen film was more polar and less rough than PSC and we can observe the formation of collagen fibrils after solvent evaporation. ASC films showed much higher tensile properties than PSC. The obtained results suggest that northern pike scales have potential as an alternative source of collagen for use in various fields. PMID:26254247

  11. Effect of Surface Chemical Functionalities on Collagen Deposition by Primary Human Dermal Fibroblasts.

    PubMed

    Bachhuka, Akash; Hayball, John; Smith, Louise E; Vasilev, K

    2015-10-28

    Surface modification has been identified as an important technique that could improve the response of the body to implanted medical devices. Collagen production by fibroblasts is known to play a vital role in wound healing and device fibrous encapsulation. However, how surface chemistry affects collagen I and III deposition by these cells has not been systematically studied. Here, we report how surface chemistry influences the deposition of collagen I and III by primary human dermal fibroblasts. Amine (NH3), carboxyl acid (COOH), and hydrocarbon (CH3) surfaces were generated by plasma deposition. This is a practically relevant tool to deposit a functional coating on any type of substrate material. We show that fibroblasts adhere better and proliferate faster on amine-rich surfaces. In addition, the initial collagen I and III production is greater on this type of coating. These data indicates that surface modification can be a promising route for modulating the rate and level of fibrous encapsulation and may be useful in informing the design of implantable biomedical devices to produce more predictable clinical outcomes. PMID:26457649

  12. Surface modification of PVDF using non-mammalian sources of collagen for enhancement of endothelial cell functionality.

    PubMed

    Wang, Jun Kit; Xiong, Gordon Minru; Luo, Baiwen; Choo, Chee Chong; Yuan, Shaojun; Tan, Nguan Soon; Choong, Cleo

    2016-03-01

    Although polyvinylidene fluoride (PVDF) is non-toxic and stable in vivo, its hydrophobic surface has limited its bio-applications due to poor cell-material interaction and thrombus formation when used in blood contacting devices. In this study, surface modification of PVDF using naturally derived non-mammalian collagen was accomplished via direct surface-initiated atom transfer radical polymerisation (SI-ATRP) to enhance its cytocompatibility and hemocompatibility. Results showed that Type I collagen was successfully extracted from fish scales and bullfrog skin. The covalent immobilisation of fish scale-derived collagen (FSCOL) and bullfrog skin-derived collagen (BFCOL) onto the PVDF surface improves the attachment and proliferation of human umbilical vein endothelial cells (HUVECs). Furthermore, both FSCOL and BFCOL had comparable anti-thrombogenic profiles to that of commercially available bovine collagen (BVCOL). Also, cell surface expression of the leukocyte adhesion molecule was lower on HUVECs cultured on non-mammalian collagen surfaces than on BVCOL, which is an indication of lower pro-inflammatory response. Overall, results from this study demonstrated that non-mammalian sources of collagen could be used to confer bioactivity to PVDF, with comparable cell-material interactions and hemocompatibility to BVCOL. Additionally, higher expression levels of Type IV collagen in HUVECs cultured on FSCOL and BFCOL were observed as compared to BVCOL, which is an indication that the non-mammalian sources of collagen led to a better pro-angiogenic properties, thus making them suitable for blood contacting applications. PMID:26758892

  13. Post-translational control of collagen fibrillogenesis in mineralizing cultures of chick osteoblasts

    NASA Technical Reports Server (NTRS)

    Gerstenfeld, L. C.; Riva, A.; Hodgens, K.; Eyre, D. R.; Landis, W. J.

    1993-01-01

    Cultured osteoblasts from chick embryo calvaria were used as a model system to investigate the post-translational extracellular mechanisms controlling the macroassembly of collagen fibrils. The results of these studies demonstrated that cultured osteoblasts secreted a collagenous extracellular matrix that assembled and mineralized in a defined temporal and spatial sequence. The assembly of collagen occurred in a polarized fashion, such that successive orthogonal arrays of fibrils formed between successive cell layers proceeding from the culture surface toward the media. Mineralization followed in the same manner, being observed first in the deepest and oldest fibril layers. Collagen fibrillogenesis, the kinetics of cross-link formation, and collagen stability in the extracellular matrix of the cultures were examined over a 30 day culture period. Between days 8 and 12 in culture, collagen fibril diameters increased from < 30 nm to an average of 30-45 nm. Thereafter, diameters ranged in size from 20 to 200 nm. Quantitation of the collagen cross-linking residues, hydroxylysyl pyridinoline (HP) and lysyl pyridinoline (LP), showed that these mature cross-links increased from undetectable levels to concentrations found in normal chick bone. Analysis of the kinetics of their formation by pulse-chase labeling the cultures with [3H]lysine showed a doubling time of approximately 5 days. The relationships between cross-link formation, fibrillogenesis, and collagen stability were examined in cultures treated with beta-aminopropionitrile (beta-APN), a potent inhibitor of lysyl oxidase and cross-link formation. In beta-APN-treated cultures, total collagen synthesis was increased twofold, with no change in mRNA levels for type I collagen, whereas the amount of collagen accumulated in the cell layer was decreased by 50% and mineral deposition was reduced. The rate of collagen retention in the matrix was assessed by pulse-chase analysis of [3H]proline over a 16 day period in

  14. Information contained in the amino acid sequence of the alpha1(I)-chain of collagen and its consequences upon the formation of the triple helix, of fibrils and crosslinks.

    PubMed

    Fietzek, P P; Kühn, K

    1975-09-30

    The molecule of type I collagen from skin consists of two alpha1(I)-chains and one alpha2-chain. The sequence of the entire alpha1-chain comprising 1052 residues is summarily presented and discussed. Apart from the 279 residues of alpha1(I)-CB8 whose sequence has been established for rat skin collagen, all sequences have been determined for calf skin collagen. In order to facilitate sequence analysis, the alpha1-chain was cleaved into defined fragments by cyanogen bromide or hydroxylamine or limited collagenase digestion. Most of the sequence was established by automated stepwise Edman degradation. The alpha1-chain contains two basically different types of sequences: the triple helical region of 1011 amino acid residues in which every third position is occupied by glycine and the N- and C-terminal regions not displaying this type of regularity. Both of these non-triple helical regions carry oxidizable lysine or hydroxylysine residues as functional sites for the intermolecular crosslink formation. Implications of the amino acid sequence for the stability of the triple helix and the fibril as well as for formation of crosslinks are discussed. Evaluation of the sequence in connection with electron microscopical investigations yielded the parameters of the axial arrangement of the molecules within the fibrils. Axial stagger of the molecules by a distance D = 670 angstrom = 233 amino acid residues results in maximal interaction of polar sequence regions of adjacent molecules and similarly of regions of hydrophobic residues. Ordered aggregation of molecules into fibrils is, therefore, regulated by electrostatic and electrophobic forces. Possible loci of intermolecular crosslinks between the alpha1-chains of adjacent molecules may be deduced from the dimensions of the axial aggregation of molecules. PMID:171554

  15. Surface modification of electrospun PLGA scaffold with collagen for bioengineered skin substitutes.

    PubMed

    Sadeghi, A R; Nokhasteh, S; Molavi, A M; Khorsand-Ghayeni, M; Naderi-Meshkin, H; Mahdizadeh, A

    2016-09-01

    In skin tissue engineering, surface feature of the scaffolds plays an important role in cell adhesion and proliferation. In this study, non-woven fibrous substrate based on poly (lactic-co-glycolic acid) (PLGA) (75/25) were hydrolyzed in various concentrations of NaOH (0.05N, 0.1N, 0.3N) to increase carboxyl and hydroxyl groups on the fiber surfaces. These functional groups were activated by EDC/NHS to create chemical bonding with collagen. To improve bioactivity, the activated substrates were coated with a collagen solution (2mg/ml) and cross-linking was carried out using the EDC/NHS in MES buffer. The effectiveness of the method was evaluated by contact angle measurements, porosimetry, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), tensile and degradation tests as well as in vitro cell attachment and cytotoxicity assays. Cell culture results of human dermal fibroblasts (HDF) and keratinocytes cell line (HaCat) revealed that the cells could attach to the scaffold. Further investigation with MTT assay showed that the cell proliferation of HaCat significantly increases with collagen coating. It seems that sufficient stability of collagen on the surface due to proper chemical bonding and cross-linking has increased the bioactivity of surface remarkably which can be promising for bioengineered skin applications. PMID:27207046

  16. Structural basis of sequence-specific collagen recognition by SPARC

    PubMed Central

    Hohenester, Erhard; Sasaki, Takako; Giudici, Camilla; Farndale, Richard W.; Bächinger, Hans Peter

    2008-01-01

    Protein interactions with the collagen triple helix play a critical role in collagen fibril formation, cell adhesion, and signaling. However, structural insight into sequence-specific collagen recognition is limited to an integrin-peptide complex. A GVMGFO motif in fibrillar collagens (O denotes 4-hydroxyproline) binds 3 unrelated proteins: von Willebrand factor (VWF), discoidin domain receptor 2 (DDR2), and the extracellular matrix protein SPARC/osteonectin/BM-40. We report the crystal structure at 3.2 Å resolution of human SPARC bound to a triple-helical 33-residue peptide harboring the promiscuous GVMGFO motif. SPARC recognizes the GVMGFO motifs of the middle and trailing collagen chains, burying a total of 720 Å2 of solvent-accessible collagen surface. SPARC binding does not distort the canonical triple helix of the collagen peptide. In contrast, a critical loop in SPARC is substantially remodelled upon collagen binding, creating a deep pocket that accommodates the phenylalanine residue of the trailing collagen chain (“Phe pocket”). This highly restrictive specificity pocket is shared with the collagen-binding integrin I-domains but differs strikingly from the shallow collagen-binding grooves of the platelet receptor glycoprotein VI and microbial adhesins. We speculate that binding of the GVMGFO motif to VWF and DDR2 also results in structural changes and the formation of a Phe pocket. PMID:19011090

  17. Collagen fibrillogenesis: fibronectin, integrins, and minor collagens as organizers and nucleators

    PubMed Central

    Kadler, Karl E; Hill, Adele; Canty-Laird, Elizabeth G

    2008-01-01

    Collagens are triple helical proteins that occur in the extracellular matrix (ECM) and at the cell–ECM interface. There are more than 30 collagens and collagen-related proteins but the most abundant are collagens I and II that exist as D-periodic (where D = 67 nm) fibrils. The fibrils are of broad biomedical importance and have central roles in embryogenesis, arthritis, tissue repair, fibrosis, tumor invasion, and cardiovascular disease. Collagens I and II spontaneously form fibrils in vitro, which shows that collagen fibrillogenesis is a selfassembly process. However, the situation in vivo is not that simple; collagen I-containing fibrils do not form in the absence of fibronectin, fibronectin-binding and collagen-binding integrins, and collagen V. Likewise, the thin collagen II-containing fibrils in cartilage do not form in the absence of collagen XI. Thus, in vivo, cellular mechanisms are in place to control what is otherwise a protein self-assembly process. This review puts forward a working hypothesis for how fibronectin and integrins (the organizers) determine the site of fibril assembly, and collagens V and XI (the nucleators) initiate collagen fibrillogenesis. PMID:18640274

  18. Impact of temperature and electrical potentials on the stability and structure of collagen adsorbed on the gold electrode

    NASA Astrophysics Data System (ADS)

    Meiners, Frank; Ahlers, Michael; Brand, Izabella; Wittstock, Gunther

    2015-01-01

    The morphology and structure of collagen type I adsorbed on gold electrodes were studied as a function of electrode potential and temperature by means of capacitance measurements, polarization modulation infrared reflection-absorption spectroscopy and scanning force microscopy at temperatures of 37 °C, 43 °C and 50 °C. The selected temperatures corresponded to the normal body temperature, temperature of denaturation of collagen molecules and denaturation of collagen fibrils, respectively. Independently of the solution temperature, collagen was adsorbed on gold electrodes in the potential range - 0.7 V < E < 0.4 V vs. Ag/AgCl, where the protein film was very stable. Fragments of collagen molecules made a direct contact to the gold surface and water was present in the film. Protein molecules were oriented preferentially with their long axis towards the gold surface. Collagen molecules in the adsorbed state preserved their native triple helical structure even at temperatures corresponding to collagen denaturation in aqueous solutions. Application of E < - 0.75 V vs. Ag/AgCl leads to the swelling of the protein film by water and desorption from the electrode surface. IR spectra provided no evidence of the thermal denaturation of adsorbed collagen molecules. A temperature increase to 50 °C leads to a distortion of the collagen film. The processes of aggregation and fibrilization were preferred over thermal denaturation for collagen adsorbed on the electrode surface and exposed to changing potentials.

  19. Amyloid fibrillation in native and chemically-modified forms of carbonic anhydrase II: role of surface hydrophobicity.

    PubMed

    Es-Haghi, Ali; Shariatizi, Sajad; Ebrahim-Habibi, Azadeh; Nemat-Gorgani, Mohsen

    2012-03-01

    Chemical modification or mutation of proteins may bring about significant changes in the net charge or surface hydrophobicity of a protein structure. Such events may be of major physiological significance and may provide important insights into the genetics of amyloid diseases. In the present study, fibrillation potential of native and chemically-modified forms of bovine carbonic anhydrase II (BCA II) were investigated. Initially, various denaturing conditions including low pH and high temperatures were tested to induce fibrillation. At a low pH of around 2.4, where the protein is totally dissociated, the apo form was found to take up a pre-molten globular (PMG) conformation with the capacity for fibril formation. Upon increasing the pH to around 3.6, a molten globular (MG) form became abundant, forming amorphous aggregates. Charge neutralization and enhancement of hydrophobicity by methylation, acetylation and propionylation of lysine residues appeared very effective in promoting fibrillation of both the apo and holo forms under native conditions, the rates and extents of which were directly proportional to surface hydrophobicity, and influenced by salt concentration and temperature. These modified structures underwent more pronounced fibrillation under native conditions, than the PMG intermediate form, observed under denaturing conditions. The nature of the fibrillation products obtained from intermediate and modified structures were characterized and compared and their possible cytotoxicity determined. Results are discussed in terms of the importance of surface net charge and hydrophobicity in controlling protein aggregation. A discussion on the physiological significance of the observations is also presented. PMID:22251892

  20. Cell Proliferation on Macro/Nano Surface Structure and Collagen Immobilization of 3D Polycaprolactone Scaffolds.

    PubMed

    Park, Young-Ouk; Myung, Sung-Woon; Kook, Min-Suk; Jung, Sang-Chul; Kim, Byung-Hoon

    2016-02-01

    In this study, 3D polycaprolactone (PCL) scaffolds were fabricated by 3D printing technique. The macro/nano morphology of, 3D PCL scaffolds surface was etched with oxygen plasma. Acrylic acid (AA) plasma-polymerization was performed to functionalize the macro/nano surface with carboxyl groups and then collagen was immobilized with plasma-polymerized 3D PCL scaffolds. After O2 plasma and AA plasma-polymerization, contact angles were decreased. The FE-SEM and AFM results showed that O2 plasma is increased the surface roughness. The MTT assay results showed that proliferation of the M3CT3-E1 cells increased on the oxygen plasma treated and collagen immobilized 3D PCL scaffolds. PMID:27433597

  1. Competition between surface adsorption and folding of fibril-forming polypeptides

    NASA Astrophysics Data System (ADS)

    Ni, Ran; Kleijn, J. Mieke; Abeln, Sanne; Cohen Stuart, Martien A.; Bolhuis, Peter G.

    2015-02-01

    Self-assembly of polypeptides into fibrillar structures can be initiated by planar surfaces that interact favorably with certain residues. Using a coarse-grained model, we systematically studied the folding and adsorption behavior of a β -roll forming polypeptide. We find that there are two different folding pathways depending on the temperature: (i) at low temperature, the polypeptide folds in solution into a β -roll before adsorbing onto the attractive surface; (ii) at higher temperature, the polypeptide first adsorbs in a disordered state and folds while on the surface. The folding temperature increases with increasing attraction as the folded β -roll is stabilized by the surface. Surprisingly, further increasing the attraction lowers the folding temperature again, as strong attraction also stabilizes the adsorbed disordered state, which competes with folding of the polypeptide. Our results suggest that to enhance the folding, one should use a weakly attractive surface. They also explain the recent experimental observation of the nonmonotonic effect of charge on the fibril formation on an oppositely charged surface [C. Charbonneau et al., ACS Nano 8, 2328 (2014), 10.1021/nn405799t].

  2. Collagen Cross-linking Increases Its Biodegradation Resistance in Wet Dentin Bonding

    PubMed Central

    Xu, Changqi; Wang, Yong

    2014-01-01

    Purpose The biodegradation of exposed dentin collagen within the adhesive/dentin (a/d) interface is one of main reasons leading to composite restoration failures and seriously affects the durability of dental restorations. In the present study, the objective was to investigate whether the inclusion of the crosslinking reagent (glutaraldehyde, GA) in the adhesive would increase collagen biodegradation resistance within the a/d interface. Materials and methods The model adhesive consisted of ~60 % monomers (HEMA/BisGMA, 45/55 wt/wt) and ~ 40 % ethanol as a solvent. 5% GA was added to the above formulation. After the dentin surfaces were etched for 15 s with 35% phosphoric acid, rinsed with water and blotted dry, adhesives both with and without GA were applied and polymerized by visible light for 20 s. These a/d specimens were immersed in the biodegradation solution (prepared by adding 160 mg collagenase in 1 liter of TESCA buffer solution) for up to 30 days after proceeding with the sectioning/fracture to expose the a/d interfaces. The specimens were analyzed using SEM and micro-Raman. Results SEM results indicated that for the adhesive without GA, there were many voids and was a loss of collagen fibrils in the a/d interface after being challenged by the biodegradation solution. The Raman spectra collected from the interface showed that the amide I of collagen at 1667 cm−1 obviously decreased, indicating a removal of collagen fibrils during the degradation process. For the adhesive containing GA, the collagen fibrils within the interface did not degrade at all, which was also confirmed by the Raman results. Conclusion The results corroborate the previous findings that by using the current adhesive system and wet bonding, the collagen fibrils in the a/d interface are largely unprotected and easily undergo biodegradation. Directly including crosslinking agents in the adhesive could protect collagen fibrils from degradation in situ within the a/d interface. PMID

  3. Controlling the nano-bio interface to build collagen-silica self-assembled networks

    NASA Astrophysics Data System (ADS)

    Aimé, Carole; Mosser, Gervaise; Pembouong, Gaëlle; Bouteiller, Laurent; Coradin, Thibaud

    2012-10-01

    Bio-hybrid networks are designed based on the self-assembly of surface-engineered collagen-silica nanoparticles. Collagen triple helices can be confined on the surface of sulfonate-modified silica particles in a controlled manner. This gives rise to hybrid building blocks with well-defined diameters and surface potentials. Taking advantage of the self-assembling properties of collagen, collagen-silica networks are further built-up in solution. The structural and specific recognition properties of the collagen fibrils are well-preserved within the hybrid assembly. A combination of calorimetry, dynamic light scattering, zetametry and microscopy studies indicates that network formation occurs via a surface-mediated mechanism where pre-organization of the protein chains on the particle surface favors the fibrillogenesis process. These results enlighten the importance of the nano-bio interface on the formation and properties of self-assembled bionanocomposites.Bio-hybrid networks are designed based on the self-assembly of surface-engineered collagen-silica nanoparticles. Collagen triple helices can be confined on the surface of sulfonate-modified silica particles in a controlled manner. This gives rise to hybrid building blocks with well-defined diameters and surface potentials. Taking advantage of the self-assembling properties of collagen, collagen-silica networks are further built-up in solution. The structural and specific recognition properties of the collagen fibrils are well-preserved within the hybrid assembly. A combination of calorimetry, dynamic light scattering, zetametry and microscopy studies indicates that network formation occurs via a surface-mediated mechanism where pre-organization of the protein chains on the particle surface favors the fibrillogenesis process. These results enlighten the importance of the nano-bio interface on the formation and properties of self-assembled bionanocomposites. Electronic supplementary information (ESI) available: XPS

  4. Libby amphibole-induced mesothelial cell autoantibodies bind to surface plasminogen and alter collagen matrix remodeling.

    PubMed

    Hanson, Robert; Evilia, Caryn; Gilmer, John; Woods, Linda; Black, Brad; Flores, Raja; Pfau, Jean C

    2016-08-01

    Lamellar pleural thickening (LPT) is a fibrotic disease induced by exposure to Libby amphibole (LA) asbestos that causes widespread scarring around the lung, resulting in deterioration of pulmonary function. Investigating the effects of autoantibodies to mesothelial cells (MCAA) present in the study populations has been a major part of the effort to understand the mechanism of pathogenesis. It has been shown in vitro that human mesothelial cells (Met5a) exposed to MCAA increase collagen deposition into the extracellular matrix (ECM). In this study, we sought to further elucidate how MCAA drive increased collagen deposition by identifying the protein targets bound by MCAA on the cellular surface using biotinylation to label and isolate surface proteins. Isolated surface protein fractions were identified as containing MCAA targets using ELISA The fractions that demonstrated binding by MCAA were then analyzed by tandem mass spectrometry (MS/MS) and MASCOT analysis. The most promising result from the MASCOT analysis, plasminogen (PLG), was tested for MCAA binding using purified human PLG in an ELISA We report that serum containing MCAA bound at an optical density (OD) 3 times greater than that of controls, and LA-exposed subjects had a high frequency of positive tests for anti-PLG autoantibodies. This work implicates the involvement of the plasminogen/plasmin system in the mechanism of excess collagen deposition in Met5a cells exposed to MCAA Elucidating this mechanism could contribute to the understanding of LPT. PMID:27519611

  5. Ventricular fibrillation

    MedlinePlus

    ... Fibrillation is an uncontrolled twitching or quivering of muscle fibers (fibrils). When it occurs in the lower chambers of the heart, it is called ventricular fibrillation. During ... the heart muscle does not get enough oxygen for any reason. ...

  6. Different Sialoside Epitopes on Collagen Film Surfaces Direct Mesenchymal Stem Cell Fate.

    PubMed

    Sgambato, Antonella; Russo, Laura; Montesi, Monica; Panseri, Silvia; Marcacci, Maurilio; Caravà, Elena; Raspanti, Mario; Cipolla, Laura

    2016-06-22

    3'-Sialyllactose and 6'-sialyllactose have been covalently linked to collagen films. Preliminary in vitro study on the behavior of mesenchymal stem cells (MSCs) in terms of cell viability, proliferation and induction of osteogenic and chondrogenic related genes has been performed. Results indicate that sialoside epitopes on collagen surface represent a suitable support for MSCs adhesion and cell proliferation, moreover, the neoglycosylation provide MSCs with different and specific stimuli, saccharide-type depending, in term of expression of osteogenic and chondrogenic related genes. In particular, 3'-sialyllactose significantly upregulate the expression of RUNX2 and ALP, well-known markers of osteogenesis, whereas 6'-sialyllactose up-regulate the expression of chondrocyte marker ACAN. Because no osteogenic or chondrogenic supplements in culture media were added, the inductive effect in terms of increased gene expression has to be ascribed uniquely to collagen surface functionalization. These results support the promising role of sialosides in the regulation of stem cells fate and open brilliant perspective for the future use of the presented approach toward osteochondral tissue engineering applications. PMID:26697920

  7. Identification of Surface Proteins from Lactobacillus casei BL23 Able to Bind Fibronectin and Collagen.

    PubMed

    Muñoz-Provencio, Diego; Pérez-Martínez, Gaspar; Monedero, Vicente

    2011-03-01

    Strains of lactobacilli show the capacity to attach to extracellular matrix proteins. Cell-wall fractions of Lactobacillus casei BL23 enriched in fibronectin, and collagen-binding proteins were isolated. Mass spectrometry analysis of their protein content revealed the presence of stress-related proteins (GroEL, ClpL), translational elongation factors (EF-Tu, EF-G), oligopeptide solute-binding proteins, and the glycolytic enzymes enolase and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The latter two enzymes were expressed in Escherichia coli and purified as glutathione-S-transferase (GST) fusion proteins, and their in vitro binding activity to fibronectin and collagen was confirmed. These results reinforce the idea that lactobacilli display on their surfaces a variety of moonlighting proteins that can be important in their adaptation to survive at intestinal mucosal sites and in the interaction with host cells. PMID:26781495

  8. Depth-Dependent Changes in Collagen Organization in the Human Peripapillary Sclera

    PubMed Central

    Pijanka, Jacek K.; Spang, Martin T.; Sorensen, Thomas; Liu, Jun; Nguyen, Thao D.; Quigley, Harry A.; Boote, Craig

    2015-01-01

    Purpose The collagen structure of the human peripapillary sclera plays a significant role in determining optic nerve head (ONH) biomechanics, and is therefore of interest in the study of glaucoma. The aim of the current work was to map the anisotropic collagen structure of the normal human peripapillary sclera as a function of tissue depth. Methods Wide-angle x-ray scattering was used to quantify collagen fibril orientation at 0.5mm intervals across six 150μm-thick serial sections through the peripapillary sclera of eight normal European-derived human eyes. Two structural parameters were measured: 1) the relative number of fibrils preferentially aligned at a given angle within the tissue plane, 2) the degree of collagen alignment (anisotropy). Results The inner-most one-third of the peripapillary scleral stroma (nearest to the choroid) was characterised by collagen fibrils either randomly arranged or preferentially aligned radially with respect to the ONH. In contrast, the outer two-thirds of the tissue was dominated by a circumferential arrangement of collagen encircling the ONH. In all tissue regions the degree of collagen anisotropy peaked in the mid-stroma and progressively decreased towards the tissue surfaces, with the largest depth variations occurring in the inferior-nasal quadrant, and the smallest occurring in the superior-nasal quadrant. Conclusions Significant, region-specific variations in collagen structure are present in the human peripapillary sclera as a function of depth. In normal eyes, the circumferential collagen fibril architecture is most prominent in the outer two-thirds of the stroma, possibly as a mechanical adaption to more effectively support the lamina cribrosa at the level of its insertion into the scleral canal wall. PMID:25714753

  9. Optimization of antioxidant activity by response surface methodology in hydrolysates of jellyfish (Rhopilema esculentum) umbrella collagen*

    PubMed Central

    Zhuang, Yong-liang; Zhao, Xue; Li, Ba-fang

    2009-01-01

    To optimize the hydrolysis conditions to prepare hydrolysates of jellyfish umbrella collagen with the highest hydroxyl radical scavenging activity, collagen extracted from jellyfish umbrella was hydrolyzed with trypsin, and response surface methodology (RSM) was applied. The optimum conditions obtained from experiments were pH 7.75, temperature (T) 48.77 °C, and enzyme-to-substrate ratio ([E]/[S]) 3.50%. The analysis of variance in RSM showed that pH and [E]/[S] were important factors that significantly affected the process (P<0.05 and P<0.01, respectively). The hydrolysates of jellyfish umbrella collagen were fractionated by high performance liquid chromatography (HPLC), and three fractions (HF-1>3000 Da, 1000 Da

  10. Evidence for the presence of collagenous domains in Candida albicans cell surface proteins.

    PubMed Central

    Sepúlveda, P; Murgui, A; López-Ribot, J L; Casanova, M; Timoneda, J; Martínez, J P

    1995-01-01

    Rabbit polyclonal antibodies (PAbs) directed towards the amino-terminal cysteine-rich 7S domain (PAb anti-7S), the major internal collagenous domain (PAb anti-type IV), and the C-terminal noncollagenous region (PAb anti-NC1) of the type IV collagen molecule were probed by indirect immunofluorescence against Candida albicans blastoconidia and germinated blastoconidia. Most nongerminating cells and mother blastoconidia from which germ tubes originated showed strong fluorescence when PAb anti-7S was used, whereas with PAb anti-type IV, fluorescence was found almost exclusively on the surface of filamentous forms. A patched fluorescent pattern rather than a homogenous confluent fluorescence was observed in all cases. No fluorescent cells were observed with PAb anti-NC1. By Western immunoblotting, PAb anti-type IV cross-reacted primarily with a polypeptide of 37 kDa present in wall extracts obtained from intact cells of both growth forms by treatment with beta-mercaptoethanol, whereas PAb anti-7S recognized a major 58-kDa antigen also present in both extracts, along with some other high-molecular-mass (> 106-kDa) polydisperse species present only in the material released from blastoconidia with beta-mercaptoethanol. No reactive bands were observed when PAb anti-NC1 was used as a probe in Western immunoblotting experiments. The sensitivities or resistances to collagenase digestion of the different polypeptides that cross-reacted with PAbs anti-type IV and anti-7S suggest the existence of cell wall components in C. albicans that contain epitopes that mimic the collagenous domains of the type IV collagen molecule. PMID:7768595

  11. Structural relations between collagen and mineral in bone as determined by high voltage electron microscopic tomography

    NASA Technical Reports Server (NTRS)

    Landis, W. J.; Hodgens, K. J.; Arena, J.; Song, M. J.; McEwen, B. F.

    1996-01-01

    Aspects of the ultrastructural interaction between collagen and mineral crystals in embryonic chick bone have been examined by the novel technique of high voltage electron microscopic tomography to obtain three-dimensional information concerning extracellular calcification in this tissue. Newly mineralizing osteoid along periosteal surfaces of mid-diaphyseal regions from normal chick tibiae was embedded, cut into 0.25 microns thick sections, and documented at 1.0 MV in the Albany AEI-EM7 high voltage electron microscope. The areas of the tissue studied contained electron dense mineral crystals associated with collagen fibrils, some marked by crystals disposed along their cylindrically shaped lengths. Tomographic reconstructions of one site with two mineralizing fibrils were computed from a 5 degrees tilt series of micrographs over a +/- 60 degrees range. Reconstructions showed that the mineral crystals were platelets of irregular shape. Their sizes were variable, measured here up to 80 x 30 x 8 nm in length, width, and thickness, respectively. The longest crystal dimension, corresponding to the c-axis crystallographically, was generally parallel to the collagen fibril long axis. Individual crystals were oriented parallel to one another in each fibril examined. They were also parallel in the neighboring but apparently spatially separate fibrils. Crystals were periodically (approximately 67 nm repeat distance) arranged along the fibrils and their location appeared to correspond to collagen hole and overlap zones defined by geometrical imaging techniques. The crystals appeared to be continuously distributed along a fibril, their size and number increasing in a tapered fashion from a relatively narrow tip containing smaller and infrequent crystals to wider regions having more densely packed and larger crystals. Defined for the first time by direct visual 3D imaging, these data describe the size, shape, location, orientation, and development of early crystals in normal

  12. Effect of nitrogen-rich cell culture surfaces on type X collagen expression by bovine growth plate chondrocytes

    PubMed Central

    2011-01-01

    Background Recent evidence indicates that osteoarthritis (OA) may be a systemic disease since mesenchymal stem cells (MSCs) from OA patients express type X collagen, a marker of late stage chondrocyte hypertrophy (associated with endochondral ossification). We recently showed that the expression of type X collagen was suppressed when MSCs from OA patients were cultured on nitrogen (N)-rich plasma polymer layers, which we call "PPE:N" (N-doped plasma-polymerized ethylene, containing up to 36 atomic percentage (at.% ) of N. Methods In the present study, we examined the expression of type X collagen in fetal bovine growth plate chondrocytes (containing hypertrophic chondrocytes) cultured on PPE:N. We also studied the effect of PPE:N on the expression of matrix molecules such as type II collagen and aggrecan, as well as on proteases (matrix metalloproteinase-13 (MMP-13) and molecules implicated in cell division (cyclin B2). Two other culture surfaces, "hydrophilic" polystyrene (PS, regular culture dishes) and nitrogen-containing cation polystyrene (Primaria®), were also investigated for comparison. Results Results showed that type X collagen mRNA levels were suppressed when cultured for 4 days on PPE:N, suggesting that type X collagen is regulated similarly in hypertrophic chondrocytes and in human MSCs from OA patients. However, the levels of type X collagen mRNA almost returned to control value after 20 days in culture on these surfaces. Culture on the various surfaces had no significant effects on type II collagen, aggrecan, MMP-13, and cyclin B2 mRNA levels. Conclusion Hypertrophy is diminished by culturing growth plate chondrocytes on nitrogen-rich surfaces, a mechanism that is beneficial for MSC chondrogenesis. Furthermore, one major advantage of such "intelligent surfaces" over recombinant growth factors for tissue engineering and cartilage repair is potentially large cost-saving. PMID:21244651

  13. Enhancing anticoagulation and endothelial cell proliferation of titanium surface by sequential immobilization of poly(ethylene glycol) and collagen

    NASA Astrophysics Data System (ADS)

    Pan, Chang-Jiang; Hou, Yan-Hua; Ding, Hong-Yan; Dong, Yun-Xiao

    2013-12-01

    In the present study, poly(ethylene glycol) (PEG) and collagen I were sequentially immobilized on the titanium surface to simultaneously improve the anticoagulation and endothelial cell proliferation. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy analysis confirmed that PEG and collagen I were successfully immobilized on the titanium surface. Water contact angle results suggested the excellent hydrophilic surface after the immobilization. The anticoagulation experiments demonstrated that the immobilized PEG and collagen I on the titanium surface could not only obviously prevent platelet adhesion and aggregation but also prolong activated partial thromboplastin time (APTT), leading to the improved blood compatibility. Furthermore, immobilization of collagen to the end of PEG chain did not abate the anticoagulation. As compared to those on the pristine and PEG-modified titanium surfaces, endothelial cells exhibited improved proliferative profiles on the surface modified by the sequential immobilization of PEG and collagen in terms of CCK-8 assay, implying that the modified titanium may promote endothelialization without abating the blood compatibility. Our method may be used to modify the surface of blood-contacting biomaterials such as titanium to promote endothelialization and improve the anticoagulation, it may be helpful for development of the biomedical devices such as coronary stents, where endothelializaton and excellent anticoagulation are required.

  14. An Equilibrium Model for the Combined Effect of Macromolecular Crowding and Surface Adsorption on the Formation of Linear Protein Fibrils

    PubMed Central

    Hoppe, Travis; Minton, Allen P.

    2015-01-01

    The formation of linear protein fibrils has previously been shown to be enhanced by volume exclusion or crowding in the presence of a high concentration of chemically inert protein or polymer, and by adsorption to membrane surfaces. An equilibrium mesoscopic model for the combined effect of both crowding and adsorption upon the fibrillation of a dilute tracer protein is presented. The model exhibits behavior that differs qualitatively from that observed in the presence of crowding or adsorption alone. The model predicts that in a crowded solution, at critical values of the volume fraction of crowder or intrinsic energy of the tracer-wall interaction, the tracer protein will undergo an extremely cooperative transition—approaching a step function—from existence as a slightly self-associated species in solution to existence as a highly self-associated and completely adsorbed species. Criteria for a valid experimental test of these predictions are presented. PMID:25692600

  15. COMP-assisted collagen secretion--a novel intracellular function required for fibrosis.

    PubMed

    Schulz, Jan-Niklas; Nüchel, Julian; Niehoff, Anja; Bloch, Wilhelm; Schönborn, Katrin; Hayashi, Shujiro; Kamper, Matthias; Brinckmann, Jürgen; Plomann, Markus; Paulsson, Mats; Krieg, Thomas; Zaucke, Frank; Eckes, Beate

    2016-02-15

    Cartilage oligomeric matrix protein (COMP) is an abundant component in the extracellular matrix (ECM) of load-bearing tissues such as tendons and cartilage. It provides adaptor functions by bridging different ECM structures. We have previously shown that COMP is also a constitutive component of healthy human skin and is strongly induced in fibrosis. It binds directly and with high affinity to collagen I and to collagen XII that decorates the surface of collagen I fibrils. We demonstrate here that lack of COMP-collagen interaction in the extracellular space leads to changes in collagen fibril morphology and density, resulting in altered skin biomechanical properties. Surprisingly, COMP also fulfills an important intracellular function in assisting efficient secretion of collagens, which were retained in the endoplasmic reticulum of COMP-null fibroblasts. Accordingly, COMP-null mice showed severely attenuated fibrotic responses in skin. Collagen secretion was fully restored by introducing wild-type COMP. Hence, our work unravels a new, non-structural and intracellular function of the ECM protein COMP in controlling collagen secretion. PMID:26746240

  16. Collagen: Biochemistry, biomechanics, biotechnology

    SciTech Connect

    Nimni, M.E.

    1988-01-01

    This book is an up-to-date reference for new ideas, information, and concepts in collagen research. The first volume emphasizes the relationship between the molecular structure and function of collagen, including descriptions of collagen types which exist in tissues as well as how these molecules organize into fibrils and the nature of the chemical crosslinks which stabilize them. In Volume II the biomechanical behavior of various specialized tissues, abnormal accumulation of collagen in the form of scars of fibrous infiltration are examined/and wound healing, tissue regulation and repair are covered in detail. Volume III explores the increasing application of collagen technology to the field of bioprosthesis, including the production of heart valve bioprosthesis, blood vessels, ligament substitutes, and bone substitutes.

  17. Engineering and Characterization of Collagen Networks Using Wet Atomic Force Microscopy and Environmental Scanning Electron Microscopy

    NASA Astrophysics Data System (ADS)

    Osborn, Jenna; Coffey, Tonya; Conrad, Brad; Burris, Jennifer; Hester, Brooke

    2014-03-01

    Collagen is an abundant protein and its monomers covalently crosslink to form fibrils which form fibers which contribute to forming macrostructures like tendon or bone. While the contribution is well understood at the macroscopic level, it is not well known at the fibril level. We wish to study the mechanical properties of collagen for networks of collagen fibers that vary in size and density. We present here a method to synthesize collagen networks from monomers and that allows us to vary the density of the networks. By using biotynilated collagen and a surface that is functionalized with avidin, we generate two-dimensional collagen networks across the surface of a silicon wafer. During network synthesis, the incubation time is varied from 30 minutes to 3 hours or temperature is varied from 25°C to 45°C. The two-dimensional collagen network created in the process is characterized using environmental atomic force microscopy (AFM) and scanning electron microscopy (SEM). The network density is measured by the number of strands in one frame using SPIP software. We expect that at body temperature (37°C) and with longer incubation times, the network density should increase.

  18. Nanoscale Mechanics of Type I Collagen

    NASA Astrophysics Data System (ADS)

    Harper, H.; Cropper, E.; Bulger, A.; Choksi, U.; Koob, T. J.; Pandit, S.; Matthews, W. G.

    2009-03-01

    Collagen is the most abundant protein in the body by mass. Type I collagen fibrils provide mechanical strength and cellular housing within tissues exhibiting a broad range of mechanical properties. This diversity in the mechanics of tissues with similar underlying components warrants detailed study of the process by which structure and mechanics develop. While collagen mechanics have been studied at the tissue level for decades, surprising little is known about collagen mechanics at the fibril and molecular level. Presented herein is a multi-scale experimental and computational investigation of collagen I mechanics, bridging the single molecule and fibril hierarchal forms. The mechanics of single collagen molecules are explored using AFM and force spectroscopy. Moreover, atomistic molecular-dynamics simulations are performed to provide structural information not accessible to the experimental system. Fibrils then are grown from molecular collagen, and the mechanics of these fibrils are investigated using AFM. Based upon the single molecule and fibril results, a coarse-grain computational model is being developed. The outcomes include a better understanding of how the mechanics of filamentous self-organizing systems are derived and how their hierarchical forms are established.

  19. Changes in corneal collagen induced by holmium:YAG laser irradiation

    NASA Astrophysics Data System (ADS)

    Timberlake, George T.; Reinke, Martin H.; Miller, Alvin

    1996-05-01

    Holmium:YAG laser thermokeratoplasty corrects hyperopia (farsightedness) by producing small areas of corneal collagen shrinkage that cause the central cornea to bulge outward, increasing optical power. Collagen shrinkage is probably caused by laser-heated corneal water, but details of the shrinkage mechanism are not known. We investigated the shrinkage mechanism by measuring changes in corneal ultrastructure, surface shrinkage, water content, and strength following Ho:YAG laser exposures. Morphological changes in collagen were documented by measurements from electron micrographs. Corneal adhesive strength was determined by measuring tearing force in a plane parallel to the corneal surface. Laser-induced water loss was measured by weighing corneal samples before and after exposure. Corneal surface shrinkage was assessed by photographing the movement of particles on the cornea. Lasered collagen fibrils increased in diameter, lost their orderly arrangement, and appeared `frayed.' The corneal surface contracted toward lasered areas with a maximal shift of approximately 190 micrometers , more than could be explained by a model based on collagen fibril changes. Water loss plays a minor role in corneal shrinkage since corneal samples lost about only about 1.4% of their weight after massive laser exposure. Despite marked changes in collagen structure, corneal adhesive force was unchanged.

  20. Ovine tendon collagen: Extraction, characterisation and fabrication of thin films for tissue engineering applications.

    PubMed

    Fauzi, M B; Lokanathan, Y; Aminuddin, B S; Ruszymah, B H I; Chowdhury, S R

    2016-11-01

    Collagen is the most abundant extracellular matrix (ECM) protein in the human body, thus widely used in tissue engineering and subsequent clinical applications. This study aimed to extract collagen from ovine (Ovis aries) Achilles tendon (OTC), and to evaluate its physicochemical properties and its potential to fabricate thin film with collagen fibrils in a random or aligned orientation. Acid-solubilized protein was extracted from ovine Achilles tendon using 0.35M acetic acid, and 80% of extracted protein was measured as collagen. SDS-PAGE and mass spectrometry analysis revealed the presence of alpha 1 and alpha 2 chain of collagen type I (col I). Further analysis with Fourier transform infrared spectrometry (FTIR), X-ray diffraction (XRD) and energy dispersive X-ray spectroscopy (EDS) confirms the presence of triple helix structure of col I, similar to commercially available rat tail col I. Drying the OTC solution at 37°C resulted in formation of a thin film with randomly orientated collagen fibrils (random collagen film; RCF). Introduction of unidirectional mechanical intervention using a platform rocker prior to drying facilitated the fabrication of a film with aligned orientation of collagen fibril (aligned collagen film; ACF). It was shown that both RCF and ACF significantly enhanced human dermal fibroblast (HDF) attachment and proliferation than that on plastic surface. Moreover, cells were distributed randomly on RCF, but aligned with the direction of mechanical intervention on ACF. In conclusion, ovine tendon could be an alternative source of col I to fabricate scaffold for tissue engineering applications. PMID:27524008

  1. The Collagen Family

    PubMed Central

    Ricard-Blum, Sylvie

    2011-01-01

    Collagens are the most abundant proteins in mammals. The collagen family comprises 28 members that contain at least one triple-helical domain. Collagens are deposited in the extracellular matrix where most of them form supramolecular assemblies. Four collagens are type II membrane proteins that also exist in a soluble form released from the cell surface by shedding. Collagens play structural roles and contribute to mechanical properties, organization, and shape of tissues. They interact with cells via several receptor families and regulate their proliferation, migration, and differentiation. Some collagens have a restricted tissue distribution and hence specific biological functions. PMID:21421911

  2. Dermal ultrastructure in collagen VI myopathy.

    PubMed

    Hermanns-Lê, Trinh; Piérard, Gérald E; Piérard-Franchimont, Claudine; Delvenne, Philippe

    2014-04-01

    The COL VI mutations are responsible for a spectrum of myopathies. The authors report cutaneous ultrastructural alterations in a patient with COL6A2 myopathy. The changes include variations in size of collagen fibrils, flower-like sections of collagen fibrils, as well as thickening of vessel and nerve basement membranes. Electron microscopy of a skin biopsy contributes to the diagnosis of COL VI myopathies. PMID:24134684

  3. Studies on the Interaction between Collagen and a Plasma Kallikrein-Like Activity EVIDENCE FOR A SURFACE-ACTIVE ENZYME SYSTEM

    PubMed Central

    Harpel, Peter C.

    1972-01-01

    This study has demonstrated that collagen particles, after exposure to platelet-poor human plasma and subsequent washing, generate a kinin-like agent when incubated with prekinin substrate. The binding of kinin-generating activity to collagen in the plasma collagen incubation mixture occurs rapidly, whereas the loss of this activity in the incubation mixture occurs relatively slowly. The Hageman factor appeared to be necessary for the surface-bound kinin-generating activity, as this activity was absent in collagen exposed to Hageman factor-deficient plasma. These studies have partially characterized the plasma-derived enzymatic activity bound to collagen. Incubation of collagen with plasma caused a concentration-dependent reduction in the kinin-producing activity which was generated by the addition of ellagic acid, a known activator of plasma kallikrein. The kinin-inducing activity bound to collagen is inhibited by soybean trypsin inhibitor, Trasylol, serum C1̄ inactivator and the plasma α2-macroglobulin, but not by lima bean trypsin inhibitor. An eluate prepared from plasma-treated collagen, when compared with purified plasma kallikrein, shared a similar inhibitor profile. Selective chemical blockage of the free carboxyl groups on the collagen molecule, or heat denaturation, inactivated the ability of the collagen to generate kinin-like activity after incubation with plasma. Removal of the collagen telopeptides or blockage of the free amino groups failed to affect the collagen-plasma interaction. The binding of partially purified plasma kallikrein to collagen was found to have similar structural and chemical requirements. These data indicate that there is a structural and chemical specificity for the activation and binding of plasma kallikrein-like activity by collagen. These studies suggest that a plasma kallikrein may function as a surface-bound enzyme system. PMID:4338122

  4. Structure of collagen adsorbed on a model implant surface resolved by polarization modulation infrared reflection-absorption spectroscopy.

    PubMed

    Brand, Izabella; Habecker, Florian; Ahlers, Michael; Klüner, Thorsten

    2015-03-01

    The polarization modulation infrared reflection-absorption spectra of collagen adsorbed on a titania surface and quantum chemical calculations are used to describe components of the amide I mode to the protein structure at a sub-molecular level. In this study, imino acid rich and poor fragments, representing the entire collagen molecule, are taken into account. The amide I mode of the collagen triple helix is composed of three absorption bands which involve: (i) (∼1690cm(-1)) the CO stretching modes at unhydrated groups, (ii) (1655-1673cm(-1)) the CO stretching at carbonyl groups at imino acids and glycine forming intramolecular hydrogen bonds with H atoms at both NH2 and, unusual for proteins, CH2 groups at glycine at a neighbouring chain and (iii) (∼1640cm(-1)) the CO stretching at carbonyl groups forming hydrogen bonds between two, often charged, amino acids as well as hydrogen bonds to water along the entire helix. The IR spectrum of films prepared from diluted solutions (c<50μgml(-1)) corresponds to solution spectra indicating that native collagen molecules interact with water adsorbed on the titania surface. In films prepared from solutions (c⩾50μgml(-1)) collagen multilayers are formed. The amide I mode is blue-shifted by 18cm(-1), indicating that intramolecular hydrogen bonds at imino acid rich fragments are weakened. Simultaneous red-shift of the amide A mode implies that the strength of hydrogen bonds at the imino acid poor fragments increases. Theoretically predicted distortion of the collagen structure upon adsorption on the titania surface is experimentally confirmed. PMID:25498816

  5. Full-Length Fibronectin Drives Fibroblast Accumulation at the Surface of Collagen Microtissues during Cell-Induced Tissue Morphogenesis.

    PubMed

    Foolen, Jasper; Shiu, Jau-Ye; Mitsi, Maria; Zhang, Yang; Chen, Christopher S; Vogel, Viola

    2016-01-01

    Generating and maintaining gradients of cell density and extracellular matrix (ECM) components is a prerequisite for the development of functionality of healthy tissue. Therefore, gaining insights into the drivers of spatial organization of cells and the role of ECM during tissue morphogenesis is vital. In a 3D model system of tissue morphogenesis, a fibronectin-FRET sensor recently revealed the existence of two separate fibronectin populations with different conformations in microtissues, i.e. 'compact and adsorbed to collagen' versus 'extended and fibrillar' fibronectin that does not colocalize with the collagen scaffold. Here we asked how the presence of fibronectin might drive this cell-induced tissue morphogenesis, more specifically the formation of gradients in cell density and ECM composition. Microtissues were engineered in a high-throughput model system containing rectangular microarrays of 12 posts, which constrained fibroblast-populated collagen gels, remodeled by the contractile cells into trampoline-shaped microtissues. Fibronectin's contribution during the tissue maturation process was assessed using fibronectin-knockout mouse embryonic fibroblasts (Fn-/- MEFs) and floxed equivalents (Fnf/f MEFs), in fibronectin-depleted growth medium with and without exogenously added plasma fibronectin (full-length, or various fragments). In the absence of full-length fibronectin, Fn-/- MEFs remained homogenously distributed throughout the cell-contracted collagen gels. In contrast, in the presence of full-length fibronectin, both cell types produced shell-like tissues with a predominantly cell-free compacted collagen core and a peripheral surface layer rich in cells. Single cell assays then revealed that Fn-/- MEFs applied lower total strain energy on nanopillar arrays coated with either fibronectin or vitronectin when compared to Fnf/f MEFs, but that the presence of exogenously added plasma fibronectin rescued their contractility. While collagen decoration of

  6. Improving the moisturizing properties of collagen film by surface grafting of chondroitin sulfate for corneal tissue engineering.

    PubMed

    Liu, Yang; Lv, Huilin; Ren, Li; Xue, Guanhua; Wang, Yingjun

    2016-06-01

    Cornea disease is the second cause of blindness and keratoplasty is the most commonly performed option for visual rehabilitation of patients with corneal blindness. However, the clinical treatment has been drastically limited due to a severe shortage of high-quality donor corneas. Although collagen film with outstanding biocompatibility has promising application in corneal tissue engineering, the moisturizing properties of collagen-based materials must be further improved to satisfy the requirements of clinical applications. This paper describes a novel collagen-based film with high moisture capacity reinforced by surface grafting of chondroitin sulfate. The collagen-chondroitin sulfate (abbreviated as Col-CS) film was analyzed by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy and its hydrophilic property, moisture retention, optical property, and mechanical performance had been tested. The moisture-retaining capacity is found to be improved with the introduction of chondroitin sulfate, and the Col-CS membrane performs better mechanical properties than the collagen film. Moreover, the modified film proves excellent biocompatibility for the proliferation of human corneal epithelial cells in vitro. This Col-CS film with good moisturizing properties can reduce the risk of xerophthalmia and is expected to increase the implant success rate in clinic patients with corneal defects. PMID:26948819

  7. Biomedical applications of collagens.

    PubMed

    Ramshaw, John A M

    2016-05-01

    Collagen-based biomedical materials have developed into important, clinically effective materials used in a range of devices that have gained wide acceptance. These devices come with collagen in various formats, including those based on stabilized natural tissues, those that are based on extracted and purified collagens, and designed composite, biosynthetic materials. Further knowledge on the structure and function of collagens has led to on-going developments and improvements. Among these developments has been the production of recombinant collagen materials that are well defined and are disease free. Most recently, a group of bacterial, non-animal collagens has emerged that may provide an excellent, novel source of collagen for use in biomaterials and other applications. These newer collagens are discussed in detail. They can be modified to direct their function, and they can be fabricated into various formats, including films and sponges, while solutions can also be adapted for use in surface coating technologies. PMID:26448097

  8. Full-Length Fibronectin Drives Fibroblast Accumulation at the Surface of Collagen Microtissues during Cell-Induced Tissue Morphogenesis

    PubMed Central

    Foolen, Jasper; Shiu, Jau-Ye; Mitsi, Maria; Zhang, Yang; Chen, Christopher S.; Vogel, Viola

    2016-01-01

    Generating and maintaining gradients of cell density and extracellular matrix (ECM) components is a prerequisite for the development of functionality of healthy tissue. Therefore, gaining insights into the drivers of spatial organization of cells and the role of ECM during tissue morphogenesis is vital. In a 3D model system of tissue morphogenesis, a fibronectin-FRET sensor recently revealed the existence of two separate fibronectin populations with different conformations in microtissues, i.e. ‘compact and adsorbed to collagen’ versus ‘extended and fibrillar’ fibronectin that does not colocalize with the collagen scaffold. Here we asked how the presence of fibronectin might drive this cell-induced tissue morphogenesis, more specifically the formation of gradients in cell density and ECM composition. Microtissues were engineered in a high-throughput model system containing rectangular microarrays of 12 posts, which constrained fibroblast-populated collagen gels, remodeled by the contractile cells into trampoline-shaped microtissues. Fibronectin’s contribution during the tissue maturation process was assessed using fibronectin-knockout mouse embryonic fibroblasts (Fn-/- MEFs) and floxed equivalents (Fnf/f MEFs), in fibronectin-depleted growth medium with and without exogenously added plasma fibronectin (full-length, or various fragments). In the absence of full-length fibronectin, Fn-/- MEFs remained homogenously distributed throughout the cell-contracted collagen gels. In contrast, in the presence of full-length fibronectin, both cell types produced shell-like tissues with a predominantly cell-free compacted collagen core and a peripheral surface layer rich in cells. Single cell assays then revealed that Fn-/- MEFs applied lower total strain energy on nanopillar arrays coated with either fibronectin or vitronectin when compared to Fnf/f MEFs, but that the presence of exogenously added plasma fibronectin rescued their contractility. While collagen

  9. Characterization of a variety of standard collagen substrates: ultrastructure, uniformity, and capacity to bind and promote growth of neurons

    SciTech Connect

    Iversen, P.L.; Partlow, L.M.; Stensaas, L.J.; Moatamed, F.

    1981-06-01

    Collagen substrates were characterized after preparation by the four methods most commonly used for tissue culture (saline precipitation, exposure to ammonium hydroxide vapor, exposure to ultraviolet light, and air drying). Although roughly equivalent percentages of collagen were precipitated by each technique (87 to 97%), marked differences were found in surface uniformity and ultrastructure. Substrates were quite uniform if precipitated by exposure to ammonium hydroxide or ultraviolet light, of intermediate uniformity if saline precipitated, and not at all uniform if air dried. Scanning electron microscopy revealed that (a) ammonium hydroxide and saline precipitation primarily resulted in formation of collagen fibrils, (b) air drying produced a small number of fibrils plus a large amount of amorphous material, and (c) exposure to ultraviolet light only resulted in the formation of globular, nonfibrillar collagen aggregates. The capacity of collagen substrates to bind and grow neurons differed markedly with the method of preparation and the amount of collagen plated per unit area. Quantifications of binding and growth of both cerebral and sympathetic neurons revealed that these are separate measures of the biocompatibility of a surface and that growth was uniformly inferior on globular collagen that had been precipitated by ultraviolet light. Long-term (greater than or equal to 2 wk) growth of sympathetic neurons was optimal on thick beds of saline-precipitated collagen, whereas short-term growth was best on thin layers of either saline or ammonium hydroxide-precipitated collagen. Cerebral neurons bound and grew optimally on thick collagen beds after both short- and long-term culture. In addition, cerebral neurons were found to be more dependent on the method of precipitation of the thin collagen substrates than were sympathetic neurons.

  10. Enhanced stabilization of collagen by furfural.

    PubMed

    Lakra, Rachita; Kiran, Manikantan Syamala; Usha, Ramamoorthy; Mohan, Ranganathan; Sundaresan, Raja; Korrapati, Purna Sai

    2014-04-01

    Furfural (2-furancarboxaldehyde), a product derived from plant pentosans, has been investigated for its interaction with collagen. Introduction of furfural during fibril formation enhanced the thermal and mechanical stability of collagen. Collagen films treated with furfural exhibited higher denaturation temperature (Td) (p<0.04) and showed a 3-fold increase in Young's modulus (p<0.04) at higher concentration. Furfural and furfural treated collagen films did not have any cytotoxic effect. Rheological characterization showed an increase in shear stress and shear viscosity with increasing shear rate for treated collagen. Circular dichroism (CD) studies indicated that the furfural did not have any impact on triple helical structure of collagen. Scanning electron microscopy (SEM) of furfural treated collagen exhibited small sized porous structure in comparison with untreated collagen. Thus this study provides an alternate ecologically safe crosslinking agent for improving the stability of collagen for biomedical and industrial applications. PMID:24468046

  11. Gene expression of markers of osteogenic differentiation of human mesenchymal cells on collagen I-modified microrough titanium surfaces.

    PubMed

    Morra, M; Cassinelli, C; Cascardo, G; Bollati, D; Baena, R Rodriguez Y

    2011-02-01

    Microrough, doubly acid etched titanium surfaces (Ti) were further modified by amination and covalent coupling of fibrillar collagen type I (ColTi). Human Mesenchymal Cells (HMC) adhesion and growth, and relevant osteogenic differentiation in nonosteogenic (basal) medium were evaluated by fluorescence microscopy, scanning electron microscopy, and RT-PCR for a three-week period. Results show strongly enhanced HMC adhesion and cell density at short experimental time on ColTi, together with complete spreading of the cell body over the microrough surface topography. RT-PCR analysis of several genes involved in osteogenesis indicate, since the first week of culturing, significant progression of HMC on ColTi along the osteogenic pathway. These results indicate that the adopted process of surface immobilization of collagen, mandatory to impart collagenase resistance in implant sites, does not impair biospecific interactions between HMC and collagen. Thus, it is possible to upgrade properties arising from the control of Ti surfaces topography by surface-chemistry driven enhanced recruitment of precursor osteogenic cells and pro-osteogenic stimula. PMID:21171164

  12. The evolution of fibrillar collagens: a sea-pen collagen shares common features with vertebrate type V collagen.

    PubMed

    Tillet, E; Franc, J M; Franc, S; Garrone, R

    1996-02-01

    The extracellular matrix of marine primitive invertebrates (sponges, polyps and jellyfishes) contains collagen fibrils with narrow diameters. From various data, it has been hypothesized that these primitive collagens could represent ancestral forms of the vertebrate minor collagens, i.e., types V or XI. Recently we have isolated a primitive collagen from the soft tissues of the sea-pen Veretillum cynomorium. This report examines whether the sea-pen collagen shares some features with vertebrate type V collagen. Rotary shadowed images of acid-soluble collagen molecules extracted from beta-APN treated animals, positive staining of segment-long-spacing crystallites precipitated from pepsinized collagen, Western blots of the pepsinized alpha1 and alpha2 chains with antibodies to vertebrate types I, III and V collagens, and in situ gold immunolabeling of ECM collagen fibrils were examined. Our results showed that the tissue form of the sea-pen collagen is a 340-nm threadlike molecule, which is close to the vertebrate type V collagen with its voluminous terminal globular domain, the distribution of most of its polar amino-acid residues, and its antigenic properties. PMID:8653581

  13. Extraction optimization of pepsin-soluble collagen from eggshell membrane by response surface methodology (RSM).

    PubMed

    Mohammadi, Reza; Mohammadifar, Mohammad Amin; Mortazavian, Amir Mohammad; Rouhi, Milad; Ghasemi, Jahan B; Delshadian, Zohre

    2016-01-01

    Response surface methodology (RSM) was used to investigate the effect of extraction-process variables on pepsin-soluble collagen (PSC) from eggshell membrane. A central composite design (CCD) was employed for experimental design and analysis of the results to obtain the best possible combination of NaOH concentration (X1: 0.4-1.2 mol/l), alkali treatment time (X2: 6-30 h), enzyme concentration (X3: 15-75 U/mg) and hydrolysis time (X4: 12-60 h) for maximum PSC extraction. The experimental data obtained were fitted to a second-order polynomial equation using multiple regression analysis and analyzed by appropriate statistical methods. According to the results, optimum extraction conditions were as follows: NaOH concentration of 0.76 mol/l, alkali treatment time of 18 h, enzyme concentration of 50 U/mg and hydrolysis time of 43.42 h. The experimental extraction yield under optimal conditions was found to be 30.049%, which is in good agreement with the predicted value of 30.054%. PMID:26212959

  14. Widespread histologic distribution of the alpha 2 beta 1 integrin cell-surface collagen receptor.

    PubMed Central

    Zutter, M. M.; Santoro, S. A.

    1990-01-01

    The alpha 2 beta 1 integrin (platelet membrane glycoprotein Ia-IIa, VLA-2, ECMR-II) functions as a cell surface receptor for collagen. The authors have determined the histologic distribution of the alpha 2 beta 1 receptor in normal tissues by immunohistochemical technique. The studies revealed that the alpha 2 beta 1 receptor was expressed on fibroblasts, endothelial cells, and epithelial cells from multiple sites including skin, tonsil, breast, sweat gland, gastrointestinal tract, lung, bladder, cervix, and prostate. Follicular dendritic cells of the lymph node, tonsil, and spleen and dendritic cells of the thymus also expressed the alpha 2 beta 1 receptor. The receptor also was present on Schwann cells of ganglia and on neuroglia. Greatly enhanced expression of the receptor in regions of proliferating epithelium suggests that enhanced expression of alpha 2 beta 1 is associated with orderly, regulated cell proliferation. The circumferential staining pattern of the alpha 2 beta 1 integrin within many epithelia is virtually identical to that observed for other adhesive receptors, such as the cadherins, which have been implicated in cell-cell adhesion. Images Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 16 Figure 17 PMID:2164774

  15. Radial organization of interstitial exchange pathway and influence of collagen in synovium.

    PubMed Central

    Price, F M; Mason, R M; Levick, J R

    1995-01-01

    The synovial intercellular space is the path by which water, nutrients, cytokines, and macromolecules enter and leave the joint cavity. In this study two structural factors influencing synovial permeability were quantified by morphometry (Delesse's principle) of synovial electronmicrographs (rabbit knee), namely interstitial volume fraction Vv.1 and the fraction of the interstitium obstructed by collagen fibrils. Mean Vv.1 across the full thickness was 0.66 +/- 0.03 SEM (n = 11); but Vv.1 actually varied systematically with depth normal to the surface, increasing nonlinearly from 0.40 +/- 0.04 (n = 5 joints) near the free surface to 0.92 +/- 0.02 near the subsynovial interface. Tending to offset this increase in transport space, however, the space "blocked" by collagen fibrils also increased nonlinearly with depth. Bundles of collagen fibrils occupied 13.6 +/- 2.4% of interstitial volume close to the free surface but 49 +/- 4.8% near the subsynovial surface (full-thickness average, 40.5 +/- 3.5%), with fibrils accounting for 48.6-57.1% of the bundle space. Because of the two counteracting compositional gradients, the space available for fibril-excluded transport (hydraulic flow and macromolecular diffusion) was relatively constant > 4 microns below the surface but constricted at the synovium-cavity interface. The space available to extracellular polymers was only 51-53% of tissue volume, raising their effective concentration and hence the lining's resistance to flow and ability to confine the synovial fluid. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 7 PMID:8534813

  16. Atrial Fibrillation

    MedlinePlus

    ... cause the atria to fibrillate. The faulty signals flood the AV node with electrical impulses. As a ... people who have permanent AF, treatment can help control symptoms and prevent complications. Treatment may include medicines, ...

  17. Ventricular fibrillation

    MedlinePlus

    ... seconds, it can lead to fainting (syncope) or cardiac arrest. Fibrillation is an uncontrolled twitching or quivering of ... pubmed/23801105 . Myerburg RJ, Castellanos A. Approach to cardiac arrest and life-threatening arrhythmias. In: Goldman L, Schafer ...

  18. Eumelanin fibrils

    NASA Astrophysics Data System (ADS)

    McQueenie, Ross; Sutter, Jens; Karolin, Jan; Birch, David J. S.

    2012-07-01

    We describe the auto-oxidation of 3, 4-dihydroxy-L-phenylalanine (L-DOPA) in the synthesis of eumelanin to spontaneously produce fibrils upon drying. The self-assembled fibrils are of characteristic diameter ~1 to 2 μm, composed of filaments, and are unidirectional, apart from branches that are formed at typically an angle of 20 to 22 deg. The fibrils are characterized using fluorescence spectroscopy, fluorescence decay times, scanning electron microscopy, atomic force microscopy, and fluorescence lifetime imaging microscopy. The fibrils mimic natural melanin in consisting of core eumelanin with efficient nonradiative properties, but they also display pockets of electronically isolated species with higher radiative rates on the nanosecond timescale. Eumelanin fibrils formed occasionally in solution are tentatively attributed to a scaffold of bacteria or fungus. Fabricating and characterizing novel synthetic eumelanin structures such as fibrils are of interest in helping to reveal a functional structure for eumelanin, in understanding its photophysics, in learning more about L-DOPA as it is used in the treatment of Parkinson's disease, and in producing novel materials which might embody some of the diverse properties of eumelanin.

  19. Acid phosphatase activity and intracellular collagen degradation by fibroblasts in vitro.

    PubMed

    Yajima, T

    1986-01-01

    Human gingival fibroblasts were cultured with collagen fibrils. The precise process of collagen phagocytosis and the relationship between acid phosphatase activity and intracellular degradation of collagen were investigated by cytochemical methods at the ultrastructural level. The collagen fibrils were first engulfed at one end by cellular processes, or the cell membrane wrapped itself around the middle of the fibrils. Collagen phagocytosis induced acid phosphatase activity in the fibroblast Golgi-endoplasmic reticulum-lysosome system. By application of the tracer lanthanum, deposits were observed in the intercellular spaces and along the fibrils being phagocytosed. At this stage, primary lysosomes were seen in close proximity to the collagen being engulfed, but no signs of fusion were observed. When the fibrils had been interiorized in whole or in part, they ultimately became enclosed within phagosomes, and no tracer was observed along the interiorized portion of the fibrils. Primary lysosomes then fused with these collagen-containing phagosomes to form phagolysosomes. Collagen degradation occurred within these bodies even though the end of a fibril might have protruded outside of the cell. These results suggest that selective and controlled phagocytosis of collagen and intracellular digestion of it may play a central role in the physiological remodeling and metabolic breakdown of the collagen of connective tissues. PMID:3742560

  20. Radiating Amyloid Fibril Formation on the Surface of Lipid Membranes through Unit-Assembly of Oligomeric Species of α-Synuclein

    PubMed Central

    Lee, Jung-Ho; Hong, Chul-Suk; Lee, Soonkoo; Yang, Jee-Eun; Park, Yong Il; Lee, Daekyun; Hyeon, Taeghwan; Jung, Seunho; Paik, Seung R.

    2012-01-01

    Background Lewy body in the substantia nigra is a cardinal pathological feature of Parkinson's disease. Despite enormous efforts, the cause-and-effect relationship between Lewy body formation and the disorder is yet to be explicitly unveiled. Methodology/Principal Findings Here, we showed that radiating amyloid fibrils (RAFs) were instantly developed on the surface of synthetic lipid membranes from the β-sheet free oligomeric species of α-synuclein through a unit-assembly process. The burgeoning RAFs were successfully matured by feeding them with additional oligomers, which led to concomitant dramatic shrinkage and disintegration of the membranes by pulling off lipid molecules to the extending fibrils. Mitochondria and lysosomes were demonstrated to be disrupted by the oligomeric α-synuclein via membrane-dependent fibril formation. Conclusion The physical structure formation of amyloid fibrils, therefore, could be considered as detrimental to the cells by affecting membrane integrity of the intracellular organelles, which might be a molecular cause for the neuronal degeneration observed in Parkinson's disease. PMID:23077644

  1. Ultrastructural quantification of collagen in human chordae tendineae.

    PubMed Central

    Berkovitz, B K; Rollinson, C

    1993-01-01

    OBJECTIVE--To determine collagen fibril diameter distributions in the chordae tendineae and to see whether there are any differences between right and left sides of the human heart. DESIGN--Collagen fibril diameters and the percentage volume occupied by collagen fibrils (as opposed to ground substance) were determined by means of a planimeter with a digitising tablet from electron micrographs printed at a magnification of 136; 000. MATERIAL--Human chordae tendineae were obtained at postmortem examination from seven subjects aged 50-75 years. MEASUREMENTS AND MAIN RESULTS--Histograms of collagen fibril diameter distributions showed a sharp, unimodal distribution. The mean collagen fibril diameters associated with the tricuspid and mitral valves were 41.1 nm and 40.5 nm respectively. The percentage volume occupied by collagen was about 39% for each valve. There were no significant differences for any of these values between the right and left sides of the heart. CONCLUSIONS--The greater pressures present on the left side of the heart are not reflected in any difference in collagen fibril diameters within the chordae tendineae compared with the right side. Images PMID:8461224

  2. Fabrication and evaluation of a biodegradable cohesive plug based on reconstituted collagen/γ-polyglutamic acid.

    PubMed

    Hsu, Fu-Yin; Cheng, Ya-Yun; Tsai, Shiao-Wen; Tsai, Wei-Bor

    2010-10-01

    In the past decade, numerous studies have been devoted to developing natural bioadhesives that have the notable capacity to adhere to wet surfaces. Collagen and γ-polyglutamic acid (γ-PGA) are well-known natural hydrophilic polymers that have both been utilized for their versatility in a wide range of biomedical applications. The aim of this study was the construction and characterization of a cohesive plug composed of γ-PGA and reconstituted collagen fibrils crosslinked with water-soluble carbodiimide. Transmission electron microscopy examinations confirmed that the collagen fibrils in the reconstituted collagen/γ-PGA gel retained their native specific D-period structure. This unique D-pattern structure of collagen plays a major role in hemostasis and is also related to several cellular behaviors. The bonding strength of the reconstituted collagen/γ-PGA adhesive was approximately 42.9 ± 4.0 KPa after 5 min of application and increased to 76.5 ± 15.1 KPa after 24 h. This was much stronger than the fibrin adhesive, whose bonding strength was 30.9 ± 0.2 KPa. Furthermore, the reconstituted collagen/γ-PGA gel degraded gradually after subcutaneous implantation in the backs of rats over a period of 8 weeks, without any severe inflammatory response. On the basis of the histological analysis, fibroblasts migrated into the gel while it degraded, which indicates that the gel is not harmful to cellular activity. Together, these findings demonstrate that using reconstituted collagen with retained D-periodicity as a component of the bioadhesive is a possibly better option to formulate effective adhesiveness and is promising as a scaffold for tissue repair. PMID:20665682

  3. High Membrane Curvature Enhances Binding, Conformational Changes, and Fibrillation of Amyloid-β on Lipid Bilayer Surfaces.

    PubMed

    Sugiura, Yuuki; Ikeda, Keisuke; Nakano, Minoru

    2015-10-27

    Aggregation of the amyloid-β (Aβ) protein and the formation of toxic aggregates are the possible pathogenic pathways in Alzheimer's disease. Accumulating evidence suggests that lipid membranes play key roles in protein aggregation, although the intermolecular forces that drive the interactions between Aβ-(1-40) and the membranes vary in different membrane systems. Here, we observed that a high positive curvature of lipid vesicles with diameters of ∼30 nm enhanced the association of Aβ with anionic phosphatidylglycerol membranes in the liquid-crystalline phase and with zwitterionic phosphatidylcholine membranes in the gel phase. The binding modes of Aβ to these membranes differ in terms of the location of the protein on the membrane and of the protein secondary structure. The fibrillation of Aβ was accelerated in the presence of the vesicles and at high protein-to-lipid ratios. Under these conditions, the protein accumulated on the surfaces, as demonstrated by a high (10(7) M(-1)) binding constant. Our findings suggest that packing defects on membranes with high curvatures, such as the intraluminal vesicles in multivesicular bodies and the exosomes, might result in the accumulation of toxic protein aggregates. PMID:26474149

  4. A continuum model for hierarchical fibril assembly

    NASA Astrophysics Data System (ADS)

    van Lith, B. S.; Muntean, A.; Storm, C.

    2014-06-01

    Most of the biological polymers that make up our cells and tissues are hierarchically structured. For biopolymers ranging from collagen, to actin, to fibrin and amyloid fibrils this hierarchy provides vitally important versatility. The structural hierarchy must be encoded in the self-assembly process, from the earliest stages onward, in order to produce the appropriate substructures. In this letter, we explore the kinetics of multistage self-assembly processes in a model system which allows comparison to bulk probes such as light scattering. We apply our model to recent turbidimetry data on the self-assembly of collagen fibrils. Our analysis suggests a connection between diffusion-limited aggregation kinetics and fibril growth, supported by slow, power-law growth at very long time scales.

  5. Changes in collagenous tissue microstructures and distributions of cathepsin L in body wall of autolytic sea cucumber (Stichopus japonicus).

    PubMed

    Liu, Yu-Xin; Zhou, Da-Yong; Ma, Dong-Dong; Liu, Yan-Fei; Li, Dong-Mei; Dong, Xiu-Ping; Tan, Ming-Qian; Du, Ming; Zhu, Bei-Wei

    2016-12-01

    The autolysis of sea cucumber (Stichopus japonicus) was induced by ultraviolet (UV) irradiation, and the changes of microstructures of collagenous tissues and distributions of cathepsin L were investigated using histological and histochemical techniques. Intact collagen fibers in fresh S. japonicus dermis were disaggregated into collagen fibrils after UV stimuli. Cathepsin L was identified inside the surface of vacuoles in the fresh S. japonicus dermis cells. After the UV stimuli, the membranes of vacuoles and cells were fused together, and cathepsin L was released from cells and diffused into tissues. The density of cathepsin L was positively correlated with the speed and degree of autolysis in different layers of body wall. Our results revealed that lysosomal cathepsin L was released from cells in response to UV stimuli, which contacts and degrades the extracellular substrates such as collagen fibers, and thus participates in the autolysis of S. japonicus. PMID:27374541

  6. Analysis of forward and backward Second Harmonic Generation images to probe the nanoscale structure of collagen within bone and cartilage.

    PubMed

    Houle, Marie-Andrée; Couture, Charles-André; Bancelin, Stéphane; Van der Kolk, Jarno; Auger, Etienne; Brown, Cameron; Popov, Konstantin; Ramunno, Lora; Légaré, François

    2015-11-01

    Collagen ultrastructure plays a central role in the function of a wide range of connective tissues. Studying collagen structure at the microscopic scale is therefore of considerable interest to understand the mechanisms of tissue pathologies. Here, we use second harmonic generation microscopy to characterize collagen structure within bone and articular cartilage in human knees. We analyze the intensity dependence on polarization and discuss the differences between Forward and Backward images in both tissues. Focusing on articular cartilage, we observe an increase in Forward/Backward ratio from the cartilage surface to the bone. Coupling these results to numerical simulations reveals the evolution of collagen fibril diameter and spatial organization as a function of depth within cartilage. PMID:26349534

  7. Magnetic Resonance Microscopy of Collagen Mineralization

    PubMed Central

    Chesnick, Ingrid E.; Mason, Jeffrey T.; Giuseppetti, Anthony A.; Eidelman, Naomi; Potter, Kimberlee

    2008-01-01

    A model mineralizing system was subjected to magnetic resonance microscopy to investigate how water proton transverse (T2) relaxation times and magnetization transfer ratios can be applied to monitor collagen mineralization. In our model system, a collagen sponge was mineralized with polymer-stabilized amorphous calcium carbonate. The lower hydration and water proton T2 values of collagen sponges during the initial mineralization phase were attributed to the replacement of the water within the collagen fibrils by amorphous calcium carbonate. The significant reduction in T2 values by day 6 (p < 0.001) was attributed to the appearance of mineral crystallites, which were also detected by x-ray diffraction and scanning electron microscopy. In the second phase, between days 6 and 13, magnetic resonance microscopy properties appear to plateau as amorphous calcium carbonate droplets began to coalesce within the intrafibrillar space of collagen. In the third phase, after day 15, the amorphous mineral phase crystallized, resulting in a reduction in the absolute intensity of the collagen diffraction pattern. We speculate that magnetization transfer ratio values for collagen sponges, with similar collagen contents, increased from 0.25 ± 0.02 for control strips to a maximum value of 0.31 ± 0.04 at day 15 (p = 0.03) because mineral crystals greatly reduce the mobility of the collagen fibrils. PMID:18487295

  8. OPTIMIZING COLLAGEN TRANSPORT THROUGH TRACK-ETCHED NANOPORES

    PubMed Central

    Bueno, Ericka M.; Ruberti, Jeffrey W.

    2008-01-01

    Polymer transport through nanopores is a potentially powerful tool for separation and organization of molecules in biotechnology applications. Our goal is to produce aligned collagen fibrils by mimicking cell-mediated collagen assembly: driving collagen monomers in solution through the aligned nanopores in track-etched membranes followed by fibrillogenesis at the pore exit. We examined type I atelo-collagen monomer transport in neutral, cold solution through polycarbonate track-etched membranes comprising 80-nm-diameter, 6-μm-long pores at 2% areal fraction. Source concentrations of 1.0, 2.8 and 7.0 mg/ml and pressure differentials of 0, 10 and 20 inH2O were used. Membrane surfaces were hydrophilized via covalent poly(ethylene-glycol) binding to limit solute-membrane interaction. Collagen transport through the nanopores was a non-intuitive process due to the complex behavior of this associating molecule in semi-dilute solution. Nonetheless, a modified open pore model provided reasonable predictions of transport parameters. Transport rates were concentration- and pressure-dependent, with diffusivities across the membrane in semi-dilute solution two-fold those in dilute solution, possibly via cooperative diffusion or polymer entrainment. The most significant enhancement of collagen transport was accomplished by membrane hydrophilization. The highest concentration transported (5.99±2.58 mg/ml) with the highest monomer flux (2.60±0.49 ×103 molecules s-1 pore-1) was observed using 2.8 mg collagen/ml, 10 inH2O and hydrophilic membranes. PMID:21394216

  9. Precipitation of Calcium Phosphates in the Presence of Collagen Type I on Four Different Bioactive Titanium Surfaces: an in Vitro Study

    PubMed Central

    Olander, Julia; Kjellin, Per; Currie, Fredrik; Sul, Young-Taeg; Anna, Arvidsson

    2015-01-01

    ABSTRACT Objectives To compare the properties of calcium phosphate precipitation on four different bioactive surface preparations and one control surface in the simulated body fluid model with added collagen type I. Material and Methods Blasted titanium discs were treated with four different surface modifications, alkali and heat, sodium fluoride, anodic oxidation and hydroxyapatite coating. The discs were divided into five groups where one group, the blasted, served as control. The discs were immersed in simulated body fluid and collagen for 24 h, 3 days, 1 week and 2 weeks and then analysed by optical interferometry, scanning electron microscopy/energy dispersive X-ray analysis and X-ray photoelectron spectroscopy. Results All surfaces show small precipitates after 3 days which with longer immersion times increase. After 2 weeks the surfaces were completely covered with precipitates, and Ca/P ratios were approximately 1.3, independently on surface preparation. The fluoridated discs showed significantly (P ≤ 0.05) higher degree of CaP after one week of immersion as compared to the other surface preparations. The collagen type I content increased with time, as reflected by increased nitrogen content. Conclusions The results from this study indicate that a fluoridated titanium surface may favour precipitation of calcium phosphate in the presence of collagen type I, as compared to the other surface treatments of the present study. PMID:26904178

  10. The effect of gamma irradiation on injectable human amnion collagen

    SciTech Connect

    Liu, B.C.; Harrell, R.; Davis, R.H.; Dresden, M.H.; Spira, M. )

    1989-08-01

    The effect of gamma irradiation on the physicochemical properties of injectable human amnion collagen was investigated. Pepsin-extracted human amnion collagen was purified, reconstituted, and irradiated with varying doses of gamma irradiation (0.25 Mrads to 2.5 Mrads). Gamma irradiation had a significant impact on the physical characteristics of the collagen. The neutral solubility of collagen in PBS at 45{degrees}C was decreased from 100% for the nonirradiated control sample to 16% for the 2.5 Mrads irradiated sample. SDS polyacrylamide gel electrophoresis also demonstrated the dose-dependent effect of gamma irradiation on collagen cross-links. Electron microscopic observation revealed that even at low irradiation dose (0.25 Mrads), collagen fibril diameter increased. The average diameter was 50 nm for nonirradiated control fibrils, while 4.4% of the irradiated collagen fibrils had a diameter greater than 100 nm. Irradiated collagen showed little evidence of damage. Well-preserved cross-striations were found in collagen fibrils at all doses of irradiation. Native amnion collagen irradiated with gamma rays demonstrated a slight increase in resistance to collagenase degradation compared with nonirradiated native collagen samples. Increased resistance to collagenase did not correlate with increasing irradiation dose. After 30 min of incubation at 37{degrees}C, both irradiated and nonirradiated collagen was completely digested by collagenase. However, gamma-irradiated collagen did become more sensitive to hydrolysis by trypsin. The higher the irradiation doses used, the greater sensitivity to trypsin was observed. At 0.25 Mrads irradiation only a slight increase was found. No marked differences in amino acid composition were noted among the high dose irradiated, low dose irradiated and control amnion collagen.

  11. Constructing bio-layer of heparin and type IV collagen on titanium surface for improving its endothelialization and blood compatibility.

    PubMed

    Zhang, Kun; Chen, Jun-ying; Qin, Wei; Li, Jing-an; Guan, Fang-xia; Huang, Nan

    2016-04-01

    The modification of cardiovascular stent surface for a better micro-environment has gradually changed to multi-molecule, multi-functional designation. In this study, heparin (Hep) and type IV collagen (IVCol) were used as the functional molecule to construct a bifunctional micro-environment of anticoagulation and promoting endothelialization on titanium (Ti). The surface characterization results (AFM, Alcian Blue 8GX Staining and fluorescence staining of IVCol) indicated that the bio-layer of Hep and IVCol were successfully fabricated on the Ti surface through electrostatic self-assembly. The APTT and platelet adhesion test demonstrated that the bionic layer possessed better blood compatibility compared with Ti surface. The adhesion, proliferation, migration and apoptosis tests of endothelial cells proved that the Hep/IVCol layer was able to enhance the endothelialization of the Ti surface. The in vivo animal implantation results manifested that the bionic surface could encourage new endothelialization. This work provides an important reference for the construction of multifunction micro-environment on the cardiovascular scaffold surface. PMID:26936367

  12. Fluorescent nanonetworks: A novel bioalley for collagen scaffolds and Tissue Engineering

    PubMed Central

    Nidhin, Marimuthu; Vedhanayagam, Mohan; Sangeetha, Selvam; Kiran, Manikantan Syamala; Nazeer, Shaiju S.; Jayasree, Ramapurath S.; Sreeram, Kalarical Janardhanan; Nair, Balachandran Unni

    2014-01-01

    Native collagen is arranged in bundles of aligned fibrils to withstand in vivo mechanical loads. Reproducing such a process under in vitro conditions has not met with major success. Our approach has been to induce nanolinks, during the self-assembly process, leading to delayed rather than inhibited fibrillogenesis. For this, a designed synthesis of nanoparticles - using starch as a template and a reflux process, which would provide a highly anisotropic (star shaped) nanoparticle, with large surface area was adopted. Anisotropy associated decrease in Morin temperature and superparamagnetic behavior was observed. Polysaccharide on the nanoparticle surface provided aqueous stability and low cytotoxicity. Starch coated nanoparticles was utilized to build polysaccharide - collagen crosslinks, which supplemented natural crosslinks in collagen, without disturbing the conformation of collagen. The resulting fibrillar lamellae showed a striking resemblance to native lamellae, but had a melting and denaturation temperature higher than native collagen. The biocompatibility and superparamagnetism of the nanoparticles also come handy in the development of stable collagen constructs for various biomedical applications, including that of MRI contrast agents. PMID:25095810

  13. Negative staining and immunoelectron microscopy of adhesion-deficient mutants of Streptococcus salivarius reveal that the adhesive protein antigens are separate classes of cell surface fibril.

    PubMed Central

    Weerkamp, A H; Handley, P S; Baars, A; Slot, J W

    1986-01-01

    The subcellular distribution of the cell wall-associated protein antigens of Streptococcus salivarius HB, which are involved in specific adhesive properties of the cells, was studied. Mutants which had lost the adhesive properties and lacked the antigens at the cell surface were compared with the parent strain. Immunoelectron microscopy of cryosections of cells labeled with affinity-purified, specific antisera and colloidal gold-protein A complexes was used to locate the antigens. Antigen C (AgC), a glycoprotein involved in attachment to host surfaces, was mainly located in the fibrillar layer outside the cell wall. A smaller amount of label was also found throughout the cytoplasmic area in the form of small clusters of gold particles, which suggests a macromolecular association. Mutant HB-7, which lacks the wall-associated AgC, accumulated AgC reactivity intracellularly. Intracellular AgC was often found associated with isolated areas of increased electron density, but sometimes seemed to fill the entire interior of the cell. Antigen B (AgB), a protein responsible for interbacterial coaggregation, was also located in the fibrillar layer, although its distribution differed from that of the wall-associated AgC since AgB was found predominantly in the peripheral areas. A very small amount of label was also found in the cytoplasmic area as discrete gold particles. Mutant HB-V5, which lacks wall-associated AgB, was not labeled in the fibrillar coat, but showed the same weak intracellular label as the parent strain. Immunolabeling with serum against AgD, another wall-associated protein but of unknown function, demonstrated its presence in the fibrillar layer of strain HB. Negatively stained preparations of whole cells of wild-type S. salivarius and mutants that had lost wall-associated AgB or AgC revealed that two classes of short fibrils are carried on the cell surface at the same time. AgB and AgC are probably located on separate classes of short, protease

  14. Increase in the water contact angle of composite film surfaces caused by the assembly of hydrophilic nanocellulose fibrils and nanoclay platelets.

    PubMed

    Wu, Chun-Nan; Saito, Tsuguyuki; Yang, Quanling; Fukuzumi, Hayaka; Isogai, Akira

    2014-08-13

    Controlling the assembly modes of different crystalline nanoparticles in composites is important for the expression of specific characteristics of the assembled structures. We report a unique procedure for increasing water contact angles (CAs) of composite film surfaces via the assembly of two different hydrophilic components, nanocellulose fibrils and nanoclay platelets. The nanocellulose fibrils and nanoclay platelets used have ionic groups on their surfaces in high densities (∼1 mmol g(-1)) and have no hydrophobic surface. The increase in the CA of the nanocellulose/nanoclay composite films was thus analyzed on the basis of the air area fractions of their nanostructured surfaces following Cassie's law. The air area fractions were geographically estimated from the atomic force microscopy height profiles of the composite film surfaces. The CAs of the composite film surfaces were found to be well described by Cassie's law. Interestingly, the composite films consisting of two hydrophilic nanoelements with different shapes exhibited CAs larger than those of the individual neat films. PMID:24977651

  15. Quantification of Collagen Organization in the Peripheral Human Cornea at Micron-Scale Resolution

    PubMed Central

    Boote, Craig; Kamma-Lorger, Christina S.; Hayes, Sally; Harris, Jonathan; Burghammer, Manfred; Hiller, Jennifer; Terrill, Nicholas J.; Meek, Keith M.

    2011-01-01

    The collagen microstructure of the peripheral cornea is important in stabilizing corneal curvature and refractive status. However, the manner in which the predominantly orthogonal collagen fibrils of the central cornea integrate with the circumferential limbal collagen is unknown. We used microfocus wide-angle x-ray scattering to quantify the relative proportion and orientation of collagen fibrils over the human corneolimbal interface at intervals of 50 μm. Orthogonal fibrils changed direction 1–1.5 mm before the limbus to integrate with the circumferential limbal fibrils. Outside the central 6 mm, additional preferentially aligned collagen was found to reinforce the cornea and limbus. The manner of integration and degree of reinforcement varied significantly depending on the direction along which the limbus was approached. We also employed small-angle x-ray scattering to measure the average collagen fibril diameter from central cornea to limbus at 0.5 mm intervals. Fibril diameter was constant across the central 6 mm. More peripherally, fibril diameter increased, indicative of a merging of corneal and scleral collagen. The point of increase varied with direction, consistent with a scheme in which the oblique corneal periphery is reinforced by chords of scleral collagen. The results have implications for the cornea's biomechanical response to ocular surgeries involving peripheral incision. PMID:21723812

  16. Anisotropic collagen fibrillogenesis within microfabricated scaffolds: implications for biomimetic tissue engineering.

    PubMed

    Jean, Aurélie; Engelmayr, George C

    2012-01-11

    Anisotropic collagen fibrillogenesis is demonstrated within the pores of an accordion-like honeycomb poly(glycerol sebacate) tissue engineering scaffold. Confocal reflectance microscopy and image analysis demonstrate increased fibril distribution order, fibril density, and alignment in accordion-like honeycomb pores compared with collagen gelled unconstrained. Finite element modeling predicts how collagen gel and scaffold mechanics couple in matching native heart muscle stiffness and anisotropy. PMID:23184695

  17. Fibrillar assembly and stability of collagen coating on titanium for improved osteoblast responses.

    PubMed

    Kim, Hae-Won; Li, Long-Hao; Lee, Eun-Jung; Lee, Su-Hee; Kim, Hyoun-Ee

    2005-12-01

    Collagen, as a major constituent of human connective tissues, has been regarded as one of the most important biomaterials. As a coating moiety on Ti hard-tissue implants, the collagen has recently attracted a great deal of attention. This article reports the effects of fibrillar assembly and crosslinking of collagen on its chemical stability and the subsequent osteoblastic responses. The fibrillar self-assembly of collagen was carried out by incubating acid-dissolved collagen in an ionic-buffered medium at 37 degrees C. The degree of assembly was varied with the incubation time and monitored by the turbidity change. The differently assembled collagen was coated on the Ti and crosslinked with a carbodiimide derivative. The partially assembled collagen contained fibrils with varying diameters as well as nonfibrillar aggregates. On the other hand, the fully assembled collagen showed the complete formation of fibrils with uniform diameters of approximately 100-200 nm with periodic stain patterns within the fibrils, which are typical of native collagen fibers. Through this fibrillar assembly, the collagen coating had significantly improved chemical stability in both the saline and collagenase media. The subsequent crosslinking step also improved the stability of the collagen coating, particularly in the unassembled collagen. The fibrillar assembly and the crosslinking of collagen significantly influenced the osteoblastic cell responses. Without the assembly, the collagen layer on Ti adversely affected the cell attachment and proliferation. However, those cellular responses were improved significantly when the collagen was assembled to fibrils and the assembly degree was increased. After crosslinking the collagen coating, these cellular responses were significantly enhanced in the case of the unassembled collagen but were not altered much in the assembled collagen. Based on these observations, it is suggested that the fibrillar assembly and the crosslinking of collagen

  18. Surface Binding of TOTAPOL Assists Structural Investigations of Amyloid Fibrils by Dynamic Nuclear Polarization NMR Spectroscopy.

    PubMed

    Nagaraj, Madhu; Franks, Trent W; Saeidpour, Siavash; Schubeis, Tobias; Oschkinat, Hartmut; Ritter, Christiane; van Rossum, Barth-Jan

    2016-07-15

    Dynamic nuclear polarization (DNP) NMR can enhance sensitivity but often comes at the price of a substantial loss of resolution. Two major factors affect spectral quality: low-temperature heterogeneous line broadening and paramagnetic relaxation enhancement (PRE) effects. Investigations by NMR spectroscopy, isothermal titration calorimetry (ITC), and EPR revealed a new substantial affinity of TOTAPOL to amyloid surfaces, very similar to that shown by the fluorescent dye thioflavin-T (ThT). As a consequence, DNP spectra with remarkably good resolution and still reasonable enhancement could be obtained at very low TOTAPOL concentrations, typically 400 times lower than commonly employed. These spectra yielded several long-range constraints that were difficult to obtain without DNP. Our findings open up new strategies for structural studies with DNP NMR spectroscopy on amyloids that can bind the biradical with affinity similar to that shown towards ThT. PMID:27147408

  19. Atrial Fibrillation and Stroke

    MedlinePlus

    ... Find People About NINDS NINDS Atrial Fibrillation and Stroke Information Page Table of Contents (click to jump ... done? Clinical Trials What is Atrial Fibrillation and Stroke? Atrial fibrillation (AF) describes the rapid, irregular beating ...

  20. Living with Atrial Fibrillation

    MedlinePlus

    ... Topics » Atrial Fibrillation » Living With Atrial Fibrillation Explore Atrial Fibrillation What Is... Types Other Names Causes Who Is at Risk Signs & Symptoms Diagnosis Treatments Prevention Living With Clinical Trials Links Related Topics Arrhythmia ...

  1. Stable immobilization of rat hepatocytes as hemispheroids onto collagen-conjugated poly-dimethylsiloxane (PDMS) surfaces: importance of direct oxygenation through PDMS for both formation and function.

    PubMed

    Nishikawa, Masaki; Yamamoto, Takatoki; Kojima, Nobuhiko; Kikuo, Komori; Fujii, Teruo; Sakai, Yasuyuki

    2008-04-15

    The highly oxygen-permeable material, poly-dimethylsiloxane (PDMS), has the potential to be applied to cell culture microdevices, but cell detachment from PDMS has been a major problem. In this study, we demonstrate that a combination of collagen covalently immobilized PDMS and an adequate oxygen supply enables the establishment of a stable, attached spheroid (hemispheroid) culture of rat hepatocytes. The bottom PDMS surfaces were first treated with oxygen plasma, then coupled with aminosilane followed by a photoreactive crosslinker, and they were finally reacted with a collagen solution. X-ray photoelectron spectroscopy (XPS) and contact angle measurements showed that the covalent immobilization of collagen on the surface occurred only where the crosslinker had been introduced. On the collagen-conjugated PDMS surface, rat hepatocytes organized themselves into hemispheroids and maintained the viability and a remarkably high albumin production at least for 2 weeks of culture. In contrast, hepatocytes on the other types of PDMS surfaces formed suspended spheroids that had low albumin production. In addition, we showed that blocking the oxygen supply through the bottom PDMS surface inhibited the formation of hemispheroids and the augmentation of hepatocellular function. These results show that appropriate surface modification of PDMS is a promising approach towards the development of liver tissue microdevices. PMID:17969156

  2. Anisotropy of Chemical Bonds in Collagen Molecules Studied by X-ray Absorption Near-Edge Structure (XANES) Spectroscopy

    PubMed Central

    Lam, Raymond S.K.; Metzler, Rebecca A.; Gilbert, Pupa U.P.A.; Beniash, Elia

    2012-01-01

    Collagen type I fibrils are the major building blocks of connective tissues. Collagen fibrils are anisotropic supra-molecular structures, and their orientation can be revealed by polarized light microscopy and vibrational microspectroscopy. We hypothesized that the anisotropy of chemical bonds in the collagen molecules, and hence their orientation, might also be detected by X-ray photoemission electron spectromicroscopy (X-PEEM) and X-ray absorption near-edge structure (XANES) spectroscopy, which use linearly polarized synchrotron light. To test this hypothesis, we analyzed sections of rat-tail tendon, composed of parallel arrays of collagen fibrils. The results clearly indicate that XANES-PEEM is sensitive to collagen fibril orientation and, more specifically, to the orientations of carbonyl and amide bonds in collagen molecules. These data suggest that XANES-PEEM is a promising technique for characterizing the chemical composition and structural organization at the nanoscale of collagen-based connective tissues, including tendons, cartilage, and bone. PMID:22148847

  3. Collagen telopeptides (cross-linking sites) play a role in collagen gel lattice contraction

    NASA Technical Reports Server (NTRS)

    Woodley, D. T.; Yamauchi, M.; Wynn, K. C.; Mechanic, G.; Briggaman, R. A.

    1991-01-01

    Solubilized interstitial collagens will form a fibrillar, gel-like lattice when brought to physiologic conditions. In the presence of human dermal fibroblasts the collagen lattice will contract. The rate of contraction can be determined by computer-assisted planemetry. The mechanisms involved in contraction are as yet unknown. Using this system it was found that the rate of contraction was markedly decreased when collagen lacking telopeptides was substituted for native collagen. Histidinohydroxylysinonorleucine (HHL) is a major stable trifunctional collagen cross-link in mature skin that involves a carboxyl terminal, telopeptide site 16c, the sixteenth amino acid residue from the carboxy terminal of the telopeptide region of alpha 1 (I) in type I collagen. Little, if any, HHL was present in native, purified, reconstituted, soluble collagen fibrils from 1% acetic acid-extracted 2-year-old bovine skin. In contrast, HHL cross-links were present (0.22 moles of cross-link per mole of collagen) in lattices of the same collagen contracted by fibroblasts. However, rat tail tendon does not contain HHL cross-links, and collagen lattices made of rat tail tendon collagen are capable of contraction. This suggests that telopeptide sites, and not mature HHL cross-links per se, are essential for fibroblasts to contract collagen lattices. Beta-aminopropionitrile fumarate (BAPN), a potent lathyrogen that perturbs collagen cross-linking by inhibition of lysyl oxidase, also inhibited the rate of lattice cell contraction in lattices composed of native collagen. However, the concentrations of BAPN that were necessary to inhibit the contraction of collagen lattices also inhibited fibroblast growth suggestive of cellular toxicity. In accordance with other studies, we found no inhibition of the rate of lattice contraction when fibronectin-depleted serum was used. Electron microscopy of contracted gels revealed typical collagen fibers with a characteristic axial periodicity. The data

  4. Nanoparticle-Templated Formation and Growth Mechanism of Curved Protein Polymer Fibrils.

    PubMed

    Pham, Thao T H; Rombouts, Wolf H; Fokkink, Remco; Stuart, Marc C A; Cohen Stuart, Martien A; Kleijn, J Mieke

    2016-07-11

    We investigated the growth of biosynthetic protein polymers with templated curvature on pluronic nanospheres. The protein has a central silk-like block containing glutamic residues (S(E)) and collagen-like end-blocks (C). The S(E) blocks stack into filaments when their charge is removed (pH <5). Indeed, at low pH curved and circular fibers are formed at the surface of the nanospheres, which keep their shape after removal of the pluronics. The data reveal the mechanism of the templated fibril-growth: The growth of protein assemblies is nucleated in solution; small protein fibrils adsorb on the nanospheres, presumably due to hydrogen bond formation between the silk-like blocks and the pluronic PEO blocks. The surface of the pluronic particles templates further growth. At relatively low protein/pluronic weight ratios, only a fraction of the nanospheres bears protein fibers, pointing to a limiting amount of nuclei in solution. Because the nanospheres capture fibrils at an early stage of growth, they can be used to separate growth and nucleation rates in protein fibril formation. Moreover, the nanoparticle-templated growth of stable curved fibers opens ways to build proteinaceous nanocapsules from designed protein polymers. PMID:27250876

  5. Atrial fibrillation or flutter

    MedlinePlus

    ... causes of atrial fibrillation include: Alcohol use (especially binge drinking) Coronary artery disease Heart attack or heart ... conditions that cause atrial fibrillation and flutter. Avoid binge drinking.

  6. Manipulation of in vitro collagen matrix architecture for scaffolds of improved physiological relevance

    NASA Astrophysics Data System (ADS)

    Hapach, Lauren A.; VanderBurgh, Jacob A.; Miller, Joseph P.; Reinhart-King, Cynthia A.

    2015-12-01

    Type I collagen is a versatile biomaterial that is widely used in medical applications due to its weak antigenicity, robust biocompatibility, and its ability to be modified for a wide array of applications. As such, collagen has become a major component of many tissue engineering scaffolds, drug delivery platforms, and substrates for in vitro cell culture. In these applications, collagen constructs are fabricated to recapitulate a diverse set of conditions. Collagen fibrils can be aligned during or post-fabrication, cross-linked via numerous techniques, polymerized to create various fibril sizes and densities, and copolymerized into a wide array of composite scaffolds. Here, we review approaches that have been used to tune collagen to better recapitulate physiological environments for use in tissue engineering applications and studies of basic cell behavior. We discuss techniques to control fibril alignment, methods for cross-linking collagen constructs to modulate stiffness, and composite collagen constructs to better mimic physiological extracellular matrix.

  7. Manipulation of in vitro collagen matrix architecture for scaffolds of improved physiological relevance.

    PubMed

    Hapach, Lauren A; VanderBurgh, Jacob A; Miller, Joseph P; Reinhart-King, Cynthia A

    2015-01-01

    Type I collagen is a versatile biomaterial that is widely used in medical applications due to its weak antigenicity, robust biocompatibility, and its ability to be modified for a wide array of applications. As such, collagen has become a major component of many tissue engineering scaffolds, drug delivery platforms, and substrates for in vitro cell culture. In these applications, collagen constructs are fabricated to recapitulate a diverse set of conditions. Collagen fibrils can be aligned during or post-fabrication, cross-linked via numerous techniques, polymerized to create various fibril sizes and densities, and copolymerized into a wide array of composite scaffolds. Here, we review approaches that have been used to tune collagen to better recapitulate physiological environments for use in tissue engineering applications and studies of basic cell behavior. We discuss techniques to control fibril alignment, methods for cross-linking collagen constructs to modulate stiffness, and composite collagen constructs to better mimic physiological extracellular matrix. PMID:26689380

  8. Role of surface layer collagen binding protein from indigenous Lactobacillus plantarum 91 in adhesion and its anti-adhesion potential against gut pathogen.

    PubMed

    Yadav, Ashok Kumar; Tyagi, Ashish; Kaushik, Jai Kumar; Saklani, Asha Chandola; Grover, Sunita; Batish, Virender Kumar

    2013-12-14

    Human feacal isolates were ascertain as genus Lactobacillus using specific primer LbLMA1/R16-1 and further identified as Lactobacillus plantarum with species specific primers Lpl-3/Lpl-2. 25 L. plantarum strains were further assessed for hydrophobicity following the microbial adhesion to hydrocarbons (MATH) method and colonization potentials based on their adherence to immobilized human collagen type-1. Surface proteins were isolated from selected L. plantarum 91(Lp91) strain. The purified collagen binding protein (Cbp) protein was assessed for its anti-adhesion activity against enteric Escherichia coli 0157:H7 pathogen on immobilized collagen. Four L. plantarum strains displayed high degree of hydrophobicity and significant adhesion to collagen. A 72 kDa protein was purified which reduced 59.71% adhesion of E. coli 0157:H7 on immobilized collagen as compared to control well during adhesion assay. Cbp protein is the major influencing factor in inhibition of E. coli 0157:H7 adhesion with extracellular matrix (ECM) components. Hydrophobicity and adhesion potential are closely linked attributes precipitating in better colonization potential of the lactobacillus strains. Cbp is substantiated as a crucial surface protein contributing in adhesion of lactobacillus strains. The study can very well be the platform for commercialization of indigenous probiotic strain once their functional attributes are clinically explored. PMID:23890721

  9. Chondroitin sulfate cluster of epiphycan from salmon nasal cartilage defines binding specificity to collagens.

    PubMed

    Tatara, Yota; Kakizaki, Ikuko; Suto, Shinichiro; Ishioka, Haruna; Negishi, Mika; Endo, Masahiko

    2015-05-01

    Epiphycan (EPY) from salmon nasal cartilage has a glycosaminoglycan (GAG) domain that is heavily modified by chondroitin 4-sulfate and chondroitin 6-sulfate. The functional role of the GAG domain has not been investigated. The interaction of EPY with collagen was examined in vitro using surface plasmon resonance analysis. EPY was found to bind to type I collagen via clustered chondroitin sulfate (CS), while a single chain of CS was unable to bind. Types I, III, VII, VIII and X collagen showed high binding affinity with EPY, whereas types II, IV, V, VI and IX showed low binding affinities. Chemical modification of lysine residues in collagen decreased the affinity with the clustered CS. These results suggest that lysine residues of collagen are involved in the interaction with the clustered CS, and the difference in lysine modification defines the binding affinity to EPY. The clustered CS was also involved in an inter-saccharide interaction, and formed self-associated EPY. CS of EPY promoted fibril formation of type I collagen. PMID:25533443

  10. Cystamine immobilization on TiO 2 film surfaces and the influence on inhibition of collagen-induced platelet activation

    NASA Astrophysics Data System (ADS)

    Zhou, Yujuan; Weng, Yajun; Zhang, Liping; Jing, Fengjuan; Huang, Nan; Chen, Junying

    2011-12-01

    Poor haemocompatibility is a main issue of artificial cardiovascular materials in clinical application. Nitric oxide (NO), produced by vascular endothelial cells, is a well known inhibitor of platelet adhesion and activation. Thus, NO-releasing biomaterials are beneficial for improving haemocompatibility of blood-contacting biomedical devices. In this paper, a novel method was developed for enhancement of haemocompatibility by exploiting endogenous NO donors. TiO 2 films were firstly synthesized on Si (1 0 0) wafers via unbalanced magnetron sputtering technology, and then polydopamine was grafted on TiO 2 films and used as a linker for further immobilization of cystamine. The obtained surfaces were characterized by scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) analysis. NO generation is evaluated by saville-griess reagents, and it shows that cystamine immobilized samples are able to catalytically generate NO by decomposing endogenous S-nitrosothiols (RSNO). In vitro platelet adhesion results reveal that cystamine modified surfaces can inhibit collagen-induced platelet activation. ELISA analysis reveals that cGMP in platelets obviously increases on cystamine immobilized surface, which suggests the reducing of platelet activation is through NO/cGMP signal channel. It can be concluded that cystamine immobilized surface shows better blood compatibility by catalyzing NO release from the endogenous NO donor. It may be a promising method for improvement of haemocompatibility of blood-contacting implants.

  11. Collagen network strengthening following cyclic tensile loading.

    PubMed

    Susilo, Monica E; Paten, Jeffrey A; Sander, Edward A; Nguyen, Thao D; Ruberti, Jeffrey W

    2016-02-01

    The bulk mechanical properties of tissues are highly tuned to the physiological loads they experience and reflect the hierarchical structure and mechanical properties of their constituent parts. A thorough understanding of the processes involved in tissue adaptation is required to develop multi-scale computational models of tissue remodelling. While extracellular matrix (ECM) remodelling is partly due to the changing cellular metabolic activity, there may also be mechanically directed changes in ECM nano/microscale organization which lead to mechanical tuning. The thermal and enzymatic stability of collagen, which is the principal load-bearing biopolymer in vertebrates, have been shown to be enhanced by force suggesting that collagen has an active role in ECM mechanical properties. Here, we ask how changes in the mechanical properties of a collagen-based material are reflected by alterations in the micro/nanoscale collagen network following cyclic loading. Surprisingly, we observed significantly higher tensile stiffness and ultimate tensile strength, roughly analogous to the effect of work hardening, in the absence of network realignment and alterations to the fibril area fraction. The data suggest that mechanical loading induces stabilizing changes internal to the fibrils themselves or in the fibril-fibril interactions. If such a cell-independent strengthening effect is operational in vivo, then it would be an important consideration in any multiscale computational approach to ECM growth and remodelling. PMID:26855760

  12. Microstructural characterization of vocal folds toward a strain-energy model of collagen remodeling.

    PubMed

    Miri, Amir K; Heris, Hossein K; Tripathy, Umakanta; Wiseman, Paul W; Mongeau, Luc

    2013-08-01

    Collagen fibrils are believed to control the immediate deformation of soft tissues under mechanical load. Most extracellular matrix proteins remain intact during frozen sectioning, which allows them to be scanned using atomic force microscopy (AFM). Collagen fibrils are distinguishable because of their periodic roughness wavelength. In the present study, the shape and organization of collagen fibrils in dissected porcine vocal folds were quantified using nonlinear laser scanning microscopy data at the micrometer scale and AFM data at the nanometer scale. Rope-shaped collagen fibrils were observed. The geometric characteristics for the fibrils were fed into a hyperelastic model to predict the biomechanical response of the tissue. The model simulates the micrometer-scale unlocking behavior of collagen bundles when extended from their unloaded configuration. Force spectroscopy using AFM was used to estimate the stiffness of collagen fibrils (1±0.5MPa). The presence of rope-shaped fibrils is postulated to change the slope of the force-deflection response near the onset of nonlinearity. The proposed model could ultimately be used to evaluate changes in elasticity of soft tissues that result from the collagen remodeling. PMID:23643604

  13. Microstructural Characterization of Vocal Folds toward a Strain-Energy Model of Collagen Remodeling

    PubMed Central

    Miri, Amir K.; Heris, Hossein K.; Tripathy, Umakanta; Wiseman, Paul W.; Mongeau, Luc

    2013-01-01

    Collagen fibrils are believed to control the immediate deformation of soft tissues under biomechanical load. Most extracellular matrix proteins remain intact during frozen sectioning, which allows them to be scanned using atomic force microscopy (AFM). Collagen fibrils are distinguishable because of their helical shape. In the present study, the shape and organization of collagen fibrils in dissected porcine vocal folds were quantified using nonlinear laser scanning microscopy data at the micrometer scale and AFM data at the nanometer scale. Rope-shape collagen fibrils were observed. Geometric characteristics for the fibrils were fed to a hyperelastic model to predict the biomechanical response of the tissue. The model simulates the micrometer-scale unlocking behavior of collagen bundles when extended from their unloaded configuration. Force spectroscopy using AFM was used to estimate the stiffness of collagen fibrils (1 ± 0.5 MPa). The presence of rope-shape fibrils is postulated to change the slope of the force-deflection response near the onset of nonlinearity. The proposed model could ultimately be used to evaluate changes in elasticity of soft tissues that result from the collagen remodeling. PMID:23643604

  14. Investigating collagen self-assembly with optical tweezers microrheology

    NASA Astrophysics Data System (ADS)

    Forde, Nancy; Shayegan, Marjan; Altindal, Tuba

    Collagen is the fundamental structural protein in vertebrates. Assembled from individual triple-helical proteins to make strong fibres, collagen is a beautiful example of a hierarchical self-assembling system. Using optical tweezers to perform microrheology measurements, we explore the dynamics of interactions between collagens responsible for their self-assembly and examine the development of heterogeneous mechanics during assembly into fibrillar gels. Telopeptides, short non-helical regions that flank the triple helix, have long been known to facilitate fibril self-assembly. We find that their removal not only slows down fibril nucleation but also results in a significant frequency-dependent reduction in the elastic modulus of collagens in solution. We interpret these results in terms of a model in which telopeptides facilitate transient intermolecular interactions, which enhance network connectivity in solution and lead to more rapid assembly in fibril-forming conditions. Current address: Department of Physics, McGill University.

  15. Nonlinear microscopy of collagen fibers

    NASA Astrophysics Data System (ADS)

    Strupler, M.; Pena, A.-M.; Hernest, M.; Tharaux, P.-L.; Fabre, A.; Marchal-Somme, J.; Crestani, B.; Débarre, D.; Martin, J.-L.; Beaurepaire, E.; Schanne-Klein, M.-C.

    2007-02-01

    We used intrinsic Second Harmonic Generation (SHG) by fibrillar collagen to visualize the three-dimensional architecture of collagen fibrosis at the micrometer scale using laser scanning nonlinear microscopy. We showed that SHG signals are highly specific to fibrillar collagen and provide a sensitive probe of the micrometer-scale structural organization of collagen in tissues. Moreover, recording simultaneously other nonlinear optical signals in a multimodal setup, we visualized the tissue morphology using Two-Photon Excited Fluorescence (2PEF) signals from endogenous chromophores such as NADH or elastin. We then compared different methods to determine accurate indexes of collagen fibrosis using nonlinear microscopy, given that most collagen fibrils are smaller than the microscope resolution and that second harmonic generation is a coherent process. In order to define a robust method to process our three-dimensional images, we either calculated the fraction of the images occupied by a significant SHG signal, or averaged SHG signal intensities. We showed that these scores provide an estimation of the extension of renal and pulmonary fibrosis in murine models, and that they clearly sort out the fibrotic mice.

  16. Human collagen produced in plants

    PubMed Central

    Shoseyov, Oded; Posen, Yehudit; Grynspan, Frida

    2014-01-01

    Consequential to its essential role as a mechanical support and affinity regulator in extracellular matrices, collagen constitutes a highly sought after scaffolding material for regeneration and healing applications. However, substantiated concerns have been raised with regard to quality and safety of animal tissue-extracted collagen, particularly in relation to its immunogenicity, risk of disease transmission and overall quality and consistency. In parallel, contamination with undesirable cellular factors can significantly impair its bioactivity, vis-a-vis its impact on cell recruitment, proliferation and differentiation. High-scale production of recombinant human collagen Type I (rhCOL1) in the tobacco plant provides a source of an homogenic, heterotrimeric, thermally stable “virgin” collagen which self assembles to fine homogenous fibrils displaying intact binding sites and has been applied to form numerous functional scaffolds for tissue engineering and regenerative medicine. In addition, rhCOL1 can form liquid crystal structures, yielding a well-organized and mechanically strong membrane, two properties indispensable to extracellular matrix (ECM) mimicry. Overall, the shortcomings of animal- and cadaver-derived collagens arising from their source diversity and recycled nature are fully overcome in the plant setting, constituting a collagen source ideal for tissue engineering and regenerative medicine applications. PMID:23941988

  17. Chondrogenic differentiation of human mesenchymal stem cells on fish scale collagen.

    PubMed

    Hsu, Han-Hsiu; Uemura, Toshimasa; Yamaguchi, Isamu; Ikoma, Toshiyuki; Tanaka, Junzo

    2016-08-01

    Fish collagen has recently been reported to be a novel biomaterial for cell and tissue culture as an alternative to conventional mammalian collagens such as bovine and porcine collagens. Fish collagen could overcome the risk of zoonosis, such as from bovine spongiform encephalopathy. Among fish collagens, tilapia collagen, the denaturing temperature of which is near 37°C, is appropriate for cell and tissue culture. In this study, we investigated chondrogenic differentiation of human mesenchymal stem cells (hMSCs) cultured on tilapia scale collagen fibrils compared with porcine collagen and non-coated dishes. The collagen fibrils were observed using a scanning electronic microscope. Safranin O staining, glycosaminoglycans (GAG) expression, and real-time PCR were examined to evaluate chondrogenesis of hMSCs on each type of collagen fibril. The results showed that hMSCs cultured on tilapia scale collagen showed stronger Safranin O staining and higher GAG expression at day 6. Results of real-time PCR indicated that hMSCs cultured on tilapia collagen showed earlier SOX9 expression on day 4 and higher AGGRECAN and COLLAGEN II expression on day 6 compared with on porcine collagen and non-coated dishes. Furthermore, low mRNA levels of bone gamma-carboxyglutamate, a specific marker of osteogenesis, showed that tilapia collagen fibrils specifically enhanced chondrogenic differentiation of hMSCs in chondrogenic medium, as well as porcine collagen. Accordingly, tilapia scale collagen may provide an appropriate collagen source for hMSC chondrogenesis in vitro. PMID:26829997

  18. Atrial fibrillation

    PubMed Central

    Munger, Thomas M.; Wu, Li-Qun; Shen, Win K.

    2014-01-01

    Atrial fibrillation is the most common arrhythmia affecting patients today. Disease prevalence is increasing at an alarming rate worldwide, and is associated with often catastrophic and costly consequences, including heart failure, syncope, dementia, and stroke. Therapies including anticoagulants, anti-arrhythmic medications, devices, and non-pharmacologic procedures in the last 30 years have improved patients' functionality with the disease. Nonetheless, it remains imperative that further research into AF epidemiology, genetics, detection, and treatments continues to push forward rapidly as the worldwide population ages dramatically over the next 20 years. PMID:24474959

  19. Bioengineered collagens

    PubMed Central

    Ramshaw, John AM; Werkmeister, Jerome A; Dumsday, Geoff J

    2014-01-01

    Mammalian collagen has been widely used as a biomedical material. Nevertheless, there are still concerns about the variability between preparations, particularly with the possibility that the products may transmit animal-based diseases. Many groups have examined the possible application of bioengineered mammalian collagens. However, translating laboratory studies into large-scale manufacturing has often proved difficult, although certain yeast and plant systems seem effective. Production of full-length mammalian collagens, with the required secondary modification to give proline hydroxylation, has proved difficult in E. coli. However, recently, a new group of collagens, which have the characteristic triple helical structure of collagen, has been identified in bacteria. These proteins are stable without the need for hydroxyproline and are able to be produced and purified from E. coli in high yield. Initial studies indicate that they would be suitable for biomedical applications. PMID:24717980

  20. Viscoelastic properties of model segments of collagen molecules.

    PubMed

    Gautieri, Alfonso; Vesentini, Simone; Redaelli, Alberto; Buehler, Markus J

    2012-03-01

    Collagen is the prime construction material in vertebrate biology, determining the mechanical behavior of connective tissues such as tendon, bone and skin. Despite extensive efforts in the investigation of the origin of collagen unique mechanical properties, a deep understanding of the relationship between molecular structure and mechanical properties remains elusive, hindered by the complex hierarchical structure of collagen-based tissues. In particular, although extensive studies of viscoelastic properties have been pursued at the macroscopic (fiber/tissue) level, fewer investigations have been performed at the smaller scales, including in particular collagen molecules and fibrils. These scales are, however, important for a complete understanding of the role of collagen as an important constituent in the extracellular matrix. Here, using an atomistic modeling approach, we perform in silico creep tests of a collagen-like peptide, monitoring the strain-time response for different values of applied external load. The results show that individual collagen molecules exhibit a nonlinear viscoelastic behavior, with a Young's modulus increasing from 6 to 16GPa (for strains up to 20%), a viscosity of 3.84.±0.38Pa·s, and a relaxation time in the range of 0.24-0.64ns. The single molecule viscosity, for the first time reported here, is several orders of magnitude lower than the viscosity found for larger-scale single collagen fibrils, suggesting that the viscous behavior of collagen fibrils and fibers involves additional mechanisms, such as molecular sliding between collagen molecules within the fibril or the effect of relaxation of larger volumes of solvent. Based on our molecular modeling results we propose a simple structural model that describes collagen tissue as a hierarchical structure, providing a bottom-up description of elastic and viscous properties form the properties of the tissue basic building blocks. PMID:22204879

  1. Development of thrombus-resistant and cell compatible crimped polyethylene terephthalate cardiovascular grafts using surface co-immobilized heparin and collagen.

    PubMed

    Al Meslmani, Bassam; Mahmoud, Gihan; Strehlow, Boris; Mohr, Eva; Leichtweiß, Thomas; Bakowsky, Udo

    2014-10-01

    Short-term patency of polyethylene terephthalate (PET) cardiovascular grafts is determined mainly by the inherent thrombogenicity and improper endothelialization following grafts implantation. The aim of the present study was to immobilize heparin to develop thrombus resistant grafts. Additionally, collagen was co-immobilized to enhance the host cell compatibility. The synthetic woven and knitted forms of crimped PET grafts were surface modified by Denier reduction to produce functional carboxyl groups. The produced groups were used as anchor sites for covalent immobilization of heparin or co-immobilization of heparin/collagen by the end-point method. The modified surface was characterized using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. The biological activity of immobilized molecules was investigated in vitro using direct blood coagulation test, and "platelet deposition under flow condition. Furthermore, the biocompatibility of modified grafts with host cells was assessed using L929 cell as model. All modified grafts showed significant resistance against fibrin and clot formation. The number of deposited platelets on heparin-immobilized woven and knitted grafts obviously decreased by 3 fold and 2.8 fold per unit surface area respectively, while the heparin/collagen co-immobilized grafts showed only a decrease by 1.7 and 1.8 fold compared to unmodified PET. Heparin-immobilized grafts reported no significant effect on L929 cells adhesion and growth (P>0.05), conversely, collagen co-immobilization considerably increased cell adhesion almost ~1.3 fold and 2 fold per unit surface area for woven and knitted grafts respectively. Our results emphasize that immobilization of heparin minimized the inherent thrombogenicity of the PET grafts. The simultaneous co-immobilization of collagen supported host cell adhesion and growth required for the grafts biocompatibility. PMID:25175248

  2. Cell-surface serglycin promotes adhesion of myeloma cells to collagen type I and affects the expression of matrix metalloproteinases.

    PubMed

    Skliris, Antonis; Labropoulou, Vassiliki T; Papachristou, Dionysios J; Aletras, Alexios; Karamanos, Nikos K; Theocharis, Achilleas D

    2013-05-01

    Serglycin (SG) is mainly expressed by hematopoetic cells as an intracellular proteoglycan. Multiple myeloma cells constitutively secrete SG, which is also localized on the cell surface in some cell lines. In this study, SG isolated from myeloma cells was found to interact with collagen type I (Col I), which is a major bone matrix component. Notably, myeloma cells positive for cell-surface SG (csSG) adhered significantly to Col I, compared to cells lacking csSG. Removal of csSG by treatment of the cells with chondroitinase ABC or blocking of csSG by an SG-specific polyclonal antibody significantly reduced the adhesion of myeloma cells to Col I. Significant up-regulation of expression of the matrix metalloproteinases MMP-2 and MMP-9 at both the mRNA and protein levels was observed when culturing csSG-positive myeloma cells on Col I-coated dishes or in the presence of soluble Col I. MMP-9 and MMP-2 were also expressed in increased amounts by myeloma cells in the bone marrow of patients with multiple myeloma. Our data indicate that csSG of myeloma cells affects key functional properties, such as adhesion to Col I and the expression of MMPs, and imply that csSG may serve as a potential prognostic factor and/or target for pharmacological interventions in multiple myeloma. PMID:23387827

  3. Collagenous microstructure of the glenoid labrum and biceps anchor

    PubMed Central

    Hill, A M; Hoerning, E J; Brook, K; Smith, C D; Moss, J; Ryder, T; Wallace, A L; Bull, A M J

    2008-01-01

    The glenoid labrum is a significant passive stabilizer of the shoulder joint. However, its microstructural form remains largely unappreciated, particularly in the context of its variety of functions. The focus of labral microscopy has often been histology and, as such, there is very little appreciation of collagen composition and arrangement of the labrum, and hence the micromechanics of the structure. On transmission electron microscopy, significant differences in diameter, area and perimeter were noted in the two gross histological groups of collagen fibril visualized; this suggests a heterogeneous collagenous composition with potentially distinct mechanical function. Scanning electron microscopy demonstrated three distinct zones of interest: a superficial mesh, a dense circumferential braided core potentially able to accommodate hoop stresses, and a loosely packed peri-core zone. Confocal microscopy revealed an articular surface fine fibrillar mesh potentially able to reduce surface friction, bundles of circumferential encapsulated fibres in the bulk of the tissue, and bone anchoring fibres at the osseous interface. Varying microstructure throughout the depth of the labrum suggests a role in accommodating different types of loading. An understanding of the labral microstructure can lead to development of hypotheses based upon an appreciation of this component of material property. This may aid an educated approach to surgical timing and repair. PMID:18429974

  4. Lysyl Oxidase Activity Is Required for Ordered Collagen Fibrillogenesis by Tendon Cells*

    PubMed Central

    Herchenhan, Andreas; Uhlenbrock, Franziska; Eliasson, Pernilla; Weis, MaryAnn; Eyre, David; Kadler, Karl E.; Magnusson, S. Peter; Kjaer, Michael

    2015-01-01

    Lysyl oxidases (LOXs) are a family of copper-dependent oxido-deaminases that can modify the side chain of lysyl residues in collagen and elastin, thereby leading to the spontaneous formation of non-reducible aldehyde-derived interpolypeptide chain cross-links. The consequences of LOX inhibition in producing lathyrism are well documented, but the consequences on collagen fibril formation are less clear. Here we used β-aminoproprionitrile (BAPN) to inhibit LOX in tendon-like constructs (prepared from human tenocytes), which are an experimental model of cell-mediated collagen fibril formation. The improvement in structure and strength seen with time in control constructs was absent in constructs treated with BAPN. As expected, BAPN inhibited the formation of aldimine-derived cross-links in collagen, and the constructs were mechanically weak. However, an unexpected finding was that BAPN treatment led to structurally abnormal collagen fibrils with irregular profiles and widely dispersed diameters. Of special interest, the abnormal fibril profiles resembled those seen in some Ehlers-Danlos Syndrome phenotypes. Importantly, the total collagen content developed normally, and there was no difference in COL1A1 gene expression. Collagen type V, decorin, fibromodulin, and tenascin-X proteins were unaffected by the cross-link inhibition, suggesting that LOX regulates fibrillogenesis independently of these molecules. Collectively, the data show the importance of LOX for the mechanical development of early collagenous tissues and that LOX is essential for correct collagen fibril shape formation. PMID:25979340

  5. Lysyl Oxidase Activity Is Required for Ordered Collagen Fibrillogenesis by Tendon Cells.

    PubMed

    Herchenhan, Andreas; Uhlenbrock, Franziska; Eliasson, Pernilla; Weis, MaryAnn; Eyre, David; Kadler, Karl E; Magnusson, S Peter; Kjaer, Michael

    2015-06-26

    Lysyl oxidases (LOXs) are a family of copper-dependent oxido-deaminases that can modify the side chain of lysyl residues in collagen and elastin, thereby leading to the spontaneous formation of non-reducible aldehyde-derived interpolypeptide chain cross-links. The consequences of LOX inhibition in producing lathyrism are well documented, but the consequences on collagen fibril formation are less clear. Here we used β-aminoproprionitrile (BAPN) to inhibit LOX in tendon-like constructs (prepared from human tenocytes), which are an experimental model of cell-mediated collagen fibril formation. The improvement in structure and strength seen with time in control constructs was absent in constructs treated with BAPN. As expected, BAPN inhibited the formation of aldimine-derived cross-links in collagen, and the constructs were mechanically weak. However, an unexpected finding was that BAPN treatment led to structurally abnormal collagen fibrils with irregular profiles and widely dispersed diameters. Of special interest, the abnormal fibril profiles resembled those seen in some Ehlers-Danlos Syndrome phenotypes. Importantly, the total collagen content developed normally, and there was no difference in COL1A1 gene expression. Collagen type V, decorin, fibromodulin, and tenascin-X proteins were unaffected by the cross-link inhibition, suggesting that LOX regulates fibrillogenesis independently of these molecules. Collectively, the data show the importance of LOX for the mechanical development of early collagenous tissues and that LOX is essential for correct collagen fibril shape formation. PMID:25979340

  6. Atrial fibrillation.

    PubMed

    Lip, Gregory Y H; Fauchier, Laurent; Freedman, Saul B; Van Gelder, Isabelle; Natale, Andrea; Gianni, Carola; Nattel, Stanley; Potpara, Tatjana; Rienstra, Michiel; Tse, Hung-Fat; Lane, Deirdre A

    2016-01-01

    Atrial fibrillation (AF) is the most common sustained cardiac rhythm disorder, and increases in prevalence with increasing age and the number of cardiovascular comorbidities. AF is characterized by a rapid and irregular heartbeat that can be asymptomatic or lead to symptoms such as palpitations, dyspnoea and dizziness. The condition can also be associated with serious complications, including an increased risk of stroke. Important recent developments in the clinical epidemiology and management of AF have informed our approach to this arrhythmia. This Primer provides a comprehensive overview of AF, including its epidemiology, mechanisms and pathophysiology, diagnosis, screening, prevention and management. Management strategies, including stroke prevention, rate control and rhythm control, are considered. We also address quality of life issues and provide an outlook on future developments and ongoing clinical trials in managing this common arrhythmia. PMID:27159789

  7. Amyloid fibrils

    PubMed Central

    Rambaran, Roma N

    2008-01-01

    Amyloid refers to the abnormal fibrous, extracellular, proteinaceous deposits found in organs and tissues. Amyloid is insoluble and is structurally dominated by β-sheet structure. Unlike other fibrous proteins it does not commonly have a structural, supportive or motility role but is associated with the pathology seen in a range of diseases known as the amyloidoses. These diseases include Alzheimer's, the spongiform encephalopathies and type II diabetes, all of which are progressive disorders with associated high morbidity and mortality. Not surprisingly, research into the physicochemical properties of amyloid and its formation is currently intensely pursued. In this chapter we will highlight the key scientific findings and discuss how the stability of amyloid fibrils impacts on bionanotechnology. PMID:19158505

  8. Generation of 3D Collagen Gels with Controlled Diverse Architectures.

    PubMed

    Doyle, Andrew D

    2016-01-01

    Rat tail collagen solutions have been used as polymerizable in vitro three dimensional (3D) extracellular matrix (ECM) gels for single and collective cell migration assays as well as spheroid formation. Factors such as ECM concentration, pH, ionic concentration, and temperature can alter collagen polymerization and ECM architecture. This unit describes how to generate 3D collagen gels that have distinct architectures ranging from a highly reticular meshwork of short thin fibrils with small pores to a loose matrix consisting of stiff, parallel-bundled long fibrils by changing collagen polymerization temperature. This permits analysis of 3D cell migration in different ECM architectures found in vivo while maintaining a similar ECM concentration. Also included are collagen labeling techniques helpful for ECM visualization during live fluorescence imaging. © 2016 by John Wiley & Sons, Inc. PMID:27580704

  9. Biosynthesis of collagen and other matrix proteins by articular cartilage in experimental osteoarthrosis.

    PubMed Central

    Eyre, D R; McDevitt, C A; Billingham, M E; Muir, H

    1980-01-01

    Osteoarthrosis was induced in one knee joint of dogs by an established surgical procedure. Changes in the articular cartilage in the biosynthesis of collagen and other proteins were sought by radiochemical labelling in vivo, with the following findings. (1) Collagen synthesis was stimulated in all cartilage surfaces of the experimental joints at 2, 8 and 24 weeks after surgery. Systemic labelling with [3H]proline showed that over 10 times more collagen was being deposited per dry weight of experimental cartilage compared with control cartilage in the unoperated knee. (2) Type-II collagen was the radiolabelled product in all samples of experimental cartilage ranging in quality from undamaged to overtly fibrillated, and was the only collagen detected chemically in the matrix of osteoarthrotic cartilage from either dog or human joints. (3) Hydroxylysine glycosylation was examined in the newly synthesized cartilage collagen by labelling dog joints in vivo with [3H]lysine. In experimental knees the new collagen was less glycosylated than in controls. However, no difference in glycosylation of the total collagen in the tissues was observed by chemical analysis. (4) Over half the protein-bound tritium was extracted by 4 M-guanidinium chloride from control cartilage labelled with [3H]proline, compared with one-quarter or less from experimental cartilage. Two-thirds of the extracted tritium separated in the upper fraction on density-gradient centrifugation in CsCl under associative conditions. Much of this ran with a single protein band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under reducing conditions. The identity of this protein was unknown, although it resembled serum albumin in mobility afte disulphide-bond cleavage. Images Fig. 3. PMID:7470037

  10. The size exclusion characteristics of type I collagen: implications for the role of noncollagenous bone constituents in mineralization.

    PubMed

    Toroian, Damon; Lim, Joo Eun; Price, Paul A

    2007-08-01

    The mineral in bone is located primarily within the collagen fibril, and during mineralization the fibril is formed first and then water within the fibril is replaced with mineral. The collagen fibril therefore provides the aqueous compartment in which mineral grows. Although knowledge of the size of molecules that can diffuse into the fibril to affect crystal growth is critical to understanding the mechanism of bone mineralization, there have been as yet no studies on the size exclusion properties of the collagen fibril. To determine the size exclusion characteristics of collagen, we developed a gel filtration-like procedure that uses columns containing collagen from tendon and bone. The elution volumes of test molecules show the volume within the packed column that is accessible to the test molecules, and therefore reveal the size exclusion characteristics of the collagen within the column. These experiments show that molecules smaller than a 6-kDa protein diffuse into all of the water within the collagen fibril, whereas molecules larger than a 40-kDa protein are excluded from this water. These studies provide an insight into the mechanism of bone mineralization. Molecules and apatite crystals smaller than a 6-kDa protein can diffuse into all water within the fibril and so can directly impact mineralization. Although molecules larger than a 40-kDa protein are excluded from the fibril, they can initiate mineralization by forming small apatite crystal nuclei that diffuse into the fibril, or can favor fibril mineralization by inhibiting apatite growth everywhere but within the fibril. PMID:17562713

  11. Fast and mild strategy, using superhydrophobic surfaces, to produce collagen/platelet lysate gel beads for skin regeneration.

    PubMed

    Lima, Ana Catarina; Mano, João F; Concheiro, Angel; Alvarez-Lorenzo, Carmen

    2015-02-01

    Platelet lysate (PL) was encapsulated in collagen (Coll) millimetric gel beads, on biomimetic superhydrophobic surfaces, under mild conditions, with the aim of obtaining easy-to-handle formulations able to provide sustained release of multiple growth factors for skin ulcers treatment. The gel particles were prepared with various concentrations of PL incorporating or not stem cells, and tested as freshly prepared or after being freeze-dried or cryopreserved. Coll + PL particles were evaluated regarding degradation in collagenase-rich environment (simulating the aggressive environment of the chronic ulcers), sustained release of total protein, PDGF-BB and VEGF, cell proliferation (using particles as the only source of growth factors), scratch wound recovery and angiogenic capability. Compared to Coll solely particles, incorporation of PL notably enhanced cell proliferation (inside and outside gels) and favored scratch wound recovery and angiogenesis. Moreover, cell-laden gel particles containing PL notably improved cell proliferation and even migration of cells from one particle towards a neighbor one, which led to cell-cell contacts and the spontaneous formation of tissue layers in which the spherical gels were interconnected by the stem cells. PMID:25120225

  12. Changes in Scleral Collagen Organization in Murine Chronic Experimental Glaucoma

    PubMed Central

    Pijanka, Jacek K.; Kimball, Elizabeth C.; Pease, Mary E.; Abass, Ahmed; Sorensen, Thomas; Nguyen, Thao D.; Quigley, Harry A.; Boote, Craig

    2014-01-01

    Purpose. The organization of scleral collagen helps to determine the eye's biomechanical response to intraocular pressure (IOP), and may therefore be important in glaucoma. This study provides a quantitative assessment of changes in scleral collagen fibril organization in bead-induced murine experimental glaucoma. Methods. Wide-angle X-ray scattering was used to study the effect of bead-induced glaucoma on posterior scleral collagen organization in one eye of 12 CD1 mice, with untreated fellow eyes serving as controls. Three collagen parameters were measured: the local preferred fibril directions, the degree of collagen anisotropy, and the total fibrillar collagen content. Results. The mouse sclera featured a largely circumferential orientation of fibrillar collagen with respect to the optic nerve head canal. Localized alteration to fibril orientations was evident in the inferior peripapillary sclera of bead-treated eyes. Collagen anisotropy was significantly (P < 0.05) reduced in bead-treated eyes in the superior peripapillary (Treated: 43 ± 8%; Control: 49 ± 6%) and midposterior (Treated: 39 ± 4%; Control: 43 ± 4%) sclera, and in the peripapillary region overall (Treated: 43 ± 6%; Control: 47 ± 3%). No significant differences in total collagen content were found between groups. Conclusions. Spatial changes in collagen fibril anisotropy occur in the posterior sclera of mice with bead-induced chronic IOP elevation and axonal damage. These results support the idea that dynamic changes in scleral form and structure play a role in the development of experimental glaucoma in mice, and potentially in human glaucoma. PMID:25228540

  13. Collagen in Human Tissues: Structure, Function, and Biomedical Implications from a Tissue Engineering Perspective

    NASA Astrophysics Data System (ADS)

    Balasubramanian, Preethi; Prabhakaran, Molamma P.; Sireesha, Merum; Ramakrishna, Seeram

    The extracellular matrix is a complex biological structure encoded with various proteins, among which the collagen family is the most significant and abundant of all, contributing 30-35% of the whole-body protein. "Collagen" is a generic term for proteins that forms a triple-helical structure with three polypeptide chains, and around 29 types of collagen have been identified up to now. Although most of the members of the collagen family form such supramolecular structures, extensive diversity exists between each type of collagen. The diversity is not only based on the molecular assembly and supramolecular structures of collagen types but is also observed within its tissue distribution, function, and pathology. Collagens possess complex hierarchical structures and are present in various forms such as collagen fibrils (1.5-3.5 nm wide), collagen fibers (50-70 nm wide), and collagen bundles (150-250 nm wide), with distinct properties characteristic of each tissue providing elasticity to skin, softness of the cartilage, stiffness of the bone and tendon, transparency of the cornea, opaqueness of the sclera, etc. There exists an exclusive relation between the structural features of collagen in human tissues (such as the collagen composition, collagen fibril length and diameter, collagen distribution, and collagen fiber orientation) and its tissue-specific mechanical properties. In bone, a transverse collagen fiber orientation prevails in regions of higher compressive stress whereas longitudinally oriented collagen fibers correlate to higher tensile stress. The immense versatility of collagen compels a thorough understanding of the collagen types and this review discusses the major types of collagen found in different human tissues, highlighting their tissue-specific uniqueness based on their structure and mechanical function. The changes in collagen during a specific tissue damage or injury are discussed further, focusing on the many tissue engineering applications for

  14. Abnormal arrangement of a collagen/apatite extracellular matrix orthogonal to osteoblast alignment is constructed by a nanoscale periodic surface structure.

    PubMed

    Matsugaki, Aira; Aramoto, Gento; Ninomiya, Takafumi; Sawada, Hiroshi; Hata, Satoshi; Nakano, Takayoshi

    2015-01-01

    Morphological and directional alteration of cells is essential for structurally appropriate construction of tissues and organs. In particular, osteoblast alignment is crucial for the realization of anisotropic bone tissue microstructure. In this article, the orientation of a collagen/apatite extracellular matrix (ECM) was established by controlling osteoblast alignment using a surface geometry with nanometer-sized periodicity induced by laser ablation. Laser irradiation induced self-organized periodic structures (laser-induced periodic surface structures; LIPSS) with a spatial period equal to the wavelength of the incident laser on the surface of biomedical alloys of Ti-6Al-4V and Co-Cr-Mo. Osteoblast orientation was successfully induced parallel to the grating structure. Notably, both the fibrous orientation of the secreted collagen matrix and the c-axis of the produced apatite crystals were orientated orthogonal to the cell direction. To the best of our knowledge, this is the first report demonstrating that bone tissue anisotropy is controllable, including the characteristic organization of a collagen/apatite composite orthogonal to the osteoblast orientation, by controlling the cell alignment using periodic surface geometry. PMID:25453944

  15. Signal Transducer and Activator of Transcription 3/MicroRNA-21 Feedback Loop Contributes to Atrial Fibrillation by Promoting Atrial Fibrosis in a Rat Sterile Pericarditis Model

    PubMed Central

    Huang, Zhengrong; Chen, Xiao-jun; Qian, Cheng; Dong, Qian; Ding, Dan; Wu, Qiong-feng; Li, Jing; Wang, Hong-fei; Li, Wei-hua; Xie, Qiang; Cheng, Xiang; Liao, Yu-hua

    2016-01-01

    Background— Postoperative atrial fibrillation is a frequent complication in cardiac surgery. The aberrant activation of signal transducer and activator of transcription 3 (STAT3) contributes to the pathogenesis of atrial fibrillation. MicroRNA-21 (miR-21) promotes atrial fibrosis. Recent studies support the existence of reciprocal regulation between STAT3 and miR-21. Here, we test the hypothesis that these 2 molecules might form a feedback loop that contributes to postoperative atrial fibrillation by promoting atrial fibrosis. Methods and Results— A sterile pericarditis model was created using atrial surfaces dusted with sterile talcum powder in rats. The inflammatory cytokines interleukin (IL)-1β, IL-6, transforming growth factor-β, and tumor necrosis factor-α, along with STAT3 and miR-21, were highly upregulated in sterile pericarditis rats. The inhibition of STAT3 by S3I-201 resulted in miR-21 downregulation, which ameliorated atrial fibrosis and decreased the expression of the fibrosis-related genes, α-smooth muscle actin, collagen-1, and collagen-3; reduced the inhomogeneity of atrial conduction; and attenuated atrial fibrillation vulnerability. Meanwhile, treatment with antagomir-21 decreased STAT3 phosphorylation, alleviated atrial remodeling, abrogated sterile pericarditis–induced inhomogeneous conduction, and prevented atrial fibrillation promotion. The culturing of cardiac fibroblasts with IL-6 resulted in progressively augmented STAT3 phosphorylation and miR-21 levels. S3I-201 blocked IL-6 induced the expression of miR-21 and fibrosis-related genes in addition to cardiac fibroblast proliferation. Transfected antagomir-21 decreased the IL-6–induced cardiac fibroblast activation and STAT3 phosphorylation. The overexpression of miR-21 in cardiac fibroblasts caused the upregulation of STAT3 phosphorylation, enhanced fibrosis-related genes, and increased cell numbers. Conclusions— Our results have uncovered a novel reciprocal loop between STAT3

  16. FXIa and platelet polyphosphate as therapeutic targets during human blood clotting on collagen/tissue factor surfaces under flow.

    PubMed

    Zhu, Shu; Travers, Richard J; Morrissey, James H; Diamond, Scott L

    2015-09-17

    Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas platelet-derived polyphosphate (polyP) may potentiate contact or thrombin-feedback pathways. The significance of these mediators in human blood under thrombotic flow conditions on tissue factor (TF) -bearing surfaces remains inadequately resolved. Human blood (corn trypsin inhibitor treated [4 μg/mL]) was tested by microfluidic assay for clotting on collagen/TF at TF surface concentration ([TF]wall) from ∼0.1 to 2 molecules per μm(2). Anti-FXI antibodies (14E11 and O1A6) or polyP-binding protein (PPXbd) were used to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-derived polyP, respectively. Fibrin formation was sensitive to 14E11 at 0 to 0.1 molecules per µm(2) and sensitive to O1A6 at 0 to 0.2 molecules per µm(2). However, neither antibody reduced fibrin generation at ∼2 molecules per µm(2) when the extrinsic pathway became dominant. Interestingly, PPXbd reduced fibrin generation at low [TF]wall (0.1 molecules per µm(2)) but not at zero or high [TF]wall, suggesting a role for polyP distinct from FXIIa activation and requiring low extrinsic pathway participation. Regardless of [TF]wall, PPXbd enhanced fibrin sensitivity to tissue plasminogen activator and promoted clot retraction during fibrinolysis concomitant with an observed PPXbd-mediated reduction of fibrin fiber diameter. This is the first detection of endogenous polyP function in human blood under thrombotic flow conditions. When triggered by low [TF]wall, thrombosis may be druggable by contact pathway inhibition, although thrombolytic susceptibility may benefit from polyP antagonism regardless of [TF]wall. PMID:26136249

  17. Quantitative Assessment of Local Collagen Matrix Remodeling in 3-D Culture: The Role of Rho Kinase

    PubMed Central

    Kim, Areum; Lakshman, Neema; Petroll, W.Matthew

    2007-01-01

    The purpose of this study was to quantitatively assess the role of Rho kinase in modulating the pattern and amount of local cell-induced collagen matrix remodeling. Human corneal fibroblasts were plated inside 100 μm thick fibrillar collagen matrices and cultured for 24 hours in media with or without the Rho kinase inhibitor Y-27632. Cells were then fixed and stained with phalloidin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) 3-D optical section images were acquired using laser confocal microscopy. Fourier transform analysis was used to assess collagen fibril alignment, and 3-D cell morphology and local collagen density were measured using MetaMorph. Culture in serum-containing media induced significant global matrix contraction, which was inhibited by blocking Rho kinase (p < 0.001). Fibroblasts generally had a bipolar morphology and intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. When Rho kinase was inhibited, cells had a more cortical f-actin distribution and dendritic morphology. Both local collagen fibril density and alignment were significantly reduced (p<0.01). Overall, the data suggests that Rho kinase dependent contractile force generation leads to co-alignment of cells and collagen fibrils along the plane of greatest resistance, and that this process contributes to global matrix contraction. PMID:16978606

  18. [Atrial fibrillation].

    PubMed

    Colín Lizalde, L J

    2001-01-01

    Atrial fibrillation (AF) is the most common sustained arrhythmia. AF has now been exhaustively studied: more is known about its mechanism and research is moving towards new forms of treatment. For chronic AF, basically the control of ventricular rate and the brain protection are the main issues. It is well known that with the identification of high risk group for embolism, oral anticoagulation should be administered. Ventricular rate control can be achieved by using betablockers or calcium channel blockers, unless these are contraindicated for the elderly. Oral anticoagulation prevents the stroke. The main mechanism of AF is the re-entry of multiple wavelets, but now it is more frequently found on patients with focal AF. Therapies are employed to bring the patient to a sinusal rhythm as soon as possible with antiarryhthmics or electric cardioversion externally or internally. The internal procedure includes 1 to 15 J and the success rate is of 91% vs 67% in relation to the external one. The introduction of the catheter ablation has opened new frontiers for the treatment of AF, first as the ablate-and-pace technique and now trying to mimic the maze procedure or with the ablation of the focal tachycardia. The stimulation for prevention of AF under research, as well as the implantable dysfibrillation for selected patients. On going studies will show the possible benefit of this type of benefits. PMID:11565343

  19. Real-time two- and three-dimensional imaging of monocyte motility and navigation on planar surfaces and in collagen matrices: roles of Rho

    PubMed Central

    Bzymek, Robert; Horsthemke, Markus; Isfort, Katrin; Mohr, Simon; Tjaden, Kerstin; Müller-Tidow, Carsten; Thomann, Marlies; Schwerdtle, Tanja; Bähler, Martin; Schwab, Albrecht; Hanley, Peter J.

    2016-01-01

    We recently found that macrophages from RhoA/RhoB double knockout mice had increased motility of the cell body, but severely impaired retraction of the tail and membrane extensions, whereas RhoA- or RhoB-deficient cells exhibited mild phenotypes. Here we extended this work and investigated the roles of Rho signaling in primary human blood monocytes migrating in chemotactic gradients and in various settings. Monocyte velocity, but not chemotactic navigation, was modestly dependent on Rho-ROCK-myosin II signaling on a 2D substrate or in a loose collagen type I matrix. Viewed by time-lapse epi-fluorescence microscopy, monocytes appeared to flutter rather than crawl, such that the 3D surface topology of individual cells was difficult to predict. Spinning disk confocal microscopy and 3D reconstruction revealed that cells move on planar surfaces and in a loose collagen matrix using prominent, curved planar protrusions, which are rapidly remodeled and reoriented, as well as resorbed. In a dense collagen type I matrix, there is insufficient space for this mode and cells adopt a highly Rho-dependent, lobular mode of motility. Thus, in addition to its role in tail retraction on 2D surfaces, Rho is critical for movement in confined spaces, but is largely redundant for motility and chemotaxis in loose matrices. PMID:27122054

  20. Real-time two- and three-dimensional imaging of monocyte motility and navigation on planar surfaces and in collagen matrices: roles of Rho.

    PubMed

    Bzymek, Robert; Horsthemke, Markus; Isfort, Katrin; Mohr, Simon; Tjaden, Kerstin; Müller-Tidow, Carsten; Thomann, Marlies; Schwerdtle, Tanja; Bähler, Martin; Schwab, Albrecht; Hanley, Peter J

    2016-01-01

    We recently found that macrophages from RhoA/RhoB double knockout mice had increased motility of the cell body, but severely impaired retraction of the tail and membrane extensions, whereas RhoA- or RhoB-deficient cells exhibited mild phenotypes. Here we extended this work and investigated the roles of Rho signaling in primary human blood monocytes migrating in chemotactic gradients and in various settings. Monocyte velocity, but not chemotactic navigation, was modestly dependent on Rho-ROCK-myosin II signaling on a 2D substrate or in a loose collagen type I matrix. Viewed by time-lapse epi-fluorescence microscopy, monocytes appeared to flutter rather than crawl, such that the 3D surface topology of individual cells was difficult to predict. Spinning disk confocal microscopy and 3D reconstruction revealed that cells move on planar surfaces and in a loose collagen matrix using prominent, curved planar protrusions, which are rapidly remodeled and reoriented, as well as resorbed. In a dense collagen type I matrix, there is insufficient space for this mode and cells adopt a highly Rho-dependent, lobular mode of motility. Thus, in addition to its role in tail retraction on 2D surfaces, Rho is critical for movement in confined spaces, but is largely redundant for motility and chemotaxis in loose matrices. PMID:27122054

  1. Atrial Fibrillation Medications

    MedlinePlus

    ... think you are pregnant If you notice red, dark brown or black urine or stools If you ... Fibrillation • Introduction • What is Atrial Fibrillation? • Why AFib Matters • Understand your Risk for AFib Children • Symptoms of ...

  2. Amyloid Fibril Solubility.

    PubMed

    Rizzi, L G; Auer, S

    2015-11-19

    It is well established that amyloid fibril solubility is protein specific, but how solubility depends on the interactions between the fibril building blocks is not clear. Here we use a simple protein model and perform Monte Carlo simulations to directly measure the solubility of amyloid fibrils as a function of the interaction between the fibril building blocks. Our simulations confirms that the fibril solubility depends on the fibril thickness and that the relationship between the interactions and the solubility can be described by a simple analytical formula. The results presented in this study reveal general rules how side-chain-side-chain interactions, backbone hydrogen bonding, and temperature affect amyloid fibril solubility, which might prove to be a powerful tool to design protein fibrils with desired solubility and aggregation properties in general. PMID:26496385

  3. Collagen Gel Contraction by Fibroblasts: The Role of Myosin 2 and Gravity Effects

    NASA Technical Reports Server (NTRS)

    Johnson-Wint, Barbara P.; Malouvier, Alexandre; Holton, Emily

    1996-01-01

    Several lines of evidence suggest that collagen organization by connective tissue cells is sensitive to force. For instance, in flight experiments on rats the collagen fibrils which were produced under weightlessness and which were immediately next to the tendon fibroblasts were shown to be oriented randomly around the cells while the older fibrils right next to these and which were produced under 1 G, were highly organized.

  4. Atrial Fibrillation in Children

    MedlinePlus

    ... Pressure High Blood Pressure Tools & Resources Stroke More Atrial Fibrillation in Children Updated:Jul 18,2016 Does your ... content was last reviewed on 04/16/14. Atrial Fibrillation • Introduction • What is Atrial Fibrillation? • Why AFib Matters • ...

  5. Fibromodulin Interacts with Collagen Cross-linking Sites and Activates Lysyl Oxidase*

    PubMed Central

    Bihan, Dominique; Bonna, Arkadiusz; Rubin, Kristofer; Farndale, Richard W.

    2016-01-01

    The hallmark of fibrotic disorders is a highly cross-linked and dense collagen matrix, a property driven by the oxidative action of lysyl oxidase. Other fibrosis-associated proteins also contribute to the final collagen matrix properties, one of which is fibromodulin. Its interactions with collagen affect collagen cross-linking, packing, and fibril diameter. We investigated the possibility that a specific relationship exists between fibromodulin and lysyl oxidase, potentially imparting a specific collagen matrix phenotype. We mapped the fibromodulin-collagen interaction sites using the collagen II and III Toolkit peptide libraries. Fibromodulin interacted with the peptides containing the known collagen cross-linking sites and the MMP-1 cleavage site in collagens I and II. Interestingly, the interaction sites are closely aligned within the quarter-staggered collagen fibril, suggesting a multivalent interaction between fibromodulin and several collagen helices. Furthermore, we detected an interaction between fibromodulin and lysyl oxidase (a major collagen cross-linking enzyme) and mapped the interaction site to 12 N-terminal amino acids on fibromodulin. This interaction also increases the activity of lysyl oxidase. Together, the data suggest a fibromodulin-modulated collagen cross-linking mechanism where fibromodulin binds to a specific part of the collagen domain and also forms a complex with lysyl oxidase, targeting the enzyme toward specific cross-linking sites. PMID:26893379

  6. Collagen-binding Microbial Surface Components Recognizing Adhesive Matrix Molecule (MSCRAMM) of Gram-positive Bacteria Inhibit Complement Activation via the Classical Pathway*

    PubMed Central

    Kang, Mingsong; Ko, Ya-Ping; Liang, Xiaowen; Ross, Caná L.; Liu, Qing; Murray, Barbara E.; Höök, Magnus

    2013-01-01

    Members of a family of collagen-binding microbial surface components recognizing adhesive matrix molecules (MSCRAMMs) from Gram-positive bacteria are established virulence factors in several infectious diseases models. Here, we report that these adhesins also can bind C1q and act as inhibitors of the classical complement pathway. Molecular analyses of Cna from Staphylococcus aureus suggested that this prototype MSCRAMM bound to the collagenous domain of C1q and interfered with the interactions of C1r with C1q. As a result, C1r2C1s2 was displaced from C1q, and the C1 complex was deactivated. This novel function of the Cna-like MSCRAMMs represents a potential immune evasion strategy that could be used by numerous Gram-positive pathogens. PMID:23720782

  7. Parathyroid hormone attenuates radiation-induced increases in collagen crosslink ratio at periosteal surfaces of mouse tibia.

    PubMed

    Oest, Megan E; Gong, Bo; Esmonde-White, Karen; Mann, Kenneth A; Zimmerman, Nicholas D; Damron, Timothy A; Morris, Michael D

    2016-05-01

    As part of our ongoing efforts to understand underlying mechanisms contributing to radiation-associated bone fragility and to identify possible treatments, we evaluated the longitudinal effects of parathyroid hormone (PTH) treatment on bone quality in a murine model of limited field irradiation. We hypothesized PTH would mitigate radiation-induced changes in the chemical composition and structure of bone, as measured by microscope-based Raman spectroscopy. We further hypothesized that collagen crosslinking would be especially responsive to PTH treatment. Raman spectroscopy was performed on retrieved tibiae (6-7/group/time point) to quantify metrics associated with bone quality, including: mineral-to-matrix ratio, carbonate-to-phosphate ratio, mineral crystallinity, collagen crosslink (trivalent:divalent) ratio, and the mineral and matrix depolarization ratios. Irradiation disrupted the molecular structure and orientation of bone collagen, as evidenced by a higher collagen crosslink ratio and lower matrix depolarization ratio (vs. non-irradiated control bones), persisting until 12weeks post-irradiation. Radiation transiently affected the mineral phase, as evidenced by increased mineral crystallinity and mineral-to-matrix ratio at 4weeks compared to controls. Radiation decreased bone mineral depolarization ratios through 12weeks, indicating increased mineral alignment. PTH treatment partially attenuated radiation-induced increases in collagen crosslink ratio, but did not restore collagen or mineral alignment. These post-radiation matrix changes are consistent with our previous studies of radiation damage to bone, and suggest that the initial radiation damage to bone matrix has extensive effects on the quality of tissue deposited thereafter. In addition to maintaining bone quality, preventing initial radiation damage to the bone matrix (i.e. crosslink ratio, matrix orientation) may be critical to preventing late-onset fragility fractures. PMID:26960578

  8. Mechanisms of disruption of the articular cartilage surface in inflammation. Neutrophil elastase increases availability of collagen type II epitopes for binding with antibody on the surface of articular cartilage.

    PubMed Central

    Jasin, H E; Taurog, J D

    1991-01-01

    We recently observed that specific antibodies to type II collagen do not bind in appreciable amounts to the intact surface of articular cartilage, whereas antibodies to the minor collagen types V, VI, and IX do. These results suggest that the outermost cartilage surface layer prevented interaction of the antibodies with the major collagen type in articular cartilage. The present studies were designed to investigate the pathogenic mechanisms involved in the disruption of the cartilage surface layer in inflammatory arthritis. Articular cartilage obtained from rabbits undergoing acute antigen-induced arthritis of 72 h duration showed a significant increase in binding of anti-type II antibody to cartilage surfaces compared with normal control cartilage (P less than 0.01). Augmentation of anti-type II binding was also observed upon in vitro incubation of bovine articular slices or intact rabbit patellar cartilage for 1 h with human polymorphonuclear neutrophils (PMN), PMN lysates, or purified human PMN elastase. This increase was not inhibited by sodium azide, nor was it enhanced by incubation of cartilage with the strong oxidant hypochlorous acid. Chondrocyte-mediated matrix proteoglycan degradation in cartilage explants cultured in the presence of cytokines failed to increase antibody binding appreciably. The augmentation in antibody binding seen with PMN lysates was inhibited by the nonspecific serine-esterase inhibitor PMSF, but not by the divalent metal chelator EDTA. The elastase-specific inhibitor AAPVCMK also inhibited most of the PMN-induced increase in antibody binding, whereas the cathepsin G-specific inhibitor GLPCMK was much less effective. Incubation of intact cartilage with purified human PMN elastase indicated that this serine esterase could account for the increase in anti-type II collagen antibody binding to intact cartilage surfaces. These studies suggest that in an inflammatory response, PMN-derived elastase degrades the outer layer of articular

  9. Biomimetic Analogs for Collagen Biomineralization

    PubMed Central

    Gu, L.; Kim, Y.K.; Liu, Y.; Ryou, H.; Wimmer, C.E.; Dai, L.; Arola, D.D.; Looney, S.W.; Pashley, D.H.; Tay, F.R.

    2011-01-01

    Inability of chemical phosphorylation of sodium trimetaphosphate to induce intrafibrillar mineralization of type I collagen may be due to the failure to incorporate a biomimetic analog to stabilize amorphous calcium phosphates (ACP) as nanoprecursors. This study investigated adsorption/desorption characteristics of hydrolyzed and pH-adjusted sodium trimetaphosphate (HPA-Na3P3O9) to collagen. Based on those results, a 5-minute treatment time with 2.8 wt% HPA-Na3P3O9 was used in a single-layer reconstituted collagen model to confirm that both the ACP-stabilization analog and matrix phosphoprotein analog must be present for intrafibrillar mineralization. The results of that model were further validated by complete remineralization of phosphoric-acid-etched dentin treated with the matrix phosphoprotein analog and lined with a remineralizing lining composite, and with the ACP-stabilization analog supplied in simulated body fluid. An understanding of the basic processes involved in intrafibrillar mineralization of reconstituted collagen fibrils facilitates the design of novel tissue engineering materials for hard tissue repair and regeneration. PMID:20940362

  10. Differences between top-down and bottom-up approaches in mineralizing thick, partially demineralized collagen scaffolds.

    PubMed

    Liu, Yan; Mai, Sui; Li, Nan; Yiu, Cynthia K Y; Mao, Jing; Pashley, David H; Tay, Franklin R

    2011-04-01

    Biominerals exhibit complex hierarchical structures derived from bottom-up self-assembly mechanisms. Type I collagen serves as the building block for mineralized tissues such as bone and dentin. In the present study, 250-300 μm thick, partially demineralized collagen scaffolds exhibiting a gradient of demineralization from the base to surface were mineralized using a classical top-down approach and a non-classical bottom-up approach. The top-down approach involved epitaxial growth over seed crystallites. The bottom-up approach utilized biomimetic analogs of matrix proteins to stabilize amorphous calcium phosphate nanoprecursors and template apatite nucleation and growth within the collagen matrix. Micro-computed tomography and transmission electron microscopy were employed to examine mineral uptake and apatite arrangement within the mineralized collagen matrix. The top-down approach could mineralize only the base of the partially demineralized scaffold, where remnant seed crystallites were abundant. Minimal mineralization was observed along the surface of the scaffold; extrafibrillar mineralization was predominantly observed. Conversely, the entire partially demineralized scaffold, including apatite-depleted collagen fibrils, was mineralized by the bottom-up approach, with evidence of both intrafibrillar and extrafibrillar mineralization. Understanding the different mechanisms involved in these two mineralization approaches is pivotal in adopting the optimum strategy for fabricating novel nanostructured materials in bioengineering research. PMID:21111071

  11. Differences between top-down and bottom-up approaches in mineralizing thick, partially-demineralized collagen scaffolds

    PubMed Central

    Liu, Yan; Mai, Sui; Li, Nan; Yiu, Cynthia K.Y.; Mao, Jing; Pashley, David H.; Tay, Franklin R.

    2010-01-01

    Biominerals exhibit complex hierarchical structures derived from bottom-up self-assembly mechanisms. Type I collagen serves as the building block for mineralized tissues such as bone and dentin. In the present study, 250–300 μm thick, partially-demineralized collagen scaffolds exhibiting a gradient of demineralization from the base to surface were mineralized using a classical top-down approach and a non-classical bottom-up approach. The top-down approach involved epitaxial growth over seed crystallites. The bottom-up approach utilized biomimetic analogs of matrix proteins to stabilize amorphous calcium phosphate nanoprecursors and template apatite nucleation and growth within the collagen matrix. Micro-computed tomography and transmission electron microscopy were employed to examine mineral uptake and apatite arrangement within the mineralized collagen matrix. The top-down approach could only mineralize the base of the partially-demineralized scaffold where remnant seed crystallites were abundant. Minimal mineralization was observed along the surface of the scaffold; extrafibrillar mineralization was predominantly observed. Conversely, the entire partially-demineralized scaffold including apatite-depleted collagen fibrils was mineralized by the bottom-up approach, with evidence of both intrafibrillar and extrafibrillar mineralization. Understanding the different mechanisms involved in these two mineralization approaches is pivotal in adopting the optimum strategy for fabricating novel nanostructured materials in bioengineering research. PMID:21111071

  12. Treatment of photoaged skin with topical tretinoin increases epidermal-dermal anchoring fibrils

    SciTech Connect

    Woodley, D.T.; Briggaman, R.A. ); Zelickson, A.S. ); Hamilton, T.A.; Weiss, J.S.; Ellis, C.N.; Voorhees, J.J. )

    1990-06-13

    Topical 0.1% tretinoin or vehicle control was applied daily to the forearm skin of six caucasian adults for 4 months. Two-millimeter punch biopsy specimens were obtained from treatment sites at the beginning and end of the study period for electron microscopy. Anchoring fibrils within the epidermal-dermal junction of skin treatment sites were quantitated by blinded, standardized, computer-assisted morphometry. After 4 months of continual daily treatment, skin sites that received topical tretinoin showed double the anchoring fibril density compared with vehicle control sites. The possible mechanism by which topical tretinoin increases anchoring fibrils in skin include the drug's property of inhibiting collagenase, a dermal enzyme that degrades anchoring fibril collagen. The authors speculate that increased numbers of collagenous anchoring fibrils within the papillary dermis of human skin is one of the connective-tissue correlates of the clinical improvement observed in photoaged skin after treatment with topical tretinoin.

  13. Ultrafast propagation of β-amyloid fibrils in oligomeric cloud

    PubMed Central

    Ogi, Hirotsugu; Fukukshima, Masahiko; Hamada, Hiroki; Noi, Kentaro; Hirao, Masahiko; Yagi, Hisashi; Goto, Yuji

    2014-01-01

    Interaction between monomer peptides and seeds is essential for clarifying the fibrillation mechanism of amyloid β (Aβ) peptides. We monitored the deposition reaction of Aβ1–40 peptides on immobilized seeds grown from Aβ1–42, which caused formation of oligomers in the early stage. The deposition reaction and fibrillation procedure were monitored throughout by novel total-internal-reflection-fluorescence microscopy with a quartz-crystal microbalance (TIRFM-QCM) system. This system allows simultaneous evaluation of the amount of deposited peptides on the surface seeds by QCM and fibril nucleation and elongation by TIRFM. Most fibrils reached other nuclei, forming the fibril network across the nucleus hubs in a short time. We found a fibril-elongation rate two-orders-of-magnitude higher in an oligomeric cloud than reported values, indicating ultrafast transition of oligomers into fibrils. PMID:25376301

  14. Atrial fibrillation.

    PubMed

    Bang, Casper N

    2013-10-01

    Atrial fibrillation (AF) is a common complication after myocardial infarction (MI) and new-onset AF has been demonstrated to be associated with adverse outcome and a large excess risk of death in both MI and aortic stenosis (AS) patients. Prevention of new-onset AF is therefore a potential therapeutic target in AS and MI patients. Lipid-lowering drugs, particularly statins, have anti-inflammatory and antioxidant properties that may prevent AF. Accordingly, statins are recommended as a class IIa recommendation for prevention of new-onset AF after coronary artery bypass grafting (CABG). However, this preventive effect has not been investigated on new-onset AF in asymptomatic patients with AS or a large scale first-time MI patient sample and data in patients not undergoing invasive cardiac interventions are limited. This PhD thesis was conducted at the Heart Centre, Rigshospitalet, Denmark, with the aim to investigate the three aforementioned questions and to add to the existing evidence of AF prevention with statins. This was done using three different settings: 1) a randomized patients sample of 1,873 from the Simvastatin and Ezetimibe in Aortic Stenosis (SEAS) study, 2) a register patient sample of 97,499 with first-time MI, and 3) all published studies until beginning of June 2011 examining statin treatment on new-onset and recurrent AF in patients not undergoing cardiac surgery. This thesis revealed that statins did not lower the incidence or the time to new-onset AF in patients with asymptomatic AS. However, statin treatment showed an independently preventive effect on new-onset AF, including type-dependent effect and a trend to dosage-dependent effect. In addition, this thesis showed that good compliance to statin treatment was important to prevent new-onset AF. Finally, the meta-analysis in this PhD thesis showed a preventive effect in the observational studies although this effect was absent in the randomized controlled trials. Based on this PhD thesis

  15. A titanium surface with nano-ordered spikes and pores enhances human dermal fibroblastic extracellular matrix production and integration of collagen fibers.

    PubMed

    Yamada, Masahiro; Kato, Eiji; Yamamoto, Akiko; Sakurai, Kaoru

    2016-02-01

    The acquisition of substantial dermal sealing determines the prognosis of percutaneous titanium-based medical devices or prostheses. A nano-topographic titanium surface with ordered nano-spikes and pores has been shown to induce periodontal-like connective tissue attachment and activate gingival fibroblastic functions. This in vitro study aimed to determine whether an alkali-heat (AH) treatment-created nano-topographic titanium surface could enhance human dermal fibroblastic functions and binding strength to the deposited collagen on the titanium surface. The surface topographies of commercially pure titanium machined discs exposed to two different AH treatments were evaluated. Human dermal fibroblastic cultures grown on the discs were evaluated in terms of cellular morphology, proliferation, extracellular matrix (ECM) and proinflammatory cytokine synthesis, and physicochemical binding strength of surface-deposited collagen. An isotropically-patterned, shaggy nano-topography with a sponge-like inner network and numerous well-organized, anisotropically-patterned fine nano-spikes and pores were observed on each nano-topographic surface type via scanning electron microscopy. In contrast to the typical spindle-shaped cells on the machined surfaces, the isotropically- and anisotropically-patterned nano-topographic titanium surfaces had small circular/angular cells containing contractile ring-like structures and elongated, multi-shaped cells with a developed cytoskeletal network and multiple filopodia and lamellipodia, respectively. These nano-topographic surfaces enhanced dermal-related ECM synthesis at both the protein and gene levels, without proinflammatory cytokine synthesis or reduced proliferative activity. Deposited collagen fibers were included in these surfaces and sufficiently bound to the nano-topographies to resist the physical, enzymatic and chemical detachment treatments, in contrast to machined surfaces. Well-organized, isotropically

  16. Study of the collagen structure in the superficial zone and physiological state of articular cartilage using a 3D confocal imaging technique

    PubMed Central

    Wu, Jian P; Kirk, Thomas B; Zheng, Ming H

    2008-01-01

    Introduction The collagen structure in the superficial zone of articular cartilage is critical to the tissue's durability. Early osteoarthritis is often characterized with fissures on the articular surface. This is closely related to the disruption of the collagen network. However, the traditional histology can not offer visualization of the collagen structure in articular cartilage because it uses conventional optical microscopy that does not have insufficient imaging resolution to resolve collagen from proteoglycans in hyaline articular cartilage. This study examines the 3D collagen network of articular cartilage scored from 0 to 2 in the scoring system of International Cartilage Repair Society, and aims to develop a 3D histology for assessing early osteoarthritis. Methods Articular cartilage was visually classified into five physiological groups: normal cartilage, aged cartilage, cartilage with artificial and natural surface disruption, and fibrillated. The 3D collagen matrix of the cartilage was acquired using a 3D imaging technique developed previously. Traditional histology was followed to grade the physiological status of the cartilage in the scoring system of International Cartilage Repair Society. Results Normal articular cartilage contains interwoven collagen bundles near the articular surface, approximately within the lamina splendens. However, its collagen fibres in the superficial zone orient predominantly in a direction spatially oblique to the articular surface. With age and disruption of the articular surface, the interwoven collagen bundles are gradually disappeared, and obliquely oriented collagen fibres change to align predominantly in a direction spatially perpendicular to the articular surface. Disruption of the articular surface is well related to the disappearance of the interwoven collagen bundles. Conclusion A 3D histology has been developed to supplement the traditional histology and study the subtle changes in the collagen network in the

  17. Comparative assessment of nonlinear metrics to quantify organization-related events in surface electrocardiograms of atrial fibrillation.

    PubMed

    Julián, M; Alcaraz, R; Rieta, J J

    2014-05-01

    Atrial fibrillation (AF) is today the most common sustained arrhythmia, its treatment being not completely satisfactory. Electrical activity organization analysis within the atria could play a key role in the improvement of current AF therapies. The application of a nonlinear regularity index, such as sample entropy (SampEn), to the atrial activity (AA) fundamental waveform has proven to be a successful noninvasive AF organization estimator. However, the use of alternative nonlinear metrics within this context is a pending issue. The present work analyzes the ability of several nonlinear indices to assess regularity of patterns and, thus, organization, in the AA signal and its fundamental waveform, defined as the main atrial wave (MAW). Precisely, Fuzzy Entropy, Spectral Entropy, Lempel-Ziv Complexity and Hurst Exponents were studied, achieving more robust and accurate AF organization estimates than SampEn. Results also provided better AF organization estimates from the MAW than from the AA signal for all the tested nonlinear metrics, which agrees with previous works only focused on SampEn. Furthermore, some of these indices reported a discriminant ability close to 95% in the classification of AF organization-dependent events, thus outperforming the diagnostic accuracy of SampEn and other widely used noninvasive estimators, such as the dominant atrial frequency (DAF). As a conclusion, these nonlinear metrics could be considered as promising estimators of noninvasive AF organization and could be helpful in making appropriate decisions on the patients' management. PMID:24642478

  18. Quantification of Collagen Ultrastructure after Penetrating Keratoplasty – Implications for Corneal Biomechanics

    PubMed Central

    Gardner, Steven J.; Kamma-Lorger, Christina S.; Hayes, Sally; Nielsen, Kim; Hjortdal, Jesper; Sorensen, Thomas; Terrill, Nicholas J.; Meek, Keith M.

    2013-01-01

    Purpose To quantify long-term changes in stromal collagen ultrastructure following penetrating keratoplasty (PK), and evaluate their possible implications for corneal biomechanics. Methods A pair of 16 mm post-mortem corneo-scleral buttons was obtained from a patient receiving bilateral penetrating keratoplasty 12 (left)/28 (right) years previously. Small-angle x-ray scattering quantified collagen fibril spacing, diameter and spatial order at 0.5 mm or 0.25 mm intervals along linear scans across the graft margin. Corresponding control data was collected from two corneo-scleral buttons with no history of refractive surgery. Wide-angle x-ray scattering quantified collagen fibril orientation at 0.25 mm (horizontal)×0.25 mm (vertical) intervals across both PK specimens. Quantification of orientation changes in the graft margin were verified by equivalent analysis of data from a 13 year post-operative right PK specimen obtained from a second patient in a previous study, and comparison made with new and published data from normal corneas. Results Marked changes to normal fibril alignment, in favour of tangentially oriented collagen, were observed around the entire graft margin in all PK specimens. The total number of meridional fibrils in the wound margin was observed to decrease by up to 40%, with the number of tangentially oriented fibrils increasing by up to 46%. As a result, in some locations the number of fibrils aligned parallel to the wound outnumbered those spanning it by up to five times. Localised increases in fibril spacing and diameter, with an accompanying reduction in matrix order, were also evident. Conclusions Abnormal collagen fibril size and spatial order within the PK graft margin are indicative of incomplete stromal wound remodelling and the long term persistence of fibrotic scar tissue. Lasting changes in collagen fibril orientation in and around PK wounds may alter corneal biomechanics and compromise the integrity of the graft-host interface in the

  19. Function of glycoprotein VI and integrin alpha2beta1 in the procoagulant response of single, collagen-adherent platelets.

    PubMed

    Heemskerk, J W; Siljander, P; Vuist, W M; Breikers, G; Reutelingsperger, C P; Barnes, M J; Knight, C G; Lassila, R; Farndale, R W

    1999-05-01

    Various collagen-based materials were used to assess the structural requirements of collagen for inducing the procoagulant response of adhering platelets, as well as the collagen receptors involved. Cross-linked or monomeric collagen-related peptide (CRP), Gly-Cys-Hyp-(Gly-Pro-Hyp)10-Gly-Cys-Hyp-Gly was highly adhesive for platelets in a glycoprotein VI-(GpVI-)dependent manner. Adhesion was followed by a prolonged increase in cytosolic [Ca2+]i, formation of membrane blebs, exposure of phosphatidylserine (PS) and generation of prothrombinase-stimulating activity. Fibrils of type-I collagen were less adhesive but, once adhered, many of the platelets presented a full procoagulant response. Monomeric type-I collagen was unable to support adhesion, unless Mg(2+)-dependent integrin alpha2beta1 interactions were facilitated by omission of Ca2+ ions. With all surfaces, however, post-addition of CaCl2 to adhering platelets resulted in a potent Ca(2+)-influx signal, followed by PS exposure and bleb formation. The procoagulant response elicited by binding to CRP was inhibited by anti-GpVI Fab fragments, but not by impeding integrin alpha2beta1-mediated events. With fibrillar collagen, it was inhibited by blocking either the GpVI- or integrin alpha2beta1-mediated interactions. This suggests that the triple-helical Gly-Pro-Hyp repeat in CRP and analogous sequences in fibrillar collagen stimulate the procoagulant response of adherent platelets by acting as ligands for GpVI. Influx of Ca2+ is required for this response, and adhesion via integrin alpha2beta1 serves to potentiate the signaling effects of GpVI. PMID:10365754

  20. Magnetite nanoparticle interactions with insulin amyloid fibrils.

    PubMed

    Chen, Yun-Wen; Chang, Chiung-Wen; Hung, Huey-Shan; Kung, Mei-Lang; Yeh, Bi-Wen; Hsieh, Shuchen

    2016-10-14

    Accumulation of amyloid fibrils is one of the likely key factors leading to the development of Alzheimer's disease and other amyloidosis associated diseases. Magnetic nanoparticles (NPs) have been developed as promising medical materials for many medical applications. In this study, we have explored the effects of Fe3O4 NPs on the fibrillogenesis process of insulin fibrils. When Fe3O4 NPs were co-incubated with insulin, Fe3O4 NPs had no effect on the structural transformation into amyloid-like fibrils but had higher affinity toward insulin fibrils. We demonstrated that the zeta potential of insulin fibrils and Fe3O4 NPs were both positive, suggesting the binding forces between Fe3O4 NPs and insulin fibrils were van der Waals forces but not surface charge. Moreover, a different amount of Fe3O4 NPs added had no effect on secondary structural changes of insulin fibrils. These results propose the potential use of Fe3O4 NPs as therapeutic agents against diseases related to protein aggregation or contrast agents for magnetic resonance imaging. PMID:27585675

  1. The streptococcal collagen-binding protein CNE specifically interferes with alphaVbeta3-mediated cellular interactions with triple helical collagen.

    PubMed

    van Wieringen, Tijs; Kalamajski, Sebastian; Lidén, Asa; Bihan, Dominique; Guss, Bengt; Heinegård, Dick; Farndale, Richard W; Rubin, Kristofer

    2010-11-12

    Collagen fibers expose distinct domains allowing for specific interactions with other extracellular matrix proteins and cells. To investigate putative collagen domains that govern integrin α(V)β(3)-mediated cellular interactions with native collagen fibers we took advantage of the streptococcal protein CNE that bound native fibrillar collagens. CNE specifically inhibited α(V)β(3)-dependent cell-mediated collagen gel contraction, PDGF BB-induced and α(V)β(3)-mediated adhesion of cells, and binding of fibronectin to native collagen. Using a Toolkit composed of overlapping, 27-residue triple helical segments of collagen type II, two CNE-binding sites present in peptides II-1 and II-44 were identified. These peptides lack the major binding site for collagen-binding β(1) integrins, defined by the peptide GFOGER. Peptide II-44 corresponds to a region of collagen known to bind collagenases, discoidin domain receptor 2, SPARC (osteonectin), and fibronectin. In addition to binding fibronectin, peptide II-44 but not II-1 inhibited α(V)β(3)-mediated collagen gel contraction and, when immobilized on plastic, supported adhesion of cells. Reduction of fibronectin expression by siRNA reduced PDGF BB-induced α(V)β(3)-mediated contraction. Reconstitution of collagen types I and II gels in the presence of CNE reduced collagen fibril diameters and fibril melting temperatures. Our data indicate that contraction proceeded through an indirect mechanism involving binding of cell-produced fibronectin to the collagen fibers. Furthermore, our data show that cell-mediated collagen gel contraction does not directly depend on the process of fibril formation. PMID:20837478

  2. A Mechanistic study of Plasma Treatment Effects on Demineralized Dentin Surfaces for Improved Adhesive/Dentin Interface Bonding

    PubMed Central

    Dong, Xiaoqing; Chen, Meng; Wang, Yong; Yu, Qingsong

    2014-01-01

    Our previous work has shown that non-thermal plasma treatment of demineralized dentin significantly (p<0.05) improved adhesive/dentin bonding strength for dental composite restoration as compared with the untreated controls. This study is to achieve mechanistic understanding of the plasma treatment effects on dentin surface through investigating the plasma treated dentin surfaces and their interaction with adhesive monomer, 2-Hydroxyethyl methacrylate (HEMA). The plasma treated dentin surfaces from human third molars were evaluated by water contact angle measurements and scanning electron microscopy (SEM). It was found that plasma-treated dentin surface with subsequent HEMA immersion (Plasma/HEMA Treated) had much lower water contact angle compared with only plasma-treated (Plasma Treated) or only HEMA immersed (HEMA Treated) dentin surfaces. With prolong water droplet deposition time, water droplets spread out completely on the Plasma/HEMA Treated dentin surfaces. SEM images of Plasma/HEMA Treated dentin surfaces verified that dentin tubules were opened-up and filled with HEMA monomers. Extracted type I collagen fibrils, which was used as simulation of the exposed dentinal collagen fibrils after acid etching step, were plasma treated and analyzed with Fourier transform infrared spectroscopy (FT-IR) and circular dichroism (CD) spectra. FT-IR spectra of the Plasma/HEMA Treated collage fibrils showed broadened amide I peak at 1660 cm−1 and amide II at 1550 cm−1, which indicate secondary structure changes of the collagen fibrils. CD spectra indicated that 67.4% collagen helix structures were denatured after plasma treatment. These experimental results demonstrate that non-thermal argon plasma treatment was very effective in loosing collagen structure and enhancing adhesive monomer penetration, which are beneficial to thicker hybrid layer and longer resin tag formation, and consequently enhance adhesive/dentin interface bonding. PMID:25267936

  3. Development and utilization of a bovine type I collagen microfibril model

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The structure of fibrous collagen, a long triple helix that self-associates in a staggered array to form a matrix of fibrils, fibers and fiber bundles, makes it uniquely suitable as a scaffold for biomaterial engineering. A major challenge for this application is to stabilize collagen structure by m...

  4. Mineralisation of reconstituted collagen using polyvinylphosphonic acid/polyacrylic acid templating matrix protein analogues in the presence of calcium, phosphate and hydroxyl ions

    PubMed Central

    Kim, Young Kyung; Gu, Li-sha; Bryan, Thomas E.; Kim, Jong Ryul; Chen, Liang; Liu, Yan; Yoon, James C.; Breschi, Lorenzo; Pashley, David H.; Tay, Franklin R.

    2010-01-01

    The complex morphologies of mineralised collagen fibrils are regulated through interactions between the collagen matrix and non-collagenous extracellular proteins. In the present study, polyvinylphosphonic acid, a biomimetic analogue of matrix phosphoproteins, was synthesised and confirmed with FTIR and NMR. Biomimetic mineralisation of reconstituted collagen fibrils devoid of natural non-collagenous proteins was demonstrated with TEM using a Portland cement-containing resin composite and a phosphate-containing fluid in the presence of polyacrylic acid as sequestration, and polyvinylphosphonic acid as templating matrix protein analogues. In the presence of these dual biomimetic analogues in the mineralisation medium, intrafibrillar and extrafibrillar mineralisation via bottom-up nanoparticle assembly based on the nonclassical crystallisation pathway could be identified. Conversely, only large mineral spheres with no preferred association with collagen fibrils were observed in the absence of biomimetic analogues in the medium. Mineral phases were evident within the collagen fibrils as early as 4 hours after the initially-formed amorphous calcium phosphate nanoprecursors were transformed into apatite nanocrystals. Selected area electron diffraction patterns of highly mineralised collagen fibrils were nearly identical to those of natural bone, with apatite crystallites preferentially aligned along the collagen fibril axes. PMID:20621767

  5. Mineralisation of reconstituted collagen using polyvinylphosphonic acid/polyacrylic acid templating matrix protein analogues in the presence of calcium, phosphate and hydroxyl ions.

    PubMed

    Kim, Young Kyung; Gu, Li-sha; Bryan, Thomas E; Kim, Jong R; Chen, Liang; Liu, Yan; Yoon, James C; Breschi, Lorenzo; Pashley, David H; Tay, Franklin R

    2010-09-01

    The complex morphologies of mineralised collagen fibrils are regulated through interactions between the collagen matrix and non-collagenous extracellular proteins. In the present study, polyvinylphosphonic acid, a biomimetic analogue of matrix phosphoproteins, was synthesised and confirmed with FTIR and NMR. Biomimetic mineralisation of reconstituted collagen fibrils devoid of natural non-collagenous proteins was demonstrated with TEM using a Portland cement-containing resin composite and a phosphate-containing fluid in the presence of polyacrylic acid as sequestration, and polyvinylphosphonic acid as templating matrix protein analogues. In the presence of these dual biomimetic analogues in the mineralisation medium, intrafibrillar and extrafibrillar mineralisation via bottom-up nanoparticle assembly based on the non-classical crystallisation pathway could be identified. Conversely, only large mineral spheres with no preferred association with collagen fibrils were observed in the absence of biomimetic analogues in the medium. Mineral phases were evident within the collagen fibrils as early as 4 h after the initially-formed amorphous calcium phosphate nanoprecursors were transformed into apatite nanocrystals. Selected area electron diffraction patterns of highly mineralised collagen fibrils were nearly identical to those of natural bone, with apatite crystallites preferentially aligned along the collagen fibril axes. PMID:20621767

  6. Pre-adsorbed type-I collagen structure-dependent changes in osteoblastic phenotype

    SciTech Connect

    Hanagata, Nobutaka . E-mail: HANAGATA.Nobutaka@nims.go.jp; Takemura, Taro; Monkawa, Akira; Ikoma, Toshiyuki; Tanaka, Junzo

    2006-06-16

    Type-I collagen is the most abundant extracellular matrix in bones and modulates various functions of osteoblasts. We prepared two different structures of type-I collagen on tissue culture grade polystylene (TCPS) surfaces, one is feltwork structure of filamentous molecules from acid solutions (ACs) and the other is network structure of fibrils from neutral solutions (NCs), to examine effects of the structures on the maturation process of osteoblast-like cells. No significant differences of cell proliferation were observed between TCPS and ACs, but NCs delayed the proliferation. In initial cell attachment, the cells on ACs had tense lamellipodia with sharp tips, while those on NCs had loose lamellipodia. No detectable differences in levels of expressed integrin {alpha}{sub 2}- and {alpha}{sub 5}-subunits were observed between the structures. Although the matrix mineralization in NCs was also delayed in comparison with TCPS and ACs, fully mineralized levels in NCs were the same as those of TCPS and ACs. In addition, although we examined the effects of densities of pre-adsorbed collagen molecules on osteoblast maturation, the effects were less serious than those of the structures. This study suggests that the structures of collagen affect proliferation and mineralization of osteoblast-like cells.

  7. Development of a nonthrombogenic collagenous blood-prosthetic interface.

    PubMed Central

    Bernhard, W F; Colo, N A; Szycher, M; Wesolowski, J S; Haudenschild, C C; Franzblau, C C; Parkman, R; Liss, R H

    1980-01-01

    Investigations to develop an implantable assist pump for prolonged circulatory support have been impeded by accumulation of friable thrombus on the prosthetic interface, with subsequent embolization. To circumvent this problem, the textured, fibril surface of a polyurethane pump chamber (mat thickness 430 microns) was inoculated with cultured bovine fetal fibroblasts (labelled with thymidine-14C) prior to animal implantation. The pneumatically actuated device (stroke volume 75 ml), maintained a pulsatile blood flow throughout each study. In 20 calf experiments, extending up to 335 days, 30 X 10(6) fibroblasts (in 50 ml media) derived from a single Holstein fetus were distributed on the urethane surface (360 +/- 50 cells/mm2) by rotation of a sealed device for three hours (12 revolutions/hour). Following connection to the circulation, cell washout was minimal. Resultant biologic linings, examined after animal sacrifice, were densely adherent to the underlying polymer matrix, and varied in thickness from 250 micron-1.5 mm. Microscopically, fibroblasts were identified from the surface to base, accompanied by numerous collagen bundles and abundant ground substance. Amino acid analysis in 10/20 pumps implanted for 31--335 days, revealed 50 +/- 5 Hydroxyproline residues/1000 residues (50% collagen) and scant elastin. Donor fibroblasts were identified by radioautography and karyotyping. Lack of immunologic response in 12 Hereford pump recipients as confirmed by serial fibroblast cytotoxicity assays. In conclusion, an induced collagenous-blood interface permitted prolonged mechanical circulatory support in animals without thromboembolic complications. Images Fig. 1. Fig. 2. Figs. 3A and B. Fig. 4A. Fig. 4B. PMID:6448027

  8. Highly nonlinear stress-relaxation response of articular cartilage in indentation: Importance of collagen nonlinearity.

    PubMed

    Mäkelä, J T A; Korhonen, R K

    2016-06-14

    Modern fibril-reinforced computational models of articular cartilage can include inhomogeneous tissue composition and structure, and nonlinear mechanical behavior of collagen, proteoglycans and fluid. These models can capture well experimental single step creep and stress-relaxation tests or measurements under small strains in unconfined and confined compression. Yet, it is known that in indentation, especially at high strain velocities, cartilage can express highly nonlinear response. Different fibril reinforced poroelastic and poroviscoelastic models were used to assess measured highly nonlinear stress-relaxation response of rabbit articular cartilage in indentation. Experimentally measured depth-dependent volume fractions of different tissue constituents and their mechanical nonlinearities were taken into account in the models. In particular, the collagen fibril network was modeled using eight separate models that implemented five different constitutive equations to describe the nonlinearity. These consisted of linear elastic, nonlinear viscoelastic and multiple nonlinear elastic representations. The model incorporating the most nonlinearly increasing Young׳s modulus of collagen fibrils as a function of strain captured best the experimental data. Relative difference between the model and experiment was ~3%. Surprisingly, the difference in the peak forces between the experiment and the model with viscoelastic collagen fibrils was almost 20%. Implementation of the measured volume fractions did not improve the ability of the model to capture the measured mechanical data. These results suggest that a highly nonlinear formulation for collagen fibrils is needed to replicate multi-step stress-relaxation response of rabbit articular cartilage in indentation with high strain rates. PMID:27130474

  9. Nucleation of amyloid fibrils

    NASA Astrophysics Data System (ADS)

    Kashchiev, Dimo; Auer, Stefan

    2010-06-01

    We consider nucleation of amyloid fibrils in the case when the process occurs by the mechanism of direct polymerization of practically fully extended protein segments, i.e., β-strands, into β-sheets. Applying the classical nucleation theory, we derive a general expression for the work to form a nanosized amyloid fibril (protofilament) constituted of successively layered β-sheets. Analysis of this expression reveals that with increasing its size, the fibril transforms from one-dimensional to two-dimensional aggregate in order to preserve the equilibrium shape corresponding to minimal formation work. We determine the size of the fibril nucleus, the fibril nucleation work, and the fibril nucleation rate as explicit functions of the concentration and temperature of the protein solution. The results obtained are applicable to homogeneous nucleation, which occurs when the solution is sufficiently pure and/or strongly supersaturated.

  10. Internal strain drives spontaneous periodic buckling in collagen and regulates remodeling

    PubMed Central

    Dittmore, Andrew; Silver, Jonathan; Sarkar, Susanta K.; Marmer, Barry; Goldberg, Gregory I.; Neuman, Keir C.

    2016-01-01

    Fibrillar collagen, an essential structural component of the extracellular matrix, is remarkably resistant to proteolysis, requiring specialized matrix metalloproteinases (MMPs) to initiate its remodeling. In the context of native fibrils, remodeling is poorly understood; MMPs have limited access to cleavage sites and are inhibited by tension on the fibril. Here, single-molecule recordings of fluorescently labeled MMPs reveal cleavage-vulnerable binding regions arrayed periodically at ∼1-µm intervals along collagen fibrils. Binding regions remain periodic even as they migrate on the fibril, indicating a collective process of thermally activated and self-healing defect formation. An internal strain relief model involving reversible structural rearrangements quantitatively reproduces the observed spatial patterning and fluctuations of defects and provides a mechanism for tension-dependent stabilization of fibrillar collagen. This work identifies internal–strain-driven defects that may have general and widespread regulatory functions in self-assembled biological filaments. PMID:27402741

  11. Pathways of tau fibrillization.

    PubMed

    Kuret, Jeff; Chirita, Carmen N; Congdon, Erin E; Kannanayakal, Theresa; Li, Guibin; Necula, Mihaela; Yin, Haishan; Zhong, Qi

    2005-01-01

    New methods for analyzing tau fibrillization have yielded insights into the biochemical transitions involved in the process. Here we review the parallels between the sequential progression of tau fibrillization observed macroscopically in Alzheimer's disease (AD) lesions and the pathway of tau aggregation observed in vitro with purified tau preparations. In addition, pharmacological agents for further dissection of fibrillization mechanism and lesion formation are discussed. PMID:15615636

  12. Preoperative study of the surface ECG for the prognosis of atrial fibrillation maze surgery outcome at discharge.

    PubMed

    Hernández, Antonio; Alcaraz, Raúl; Hornero, Fernando; Rieta, José Joaquín

    2014-07-01

    The Cox-maze surgery is an effective procedure for terminating atrial fibrillation (AF) in patients requiring open-heart surgery associated with another heart disease. After the intervention, regardless of the patient's rhythm, all are treated with oral anticoagulants and antiarrhythmic drugs prior to discharge. Furthermore, patients maintaining AF before discharge could also be treated with electrical cardioversion (ECV). In view of this, a preoperative prognosis of the patient's rhythm at discharge would be helpful for optimizing drug therapy planning as well as for advancing ECV therapy. This work analyzes 30 preoperative electrocardiograms (ECGs) from patients suffering from AF in order to predict the Cox-maze surgery outcome at discharge. Two different characteristics of the AF pattern have been studied. On the one hand, the atrial activity (AA) organization, which provides information about the number of propagating wavelets in the atria, was investigated. AA organization has been successfully used in previous studies related to spontaneous reversion of paroxysmal AF and to the outcome of ECV. To assess organization, the dominant atrial frequency (DAF) and sample entropy (SampEn) have been computed. On the other hand, the second characteristic studied was the fibrillatory wave (f-wave) amplitude, which has been demonstrated to be a valuable indicator of the Cox-maze surgery outcome in previous studies. Moreover, this parameter has been obtained through a new methodology, based on computing the f-wave average power (fWP). Finally, all the computed indices were combined in a decision tree in order to improve prediction capability. Results for the DAF yielded a sensitivity (Se), a specificity (Sp) and an accuracy (Acc) of 61.54%, 82.35% and 73.33%, respectively. For SampEn the values were 69.23%, 76.00% and 73.33%, respectively, and for fWP they were 92.31%, 82.35% and 86.67%, respectively. Finally, the decision tree combining the three parameters analyzed

  13. Biomimetic remineralization as a progressive dehydration mechanism of collagen matrices – implications in the aging of resin-dentin bonds

    PubMed Central

    Kim, Young Kyung; Mai, Sui; Mazzoni, Annalisa; Liu, Yan; Tezvergil-Mutluay, Arzu; Takahashi, Kei; Zhang, Kai; Pashley, David H.; Tay, Franklin R.

    2010-01-01

    Biomineralization is a dehydration process in which water from the intrafibrillar compartments of collagen fibrils are progressively replaced by apatites. As water is an important element that precipitates the lack of durability of resin-dentin bonds, this study examined the use of a biomimetic remineralization strategy as a progressive dehydration mechanism for preserving joint integrity and maintaining adhesive strength after aging. Human dentin surfaces were bonded with dentin adhesives, restored with resin composites and sectioned into sticks containing the adhesive joint. Experimental specimens were aged in a biomimetic analog-containing remineralizing medium and control specimens in simulated body fluid for up to 12 months. Specimens retrieved from the designated periods were examined by transmission electron microscopy for manifestation of water-rich regions using a silver tracer and for collagen degradation within the adhesive joints. Tensile testing was performed to determine the potential loss of bond integrity after aging. Control specimens exhibited severe collagen degradation within the adhesive joint after aging. Remineralized specimens exhibited progressive dehydration as manifested by silver tracer reduction and partial remineralization of water-filled micro-channels within the adhesive joint, as well as intrafibrillar remineralization of collagen fibrils that were demineralized initially as part of the bonding procedure. Biomimetic remineralization as a progressive dehydration mechanism of water-rich, resin-sparse collagen matrices enables those adhesive joints to resist degradation over the 12-month aging period, as verified by the conservation of their tensile bond strengths. The ability of the proof-of-concept biomimetic remineralization strategy to prevent bond degradation warrants further development of clinically-relevant delivery systems. PMID:20304110

  14. Exposure to Mimivirus Collagen Promotes Arthritis

    PubMed Central

    Shah, Nikunj; Hülsmeier, Andreas J.; Hochhold, Nina; Neidhart, Michel; Gay, Steffen

    2014-01-01

    Collagens, the most abundant proteins in animals, also occur in some recently described nucleocytoplasmic large DNA viruses such as Mimiviridae, which replicate in amoebae. To clarify the impact of viral collagens on the immune response of animals exposed to Mimiviridae, we have investigated the localization of collagens in Acanthamoeba polyphaga mimivirus particles and the response of mice to immunization with mimivirus particles. Using protein biotinylation, we have first shown that viral collagen encoded by open reading frame L71 is present at the surface of mimivirus particles. Exposure to mimivirus collagens elicited the production of anti-collagen antibodies in DBA/1 mice immunized intradermally with mimivirus protein extracts. This antibody response also targeted mouse collagen type II and was accompanied by T-cell reactivity to collagen and joint inflammation, as observed in collagen-induced arthritis following immunization of mice with bovine collagen type II. The broad distribution of nucleocytoplasmic large DNA viruses in the environment suggests that humans are constantly exposed to such large virus particles. A survey of blood sera from healthy human subjects and from rheumatoid arthritis patients indeed demonstrated that 30% of healthy-subject and 36% of rheumatoid arthritis sera recognized the major mimivirus capsid protein L425. Moreover, whereas 6% of healthy-subject sera recognized the mimivirus collagen protein L71, 22% of rheumatoid arthritis sera were positive for mimivirus L71. Accordingly, our study shows that environmental exposure to mimivirus represents a risk factor in triggering autoimmunity to collagens. PMID:24173233

  15. Fibronectin provides a conduit for fibroblast transmigration from collagenous stroma into fibrin clot provisional matrix.

    PubMed

    Greiling, D; Clark, R A

    1997-04-01

    After injury, the wound space is filled with a fibrin/fibronectin clot containing growth factors released by platelets and monocytes. In response to these factors, fibroblasts migrate into the fibrin clot and contribute to the formation of granulation tissue. The functional mechanisms allowing fibroblasts to leave the collagenous matrix of normal connective tissue and invade the provisional matrix of the fibrin clot have not been fully defined. To investigate these mechanisms we established a new in vitro model which simulates specific aspects of early wound healing, that is, the migration of fibroblasts from a three-dimensional collagen matrix into a fibrin clot. This transmigration could be induced by physiological concentrations of platelet releasate or platelet-derived growth factor BB (PDGF-BB) in a concentration-dependent manner. At 24 hours irradiated fibroblasts invaded the fibrin gel almost as well as non-irradiated cells, indicating that transmigration was independent of proliferation. Plasminogen and its activators appear to be necessary for invasion of the fibrin clot since protease inhibitors decreased the amount of migration. These serine proteases, however, were not necessary for exit from the collagen gel as fibroblasts migrated out of the collagen gel onto a surface coated with fibrin fibrils even in the presence of inhibitors. Removal of fibronectin (FN) from either the collagen gel or the fibrin gel markedly decreased the number of migrating cells, suggesting that FN provides a conduit for transmigration. Cell movement in the in vitro model was inhibited by RGD peptide, and by monoclonal antibodies against the subunits of the alpha5 beta1 and alpha v beta3 integrin receptor. Thus, the functional requirements for fibroblast transmigration from collagen-rich to fibrin-rich matrices, such as occurs in early wound healing, have been partially defined using an in vitro paradigm of this important biologic process. PMID:9133673

  16. Biomimetic silicification of demineralized hierarchical collagenous tissues

    PubMed Central

    Ryou, Heonjune; Diogenes, Anibal; Yiu, Cynthia K.Y.; Mazzoni, Annalisa; Chen, Ji-hua; Arola, Dwayne D.; Hargreaves, Kenneth M.; Pashley, David H.; Tay, Franklin R.

    2013-01-01

    Unlike man-made composite materials, natural biominerals containing composites usually demonstrate different levels of sophisticated hierarchical structures which are responsible for their mechanical properties and other metabolic functions. However, the complex spatial organizations of the organic-inorganic phases are far beyond what they be achieved by contemporary engineering techniques. Here, we demonstrate that carbonated apatite present in collagen matrices derived from fish scale and bovine bone may be replaced by amorphous silica, using an approach that simulates what is utilized by phylogenetically ancient glass sponges. The structural hierarchy of these collagen-based biomaterials is replicated by the infiltration and condensation of fluidic polymer-stabilized silicic acid precursors within the intrafibrillar milieu of type I collagen fibrils. This facile biomimetic silicification strategy may be used for fabricating silica-based, three-dimensional functional materials with specific morphological and hierarchical requirements. PMID:23586938

  17. Nucleation of protein fibrillation by nanoparticles

    PubMed Central

    Linse, Sara; Cabaleiro-Lago, Celia; Xue, Wei-Feng; Lynch, Iseult; Lindman, Stina; Thulin, Eva; Radford, Sheena E.; Dawson, Kenneth A.

    2007-01-01

    Nanoparticles present enormous surface areas and are found to enhance the rate of protein fibrillation by decreasing the lag time for nucleation. Protein fibrillation is involved in many human diseases, including Alzheimer's, Creutzfeld-Jacob disease, and dialysis-related amyloidosis. Fibril formation occurs by nucleation-dependent kinetics, wherein formation of a critical nucleus is the key rate-determining step, after which fibrillation proceeds rapidly. We show that nanoparticles (copolymer particles, cerium oxide particles, quantum dots, and carbon nanotubes) enhance the probability of appearance of a critical nucleus for nucleation of protein fibrils from human β2-microglobulin. The observed shorter lag (nucleation) phase depends on the amount and nature of particle surface. There is an exchange of protein between solution and nanoparticle surface, and β2-microglobulin forms multiple layers on the particle surface, providing a locally increased protein concentration promoting oligomer formation. This and the shortened lag phase suggest a mechanism involving surface-assisted nucleation that may increase the risk for toxic cluster and amyloid formation. It also opens the door to new routes for the controlled self-assembly of proteins and peptides into novel nanomaterials. PMID:17485668

  18. The nucleation and growth of calcium phosphate crystals at protein and phosphatidylserine liposome surfaces.

    PubMed

    Nancollas, G H; Tsortos, A; Zieba, A

    1996-01-01

    The kinetics of calcium phosphate crystal growth at the surfaces of proteins and phospholipids has been investigated using free drift and constant composition methods in supersaturated calcium phosphate solutions (relative supersaturations: with respect to hydroxyapatite, HAP, sigma HAP = 15.0, and with respect to octacalcium phosphate, OCP, sigma OCP = 1.9). Fibrinogen and collagen molecules adsorbed at hydrophobic surfaces as well as uncross-linked collagen fibrils induce ion binding and subsequent nucleation of calcium phosphate. The formation of OCP on phosphatidylserine vesicles introduced to highly supersaturated calcium phosphate solutions probably involves the interaction of the calcium ions with the ionized carboxylic groups of the phospholipid. PMID:9813627

  19. Nanostructure and mechanics of mummified type I collagen from the 5300-year-old Tyrolean Iceman

    PubMed Central

    Janko, Marek; Zink, Albert; Gigler, Alexander M.; Heckl, Wolfgang M.; Stark, Robert W.

    2010-01-01

    Skin protects the body from pathogens and degradation. Mummified skin in particular is extremely resistant to decomposition. External influences or the action of micro-organisms, however, can degrade the connective tissue and lay the subjacent tissue open. To determine the degree of tissue preservation in mummified human skin and, in particular, the reason for its durability, we investigated the structural integrity of its main protein, type I collagen. We extracted samples from the Neolithic glacier mummy known as ‘the Iceman’. Atomic force microscopy (AFM) revealed collagen fibrils that had characteristic banding patterns of 69 ± 5 nm periodicity. Both the microstructure and the ultrastructure of dermal collagen bundles and fibrils were largely unaltered and extremely well preserved by the natural conservation process. Raman spectra of the ancient collagen indicated that there were no significant modifications in the molecular structure. However, AFM nanoindentation measurements showed slight changes in the mechanical behaviour of the fibrils. Young's modulus of single mummified fibrils was 4.1 ± 1.1 GPa, whereas the elasticity of recent collagen averages 3.2 ± 1.0 GPa. The excellent preservation of the collagen indicates that dehydration owing to freeze-drying of the collagen is the main process in mummification and that the influence of the degradation processes can be addressed, even after 5300 years. PMID:20356896

  20. Thermal Destabilization of Collagen Matrix Hierarchical Structure by Freeze/Thaw

    PubMed Central

    Ozcelikkale, Altug; Han, Bumsoo

    2016-01-01

    This study aims to characterize and understand the effects of freezing on collagen structures and functionality. Specifically, thermodynamic destabilization of collagen at molecular- and fibril-levels by combination of low temperatures and freezing were experimentally characterized using modulated differential scanning calorimetry. In order to delineate the effects of sub-zero temperature and water-ice phase change, we hypothesized that the extent of destabilization can be determined based on post-thaw heat induced thermal denaturation of collagen. It is found that thermal denaturation temperature of collagen in hydrogel decreases by 1.4–1.6°C after freeze/thaw while no such decrease is observed in the case of molecular solution. The destabilization is predominantly due to ice formation. Exposure to low temperatures in the absence of ice has only minimal effect. Calorimetry measurements combined with morphological examination of collagen matrices by scanning electron microscopy suggest that freezing results in destabilization of collagen fibrils due to expansion of intrafibrillar space by ice formation. This fibril-level damage can be alleviated by use of cryoprotectant DMSO at concentrations as low as 0.5 M. A theoretical model explaining the change in collagen post-thaw thermal stability by freezing-induced fibril expansion is also proposed. PMID:26765741

  1. In vitro mineralization of dense collagen substrates: a biomimetic approach toward the development of bone-graft materials.

    PubMed

    Thula, Taili T; Rodriguez, Douglas E; Lee, Myong Hwa; Pendi, Laura; Podschun, Jacob; Gower, Laurie B

    2011-08-01

    Bone is an organic-inorganic composite which has hierarchical structuring that leads to high strength and toughness. The nanostructure of bone consists of nanocrystals of hydroxyapatite embedded and aligned within the interstices of collagen fibrils. This unique nanostructure leads to exceptional properties, both mechanical and biological, making it difficult to emulate bone properties without having a bone-like nanostructured material. A primary goal of our group's work is to use biomimetic processing techniques that lead to bone-like structures. In our prior studies, we demonstrated that intrafibrillar mineralization of porous collagen sponges, leading to a bone-like nanostructure, can be achieved using a polymer-induced liquid precursor (PILP) mineralization process. The objective of this study was to investigate the use of this polymer-directed crystallization process to mineralize dense collagen substrates. To examine collagen scaffolds that truly represent the dense-packed matrix of bone, manatee bone was demineralized to isolate its collagen matrix, consisting of a dense, lamellar osteonal microstructure. This biogenic collagen scaffold was then remineralized using polyaspartate to direct the mineralization process through an amorphous precursor pathway. The various conditions investigated included polymer molecular weight, substrate dimension and mineralization time. Mineral penetration depths of up to 100 μms were achieved using this PILP process, compared to no penetration with only surface precipitates observed for the conventional crystallization process. Electron microscopy, wide-angle X-ray diffraction and thermal analysis were used to characterize the resulting hydroxyapatite/collagen composites. These studies demonstrate that the original interpenetrating bone nanostructure and osteonal microstructure could be recovered in a biogenic matrix using the PILP process. PMID:21550424

  2. Nucleation-fibrillation dynamics of Aβ1-40 peptides on liquid-solid surface studied by total-internal-reflection fluorescence microscopy coupled with quartz-crystal microbalance biosensor

    NASA Astrophysics Data System (ADS)

    Hamada, Hiroki; Ogi, Hirotsugu; Noi, Kentaro; Yagi, Hisashi; Goto, Yuji; Hirao, Masahiko

    2015-07-01

    We have successfully developed the total-internal-reflection-fluorescence microscopy combined with a quartz-crystal microbalance (TIRFM-QCM) biosensor, and monitored the nucleation-fibrillation phenomenon of amyloid β1-40 peptide on the naked quartz surface. The cross-β-sheet structures were visualized with the TIRFM using the thioflavin-T (Th-T) label, and other unlabeled aggregates were detected through the frequency change of the 58-MHz wireless-electrodeless QCM throughout the aggregation reaction. The QCM response indicates significant adsorption of the peptides on the quartz surface at the early stage, which is followed by fibrillation. The non-cross-β-sheet oligomers are first formed, and nuclei appear in the oligomer region, from which fibrils originate and elongate. The two-color TIRFM observation was performed after the aggregation reaction with the Nile-red label as well as the ThT label for identifying nucleation from non-β-sheet regions. An aggregation model is proposed.

  3. Atomic force microscopy reveals a dual collagen-binding activity for the staphylococcal surface protein SdrF.

    PubMed

    Herman-Bausier, Philippe; Dufrêne, Yves F

    2016-02-01

    Staphylococcus epidermidis causes nosocomial infections by colonizing and forming biofilms on indwelling medical devices. This process involves specific interactions between cell wall-anchored (CWA) proteins and host proteins adsorbed onto the biomaterial. Here, we have explored the molecular forces by which the S. epidermidis CWA protein serine-aspartate repeat protein F (SdrF) binds to type I collagen, by means of advanced atomic force microscopy (AFM) techniques. Using single-cell force spectroscopy, we found that SdrF mediates bacterial adhesion to collagen-coated substrates through both weak and strong bonds. Single-molecule force spectroscopy demonstrated that these bonds involve the A and B regions of SdrF, thus revealing that the protein is capable of dual ligand-binding activity. Both weak and strong bonds showed high dissociation rates, indicating they are much less stable than those formed by the well-characterized 'dock, lock and latch' mechanism. Collectively, our results show that CWA proteins can bind to ligands by novel mechanisms. We anticipate that AFM will greatly contribute to the identification of novel binding partners and binding mechanisms in staphylococcal CWA proteins. PMID:26481199

  4. Fibril Fragmentation Enhances Amyloid Cytotoxicity*♦

    PubMed Central

    Xue, Wei-Feng; Hellewell, Andrew L.; Gosal, Walraj S.; Homans, Steve W.; Hewitt, Eric W.; Radford, Sheena E.

    2009-01-01

    Fibrils associated with amyloid disease are molecular assemblies of key biological importance, yet how cells respond to the presence of amyloid remains unclear. Cellular responses may not only depend on the chemical composition or molecular properties of the amyloid fibrils, but their physical attributes such as length, width, or surface area may also play important roles. Here, we report a systematic investigation of the effect of fragmentation on the structural and biological properties of amyloid fibrils. In addition to the expected relationship between fragmentation and the ability to seed, we show a striking finding that fibril length correlates with the ability to disrupt membranes and to reduce cell viability. Thus, despite otherwise unchanged molecular architecture, shorter fibrillar samples show enhanced cytotoxic potential than their longer counterparts. The results highlight the importance of fibril length in amyloid disease, with fragmentation not only providing a mechanism by which fibril load can be rapidly increased but also creating fibrillar species of different dimensions that can endow new or enhanced biological properties such as amyloid cytotoxicity. PMID:19808677

  5. Sodium hypochlorite alterations of dentin and dentin collagen

    NASA Astrophysics Data System (ADS)

    Marshall, G. W.; Yücel, N.; Balooch, M.; Kinney, J. H.; Habelitz, S.; Marshall, S. J.

    2001-10-01

    NaOCl aq is used as a cleansing and non-specific deproteinizing agent in endodontic treatment, as a component of new chemomechanical caries treatment, and is under study for its alterations of dentin bonding characteristics. We sought to determine the microstructural and nanomechanical changes with such treatments and to test if NaOCl aq removed dentin collagen without microstructural or nanomechanical alteration of underlying mineralized dentin. Polished human dentin disks were prepared with a double reference technique that allowed changes to be determined following 10% citric acid etching for 15 s and subsequent treatment of the etched and unetched portions of the sample with 6.5% NaOCl aq, using atomic force microscopy (AFM) (Nanoscope III, Digital Instruments, Santa Barbara, CA). Images and measurements were made at intervals up to 1800 s. A Triboscope (Hysitron, Minneapolis, MN) on the AFM was used to measure nanohardness and the reduced elastic modulus. The double reference method allowed measurements immediately following etching and at intervals during deproteinization. Etching caused deep peritubular dentin removal and a small depth change of hydrated intertubular dentin as mineral was removed and left a remnant collagen matrix. NaOCl aq removed collagen over time, during which individual fibrils could be resolved; the underlying mineralized dentin was left with a unique porous surface containing numerous channels that are not normally observed in etched or fractured dentin. This could provide an attractive bonding substrate because of the increased surface area and high mineral content, if toughness is not reduced too much. Nanomechanical measurements showed that the reduced elastic modulus and hardness were 75% of original values after removal of the exposed collagen. Current dentin bonding systems rely on hybrid layer formation in which hydrophilic primers/polymers penetrate the opened collagen matrix exposed by etching. However some research suggests

  6. Collagen type IX from human cartilage: a structural profile of intermolecular cross-linking sites.

    PubMed Central

    Diab, M; Wu, J J; Eyre, D R

    1996-01-01

    Type IX collagen, a quantitatively minor collagenous component of cartilage, is known to be associated with and covalently cross-linked to type II collagen fibrils in chick and bovine cartilage. Type IX collagen molecules have also been shown to form covalent cross-links with each other in bovine cartilage. In the present study we demonstrate by structural analysis and location of cross-linking sites that, in human cartilage, type IX collagen is covalently cross-linked to type II collagen and to other molecules of type IX collagen. We also present evidence that, if the proteoglycan form of type IX collagen is present in human cartilage, it can only be a minor component of the matrix, similar to findings with bovine cartilage. PMID:8660302

  7. In vitro phagocytosis of exogenous collagen by fibroblasts from the periodontal ligament: an electron microscopic study.

    PubMed Central

    Svoboda, E L; Brunette, D M; Melcher, A H

    1979-01-01

    There have been numerous electron microscopic reports of apparent phagocytosis of collagen by fibroblasts and other cells in vivo. We have developed an in vitro system which, to the best of our knowledge, will permit for the first time the study of regulatory mechanisms governing phagocytosis and digestion of collagen fibres. Cells were cultured from explants of monkey periodontal ligament, subcultured, and grown to confluence in alpha-MEM plus 15% fetal calf serum plus antibiotics. The confluent cells were then cultured together with minced rat tail tendon collagen in alpha-MEM lacking proline, lysine, glycine and fetal calf serum for up to 7 days, after which they were processed for electron microscopy. Intracellular collagen profiles could be seen in cultured cells that were associated with exogenous collagen fibrils as early as 24 hours after addition of the collagen. Through electron microscopic examination of serial sections of the culture, we have demonstrated: (1) that fibroblasts can phagocytose collagen; (2) that the observed intracellular collagen is not the result of aggregation of endogenous synthesized collagen; (3) that it is not possible to base a decision as to whether a collagen fibril has been phagocytosed in whole or in part by the type of vesicle with which it is associated; (4) that cleavage of collagen into small pieces may not be a necessary prelude to its phagocytosis. Images Fig. 1 Fig. 2 Fig. 4 (cont.) Fig. 4 Fig. 6 (cont.) Fig. 6 Fig. 7 Fig. 8 Fig. 9 PMID:108237

  8. Controlling collagen fiber microstructure in three-dimensional hydrogels using ultrasound

    PubMed Central

    Garvin, Kelley A.; VanderBurgh, Jacob; Hocking, Denise C.; Dalecki, Diane

    2013-01-01

    Type I collagen is the primary fibrillar component of the extracellular matrix, and functional properties of collagen arise from variations in fiber structure. This study investigated the ability of ultrasound to control collagen microstructure during hydrogel fabrication. Under appropriate conditions, ultrasound exposure of type I collagen during polymerization altered fiber microstructure. Scanning electron microscopy and second-harmonic generation microscopy revealed decreased collagen fiber diameters in response to ultrasound compared to sham-exposed samples. Results of mechanistic investigations were consistent with a thermal mechanism for the effects of ultrasound on collagen fiber structure. To control collagen microstructure site-specifically, a high frequency, 8.3-MHz, ultrasound beam was directed within the center of a large collagen sample producing dense networks of short, thin collagen fibrils within the central core of the gel and longer, thicker fibers outside the beam area. Fibroblasts seeded onto these gels migrated rapidly into small, circularly arranged aggregates only within the beam area, and clustered fibroblasts remodeled the central, ultrasound-exposed collagen fibrils into dense sheets. These investigations demonstrate the capability of ultrasound to spatially pattern various collagen microstructures within an engineered tissue noninvasively, thus enhancing the level of complexity of extracellular matrix microenvironments and cellular functions achievable within three-dimensional engineered tissues. PMID:23927189

  9. Collagen Homeostasis and Metabolism.

    PubMed

    Magnusson, S Peter; Heinemeier, Katja M; Kjaer, Michael

    2016-01-01

    The musculoskeletal system and its collagen rich tissue is important for ensuring architecture of skeletal muscle, energy storage in tendon and ligaments, joint surface protection, and for ensuring the transfer of muscular forces into resulting limb movement. Structure of tendon is stable and the metabolic activity is low, but mechanical loading and subsequent mechanotransduction and molecular anabolic signaling can result in some adaptation of the tendon especially during youth and adolescence. Within short time, tendon will get stiffer with training and lack of mechanical tissue loading through inactivity or immobilization of the human body will conversely result in a dramatic loss in tendon stiffness and collagen synthesis. 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. Adaptive responses may vary along the tendon, and differ between mid-substance and insertional areas of the tendon. PMID:27535245

  10. Type V Collagen in Health, Disease, and Fibrosis.

    PubMed

    Mak, Ki M; Png, Chien Yi M; Lee, Danielle J

    2016-05-01

    Type V collagen (COLV) is a regulatory fibril-forming collagen. It has at least three different molecular isoforms-α1(V)2 α2(V), α1(V)3, and α1(V)α2(V)α3(V)-formed by combinations of three different polypeptide α chains-α1(V), α2(V), and α3(V). COL V is a relatively minor collagen of the extracellular matrix (ECM). Morphologically, COLV occurs as heterotypic fibrils with type I collagen (COLI), microfilaments, or 12-nm-thick fibrils. COLV is synthesized in various mesenchymal cells and its gene expression is modulated by TGF-β and growth factors. While resistant to digestion by interstitial collagenases, native and denatured COLV are degraded by metalloproteinases and gelatinases, thereby promoting ECM remodeling. COLV interacts with matrix collagens and structural proteins, conferring structural integrity to tissue scaffolds. It binds matrix macromolecules, modulating cellular behavior, and functions. COLV co-assembles with COLI into heterotypic fibrils in the cornea and skin dermis, acting as a dominant regulator of collagen fibrillogenesis. COLV deficiency is associated with loss of corneal transparency and classic Ehlers-Danlos syndrome, while COLV overexpression is found in cancer, granulation tissue, inflammation, atherosclerosis, and fibrosis of lungs, skin, kidneys, adipose tissue, and liver. COLV isoform containing the α3(V) chain is involved in mediating pancreatic islet cell functions. In the liver, COLV is a minor but regular component of the ECM. Increases in COLV are associated with both early and advanced hepatic fibrosis. The neoepitopes of COLV have been shown to be a useful noninvasive serum biomarker for assessing fibrotic progression and resolution in experimental hepatic fibrosis. COLV is multifunctional in health, disease, and fibrosis. PMID:26910848

  11. Atrial fibrillation ablation.

    PubMed

    Pappone, Carlo; Santinelli, Vincenzo

    2012-06-01

    Atrial fibrillation is the commonest cardiac arrhythmia, with significant morbidity related to symptoms, heart failure, and thromboembolism, which is associated with excess mortality. Over the past 10 years, many centers worldwide have reported high success rates and few complications after a single ablation procedure in patients with paroxysmal atrial fibrillation. Recent studies indicate a short-term and long-term superiority of catheter ablation as compared with conventional antiarrhythmic drug therapy in terms of arrhythmia recurrence, quality of life, and arrhythmia progression. As a result, catheter ablation is evolving to a front-line therapy in many patients with atrial fibrillation. However, in patients with persistent long-standing atrial fibrillation catheter ablation strategy is more complex and time-consuming, frequently requiring repeat procedures to achieve success rates as high as in paroxysmal atrial fibrillation. In the near future, however, with growing experience and evolving technology, catheter ablation of atrial fibrillation may be extended also to patients with long-standing atrial fibrillation. PMID:22541284

  12. Development of collagen-hydroxyapatite nanostructured composites via a calcium phosphate precursor mechanism

    NASA Astrophysics Data System (ADS)

    Jee, Sang Soo

    Bone is an interpenetrating inorganic/organic composite that consists of mineralized collagen fibrils, which is hierarchically organized into various structures. The structure of mineralized collagen fibril, in which nano-crystals of hydroxyapatite are embedded within the collagen fibrils, provides remarkable mechanical and bio-resorptive properties. Therefore, there have been many attempts to produce collagen-hydroxyapatite composites having a bone-like structure. However, duplication of even the most fundamental level of bone structure has not been easily achieved by conventional nucleation and growth techniques, which are based on the most widely accepted hypothesis of bone mineralization. In nature, the collagen fibril is mineralized via intrafibrillar mineralization, which produces preferentially oriented hydroxyapatite nano-crystals occupying the interstices in collagen fibrils. Our group has demonstrated that intrafibrillar mineralization can be achieved by using a new method based on the Polymer-Induced Liquid-Precursor (PILP) mineralization process. In the PILP process, a poly-anionic additive can produce an amorphous calcium phosphate precursor which enables us to achieve intrafibrillar mineralization of collagen. It is thought that the precursor is pulled into the interstices of the collagen fibrils via capillary forces, and upon solidification and crystallization of the precursor produces an interpenetrating composite with the nanostructured architecture of bone. In this dissertation, to demonstrate the effectiveness of the PILP process on the intrafibrillar mineralization of collagen fibril, various collagen scaffolds, such as turkey tendon, bovine tendon and synthetic collagen sponge, were mineralized by the PILP process. Various poly-aspartates with different molecular weight were also used for the optimization of the PILP process for the mineralization of the collagen scaffolds. With the systematic researches, we discovered that the molecular weight

  13. Cell Alignment Driven by Mechanically Induced Collagen Fiber Alignment in Collagen/Alginate Coatings.

    PubMed

    Chaubaroux, Christophe; Perrin-Schmitt, Fabienne; Senger, Bernard; Vidal, Loïc; Voegel, Jean-Claude; Schaaf, Pierre; Haikel, Youssef; Boulmedais, Fouzia; Lavalle, Philippe; Hemmerlé, Joseph

    2015-09-01

    For many years it has been a major challenge to regenerate damaged tissues using synthetic or natural materials. To favor the healing processes after tendon, cornea, muscle, or brain injuries, aligned collagen-based architectures are of utmost interest. In this study, we define a novel aligned coating based on a collagen/alginate (COL/ALG) multilayer film. The coating exhibiting a nanofibrillar structure is cross-linked with genipin for stability in physiological conditions. By stretching COL/ALG-coated polydimethylsiloxane substrates, we developed a versatile method to align the collagen fibrils of the polymeric coating. Assays on cell morphology and alignment were performed to investigate the properties of these films. Microscopic assessments revealed that cells align with the stretched collagen fibrils of the coating. The degree of alignment is tuned by the stretching rate (i.e., the strain) of the COL/ALG-coated elastic substrate. Such coatings are of great interest for strategies that require aligned nanofibrillar biological material as a substrate for tissue engineering. PMID:25658028

  14. Comparative SEM study on the effect of different demineralization methods with tetracycline HCl on healthy root surfaces.

    PubMed

    Işik, G; Ince, S; Sağlam, F; Onan, U

    1997-09-01

    Periodontal regeneration through the use of root demineralization received a lot of interest in periodontology. Topical application of acid to dentin surfaces produced a zone of demineralization, exposing dentin collagen fibrils and opening dentin tubules. In this study, the in vitro effects of different tetracycline HCl application techniques were investigated. According to the results of this SEM study, it may be desirable to apply tetracycline HCl using burnishing technique to expose maximum intertubular fibrils and for the tubular openings. However, this technique should be studied when placed in an in vivo system. PMID:9378828

  15. Kinetics of protein fibrillation controlled by fibril elongation.

    PubMed

    Kashchiev, Dimo

    2014-09-01

    Numerous proteins have the ability to assemble into fibrillar aggregates which are of great interest, because they feature in scores of human diseases and many technological products. In the present work, we analyze the kinetics of protein fibrillation when the process is governed solely by elongation of initially appeared fibrils in the protein solution. We derive exact expressions for the time dependences of the fibrillation degree, the concentration of monomeric protein in the solution, and the average fibril size. Furthermore, we present formulas for the initial fibrillation rate and the half-fibrillation time in terms of experimentally controllable quantities. The results obtained provide a mechanistic insight into the kinetics of protein fibrillation mediated by fibril elongation. We confront theory with experiment and find that it allows a good description of available experimental data for fibrillation of the Alzheimer's disease-associated protein Aβ(1-40) and the yeast prion protein Sup35. PMID:24753319

  16. Aromatic Interactions Promote Self-association of Collagen Triple-helical Peptides to Higher Order Structures

    PubMed Central

    Kar, Karunakar; Ibrar, Sajjad; Nanda, Vikas; Getz, Todd M.; Kunapuli, Satya P.; Brodsky, Barbara

    2009-01-01

    Aromatic residues are relatively rare within the collagen triple-helix, but they appear to play a specialized role in higher order structure and function. The role of aromatic amino acids in the self-assembly of triple-helical peptides was investigated in terms of the kinetics of self-association, the nature of aggregated species formed, and the ability of these species to activate platelet aggregation. The presence of aromatic residues on both ends of a type IV collagen model peptide is observed to greatly accelerate the kinetics of self-association, decreasing the lag time and leading to insoluble, well defined linear fibrils as well as small soluble aggregates. Both macroscopic visible aggregates and small multi-molecular complexes in solution are capable of inducing platelet aggregation through the glycoprotein VI receptor on platelets. Proline-aromatic CH⋯π interactions are often observed within globular proteins and in protein complexes, and examination of molecular packing in the crystal structure of the integrin binding collagen peptide shows Phe interacts with Pro/Hyp in a neighboring triple-helical molecule. An intermolecular interaction between aromatic amino acids and imino acids within the triple-helix is also supported by the observed inhibitory effect of isolated Phe amino acids on the self-association of (Pro-Hyp-Gly)10. Given the high fraction of Pro and Hyp residues on the surface of collagen molecules, it is likely that imino acid-aromatic CH⋯π interactions are important in formation of higher order structure. It is suggested that the catalysis of type I collagen fibrillogenesis by non-helical telopeptides is due to specific intermolecular CH⋯π interactions between aromatic residues in the telopeptides and Pro/Hyp residues within the triple-helix. PMID:19610672

  17. Three dimensional patient-specific collagen architecture modulates cartilage responses in the knee joint during gait.

    PubMed

    Räsänen, Lasse P; Mononen, Mika E; Lammentausta, Eveliina; Nieminen, Miika T; Jurvelin, Jukka S; Korhonen, Rami K

    2016-08-01

    Site-specific variation of collagen fibril orientations can affect cartilage stresses in knee joints. However, this has not been confirmed by 3-D analyses. Therefore, we present a novel method for evaluation of the effect of patient-specific collagen architecture on time-dependent mechanical responses of knee joint cartilage during gait. 3-D finite element (FE) models of a human knee joint were created with the collagen architectures obtained from T2 mapped MRI (patient-specific model) and from literature (literature model). The effect of accuracy of the implementation of collagen fibril architecture into the model was examined by using a submodel with denser FE mesh. Compared to the literature model, fibril strains and maximum principal stresses were reduced especially in the superficial/middle regions of medial tibial cartilage in the patient-specific model after the loading response of gait (up to -413 and -26%, respectively). Compared to the more coarsely meshed joint model, the patient-specific submodel demonstrated similar strain and stress distributions but increased values particularly in the superficial cartilage regions (especially stresses increased >60%). The results demonstrate that implementation of subject-specific collagen architecture of cartilage in 3-D modulates location- and time-dependent mechanical responses of human knee joint cartilage. Submodeling with more accurate implementation of collagen fibril architecture alters cartilage stresses particularly in the superficial/middle tissue. PMID:26714834

  18. What Is Atrial Fibrillation?

    MedlinePlus

    ... regular beat. Certain cells in your heart make electric signals that cause the heart to contract and ... read your ECG to find out if the electric signals are normal. In atrial fibrillation (AFib), the ...

  19. Tenascin-X deficiency mimics Ehlers-Danlos syndrome in mice through alteration of collagen deposition.

    PubMed

    Mao, Jau Ren; Taylor, Glen; Dean, Willow B; Wagner, Diane R; Afzal, Veena; Lotz, Jeffrey C; Rubin, Edward M; Bristow, James

    2002-04-01

    Tenascin-X is a large extracellular matrix protein of unknown function. Tenascin-X deficiency in humans is associated with Ehlers-Danlos syndrome, a generalized connective tissue disorder resulting from altered metabolism of the fibrillar collagens. Because TNXB is the first Ehlers-Danlos syndrome gene that does not encode a fibrillar collagen or collagen-modifying enzyme, we suggested that tenascin-X might regulate collagen synthesis or deposition. To test this hypothesis, we inactivated Tnxb in mice. Tnxb-/- mice showed progressive skin hyperextensibility, similar to individuals with Ehlers-Danlos syndrome. Biomechanical testing confirmed increased deformability and reduced tensile strength of their skin. The skin of Tnxb-/- mice was histologically normal, but its collagen content was significantly reduced. At the ultrastructural level, collagen fibrils of Tnxb-/- mice were of normal size and shape, but the density of fibrils in their skin was reduced, commensurate with the reduction in collagen content. Studies of cultured dermal fibroblasts showed that although synthesis of collagen I by Tnxb-/- and wildtype cells was similar, Tnxb-/- fibroblasts failed to deposit collagen I into cell-associated matrix. This study confirms a causative role for TNXB in human Ehlers-Danlos syndrome and suggests that tenascin-X is an essential regulator of collagen deposition by dermal fibroblasts. PMID:11925569

  20. Tenascin-x deficiency mimics ehlers-danlos syndrome in mice through alteration of collagen deposition

    SciTech Connect

    Mao, J.R.; Taylor, G.; Dean, W.B.; Wagner, D.R.; Afzal, V.; Lotz, J.C.; Rubin, E.M.; Bristow, J.

    2002-03-01

    Tenascin-X is a large extracellular matrix protein of unknown function1-3. Tenascin-X deficiency in humans is associated with Ehlers-Danlos syndrome4,5, a generalized connective tissue disorder resulting from altered metabolism of the fibrillar collagens6. Because TNXB is the first Ehlers-Danlos syndrome gene that does not encode a fibrillar collagen or collagen-modifying enzyme7-14, we suggested that tenascin-X might regulate collagen synthesis or deposition15. To test this hypothesis, we inactivated Tnxb in mice. Tnxb-/- mice showed progressive skin hyperextensibility, similar to individuals with Ehlers-Danlos syndrome. Biomechanical testing confirmed increased deformability and reduced tensile strength of their skin. The skin of Tnxb-/- mice was histologically normal, but its collagen content was significantly reduced. At the ultrastructural level, collagen fibrils of Tnxb-/- mice were of normal size and shape, but the density of fibrils in their skin was reduced, commensurate with the reduction in collagen content. Studies of cultured dermal fibroblasts showed that although synthesis of collagen I by Tnxb-/- and wildtype cells was similar, Tnxb-/- fibroblasts failed to deposit collagen I into cell-associated matrix. This study confirms a causative role for TNXB in human Ehlers-Danlos syndrome and suggests that tenascin-X is an essential regulator of collagen deposition by dermal fibroblasts.

  1. Structural response of human corneal and scleral tissues to collagen cross-linking treatment with riboflavin and ultraviolet A light.

    PubMed

    Choi, Samjin; Lee, Seung-Chan; Lee, Hui-Jae; Cheong, Youjin; Jung, Gyeong-Bok; Jin, Kyung-Hyun; Park, Hun-Kuk

    2013-09-01

    High success rates in clinical trials on keratoconic corneas suggest the possibility of efficient treatment against myopic progression. This study quantitatively investigated the in vitro ultrastructural effects of a photooxidative collagen cross-linking treatment with photosensitizer riboflavin and UVA light in human corneo-scleral collagen fibrils. A total of 30.8 × 2 mm corneo-scleral strips from donor tissue were sagittally dissected using a scalpel. The five analytic parameters namely fibril density, fibril area, corneo-scleral thickness, fibril diameter, and fibril arrangement were investigated before and after riboflavin-UVA-catalyzed collagen cross-linking treatment. Collagen cross-linking effects were measured at the corneo-scleral stroma and were based on clinical corneal cross-linking procedures. The structural response levels were assessed by histology, digital mechanical caliper measurement, scanning electron microscopy, and atomic force microscopy. Riboflavin-UVA-catalyzed collagen cross-linking treatment led to an increase in the area, density, and diameters of both corneal (110, 112, and 103 %) and scleral (133, 133, and 127 %) stromal collagens. It also led to increases in corneal (107 %) and scleral (105 %) thickness. Collagen cross-linking treatment through riboflavin-sensitized photoreaction may cause structural property changes in the collagen fibril network of the cornea and sclera due to stromal edema and interfibrillar spacing narrowing. These changes were particularly prominent in the sclera. This technique can be used to treat progressive keratoconus in the cornea as well as progressive myopia in the sclera. Long-term collagen cross-linking treatment of keratoconic and myopic progression dramatically improves weakened corneo-scleral tissues. PMID:23179311

  2. Evaluation of collagen immobilized to silicon plates by ion beam

    NASA Astrophysics Data System (ADS)

    Yokoyama, Y.; Kobayashi, T.; Iwaki, M.

    2006-01-01

    A study has been made of immobilization of collagen coated on the substrate by ion beam in order to elucidate the effects of ion bombardment on cell adhesion strength. Substrates used were silicon plates, on which 0.3% type-I collagen solution was coated using a spin coater. The collagen-coated silicon was bombarded with 50 keV He+ ions at doses from 1 × 1013 to 1 × 1015 ions/cm2 using a RIKEN TK-100 ion implanter. The collagen-immobilized specimens were mounted on a parallel-plate flow chamber to perform the collagen adhesion tests with a flowing shear stress. Morphological observations of collagen were performed by scanning transmission electron microscopy (STEM). The chemical condition of collagen was detected by X-ray photoelectron spectroscopy (XPS). The collagen layer in the non-bombarded specimen was about 20 nm in thickness. STEM micrographs showed that collagen layer has thinned due to contraction by ion bombardment as the dose increased. After the collagen adhesion test, collagen layer surface with the non-bombarded specimen was peeled off by shear stress. As the dose increased, the detachment of collagen was suppressed. Detachment of collagen was hardly observed for the dose of 1 × 1015 ions/cm2. The XPS results of collagen structures showed that ion bombardment generated new bonds between collagen molecules in the collagen layer. It is concluded that the increase of collagen adhesion at higher doses is due to the ion-beam immobilization of collagen molecules resulting from new bond generation by displaced atoms and excited atoms between collagen molecules in the collagen layer.

  3. Measurement of Elastic Modulus of Collagen Type I Single Fiber.

    PubMed

    Dutov, Pavel; Antipova, Olga; Varma, Sameer; Orgel, Joseph P R O; Schieber, Jay D

    2016-01-01

    Collagen fibers are the main components of the extra cellular matrix and the primary contributors to the mechanical properties of tissues. Here we report a novel approach to measure the longitudinal component of the elastic moduli of biological fibers under conditions close to those found in vivo and apply it to type I collagen from rat tail tendon. This approach combines optical tweezers, atomic force microscopy, and exploits Euler-Bernoulli elasticity theory for data analysis. This approach also avoids drying for measurements or visualization, since samples are freshly extracted. Importantly, strains are kept below 0.5%, which appear consistent with the linear elastic regime. We find, surprisingly, that the longitudinal elastic modulus of type I collagen cannot be represented by a single quantity but rather is a distribution that is broader than the uncertainty of our experimental technique. The longitudinal component of the single-fiber elastic modulus is between 100 MPa and 360 MPa for samples extracted from different rats and/or different parts of a single tail. Variations are also observed in the fibril-bundle/fibril diameter with an average of 325±40 nm. Since bending forces depend on the diameter to the fourth power, this variation in diameter is important for estimating the range of elastic moduli. The remaining variations in the modulus may be due to differences in composition of the fibril-bundles, or the extent of the proteoglycans constituting fibril-bundles, or that some single fibrils may be of fibril-bundle size. PMID:26800120

  4. Measurement of Elastic Modulus of Collagen Type I Single Fiber

    PubMed Central

    Dutov, Pavel; Antipova, Olga; Varma, Sameer; Orgel, Joseph P. R. O.; Schieber, Jay D.

    2016-01-01

    Collagen fibers are the main components of the extra cellular matrix and the primary contributors to the mechanical properties of tissues. Here we report a novel approach to measure the longitudinal component of the elastic moduli of biological fibers under conditions close to those found in vivo and apply it to type I collagen from rat tail tendon. This approach combines optical tweezers, atomic force microscopy, and exploits Euler-Bernoulli elasticity theory for data analysis. This approach also avoids drying for measurements or visualization, since samples are freshly extracted. Importantly, strains are kept below 0.5%, which appear consistent with the linear elastic regime. We find, surprisingly, that the longitudinal elastic modulus of type I collagen cannot be represented by a single quantity but rather is a distribution that is broader than the uncertainty of our experimental technique. The longitudinal component of the single-fiber elastic modulus is between 100 MPa and 360 MPa for samples extracted from different rats and/or different parts of a single tail. Variations are also observed in the fibril-bundle / fibril diameter with an average of 325±40 nm. Since bending forces depend on the diameter to the fourth power, this variation in diameter is important for estimating the range of elastic moduli. The remaining variations in the modulus may be due to differences in composition of the fibril-bundles, or the extent of the proteoglycans constituting fibril-bundles, or that some single fibrils may be of fibril-bundle size. PMID:26800120

  5. Effect of alcohols and neutral salt on the thermal stability of soluble and precipitated acid-soluble collagen

    PubMed Central

    Russell, Allan E.

    1973-01-01

    The effects of mono- and poly-hydric alcohols in the presence of KCl on the intrinsic stability of collagen molecules in dilute acid solution were compared with corresponding solvent and salt effects on the increased stability of the aggregated molecules in salt-precipitated fibrils. Salt addition decreased solubility and increased the thermal stability of fibrils, but progressively decreased the stability of collagen molecules in solution. In contrast, the alcohols enhanced solubility and decreased fibril stability, the effects increasing with solvent hydrocarbon chain length and with decreasing hydroxyl/methylene-group ratio. Molar destabilization of dissolved collagen by alcohols was lower than for fibrils, and at low salt concentration, both ethylene glycol and glycerol were structural stabilizers. Electron-micrograph studies indicated that salt-precipitated fibrils tended to adopt the native aggregation mode, and qualitatively similar solvent effects were observed in insoluble collagens. Implications of the experimental findings are discussed in terms of a model in which electrostatic and apolar interactions mainly govern the excess of stability in collagen fibrils whereas intrinsic stability of single molecules is a function of polar interactions and polypeptide-chain rigidity. PMID:4737319

  6. Microstructural modeling of collagen network mechanics and interactions with the proteoglycan gel in articular cartilage.

    PubMed

    Quinn, T M; Morel, V

    2007-01-01

    Cartilage matrix mechanical function is largely determined by interactions between the collagen fibrillar network and the proteoglycan gel. Although the molecular physics of these matrix constituents have been characterized and modern imaging methods are capable of localized measurement of molecular densities and orientation distributions, theoretical tools for using this information for prediction of cartilage mechanical behavior are lacking. We introduce a means to model collagen network contributions to cartilage mechanics based upon accessible microstructural information (fibril density and orientation distributions) and which self-consistently follows changes in microstructural geometry with matrix deformations. The interplay between the molecular physics of the collagen network and the proteoglycan gel is scaled up to determine matrix material properties, with features such as collagen fibril pre-stress in free-swelling cartilage emerging naturally and without introduction of ad hoc parameters. Methods are developed for theoretical treatment of the collagen network as a continuum-like distribution of fibrils, such that mechanical analysis of the network may be simplified by consideration of the spherical harmonic components of functions of the fibril orientation, strain, and stress distributions. Expressions for the collagen network contributions to matrix stress and stiffness tensors are derived, illustrating that only spherical harmonic components of orders 0 and 2 contribute to the stress, while orders 0, 2, and 4 contribute to the stiffness. Depth- and compression-dependent equilibrium mechanical properties of cartilage matrix are modeled, and advantages of the approach are illustrated by exploration of orientation and strain distributions of collagen fibrils in compressed cartilage. Results highlight collagen-proteoglycan interactions, especially for very small physiological strains where experimental data are relatively sparse. These methods for

  7. Cross-linking and the molecular packing of corneal collagen

    NASA Technical Reports Server (NTRS)

    Yamauchi, M.; Chandler, G. S.; Tanzawa, H.; Katz, E. P.

    1996-01-01

    We have quantitatively characterized, for the first time, the cross-linking in bovine cornea collagen as a function of age. The major iminium reducible cross-links were dehydro-hydroxylysinonorleucine (deH-HLNL) and dehydro-histidinohydroxymerodesmosine (deH-HHMD). The former rapidly diminished after birth; however, the latter persisted in mature animals at a level of 0.3 - 0.4 moles/mole of collagen. A nonreducible cross-link, histidinohydroxylysinonorleucine (HHL), previously found only in skin, was also found to be a major mature cross-link in cornea. The presence of HHL indicates that cornea fibrils have a molecular packing similar to skin collagen. However, like deH-HHMD, the HHL content in corneal fibrils only reaches a maximum value with time about half that of skin. These data suggest that the corneal fibrils are comprised of discrete filaments that are internally stabilized by HHL and deH-HHMD cross-links. This pattern of intermolecular cross-linking would facilitate the special collagen swelling property required for corneal transparency.

  8. Ultrastructural and tissue-culture studies on the role of fibronectin, collagen and glycosaminoglycans in the migration of neural crest cells in the fowl embryo.

    PubMed

    Newgreen, D F; Gibbins, I L; Sauter, J; Wallenfels, B; Wütz, R

    1982-01-01

    The initial migration of neural crest (NC) cells into cell-free space was studied by transmission electron microscopy at trunk levels of fowl embryos, some of which were fixed in the presence of ruthenium red. Migrating NC cells occurred in zones which contained fewer ruthenium-red stained 15-40nm diameter granules than other regions. The ruthenium-red stained granules were linked by similarly stained thin (greater than 3nm diameter) microfibrils. The granules resemble proteoglycan and the microfibrils may be hyaluronate. NC cells contacted thicker (greater than 10 nm diameter) fibrils and interstitial bodies, which did not require ruthenium red for visualization. Cytoplasmic microfilaments were sometimes aligned at the point of contact with the extracellular fibrils, which may be fibronectin and collagen. Phase-contrast time-lapse videotaping and scanning electron microscopy showed that NC cells of the fowl embryo in vitro migrated earlier and more extensively on glass coated with fibronectin-rich fibrous material and adsorbed fibronectin molecules than on glass coated with collagen type I (fibres and adsorbed molecules). NC cells became completely enmeshed in fibronectin-rich fibres, but generally remained on the surface of collagen-fibre gels. When given a choice, NC cells strongly preferred fibronectin coatings to plain glass, and plain glass to dried collagen gels. NC cells showed a slight preference for plain glass over glass to which collagen was adsorbed. Addition to the culture medium of hyaluronate (initial conc. 20 mg/ml), chondroitin (5 mg/ml) and fully sulphated chondroitin sulphate and dermatan sulphate (up to 10 mg/ml) did not drastically alter NC cell migration on fibronectin-rich fibrous substrates. PMID:7034954

  9. Enhancing collagen stability through nanostructures containing chromium(III) oxide.

    PubMed

    Sangeetha, Selvam; Ramamoorthy, Usha; Sreeram, Kalarical Janardhanan; Nair, Balachandran Unni

    2012-12-01

    Stabilization of collagen for various applications employs chemicals such as aldehydes, metal ions, polyphenols, etc. Stability against enzymatic, thermal and mechanical degradation is required for a range of biomedical applications. The premise of this research is to explore the use of nanoparticles with suitable functionalization/encapsulation to crosslink with collagen, such that the three dimensional architecture had the desired stability. Collagen solution prepared as per standard protocols is treated with chromium(III) oxide nanoparticules encapsulated within a polymeric matrix (polystyrene-block-polyacrylic acid copolymer). Selectivity towards encapsulation was ensured by the reaction in dimethyl sulfoxide, where the PS groups popped out and encapsulated the Cr(2)O(3). Subsequently when immersed in aqueous solution, PAA units popped up to react with functional groups of collagen. The interaction with collagen was monitored through techniques such as CD, FTIR, viscosity measurements, stress analysis. CD studies and FTIR showed no degradation of collagen. Thermal stability was enhanced upon interaction of nanostructures with collagen. Self-assembly of collagen was delayed but not inhibited, indicating a compete binding of the metal oxide encapsulated polymer to collagen. Metal oxide nanoparticles encapsulated within a polymeric matrix could provide thermal and mechanical stability to collagen. The formed fibrils of collagen could serve as ideal material for various smart applications such as slow/sustained drug release. The study is also relevant to the leather industry in that the nanostructures can diffuse through the highly networked collagen fibre bundles in skin matrix easily, thus overcoming the rate limiting step of diffusion. PMID:22766281

  10. Collagen deposition and mechanical strength of colon anastomoses and skin incisional wounds of rats.

    PubMed

    Oxlund, H; Christensen, H; Seyer-Hansen, M; Andreassen, T T

    1996-11-01

    The mechanical strength development of healing wounds depends on the formation of collagen fibrils bridging the wound cleft. A considerable deposition, degradation, and remodeling of these fibrils takes place influencing the mechanical strength of the healing wounds. A method for studies of wound collagen metabolism in vivo is delineated, enabling determination of collagen deposition per hour in rat colon anastomoses and skin incisional wounds. Labeled proline was incorporated into wound collagen with a flooding dose of unlabeled proline, reducing errors introduced by proline recycling and proline de novo synthesis. The mechanical strength was determined by a materials testing machine. In both colon anastomoses and skin wounds a substantial increase in collagen deposition was observed at Day 2, reached a maximum at Day 6, and was still relatively high at Day 12 during the remodeling of collagen fibers in the wound cleft. The collagen deposition in colon anastomoses at Day 6, however, was 10-fold higher compared with that of the skin incisional wounds. The time course of the collagen deposition was much alike in colon anastomoses and skin incisional wounds reaching a maximum at Day 6. The mechanical strength of these two rather different types of wounds was increased correspondingly and to the same level during the 1st week of healing. The measurements of collagen deposition, collagen content, and biomechanical strength indicated a substantial turnover of newly synthesized and deposited collagen during the early phases of wound healing. On the basis of this, it seems obvious that even small disturbances to the balance between collagen synthesis, deposition, collagen cross-linking, and collagen degradation/remodeling may result in defective wound healing. PMID:8954827

  11. Human recombinant type I collagen produced in plants.

    PubMed

    Shoseyov, Oded; Posen, Yehudit; Grynspan, Frida

    2013-07-01

    As a central element of the extracellular matrix, collagen is intimately involved in tissue development, remodeling, and repair and confers high tensile strength to tissues. Numerous medical applications, particularly, wound healing, cell therapy, bone reconstruction, and cosmetic technologies, rely on its supportive and healing qualities. Its synthesis and assembly require a multitude of genes and post-translational modifications, where even minor deviations can be deleterious or even fatal. Historically, collagen was always extracted from animal and human cadaver sources, but bare risk of contamination and allergenicity and was subjected to harsh purification conditions resulting in irreversible modifications impeding its biofunctionality. In parallel, the highly complex and stringent post-translational processing of collagen, prerequisite of its viability and proper functioning, sets significant limitations on recombinant expression systems. A tobacco plant expression platform has been recruited to effectively express human collagen, along with three modifying enzymes, critical to collagen maturation. The plant extracted recombinant human collagen type I forms thermally stable helical structures, fibrillates, and demonstrates bioactivity resembling that of native collagen. Deployment of the highly versatile plant-based biofactory can be leveraged toward mass, rapid, and low-cost production of a wide variety of recombinant proteins. As in the case of collagen, proper planning can bypass plant-related limitations, to yield products structurally and functionally identical to their native counterparts. PMID:23252967

  12. The role of collagen in bone apatite formation in the presence of hydroxyapatite nucleation inhibitors

    PubMed Central

    Nudelman, Fabio; Pieterse, Koen; George, Anne; Bomans, Paul H. H.; Friedrich, Heiner; Brylka, Laura J.; Hilbers, Peter A. J.; de With, Gijsbertus; Sommerdijk, Nico A. J. M.

    2011-01-01

    Bone is a composite material, in which collagen fibrils form a scaffold for a highly organized arrangement of uniaxially oriented apatite crystals1,2. In the periodic 67 nm cross-striated pattern of the collagen fibril3–5, the less dense 40-nm-long gap zone has been implicated as the place where apatite crystals nucleate from an amorphous phase, and subsequently grow6–9. This process is believed to be directed by highly acidic non-collagenous proteins6,7,9–11; however, the role of the collagen matrix12–14 during bone apatite mineralization remains unknown. Here, combining nanometre-scale resolution cryogenic transmission electron microscopy and cryogenic electron tomography15 with molecular modelling, we show that collagen functions in synergy with inhibitors of hydroxyapatite nucleation to actively control mineralization. The positive net charge close to the C-terminal end of the collagen molecules promotes the infiltration of the fibrils with amorphous calcium phosphate (ACP). Furthermore, the clusters of charged amino acids, both in gap and overlap regions, form nucleation sites controlling the conversion of ACP into a parallel array of oriented apatite crystals. We developed a model describing the mechanisms through which the structure, supramolecular assembly and charge distribution of collagen can control mineralization in the presence of inhibitors of hydroxyapatite nucleation. PMID:20972429

  13. Thermal Denaturation Studies of Collagen by Microthermal Analysis and Atomic Force Microscopy

    PubMed Central

    Bozec, Laurent; Odlyha, Marianne

    2011-01-01

    The structural properties of collagen have been the subject of numerous studies over past decades, but with the arrival of new technologies, such as the atomic force microscope and related techniques, a new era of research has emerged. Using microthermal analysis, it is now possible to image samples as well as performing localized thermal measurements without damaging or destroying the sample itself. This technique was successfully applied to characterize the thermal response between native collagen fibrils and their denatured form, gelatin. Thermal transitions identified at (150 ± 10)°C and (220 ± 10)°C can be related to the process of gelatinization of the collagen fibrils, whereas at higher temperatures, both the gelatin and collagen samples underwent two-stage transitions with a common initial degradation temperature at (300 ± 10)°C and a secondary degradation temperature of (340 ± 10)°C for the collagen and of (420 ± 10)°C for the gelatin, respectively. The broadening and shift in the secondary degradation temperature was linked to the spread of thermal degradation within the gelatin and collagen fibrils matrix further away from the point of contact between probe and sample. Finally, similar measurements were performed inside a bone resorption lacuna, suggesting that microthermal analysis is a viable technique for investigating the thermomechanical response of collagen for in situ samples that would be, otherwise, too challenging or not possible using bulk techniques. PMID:21723833

  14. Strain stiffening in collagen I networks.

    PubMed

    Motte, Stéphanie; Kaufman, Laura J

    2013-01-01

    Biopolymer gels exhibit strain stiffening that is generally not seen in synthetic gels. Here, we investigate the strain-stiffening behavior in collagen I gels that demonstrate elasticity derived from a variety of sources including crosslinking through telopeptides, bundling through low-temperature gelation, and exogenous crosslinking with genipin. In all cases, it is found that these gels exhibit strain stiffening; in general, onset of strain stiffening occurs earlier, yield strain is lower, and degree of strain stiffening is smaller in higher concentration gels and in those displaying thick fibril bundles. Recovery after exposure to high strains is substantial and similar in all gels, suggesting that much of the stiffening comes from reversible network deformations. A key finding of this study is that collagen I gels of identical storage and loss moduli may display different nonlinear responses and different capacities to recover from high strain. PMID:23097228

  15. Targeted Deletion of Collagen V in Tendons and Ligaments Results in a Classic Ehlers-Danlos Syndrome Joint Phenotype

    PubMed Central

    Sun, Mei; Connizzo, Brianne K.; Adams, Sheila M.; Freedman, Benjamin R.; Wenstrup, Richard J.; Soslowsky, Louis J.; Birk, David E.

    2016-01-01

    Collagen V mutations underlie classic Ehlers-Danlos syndrome, and joint hypermobility is an important clinical manifestation. We define the function of collagen V in tendons and ligaments, as well as the role of alterations in collagen V expression in the pathobiology in classic Ehlers-Danlos syndrome. A conditional Col5a1flox/flox mouse model was bred with Scleraxis-Cre mice to create a targeted tendon and ligament Col5a1-null mouse model, Col5a1Δten/Δten. Targeting was specific, resulting in collagen V–null tendons and ligaments. Col5a1Δten/Δten mice demonstrated decreased body size, grip weakness, abnormal gait, joint laxity, and early-onset osteoarthritis. These gross changes were associated with abnormal fiber organization, as well as altered collagen fibril structure with increased fibril diameters and decreased fibril number that was more severe in a major joint stabilizing ligament, the anterior cruciate ligament (ACL), than in the flexor digitorum longus tendon. The ACL also had a higher collagen V content than did the flexor digitorum longus tendon. The collagen V–null ACL and flexor digitorum longus tendon both had significant alterations in mechanical properties, with ACL exhibiting more severe changes. The data demonstrate critical differential regulatory roles for collagen V in tendon and ligament structure and function and suggest that collagen V regulatory dysfunction is associated with an abnormal joint phenotype, similar to the hypermobility phenotype in classic Ehlers-Danlos syndrome. PMID:25797646

  16. Targeted deletion of collagen V in tendons and ligaments results in a classic Ehlers-Danlos syndrome joint phenotype.

    PubMed

    Sun, Mei; Connizzo, Brianne K; Adams, Sheila M; Freedman, Benjamin R; Wenstrup, Richard J; Soslowsky, Louis J; Birk, David E

    2015-05-01

    Collagen V mutations underlie classic Ehlers-Danlos syndrome, and joint hypermobility is an important clinical manifestation. We define the function of collagen V in tendons and ligaments, as well as the role of alterations in collagen V expression in the pathobiology in classic Ehlers-Danlos syndrome. A conditional Col5a1(flox/flox) mouse model was bred with Scleraxis-Cre mice to create a targeted tendon and ligament Col5a1-null mouse model, Col5a1(Δten/Δten). Targeting was specific, resulting in collagen V-null tendons and ligaments. Col5a1(Δten/Δten) mice demonstrated decreased body size, grip weakness, abnormal gait, joint laxity, and early-onset osteoarthritis. These gross changes were associated with abnormal fiber organization, as well as altered collagen fibril structure with increased fibril diameters and decreased fibril number that was more severe in a major joint stabilizing ligament, the anterior cruciate ligament (ACL), than in the flexor digitorum longus tendon. The ACL also had a higher collagen V content than did the flexor digitorum longus tendon. The collagen V-null ACL and flexor digitorum longus tendon both had significant alterations in mechanical properties, with ACL exhibiting more severe changes. The data demonstrate critical differential regulatory roles for collagen V in tendon and ligament structure and function and suggest that collagen V regulatory dysfunction is associated with an abnormal joint phenotype, similar to the hypermobility phenotype in classic Ehlers-Danlos syndrome. PMID:25797646

  17. Light Chain Amyloid Fibrils Cause Metabolic Dysfunction in Human Cardiomyocytes

    SciTech Connect

    McWilliams-Koeppen, Helen P.; Foster, James S.; Hackenbrack, Nicole; Ramirez-Alvarado, Marina; Donohoe, Dallas; Williams, Angela; Macy, Sallie; Wooliver, Craig; Wortham, Dale; Morrell-Falvey, Jennifer; Foster, Carmen M.; Kennel, Stephen J.; Wall, Jonathan S.

    2015-09-22

    Light chain (AL) amyloidosis is the most common form of systemic amyloid disease, and cardiomyopathy is a dire consequence, resulting in an extremely poor prognosis. AL is characterized by the production of monoclonal free light chains that deposit as amyloid fibrils principally in the heart, liver, and kidneys causing organ dysfunction. We have studied the effects of amyloid fibrils, produced from recombinant λ6 light chain variable domains, on metabolic activity of human cardiomyocytes. The data indicate that fibrils at 0.1 μM, but not monomer, significantly decrease the enzymatic activity of cellular NAD(P)H-dependent oxidoreductase, without causing significant cell death. The presence of amyloid fibrils did not affect ATP levels; however, oxygen consumption was increased and reactive oxygen species were detected. Confocal fluorescence microscopy showed that fibrils bound to and remained at the cell surface with little fibril internalization. Ultimately, these data indicate that AL amyloid fibrils severely impair cardiomyocyte metabolism in a dose dependent manner. These data suggest that effective therapeutic intervention for these patients should include methods for removing potentially toxic amyloid fibrils.

  18. Light Chain Amyloid Fibrils Cause Metabolic Dysfunction in Human Cardiomyocytes

    DOE PAGESBeta

    McWilliams-Koeppen, Helen P.; Foster, James S.; Hackenbrack, Nicole; Ramirez-Alvarado, Marina; Donohoe, Dallas; Williams, Angela; Macy, Sallie; Wooliver, Craig; Wortham, Dale; Morrell-Falvey, Jennifer; et al

    2015-09-22

    Light chain (AL) amyloidosis is the most common form of systemic amyloid disease, and cardiomyopathy is a dire consequence, resulting in an extremely poor prognosis. AL is characterized by the production of monoclonal free light chains that deposit as amyloid fibrils principally in the heart, liver, and kidneys causing organ dysfunction. We have studied the effects of amyloid fibrils, produced from recombinant λ6 light chain variable domains, on metabolic activity of human cardiomyocytes. The data indicate that fibrils at 0.1 μM, but not monomer, significantly decrease the enzymatic activity of cellular NAD(P)H-dependent oxidoreductase, without causing significant cell death. The presencemore » of amyloid fibrils did not affect ATP levels; however, oxygen consumption was increased and reactive oxygen species were detected. Confocal fluorescence microscopy showed that fibrils bound to and remained at the cell surface with little fibril internalization. Ultimately, these data indicate that AL amyloid fibrils severely impair cardiomyocyte metabolism in a dose dependent manner. These data suggest that effective therapeutic intervention for these patients should include methods for removing potentially toxic amyloid fibrils.« less

  19. Light Chain Amyloid Fibrils Cause Metabolic Dysfunction in Human Cardiomyocytes

    PubMed Central

    McWilliams-Koeppen, Helen P.; Foster, James S.; Hackenbrack, Nicole; Ramirez-Alvarado, Marina; Donohoe, Dallas; Williams, Angela; Macy, Sallie; Wooliver, Craig; Wortham, Dale; Morrell-Falvey, Jennifer; Foster, Carmen M.; Kennel, Stephen J.; Wall, Jonathan S.

    2015-01-01

    Light chain (AL) amyloidosis is the most common form of systemic amyloid disease, and cardiomyopathy is a dire consequence, resulting in an extremely poor prognosis. AL is characterized by the production of monoclonal free light chains that deposit as amyloid fibrils principally in the heart, liver, and kidneys causing organ dysfunction. We have studied the effects of amyloid fibrils, produced from recombinant λ6 light chain variable domains, on metabolic activity of human cardiomyocytes. The data indicate that fibrils at 0.1 μM, but not monomer, significantly decrease the enzymatic activity of cellular NAD(P)H-dependent oxidoreductase, without causing significant cell death. The presence of amyloid fibrils did not affect ATP levels; however, oxygen consumption was increased and reactive oxygen species were detected. Confocal fluorescence microscopy showed that fibrils bound to and remained at the cell surface with little fibril internalization. These data indicate that AL amyloid fibrils severely impair cardiomyocyte metabolism in a dose dependent manner. These data suggest that effective therapeutic intervention for these patients should include methods for removing potentially toxic amyloid fibrils. PMID:26393799

  20. Anticoagulation in atrial fibrillation

    PubMed Central

    Piccini, Jonathan P

    2014-01-01

    Atrial fibrillation increases the risk of stroke, which is a leading cause of death and disability worldwide. The use of oral anticoagulation in patients with atrial fibrillation at moderate or high risk of stroke, estimated by established criteria, improves outcomes. However, to ensure that the benefits exceed the risks of bleeding, appropriate patient selection is essential. Vitamin K antagonism has been the mainstay of treatment; however, newer drugs with novel mechanisms are also available. These novel oral anticoagulants (direct thrombin inhibitors and factor Xa inhibitors) obviate many of warfarin’s shortcomings, and they have demonstrated safety and efficacy in large randomized trials of patients with non-valvular atrial fibrillation. However, the management of patients taking warfarin or novel agents remains a clinical challenge. There are several important considerations when selecting anticoagulant therapy for patients with atrial fibrillation. This review will discuss the rationale for anticoagulation in patients with atrial fibrillation; risk stratification for treatment; available agents; the appropriate implementation of these agents; and additional, specific clinical considerations for treatment. PMID:24733535

  1. Anticoagulation in atrial fibrillation.

    PubMed

    Steinberg, Benjamin A; Piccini, Jonathan P

    2014-01-01

    Atrial fibrillation increases the risk of stroke, which is a leading cause of death and disability worldwide. The use of oral anticoagulation in patients with atrial fibrillation at moderate or high risk of stroke, estimated by established criteria, improves outcomes. However, to ensure that the benefits exceed the risks of bleeding, appropriate patient selection is essential. Vitamin K antagonism has been the mainstay of treatment; however, newer drugs with novel mechanisms are also available. These novel oral anticoagulants (direct thrombin inhibitors and factor Xa inhibitors) obviate many of warfarin's shortcomings, and they have demonstrated safety and efficacy in large randomized trials of patients with non-valvular atrial fibrillation. However, the management of patients taking warfarin or novel agents remains a clinical challenge. There are several important considerations when selecting anticoagulant therapy for patients with atrial fibrillation. This review will discuss the rationale for anticoagulation in patients with atrial fibrillation; risk stratification for treatment; available agents; the appropriate implementation of these agents; and additional, specific clinical considerations for treatment. PMID:24733535

  2. Protein Fibrils Induce Emulsion Stabilization.

    PubMed

    Peng, Jinfeng; Simon, Joana Ralfas; Venema, Paul; van der Linden, Erik

    2016-03-01

    The behavior of an oil-in-water emulsion was studied in the presence of protein fibrils for a wide range of fibril concentrations by using rheology, diffusing wave spectroscopy, and confocal laser scanning microscopy. Results showed that above a minimum fibril concentration depletion flocculation occurred, leading to oil droplet aggregation and enhanced creaming of the emulsion. Upon further increasing the concentration of the protein fibrils, the emulsions were stabilized. In this stable regime both aggregates of droplets and single droplets are present, and these aggregates are smaller than the aggregates in the flocculated emulsion samples at the lower fibril concentrations. The size of the droplet aggregates in the stabilized emulsions is independent of fibril concentration. In addition, the droplet aggregation was reversible upon dilution both by a pH 2 HCl solution and by a fibril solution at the same concentration. The viscosity of the emulsions containing fibrils was comparable to that of the pure fibril solution. Neither fibril networks nor droplet gel networks were observed in our study. The stabilization mechanism of emulsions containing long protein fibrils at high protein fibril concentrations points toward the mechanism of a kinetic stabilization. PMID:26882086

  3. Atrial Fibrillation (AF or AFib)

    MedlinePlus

    ... Pressure High Blood Pressure Tools & Resources Stroke More Atrial Fibrillation (AF or AFib) Updated:Feb 10,2016 What ... to the Terms and Conditions and Privacy Policy Atrial Fibrillation • Introduction • What is Atrial Fibrillation? • Why AFib Matters • ...

  4. Collagen vascular disease

    MedlinePlus

    ... this page: //medlineplus.gov/ency/article/001223.htm Collagen vascular disease To use the sharing features on ... were previously said to have "connective tissue" or "collagen vascular" disease. We now have names for many ...

  5. Evaluation of bioprosthetic heart valve failure using a matrix-fibril shear stress transfer approach.

    PubMed

    Anssari-Benam, Afshin; Barber, Asa H; Bucchi, Andrea

    2016-02-01

    A matrix-fibril shear stress transfer approach is devised and developed in this paper to analyse the primary biomechanical factors which initiate the structural degeneration of the bioprosthetic heart valves (BHVs). Using this approach, the critical length of the collagen fibrils l c and the interface shear acting on the fibrils in both BHV and natural aortic valve (AV) tissues under physiological loading conditions are calculated and presented. It is shown that the required critical fibril length to provide effective reinforcement to the natural AV and the BHV tissue is l c  = 25.36 µm and l c  = 66.81 µm, respectively. Furthermore, the magnitude of the required shear force acting on fibril interface to break a cross-linked fibril in the BHV tissue is shown to be 38 µN, while the required interfacial force to break the bonds between the fibril and the surrounding extracellular matrix is 31 µN. Direct correlations are underpinned between these values and the ultimate failure strength and the failure mode of the BHV tissue compared with the natural AV, and are verified against the existing experimental data. The analyses presented in this paper explain the role of fibril interface shear and critical length in regulating the biomechanics of the structural failure of the BHVs, for the first time. This insight facilitates further understanding into the underlying causes of the structural degeneration of the BHVs in vivo. PMID:26715134

  6. Biochemical and biophysical characterization of collagens of marine sponge, Ircinia fusca (Porifera: Demospongiae: Irciniidae).

    PubMed

    Pallela, Ramjee; Bojja, Sreedhar; Janapala, Venkateswara Rao

    2011-07-01

    Collagens were isolated and partially characterized from the marine demosponge, Ircinia fusca from Gulf of Mannar (GoM), India, with an aim to develop potentially applicable collagens from unused and under-used resources. The yield of insoluble, salt soluble and acid soluble forms of collagens was 31.71 ± 1.59, 20.69 ± 1.03, and 17.38 ± 0.87 mg/g dry weight, respectively. Trichrome staining, Scanning & Transmission Electron microscopic (SEM & TEM) studies confirmed the presence of collagen in the isolated, terminally globular irciniid filaments. The partially purified (gel filtration chromatography), non-fibrillar collagens appeared as basement type collagenous sheets under light microscopy whereas the purified fibrillar collagens appeared as fibrils with a repeated band periodicity of 67 nm under Atomic Force Microscope (AFM). The non-fibrillar and fibrillar collagens were seen to have affinity for anti-collagen type IV and type I antibodies raised against human collagens, respectively. The macromolecules, i.e., total protein, carbohydrate and lipid contents within the tissues were also quantified. The present information on the three characteristic irciniid collagens (filamentous, fibrillar and non-fibrillar) could assist the future attempts to unravel the therapeutically important, safer collagens from marine sponges for their use in pharmaceutical and cosmeceutical industries. PMID:21501629

  7. The Effect of Riboflavin/UVA Collagen Cross-linking Therapy on the Structure and Hydrodynamic Behaviour of the Ungulate and Rabbit Corneal Stroma

    PubMed Central

    Hayes, Sally; Kamma-Lorger, Christina S.; Boote, Craig; Young, Robert D.; Quantock, Andrew J.; Rost, Anika; Khatib, Yasmeen; Harris, Jonathan; Yagi, Naoto; Terrill, Nicholas; Meek, Keith M.

    2013-01-01

    Purpose To examine the effect of riboflavin/UVA corneal crosslinking on stromal ultrastructure and hydrodynamic behaviour. Methods One hundred and seventeen enucleated ungulate eyes (112 pig and 5 sheep) and 3 pairs of rabbit eyes, with corneal epithelium removed, were divided into four treatment groups: Group 1 (28 pig, 2 sheep and 3 rabbits) were untreated; Group 2 (24 pig) were exposed to UVA light (3.04 mW/cm2) for 30 minutes and Group 3 (29 pig) and Group 4 (31 pig, 3 sheep and 3 rabbits) had riboflavin eye drops applied to the corneal surface every 5 minutes for 35 minutes. Five minutes after the initial riboflavin instillation, the corneas in Group 4 experienced a 30 minute exposure to UVA light (3.04 mW/cm2). X-ray scattering was used to obtain measurements of collagen interfibrillar spacing, spatial order, fibril diameter, D-periodicity and intermolecular spacing throughout the whole tissue thickness and as a function of tissue depth in the treated and untreated corneas. The effect of each treatment on the hydrodynamic behaviour of the cornea (its ability to swell in saline solution) and its resistance to enzymatic digestion were assessed using in vitro laboratory techniques. Results Corneal thickness decreased significantly following riboflavin application (p<0.01) and also to a lesser extent after UVA exposure (p<0.05). With the exception of the spatial order factor, which was higher in Group 4 than Group 1 (p<0.01), all other measured collagen parameters were unaltered by cross-linking, even within the most anterior 300 microns of the cornea. The cross-linking treatment had no effect on the hydrodynamic behaviour of the cornea but did cause a significant increase in its resistance to enzymatic digestion. Conclusions It seems likely that cross-links formed during riboflavin/UVA therapy occur predominantly at the collagen fibril surface and in the protein network surrounding the collagen. PMID:23349690

  8. Rapid Fabrication of Living Tissue Models by Collagen Plastic Compression: Understanding Three-Dimensional Cell Matrix Repair In Vitro

    PubMed Central

    Cheema, Umber; Brown, Robert A.

    2013-01-01

    Objective To produce biomimetic collagen scaffolds for tissue modeling and as tissue-engineered implants. Approach Control of collagen fibril material parameters in collagen hydrogel scaffolds by using plastic compression (PC), resulting in direct control of cell proliferation, cell migration, and cell–cell interaction. Results We were able to control the density of collagen in such scaffolds from between 0.2% and 30%, and controllably layer the fibrils in the Z-plane. Cell migration was observed in gels where a gradient of collagen density was present. In these gels, cells preferentially migrated toward the collagen-dense areas. Cell proliferation rates were measurably higher in dense collagen gels. Innovation The use of PC to control material properties of collagen hydrogels results in collagen scaffolds that are biomimetic. These collagen gels reproduce the relevant matrix-mechanical environment in which behavior is more representative of that found in vivo. Conclusion The material properties of native collagen type I gels can be engineered to match those found in tissues in vivo to elicit more biomimetic cell behavior. PMID:24527341

  9. Transverse depth-dependent changes in corneal collagen lamellar orientation and distribution.

    PubMed

    Abass, Ahmed; Hayes, Sally; White, Nick; Sorensen, Thomas; Meek, Keith M

    2015-03-01

    It is thought that corneal surface topography may be stabilized by the angular orientation of out-of plane lamellae that insert into the anterior limiting membrane. In this study, micro-focus X-ray scattering data were used to obtain quantitative information about lamellar inclination (with respect to the corneal surface) and the X-ray scatter intensity throughout the depth of the cornea from the centre to the temporal limbus. The average collagen inclination remained predominantly parallel to the tissue surface at all depths. However, in the central cornea, the spread of inclination angles was greatest in the anterior-most stroma (reflecting the increased lamellar interweaving in this region), and decreased with tissue depth; in the peripheral cornea inclination angles showed less variation throughout the tissue thickness. Inclination angles in the deeper stroma were generally higher in the peripheral cornea, suggesting the presence of more interweaving in the posterior stroma away from the central cornea. An increase in collagen X-ray scatter was identified in a region extending from the sclera anteriorly until about 2 mm from the corneal centre. This could arise from the presence of larger diameter fibrils, probably of scleral origin, which are known to exist in this region. Incorporation of this quantitative information into finite-element models will further improve the accuracy with which they can predict the biomechanical response of the cornea to pathology and refractive procedures. PMID:25631562

  10. Transverse depth-dependent changes in corneal collagen lamellar orientation and distribution

    PubMed Central

    Abass, Ahmed; Hayes, Sally; White, Nick; Sorensen, Thomas; Meek, Keith M.

    2015-01-01

    It is thought that corneal surface topography may be stabilized by the angular orientation of out-of plane lamellae that insert into the anterior limiting membrane. In this study, micro-focus X-ray scattering data were used to obtain quantitative information about lamellar inclination (with respect to the corneal surface) and the X-ray scatter intensity throughout the depth of the cornea from the centre to the temporal limbus. The average collagen inclination remained predominantly parallel to the tissue surface at all depths. However, in the central cornea, the spread of inclination angles was greatest in the anterior-most stroma (reflecting the increased lamellar interweaving in this region), and decreased with tissue depth; in the peripheral cornea inclination angles showed less variation throughout the tissue thickness. Inclination angles in the deeper stroma were generally higher in the peripheral cornea, suggesting the presence of more interweaving in the posterior stroma away from the central cornea. An increase in collagen X-ray scatter was identified in a region extending from the sclera anteriorly until about 2 mm from the corneal centre. This could arise from the presence of larger diameter fibrils, probably of scleral origin, which are known to exist in this region. Incorporation of this quantitative information into finite-element models will further improve the accuracy with which they can predict the biomechanical response of the cornea to pathology and refractive procedures. PMID:25631562

  11. A collagen and elastic network in the wing of the bat.

    PubMed Central

    Holbrook, K A; Odland, G F

    1978-01-01

    Bundles of collagen fibrils, elastic fibres and fibroblasts are organized into a network that lies in the plane of a large portion of the bat wing. By ultrastructural (TEM and SEM) and biochemical analyses it was found that individual bundles of the net are similar to elastic ligaments. Although elastic fibres predominate, they are integrated and aligned in parallel with small bundles of collagen. A reticulum of fibroblasts, joined by focal junctions, forms a cellular framework throughout each bundle. Because of the unique features of the fibre bundles of the bat's wing, in particular their accessibility, and the parallel alignment of the collagen fibrils and elastic fibres in each easily isolatable fibre bundle, they should prove a most valuable model for connective tissue studies, particularly for the study of collagen-elastin interactions. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 PMID:649500

  12. Are amyloid fibrils molecular spandrels?

    PubMed

    Hane, Francis

    2013-11-15

    Amyloid-β, the protein implicated in Alzheimer's disease, along with a number of other proteins, has been shown to form amyloid fibrils. Fibril forming proteins share no common primary structure and have little known function. Furthermore, all proteins have the ability to form amyloid fibrils under certain conditions as the fibrillar structure lies at the global free energy minimum of proteins. This raises the question of the mechanism of the evolution of the amyloid fibril structure. Experimental evidence supports the hypothesis that the fibril structure is a by-product of the forces of protein folding and lies outside the bounds of evolutionary pressures. PMID:24140343

  13. GM1 cluster mediates formation of toxic Aβ fibrils by providing hydrophobic environments.

    PubMed

    Fukunaga, Saori; Ueno, Hiroshi; Yamaguchi, Takahiro; Yano, Yoshiaki; Hoshino, Masaru; Matsuzaki, Katsumi

    2012-10-16

    The conversion of soluble, nontoxic amyloid β-proteins (Aβ) to aggregated, toxic forms rich in β-sheets is considered to be a key step in the development of Alzheimer's disease. Accumulating evidence suggests that lipid-protein interactions play a crucial role in the aggregation of amyloidogenic proteins like Aβ. Our group has previously reported that amyloid fibrils of Aβ formed on membranes containing clusters of GM1 ganglioside (M-fibrils) exhibit greater cytotoxicity than fibrils formed in aqueous solution (W-fibrils) [ Okada ( 2008 ) J. Mol. Biol. 382 , 1066 - 1074 ]. W-fibrils are considered to consist of in-register parallel β-sheets. However, the precise molecular structure of M-fibrils and force driving the formation of toxic fibrils remain unclear. In this study, we hypothesized that low-polarity environments provided by GM1 clusters drive the formation of toxic fibrils and compared the structure and cytotoxicity of W-fibrils, M-fibrils, and aggregates formed in a low-polarity solution mimicking membrane environments. First, we determined solvent conditions which mimic the polarity of raftlike membranes using Aβ-(1-40) labeled with the 7-diethylaminocoumarin-3-carbonyl dye. The polarity of a mixture of 80% 1,4-dioxane and 20% water (v/v) was found to be close to that of raftlike membranes. Aβ-(1-40) formed amyloid fibrils within several hours in 80% dioxane (D-fibrils) or in the presence of raftlike membranes, whereas a much longer incubation time was required for fibril formation in a conventional buffer. D-fibrils were morphologically similar to M-fibrils. Fourier-transform infrared spectroscopy suggested that M-fibrils and D-fibrils contained antiparallel β-sheets. These fibrils had greater surface hydrophobicity and exhibited significant toxicity against human neuroblastoma SH-SY5Y cells, whereas W-fibrils with less surface hydrophobicity were not cytotoxic. We concluded that ganglioside clusters mediate the formation of toxic amyloid fibrils

  14. Teleost fish scales amongst the toughest collagenous materials.

    PubMed

    Khayer Dastjerdi, A; Barthelat, F

    2015-12-01

    Fish scales from modern teleost fish are high-performance materials made of cross-plies of collagen type I fibrils reinforced with hydroxyapatite. Recent studies on this material have demonstrated the remarkable performance of this material in tension and against sharp puncture. Although it is known that teleost fish scales are extremely tough, actual measurements of fracture toughness have so far not been reported because it is simply not possible to propagate a crack in this material using standard fracture testing configurations. Here we present a new fracture test setup where the scale is clamped between two pairs of miniature steel plates. The plates transmit the load uniformly, prevent warping of the scale and ensure a controlled crack propagation. We report a toughness of 15 to 18kJm(-2) (depending on the direction of crack propagation), which confirms teleost fish scales as one of the toughest biological material known. We also tested the individual bony layers, which we found was about four times less tough than the collagen layer because of its higher mineralization. The mechanical response of the scales also depends on the cohesion between fibrils and plies. Delamination tests show that the interface between the collagen fibrils is three orders of magnitude weaker than the scale, which explains the massive delamination and defibrillation observed experimentally. Finally, simple fracture mechanics models showed that process zone toughening is the principal source of toughening for the scales, followed by bridging by delaminated fibrils. These findings can guide the design of cross-ply composites and engineering textiles for high-end applications. This study also hints on the fracture mechanics and performance of collagenous materials with similar microstructures: fish skin, lamellar bone or tendons. PMID:25457170

  15. Nucleation of polymorphic amyloid fibrils.

    PubMed

    Auer, Stefan

    2015-03-10

    One and the same protein can self-assemble into amyloid fibrils with different morphologies. The phenomenon of fibril polymorphism is relevant biologically because different fibril polymorphs can have different toxicity, but there is no tool for predicting which polymorph forms and under what conditions. Here, we consider the nucleation of polymorphic amyloid fibrils occurring by direct polymerization of monomeric proteins into fibrils. We treat this process within the framework of our newly developed nonstandard nucleation theory, which allows prediction of the concentration dependence of the nucleation rate for different fibril polymorphs. The results highlight that the concentration dependence of the nucleation rate is closely linked with the protein solubility and a threshold monomer concentration below which fibril formation becomes biologically irrelevant. The relation between the nucleation rate, the fibril solubility, the threshold concentration, and the binding energies of the fibril building blocks within fibrils might prove a valuable tool for designing new experiments to control the formation of particular fibril polymorphs. PMID:25762329

  16. Prolyl hydroxylation of collagen type I is required for efficient binding to integrin alpha 1 beta 1 and platelet glycoprotein VI but not to alpha 2 beta 1.

    PubMed

    Perret, Stéephanie; Eble, Johannes A; Siljander, Pia R-M; Merle, Christine; Farndale, Richard W; Theisen, Manfred; Ruggiero, Florence

    2003-08-01

    Collagen is a potent adhesive substrate for cells, an event essentially mediated by the integrins alpha 1 beta 1 and alpha 2 beta 1. Collagen fibrils also bind to the integrin alpha 2 beta 1 and the platelet receptor glycoprotein VI to activate and aggregate platelets. The distinct triple helical recognition motifs for these receptors, GXOGER and (GPO)n, respectively, all contain hydroxyproline. Using unhydroxylated collagen I produced in transgenic plants, we investigated the role of hydroxyproline in the receptor-binding properties of collagen. We show that alpha 2 beta 1 but not alpha 1 beta 1 mediates cell adhesion to unhydroxylated collagen. Soluble recombinant alpha 1 beta 1 binding to unhydroxylated collagen is considerably reduced compared with bovine collagens, but binding can be restored by prolyl hydroxylation of recombinant collagen. We also show that platelets use alpha 2 beta 1 to adhere to the unhydroxylated recombinant molecules, but the adhesion is weaker than on fully hydroxylated collagen, and the unhydroxylated collagen fibrils fail to aggregate platelets. Prolyl hydroxylation is thus required for binding of collagen to platelet glycoprotein VI and to cells by alpha 1 beta 1. These observations give new insights into the molecular basis of collagen-receptor interactions and offer new selective applications for the recombinant unhydroxylated collagen I. PMID:12771137

  17. A Mouse Model for Dominant Collagen VI Disorders

    PubMed Central

    Pan, Te-Cheng; Zhang, Rui-Zhu; Arita, Machiko; Bogdanovich, Sasha; Adams, Sheila M.; Gara, Sudheer Kumar; Wagener, Raimund; Khurana, Tejvior S.; Birk, David E.; Chu, Mon-Li

    2014-01-01

    Dominant and recessive mutations in collagen VI genes, COL6A1, COL6A2, and COL6A3, cause a continuous spectrum of disorders characterized by muscle weakness and connective tissue abnormalities ranging from the severe Ullrich congenital muscular dystrophy to the mild Bethlem myopathy. Herein, we report the development of a mouse model for dominant collagen VI disorders by deleting exon 16 in the Col6a3 gene. The resulting heterozygous mouse, Col6a3+/d16, produced comparable amounts of normal Col6a3 mRNA and a mutant transcript with an in-frame deletion of 54 bp of triple-helical coding sequences, thus mimicking the most common molecular defect found in dominant Ullrich congenital muscular dystrophy patients. Biosynthetic studies of mutant fibroblasts indicated that the mutant α3(VI) collagen protein was produced and exerted a dominant-negative effect on collagen VI microfibrillar assembly. The distribution of the α3(VI)-like chains of collagen VI was not altered in mutant mice during development. The Col6a3+/d16 mice developed histopathologic signs of myopathy and showed ultrastructural alterations of mitochondria and sarcoplasmic reticulum in muscle and abnormal collagen fibrils in tendons. The Col6a3+/d16 mice displayed compromised muscle contractile functions and thereby provide an essential preclinical platform for developing treatment strategies for dominant collagen VI disorders. PMID:24563484

  18. Variation in the helical structure of native collagen.

    PubMed

    Orgel, Joseph P R O; Persikov, Anton V; Antipova, Olga

    2014-01-01

    The structure of collagen has been a matter of curiosity, investigation, and debate for the better part of a century. There has been a particularly productive period recently, during which much progress has been made in better describing all aspects of collagen structure. However, there remain some questions regarding its helical symmetry and its persistence within the triple-helix. Previous considerations of this symmetry have sometimes confused the picture by not fully recognizing that collagen structure is a highly complex and large hierarchical entity, and this affects and is effected by the super-coiled molecules that make it. Nevertheless, the symmetry question is not trite, but of some significance as it relates to extracellular matrix organization and cellular integration. The correlation between helical structure in the context of the molecular packing arrangement determines which parts of the amino acid sequence of the collagen fibril are buried or accessible to the extracellular matrix or the cell. In this study, we concentrate primarily on the triple-helical structure of fibrillar collagens I and II, the two most predominant types. By comparing X-ray diffraction data collected from type I and type II containing tissues, we point to evidence for a range of triple-helical symmetries being extant in the molecules native environment. The possible significance of helical instability, local helix dissociation and molecular packing of the triple-helices is discussed in the context of collagen's supramolecular organization, all of which must affect the symmetry of the collagen triple-helix. PMID:24586843

  19. X-ray scattering used to map the preferred collagen orientation in the human cornea and limbus.

    PubMed

    Aghamohammadzadeh, Hossein; Newton, Richard H; Meek, Keith M

    2004-02-01

    Many properties of connective tissues are governed by the organization of the constituent collagen. For example, the organization of collagen in the cornea and the limbus, where the cornea and sclera meet, is an important determinant of corneal curvature and hence of the eye's focusing power. We have used synchrotron X-ray scattering to map the orientation of the collagen fibrils throughout the human cornea, limbus, and adjacent sclera. We demonstrate a preferred orientation of collagen in the vertical and horizontal directions that is maintained to within about 1 mm from the limbus, where a circular or tangential disposition of fibrils occurs. The data are also used to map the relative distribution of both the total and the preferentially aligned collagen in different parts of the tissue, revealing considerable anisotropy. The detailed structural information provided is an important step toward understanding the shape and the mechanical properties of the tissue. PMID:14962385

  20. Micro-mechanical model for the tension-stabilized enzymatic degradation of collagen tissues

    NASA Astrophysics Data System (ADS)

    Nguyen, Thao; Ruberti, Jeffery

    We present a study of how the collagen fiber structure influences the enzymatic degradation of collagen tissues. Experiments of collagen fibrils and tissues show that mechanical tension can slow and halt enzymatic degradation. Tissue-level experiments also show that degradation rate is minimum at a stretch level coincident with the onset of strain-stiffening in the stress response. To understand these phenomena, we developed a micro-mechanical model of a fibrous collagen tissue undergoing enzymatic degradation. Collagen fibers are described as sinusoidal elastica beams, and the tissue is described as a distribution of fibers. We assumed that the degradation reaction is inhibited by the axial strain energy of the crimped collagen fibers. The degradation rate law was calibrated to experiments on isolated single fibrils from bovine sclera. The fiber crimp and properties were fit to uniaxial tension tests of tissue strips. The fibril-level kinetic and tissue-level structural parameters were used to predict tissue-level degradation-induced creep rate under a constant applied force. We showed that we could accurately predict the degradation-induce creep rate of the pericardium and cornea once we accounted for differences in the fiber crimp structure and properties.

  1. Lumican deficiency results in cardiomyocyte hypertrophy with altered collagen assembly.

    PubMed

    Dupuis, Loren E; Berger, Matthew G; Feldman, Samuel; Doucette, Lorna; Fowlkes, Vennece; Chakravarti, Shukti; Thibaudeau, Sarah; Alcala, Nicolas E; Bradshaw, Amy D; Kern, Christine B

    2015-07-01

    The ability of the heart to adapt to increased stress is dependent on the modification of its extracellular matrix (ECM) architecture that is established during postnatal development as cardiomyocytes differentiate, a process that is poorly understood. We hypothesized that the small leucine-rich proteoglycan (SLRP) lumican (LUM), which binds collagen and facilitates collagen assembly in other tissues, may play a critical role in establishing the postnatal murine myocardial ECM. Although previous studies suggest that LUM deficient mice (lum(-/-)) exhibit skin anomalies consistent with Ehlers-Danlos syndrome, lum(-/-) hearts have not been evaluated. These studies show that LUM was immunolocalized to non-cardiomyocytes of the cardiac ventricles and its expression increased throughout development. Lumican deficiency resulted in significant (50%) perinatal death and further examination of the lum(-/-) neonatal hearts revealed an increase in myocardial tissue without a significant increase in cell proliferation. However cardiomyocytes from surviving postnatal day 0 (P0), 1 month (1 mo) and adult (4 mo) lum(-/-) hearts were significantly larger than their wild type (WT) littermates. Immunohistochemistry revealed that the increased cardiomyocyte size in the lum(-/-) hearts correlated with alteration of the cardiomyocyte pericellular ECM components collagenα1(I) and the class I SLRP decorin (DCN). Western blot analysis demonstrated that the ratio of glycosaminoglycan (GAG) decorated DCN to core DCN was reduced in P0 and 1 mo lum(-/-) hearts. There was also a reduction in the β and γ forms of collagenα1(I) in lum(-/-) hearts. While the total insoluble collagen content was significantly reduced, the fibril size was increased in lum(-/-) hearts, indicating that LUM may play a role in collagen fiber stability and lateral fibril assembly. These results suggest that LUM controls cardiomyocyte growth by regulating the pericellular ECM and also indicates that LUM may coordinate

  2. Developmental Stage-dependent Regulation of Prolyl 3-Hydroxylation in Tendon Type I Collagen.

    PubMed

    Taga, Yuki; Kusubata, Masashi; Ogawa-Goto, Kiyoko; Hattori, Shunji

    2016-01-01

    3-Hydroxyproline (3-Hyp), which is unique to collagen, is a fairly rare post-translational modification. Recent studies have suggested a function of prolyl 3-hydroxylation in fibril assembly and its relationships with certain disorders, including recessive osteogenesis imperfecta and high myopia. However, no direct evidence for the physiological and pathological roles of 3-Hyp has been presented. In this study, we first estimated the overall alterations in prolyl hydroxylation in collagens purified from skin, bone, and tail tendon of 0.5-18-month-old rats by LC-MS analysis with stable isotope-labeled collagen, which was recently developed as an internal standard for highly accurate collagen analyses. 3-Hyp was found to significantly increase in tendon collagen until 3 months after birth and then remain constant, whereas increased prolyl 3-hydroxylation was not observed in skin and bone collagen. Site-specific analysis further revealed that 3-Hyp was increased in tendon type I collagen in a specific sequence region, including a previously known modification site at Pro(707) and newly identified sites at Pro(716) and Pro(719), at the early ages. The site-specific alterations in prolyl 3-hydroxylation with aging were also observed in bovine Achilles tendon. We postulate that significant increases in 3-Hyp at the consecutive modification sites are correlated with tissue development in tendon. The present findings suggest that prolyl 3-hydroxylation incrementally regulates collagen fibril diameter in tendon. PMID:26567337

  3. On the Presence of Affine Fibril and Fiber Kinematics in the Mitral Valve Anterior Leaflet

    PubMed Central

    Lee, Chung-Hao; Zhang, Will; Liao, Jun; Carruthers, Christopher A.; Sacks, Jacob I.; Sacks, Michael S.

    2015-01-01

    In this study, we evaluated the hypothesis that the constituent fibers follow an affine deformation kinematic model for planar collagenous tissues. Results from two experimental datasets were utilized, taken at two scales (nanometer and micrometer), using mitral valve anterior leaflet (MVAL) tissues as the representative tissue. We simulated MVAL collagen fiber network as an ensemble of undulated fibers under a generalized two-dimensional deformation state, by representing the collagen fibrils based on a planar sinusoidally shaped geometric model. The proposed approach accounted for collagen fibril amplitude, crimp period, and rotation with applied macroscopic tissue-level deformation. When compared to the small angle x-ray scattering measurements, the model fit the data well, with an r2 = 0.976. This important finding suggests that, at the homogenized tissue-level scale of ∼1 mm, the collagen fiber network in the MVAL deforms according to an affine kinematics model. Moreover, with respect to understanding its function, affine kinematics suggests that the constituent fibers are largely noninteracting and deform in accordance with the bulk tissue. It also suggests that the collagen fibrils are tightly bounded and deform as a single fiber-level unit. This greatly simplifies the modeling efforts at the tissue and organ levels, because affine kinematics allows a straightforward connection between the macroscopic and local fiber strains. It also suggests that the collagen and elastin fiber networks act independently of each other, with the collagen and elastin forming long fiber networks that allow for free rotations. Such freedom of rotation can greatly facilitate the observed high degree of mechanical anisotropy in the MVAL and other heart valves, which is essential to heart valve function. These apparently novel findings support modeling efforts directed toward improving our fundamental understanding of tissue biomechanics in healthy and diseased conditions

  4. Polyvinyl alcohol-graft-polyethylene glycol hydrogels improve utility and biofunctionality of injectable collagen biomaterials.

    PubMed

    Hartwell, Ryan; Chan, Ben; Elliott, Keenan; Alnojeidi, Hatem; Ghahary, Aziz

    2016-01-01

    Collagen-based materials have become a staple in both research and the clinic. In wound care, collagen-based materials comprise a core gamut of biological dressings and therapeutic strategies. In research, collagen-based materials are employed in everything from 3D cultures to bioprinting. Soluble collagen is well characterized to undergo fibrillation at neutral pH and 37 °C. To remain stable, a neutralized collagen solution must be maintained at 4 °C. These physical characteristics of collagen impose limitations on its utility. In our previous work, we identified that the incorporation of a simple polyvinyl alcohol:borate hydrogel could improve the rate of collagen gel fibrillation. In this work we sought to further investigate the interactions of polyvinyl alcohol blend variants, as surfactant-like polymers, in comparison with known non-polymer surfactants. To conduct our investigations scaffold variants were created using increasing concentrations of polyvinyl alcohol, differing combinations of polymers, and non-polymer surfactants Tweens 20 and 80, and TritonX-100. Activation energy for collagen fibrillation was found to significantly decrease in the presence of polyvinyl alcohols (p  <  0.01) at and above 0.4%w/v concentration. Further, addition of polyvinyl alcohol-graft-polyethylene glycol had the greatest enhancement (2.02 fold) on the fibrillation kinetics (p  <  0.01), wetting properties and the stability of the collagen scaffolds post-freeze drying. Our results demonstrated that the addition of polyvinyl alcohol hydrogels to a collagen solution could stabilize collagen solution such that the solution could easily be lyophilized (at pH 7) and then reconstituted with water. Cells cultured in polyvinyl alcohol scaffolds also exhibited more organized F-actin, as well as a reduced abundance of pro-collagen and α-smooth actin. In conclusion, our results demonstrate for the first time that polyvinyl alcohol, preferably polyvinyl alcohol

  5. A Colloidal Description of Intermolecular Interactions Driving Fibril-Fibril Aggregation of a Model Amphiphilic Peptide.

    PubMed

    Owczarz, Marta; Motta, Anna C; Morbidelli, Massimo; Arosio, Paolo

    2015-07-14

    effects are mainly related to the binding to the positive groups of the fibril surface and to the resulting decrease of the surface charge, cation effects are more complex and involve additional solvation forces. PMID:26125620

  6. Lung response to ultrafine Kevlar aramid synthetic fibrils following 2-year inhalation exposure in rats.

    PubMed

    Lee, K P; Kelly, D P; O'Neal, F O; Stadler, J C; Kennedy, G L

    1988-07-01

    Four groups of 100 male and 100 female rats were exposed to ultrafine Kevlar fibrils at concentrations of 0, 2.5, 25, and 100 fibrils/cc for 6 hr/day, 5 days/week for 2 years. One group was exposed to 400 fibrils/cc for 1 year and allowed to recover for 1 year. At 2.5 fibrils/cc, the lungs had normal alveolar architecture with a few dust-laden macrophages (dust cell response) in the alveolar airspaces. At 25 fibrils/cc, the lungs showed a dust cell response, slight Type II pneumocyte hyperplasia, alveolar bronchiolarization, and a negligible amount of collagenized fibrosis in the alveolar duct region. At 100 fibrils/cc, the same pulmonary responses were seen as at 25 fibrils/cc. In addition, cystic keratinizing squamous cell carcinoma (CKSCC) was found in 4 female rats, but not in male rats. Female rats had more prominent foamy alveolar macrophages, cholesterol granulomas, and alveolar bronchiolarization. These pulmonary lesions were related to the development of CKSCC. The lung tumors were derived from metaplastic squamous cells in areas of alveolar bronchiolarization. At 400 fibrils/cc following 1 year of recovery, the lung dust content, average fiber length, and the pulmonary lesions were markedly reduced, but slight centriacinar emphysema and minimal collagenized fibrosis were found in the alveolar duct region. One male and 6 female rats developed CKSCC. The lung tumors were a unique type of experimentally induced tumors in the rats and have not been seen as spontaneous tumors in man or animals. Therefore, the relevance of this type of lung tumor to the human situation is minimal. PMID:3209007

  7. Adhesion forces between AFM tips and superficial dentin surfaces.

    PubMed

    Pelin, I M; Piednoir, A; Machon, D; Farge, P; Pirat, C; Ramos, S M M

    2012-06-15

    In this work, we study the adhesion forces between atomic force microscopy (AFM) tips and superficial dentin etched with phosphoric acid. Initially, we quantitatively analyze the effect of acid etching on the surface heterogeneity and the surface roughness, two parameters that play a key role in the adhesion phenomenon. From a statistical study of the force-distance curves, we determine the average adhesion forces on the processed substrates. Our results show that the average adhesion forces, measured in water, increase linearly with the acid exposure time. The highest values of such forces are ascribed to the high density of collagen fibers on the etched surfaces. The individual contribution of exposed collagen fibrils to the adhesion force is highlighted. We also discuss in this paper the influence of the environmental medium (water/air) in the adhesion measurements. We show that the weak forces involved require working in the aqueous medium. PMID:22472512

  8. Structural aspects of the calcification process of lower vertebrate collagen.

    PubMed

    Bigi, A; Koch, M H; Panzavolta, S; Roveri, N; Rubini, K

    2000-01-01

    In order to investigate the structural relationship between inorganic phase and collagen fibrils in the calcified tissues of lower vertebrates we have carried out a wide and small angle X-ray diffraction investigation on carp scales and bone samples. The small angle patterns from decalcified bone and scales, as well as uncalcified tendon samples from carp are very similar to that of type I collagen from higher vertebrates. The D-axial period, 67 nm, is the same as that of higher vertebrate type I collagen, while the most significant difference is the relatively low intensity of the first order reflection, which is, however, the most intense. The relative intensity distributions of the meridional reflections recorded from fish bone and scales are in agreement with an electron density distribution according to a step function. The calculated step length is very close to the values previously reported for calcified tissues from higher vertebrates. The small angle reflections from calcified, as well as decalcified, scales display different directions of orientation, which could be in agreement with a plywood arrangement of collagen fibrils in successive sheets parallel to the plane of the scale. PMID:10826707

  9. Phase matching of backward second harmonic generation assisted by lattice structure in collagen tissues.

    PubMed

    Shen, Mengzhe; Zeng, Haishan; Tang, Shuo

    2015-10-01

    Phase matching of backward second harmonic generation (SHG) in a periodic structure of collagen fibrils is investigated through theoretical modeling, simulation, and experiments. The lattice structure of collagen fibrils is considered to provide a virtual momentum for assisting the phase matching of backward SHG. Phase matching over a relatively wide excitation wavelength range is achieved by tilting the angle of the fundamental excitation and SHG wave vectors. The SHG intensity in the periodic structure is simulated to quantify the phase matching effect. The effect of the fundamental excitation and the SHG emission angles on the peak excitation wavelength of the SHG excitation spectrum is further validated in experiments, where the excitation and emission angles are controlled by spatial filtering. It is found that an optimized excitation wavelength exists for a certain collagen fibril structure, which shifts toward a shorter wavelength when the excitation and emission angles are increased. Our results show that the lattice structure of collagen fibrils can assist the phase matching, providing a mechanism for generating backward SHG in multiphoton microscopy. PMID:26502229

  10. Phase matching of backward second harmonic generation assisted by lattice structure in collagen tissues

    NASA Astrophysics Data System (ADS)

    Shen, Mengzhe; Zeng, Haishan; Tang, Shuo

    2015-10-01

    Phase matching of backward second harmonic generation (SHG) in a periodic structure of collagen fibrils is investigated through theoretical modeling, simulation, and experiments. The lattice structure of collagen fibrils is considered to provide a virtual momentum for assisting the phase matching of backward SHG. Phase matching over a relatively wide excitation wavelength range is achieved by tilting the angle of the fundamental excitation and SHG wave vectors. The SHG intensity in the periodic structure is simulated to quantify the phase matching effect. The effect of the fundamental excitation and the SHG emission angles on the peak excitation wavelength of the SHG excitation spectrum is further validated in experiments, where the excitation and emission angles are controlled by spatial filtering. It is found that an optimized excitation wavelength exists for a certain collagen fibril structure, which shifts toward a shorter wavelength when the excitation and emission angles are increased. Our results show that the lattice structure of collagen fibrils can assist the phase matching, providing a mechanism for generating backward SHG in multiphoton microscopy.

  11. Genetics Home Reference: familial atrial fibrillation

    MedlinePlus

    ... fibrillation also increases the risk of stroke and sudden death. Complications of familial atrial fibrillation can occur at ... beats , increasing the risk of syncope, stroke, and sudden death. Most cases of atrial fibrillation are not caused ...

  12. Reinforcement of polymeric structures with asbestos fibrils

    NASA Technical Reports Server (NTRS)

    Rader, C. A.; Schwartz, A. M.

    1970-01-01

    Investigation determines structural potential of asbestos fibrils. Methods are developed for dispersing macrofibers of the asbestos into colloidal-sized ultimate fibrils and incorporating these fibrils in matrices without causing reagglomeration.

  13. Forward versus backward polarization-resolved SHG imaging of collagen structure in tissues

    NASA Astrophysics Data System (ADS)

    Teulon, Claire; Gusachenko, Ivan; Latour, Gaël.; Schanne-Klein, Marie-Claire

    2016-03-01

    Second harmonic generation (SHG) is a powerful technique to observe fibrillar collagen without any staining and with a good contrast. More information about the molecular structure of collagen fibrils in tissues and their 3D distribution can be gained with polarization-resolved SHG imaging. Nevertheless, strong focusing is required for effective imaging and light propagation in tissues is complex and not thoroughly understood yet, preventing accurate and reproducible measurements. Theoretical analysis, vectorial numerical simulations and experiments were implemented to understand how the SHG signal builds up and how geometrical parameters affect polarization-resolved measurements in homogeneous collagen-rich tissues.

  14. Atrial fibrillation and inflammation

    PubMed Central

    Ozaydin, Mehmet

    2010-01-01

    Atrial fibrillation (AF) is the most common clinical arrhythmia. Recent investigations have suggested that inflammation might have a role in the pathophysiology of AF. In this review, the association between inflammation and AF, and the effects of several agents that have anti-inflammatory actions, such as statins, polyunsaturated fatty acids, corticosteroids and angiotensin-converting enzyme inhibitors and angiotensin receptor blockers, have been investigated. PMID:21160591

  15. Atrial fibrillation case study.

    PubMed

    Johnson, Sarah; Wilson, Tracey

    2016-03-01

    This article discusses the irregular heart rhythm caused by atrial fibrillation (AF). A brief overview of the pathophysiology will be provided. A case study is discussed to highlight the treatment and management of AF. The care provision describes common signs and symptoms and also the treatment and management of AF within the maternity care setting. The importance of maintaining the mother-baby dyad is highlighted. For the purpose of maintaining confidentiality the woman will be referred to as Shama. PMID:27044188

  16. Intrafibrillar plasticity through mineral/collagen sliding is the dominant mechanism for the extreme toughness of antler bone.

    PubMed

    Gupta, H S; Krauss, S; Kerschnitzki, M; Karunaratne, A; Dunlop, J W C; Barber, A H; Boesecke, P; Funari, S S; Fratzl, P

    2013-12-01

    The inelastic deformability of the mineralised matrix in bones is critical to their high toughness, but the nanoscale mechanisms are incompletely understood. Antler is a tough bone type, with a nanostructure composed of mineralised collagen fibrils ∼100nm diameter. We track the fibrillar deformation of antler tissue during cyclic loading using in situ synchrotron small-angle X-ray diffraction (SAXD), finding that residual strain remains in the fibrils after the load was removed. During repeated unloading/reloading cycles, the fibril strain shows minimal hysteresis when plotted as a function of tissue strain, indicating that permanent plastic strain accumulates inside the fibril. We model the tensile response of the mineralised collagen fibril by a two - level staggered model - including both elastic - and inelastic regimes - with debonding between mineral and collagen within fibrils triggering macroscopic inelasticity. In the model, the subsequent frictional sliding at intrafibrillar mineral/collagen interfaces accounts for subsequent inelastic deformation of the tissue in tension. The model is compared to experimental measurements of fibrillar and mineral platelet strain during tensile deformation, measured by in situ synchrotron SAXD and wide-angle X-ray diffraction (WAXD) respectively, as well as macroscopic tissue stress and strain. By fitting the model predictions to experimentally observed parameters like the yield point, elastic modulus and post-yield slope, extremely good agreement is found between the model and experimental data at both the macro- and at the nanoscale. Our results provide strong evidence that intrafibrillar sliding between mineral and collagen leads to permanent plastic strain at both the fibril and the tissue level, and that the energy thus dissipated is a significant factor behind the high toughness of antler bone. PMID:23707600

  17. Identification of cooked bone using TEM imaging of bone collagen.

    PubMed

    Koon, Hannah E C

    2012-01-01

    Mild heating (≤100° C, 1 h)-typical of cooking-does not lead to detectable changes in any biochemical parameter yet measured; consequently bones that have been cooked, but which have not reached a temperature that will induce charring go undetected. We have used a microscopy based approach to investigate changes in the organization of the bone protein, collagen, as it is heated, using bone from heating experiments, short term burials, and archaeological sites. The work has revealed that the presence of a mineral matrix stabilizes the collagen enabling the damage to accumulate, but preventing it from causing immediate gelatinization. Once the mineral is removed, the damage can be observed using appropriate visualization methods. This chapter describes the transmission electron microscopy (TEM) technique that has been used to detect cooked bone by visualizing minor heat-induced damage at the level of the collagen fibril. PMID:22907413

  18. Lysyl Hydroxylase 3-mediated Glucosylation in Type I Collagen

    PubMed Central

    Sricholpech, Marnisa; Perdivara, Irina; Yokoyama, Megumi; Nagaoka, Hideaki; Terajima, Masahiko; Tomer, Kenneth B.; Yamauchi, Mitsuo

    2012-01-01

    Recently, by employing the short hairpin RNA technology, we have generated MC3T3-E1 (MC)-derived clones stably suppressing lysyl hydroxylase 3 (LH3) (short hairpin (Sh) clones) and demonstrated the LH3 function as glucosyltransferase in type I collagen (Sricholpech, M., Perdivara, I., Nagaoka, H., Yokoyama, M., Tomer, K. B., and Yamauchi, M. (2011) Lysyl hydroxylase 3 glucosylates galactosylhydroxylysine residues in type I collagen in osteoblast culture. J. Biol. Chem. 286, 8846–8856). To further elucidate the biological significance of this modification, we characterized and compared type I collagen phenotypes produced by Sh clones and two control groups, MC and those transfected with empty vector. Mass spectrometric analysis identified five glycosylation sites in type I collagen (i.e. α1,2-87, α1,2-174, and α2-219. Of these, the predominant glycosylation site was α1-87, one of the major helical cross-linking sites. In Sh collagen, the abundance of glucosylgalactosylhydroxylysine was significantly decreased at all of the five sites with a concomitant increase in galactosylhydroxylysine at four of these sites. The collagen cross-links were significantly diminished in Sh clones, and, for the major cross-link, dihydroxylysinonorleucine (DHLNL), glucosylgalactosyl-DHLNL was diminished with a concomitant increase in galactosyl-DHLNL. When subjected to in vitro incubation, in Sh clones, the rate of decrease in DHLNL was lower, whereas the rate of increase in its maturational cross-link, pyridinoline, was comparable with controls. Furthermore, in Sh clones, the mean diameters of collagen fibrils were significantly larger, and the onset of mineralized nodule formation was delayed when compared with those of controls. These results indicate that the LH3-mediated glucosylation occurs at the specific molecular loci in the type I collagen molecule and plays critical roles in controlling collagen cross-linking, fibrillogenesis, and mineralization. PMID:22573318

  19. Enigmatic insight into collagen.

    PubMed

    Deshmukh, Shrutal Narendra; Dive, Alka M; Moharil, Rohit; Munde, Prashant

    2016-01-01

    Collagen is a unique, triple helical molecule which forms the major part of extracellular matrix. It is the most abundant protein in the human body, representing 30% of its dry weight. It is the fibrous structural protein that makes up the white fibers (collagen fibers) of skin, tendons, bones, cartilage and all other connective tissues. Collagens are not only essential for the mechanical resistance and resilience of multicellular organisms, but are also signaling molecules defining cellular shape and behavior. The human body has at least 16 types of collagen, but the most prominent types are I, II and III. Collagens are produced by several cell types and are distinguishable by their molecular compositions, morphologic characteristics, distribution, functions and pathogenesis. This is the major fibrous glycoprotein present in the extracellular matrix and in connective tissue and helps in maintaining the structural integrity of these tissues. It has a triple helical structure. Various studies have proved that mutations that modify folding of the triple helix result in identifiable genetic disorders. Collagen diseases share certain similarities with autoimmune diseases, because autoantibodies specific to each collagen disease are produced. Therefore, this review highlights the role of collagen in normal health and also the disorders associated with structural and functional defects in collagen. PMID:27601823

  20. Collagen and gelatin.

    PubMed

    Liu, Dasong; Nikoo, Mehdi; Boran, Gökhan; Zhou, Peng; Regenstein, Joe M

    2015-01-01

    Collagen and gelatin have been widely used in the food, pharmaceutical, and cosmetic industries due to their excellent biocompatibility, easy biodegradability, and weak antigenicity. Fish collagen and gelatin are of renewed interest, owing to the safety and religious concerns of their mammalian counterparts. The structure of collagen has been studied using various modern technologies, and interpretation of the raw data should be done with caution. The structure of collagen may vary with sources and seasons, which may affect its applications and optimal extraction conditions. Numerous studies have investigated the bioactivities and biological effects of collagen, gelatin, and their hydrolysis peptides, using both in vitro and in vivo assay models. In addition to their established nutritional value as a protein source, collagen and collagen-derived products may exert various potential biological activities on cells in the extracellular matrix through the corresponding food-derived peptides after ingestion, and this might justify their applications in dietary supplements and pharmaceutical preparations. Moreover, an increasing number of novel applications have been found for collagen and gelatin. Therefore, this review covers the current understanding of the structure, bioactivities, and biological effects of collagen, gelatin, and gelatin hydrolysates as well as their most recent applications. PMID:25884286

  1. Enigmatic insight into collagen

    PubMed Central

    Deshmukh, Shrutal Narendra; Dive, Alka M; Moharil, Rohit; Munde, Prashant

    2016-01-01

    Collagen is a unique, triple helical molecule which forms the major part of extracellular matrix. It is the most abundant protein in the human body, representing 30% of its dry weight. It is the fibrous structural protein that makes up the white fibers (collagen fibers) of skin, tendons, bones, cartilage and all other connective tissues. Collagens are not only essential for the mechanical resistance and resilience of multicellular organisms, but are also signaling molecules defining cellular shape and behavior. The human body has at least 16 types of collagen, but the most prominent types are I, II and III. Collagens are produced by several cell types and are distinguishable by their molecular compositions, morphologic characteristics, distribution, functions and pathogenesis. This is the major fibrous glycoprotein present in the extracellular matrix and in connective tissue and helps in maintaining the structural integrity of these tissues. It has a triple helical structure. Various studies have proved that mutations that modify folding of the triple helix result in identifiable genetic disorders. Collagen diseases share certain similarities with autoimmune diseases, because autoantibodies specific to each collagen disease are produced. Therefore, this review highlights the role of collagen in normal health and also the disorders associated with structural and functional defects in collagen. PMID:27601823

  2. Cell Adhesion on Amyloid Fibrils Lacking Integrin Recognition Motif.

    PubMed

    Jacob, Reeba S; George, Edna; Singh, Pradeep K; Salot, Shimul; Anoop, Arunagiri; Jha, Narendra Nath; Sen, Shamik; Maji, Samir K

    2016-03-01

    Amyloids are highly ordered, cross-β-sheet-rich protein/peptide aggregates associated with both human diseases and native functions. Given the well established ability of amyloids in interacting with cell membranes, we hypothesize that amyloids can serve as universal cell-adhesive substrates. Here, we show that, similar to the extracellular matrix protein collagen, amyloids of various proteins/peptides support attachment and spreading of cells via robust stimulation of integrin expression and formation of integrin-based focal adhesions. Additionally, amyloid fibrils are also capable of immobilizing non-adherent red blood cells through charge-based interactions. Together, our results indicate that both active and passive mechanisms contribute to adhesion on amyloid fibrils. The present data may delineate the functional aspect of cell adhesion on amyloids by various organisms and its involvement in human diseases. Our results also raise the exciting possibility that cell adhesivity might be a generic property of amyloids. PMID:26742841

  3. Electroactive biomimetic collagen-silver nanowire composite scaffolds

    NASA Astrophysics Data System (ADS)

    Wickham, Abeni; Vagin, Mikhail; Khalaf, Hazem; Bertazzo, Sergio; Hodder, Peter; Dånmark, Staffan; Bengtsson, Torbjörn; Altimiras, Jordi; Aili, Daniel

    2016-07-01

    Electroactive biomaterials are widely explored as bioelectrodes and as scaffolds for neural and cardiac regeneration. Most electrodes and conductive scaffolds for tissue regeneration are based on synthetic materials that have limited biocompatibility and often display large discrepancies in mechanical properties with the surrounding tissue causing problems during tissue integration and regeneration. This work shows the development of a biomimetic nanocomposite material prepared from self-assembled collagen fibrils and silver nanowires (AgNW). Despite consisting of mostly type I collagen fibrils, the homogeneously embedded AgNWs provide these materials with a charge storage capacity of about 2.3 mC cm-2 and a charge injection capacity of 0.3 mC cm-2, which is on par with bioelectrodes used in the clinic. The mechanical properties of the materials are similar to soft tissues with a dynamic elastic modulus within the lower kPa range. The nanocomposites also support proliferation of embryonic cardiomyocytes while inhibiting the growth of both Gram-negative Escherichia coli and Gram-positive Staphylococcus epidermidis. The developed collagen/AgNW composites thus represent a highly attractive bioelectrode and scaffold material for a wide range of biomedical applications.Electroactive biomaterials are widely explored as bioelectrodes and as scaffolds for neural and cardiac regeneration. Most electrodes and conductive scaffolds for tissue regeneration are based on synthetic materials that have limited biocompatibility and often display large discrepancies in mechanical properties with the surrounding tissue causing problems during tissue integration and regeneration. This work shows the development of a biomimetic nanocomposite material prepared from self-assembled collagen fibrils and silver nanowires (AgNW). Despite consisting of mostly type I collagen fibrils, the homogeneously embedded AgNWs provide these materials with a charge storage capacity of about 2.3 mC cm-2

  4. Mechano-regulation of Collagen Biosynthesis in Periodontal Ligament

    PubMed Central

    Kaku, Masaru; Yamauchi, Mitsuo

    2014-01-01

    Purpose Periodontal ligament (PDL) plays critical roles in the development and maintenance of periodontium such as tooth eruption and dissipation of masticatory force. The mechanical properties of PDL are mainly derived from fibrillar type I collagen, the most abundant extracellular component. Study selection The biosynthesis of type I collagen is a long, complex process including a number of intra- and extracellular post-translational modifications. The final modification step is the formation of covalent intra- and intermolecular cross-links that provide collagen fibrils with stability and connectivity. Results It is now clear that collagen post-translational modifications are regulated by groups of specific enzymes and associated molecules in a tissue-specific manner; and these modifications appear to change in response to mechanical force. Conclusions This review focuses on the effect of mechanical loading on collagen biosynthesis and fibrillogenesis in PDL with emphasis on the post-translational modifications of collagens, which is an important molecular aspect to understand in the field of prosthetic dentistry. PMID:25311991

  5. Structural basis for collagen recognition by the immune receptor OSCAR

    PubMed Central

    Zhou, Long; Hinerman, Jennifer M.; Blaszczyk, Michal; Miller, Jeanette L. C.; Conrady, Deborah G.; Barrow, Alexander D.; Chirgadze, Dimitri Y.; Bihan, Dominique; Farndale, Richard W.

    2016-01-01

    The osteoclast-associated receptor (OSCAR) is a collagen-binding immune receptor with important roles in dendritic cell maturation and activation of inflammatory monocytes as well as in osteoclastogenesis. The crystal structure of the OSCAR ectodomain is presented, both free and in complex with a consensus triple-helical peptide (THP). The structures revealed a collagen-binding site in each immunoglobulin-like domain (D1 and D2). The THP binds near a predicted collagen-binding groove in D1, but a more extensive interaction with D2 is facilitated by the unusually wide D1-D2 interdomain angle in OSCAR. Direct binding assays, combined with site-directed mutagenesis, confirm that the primary collagen-binding site in OSCAR resides in D2, in marked contrast to the related collagen receptors, glycoprotein VI (GPVI) and leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1). Monomeric OSCAR D1D2 binds to the consensus THP with a KD of 28 µM measured in solution, but shows a higher affinity (KD 1.5 μM) when binding to a solid-phase THP, most likely due to an avidity effect. These data suggest a 2-stage model for the interaction of OSCAR with a collagen fibril, with transient, low-affinity interactions initiated by the membrane-distal D1, followed by firm adhesion to the primary binding site in D2. PMID:26552697

  6. Tumor matrix protein collagen XIα1 in cancer

    PubMed Central

    Raglow, Zoe; Thomas, Sufi M

    2015-01-01

    The extracellular matrix is increasingly recognized as an essential player in cancer development and progression. Collagens are one of the most important components of the extracellular matrix, and have themselves been implicated in many aspects of neoplastic transformation. Collagen XI is a minor collagen whose main physiologic function is to regulate the diameter of major collagen fibrils. The α1 chain of collagen XI (colXIα1), has known pathogenic roles in several musculoskeletal disorders. Recent research has highlighted the importance of colXIα1 in many types of cancer, including its roles in metastasis, angiogenesis, and drug resistance, as well as its potential utility in screening tests and as a therapeutic target. High levels of colXIα1 overexpression have been reported in multiple expression profile studies examining differences between cancerous and normal tissue, and between beginning and advanced stage cancer. Its expression has been linked to poor progression-free and overall survival. The consistency of this data across cancer types is particularly striking, including colorectal, ovarian, breast, head and neck, lung, and brain cancers. This review discusses the role of collagen XIα1 in cancer and its potential as a target for cancer therapy. PMID:25511741

  7. Structural basis for collagen recognition by the immune receptor OSCAR.

    PubMed

    Zhou, Long; Hinerman, Jennifer M; Blaszczyk, Michal; Miller, Jeanette L C; Conrady, Deborah G; Barrow, Alexander D; Chirgadze, Dimitri Y; Bihan, Dominique; Farndale, Richard W; Herr, Andrew B

    2016-02-01

    The osteoclast-associated receptor (OSCAR) is a collagen-binding immune receptor with important roles in dendritic cell maturation and activation of inflammatory monocytes as well as in osteoclastogenesis. The crystal structure of the OSCAR ectodomain is presented, both free and in complex with a consensus triple-helical peptide (THP). The structures revealed a collagen-binding site in each immunoglobulin-like domain (D1 and D2). The THP binds near a predicted collagen-binding groove in D1, but a more extensive interaction with D2 is facilitated by the unusually wide D1-D2 interdomain angle in OSCAR. Direct binding assays, combined with site-directed mutagenesis, confirm that the primary collagen-binding site in OSCAR resides in D2, in marked contrast to the related collagen receptors, glycoprotein VI (GPVI) and leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1). Monomeric OSCAR D1D2 binds to the consensus THP with a KD of 28 µM measured in solution, but shows a higher affinity (KD 1.5 μM) when binding to a solid-phase THP, most likely due to an avidity effect. These data suggest a 2-stage model for the interaction of OSCAR with a collagen fibril, with transient, low-affinity interactions initiated by the membrane-distal D1, followed by firm adhesion to the primary binding site in D2. PMID:26552697

  8. Human collagen produced in plants: more than just another molecule.

    PubMed

    Shoseyov, Oded; Posen, Yehudit; Grynspan, Frida

    2014-01-01

    Consequential to its essential role as a mechanical support and affinity regulator in extracellular matrices, collagen constitutes a highly sought after scaffolding material for regeneration and healing applications. However, substantiated concerns have been raised with regard to quality and safety of animal tissue-extracted collagen, particularly in relation to its immunogenicity, risk of disease transmission and overall quality and consistency. In parallel, contamination with undesirable cellular factors can significantly impair its bioactivity, vis-a-vis its impact on cell recruitment, proliferation and differentiation. High-scale production of recombinant human collagen Type I (rhCOL1) in the tobacco plant provides a source of an homogenic, heterotrimeric, thermally stable "virgin" collagen which self assembles to fine homogenous fibrils displaying intact binding sites and has been applied to form numerous functional scaffolds for tissue engineering and regenerative medicine. In addition, rhCOL1 can form liquid crystal structures, yielding a well-organized and mechanically strong membrane, two properties indispensable to extracellular matrix (ECM) mimicry. Overall, the shortcomings of animal- and cadaver-derived collagens arising from their source diversity and recycled nature are fully overcome in the plant setting, constituting a collagen source ideal for tissue engineering and regenerative medicine applications. PMID:23941988

  9. Cervical Softening During Pregnancy: Regulated Changes in Collagen Cross-Linking and Composition of Matricellular Proteins in the Mouse1

    PubMed Central

    Akins, Meredith L.; Luby-Phelps, Katherine; Bank, Ruud A.; Mahendroo, Mala

    2011-01-01

    A greater understanding of the parturition process is essential in the prevention of preterm birth, which occurs in 12.7% of infants born in the United States annually. Cervical remodeling is a critical component of this process. Beginning early in pregnancy, remodeling requires cumulative, progressive changes in the cervical extracellular matrix (ECM) that result in reorganization of collagen fibril structure with a gradual loss of tensile strength. In the current study, we undertook a detailed biochemical analysis of factors in the cervix that modulate collagen structure during early mouse pregnancy, including expression of proteins involved in processing of procollagen, assembly of collagen fibrils, cross-link formation, and deposition of collagen in the ECM. Changes in these factors correlated with changes in the types of collagen cross-links formed and packing of collagen fibrils as measured by electron microscopy. Early in pregnancy there is a decline in expression of two matricellular proteins, thrombospondin 2 and tenascin C, as well as a decline in expression of lysyl hydroxylase, which is involved in cross-link formation. These changes are accompanied by a decline in both HP and LP cross-links by gestation Days 12 and 14, respectively, as well as a progressive increase in collagen fibril diameter. In contrast, collagen abundance remains constant over the course of pregnancy. We conclude that early changes in tensile strength during cervical softening result in part from changes in the number and type of collagen cross-links and are associated with a decline in expression of two matricellular proteins thrombospondin 2 and tenascin C. PMID:21248285

  10. Fibrillization kinetics of insulin solution in an interfacial shearing flow

    NASA Astrophysics Data System (ADS)

    Balaraj, Vignesh; McBride, Samantha; Hirsa, Amir; Lopez, Juan

    2015-11-01

    Although the association of fibril plaques with neurodegenerative diseases like Alzheimer's and Parkinson's is well established, in-depth understanding of the roles played by various physical factors in seeding and growth of fibrils is far from well known. Of the numerous factors affecting this complex phenomenon, the effect of fluid flow and shear at interfaces is paramount as it is ubiquitous and the most varying factor in vivo. Many amyloidogenic proteins have been found to denature upon contact at hydrophobic interfaces due to the self-assembling nature of protein in its monomeric state. Here, fibrillization kinetics of insulin solution is studied in an interfacial shearing flow. The transient surface rheological response of the insulin solution to the flow and its effect on the bulk fibrillization process has been quantified. Minute differences in hydrophobic characteristics between two variants of insulin- Human recombinant and Bovine insulin are found to result in very different responses. Results presented will be in the form of fibrillization assays, images of fibril plaques formed, and changes in surface rheological properties of the insulin solution. The interfacial velocity field, measured from images (via Brewster Angle Microscopy), is compared with computations. Supported by NNX13AQ22G, National Aeronautics and Space Administration.

  11. Polarization-Modulated Second Harmonic Generation Microscopy in Collagen

    SciTech Connect

    Stoller, P C

    2002-09-30

    Collagen is a key structural protein in the body; several pathological conditions lead to changes in collagen. Among imaging modalities that can be used in vivo, second harmonic generation (SHG) microscopy has a key advantage: it provides {approx}1 {micro}m resolution information about collagen structure as a function of depth. A new technique--polarization-modulated SHG--is presented: it permits simultaneous measurement of collagen orientation, of a lower bound on the magnitude of the second order nonlinear susceptibility tensor, and of the ratio of the two independent elements in this tensor. It is applied to characterizing SHG in collagen and to determining effects of biologically relevant changes in collagen structure. The magnitude of the second harmonic signal in two dimensional images varies with position even in structurally homogeneous tissue; this phenomenon is due to interference between second harmonic light generated by neighboring fibrils, which are randomly oriented parallel or anti-parallel to each other. Studies in which focal spot size was varied indicated that regions where fibrils are co-oriented are less than {approx}1.5 {micro}m in diameter. A quartz reference was used to determine the spot size as well as a lower limit (d{sub xxx} > 0.3 pm/V) for the magnitude of the second order nonlinear susceptibility. The ratio of the two independent tensor elements ranged between d{sub XYY}/d{sub XXX} = 0.60 and 0.75. SHG magnitude alone was not useful for identifying structural anomalies in collagenous tissue. Instead, changes in the polarization dependence of SHG were used to analyze biologically relevant perturbations in collagen structure. Changes in polarization dependence were observed in dehydrated samples, but not in highly crosslinked samples, despite significant alterations in packing structure. Complete thermal denaturation and collagenase digestion produced samples with no detectable SHG signal. Collagen orientation was measured in thin

  12. Experimental and Modeling Study of Collagen Scaffolds with the Effects of Crosslinking and Fiber Alignment

    PubMed Central

    Xu, Bin; Chow, Ming-Jay; Zhang, Yanhang

    2011-01-01

    Collagen type I scaffolds are commonly used due to its abundance, biocompatibility, and ubiquity. Most applications require the scaffolds to operate under mechanical stresses. Therefore understanding and being able to control the structural-functional integrity of collagen scaffolds becomes crucial. Using a combined experimental and modeling approach, we studied the structure and function of Type I collagen gel with the effects of spatial fiber alignment and crosslinking. Aligned collagen scaffolds were created through the flow of magnetic particles enmeshed in collagen fibrils to mimic the anisotropy seen in native tissue. Inter- and intra- molecular crosslinking was modified chemically with Genipin to further improve the stiffness of collagen scaffolds. The anisotropic mechanical properties of collagen scaffolds were characterized using a planar biaxial tensile tester and parallel plate rheometer. The tangent stiffness from biaxial tensile test is two to three orders of magnitude higher than the storage moduli from rheological measurements. The biphasic nature of collagen gel was discussed and used to explain the mechanical behavior of collagen scaffolds under different types of mechanical tests. An anisotropic hyperelastic constitutive model was used to capture the characteristics of the stress-strain behavior exhibited by collagen scaffolds. PMID:21876695

  13. FTIR spectro-imaging of collagen scaffold formation during glioma tumor development.

    PubMed

    Noreen, Razia; Chien, Chia-Chi; Chen, Hsiang-Hsin; Bobroff, Vladimir; Moenner, Michel; Javerzat, Sophie; Hwu, Yeukuang; Petibois, Cyril

    2013-11-01

    Evidence has recently emerged that solid and diffuse tumors produce a specific extracellular matrix (ECM) for division and diffusion, also developing a specific interface with microvasculature. This ECM is mainly composed of collagens and their scaffolding appears to drive tumor growth. Although collagens are not easily analyzable by UV-fluorescence means, FTIR imaging has appeared as a valuable tool to characterize collagen contents in tissues, specially the brain, where ECM is normally devoid of collagen proteins. Here, we used FTIR imaging to characterize collagen content changes in growing glioma tumors. We could determine that C6-derived solid tumors presented high content of triple helix after 8-11 days of growth (typical of collagen fibrils formation; 8/8 tumor samples; 91 % of total variance), and further turned to larger α-helix (days 12-15; 9/10 of tumors; 94 % of variance) and β-turns (day 18-21; 7/8 tumors; 97 % of variance) contents, which suggest the incorporation of non-fibrillar collagen types in ECM, a sign of more and more organized collagen scaffold along tumor progression. The growth of tumors was also associated to the level of collagen produced (P < 0.05). This study thus confirms that collagen scaffolding is a major event accompanying the angiogenic shift and faster tumor growth in solid glioma phenotypes. PMID:24068168

  14. Collagen-platelet interactions: recognition and signalling.

    PubMed

    Farndale, Richard W; Siljander, Pia R; Onley, David J; Sundaresan, Pavithra; Knight, C Graham; Barnes, Michael J

    2003-01-01

    The collagen-platelet interaction is central to haemostasis and may be a critical determinant of arterial thrombosis, where subendothelium is exposed after rupture of atherosclerotic plaque. Recent research has capitalized on the cloning of an important signalling receptor for collagen, glycoprotein VI, which is expressed only on platelets, and on the use of collagen-mimetic peptides as specific tools for both glycoprotein VI and integrin alpha 2 beta 1. We have identified sequences, GPO and GFOGER (where O denotes hydroxyproline), within collagen that are recognized by the collagen receptors glycoprotein VI and integrin alpha 2 beta 1 respectively, allowing their signalling properties and specific functional roles to be examined. Triple-helical peptides containing these sequences were used to show the signalling potential of integrin alpha 2 beta 1, and to confirm its important contribution to platelet adhesion. Glycoprotein VI appears to operate functionally on the platelet surface as a dimer, which recognizes GPO motifs that are separated by four triplets of collagen sequence. These advances will allow the relationship between the structure of collagen and its haemostatic activity to be established. PMID:14587284

  15. Alterations of Dermal Connective Tissue Collagen in Diabetes: Molecular Basis of Aged-Appearing Skin

    PubMed Central

    Argyropoulos, Angela J.; Robichaud, Patrick; Balimunkwe, Rebecca Mutesi; Fisher, Gary J.; Hammerberg, Craig; Yan, Yan

    2016-01-01

    Alterations of the collagen, the major structural protein in skin, contribute significantly to human skin connective tissue aging. As aged-appearing skin is more common in diabetes, here we investigated the molecular basis of aged-appearing skin in diabetes. Among all known human matrix metalloproteinases (MMPs), diabetic skin shows elevated levels of MMP-1 and MMP-2. Laser capture microdissection (LCM) coupled real-time PCR indicated that elevated MMPs in diabetic skin were primarily expressed in the dermis. Furthermore, diabetic skin shows increased lysyl oxidase (LOX) expression and higher cross-linked collagens. Atomic force microscopy (AFM) further indicated that collagen fibrils were fragmented/disorganized, and key mechanical properties of traction force and tensile strength were increased in diabetic skin, compared to intact/well-organized collagen fibrils in non-diabetic skin. In in vitro tissue culture system, multiple MMPs including MMP-1 and MM-2 were induced by high glucose (25 mM) exposure to isolated primary human skin dermal fibroblasts, the major cells responsible for collagen homeostasis in skin. The elevation of MMPs and LOX over the years is thought to result in the accumulation of fragmented and cross-linked collagen, and thus impairs dermal collagen structural integrity and mechanical properties in diabetes. Our data partially explain why old-looking skin is more common in diabetic patients. PMID:27104752

  16. Second-harmonic generation scattering directionality predicts tumor cell motility in collagen gels

    NASA Astrophysics Data System (ADS)

    Burke, Kathleen A.; Dawes, Ryan P.; Cheema, Mehar K.; Van Hove, Amy; Benoit, Danielle S. W.; Perry, Seth W.; Brown, Edward

    2015-05-01

    Second-harmonic generation (SHG) allows for the analysis of tumor collagen structural changes throughout metastatic progression. SHG directionality, measured through the ratio of the forward-propagating to backward-propagating signal (F/B ratio), is affected by collagen fibril diameter, spacing, and disorder of fibril packing within a fiber. As tumors progress, these parameters evolve, producing concurrent changes in F/B. It has been recently shown that the F/B of highly metastatic invasive ductal carcinoma (IDC) breast tumors is significantly different from less metastatic tumors. This suggests a possible relationship between the microstructure of collagen, as measured by the F/B, and the ability of tumor cells to locomote through that collagen. Utilizing in vitro collagen gels of different F/B ratios, we explored the relationship between collagen microstructure and motility of tumor cells in a "clean" environment, free of the myriad cells, and signals found in in vivo. We found a significant relationship between F/B and the total distance traveled by the tumor cell, as well as both the average and maximum velocities of the cells. Consequently, one possible mechanism underlying the observed relationship between tumor F/B and metastatic output in IDC patient samples is a direct influence of collagen structure on tumor cell motility.

  17. Second-harmonic generation scattering directionality predicts tumor cell motility in collagen gels.

    PubMed

    Burke, Kathleen A; Dawes, Ryan P; Cheema, Mehar K; Van Hove, Amy; Benoit, Danielle S W; Perry, Seth W; Brown, Edward

    2015-05-01

    Second-harmonic generation (SHG) allows for the analysis of tumor collagen structural changes throughout metastatic progression. SHG directionality, measured through the ratio of the forward-propagating to backward-propagating signal (F/B ratio), is affected by collagen fibril diameter, spacing, and disorder of fibril packing within a fiber. As tumors progress, these parameters evolve, producing concurrent changes in F/B. It has been recently shown that the F/B of highly metastatic invasive ductal carcinoma (IDC) breast tumors is significantly different from less metastatic tumors. This suggests a possible relationship between the microstructure of collagen, as measured by the F/B, and the ability of tumor cells to locomote through that collagen. Utilizing in vitro collagen gels of different F/B ratios, we explored the relationship between collagen microstructure and motility of tumor cells in a “clean” environment, free of the myriad cells, and signals found in in vivo. We found a significant relationship between F/B and the total distance traveled by the tumor cell, as well as both the average and maximum velocities of the cells. Consequently, one possible mechanism underlying the observed relationship between tumor F/B and metastatic output in IDC patient samples is a direct influence of collagen structure on tumor cell motility. PMID:25625899

  18. Nano-imaging collagen by atomic force, near-field and nonlinear microscope

    NASA Astrophysics Data System (ADS)

    Lim, Ken Choong; Tang, Jinkai; Li, Hao; Ng, Boon Ping; Kok, Shaw Wei; Wang, Qijie; Zhang, Ying

    2015-03-01

    As the most abundant protein in the human body, collagen has a very important role in vast numbers of bio-medical applications. The unique second order nonlinear properties of fibrillar collagen make it a very important index in nonlinear optical imaging based disease diagnosis of the brain, skin, liver, colon, kidney, bone, heart and other organs in the human body. The second-order nonlinear susceptibility of collagen has been explored at the macroscopic level and was explained as a volume-averaged molecular hyperpolarizability. However, details about the origin of optical second harmonic signals from collagen fibrils at the molecular level are still not clear. Such information is necessary for accurate interpolation of bio-information from nonlinear optical imaging techniques. The later has shown great potential in collagen based disease diagnosis methodologies. In this paper, we report our work using an atomic force microscope (AFM), near field (SNOM) and nonlinear laser scanning microscope (NLSM) to study the structure of collagen fibrils and other pro-collagen structures.

  19. Collagen V-heterozygous and -null supraspinatus tendons exhibit altered dynamic mechanical behaviour at multiple hierarchical scales.

    PubMed

    Connizzo, Brianne K; Han, Lin; Birk, David E; Soslowsky, Louis J

    2016-02-01

    Tendons function using a unique set of mechanical properties governed by the extracellular matrix and its ability to respond to varied multi-axial loads. Reduction of collagen V expression, such as in classic Ehlers-Danlos syndrome, results in altered fibril morphology and altered macroscale mechanical function in both clinical and animal studies, yet the mechanism by which changes at the fibril level lead to macroscale functional changes has not yet been investigated. This study addresses this by defining the multiscale mechanical response of wild-type, collagen V-heterozygous and -null supraspinatus tendons. Tendons were subjected to mechanical testing and analysed for macroscale properties, as well as microscale (fibre re-alignment) and nanoscale (fibril deformation and sliding) responses. In many macroscale parameters, results showed a dose-dependent response with severely decreased properties in the null group. In addition, both heterozygous and null groups responded to load faster than in wild-type tendons via earlier fibre re-alignment and fibril stretch. However, the heterozygous group exhibited increased fibril sliding, while the null group exhibited no fibril sliding. These studies demonstrate that dynamic responses play an important role in determining overall function and that collagen V is a critical regulator in the development of these relationships. PMID:26855746

  20. Effect of Collagen Nanotopography on Keloid Fibroblast Proliferation and Matrix Synthesis: Implications for Dermal Wound Healing

    PubMed Central

    Muthusubramaniam, Lalitha; Zaitseva, Tatiana; Paukshto, Michael; Martin, George

    2014-01-01

    Keloids are locally exuberant dermal scars characterized by excessive fibroblast proliferation and matrix accumulation. Although treatment strategies include surgical removal and intralesional steroid injections, an effective regimen is yet to be established due to a high rate of recurrence. The regressing center and growing margin of the keloid have different collagen architecture and also differ in the rate of proliferation. To investigate whether proliferation is responsive to collagen topography, keloid, scar, and dermal fibroblasts were cultured on nanopatterned scaffolds varying in collagen fibril diameter and alignment-small and large diameter, aligned and random fibrils, and compared to cells grown on flat collagen-coated substrates, respectively. Cell morphology, proliferation, and expression of six genes related to proliferation (cyclin D1), phenotype (α-smooth muscle actin), and matrix synthesis (collagens I and III, and matrix metalloproteinase-1 and -2) were measured to evaluate cell response. Fibril alignment was shown to reduce proliferation and matrix synthesis in all three types of fibroblasts. Further, keloid cells were found to be most responsive to nanotopography. PMID:24724556

  1. Type VII Collagen Replacement Therapy in Recessive Dystrophic Epidermolysis Bullosa-How Much, How Often?

    PubMed

    South, Andrew P; Uitto, Jouni

    2016-06-01

    Recessive dystrophic epidermolysis bullosa is a devastating blistering disease caused by mutations in the COL7A1 gene, which encodes type VII collagen, the major component of anchoring fibrils. The anchoring fibrils in patients with recessive dystrophic epidermolysis bullosa can be morphologically altered, reduced in number, or absent entirely. There is no specific treatment for this disease, but recent advances in gene, protein replacement, or cell-based therapies, with the purpose of delivering functional type VII collagen to the skin, have shown encouraging results in both preclinical and clinical settings. One critical issue is the stability of type VII collagen in anchoring fibrils, which will ultimately determine the dose and frequency of administration of the missing protein. Kühl et al. attempted to determine the half-life of type VII collagen in the skin, tongue, and esophagus of genetically altered mice that express type VII collagen constitutively, but with its expression abrogated by genetic manipulation. Their results revealed a half-life much shorter than previously anticipated, some 30 days. These findings have implications for strategies to be used for protein replacement therapy, and they also suggest that the basement membrane components at the dermal-epidermal junction are subject to ongoing remodeling and turnover. PMID:27212645

  2. Dynamic mechanical behavior of human dentin and collagen: Methods and properties

    NASA Astrophysics Data System (ADS)

    Ryou, Heonjune

    Experimental evaluations of human coronal dentin and its collagen fibrils were performed by Dynamic Mechanical Analysis (DMA) using nanoindentation and Atomic Force Microscopy (AFM). The primary objectives were to quantify the changes in mechanical behavior of intertubular and peritubular dentin with age, and to evaluate the nanostructure and mechanical behavior of the collagen fibrils. Specimens of coronal dentin were evaluated by nanoDMA using single indents and in scanning mode via scanning probe microscopy. Collagen fibrils from coronal dentin were evaluated using Pulse-Force Mode (PFM) AFM (Peakforce QNM). Nanoindentation results showed that there were no significant differences in the storage modulus or complex modulus between the two age groups (18-25 versus 54-83 yrs) for either the intertubular or peritubular dentin. However, there were significant differences in the dampening behavior between the young and old tissues, as represented in the loss modulus and tanϕ responses. For both the intertubular and peritubular components, the capacity for dampening was significantly lower in the old group. Scanning based nanoDMA showed that the tubules of old dentin exhibit a gradient in elastic behavior, with decrease in elastic modulus from the cuff to the center of tubules filled with newly deposited mineral. AFM results showed that the stiffness of the old dentin fibrils in the peak and trough regions were greater than the young dentin fibrils. In addition, there were significant differences in the dampening behavior between the young and old dentin fibrils, as represented in the energy dissipation, phase angle and loss modulus responses. For both the peak and trough regions, the dissipative capacity was significantly lower in the old dentin fibrils.

  3. The role of polymorphisms of genes encoding collagen IX and XI in lumbar disc disease.

    PubMed

    Janeczko, Łukasz; Janeczko, Magdalena; Chrzanowski, Robert; Zieliński, Grzegorz

    2014-01-01

    The intervertebral disc disease (IDD) is one of the most common musculoskeletal disorders. A number of environment and anthropometric risk factors may contribute to it. The recent reports have suggested the importance of genetic factors, especially these which encode collagen types IX and XI. The allelic variants in the collagen IX genes - COL9A2 (Trp2) and COL9A3 (Trp3) have been identified as genetic risk factors for IDD, because they interfere the cross-linking between collagen types II, IX and XI and result in decreased stability of intervertebral discs. Type XI collagen is a minor component of cartilage collagen fibrils, but it is present in the annulus fibrosus and nucleus pulposus of intervertebral discs. Some studies have shown the association between gene COL11A1 polymorphism c.4603C>T and IDD. The frequency of 4603T allele was significantly higher in the patients with IDD than in the healthy controls. PMID:24636772

  4. Interactions between collagen IX and biglycan measured by atomic force microscopy

    SciTech Connect

    Chen, C.-H.; Yeh, M.-L.; Geyer, Mark; Wang, Gwo-Jaw; Huang, M.-H.; Heggeness, Michael H.; Hoeoek, Magnus; Luo, Z.-P. . E-mail: luo@bcm.tmc.edu

    2006-01-06

    The stability of the lattice-like type II collagen architecture of articular cartilage is paramount to its optimal function. Such stability not only depends on the rigidity of collagen fibrils themselves, but more importantly, on their interconnections. One known interconnection is through type IX and biglycan molecules. However, the mechanical properties of this interaction and its role in the overall stability remain unrevealed. Using atomic force microscopy, this study directly measured the mechanical strength (or the rupture force) of a single bond between collagen IX and biglycan. The results demonstrated that the rupture force of this single bond was 15 pN, which was significantly smaller than those of other known molecule interactions to date. This result suggested that type IX collagen and biglycan interaction may be the weak link in the cartilage collagen architecture, vulnerable to abnormal joint force and associated with disorders such as osteoarthritis.

  5. Oxidative damage to collagen.

    PubMed

    Monboisse, J C; Borel, J P

    1992-01-01

    Extracellular matrix molecules, such as collagens, are good targets for oxygen free radicals. Collagen is the only protein susceptible to fragmentation by superoxide anion as demonstrated by the liberation of small 4-hydroxyproline-containing-peptides. It seems likely that hydroxyl radicals in the presence of oxygen cleave collagen into small peptides, and the cleavage seems to be specific to proline or 4-hydroxyproline residues. Hydroxyl radicals in the absence of oxygen or hypochlorous acid do not induce fragmentation of collagen molecules, but they trigger a polymerization of collagen through the formation of new cross-links such as dityrosine or disulfure bridges. Moreover, these cross-links can not explain the totality of high molecular weight components generated under these experimental conditions, and the nature of new cross-links induced by hydroxyl radicals or hypochlorous acid remains unclear. PMID:1333311

  6. Evaluation of cell binding to collagen and gelatin: a study of the effect of 2D and 3D architecture and surface chemistry.

    PubMed

    Davidenko, Natalia; Schuster, Carlos F; Bax, Daniel V; Farndale, Richard W; Hamaia, Samir; Best, Serena M; Cameron, Ruth E

    2016-10-01

    Studies of cell attachment to collagen-based materials often ignore details of the binding mechanisms-be they integrin-mediated or non-specific. In this work, we have used collagen and gelatin-based substrates with different dimensional characteristics (monolayers, thin films and porous scaffolds) in order to establish the influence of composition, crosslinking (using carbodiimide) treatment and 2D or 3D architecture on integrin-mediated cell adhesion. By varying receptor expression, using cells with collagen-binding integrins (HT1080 and C2C12 L3 cell lines, expressing α2β1, and Rugli expressing α1β1) and a parent cell line C2C12 with gelatin-binding receptors (αvβ3 and α5β1), the nature of integrin binding sites was studied in order to explain the bioactivity of different protein formulations. We have shown that alteration of the chemical identity, conformation and availability of free binding motifs (GxOGER and RGD), resulting from addition of gelatin to collagen and crosslinking, have a profound effect on the ability of cells to adhere to these formulations. Carbodiimide crosslinking ablates integrin-dependent cell activity on both two-dimensional and three-dimensional architectures while the three-dimensional scaffold structure also leads to a high level of non-specific interactions remaining on three-dimensional samples even after a rigorous washing regime. This phenomenon, promoted by crosslinking, and attributed to cell entrapment, should be considered in any assessment of the biological activity of three-dimensional substrates. Spreading data confirm the importance of integrin-mediated cell engagement for further cell activity on collagen-based compositions. In this work, we provide a simple, but effective, means of deconvoluting the effects of chemistry and dimensional characteristics of a substrate, on the cell activity of protein-derived materials, which should assist in tailoring their biological properties for specific tissue engineering

  7. Newly identified interfibrillar collagen crosslinking suppresses cell proliferation and remodelling.

    PubMed

    Marelli, Benedetto; Le Nihouannen, Damien; Hacking, S Adam; Tran, Simon; Li, Jingjing; Murshed, Monzur; Doillon, Charles J; Ghezzi, Chiara E; Zhang, Yu Ling; Nazhat, Showan N; Barralet, Jake E

    2015-06-01

    Copper is becoming recognised as a key cation in a variety of biological processes. Copper chelation has been studied as a potential anti-angiogenic strategy for arresting tumour growth. Conversely the delivery of copper ions and complexes in vivo can elicit a pro-angiogenic effect. Previously we unexpectedly found that copper-stimulated intraperitoneal angiogenesis was accompanied by collagen deposition. Here, in hard tissue, not only was healing accelerated by copper, but again enhanced deposition of collagen was detected at 2 weeks. Experiments with reconstituted collagen showed that addition of copper ions post-fibrillogenesis rendered plastically-compressed gels resistant to collagenases, enhanced their mechanical properties and increased the denaturation temperature of the protein. Unexpectedly, this apparently interfibrillar crosslinking was not affected by addition of glucose or ascorbic acid, which are required for crosslinking by advanced glycation end products (AGEs). Fibroblasts cultured on copper-crosslinked gels did not proliferate, whereas those cultured with an equivalent quantity of copper on either tissue culture plastic or collagen showed no effect compared with controls. Although non-proliferative, fibroblasts grown on copper-cross-linked collagen could migrate, remained metabolically active for at least 14 days and displayed a 6-fold increase in Mmps 1 and 3 mRNA expression compared with copper-free controls. The ability of copper ions to crosslink collagen fibrils during densification and independently of AGEs or Fenton type reactions is previously unreported. The effect on MMP susceptibility of collagen and the dramatic change in cell behaviour on this crosslinked ECM may contribute to shedding some light on unexplained phenomena as the apparent benefit of copper complexation in fibrotic disorders or the enhanced collagen deposition in response to localised copper delivery. PMID:25907046

  8. Characterizing matrix remodeling in collagen gels using optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Levitz, David; Hinds, Monica T.; Hanson, Stephen R.; Jacques, Steven L.

    2010-02-01

    Optical coherence tomography (OCT) has shown promise at non-destructively characterizing engineered tissues such as collagen gels. However, as the collagen gels develop, the OCT images lose contrast of structures as the gels develop, making visual assessment difficult. Our group proposed quantitatively characterizing these gels by fitting the optical properties from the OCT signals. In this paper, we imaged collagen gels seeded with smooth muscle cells (SMCs) over a 5-day period and used the data to measure their optical properties. Our results showed that over time, the reflectivity of the samples increased 10-fold, corresponding to a decrease in anisotropy factor g, without much change in the scattering coefficient μs. Overall, the optical properties appeared to be dominated by scattering from the collagen matrix, not the cells. However, SMCs remodeled the collagen matrix, and this collagen remodeling by the cells is what causes the observed changes in optical properties. Moreover, the data showed that the optical properties were sensitive to the activity of matrix metalloproteinases (MMPs), enzymes that break down local collagen fibrils into smaller fragments. Blocking MMPs in the SMC gels greatly impeded both the remodeling process and change in optical properties at day 5. Treating day 1 acellular gels with MMP-8 for 3 hr managed to partially reproduce the remodeling observed in SMC gels at day 5. Altogether, we conclude that matrix remodeling in general, and MMPs specifically, greatly affect the local optical properties of the sample, and OCT is a unique tool that can assess MMP activity in collagen gels both non-destructively and label free.

  9. [Anticoagulation in atrial fibrillation].

    PubMed

    Schwarz, M; Bode, Ch

    2008-10-01

    In this overview the actual guideline-recommendations for anticoagulation in atrial fibrillation and the problems of the currently available therapy are discussed. Furthermore an outlook over future developments in this field is given. Effective anticoagulation can prohibit thrombembolic events and is thus essential for the prognosis of patients suffering from atrial fibrillation. Until now vitamin-K-antagonists (VKAs) and acetylsalicylic acid (ASA) are available for oral anticoagulation in these patients. VKAs demonstrate a satisfying efficiency combined with rather high bleeding hazard. ASA on the other hand allows only moderate risk reduction with minimal side effects. Thus the guidelines recommend anticoagulation tailored to the individual risk, which can be evaluated by the CHADS2-Score. New therapeutic strategies, like the factor Xa inhibitor rivaroxaban or the factor II inhibitor dabigatran, are actually evaluated in phase III studies. These drugs bear the hope of higher efficiency combined with improved safety and much more comfortable use in the daily practice (e. g. no need for INR measurement, no dose adaptation). PMID:18836647

  10. Hyperuricemia and Atrial Fibrillation.

    PubMed

    Maharani, Nani; Kuwabara, Masanari; Hisatome, Ichiro

    2016-07-27

    The importance of atrial fibrillation (AF) as a cause of mortality and morbidity has prompted research on its pathogenesis and treatment. Recognition of AF risk factors is essential to prevent it and reduce the risk of death. Hyperuricemia has been widely accepted to be associated with the incidence of paroxysmal or persistent AF, as well as to the risk of AF in post cardiovascular surgery patients. The possible explanations for this association have been based on their relation with either oxidative stress or inflammation. To investigate the link between hyperuricemia and AF, it is necessary to refer to hyperuricemia-induced atrial remodeling. So far, both ionic channel and structural remodeling caused by hyperuricemia might be plausible explanations for the occurrence of AF. Inhibition of xanthine oxidase and nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase, or the use of antioxidants, along with serum uric acid (SUA) level reduction to prevent inflammation, might be useful. Uric acid transporters (UATs) play a key role in the regulation of intracellular uric acid concentration. Intracellular rather than serum uric acid level is considered more important for the pathogenesis of AF. Identification of UATs expressed in cells is thus important, and targeting UATs might become a potential strategy to reduce the risk of hyperuricemia-induced atrial fibrillation. PMID:27396561

  11. Type VII Collagen Expression in the Human Vitreoretinal Interface, Corpora Amylacea and Inner Retinal Layers

    PubMed Central

    Wullink, Bart; Pas, Hendri H.; Van der Worp, Roelofje J.; Kuijer, Roel; Los, Leonoor I.

    2015-01-01

    Type VII collagen, as a major component of anchoring fibrils found at basement membrane zones, is crucial in anchoring epithelial tissue layers to their underlying stroma. Recently, type VII collagen was discovered in the inner human retina by means of immunohistochemistry, while proteomic investigations demonstrated type VII collagen at the vitreoretinal interface of chicken. Because of its potential anchoring function at the vitreoretinal interface, we further assessed the presence of type VII collagen at this site. We evaluated the vitreoretinal interface of human donor eyes by means of immunohistochemistry, confocal microscopy, immunoelectron microscopy, and Western blotting. Firstly, type VII collagen was detected alongside vitreous fibers6 at the vitreoretinal interface. Because of its known anchoring function, it is likely that type VII collagen is involved in vitreoretinal attachment. Secondly, type VII collagen was found within cytoplasmic vesicles of inner retinal cells. These cells resided most frequently in the ganglion cell layer and inner plexiform layer. Thirdly, type VII collagen was found in astrocytic cytoplasmic inclusions, known as corpora amylacea. The intraretinal presence of type VII collagen was confirmed by Western blotting of homogenized retinal preparations. These data add to the understanding of vitreoretinal attachment, which is important for a better comprehension of common vitreoretinal attachment pathologies. PMID:26709927

  12. Collagenous Colitis and Spondylarthropathy

    PubMed Central

    Ben Abdelghani, Kaouther; Sahli, Hana; Souabni, Leila; Chekili, Selma; Belhadj, Salwa; Kassab, Selma; Laatar, Ahmed; Zakraoui, Leith

    2012-01-01

    Collagenous colitis is a recent cause of chronic diarrhea. Cooccurrence with spondylarthropathy is rare. We describe two cases: one man and one woman of 33 and 20 years old were suffering from spondylarthropathy. They then developed collagenous colitis, 4 and 14 years after the onset of spondylarthropathy. The diagnosis was based on histological features. A sicca syndrome and vitiligo were observed with the female cas