Elastin is Localised to the Interfascicular Matrix of Energy Storing Tendons and Becomes Increasingly Disorganised With Ageing

Tendon is composed of fascicles bound together by the interfascicular matrix (IFM). Energy storing tendons are more elastic and extensible than positional tendons; behaviour provided by specialisation of the IFM to enable repeated interfascicular sliding and recoil. With ageing, the IFM becomes stiffer and less fatigue resistant, potentially explaining why older tendons become more injury-prone. Recent data indicates enrichment of elastin within the IFM, but this has yet to be quantified. We hypothesised that elastin is more prevalent in energy storing than positional tendons, and is mainly localised to the IFM. Further, we hypothesised that elastin becomes disorganised and fragmented, and decreases in amount with ageing, especially in energy storing tendons. Biochemical analyses and immunohistochemical techniques were used to determine elastin content and organisation, in young and old equine energy storing and positional tendons. Supporting the hypothesis, elastin localises to the IFM of energy storing tendons, reducing in quantity and becoming more disorganised with ageing. These changes may contribute to the increased injury risk in aged energy storing tendons. Full understanding of the processes leading to loss of elastin and its disorganisation with ageing may aid in the development of treatments to prevent age related tendinopathy

Spatial and temporal changes in extracellular elastin and laminin distribution during lung alveolar development

Lung alveolarization requires precise coordination of cell growth with extracellular matrix (ECM) synthesis and deposition. The role of extracellular matrices in alveogenesis is not fully understood, because prior knowledge is largely extrapolated from two-dimensional structural analysis. Herein, we studied temporospatial changes of two important ECM proteins, laminin and elastin that are tightly associated with alveolar capillary growth and lung elastic recoil respectively, during both mouse and human lung alveolarization. By combining protein immunofluorescence staining with two- and three-dimensional imaging, we found that the laminin network was simplified along with the thinning of septal walls during alveogenesis, and more tightly associated with alveolar endothelial cells in matured lung. In contrast, elastin fibers were initially localized to the saccular openings of nascent alveoli, forming a ring-like structure. Then, throughout alveolar growth, the number of such alveolar mouth ring-like structures increased, while the relative ring size decreased. These rings were interconnected via additional elastin fibers. The apparent patches and dots of elastin at the tips of alveolar septae found in two-dimensional images were cross sections of elastin ring fibers in the three-dimension. Thus, the previous concept that deposition of elastin at alveolar tips drives septal inward growth may potentially be conceptually challenged by our data

The changes in chemical composition during development of the bovine nuchal ligament

Whole bovine nuchal ligaments, or portions thereof (in the case of commercially valuable animals), were obtained from 45 animals (28 fetal and 17 postnatal) ranging in age from 110 days of gestation to 10 yr. Insoluble elastin was quantitatively prepared from the fresh ligaments by extraction with hot alkali and by a combination of multiple extractions with alkaline buffer and then repeated autoclaving. When adult samples were examined, the yields of insoluble residue by these two methods were very similar, but with young fetal samples the second method gave significantly higher values, because of incomplete purification of the elastin residue. The changes in the concentration of collagen, alkali-insoluble elastin, and DNA have been examined. DNA concentration, and, thus, cell population density, fell progressively during the fetal period of development, to reach a steady value soon after birth. Collagen appeared in appreciable quantities before elastin, but its concentration was rapidly halved at about the time of birth. Insoluble elastin concentration was low until the end of the 7th fetal month, at which time it began to rise rapidly. The rate of increase in elastin concentration remained high throughout the next 10–12 wk, by which time the adult value had been reached. Quantitative studies, on the basis of the whole ligament, showed that the total cell content rises to a maximum at birth, but falls soon after to a level about half that at birth. Total collagen production and elastin deposition continue at a steady, maximal rate over the interval from 235 days of gestation to the end of the 1st postnatal month. It is concluded that the immediate postnatal period would be the most favorable phase in which to attempt the isolation of the soluble precursor elastin

Plasma Soluble Human Elastin Fragments as an Intra-Aneurysmal Localized Biomarker for Ruptured Intracranial Aneurysm

Background—Fragmentation of the tunica media is a hallmark of intracranial aneurysm formation, often leading to aneurysmal progression and subsequent rupture. The objective of this study is to determine the plasma level of elastin fragments in the lumen of ruptured versus unruptured human intracranial aneurysms. Methods and Results—One hundred consecutive patients with/without ruptured saccular intracranial aneurysms undergoing endovascular coiling or stent-assisted coiling were recruited. Blood samples were collected from the lumen of intracranial aneurysm using a microcatheter. The tip of the microcatheter was placed inside the aneurysm’s sac in close proximity to the inner wall of the dome. Plasma levels of elastin fragments were measured using an ELISA-based method. Mean plasma level of soluble human elastin fragments was significantly greater in ruptured aneurysms when compared with nonruptured aneurysms (102.0±15.5 versus 39.3±9.6 ng/mL; P\u3c0.001). Mean plasma level of soluble human elastin fragments did not have significant correlation with age, sex, size, or aneurysm location. Conclusions—The present study revealed that a significantly higher concentration of soluble human elastin fragments in the lumen of ruptured intracranial aneurysms when compared with nonruptured ones. © 2018 The Authors

Fibulin-4 is essential for maintaining arterial wall integrity in conduit but not muscular arteries

Homozygous or compound heterozygous mutations in fibulin-4 (FBLN4) lead to autosomal recessive cutis laxa type 1B (ARCL1B), a multisystem disorder characterized by significant cardiovascular abnormalities, including abnormal elastin assembly, arterial tortuosity, and aortic aneurysms. We sought to determine the consequences of a human disease-causing mutation in FBLN4 (E57K) on the cardiovascular system and vascular elastic fibers in a mouse model of ARCL1B. Fbln4E57K/E57K mice were hypertensive and developed arterial elongation, tortuosity, and ascending aortic aneurysms. Smooth muscle cell organization within the arterial wall of large conducting vessels was abnormal, and elastic fibers were fragmented and had a moth-eaten appearance. In contrast, vessel wall structure and elastic fiber integrity were normal in resistance/muscular arteries (renal, mesenteric, and saphenous). Elastin cross-linking and total elastin content were unchanged in large or small arteries, whereas elastic fiber architecture was abnormal in large vessels. While the E57K mutation did not affect Fbln4 mRNA levels, FBLN4 protein was lower in the ascending aorta of mutant animals compared to wild-type arteries but equivalent in mesenteric arteries. We found a differential role of FBLN4 in elastic fiber assembly, where it functions mainly in large conduit arteries. These results suggest that elastin assembly has different requirements depending on vessel type. Normal levels of elastin cross-links in mutant tissue call into question FBLN4\u27s suggested role in mediating lysyl oxidase-elastin interactions. Future studies investigating tissuespecific elastic fiber assembly may lead to novel therapeutic interventions for ARCL1B and other disorders of elastic fiber assembly. 2017 © The Authors, some rights reserved

Physiologic compliance in engineered small-diameter arterial constructs based on an elastomeric substrate.

Compliance mismatch is a significant challenge to long-term patency in small-diameter bypass grafts because it causes intimal hyperplasia and ultimately graft occlusion. Current engineered grafts are typically stiff with high burst pressure but low compliance and low elastin expression. We postulated that engineering small arteries on elastomeric scaffolds under dynamic mechanical stimulation would result in strong and compliant arterial constructs. This study compares properties of engineered arterial constructs based on biodegradable polyester scaffolds composed of either rigid poly(lactide-co-glycolide) (PLGA) or elastomeric poly(glycerol sebacate) (PGS). Adult baboon arterial smooth muscle cells (SMCs) were cultured in vitro for 10 days in tubular, porous scaffolds. Scaffolds were significantly stronger after culture regardless of material, but the elastic modulus of PLGA constructs was an order of magnitude greater than that of porcine carotid arteries and PGS constructs. Deformation was elastic in PGS constructs and carotid arteries but plastic in PLGA constructs. Compliance of arteries and PGS constructs were equivalent at pressures tested. Altering scaffold material from PLGA to PGS significantly decreased collagen content and significantly increased insoluble elastin content in constructs without affecting soluble elastin concentration in the culture medium. PLGA constructs contained no appreciable insoluble elastin. This research demonstrates that: (1) substrate stiffness directly affects in vitro tissue development and mechanical properties; (2) rigid materials likely inhibit elastin incorporation into the extracellular matrix of engineered arterial tissues; and (3) grafts with physiologic compliance and significant elastin content can be engineered in vitro after only days of cell culture

Dill extract induces elastic fiber neosynthesis and functional improvement in the ascending aorta of aged mice with reversal of age-dependent cardiac hypertrophy and involvement of lysyl oxidase-like-1

Elastic fibers (90% elastin, 10% fibrillin-rich microfibrils) are synthesized only in early life and adolescence mainly by the vascular smooth muscle cells through the cross-linking of its soluble precursor, tropoelastin. Elastic fibers endow the large elastic arteries with resilience and elasticity. Normal vascular aging is associated with arterial remodeling and stiffening, especially due to the end of production and degradation of elastic fibers, leading to altered cardiovascular function. Several pharmacological treatments stimulate the production of elastin and elastic fibers. In particular, dill extract (DE) has been demonstrated to stimulate elastin production in vitro in dermal equivalent models and in skin fibroblasts to increase lysyl oxidase-like-1 (LOXL-1) gene expression, an enzyme contributing to tropoelastin crosslinking and elastin formation. Here, we have investigated the effects of a chronic treatment (three months) of aged male mice with DE (5% or 10

Analysis of the molecular mobility of collagen and elastin in safe, atheromatous and aneurysmal aortas

Aim of the study : In this study, we propose to use a thermal technique, Differential Scanning Calorimetry (DSC) to follow the evolution of elastin and collagen in safe and pathological cardiovascular tissues. Patients and methods : The first part of this study deals with the analysis of the elastin network and associated proteins during ageing (from children to old persons) in aortic walls. The second part is devoted to the characterization of the collagenic phase in aneurysms. In both cases, physical data are correlated with biochemical analyses. Results and conclusion : For old persons aortas with atheromatous stades, elastin and associated proteins are found to interpenetrate to form a homogenous phase. Abdominal aortic aneurysms (AAA) are characterized by structural alterations of the aortic wall resulting from the degradation of elastic fibers and an increase of collagen/elastin ratio. Notable modifications are evidenced between collagen from control tissue and collagen from AAA, particularly concerning the thermal denaturation. Biochemical and thermal results are compatible with the increase of new collagen deposition and/or impairment of the collagen phase stability in the extracellular matrix of AAAs

Changes in the physical structure and chain dynamics of elastin network in homocysteine-cultured arteries

The thermal and dielectric properties of the elastin network were investigated in arteries cultured with physiological and pathological concentrations of homocysteine, an aminoacid responsible of histological impairments in human arteries. The physical structure of this amorphous protein was investigated by differential scanning calorimetry (DSC). To explore the molecular dynamics of the elastin network in the nanometer range, we used thermally stimulated currents (TSC), a dielectric technique running at low frequency, and measuring the dipolar reorientations in proteins subjected to a static electrical field. Combining DSC and TSC experiments reveals the molecular mobility of the proteins, both in the glassy state and in the liquid state. Significant differences are evidenced in the physical structure and relaxation behavior of elastin network in cultured arteries (physiological and pathological concentrations of homocysteine) and discussed

Influence of homocysteine on the physical structure and molecular mobility of elastin network in cultured arteries

The thermal and dielectric properties of the elastin network were investigated in arteries cultured with physiological and pathological concentrations of homocysteine, an aminoacid responsible of histological impairments in human arteries. The glass transition of this amorphous protein was investigated by Differential Scanning Calorimetry (DSC). To explore the molecular dynamics of the elastin network in the nanometer range, we used Thermally Stimulated Currents (TSC), a dielectric technique running at low frequency and measuring the dipolar reorientations in proteins subjected to a static electrical field. Combining TSC and DSC experiments with determination of the activation parameters of relaxation times reveals the molecular mobility of the proteins. The major differences in the relaxation behavior of elastin between arteries cultured with physiological and pathological concentrations of homocysteine are discussed