Arrangement of the collagen and elastic fibers in the upper human duodenum

Os autores estudaram a disposicao das fibras elasticas e colagenas na porcao superior do duodeno. Estas fibras dispoe-se paralelamente as fibras musculares. Na submucosa, ambos os tipos de fibras formam uma rede pantografica com seus angulos de cruzamento aumentando em direcao oral-aboral. As fibras colagenas e elasticas apresentam uma orientacao polar entre as fibras musculares.

[Elastic system fibers of healthy human gingiva]. / Les fibres du systeme elastique de la gencive saine chez l'homme.

The elastic system fibers, i.e. oxytalan, elaunin and elastic fibers have respectively a fibrillar structure (oxytalan fibers), an amorphous structure (elastic fibers), or a mixed structure (elaunin fibers). The morphological distribution of these fibers is characterized by the presence of oxytalan, elaunin and elastic fibers in the upper medium and deep layers of gingival connective tissue. If the amorphous component is made up of elastin the microfibrillar component consist of structural glycoproteins containing aminoacids different of those found in elastin. Elastin is synthesized by gingival fibroblasts in the form of a precursor, tropoelastin, then disposed at the surface of the microfibrillar component and incorporated in the amorphous component.

The presence of a type IV collagen skeleton associated with periductal elastosis in breast cancer.

Using serial sections of frozen and AFA-fixed tissues from 34 breast cancers, we studied the presence of basement membrane material in the areas of elastosis. Various amounts of type IV collagen but not of laminin were demonstrated in areas of periductal elastosis. In some tumors, type IV collagen accumulated beneath the basement membrane. Periductal elastosis in areas of extensive fibrosis showed focal type IV collagen immunoreactivity, indicating remnants of ducts. Interstitial elastosis corresponded with weak type IV collagen reactivity. Each tumor showed type IV collagen immunostaining of the elastotic areas, with various degrees of intensity. Negative crossreactivity of the type IV collagen antibody with elastin was verified in skin biopsies with solar elastosis. Pre-incubation of the antibody with large amounts of elastin demonstrated an identical immunoreactivity. The specificity of the antibody was confirmed by ELISA and by Western blot analysis. To explain the periductal elastosis, we propose the following hypothesis. Excessive production of basement membrane material by the epithelial cells of the ducts leads to formation of a type IV collagen skeleton. This skeleton can act as the matrix for a secondary deposition of elastic material.

Ultrastructural and cytochemical study of elastic fibers in the ventral aorta of a teleost, Anguilla japonica.

Previous studies have revealed that amorphous elastin and microfibrils are structural entities of mammalian elastic fibers. Elastin shows a wide phylogenetic distribution, but the presence of elastin-associated microfibrils has not been demonstrated in teleost aorta. Thus, we have ultrastructurally and cytochemically examined elastic fibers in the ventral aorta of eel, a teleost, by utilizing routine uranyl acetate and lead double staining, the tannic acid (pH 7.0)-uranyl acetate (TA-UA) method, elastase en bloc digestion, Thiéry's periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method, and the horseradish-peroxidase-labeled concanavalin A (Con A) method. In the ventral aorta of eel, a little ultrastructural difference between elastic fibers in the intima and media and those in the adventitia was noticed, but in either tunic each elastic fiber was basically composed of a "fibrillar core" and surrounding microfibrils. The fibrillar core was a collection of fibrils which showed a tendency to coalesce with each other, and these constituent fibrils were TA-UA positive and elastase-sensitive, representing their nature of elastin. By contrast, microfibrils associated with the fibrillar core were TA-UA negative and elastase-resistant, and their glycoproteinaceous nature was demonstrated by PA-TCH-SP and Con A methods. Thus, this study provides evidence for the presence of elastin-associated microfibrils in teleost aorta. These results are discussed in relation to the topographical difference of elastic fibers in eel aortic wall.

Fine structure of cartilage elastic system fibers, in particular those of the mandibular condyle.

Light microscopy of the mandibular joint tissues from fetal mice show a distribution of fibrillar structures in the articular fibrous capsule covering the condylar head. Further SEM and TEM studies were conducted on autoclaved xiphoid and mandibular condylar processes of the fetuses for observation of the elastic system fibers in these cartilaginous tissues. SEM showed that non-collaginous fibers branched and united to form a complicated network in the cartilage. A fine structure study on diameter distribution of the fibers indicated elastogenesis in the differentiating cell layer and fiber maturation in the articular surfaces and calcification layer, thus suggesting a sequential development, growth, and degeneration of the cellular and fibrillar components in the cartilage, as well as bidirectional cell differentiation in the growing mandibular joint. A further TEM study on these autoclaved connective tissues showed the elastic system fibers in the network to be composed of fine microfibrils and amorphous elastin. The elastic fibers in the condylar cartilage were a loose network having many tortuous main and oblique elastic fibers, and coiling oxytalan fibers.

Isolation and characterization of a new 36-kDa microfibril-associated glycoprotein from porcine aorta.

A new connective tissue protein of 36 kDa has been purified from porcine aorta. The biochemical and immunological properties of the protein are distinct from those of microfibril-associated proteins reported previously such as lysyl oxidase, 31-kDa microfibril-associated glycoprotein, and fibrillin. It could bind to concanavalin A-Sepharose and gelatin-Sepharose. The protein contained the sequence Arg-Gly-Asp-Ala in the amino-terminal region, which is the site for the association with cell and extracellular matrix. Using specific antibody raised to the protein, we demonstrated its restricted localization in aorta adventitia. Immunoelectron microscopy specified its location to a class of extracellular structural elements described as elastin-microfibrils.

The tissue distribution of microfibrils reacting with a monospecific antibody to MAGP, the major glycoprotein antigen of elastin-associated microfibrils.

Elastic tissue, when viewed in the electron microscope, consists of an amorphous component that is immunoreactive with anti-tropoelastin (TE) antibodies and microfibrils, that react with monospecific antibodies against a 31 kDa microfibrillar glycoprotein constituent, called MAGP. A detailed study of the tissue distribution of microfibrils and of the two elastic tissue antibodies has been carried out, using single and double-labeled immunogold techniques in high resolution electron microscopy. Microfibrils similar in appearance to those associated with elastic tissue and immunoreactive with the anti-MAGP antibody, have been demonstrated in many tissues in the absence of amorphous elastic tissue. In the majority of these tissues, specific anti-TE antibody localization was demonstrated in the immediate vicinity of the microfibrils, or alternatively, the microfibrils were shown to be in direct continuity with microfibrils of similar morphology, which were associated with material immunoreactive with anti-TE antibody. The diameter of these microfibrils varied between 8 nm and 16 nm. They were unbranched structures of indefinite length, with a tubular profile on cross section and periodic staining in longitudinal section. In some tissues, notably in the ciliary zonule and in the mesangial region of the renal glomerulus, microfibrils of similar morphology were demonstrated which were immunoreactive with anti-MAGP antibody, but which were unrelated to amorphous elastic tissue and with which anti-TE antibody localization could not be demonstrated. The evidence available supports the conclusion that all these microfibrils are members of a single class of structures, which are widely distributed in the tissues and which are secreted by a range of cell types. Attention is directed to the close relationship between these microfibrils and the basement membrane of the glomerulus, of uterine smooth muscle, of the basal cells of the epidermis and of the reticulum cells of the spleen.

Changes in lung elastic fiber structure and concentration associated with hyperoxic exposure in the developing rat lung.

Prolonged hyperoxic exposure is associated with impaired alveolarization of the lung in both the rat and the human neonate. Elastin is currently thought to play a pivotal role in the alveolarization of the lung by providing the structural framework around which new alveoli will develop. Previous studies in both the rat and the human neonate have demonstrated a risk for proteolytic destruction of lung elastin associated with prolonged hyperoxic exposure. The present study was undertaken to determine whether continuous exposure to 100% oxygen during the period of alveolar development in the rat (Days 4 to 13) would alter lung elastin. Parenchymal lung elastic fiber length, volume density of parenchyma, mean linear intercept, and internal surface area were quantitated using morphometric techniques, and the values were compared in control, oxygen-exposed, and malnourished rat pups. Stereologic measurements indicated that total elastic fiber length was significantly greater in lungs of control pups than in lungs of either the oxygen-exposed or the malnourished pups. Examination of sections of lung tissue 20 to 30 microns thick indicated altered elastic fiber structure and numerous alveolar fenestrae only in the hyperoxic pups. The results of these studies demonstrated that hyperoxic exposure during alveolarization alters both total length and structure of lung elastic fibers and suggest that impaired lung development might be due in part to these observed changes.

[Elastic system in human gingiva in health and periodontal disease]. / Sistema elástico en encias humanas con salud y enfermedad paradencial.

We processed 28 samples of human gingiva in health or paradential illness, of 15 patients between 20 to 67 years old. In 306 histologies cuts we made different staining to light microscope. We analyzed presence, distribution, diameter and relations of the elastic, elaunin and oxytalan fibers. The patron of distribution fibrilar of the elastic system gingival was different than other zones of the body. The description had made in three zones of the gingival connective tissue: deep, subpapilar and papilar zones. The elastic fibers had less numerous and larger diameter than the elaunin y oxytalan ones. Their repair was irregular. The elaunin followed the colagen fibers. The oxytalan formed delicates nets in the lodge of connective tissue, inflamatories infiltrates and interfibrilar spaces. They came to the basal membrane perpendicularly or oblicualy, isolated or in little bundles. The fibers of elastic system increased principaly in inflamed gingiva of teeth with overburden, though they supported the low proportion that exist at health gingiva. They appeared at the little vases wall and were increased at zones next to edentate spaces. The analogy and close entail between the elaunin and oxytalan fibers to colagen ones, made think in a relation of reenforcement of colagen, increased at paradential illness.

Structural proteins of the neonatal and adult lamina cribrosa.

Optic nerve heads from three premature infants and six adults were studied immunohistochemically to compare the extracellular proteins in the lamina cribrosa of young and old human eyes. In both age groups, antibodies to the basement membrane components laminin and collagen type IV were associated with blood vessels and laminar beam margins. In the adult eyes, interstitial collagen types I and III were heavily distributed within the laminar beams. Antibodies to fibrillin, the microfibrillar portion of elastin, labeled discrete, heavy bands oriented longitudinally within these beams. The beams of the neonatal lamina cribrosa contained much less interstitial collagen, with a predominance of collagen type III. Neonatal elastic tissue bands were less numerous and distinct within the laminar beams. These biochemical differences between the young and old lamina cribrosa may, in part, explain different clinical behaviors of the optic nerve head in congenital and adult glaucoma.

Ganglioside content and composition of cells from normal and atherosclerotic human aorta.

The ganglioside content and composition of cells obtained by enzyme digestion of 2 layers of human aortic intima were investigated. Five gangliosides were identified in cells isolated from the external musculo-elastic intimal layer adjacent to the media: GM3, GM1, GD3, GD1a, and GT1b. The same gangliosides plus ganglioside Gx, the chromatographic mobility of which corresponded to the mobility of ganglioside GD1b from human brain, were found in cells from the internal elastic-hyperplastic intimal layer adjacent to the vessel lumen. In both layers, the major cellular ganglioside was GM3 which represented 60% of the total cellular ganglioside content. The ganglioside content was lower in cells obtained from fatty streaks compared to cells isolated from unaffected intima. The amount of di- and trisialogangliosides in atherosclerotic plaque cells was lower, and that of monosialogangliosides higher than in cells isolated from unaffected intima. The amount of GM3 was mainly responsible for the difference in the total ganglioside content of cells obtained from different lesion types. On the whole, cells from fatty streaks contained smaller amounts of total gangliosides, whereas cells from plaques had greater total ganglioside content, than cells from unaffected intima.

Immunohistochemical demonstration of age-related deposition of vitronectin (S-protein of complement) and terminal complement complex on dermal elastic fibers.

Immunoreactivity of vitronectin was investigated in 100 skin specimens from different body regions in 87 individuals of different ages using monoclonal and polyclonal anti-vitronectin antibodies in an avidin-biotin-peroxidase complex technique. Vitronectin immunoreactivity was found in conjunction with dermal elastic fibers in all subjects older than 13 years. No vitronectin immunostaining was detected in subjects younger than six years, suggesting deposition of vitronectin during late childhood or early adolescence. Using an immunogold staining procedure, vitronectin immunoreactivity was ultrastructurally localized to the periphery of elastic fibers. The blood level of vitronectin in 20 healthy newborns was 67% of the adult level, suggesting active biosynthesis already in the fetus. To investigate whether vitronectin is deposited as part of the SC5b-9 complex or as uncomplexed protein, the immunoreactivity of vitronectin was compared with that of C9, using monoclonal and polyclonal antibodies against the C9 neoantigen. Distinct C9 neoantigen immunoreactivity was demonstrated in association with dermal elastic fibers in human skin in adults but only in subjects older than 30 years. The intensity of C9 neoantigen immunoreactivity appeared to increase with age and was found to be stronger in sun-exposed skin than in sun-protected skin. These findings indicate that uncomplexed vitronectin is deposited during childhood or early adolescence and that terminal complement complexes (C5b-9 and/or SC5b-9) are deposited on elastic fibers later on in life. Hypothetically, the tissue form of vitronectin may be involved in the prevention of tissue damage in proximity to local complement activation. In addition, it may be physiologically important as substratum for cells, stimulating cell migration and anchorage.

The protein components of the 12-nanometer microfibrils of elastic and nonelastic tissues.

A procedure has been developed which is much more specific for the solubilization of the elastin-associated microfibrils from fetal bovine nuchal ligament using treatment with reductive saline in place of reductive guanidine hydrochloride buffer. When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reductive saline extracts were shown to contain only five major protein bands with Mrs of 340,000, 78,000, 70,000, 31,000, and 25,000. The 31-kDa species was identified immunologically as the previously described macromolecule named microfibril-associated glycoprotein (MAGP) (Gibson, M. A., Hughes, J. L., Fanning, J. C., and Cleary, E. G. (1986) J. Biol. Chem. 261, 11429-11436). The proteins were purified by gel permeation, ion exchange, and affinity chromatography. Amino acid analyses showed that each protein had a profile which was distinct from that of MAGP although each was also high in acidic amino acids and cystine. The 340- and 78-kDa species were each demonstrated by immunoelectron microscopy with affinity-purified antibodies to be derived from the elastin-associated microfibris, and these were provisionally named microfibrillar protein 340 (MP340) and microfibrillar protein 78 (MP78), respectively. Each of the above antibodies gave a tissue distribution identical to that of anti-MAGP antibodies, and thus MP340 and MP78 also were identified with the 12-nm microfibrils of nonelastic tissues. MP340 was shown to absorb out completely the microfibrillar immunoreactivity of anti-(reductive guanidine hydrochloride extract) antibodies, indicating that MP340 was (a) the major microfibrillar constituent in these extracts and (b) the second unidentified microfibrillar antigen described previously. The relationship of the 70- and 25-kDa proteins to microfibrils is yet to be established. Immunoblot and immunoabsorption studies showed that MAGP and MP78 were immunologically related to MP340 but not to each other. Cyanogen bromide peptide mapping indicated that MAGP was structurally related to MP340. It is postulated that MAGP and MP78 are constituents of MP340 which in turn is the subunit of which the 12-nm microfibrils are composed.

Elastic fibres in patients with systemic sclerosis. A morphological study.

Dermal elastic fibres in biopsies taken from sun-exposed involved digital skin and sun-protected uninvolved skin on the medial aspect of the upper arms from 13 patients with systemic sclerosis were examined by light and transmission electron microscopy. For controls, biopsies were taken from similar sites from 4 age- and sex-matched healthy volunteers and 4 patients with primary Raynaud's phenomenon. On light microscopy only the control digital biopsies showed mild actinic changes of the elastic fibres whereas in all the biopsies from patients with systemic sclerosis identical changes of thickening, clumping and fragmentation of the elastic fibres were observed. Quantitative assessment of the dermal elastic fibres using microdensitometry and video image analysis showed no significant difference between the patients and controls. On electron microscopy more advanced abnormalities similar to those seen in actinic damage and chronological aging were found in the biopsies from all the patients with systemic sclerosis compared to the controls.

Post-embedding methods for immunolocalization of elastin and related components in tissues.

Elastic tissue is composed of amorphous-appearing elastin and 12-nm diameter microfibrils, one component of which has recently been isolated and characterized as the 31 KD microfibril-associated glycoprotein MAGP. Monospecific antibodies to each of these components have been developed in this laboratory. The parameters that determine optimal localization of colloidal gold probes for post-embedding immunolabeling of elastic tissue components have been systematically studied in a variety of normal and developing tissues in mammals and birds. Protein A-gold probes stabilized with dextran have been shown to provide complexes that remain stable after more than 2 years. Conditions have been defined that permit precise localization within the extracellular matrix of antibodies to MAGP and to elastin, singly and together. Best results were obtained with acrylic resins (Lowicryl K4M or LR White). Fixation in glutaraldehyde or other aldehydic fixatives, with or without osmium, did not affect the immunostaining of elastic tissue with affinity-purified antibodies to tropoelastin, or to anti-[alpha-elastin] or anti-[alkali-insoluble elastin]. Immunostaining with the anti-MAGP antibody was less robust and was possible in tissues which had been fixed only lightly before embedding in Lowicryl K4M or LR White. This staining was enhanced by metaperiodate oxidation of the sections as well as by reduction of the tissues with sodium borohydride en bloc, followed by hyaluronidase digestion of the sections. The effects on immunostaining of a range of enzyme digestions have also been examined. Conditions have thus been defined that make possible detailed study of the relationship between elastic tissue, elastin-associated microfibrils, and other microfibrillar structures in normal and abnormal tissues during development and aging.

Decay-accelerating factor in human skin is associated with elastic fibers.

Recently a complement inhibitor, decay-accelerating factor (DAF), has been found in association with uncharacterized fibers in the extracellular matrix of human dermis. Here we show by immunohistochemistry and immunoelectronmicroscopy that DAF is on the periphery of elastic fibers, and that it appears to be associated with some microfibrillar elements that cover the fibers. That DAF is a component of these microfibrils is also suggested by studies of lesional skin from anetoderma, a disease characterized by destruction of elastic fibers. In two patients we found a network of residual fine fibers in the dermis that stain with antibodies against DAF and fibrillin (one of the proteins known to be present in the microfibrils of elastin), but do not stain with antibodies to elastin. Western blot analysis of dermal extracts with monoclonal antibodies to DAF identified a 67 kDa molecule, slightly smaller than membrane DAF, and similar in size to soluble DAF found in secretions. It is possible that together with vitronectin, an inhibitor of the membrane attack complex recently identified in association with elastin, DAF prevents damage of elastic fibers by complement.