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At least 451 records · Page 25Linked to original sources
PubMed1989

Ultrastructural immunocytochemical localization of elastin in normal human trabecular meshwork.

Previous studies have suggested that hydrophobic moieties within the aqueous outflow channels might interact with certain aqueous components to retard outflow. While elastin is among the most hydrophobic proteins in the trabecular meshwork, it reacts poorly with conventional ultrastructural staining methods, so its potential role in regulating outflow could not be assessed. It was our goal to specifically localize elastin ultrastructurally using polyclonal antibodies against alpha elastin and its soluble precursor, tropoelastin. Human aorta served as a positive control. Preadsorption of the primary antibodies or their substitution with either normal rabbit serum or Tris buffer resulted in negligible labelling. With either antibody, only the electron-lucent elements in the center of elastic fibers of the trabecular meshwork were labelled, indicating that only these elements truly represent elastin. The pattern of elastin distribution within these fibers is most consistent with that found in tendons elsewhere in the body.

Adolescent↗
PubMed1981

Identification of elastin in developing aortic tissue by immunological methods.

Antibodies raised in rabbits to tropoelastin isolated from lathyritic chick aorta were conjugated with ferritin and used to identify and locate elastin-containing elements in sections of aortic tissue from developing chicks. The ferritin-antibody conjugate was associated with a network of fine filaments with a diameter of 3-5 nm which, in the 8-10 day old chick embryo aorta, was distributed between and around small but recognizable elastic fibers. In older aortic tissue this ferritin-labelled material was seen principally in loose association with the periphery of developing elastic fibers. The microfibrils did not interact with the ferritin-antibody conjugate. Treatment of chicks with beta aminopropionitrile did not interfere with the accumulation of ferritin-labelled filaments on these small elastic fibers, suggesting that the mechanism of addition of new material onto growing fibers may be independent of cross-linking.

Amino Acids↗
PubMed1981

In vitro studies of elastin metabolism.

Neonatal pig and rat aortas were studied for their ability to synthesize elastin in an in vitro situation. Smooth muscle cells from the rat aorta produced excellent multilayered cultures and produced soluble elastin (tropoelastin), insoluble elastin, and small amounts of collagen. BAPN proved to be toxic to these cells, adversely affecting the level of extracellular protein production. Tissue minces from pig aorta continued to synthesize elastin for two hours after removal. However, a 24 hour study indicated that elastin synthesis had almost completely shut down and that collagen synthesis continued in an apparently normal fashion. It is concluded that in vitro elastin synthesis is an extremely sensitive process easily altered by culture conditions and the addition of extraneous substances such as BAPN, and also highly influenced by the past history of the smooth muscle cells involved.

Animals↗
PubMed2002

Accelerated calcification represses the expression of elastic fiber components and lysyl oxidase in cultured bovine aortic smooth muscle cells.

Vascular calcification is a common feature of advanced atherosclerosis resulting in reduced elasticity of elastic arteries. However, the relationship between elastic fibers and vascular calcification at the molecular and cellular levels remains unknown. We investigated the expression of major elastic fiber components such as tropoelastin (TE) and fibrillin-1 (FBN1) and elastin-related enzyme, lysyl oxidase (LO), in a calcification model using beta-glycerophosphate (beta-GP) in cultured bovine aortic smooth muscle cells (BASMCs). Ten mM of beta-GP stimulated calcium deposition in a time-dependent manner. As determined by Western blot analysis, 10 mM of beta-GP time-dependently decreased TE and FBN1 protein levels. TE, FBN1, and LO mRNA levels, assessed by reverse transcription-polymerase chain reaction, were also decreased by exposure to 10 mM beta-GP. Furthermore, we investigated whether the processes of calcification in BASMCs directly control these regulations. In experiments using levamisole, an alkaline phosphatase inhibitor, and DMDP, a bisphosphonate, both inhibitors inhibited down-regulation during beta-GP-induced calcification, suggesting that the down-regulation of TE, FBN1, and LO directly relates to calcium deposition. In cases of vascular calcification, the decreased expression of TE, FBN1, and LO may be partially responsible for decreased vascular elasticity and also for the decreased formation of new elastic fibers.

Alkaline Phosphatase↗
PubMed2001

[Effect of elastin peptides on the production of matrix metalloproteinase 2 by human skin fibroblasts in culture].

Soluble elastin-derived peptides from alkaline or elastase hydrolysis of insoluble elastin, as well as tropoelastin, increase matrix metalloproteinase-2 (MMP-2) production by human skin fibroblasts in culture as determined by gelatin zymography and ELISA. Such an effect is time and concentration dependent; it can be reproduced by synthetic elastin: VGVAPG, PGAIPG, and laminin: LGTIPG, hexapeptides and inhibited by lactose and is therefore elastin receptor-mediated. The steady state levels of MMP-2 mRNAs are invariant following elastin-fibroblasts interaction. Inhibition of phospholipase C (D-609), ADP-ribosylation factor (brefeldin), protein kinase C (RO-318220) and phospholipase D (1-propanol) totally abolished the elastin-mediated increase of MMP-2 production. It suggested that the post-transcriptional mechanism controlling the elastin-mediated overproduction of MMP-2 involved a cascade leading to phospholipase D activation.

ADP-Ribosylation Factors↗
PubMed1990

Elastin gene expression in elastotic human breast cancers and epithelial cell lines.

Elastosis is a prominent feature of the desmoplastic reaction in many invasive breast cancers. It is widely held that the elastic tissue is produced by fibroblastic cells of the breast stroma, but several studies have suggested that it derives from breast cancer epithelium. In studies directed to examining the mechanisms regulating desmoplasia in breast cancers, cell lines of human breast cancer derivation have been shown to synthesize immunoreactive tropoelastin in cell culture. Stromal fibroblasts, grown out from breast cancers, produced as much elastin as did nuchal ligament fibroblasts at similar passages. The human breast cancer cell lines, grown under similar conditions, produced elastin in culture at rates equivalent to 1.6-15% of those of the control fibroblastic cells. These included two estrogen receptor positive and one estrogen receptor negative cell types. Northern blot analysis of total RNA showed the presence, under high stringency conditions, of a 3.5-kilobase elastin mRNA band in both the fibroblastic cells and the cancer cell lines. In situ hybridization, with an elastin complementary RNA probe (prepared from a short segment of the translated region of human elastin mRNA), has been carried out on a selection of 21 invasive ductal breast cancers and 9 normal breast samples. It has been found that, while fibroblastic cells of the stroma and of the periductal region are responsible for elastin synthesis in most breast cancers, the malignant epithelium is a source of the elastin in the desmoplastic tissue of a significant proportion of such neoplasms. Vascular endothelium also expresses the elastin gene in some breast cancers. The elastotic elastin may have different cellular origins in different portions of a single ductal breast cancer. The results indicate that elastosis in breast cancers is very likely to be a complex process with multifactorial regulatory mechanisms. Subclassifying cancers according to the cellular source of the desmoplastic elastin, on the basis of in situ hybridization of elastin mRNA, may provide insights into the prognostic significance of elastosis in breast cancers.

Adult↗
PubMed1990

Effect of hypertension on fibronectin expression in the rat aorta.

Interactions between extracellular fibronectin and vascular cells are thought to influence the phenotype of those cells. To determine if changes in fibronectin expression accompany the phenotypic changes of vascular tissue characteristic of experimental hypertension, steady state mRNA levels for fibronectin were determined in aortae of normotensive and hypertensive rats. A 3-6-fold increase in fibronectin mRNA was observed in aortic tissue of hypertensive rats following 3 weeks of treatment with deoxycorticosterone and salt, whereas if rats were treated only with deoxycorticosterone or salt alone, no changes occurred. The changes were reversed by normalization of blood pressure. The increases observed were localized to aorta and not to the periaortic tissue. Angiotensin II infusion using osmotic minipumps also caused an increase in fibronectin expression. Age-dependent increases in aortic fibronectin mRNA occurred in several rat strains, and the combined effects of hypertension and aging were greater than either variable alone. A clear distinction between the expression of fibronectin mRNA and that for collagen or tropoelastin were found in hypertensive and aging models. Aortic fibronectin was also increased in the hypertensive rats as determined by Western blot analysis. The findings indicate that elevation in blood pressure increases fibronectin expression in rat aorta and suggest that such changes may influence the aortic cellular responses to hypertension.

Adipose Tissue, Brown↗
PubMed1989

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.

Animals↗
PubMed1987

Changes in the blood vessel wall elastin of spontaneously hypertensive (SHR) rats.

It has been demonstrated that both elastin and tropoelastin preparations obtained from aortae of spontaneously hypertensive rats at the stage of established hypertension differ in their amino acid composition from age-matching controls. The differences refer to an increased proportion of polar amino acids, particularly aspartic and glutamic acid (about a two-fold increase compared to the controls) and arginine and tyrosine (1.5 times the control value). On the other hand, this increase is compensated for by a decrease in the valine concentration. Furthermore, direct estimation of the number of val-pro sequence in different elastin preparations indicated a drop from 49.3 to 29.2 per 1,000 residues in normotensive controls and preparations obtained from spontaneously hypertensive rats respectively.

Amino Acids↗
PubMed1985

Elastin covalent structure as determined by solid phase amino acid sequencing.

The amino acid sequences of 16 large tryptic fragments of aortic tropoelastin have been determined establishing the presence of several repeating structures: GVP, GGVP, PGVGV, PGVGVA, and AGVPGFGVG. The methodologies for achieving these results by solid phase sequencing are reviewed and also the possible biologic significance of the unusual primary structures of elastin are discussed.

Amino Acid Sequence↗
PubMed1978

Cyclic analog of elastin polyhexapeptide exhibits an inverse temperature transition leading to crystallization.

The cyclic dodecapeptide analog of the linear polyhexapeptide of tropoelastin crystallizes from water on raising of the temperature and thereby demonstrates an inverse temperature transition implying dominant intermolecular hydrophobic interactions. The temperature profiles of turbidity (TP tau) of the cyclododecapeptide are analogous to those of the polyhexapeptide where increases in concentration lead to translations of the profiles to lower temperature without sharpening of the transition. The demonstration of increase in order with increase in temperature for the cyclododecapeptide in water and the similarity of TPtau's lends credence to the view that increases in temperature lead to increases in order, specifically, for the linear polyhexapeptide and, generally, for the precursor protein of the elastic fiber wherein the repeating hexapeptide occurs.

Crystallization↗
PubMed1994

Consequences of prolonged inhalation of ozone on F344/N rats: collaborative studies. Part IV: Effects on expression of extracellular matrix genes.

Increased deposition of lung extracellular matrix in terminal airways is associated with chronic ozone exposure. In situ hybridization was used to assess whether long-term ozone exposure causes elevated and continued expression of genes coding for connective tissue proteins. Accessory lobes were removed from the animals exposed to 0, 0.12, 0.5, or 1.0 parts per million (ppm)* ozone for 20 months as part of the National Toxicology Program (NTP)/HEI Collaborative Ozone Project. The lungs were perfused fixed under physiologic pressure and processed for in situ hybridization. Sections were hybridized with 35S-labeled probes for messenger RNA (mRNA) coding for various matrix proteins, including collagen types I and III, elastin, and fibronectin, and for interstitial collagenase, a matrix metalloproteinase. Fetal rat lung was used as a positive control for hybridization. No signal for any mRNA was detected in terminal airway stromal cells of lungs from animals exposed to ozone for 20 months or control animals breathing clean air. In all samples from animals exposed to ozone for 20 months and control animals, only a very weak signal was seen in occasional cells within the interstitial spaces around large airways and blood vessels. In contrast, a strong signal for matrix-related mRNA was detected in fetal lung tissue. These findings indicate that active or enhanced matrix production is turned off in the adult animals used in the ozone studies, suggesting that the increase in matrix deposition results from a transient and early fibrotic response. Indeed, signal for type I procollagen and tropoelastin mRNAs was seen in alveolar septal cells in lungs of rats exposed to ozone for two months. No signal was seen in alveolar cells of age-matched control animals. (These animals, exposed for two months, and age-matched controls were from earlier studies supported by the HEI.) These findings indicate that ozone mediates a transient fibrotic response that results in a sustained increase in lung extracellular matrix. Confirmation of this hypothesis would require additional studies using animals exposed to ozone for shorter times.

Administration, Inhalation↗
PubMed1983

Microfibrillar protein and phospholipid in granular corneal dystrophy.

Keratoplasty specimens from eight patients with granular corneal dystrophy (GCD) and age-matched control subjects were examined by combinations of immunohistological stains, transmission electron microscopy (TEM), and sodium dodecyl sulfate gel electrophoresis. Fresh frozen sections from corneas with GCD stained positively with antibodies to microfibrillar protein by immunofluorescence. Routine TEM disclosed that the granules had central electron-dense areas partially surrounded by 9- to 10-nm tubular microfibrils. Material eluted from corneas with GCD showed denser peptide bands at 65 and 110 kilo than in normal corneas. Stains were negative for elastin, amyloid, neutral lipids, cholesterol, and glycosaminoglycan. Luxol fast blue MBSN stain was strongly positive in the granules in all cases examined. Immunofluorescent stains were negative with antibodies to plasma fibronectin (cold insoluble globulin), laminin, collagens I to V, basement membrane proteoglycan, tropoelastin, and keratin. In two corneas with GCD an increased lipid content was found in every phospholipid class, although cholesterol content was unchanged. Alterations in the fatty acid profiles of phospholipids were also observed.

Contractile Proteins↗
PubMed1996

Immunolocalization of vinculin in the heart of the early developing rat embryo.

BACKGROUND: The interaction of cells and extracellular matrix (ECM) components is important in the morphogenesis of the developing heart and is thought be mediated in part by adhesion plaques associated with vinculin, paxillin, talin, integrin, and other proteins. We investigated the patterns of expression of vinculin in the early embryonic rat heart to evaluate the role of vinculin in cardiac morphogenesis. METHODS: Vinculin expression was studied immunohistochemically in developing Sprague-Dawley embryonic rat hearts between days 11.5 and 15.5. RESULTS: Vinculin expression was transient and specific in the aorticopulmonary septum on day 13.5 and in the conal septum on day 14.5, when the respective septations were complete. Less vinculin immunoreactivity was detected in the atrioventricular cushion or ventricular septum, where obvious morphological alteration takes place. CONCLUSIONS: Sites that were immunoreactive for vinculin in the present study are reportedly associated with the distribution of neural cell adhesion molecules (N-CAM) or of soluble tropoelastin. Thus vinculin appears to play a key role in aorticopulmonary septation, where neural crest cells are transformed into ectomesenchyme. Vinculin appears to be involved in elastogenesis and is contributed by ectomesenchyme derived from the neural crest cells.

Animals↗
PubMed2000

Characterizations of critical processes in liquid-liquid phase separation of the elastomeric protein-water system: microscopic observations and light scattering measurements.

Biological self-assembly process of tropoelastin in an extracellular space, viewed as a key step of the elastogenesis, can be mimicked by the temperature-dependent coacervation of the elastin-related polypeptide-water system. Early and late stages of the phase separation behavior of the bovine neck ligamental alpha-elastin-water system were examined respectively by the laser light scattering photometry and phase contrast microscopy. Changes in the hydrodynamic size of molecular assemblies and visible microcoacervate droplet size were traced as a function of the concentration of alpha-elastin and temperature. Near the critical point, alpha-elastin concentration of 0.11 mg/mL and temperature of 21.5 degrees C, the phase separation was initiated after fast increase of the hydrodynamic size of primary aggregates as scattering particles and followed by the appearance of larger microcoacervate droplets with a broad size distribution. Whereas in the off-critical region, slow decrease of the hydrodynamic size of primary particles induced phase separation with smaller droplets of a narrow size distribution. Observation of the phase separation processes in the alpha-elastin-water system with metal chlorides and hydrophobic synthetic model polypeptide-water system indicated that the fast and slow molecular assembly processes were based on the fundamental hydrophobic interactions and involvements of electrostatic interactions between charged amino acid residues, respectively.

Animals↗
PubMed1999

Galectin-3 regulates the adhesive interaction between breast carcinoma cells and elastin.

Galectin-3 is a beta-galactoside binding lectin whose precise physiological role is not yet defined. In the present studies, we questioned whether galectin-3 plays a role in the adhesion of breast carcinoma cells to elastin. The impetus for this analysis was the initial observation that the cellular receptor for elastin, the 67 kDa elastin/laminin protein may have galectin-like properties (Mecham et al. [1989] J. Biol. Chem. 264:16652-16657). We therefore analyzed the adhesion of breast carcinoma cells to microtiter wells coated with elastin under conditions which eliminate integrin participation in adhesion. The adhesion assay was done in the absence and presence of purified recombinant galectin-3. We hereby demonstrate that high concentrations of galectin-3 ligate breast carcinoma cells to microtiter wells coated with elastin. Galectin-3 also demonstrated a specific binding interaction with purified elastin in a dose and lactose dependent manner. Furthermore we demonstrated by immunoprecipitation that endogenous galectin-3 in breast carcinoma cells is associated with tropoelastin. Lastly, the breast carcinoma cells which expressed galectin-3 on their surface, demonstrated enhanced cellular proliferation on elastin compared to galectin-3 null expressing cells. These studies suggest that galectin-3 is capable of regulating the interactions between cells and elastin.

Antigens, Differentiation↗
PubMed1999

Astrocytoma cell interaction with elastin substrates: implications for astrocytoma invasive potential.

Elastin has been identified within the meninges and the microvasculature of the normal human brain. However, the role that elastin plays in either facilitating astrocytoma cell attachment to these structures or modulating astrocytoma invasion has not been previously characterized. We have recently shown that astrocytoma cell lines and specimens produce tropoelastin, and express the 67 kDa elastin binding protein (EBP). In the present report, we have established that astrocytoma cells attach to elastin as a substrate in vitro. The U87 MG astrocytoma cell line demonstrated the greatest degree of adhesion. In addition, all astrocytoma cell lines examined were capable of penetrating and migrating through an intact elastin membrane, and of degrading tritiated-elastin, a process that could be prevented by the pre-incubation of astrocytoma cells with EDTA, but not with alpha1-antitrypsin. Astrocytoma cells were also capable of penetrating 1 mm sections of human brain tissue maintained as organotypic cultures. Interestingly, the invasive potential of cultured astrocytoma cells plated on organotypic cultures of human brain was significantly increased after exposure to elastin degradation products (kappa-elastin), which interact with astrocytoma cell surface EBP. Our data show that astrocytoma cells express a functional 67 kDa EBP, enabling them to potentially recognize and attach to elastin as a substrate. These data also suggest that this elastin receptor may be involved in processes which regulate regional astrocytoma invasion.

Astrocytoma↗
PubMed2003

Recombinant human elastin polypeptides self-assemble into biomaterials with elastin-like properties.

Processes involving self-assembly of monomeric units into organized polymeric arrays are currently the subject of much attention, particularly in the areas of nanotechnology and biomaterials. One biological example of a protein polymer with potential for self-organization is elastin. Elastin is the extracellular matrix protein that imparts the properties of extensibility and elastic recoil to large arteries, lung parenchyma, and other tissues. Tropoelastin, the approximately 70 kDa soluble monomeric form of elastin, is highly nonpolar in character, consisting essentially of 34 alternating hydrophobic and crosslinking domains. Crosslinking domains contain the lysine residues destined to form the covalent intermolecular crosslinks that stabilize the polymer. We and others have suggested that the hydrophobic domains are sites of interactions that contribute to juxtaposition of lysine residues in preparation for crosslink formation. Here, using recombinant polypeptides based on sequences in human elastin, we demonstrate that as few as three hydrophobic domains flanking two crosslinking domains are sufficient to support a self-assembly process that aligns lysines for zero-length crosslinking, resulting in formation of the crosslinks of native elastin. This process allows fabrication of a polymeric matrix with solubility and mechanical properties similar to those of native elastin.

Biocompatible Materials↗