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Fibrils
Kinds of Fibrils Terms modified by Fibrils Selected AbstractsAssembly of 1D Nanostructures into Sub-micrometer Diameter Fibrils with Controlled and Variable Length by Dielectrophoresis,ADVANCED MATERIALS, Issue 16 2003J. Tang A dielectrophoresis method for manipulation, alignment, and assembly of 1D nanostructures is reported. Processed carbon nanotubes dispersed in water are assembled into micro-electrodes and fibrils of variable and controllable length from ,,1 ,m to over 1 cm (see Figure). The method allows of parallel fabrication, and the fibrils could find applications as probes for scanning probe microscopes, electrodes, and conducting wires. [source] Assemblies of Metal Nanoparticles and Self-Assembled Peptide Fibrils,Formation of Double Helical and Single-Chain Arrays of Metal Nanoparticles,ADVANCED MATERIALS, Issue 11 2003X. Fu Double helical and single-chain arrays of Au and Pd metal nanoclusters have been achieved by assembling the corresponding metal nanoparticles on peptide nanofibril templates at different pH values. The assembled metal nanoparticles are useful for studying the superstructures of the peptide nanofibrils. Pd nanowires of different shapes have also been prepared by varying the amount of metal deposition. [source] Soluble protein oligomers as emerging toxins in alzheimer's and other amyloid diseasesIUBMB LIFE, Issue 4-5 2007Sergio T. Ferreira Abstract Amyloid diseases are a group of degenerative disorders characterized by cell/tissue damage caused by toxic protein aggregates. Abnormal production, processing and/or clearance of misfolded proteins or peptides may lead to their accumulation and to the formation of amyloid aggregates. Early histopathological investigation of affected organs in different amyloid diseases revealed the ubiquitous presence of fibrillar protein aggregates forming large deposits known as amyloid plaques. Further in vitro biochemical and cell biology studies, as well as studies using transgenic animal models, provided strong support to what initially seemed to be a solid concept, namely that amyloid fibrils played crucial roles in amyloid pathogenesis. However, recent studies describing tissue-specific accumulation of soluble protein oligomers and their strong impact on cell function have challenged the fibril hypothesis and led to the emergence of a new view: Fibrils are not the only toxins derived from amyloidogenic proteins and, quite possibly, not the most important ones with respect to disease etiology. Here, we review some of the recent findings and concepts in this rapidly developing field, with emphasis on the involvement of soluble oligomers of the amyloid-, peptide in the pathogenesis of Alzheimer's disease. Recent studies suggesting that soluble oligomers from different proteins may share common mechanisms of cytotoxicity are also discussed. Increased understanding of the cellular toxic mechanisms triggered by protein oligomers may lead to the development of rational, effective treatments for amyloid disorders. IUBMB Life, 59: 332-345, 2007 [source] Modeling of Bovine Type-I Collagen Fibrils: Interaction with Pickling and Retanning AgentsMACROMOLECULAR BIOSCIENCE, Issue 2 2007Rosa E. Bulo Abstract 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, CaCl2, and Na2SO4) 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 CaCl2 and Na2SO4, due to weak Ca2+/Cl, and strong SO/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. [source] Probing Dynamics within Amyloid Fibrils Using a Novel Capping Method,ANGEWANDTE CHEMIE, Issue 31 2009Geoffrey Twist and shout: Die Dynamik von Amyloidfibrillen wird mit Lösungs-NMR-Spektroskopie analysiert (siehe 1H- 15N-HSQC-NMR-Spektren von Wildtyp-,2 -Mikroglobulin und einer Variante mit verlängerter N-terminaler Region). Ein neuartiges Verfahren wird angewendet, das die Herkunft der NMR-Signale sicherstellt und das Molekülrecycling von Fibrillen berücksichtigt. [source] Characterization of Amyloid Fibrils of Human ,-2-Microglobulin by High-Resolution Magic-Angle Spinning NMRCHEMBIOCHEM, Issue 13 2010Lukasz Skora Dr. It's a kind of magic: By using high-resolution magic-angle spinning NMR spectroscopy in combination with hydrogen/deuterium exchange measurements we have shown that at least 18 residues at the N and C termini of ,-2-microglobulin aggregated into amyloid fibrils retain a large degree of mobility occurring on different timescales. This study provides insight into the structural architecture of amyloid fibrils of human ,-2-microglobulin. [source] ChemInform Abstract: [18F]-Labeled Isoindol-1-one and Isoindol-1,3-dione Derivatives as Potential PET Imaging Agents for Detection of ,-Amyloid Fibrils.CHEMINFORM, Issue 10 2009Ji Hoon Lee Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source] ChemInform Abstract: Synthesis of Functionalized Benzoxazoles and Their Binding Affinities to A,42 Fibrils.CHEMINFORM, Issue 5 2009Young Shin Chun Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source] Self-Assembly of Amylin(20,29) Amide-Bond Derivatives into Helical Ribbons and Peptide Nanotubes rather than FibrilsCHEMISTRY - A EUROPEAN JOURNAL, Issue 14 2006Ronald C. Elgersma Abstract Uncontrolled aggregation of proteins or polypeptides can be detrimental for normal cellular processes in healthy organisms. Proteins or polypeptides that form these amyloid deposits differ in their primary sequence but share a common structural motif: the (anti)parallel , sheet. A well-accepted approach for interfering with ,-sheet formation is the design of soluble ,-sheet peptides to disrupt the hydrogen-bonding network; this ultimately leads to the disassembly of the aggregates or fibrils. Here, we describe the synthesis, spectroscopic analysis, and aggregation behavior, imaged by electron microscopy, of several backbone-modified amylin(20,29) derivatives. It was found that these amylin derivatives were not able to form fibrils and to some extent were able to inhibit fibril growth of native amylin(20,29). However, two of the amylin peptides were able to form large supramolecular assemblies, like helical ribbons and peptide nanotubes, in which ,-sheet formation was clearly absent. This was quite unexpected since these peptides have been designed as soluble ,-sheet breakers for disrupting the characteristic hydrogen-bonding network of (anti)parallel , sheets. The increased hydrophobicity and the presence of essential amino acid side chains in the newly designed amylin(20,29) derivatives were found to be the driving force for self-assembly into helical ribbons and peptide nanotubes. This example of controlled and desired peptide aggregation may be a strong impetus for research on bionanomaterials in which special shapes and assemblies are the focus of interest. [source] Structural disorder in amyloid fibrils: its implication in dynamic interactions of proteinsFEBS JOURNAL, Issue 19 2009P. Tompa Proteins are occasionally converted from their normal soluble state to highly ordered fibrillar aggregates (amyloids), which give rise to pathological conditions that range from neurodegenerative disorders to systemic amyloidoses. Recent methodological advances in solid-state NMR and EPR spectroscopy have enabled determination of the 3D structure of several amyloids at residue-level resolution. The general picture that emerges is that amyloids constitute parallel , sheets, in which individual polypeptide chains run roughly perpendicular to the major axis of the fibril and are stacked in-register. Thus, the unifying theme of amyloid formation is the structural transition from an initial globular or intrinsically disordered state to a highly ordered regular form. In this minireview, we show that this description is somewhat oversimplified, because part of the polypeptide chain in the amyloid remains intrinsically disordered and does not become part of the ordered core. As demonstrated through examples such as the amyloids of ,-synuclein and A, peptide and the yeast prions HET-s and Ure2p, these disordered segments are depleted in amino acids NQFYV and are enriched in DEKP. They are also significantly more charged and have a higher predicted disordered value than segments in the cross-, core. We suggest that structural disorder in amyloid is a special case of ,fuzziness', i.e. disorder in the bound state that may serve different functions, such as the accommodation of destabilizing residues and the mediation of secondary interactions between protofibrils. [source] Consortium for osteogenesis imperfecta mutations in the helical domain of type I collagen: regions rich in lethal mutations align with collagen binding sites for integrins and proteoglycans,,HUMAN MUTATION, Issue 3 2007Joan C. Marini Abstract Osteogenesis imperfecta (OI) is a generalized disorder of connective tissue characterized by fragile bones and easy susceptibility to fracture. Most cases of OI are caused by mutations in type I collagen. We have identified and assembled structural mutations in type I collagen genes (COL1A1 and COL1A2, encoding the pro,1(I) and pro,2(I) chains, respectively) that result in OI. Quantitative defects causing type I OI were not included. Of these 832 independent mutations, 682 result in substitution for glycine residues in the triple helical domain of the encoded protein and 150 alter splice sites. Distinct genotype,phenotype relationships emerge for each chain. One-third of the mutations that result in glycine substitutions in ,1(I) are lethal, especially when the substituting residues are charged or have a branched side chain. Substitutions in the first 200 residues are nonlethal and have variable outcome thereafter, unrelated to folding or helix stability domains. Two exclusively lethal regions (helix positions 691,823 and 910,964) align with major ligand binding regions (MLBRs), suggesting crucial interactions of collagen monomers or fibrils with integrins, matrix metalloproteinases (MMPs), fibronectin, and cartilage oligomeric matrix protein (COMP). Mutations in COL1A2 are predominantly nonlethal (80%). Lethal substitutions are located in eight regularly spaced clusters along the chain, supporting a regional model. The lethal regions align with proteoglycan binding sites along the fibril, suggesting a role in fibril,matrix interactions. Recurrences at the same site in ,2(I) are generally concordant for outcome, unlike ,1(I). Splice site mutations comprise 20% of helical mutations identified in OI patients, and may lead to exon skipping, intron inclusion, or the activation of cryptic splice sites. Splice site mutations in COL1A1 are rarely lethal; they often lead to frameshifts and the mild type I phenotype. In ,2(I), lethal exon skipping events are located in the carboxyl half of the chain. Our data on genotype,phenotype relationships indicate that the two collagen chains play very different roles in matrix integrity and that phenotype depends on intracellular and extracellular events. Hum Mutat 28(3), 209,221, 2007. Published 2006 Wiley-Liss, Inc. [source] Pretibial epidermolysis bullosa: is this case a new subtype with loss of types IV and VII collagen?INTERNATIONAL JOURNAL OF DERMATOLOGY, Issue 8 2009Hong-sun Lee MD Pretibial epidermolysis bullosa (PEB) is an extremely rare subtype of dominant dystrophic epidermolysis bullosa (DDEB), in which recurrent blistering with scarring predominantly involves the pretibial skin. Nail dystrophy, albopapuloid lesions, and hypertrophic scars may also occur. In PEB, immunohistochemical and electron microscopic studies demonstrate the complete or partial loss of the anchoring fibril (AF) in the basement membrane zone, suggesting disturbed synthesis or excessive degradation of collagen VII, the main component of AF. Interestingly, we report a case of PEB with unusual results of joint loss of types IV and VII collagen. [source] Sub-micron spongiform porosity is the major ultra-structural alteration occurring in archaeological boneINTERNATIONAL JOURNAL OF OSTEOARCHAEOLOGY, Issue 6 2002G. Turner-Walker Abstract Total pore volume and pore size distribution are indicators of the degree of post-mortem modification of bone. Direct measurements of pore size distribution in archaeological bones using mercury intrusion porosimetry (HgIP) and back scattered scanning electron microscopy (BSE-SEM) reveal a common pattern in the changes seen in degraded bone as compared to modern samples. The estimates of pore size distribution from HgIP and direct measurement from the BSE-SEM images show remarkable correspondence. The coupling of these two independent approaches has allowed the diagenetic porosity changes in human archaeological bone in the >0.01 µm range to be directly imaged, and their relationship to pre-existing physiological pores to be explored. The increase in porosity in the archaeological bones is restricted to two discrete pore ranges. The smaller of these two ranges (0.007,0.1 µm) lies in the range of the collagen fibril (0.1 µm diameter) and is presumably formed by the loss of collagen, whereas the larger pore size distribution is evidence of direct microbial alteration of the bone. HgIP has great potential for the characterization of microbial and chemical alteration of bone. Copyright © 2002 John Wiley & Sons, Ltd. [source] Unraveling the mysteries of protein folding and misfoldingIUBMB LIFE, Issue 12 2008Heath Ecroyd Abstract This mini-review focuses on the processes and consequences of protein folding and misfolding. The latter process often leads to protein aggregation and precipitation with the aggregates adopting either highly ordered (amyloid fibril) or disordered (amorphous) forms. In particular, the amyloid fibril is discussed because this form has gained considerable notoriety due to its close links to a variety of debilitating diseases including Alzheimer's, Parkinson's, Huntington's, and Creutzfeldt-Jakob diseases, and type-II diabetes. In each of these diseases a different protein forms fibrils, yet the fibrils formed have a very similar structure. The mechanism by which fibrils form, fibril structure, and the cytotoxicity associated with fibril formation are discussed. The generic nature of amyloid fibril structure suggests that a common target may be accessible to treat amyloid fibril-associated diseases. As such, the ability of some molecules, for example, the small heat-shock family of molecular chaperone proteins, to inhibit fibril formation is of interest due to their therapeutic potential. © 2008 IUBMB IUBMB Life, 60(12): 769,774, 2008 [source] pH dependent self assembly of ,-amyloid(10-35) and ,-amyloid(10-35)-PEG3000JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 3-1 2000P. Thiyagarajan Small angle neutron and x-ray scattering (SANS/SAXS) studies were conducted on the structure of the aggregates formed from both the truncated model peptide ,-Amyloid(10-35) (A,10-35) and a block copolymer ,-Amyloid(10-35)-PEG3000 (A,10-35 -PEG) in D2O at pHs from 3.0 to 7.0. These studies indicate that A,10-35 aggregates into rod-like particles (fibril) and their radii are strongly dependent on the pH of the solution. The fibril-fibril association in A,10-35 solutions is less at pH < 5.6, but becomes larger at higher pH. A,10-35 -PEG also assembles into rod-like particles whose radius is larger by about 30 Å than that for A,10-35 fibril at pH 4.2, while it is about 23 Å larger at higher pH. Contrast matching SAXS/SANS experiments that eliminate the coherent scattering from PEG reveal that PEG moiety is located at the periphery of the fibril. Also the mass per unit length of the peptide portion is similar for both A,10-35 and A,10-35 fibrils at pH 5.6. The mass per unit length of the rods from SANS provides key information on the packing of A,10-35 peptides in the fibril. [source] Pyridinium Cross-Links in Bone of Patients with Osteogenesis Imperfecta: Evidence of a Normal Intrafibrillar Collagen PackingJOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2000Ruud A. Bank Ph.D. Abstract The brittleness of bone in patients with osteogenesis imperfecta (OI) has been attributed to an aberrant collagen network. However, the role of collagen in the loss of tissue integrity has not been well established. To gain an insight into the biochemistry and structure of the collagen network, the cross-links hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP) and the level of triple helical hydroxylysine (Hyl) were determined in bone of OI patients (types I, III, and IV) as well as controls. The amount of triple helical Hyl was increased in all patients. LP levels in OI were not significantly different; in contrast, the amount of HP (and as a consequence the HP/LP ratio and the total pyridinoline level) was significantly increased. There was no relationship between the sum of pyridinolines and the amount of triple helical Hyl, indicating that lysyl hydroxylation of the triple helix and the telopeptides are under separate control. Cross-linking is the result of a specific three-dimensional arrangement of collagens within the fibril; only molecules that are correctly aligned are able to form cross-links. Inasmuch as the total amount of pyridinoline cross-links in OI bone is similar to control bone, the packing geometry of intrafibrillar collagen molecules is not disturbed in OI. Consequently, the brittleness of bone is not caused by a disorganized intrafibrillar collagen packing and/or loss of cross-links. This is an unexpected finding, because mutant collagen molecules with a random distribution within the fibril are expected to result in disruptions of the alignment of neighboring collagen molecules. Pepsin digestion of OI bone revealed that collagen located at the surface of the fibril had lower cross-link levels compared with collagen located at the inside of the fibril, indicating that mutant molecules are not distributed randomly within the fibril but are located preferentially at the surface of the fibril. [source] Modeling of Bovine Type-I Collagen Fibrils: Interaction with Pickling and Retanning AgentsMACROMOLECULAR BIOSCIENCE, Issue 2 2007Rosa E. Bulo Abstract 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, CaCl2, and Na2SO4) 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 CaCl2 and Na2SO4, due to weak Ca2+/Cl, and strong SO/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. [source] Microstructure analysis of high performance fibers in compressionPOLYMER ENGINEERING & SCIENCE, Issue 3 2003Wansoo Huh In order to understand the nature of kink band formation in high performance fibers, the compression behavior of pitch and PAN-based carbon fibers (P75S, T-50, T-300, and GY-70), and of polymeric fibers (DuPont Kevlar; PBZT; and PBO) was measured using a micro-scale compression apparatus in an optical microscope. With increasing compressive strain, kink band formation was observed and the number of kink bands per unit length (referred to as kink band density) was determined. By extrapolating to zero kink band density, the critical compressive strain for rigid-rod polymeric fibers was obtained and compared to that of the carbon fiber. Using the Euler buckling equation, a fundamental dimension of the buckling element for the compression of PBO and PBZT fibers was calculated to be a 0.42,0.57 µm diameter fibril, and not the smaller diameter microfibrils. [source] Cooperative hydrogen bonding in amyloid formationPROTEIN SCIENCE, Issue 4 2007Kiril Tsemekhman Abstract Amyloid diseases, including Alzheimer's and prion diseases, are each associated with unbranched protein fibrils. Each fibril is made of a particular protein, yet they share common properties. One such property is nucleation-dependent fibril growth. Monomers of amyloid-forming proteins can remain in dissolved form for long periods, before rapidly assembly into fibrils. The lag before growth has been attributed to slow kinetics of formation of a nucleus, on which other molecules can deposit to form the fibril. We have explored the energetics of fibril formation, based on the known molecular structure of a fibril-forming peptide from the yeast prion, Sup35, using both classical and quantum (density functional theory) methods. We find that the energetics of fibril formation for the first three layers are cooperative using both methods. This cooperativity is consistent with the observation that formation of amyloid fibrils involves slow nucleation and faster growth. [source] Helical polyacetylene,Origins and synthesisTHE CHEMICAL RECORD, Issue 6 2008Kazuo Akagi Abstract We present the origins and synthesis of helical polyacetylene (H-PA) by focusing on its peculiar spiral morphology. Interfacial polymerization of acetylene was carried out in an asymmetric reaction field consisting of chiral nematic liquid crystal (N*-LC) and Ziegler,Natta catalyst. As the N*-LC is composed of nematic liquid crystal and a chiral compound such as a binaphthyl derivative with either the R - or S -configuration, the screw directions of the polyacetylene chain and fibril bundle,and even the spiral morphology,are rigorously controlled by the chirality of the selected compound. Interestingly, the screw directions of the fibril and the bundle in H-PA were found to be opposite to that of N*-LC. It is worthwhile to emphasize that the hierarchical spiral morphology involving the primary to higher order structure is generated in a synthetic polymer such as polyacetylene by using N*-LC as an asymmetric polymerization solvent. © 2008 The Japan Chemical Journal Forum and Wiley Periodicals, Inc. Chem Rec 8: 395,406; 2008: Published online in Wiley InterScience (www.interscience.wiley.com) DOI 10.1002/tcr.20163 [source] Combined role of type IX collagen and cartilage oligomeric matrix protein in cartilage matrix assembly: Cartilage oligomeric matrix protein counteracts type IX collagen,induced limitation of cartilage collagen fibril growth in mouse chondrocyte culturesARTHRITIS & RHEUMATISM, Issue 12 2009K. Blumbach Objective Defects in the assembly and composition of cartilage extracellular matrix are likely to result in impaired matrix integrity and increased susceptibility to cartilage degeneration. The aim of this study was to determine the functional interaction of the collagen fibril,associated proteins type IX collagen and cartilage oligomeric matrix protein (COMP) during cartilage matrix formation. Methods Primary chondrocytes from mice deficient in type IX collagen and COMP (double-deficient) were cultured in monolayer or alginate beads. Anchorage of matrix proteins, proteoglycan and collagen content, collagen crosslinks, matrix metalloproteinase activity, and mechanical properties of the matrix were measured. Electron microscopy was used to study the formation of fibrillar structures. Results In cartilage lacking both type IX collagen and COMP, matrilin 3 showed decreased matrix anchorage. Less matrilin 3 was deposited in the matrix of double-deficient chondrocytes, while larger amounts were secreted into the medium. Proteoglycans were less well retained in the matrix formed in alginate cultures, while collagen deposition was not significantly affected. Electron microscopy revealed similar cartilage collagen fibril diameters in the cultures of double-deficient and wild-type chondrocytes. In contrast, a larger fibril diameter was observed in the matrix of chondrocytes deficient in only type IX collagen. Conclusion Our results show that type IX collagen and COMP are involved in matrix assembly by mediating the anchorage and regulating the distribution of other matrix macromolecules such as proteoglycans and matrilins and have counteracting effects on collagen fibril growth. Loss of type IX collagen and COMP leads to matrix aberrations that may make cartilage more susceptible to degeneration. [source] A chromospheric dark-cored fibril in Ca II IR spectraASTRONOMISCHE NACHRICHTEN, Issue 6 2010C. Beck Abstract We investigate the thermodynamical and magnetic properties of a "dark-cored" fibril seen in the chromospheric Ca II IR line at 854.2 nm to determine the physical process behind its appearance. We analyse a time series of spectropolarimetric observations obtained in the Ca II IR line at 854.2 nm and the photospheric Fe I line at 630.25 nm. We simultaneously invert the spectra in both wavelength ranges with the SIR code to obtain the temperature and velocity stratification with height in the solar atmosphere and the magnetic field properties in the photosphere. The structure can be clearly traced in the line-of-sight (LOS) velocity and the temperature maps. It connects from a small pore with kG fields to a region with lower field strength. The flow velocity and the temperature indicate that the height of the structure increases with increasing distance from the inner footpoint. The Stokes V signal of 854.2 nm shows a Doppler-shifted polarization signal with the same displacement as in the intensity profile, indicating that the supersonic flow seen in the LOS velocity is located within magnetized plasma. We conclude that the chromospheric dark-cored fibril traces a siphon flow along magnetic field lines, driven by the gas pressure difference caused by the higher magnetic field strength at the inner footpoint. We suggest that fast flows guided by the magnetic field lead to the appearance of "dark-cored" fibrils in intensity images. Although the observations included the determination of the polarization signal in the chromospheric Ca II IR line, the signal could not be analysed quantitatively due to the low S/N. Chromospheric polarimetry will thus require telescopes of larger aperture able to collect a sufficient number of photons for a reliable determination of polarization in deep and only weakly polarized spectral lines (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] 2142: Ulrastructural features of keratoconus cornea after cross-linking by riboflavin/UVAACTA OPHTHALMOLOGICA, Issue 2010S AKHTAR Purpose In the present studies we assess the effects of collagen cross-linking on ultrastructure organisation of the corneal stroma of keratoconus human corneas. Methods One normal, one keratoconus (KC) and three cross-linked keratoconus corneas were analysed. One was treated with standard cross linking (SXL) and two with trans-epithelial collagen cross linking (TEXL). Penetrating keratoplasty was performed three months after treatment. All samples were fixed in 2.5% glutaraldehyde containing cuprolinic blue in sodium acetate buffer and processed for electron microscopy. Results The structure of SXL corneas was very similar to normal corneas in their hemidesmosomes, basement membrane (BM), Bowman's layer (BW) and stromal lamellae that were not undulated. The architecture of TEXL corneas presented some differences. The BM was thick with degenerated hemidesmosomes. Bowman's layer was disorganised at some places and replaced by thin filaments forming pannus. There were thin undulating lamellae in anterior, middle and posterior stroma. The keratocytes were embedded between undulating lamellae. Large amounts of abnormal PGs were attached around collagen fibrils. The parallel running lamellae were very thin. In some parts of the anterior stroma collagen fibrils were oriented (running) in random directions instead of running parallel. There were some parts of the stroma which showed a normal appearance. Conclusion The present studies demonstrate that corneal cross-linking leads to modifications in keratocytes and in the organisation of collagen fibril. The morphological changes might be correlated to the process of increase in biomechanical stability although there are differences between stromal structures treated by standard and trans [source] Remodelling of collagen fibrils and proteoglycans in the zebrafish cornea during developmentACTA OPHTHALMOLOGICA, Issue 2007S AKHTAR Purpose: Collagen fibrils and proteoglycans are the main components of the corneal extracellular matrix and corneal transparency depends crucially on their proper organisation. We investigated their formation and arrangement in the developing cornea of the zebrafish, a major model of vertebrate development and genetic disease. Methods: We employed thin-section electron microscopy to investigate the ultrastructure of the zebrafish cornea at different stages of development. Results: Layering of the zebrafish cornea into an epithelium, Bowman's layer, stroma and endothelium was observed by 72 hours post-fertilization. At this stage, the stroma contained orthogonally arranged collagen fibrils and small proteoglycans. The density of proteoglycans increased gradually throughout subsequent development. In the stroma of 2 week old larvae, the collagen fibrils were organized into thin lamellae for the first time and were separated by very large, randomly distributed proteoglycans. At 4 weeks, a regular arrangement of proteoglycans around the collagen fibrils was observed for the first time and the lamellae also thickened. Conclusions: This is the first report of collagen fibril and proteoglycan development in the zebrafish cornea and it directly correlates collagen fibril and proteoglycan organisation of the zebrafish cornea with that of the human cornea. The similarities between the two species, including the possession of a Bowman layer, suggest that the zebrafish could serve as a model for the genetics of human corneal development and inherited disease. [source] Rho plays a central role in regulating local cell-matrix mechanical interactions in 3D cultureCYTOSKELETON, Issue 6 2007N. Lakshman Abstract The purpose of this study was to assess quantitatively the role of the small GTPase Rho on cell morphology, f-actin organization, and cell-induced matrix remodeling in 3D culture. Human corneal fibroblasts (HTK) were infected with adenoviruses that express green fluorescent protein (GFP) or GFP-N19Rho (dominant negative Rho). One day later cells were plated inside collagen matrices and allowed to spread for 24 h. Cells were fixed and stained for f-actin. Fluorescent (for f-actin) and reflected light (for collagen fibrils) images were acquired using confocal microscopy. Fourier transform analysis was used to assess local collagen fibril alignment, and changes in cell morphology and collagen density were measured using MetaMorph. The decrease in matrix height was used as an indicator of global matrix contraction. HTK and HTK-GFP cells induced significant global matrix contraction; this was inhibited by N19Rho. HTK and HTK-GFP fibroblasts generally had a bipolar morphology and occasional intracellular stress fibers. Collagen fibrils were compacted and aligned parallel to stress fibers and pseudopodia. In contrast, HTK-GFPN19 cells were elongated, and had a more cortical f-actin distribution. Numerous small extensions were also observed along the cell body. In addition, both local collagen fibril density and alignment were significantly reduced. Rho plays a key role in regulating both the morphology and mechanical behavior of corneal fibroblasts in 3D culture. Overall, the data suggest that Rho-kinase dependent cell contractility contributes to global and local matrix remodeling, whereas Rho dependent activation of mDia and/or other downstream effectors regulates the structure and number of cell processes. Cell Motil. Cytoskeleton 2007. © 2007 Wiley-Liss, Inc. [source] Development of the corneal stroma, and the collagen,proteoglycan associations that help define its structure and functionDEVELOPMENTAL DYNAMICS, Issue 10 2008Andrew J. Quantock Abstract The cornea of the eye is a unique, transparent connective tissue. It is comprised predominantly of collagen fibrils, remarkably uniform in diameter and regularly spaced, organized into an intricate lamellar array. Its establishment involves a precisely controlled sequence of developmental events in which the embryonic cornea undergoes major structural transformations that ultimately determine tissue form and function. In this article, we will review corneal developmental dynamics from a structural perspective, consider the roles and interrelationships of collagens and proteoglycans, and comment on contemporary concepts and current challenges pertinent to developmental processes that result in an optically clear, mature cornea. Developmental Dynamics 237:2607,2621, 2008. © 2008 Wiley-Liss, Inc. [source] Human primary corneal fibroblasts synthesize and deposit proteoglycans in long-term 3-D culturesDEVELOPMENTAL DYNAMICS, Issue 10 2008R. Ren Abstract Our goal was to develop a 3-D multi-cellular construct using primary human corneal fibroblasts cultured on a disorganized collagen substrate in a scaffold-free environment and to use it to determine the regulation of proteoglycans over an extended period of time (11 weeks). Electron micrographs revealed multi-layered constructs with cells present in between alternating parallel and perpendicular arrays of fibrils. Type I collagen increased 2,4-fold. Stromal proteoglycans including lumican, syndecan4, decorin, biglycan, mimecan, and perlecan were expressed. The presence of glycosaminoglycan chains was demonstrated for a subset of the core proteins (lumican, biglycan, and decorin) using lyase digestion. Cuprolinic blue,stained cultures showed that sulfated proteoglycans were present throughout the construct and most prominent in its mid-region. The size of the Cuprolinic-positive filaments resembled those previously reported in a human corneal stroma. Under the current culture conditions, the cells mimic a development or nonfibrotic repair phenotype. Developmental Dynamics 237:2705,2715, 2008. © 2008 Wiley-Liss, Inc. [source] Polyacrylamide gel electrophoresis followed by sodium dodecyl sulfate gradient polyacrylamide gel electrophoresis for the study of the dimer to monomer transition of human transthyretinELECTROPHORESIS, Issue 14 2003Klaus Altland Abstract Familial amyloidotic polyneuropathy (FAP) is caused by mutations which destabilize transthyretin (TTR) and facilitate the aggregation into extracellular amyloid fibrils preferentially in peripheral nerve and heart tissues. Therapeutic and preventive trials for FAP at the plasma TTR level require a careful study of the destabilization of TTR under variable conditions. We have developed a simple double one-dimensional (D1-D) electrophoretic procedure with polyacrylamide gel electrophoresis (PAGE) followed by sodium dodecylsulfate (SDS) gradient PAGE to study the dimer to monomer transition. TTR is first isolated by PAGE from other plasma proteins. The gel strip containing the TTR fraction is incubated in 2% SDS under varying conditions of temperature, buffer composition, pH, and additives like urea and/or a sulfhydryl-reactive agent, followed by SDS-gradient PAGE for the separation of TTR dimers and monomers. We demonstrate that an unidirectional dimer to monomer transition of normal TTR is achieved at 70,80°C in neutral to mild alkaline buffers or at 37°C and slightly acidic pH (6,7). Addition of urea favors the transition into monomers. Amyloidogenic mutations like amyloidogenic TTR (ATTR)-V30M or ATTR-I107V favor the transition into monomers in buffer systems close to the physiological pH of human plasma. We conclude that this finding has to be considered by any hypothesis on ATTR-derived amyloidogenesis. [source] Mechanisms and consequences of bladder cell invasion by uropathogenic Escherichia coliEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 2008B. K. Dhakal ABSTRACT Strains of uropathogenic Escherichia coli (UPEC) are the major cause of urinary tract infections worldwide. Multiple studies over the past decade have called into question the dogmatic view that UPEC strains act as strictly extracellular pathogens. Rather, bacterial expression of filamentous adhesive organelles known as type 1 pili and Afa/Dr fibrils enable UPEC to invade host epithelial cells within the urinary tract. Entry into bladder epithelial cells provides UPEC with a protected niche where the bacteria can persist quiescently for long periods, unperturbed by host defences and protected from many antibiotic treatments. Alternately, internalized UPEC can rapidly multiply, forming large intracellular inclusions that can contain several thousand bacteria. Initial work aimed at defining the host and bacterial factors that modulate the entry, intracellular trafficking, and eventual resurgence of UPEC suggests a high degree of host-pathogen crosstalk. Targeted disruption of these processes may provide a novel means to prevent and treat recurrent, relapsing and chronic infections within the urinary tract. [source] Biomorphic Silicon Carbide Coated with an Electrodeposition of Nanostructured Hydroxyapatite/Collagen as Biomimetic Bone Filler and Scaffold,ADVANCED ENGINEERING MATERIALS, Issue 8 2010M. Lelli Abstract The paper describes the method of preparation and chemical/physical characterization of a new biomaterial to be used as a bone substitute and bone-tissue engineering scaffold, which synergistically joins a porous bio-inspired morphology and the mechanical properties of biomorphic silicon carbide (BioSiC) with the surface bioactivity of a nanostructured hydroxyapatite/collagen biomimetic coating. FT-IR spectroscopy and XRD techniques are utilized to determine the chemical coating's composition. The morphology and size of the inorganic and protein components are investigated by TEM. The characteristic morphology of BioSiC channels and pores, which differ as a function of the transversal or longitudinal cross-section and with etching time, are investigated by SEM. Natural wood transformed into BioSiC acts as a cathode in an electrochemically assisted process that produces on its surface a biomimetic coating of hydroxyapatite nanocrystals and reconstituted type I collagen fibrils, producing an innovative apatite/collagen biomimetic porous bone filler and scaffold for tissue engineering. [source] | ||||||||||||||||||||||||||||||||||||||||