Microfibrils: Components of the extracellular matrix consisting primarily of fibrillin. They are essential for the integrity of elastic fibers.Microfilament Proteins: Monomeric subunits of primarily globular ACTIN and found in the cytoplasmic matrix of almost all cells. They are often associated with microtubules and may play a role in cytoskeletal function and/or mediate movement of the cell or the organelles within the cell.Elastic Tissue: Connective tissue comprised chiefly of elastic fibers. Elastic fibers have two components: ELASTIN and MICROFIBRILS.Cellulose: A polysaccharide with glucose units linked as in CELLOBIOSE. It is the chief constituent of plant fibers, cotton being the purest natural form of the substance. As a raw material, it forms the basis for many derivatives used in chromatography, ion exchange materials, explosives manufacturing, and pharmaceutical preparations.Marfan Syndrome: An autosomal dominant disorder of CONNECTIVE TISSUE with abnormal features in the heart, the eye, and the skeleton. Cardiovascular manifestations include MITRAL VALVE PROLAPSE, dilation of the AORTA, and aortic dissection. Other features include lens displacement (ectopia lentis), disproportioned long limbs and enlarged DURA MATER (dural ectasia). Marfan syndrome is associated with mutations in the gene encoding fibrillin, a major element of extracellular microfibrils of connective tissue.Contractile Proteins: Proteins which participate in contractile processes. They include MUSCLE PROTEINS as well as those found in other cells and tissues. In the latter, these proteins participate in localized contractile events in the cytoplasm, in motile activity, and in cell aggregation phenomena.Extracellular Matrix Proteins: Macromolecular organic compounds that contain carbon, hydrogen, oxygen, nitrogen, and usually, sulfur. These macromolecules (proteins) form an intricate meshwork in which cells are embedded to construct tissues. Variations in the relative types of macromolecules and their organization determine the type of extracellular matrix, each adapted to the functional requirements of the tissue. The two main classes of macromolecules that form the extracellular matrix are: glycosaminoglycans, usually linked to proteins (proteoglycans), and fibrous proteins (e.g., COLLAGEN; ELASTIN; FIBRONECTINS; and LAMININ).ElastinLatent TGF-beta Binding Proteins: A family of secreted multidomain proteins that were originally identified by their association with the latent form of TRANSFORMING GROWTH FACTORS. They interact with a variety of EXTRACELLULAR MATRIX PROTEINS and may play a role in the regulation of TGB-beta bioavailability.Ligaments: Shiny, flexible bands of fibrous tissue connecting together articular extremities of bones. They are pliant, tough, and inextensile.Tropoelastin: A salt-soluble precursor of elastin. Lysyl oxidase is instrumental in converting it to elastin in connective tissue.Microscopy, Electron: Microscopy using an electron beam, instead of light, to visualize the sample, thereby allowing much greater magnification. The interactions of ELECTRONS with specimens are used to provide information about the fine structure of that specimen. In TRANSMISSION ELECTRON MICROSCOPY the reactions of the electrons that are transmitted through the specimen are imaged. In SCANNING ELECTRON MICROSCOPY an electron beam falls at a non-normal angle on the specimen and the image is derived from the reactions occurring above the plane of the specimen.Collagen Type VI: A non-fibrillar collagen that forms a network of MICROFIBRILS within the EXTRACELLULAR MATRIX of CONNECTIVE TISSUE. The alpha subunits of collagen type VI assemble into antiparallel, overlapping dimers which then align to form tetramers.Cell Wall: The outermost layer of a cell in most PLANTS; BACTERIA; FUNGI; and ALGAE. The cell wall is usually a rigid structure that lies external to the CELL MEMBRANE, and provides a protective barrier against physical or chemical agents.Connective Tissue: Tissue that supports and binds other tissues. It consists of CONNECTIVE TISSUE CELLS embedded in a large amount of EXTRACELLULAR MATRIX.Apium graveolens: A plant species of the family APIACEAE. The stalks are a food source.Cetomacrogol: Non-ionic surfactant of the polyethylene glycol family. It is used as a solubilizer and emulsifying agent in foods, cosmetics, and pharmaceuticals, often as an ointment base, and also as a research tool.Gluconacetobacter xylinus: A species of acetate-oxidizing bacteria, formerly known as Acetobacter xylinum.Ectopia Lentis: Congenital displacement of the lens resulting from defective zonule formation.Gold Colloid: A suspension of metallic gold particles.Sulfanilamides: Compounds based on 4-aminobenzenesulfonamide. The '-anil-' part of the name refers to aniline.Extracellular Matrix: A meshwork-like substance found within the extracellular space and in association with the basement membrane of the cell surface. It promotes cellular proliferation and provides a supporting structure to which cells or cell lysates in culture dishes adhere.Hypocotyl: The region of the stem beneath the stalks of the seed leaves (cotyledons) and directly above the young root of the embryo plant. It grows rapidly in seedlings showing epigeal germination and lifts the cotyledons above the soil surface. In this region (the transition zone) the arrangement of vascular bundles in the root changes to that of the stem. (From Concise Dictionary of Biology, 1990)Microscopy, Electron, Scanning Transmission: A type of TRANSMISSION ELECTRON MICROSCOPY in which the object is examined directly by an extremely narrow electron beam scanning the specimen point-by-point and using the reactions of the electrons that are transmitted through the specimen to create the image. It should not be confused with SCANNING ELECTRON MICROSCOPY.Dermis: A layer of vascularized connective tissue underneath the EPIDERMIS. The surface of the dermis contains innervated papillae. Embedded in or beneath the dermis are SWEAT GLANDS; HAIR FOLLICLES; and SEBACEOUS GLANDS.Ciliary Body: A ring of tissue extending from the scleral spur to the ora serrata of the RETINA. It consists of the uveal portion and the epithelial portion. The ciliary muscle is in the uveal portion and the ciliary processes are in the epithelial portion.Cattle: Domesticated bovine animals of the genus Bos, usually kept on a farm or ranch and used for the production of meat or dairy products or for heavy labor.Desmosine: A rare amino acid found in elastin, formed by condensation of four molecules of lysine into a pyridinium ring.Xylans: Polysaccharides consisting of xylose units.Versicans: HYALURONAN-containing proteoglycans found in the EXTRACELLULAR MATRIX of a variety of tissues and organs. Several versican isoforms exist due to multiple ALTERNATIVE SPLICING of the versican MESSENGER RNA.Plant Epidermis: A thin layer of cells forming the outer integument of seed plants and ferns. (Random House Unabridged Dictionary, 2d ed)Acetobacter: A species of gram-negative bacteria of the family ACETOBACTERACEAE found in FLOWERS and FRUIT. Cells are ellipsoidal to rod-shaped and straight or slightly curved.X-Ray Diffraction: The scattering of x-rays by matter, especially crystals, with accompanying variation in intensity due to interference effects. Analysis of the crystal structure of materials is performed by passing x-rays through them and registering the diffraction image of the rays (CRYSTALLOGRAPHY, X-RAY). (From McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)Microscopy, Immunoelectron: Microscopy in which the samples are first stained immunocytochemically and then examined using an electron microscope. Immunoelectron microscopy is used extensively in diagnostic virology as part of very sensitive immunoassays.Plant Cells: Basic functional unit of plants.Rosaceae: The rose plant family in the order ROSALES and class Magnoliopsida. They are generally woody plants. A number of the species of this family contain cyanogenic compounds.Glucans: Polysaccharides composed of repeating glucose units. They can consist of branched or unbranched chains in any linkages.Collagen: A polypeptide substance comprising about one third of the total protein in mammalian organisms. It is the main constituent of SKIN; CONNECTIVE TISSUE; and the organic substance of bones (BONE AND BONES) and teeth (TOOTH).Glucosyltransferases: Enzymes that catalyze the transfer of glucose from a nucleoside diphosphate glucose to an acceptor molecule which is frequently another carbohydrate. EC 2.4.1.-.Actin Cytoskeleton: Fibers composed of MICROFILAMENT PROTEINS, which are predominately ACTIN. They are the smallest of the cytoskeletal filaments.Skin Abnormalities: Congenital structural abnormalities of the skin.Dinitrobenzenes: Benzene derivatives which are substituted with two nitro groups in the ortho, meta or para positions.Microscopy, Electron, Scanning: Microscopy in which the object is examined directly by an electron beam scanning the specimen point-by-point. The image is constructed by detecting the products of specimen interactions that are projected above the plane of the sample, such as backscattered electrons. Although SCANNING TRANSMISSION ELECTRON MICROSCOPY also scans the specimen point by point with the electron beam, the image is constructed by detecting the electrons, or their interaction products that are transmitted through the sample plane, so that is a form of TRANSMISSION ELECTRON MICROSCOPY.Cardiovascular Abnormalities: Congenital, inherited, or acquired anomalies of the CARDIOVASCULAR SYSTEM, including the HEART and BLOOD VESSELS.Fibroblasts: Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules.Skin: The outer covering of the body that protects it from the environment. It is composed of the DERMIS and the EPIDERMIS.Biopolymers: Polymers synthesized by living organisms. They play a role in the formation of macromolecular structures and are synthesized via the covalent linkage of biological molecules, especially AMINO ACIDS; NUCLEOTIDES; and CARBOHYDRATES.
Microfibril: A microfibril is a very fine fibril, or fiber-like strand, consisting of glycoproteins and cellulose. It is usually, but not always, used as a general term in describing the structure of protein fiber, e.Actin-binding protein: Actin-binding proteins (also known as ABP) are proteins that bind to actin. This may mean ability to bind actin monomers, or polymers, or both.Internal elastic laminaCellulose fiber: Cellulose fibers () are fibers made with ether or esters of cellulose, which can be obtained from the bark, wood or leaves of plants, or from a plant-based material. Besides cellulose, these fibers are compound of hemicellulose and lignin, and different percentages of these components are responsible for different mechanical properties observed.Antoine MarfanPUR4: pUR4 is a recombinant peptide that is known to inhibit the polymerization of fibronectin in a number of cell types including fibroblasts and endothelial cells. Fibronectin is an essential component of the extracellular matrix that acts as a mediator between the extracellular matrix and the cells that reside within the matrix.Elastin: Elastin is a highly elastic protein in connective tissue and allows many tissues in the body to resume their shape after stretching or contracting. Elastin helps skin to return to its original position when it is poked or pinched.TGF beta Activation: Transforming growth factor beta (TGF-β) is a potent cell regulatory polypeptide homodimer of 25kD.Roberts, A.Degenerative suspensory ligament desmitis: Degenerative Suspensory Ligament Desmitis commonly called DSLD, also known as Equine Systemic Proteoglycan Accumulation (ESPA) is a systemicTropoelastin: Tropoelastin is a water-soluble molecule with a molecular weight of approximately 72,000 daltons. Multiple tropoelastin molecules covalently bind together with crosslinks to form the protein elastin that is very prevalent in the body.Low-voltage electron microscope: Low-voltage electron microscope (LVEM) is an electron microscope which operates at accelerating voltages of a few kiloelectronvolts or less. While the low voltage electron microscopy technique will never replace conventional high voltage electron microscopes, it is quickly becoming appreciated for many different disciplines.Bethlem myopathy: Bethlem myopathy is an autosomal dominant myopathy, classified as a congenital form of muscular dystrophy, that is caused by a mutation in one of the three genes coding for type VI collagen. These include COL6A1, COL6A2, and COL6A3.Cell envelope: The cell envelope comprises the inner cell membrane and the cell wall of a bacterium, if present, plus a bacterial outer membrane, if one is present (i.e.Dense connective tissue: Dense connective tissue, also called dense fibrous tissue, has fibers as its main matrix element.Platynota stultana: The Omnivorous Leafroller (Platynota stultana) is a moth of the Tortricidae family. It is found in Mexico, California, Arizona, Texas, Florida and Hawaii.Cetomacrogol 1000: Cetomacrogol 1000 is a non-ionic surfactant of the polyethylene glycol family. It is used as a solubilizer and emulsifying agent in foods, cosmetics, and pharmaceuticals, often as an ointment base, and also as a research tool.Karl Stellwag von Carion: Karl Stellwag von Carion (January 28, 1823 - November 21, 1904) was an Austrian ophthalmologist who was a native of Langendorf, a village in Moravia.OryzalinExtracellular matrixEpicotyl: An Epicotyl is important for the beginning stages of a plants life. It is the region of a seedling stem above the stalks of the seed leaves of an embryo plant.Papillary dermis: The papillary dermis is the uppermost layer of the dermis. It intertwines with the rete ridges of the epidermis and is composed of fine and loosely arranged collagen fibers.Ciliary body: The ciliary body is a part of the eye that includes the ciliary muscle, which controls the shape of the lens, and the ciliary epithelium, which produces the aqueous humor. The ciliary body is part of the uvea, the layer of tissue that delivers oxygen and nutrients to the eye tissues.Beef cattle: Beef cattle are cattle raised for meat production (as distinguished from dairy cattle, used for milk production). The meat of adult cattle is known as beef.DesmosineGlucuronoxylan: Glucuronoxylans are the primary components of hemicellulose as found in hardwood trees, for example birch.http://www.The Werewolf (1956 film): The Werewolf is a low-budget American 1956 science fiction horror film, produced by Sam Katzman and directed by Fred F. Sears from a script by Robert E.Sobrietol: Sobrietol is a North American brand of nutritional supplement marketed as a remedy for hangovers and to prevent symptoms associated with alcohol flush reaction. The list of ingredients includes the enzymes quinoptotein alcohol dehydrogenase (QADH) and quinoprotein aldehyde dehydrogenase (QALDH) from Glucanobacter suboxydans or Acetobacter suboxydans or oxydans, either in purified form or as cell extracts, together with buffering agents and protectants designed to ensure that the enzymes remain biologically active after oral ingestion, and "a source of oxygen in an amount sufficient for the enzymes to metabolize ethanol after administration to a patient.Powder diffraction: Powder diffraction is a scientific technique using X-ray, neutron, or electron diffraction on powder or microcrystalline samples for structural characterization of materials.B.Immunostaining: Immunostaining is a general term in biochemistry that applies to any use of an antibody-based method to detect a specific protein in a sample. The term immunostaining was originally used to refer to the immunohistochemical staining of tissue sections, as first described by Albert Coons in 1941.Amelanchier alnifolia: Amelanchier alnifolia, the saskatoon, Pacific serviceberry, western serviceberry, alder-leaf shadbush, dwarf shadbush, chuckley pear, or western juneberry,Germplasm Resources Information Network: Amelanchier alnifolia is a shrub with edible berry-like fruit, native to North America from Alaska across most of western Canada and in the western and north-central United States. Historically, it was also called pigeon berry.Decyl polyglucose: Decyl polyglucose is a mild non-ionic synthetic surfactant. It is a type of alkylpolyglycoside derived from glucose or starch and the fatty alcohol decanol.CollagenAbscisate beta-glucosyltransferase: Abscisate beta-glucosyltransferase (, ABA-glucosyltransferase, ABA-GTase, AOG) is an enzyme with system name UDP-D-glucose:abscisate beta-D-glucosyltransferase. This enzyme catalyses the following chemical reactionField emission probesDouble aortic arch: Double aortic arch (DAA) is a relatively rare congenital cardiovascular malformation. DAA is an anomaly of the aortic arch in which two aortic arches form a complete vascular ring that can compress the trachea and/or esophagus.Dermal fibroblast: Dermal fibroblasts are cells within the dermis layer of skin which are responsible for generating connective tissue and allowing the skin to recover from injury. Using organelles (particularly the rough endoplasmic reticulum), dermal fibroblasts generate and maintain the connective tissue which unites separate cell layers.Dermal equivalent: The dermal equivalent is an in vitro model of the dermal layer of skin. It is constructed by seeding dermal fibroblasts into a collagen gel.Biopolymer: Biopolymers are polymers produced by living organisms; in other words, they are polymeric biomolecules. Since they are polymers, biopolymers contain monomeric units that are covalently bonded to form larger structures.
(1/184) The supramolecular organisation of fibrillin-rich microfibrils determines the mechanical properties of bovine zonular filaments.
The zonular filaments from the eyes of cows are rich in microfibrils containing fibrillin. Tensile tests, stress-relaxation tests and X-ray diffraction studies were used to study the relationship between the mechanical behaviour of zonular filaments and the molecular packing and structure of the fibrillin-rich microfibrils. Zonular filaments show a non-linear (J-shaped) stress-strain curve and appreciable stress-relaxation. It is proposed that the non-linear properties are due to local variations in waviness in the microfibrils or assemblies of microfibrils, which straighten out and become more regularly aligned with strain. Previous and current X-ray diffraction results consistently show a partial ordering of microfibrils in zonular filaments into staggered aggregates which become more ordered and laterally aligned on stretching. Although the removal and re-addition of Ca(2+) is known to change the molecular structure of fibrillin, no effect was observed on the tensile properties of the zonular filaments. It is hypothesised that strain-induced deformation in the supramolecular aggregate packing may not be Ca(2+)-sensitive but could dominate the mechanical behaviour of microfibrillar arrays in zonular filaments. (+info)
(2/184) 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.
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 (S(v) 0.030 nm2/nm3) 15% higher (P < 0.05) than values obtained by the indirect IUR method (V(v) 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 utilisation of these estimates, is here regarded as representing a 'gold standard'. The sensitivity of these 3 methods was also tested with cartilage from an in vitro loading experiment which caused tissue compression. In the superficial zone of articular cartilage Vv and Sv of collagen fibrils increased (P < 0.05). This difference in the stereological estimates was only detected by the indirect IUR estimation but not by the direct VS or direct IUR methods. This indicated that the indirect IUR estimation was more sensitive than the direct VS or direct IUR estimations. On the basis of these observations, the indirect zonal IUR estimation can be regarded as the technique of choice in the electron microscopic stereology of cartilage collagen. (+info)
(3/184) Characterization of an in vitro model of elastic fiber assembly.
Elastic fibers consist of two morphologically distinct components: elastin and 10-nm fibrillin-containing microfibrils. During development, the microfibrils form bundles that appear to act as a scaffold for the deposition, orientation, and assembly of tropoelastin monomers into an insoluble elastic fiber. Although microfibrils can assemble independent of elastin, tropoelastin monomers do not assemble without the presence of microfibrils. In the present study, immortalized ciliary body pigmented epithelial (PE) cells were investigated for their potential to serve as a cell culture model for elastic fiber assembly. Northern analysis showed that the PE cells express microfibril proteins but do not express tropoelastin. Immunofluorescence staining and electron microscopy confirmed that the microfibril proteins produced by the PE cells assemble into intact microfibrils. When the PE cells were transfected with a mammalian expression vector containing a bovine tropoelastin cDNA, the cells were found to express and secrete tropoelastin. Immunofluorescence and electron microscopic examination of the transfected PE cells showed the presence of elastic fibers in the matrix. Biochemical analysis of this matrix showed the presence of cross-links that are unique to mature insoluble elastin. Together, these results indicate that the PE cells provide a unique, stable in vitro system in which to study elastic fiber assembly. (+info)
(4/184) Initial steps in assembly of microfibrils. Formation of disulfide-cross-linked multimers containing fibrillin-1.
Fibrillins are the major constituents of extracellular microfibrils. How fibrillin molecules assemble into microfibrils is not known. Sequential extractions and pulse-chase labeling of organ cultures of embryonic chick aortae revealed rapid formation of disulfide-cross-linked aggregates containing fibrillin-1. These results demonstrated that intermolecular disulfide bond formation is an initial step in the assembly process. To identify free cysteine residues available for intermolecular cross-linking, small recombinant peptides of fibrillin-1 harboring candidate cysteine residues were analyzed. Results revealed that the first four cysteine residues in the unique N terminus form intramolecular disulfide bonds. One cysteine residue (Cys(204)) in the first hybrid domain of fibrillin-1 was found to occur as a free thiol and is therefore a good candidate for intermolecular disulfide bonding in initial steps of the assembly process. Furthermore, evidence indicated that the comparable cysteine residue in fibrillin-2 (Cys(233)) also occurs as a free thiol. These free cysteine residues in fibrillins are readily available for intermolecular disulfide bond formation, as determined by reaction with Ellman's reagent. In addition to these major results, the cleavage site of the fibrillin-1 signal peptide and the N-terminal sequence of monomeric authentic fibrillin-1 from conditioned fibroblast medium were determined. (+info)
(5/184) Rho and Rac exert antagonistic functions on spreading of macrophage-derived multinucleated cells and are not required for actin fiber formation.
Multinucleated giant cells (MNGC) derived from avian blood monocytes present, like osteoclasts, an unusual cytoskeletal organization characterized by (1) cortical rings of actin filaments, (2) unique adhesion structures called podosomes and (3) vinculin containing focal complexes which are not visibly connected to F-actin structures. The Rho family of small GTPases plays an essential role in the regulation and organization of cellular cytoskeletal structures including F-actin and vinculin associated structures. Using bacterial toxins such as modified exoenzyme C3 (C3B) and toxin B or overexpression of constitutively active Rac and Rho proteins fused to the green fluorescent protein (GFP), we show that Rac and Rho play antagonistic roles in regulating the morphology of osteoclast-like cells. Inhibition of Rho by C3B triggered MNGC spreading whereas activated Rho promoted cell retraction. However, inhibition or activation of Rho led to complete disorganization of fibrillar actin structures, including podosomes. Toxin B inhibition of Rho, Rac and Cdc42 induced a time dependent F-actin and vinculin reorganization. Initially, actin fibers with associated adhesion plaques formed and disappeared subsequently. Finally, only small focal complexes remained at the MNGC periphery before retracting. At the time when actin fibers formed, we observed that Rac was already inhibited by toxin B. By combining C3B treatment and overexpression of a dominant negative form of Rac (N17Rac), we show that the formation of these focal adhesion and actin fiber structures required neither Rho nor Rac activity. Moreover, our results show that podosomes are extremely unstable structures since any modifications of Rho or Rac activity resulted in their dissociation. (+info)
(6/184) Collagen XI nucleates self-assembly and limits lateral growth of cartilage fibrils.
Fibrils of embryonic cartilage are heterotypic alloys formed by collagens II, IX, and XI and have a uniform diameter of approximately 20 nm. The molecular basis of this lateral growth control is poorly understood. Collagen II subjected to fibril formation in vitro produced short and tapered tactoids with strong D-periodic banding. The maximal width of these tactoids varied over a broad range. By contrast, authentic mixtures of collagens II, IX, and XI yielded long and weakly banded fibrils, which, strikingly, had a uniform width of about 20 nm. The same was true for mixtures of collagens II and XI lacking collagen IX as long as the molar excess of collagen II was less than 8-fold. At higher ratios, the proteins assembled into tactoids coexisting with cartilage-like fibrils. Therefore, diameter control is an inherent property of appropriate mixtures of collagens II and XI. Collagen IX is not essential for this feature but strongly increases the efficiency of fibril formation. Therefore, this protein may be an important stabilizing factor of cartilage fibrils. (+info)
(7/184) Aprotinin binding to amyloid fibrils.
Different low molecular mass ligands have been used to identify amyloid deposits. Among these markers, the dyes Thioflavin T and Congo Red interact specifically with the beta-sheet structure arranged in a cross-beta conformation, which is characteristic of amyloid. However, the molecular details of this interaction remain unknown. When labelled with technetium-99m, the proteinase inhibitor aprotinin has been shown to represent a very important radiopharmaceutical agent for in vivo imaging of extra-abdominal deposition of amyloid in amyloidosis of the immunoglobulin type. However, no information is available as to whether aprotinin binds other types of amyloid fibrils and on the nature and characteristics of the interaction. The present work shows aprotinin binding to insulin, transthyretin, beta-amyloid peptide and immunoglobulin synthetic amyloid fibrils by a specific dot-blot ligand-binding assay. Aprotinin did not bind amorphous precipitates and/or the soluble fibril precursors. A Ka of 2.9 microM-1 for the binding of aprotinin to insulin amyloid fibrils was determined by Scatchard analysis. In competition experiments, analogues such as an aprotinin variant, a spermadhesin and the soybean trypsin inhibitor were tested and results suggest that both aprotinin and the spermadhesin interact with amyloid fibrils through pairing of beta-sheets of the ligands with exposed structures of the same type at the surface of amyloid deposits. An electrostatic component may also be involved in the binding of aprotinin to amyloid fibrils because important differences in binding constants are observed when substitutions V15L17E52 are introduced in aprotinin; on the other hand beta-sheet containing acidic proteins, such as the soybean trypsin inhibitor, are unable to bind amyloid fibrils. (+info)
(8/184) Intracellular biogenesis of collagen fibrils in 'activated fibroblasts' of tendo Achillis. An ultrastructural study in the New Zealand rabbit.
We have studied the formation of collagen fibrils in 'activated fibroblasts' of tendo Achillis of rabbits. The tendon was in the process of regeneration after experimental partial tenotomy. Samples were taken from the peri-incisional region and analysed by transmission electron microscopy. Ultrastructural examination showed the presence of a 'fine dense granular substance' inside the rough endoplasmic reticulum and procollagen filaments. These come together to form collagen fibrils in the dilated vacuoles of the rough endoplasmic reticulum. The possible intra- and extracellular origin of collagen fibrils is suggested. Within the cell biosynthesis of collagen fibrils take place with the formation of collagen substance which gives rise to procollagen filaments. These make contact in parallel apposition to produce striated 'spindle-shaped bodies' which elongate by the longitudinal attachment of more procollagen filaments and form intracellular nascent collagen fibrils. (+info)