ANTHONY WEISS

Aggrecan expression is substantially and abnormally upregulated in Hutchinson-Gilford Progeria Syndrome dermal fibroblasts

Publisher: Elsevier BV

Date: 08-2006

DOI: 10.1016/J.MAD.2006.03.004

Abstract: Hutchinson-Gilford Progeria syndrome (HGPS) is a rare genetic disorder that displays features of segmental aging. It is manifested predominantly in connective tissue, with most prominent histological changes occurring in the skin, cartilage, bone and cardiovascular tissues. Detailed quantitative real time reverse-transcription polymerase chain reaction studies confirmed the previous observation that platelet-derived growth factor A-chain transcripts are consistently elevated 11+/-2- to 13+/-2-fold in two HGPS dermal fibroblast lines compared with age-matched controls. Furthermore, we identified two additional genes with substantially altered transcript levels. Nucleotide pyrophosphatase transcription was virtually shut down with decreased expression of 13+/-3- to 59+/-3-fold in HGPS, whereas aggrecan mRNA was elevated to 24+/-5 times to 41+/-4 times that of chronologically age-matched controls. Aggrecan, normally a component of cartilage and not always detectable in normal fibroblasts cultures, was secreted by HGPS fibroblast lines and was produced as a proteoglycan. This demonstrates that elevated aggrecan expression and its secretion are aberrant features of HGPS. We conclude that HGPS cells can display massively altered transcript levels leading to the secretion of inappropriate protein species.

Zipping up transcription factors: Rational design of anti-Jun and anti-Fos peptides

Publisher: Springer Science and Business Media LLC

Date: 04-1997

DOI: 10.1007/BF02443517

Covalent Biofunctionalization of the Inner Surfaces of a Hollow-Fiber Capillary Bundle Using Packed-Bed Plasma Ion Implantation

Publisher: American Chemical Society (ACS)

Date: 12-06-2020

DOI: 10.1021/ACSAMI.0C07070

Applications of Engineering Techniques in Microvasculature Design

Publisher: Frontiers Media SA

Date: 26-04-2021

DOI: 10.3389/FCVM.2021.660958

Abstract: Achieving successful microcirculation in tissue engineered constructs in vitro and in vivo remains a challenge. Engineered tissue must be vascularized in vitro for successful inosculation post-implantation to allow instantaneous perfusion. To achieve this, most engineering techniques rely on engineering channels or pores for guiding angiogenesis and capillary tube formation. However, the chosen materials should also exhibit properties resembling the native extracellular matrix (ECM) in providing mechanical and molecular cues for endothelial cells. This review addresses techniques that can be used in conjunction with matrix-mimicking materials to further advance microvasculature design. These include electrospinning, micropatterning and bioprinting. Other techniques implemented for vascularizing organoids are also considered for their potential to expand on these approaches.

Mechanical Properties of Plasma Immersion Ion Implanted PEEK for Bioactivation of Medical Devices

Publisher: American Chemical Society (ACS)

Date: 07-10-2015

DOI: 10.1021/ACSAMI.5B06395

Abstract: Plasma immersion ion implantation (PIII) is used to modify the surface properties of polyether ether ketone for biomedical applications. Modifications to the mechanical and chemical properties are characterized as a function of ion fluence (treatment time) to determine the suitability of the treated surfaces for biological applications. Young's modulus and elastic recovery were found to increase with respect to treatment time at the surface from 4.4 to 5.2 MPa and from 0.49 to 0.68, respectively. The mechanical properties varied continuously with depth, forming a graded layer where the mechanical properties returned to untreated values deep within the layer. The treated surface layer exhibited cracking under cyclical loads, associated with an increased modulus due to dehydrogenation and cross-linking however, it did not show any sign of delamination, indicating that the modified layer is well integrated with the substrate, a critical factor for bioactive surface coatings. The oxygen concentration remained unchanged at the surface however, in contrast to ion implanted polymers containing only carbon and hydrogen, the oxygen concentration within the treated layer was found to decrease. This effect is attributed to UV exposure and suggests that PIII treatments can modify the surface to far greater depths than previously reported. Protein immobilization on PIII treated surfaces was found to be independent of treatment time, indicating that the surface mechanical properties can be tuned for specific applications without affecting the protein coverage. Our findings on the mechanical properties demonstrate such treatments render PEEK well suited for use in orthopedic implantable devices.

Restriction map of DNA spanning the replication terminus of the Bacillus subtilis chromosome

Publisher: Elsevier BV

Date: 12-1983

DOI: 10.1016/S0022-2836(83)80349-0

Abstract: The Bacillus subtilis 168 dna-1 chromosome was labelled during sporulation with [3H]thymine for five minutes immediately before termination of replication. The isolated radioactive DNA was cleaved with BamHI (or SalI) and the resulting restriction fragments separated by agarose gel electrophoresis. The in idual fragments, fractionated into a series of slices cut from the gel, were then cleaved with SalI (or BamHI) and the double-digest fragments identified by electrophoresis and fluorography. All major fragments and most minor ones present in a whole double-digest were assigned to BamHI and SalI parents. Such information enabled the construction of an unambiguous restriction map of 150 X 10(3) bases of the approximately 250 X 10(3) bases of DNA labelled in the five minutes. In conjunction with published data on the order of replication of restriction fragments as termination is approached, it was clear that most (105 X 10(3) bases) of the mapped DNA was replicated by a major fork moving in one direction towards a BamHI 24.8 X 10(3) base fragment. The 45 X 10(3) bases extending to the other side of this region were labelled only slightly, and presumably was replicated by a fork that approached the other in an opposite direction until its progress was blocked or severely impeded within this region at a site, referred to as terC, sometime (less than 5 min) earlier. The regions of the map replicated in the final 2.5 and 1.0 minute by the major fork were also identified.

Characterization of an acetylcholine receptor gene of haemonchus contortus in relation to levamisole resistance

Publisher: Elsevier BV

Date: 02-1997

DOI: 10.1016/S0166-6851(96)02793-4

Abstract: The anthelminitic drug levamisole is thought to bind to nicotinic acetylcholine receptors of nematodes. It is possible that resistance to this drug is associated with either a change in binding characteristics or a reduction in the number of nicotinic acetylcholine receptors. Therefore, the molecular mechanism of levamisole resistance in the parasitic nematode Haemonchus contortus was studied by isolating and characterising cDNA clones encoding a putative ligand binding nicotinic acetylcholine receptor subunit, HCAl, of two susceptible and one levamisole resistant population. Hcal is related to unc-38, a nicotinic acetylcholine receptor subunit gene associated with levamisole resistance in Caenorhabditis elegans. Although extensive sequence analyses of hcal sequences revealed polymorphism at amino acid level, no association with levamisole resistance could be detected. Restriction fragment length polymorphism analyses confirmed that, although polymorphism was detected, no selection of a specific allele of hcal has taken place during selection for levamisole resistance in various levamisole resistant populations.

Transposon-mediated restriction mapping of the Bacillus subtilis chromosome

Publisher: Elsevier BV

Date: 05-1989

DOI: 10.1016/0378-1119(89)90311-9

Abstract: Analysis of chromosomal DNA depends upon a knowledge of the locations of restriction sites over several thousand kilobases (kb). However determination of even a subset of these sites can be time-consuming, and it can be difficult to link genetic and physical maps. We describe here a significant improvement which can be used in concert with genetically mapped chromosomal insertions. The circular chromosome of Bacillus subtilis 168 was physically examined on contour-cl ed homogeneous electric field (CHEF) gels using the restriction enzyme NotI. Restriction mapping of the 4.7-megabase (Mb) DNA was accomplished using a novel technique involving the transposon Tn917, which linked the genetic and physical maps and also significantly increased the rate at which this was performed. The DNA of 54 strains which contained Tn917 at genetically determined locations was cleaved with NotI and used to determine the approximate positions of 31 restriction fragments with sizes between 45 kb and 290 kb, totalling 3589 kb. This information should greatly assist in the construction of a more detailed map using standard methodology.

Comparison of the morphology and histopathology of large nonpedunculated colorectal polyps in the rectum and colon: implications for endoscopic treatment

Publisher: Elsevier BV

Date: 07-2022

DOI: 10.1016/J.GIE.2022.02.022

Abstract: The risk of cancer in large nonpedunculated colorectal polyps ≥20 mm (LNPCPs) in the rectum relative to the remainder of the colon is unknown. We aimed to describe differences between rectal and colonic LNPCPs to better inform treatment decisions. Patients with LNPCPs referred to tertiary centers for endoscopic resection within a prospective, multicenter, observational cohort were evaluated. Data recorded were participant demographics, LNPCP location, morphology, resection modality, and histopathologic data. Multiple logistic regression analysis was used to identify those variables independently associated with rectal versus nonrectal location in the colon. Patients with LNPCPs referred for endoscopic resection between July 2008 and July 2021 were included. Rectal LNPCPs (n = 618) were larger (median size, 40 mm vs 30 mm P < .001) and more likely to be granular (79% vs 50%, P < .001) with a nodular component (53% vs 17%, P < .001) compared with nonrectal LNPCPs (n = 2787). Rectal LNPCPs were more likely to have tubulovillous histopathology (72% vs 47%, P < .001) and contain cancer (15% vs 6%, P < .001). After adjusting for the other features independently associated with location, cancer was more common in the rectum compared with the colon (odds ratio, 1.77 95% confidence interval, 1.25-2.53). This study suggests that compared with LNPCPs in the rest of the colon, rectal LNPCPs are more likely to be larger and contain more advanced pathology. These findings have implications for curative endoscopic resection techniques particularly where early cancer is present. (Clinical trial registration numbers: NCT01368289 and NCT02000141.).

Silk-tropoelastin protein films for nerve guidance.

Publisher: Elsevier BV

Date: 03-2015

DOI: 10.1016/J.ACTBIO.2014.11.045

Domains 12 to 16 of tropoelastin promote cell attachment and spreading through interactions with glycosaminoglycan and integrins alphaV and alpha5beta1

Publisher: Wiley

Date: 26-01-2021

DOI: 10.1111/FEBS.15702

Abstract: Elastin is an extracellular matrix component with key structural and biological roles in elastic tissues. Interactions between resident cells and tropoelastin, the monomer of elastin, underpin elastin's regulation of cellular processes. However, the nature of tropoelastin–cell interactions and the contributions of in idual tropoelastin domains to these interactions are only partly elucidated. In this study, we identified and characterized novel cell‐adhesive sites in the tropoelastin N‐terminal region between domains 12 and 16. We found that this region interacts with αV and α5β1 integrin receptors, which mediate cell attachment and spreading. A peptide sequence from within this region, spanning domains 14 to mid‐domain 16, binds heparan sulfate through electrostatic interactions with peptide lysine residues and induces conformational ordering of the peptide. We propose that domains 14–16 direct initial cell attachment through cell‐surface heparan sulfate glycosaminoglycans, followed by αV and α5β1 integrin‐promoted attachment and spreading on domains 12–16 of tropoelastin. These findings advance our mechanistic understanding of elastin matrix biology, with the potential to enhance tissue regenerative outcomes of elastin‐based materials.

Synthetic elastin hydrogels derived from massive elastic assemblies of self-organized human protein monomers

Publisher: Elsevier BV

Date: 09-2004

DOI: 10.1016/J.BIOMATERIALS.2004.01.055

Synthetic elastin hydrogels that are coblended with heparin display substantial swelling, increased porosity, and improved cell penetration

Publisher: Wiley

Date: 11-10-2010

DOI: 10.1002/JBM.A.32950

Abstract: Synthetic elastin hydrogels are useful tissue engineering scaffolds because they present cell binding sequences and display physical performance similar to that of human elastic tissue. Small pores and a low porosity can limit cellular penetration into elastin scaffolds. To overcome this problem, glycosaminoglycans were coblended with tropoelastin during the formation of synthetic elastin hydrogels. Heparin and dermatan sulfate increased the pore size and porosity of the hydrogels. Heparin was particularly effective as it enlarged the pore size from 6.6 ± 2.1 μm to 23.8 ± 8.5 μm, and generated structures occasionally separated by finely fenestrated thin walls, which allowed human dermal fibroblast cells to migrate as deep as ∼300 μm into the hydrogel under diffusion-limiting static culture conditions. Most cells displayed spindle-like morphology, appeared histologically normal and presented intact nuclei, as expected for a viable population. Hydrogel swelling studies showed that each of the hydrogels contracted as the temperature was raised from 4°C to 37°C synthetic elastin-heparin was least affected by temperature with a contraction of only 22.4 ± 1.2%, which would facilitate its transition from cold storage to body temperature. All hydrogels displayed similar compression moduli of 5.5 ± 0.4 to 6.9 ± 0.6 kPa. Compressive elastic energy losses for synthetic elastin-heparin and synthetic elastin were 33.7 ± 1.3% and 31.7 ± 2.2% respectively.

Free radical functionalization of surfaces to prevent adverse responses to biomedical devices

Publisher: Proceedings of the National Academy of Sciences

Date: 15-08-2011

DOI: 10.1073/PNAS.1103277108

Abstract: Immobilizing a protein, that is fully compatible with the patient, on the surface of a biomedical device should make it possible to avoid adverse responses such as inflammation, rejection, or excessive fibrosis. A surface that strongly binds and does not denature the compatible protein is required. Hydrophilic surfaces do not induce denaturation of immobilized protein but exhibit a low binding affinity for protein. Here, we describe an energetic ion-assisted plasma process that can make any surface hydrophilic and at the same time enable it to covalently immobilize functional biological molecules. We show that the modification creates free radicals that migrate to the surface from a reservoir beneath. When they reach the surface, the radicals form covalent bonds with biomolecules. The kinetics and number densities of protein molecules in solution and free radicals in the reservoir control the time required to form a full protein monolayer that is covalently bound. The shelf life of the covalent binding capability is governed by the initial density of free radicals and the depth of the reservoir. We show that the high reactivity of the radicals renders the binding universal across all biological macromolecules. Because the free radical reservoir can be created on any solid material, this approach can be used in medical applications ranging from cardiovascular stents to heart-lung machines.

The action of neutrophil serine proteases on elastin and its precursor

Publisher: Elsevier BV

Date: 2012

DOI: 10.1016/J.BIOCHI.2011.10.006

Abstract: This study aimed to investigate the degradation of the natural substrates tropoelastin and elastin by the neutrophil-derived serine proteases human leukocyte elastase (HLE), proteinase 3 (PR3) and cathepsin G (CG). Focus was placed on determining their cleavage site specificities using mass spectrometric techniques. Moreover, the release of bioactive peptides from elastin by the three proteases was studied. Tropoelastin was comprehensively degraded by all three proteases, whereas less cleavage occurred in mature cross-linked elastin. An analysis of the cleavage site specificities of the three proteases in tropoelastin and elastin revealed that HLE and PR3 similarly tolerate hydrophobic and/or aliphatic amino acids such as Ala, Gly and Val at P(1), which are also preferred by CG. In addition, CG prefers the bulky hydrophobic amino acid Leu and accepts the bulky aromatic amino acids Phe and Tyr. CG shows a strong preference for the charged amino acid Lys at P(1) in tropoelastin, whereas Lys was not identified at P(1) in CG digests of elastin due to extensive cross-linking at Lys residues in mature elastin. All three serine proteases showed a clear preference for Pro at P(2) and P(4)'. With respect to the liberation of potentially bioactive peptides from elastin, the study revealed that all three serine proteases have a similar ability to release bioactive sequences, with CG producing the highest number of these peptides. In bioactivity studies, potentially bioactive peptides that have not been investigated on their bioactivity to date, were tested. Three new bioactive GxxPG motifs were identified GVYPG, GFGPG and GVLPG.

A potential role for endogenous proteins as sacrificial sunscreens and antioxidants in human tissues

Publisher: Elsevier BV

Date: 08-2015

DOI: 10.1016/J.REDOX.2015.04.003

Plasma treatment in air at atmospheric pressure that enables reagent-free covalent immobilization of biomolecules on polytetrafluoroethylene (PTFE)

Publisher: Elsevier BV

Date: 07-2020

DOI: 10.1016/J.APSUSC.2020.146128

Impediment to replication fork movement in the terminus region of the Bacillus subtilis chromosome.

Publisher: Elsevier BV

Date: 11-1984

DOI: 10.1016/0022-2836(84)90165-7

Abstract: The terminus regions of the chromosomes of three strains of Bacillus subtilis 168 were radioactively labelled by supplying [3H]thymine towards the end of a round of replication. These strains lacked or contained the prophage SP beta c2. Following restriction endonuclease digestion of the purified DNA and fluorography, an SP beta c2-related perturbation of the terminus-labelling profile was observed, which was completely consistent with the previously suggested existence of an impediment to replication fork movement (terC) within a BamHI 24.8 X 10(3) base fragment (Weiss & Wake, 1983). The present data suggest that terC is located within the 11.4 X 10(3) base BamHI + SalI double-digest portion of this BamHI fragment.

Elastin Biomaterials in Dermal Repair

Publisher: Elsevier BV

Date: 03-2020

DOI: 10.1016/J.TIBTECH.2019.08.005

Abstract: Wound healing has historically relied on endogenous processes, but engineered materials are increasingly being used to assist tissue repair. Elastin is an essential functional component of the dermal extracellular matrix and is an important part of skin wound repair that encompasses an elastic dermis. Advances in modern technology have better elucidated the specific signaling factors and cells that contribute to the physiological process and have led to new developments in wound care technology. We review elastin-based materials that are used to encourage wound repair. Elastin-related biomaterials, particularly those based on tropoelastin, are particularly promising because tropoelastin is assembled to make elastin. We present insights into the roles of elastin-related biomaterials and their associated in vitro and in vivo benefits on wound healing.

Hydrophobic Domains of Human Tropoelastin Interact in a Context-dependent Manner

Publisher: Elsevier BV

Date: 11-2001

DOI: 10.1074/JBC.M107920200

Highly Elastic and Conductive Human‐Based Protein Hybrid Hydrogels

Publisher: Wiley

Date: 09-11-2015

DOI: 10.1002/ADMA.201503255

Covalent attachment of functional protein to polymer surfaces: a novel one-step dry process.

Publisher: The Royal Society

Date: 19-02-2008

DOI: 10.1098/RSIF.2007.1352

Abstract: The attachment of bioactive protein to surfaces underpins the development of biosensors and diagnostic microarrays. We present a surface treatment using plasma immersion ion implantation (PIII) to create stable covalent binding sites for the attachment of functional soya-bean peroxidase (SBP). Fourier transform infrared spectra of the surfaces show that protein is retained on the surface after boiling in sodium dodecyl sulphate and sodium hydroxide, which is indicative of covalent attachment. The activity of SBP on the treated surfaces remains high in comparison with SBP attached to control surfaces over the course of 11 days. Surface plasmon resonance was used to show that the surface coverage of the attached protein is close to a monolayer. We describe the potential of the PIII treatment method to be used as a one-step dry process to create surfaces for large-scale protein micro- or nanopatterning.

Alignment of human vascular smooth muscle cells on parallel electrospun synthetic elastin fibers

Publisher: Wiley

Date: 14-10-2011

DOI: 10.1002/JBM.A.33255

Abstract: We generated parallel elastic fibers from synthetic elastin (SE) as a model of the arterial media and assessed the alignment of smooth muscle cells (SMCs). SE utilized crosslinked electrospun human tropoelastin to form aligned fibers that mimicked the topography and elastin-rich content of the medial extracellular matrix. Bundled parallel fibers were anisotropically more elastic than randomly arranged scaffolds (111 ± 25 kPa vs. 265 ± 17 kPa) in the direction of the fibers. Aligned and random fiber scaffolds each supported SMC growth. Following attachment, SMCs proliferated longitudinally on the parallel fibers and expressed native α-smooth muscle actin.

A negatively charged residue stabilizes the tropoelastin N-terminal region for elastic fiber assembly

Publisher: Elsevier BV

Date: 12-2014

DOI: 10.1074/JBC.M114.606772

Engineered Tropoelastin and Elastin-Based Biomaterials

Publisher: Elsevier

Date: 2009

DOI: 10.1016/S1876-1623(08)78001-5

Force fields for simulating the interaction of surfaces with biological molecules.

Publisher: The Royal Society

Date: 06-02-2016

DOI: 10.1098/RSFS.2015.0045

Abstract: The interaction of biomolecules with solid interfaces is of fundamental importance to several emerging biotechnologies such as medical implants, anti-fouling coatings and novel diagnostic devices. Many of these technologies rely on the binding of peptides to a solid surface, but a full understanding of the mechanism of binding, as well as the effect on the conformation of adsorbed peptides, is beyond the resolution of current experimental techniques. Nanoscale simulations using molecular mechanics offer potential insights into these processes. However, most models at this scale have been developed for aqueous peptide and protein simulation, and there are no proven models for describing biointerfaces. In this review, we detail the current research towards developing a non-polarizable molecular model for peptide–surface interactions, with a particular focus on fitting the model parameters as well as validation by choice of appropriate experimental data.

Development of a sensitive peptide-based immunoassay: Application to detection of the Jun and Fos oncoproteins

Publisher: American Chemical Society (ACS)

Date: 1996

DOI: 10.1021/BI952817O

Tropoelastin modulates TGF-β1-induced expression of VEGF and CTGF in airway smooth muscle cells

Publisher: Elsevier BV

Date: 10-2013

DOI: 10.1016/J.MATBIO.2013.04.003

Resolving nitrogen-15 and proton chemical shifts for mobile segments of elastin with two-dimensional NMR spectroscopy

Publisher: Elsevier BV

Date: 05-2012

DOI: 10.1074/JBC.M111.285163

BREWING YEAST IDENTIFICATION AND CHROMOSOME ANALYSIS USING HIGH RESOLUTION CHEF GEL ELECTROPHORESIS

Publisher: The Institute of Brewing & Distilling

Date: 06-05-1991

DOI: 10.1002/J.2050-0416.1991.TB01061.X

Design of an elastin-layered dermal regeneration template

Publisher: Elsevier BV

Date: 04-2017

DOI: 10.1016/J.ACTBIO.2016.11.054

Elastin signaling in wound repair

Publisher: Wiley

Date: 09-2012

DOI: 10.1002/BDRC.21016

Abstract: Skin is an important organ to the human body as it functions as an interface between the body and environment. Cutaneous injury elicits a complex wound healing process, which is an orchestration of cells, matrix components, and signaling factors that re-establishes the barrier function of skin. In adults, an unavoidable consequence of wound healing is scar formation. However, in early fetal development, wound healing is scarless. This phenomenon is characterized by an attenuated inflammatory response, differential expression of signaling factors, and regeneration of normal skin architecture. Elastin endows a range of mechanical and cell interactive properties to skin. In adult wound healing, elastin is severely lacking and only a disorganized elastic fiber network is present after scar formation. The inherent properties of elastin make it a desirable inclusion to adult wound healing. Elastin imparts recoil and resistance and induces a range of cell activities, including cell migration and proliferation, matrix synthesis, and protease production. The effects of elastin align with the hallmarks of fetal scarless wound healing. Elastin synthesis is substantial in late stage in utero and drops to a trickle in adults. The physical and cell signaling advantages of elastin in a wound healing context creates a parallel with the innate features of fetal skin that can allow for scarless healing.

The effect of elastin on chondrocyte adhesion and proliferation on poly (ɛ-caprolactone)/elastin composites

Publisher: Elsevier BV

Date: 02-2011

DOI: 10.1016/J.BIOMATERIALS.2010.10.024

Abstract: The aim of this study was to demonstrate the effect of elastin on chondrocyte adhesion and proliferation within the structure of poly (ɛ-caprolactone) (PCL)/elastin composites. The homogenous 3D structure composites were constructed by using high pressure CO(2) in two stages. Porous PCL structures with average pore sizes of 540 ± 21 μm and a high degree of interconnectivity were produced using gas foaming/salt leaching. The PCL scaffolds were then impregnated with elastin and cross-linked with glutaraldehyde (GA) under high pressure CO(2). The effects of elastin and cross-linker concentrations on the characteristics of composites were investigated. Increasing the elastin concentration from 25mg/ml to 100mg/ml elevated the amount of cross-linked elastin inside the macropores of PCL. Fourier transform infrared (FTIR) analysis showed that elastin was homogeneously distributed throughout the 3D structure of all composites. The weight gain of composites increased 2-fold from 15.8 ± 0.3 to 38.3 ± 0.7 (w/w) % by increasing the elastin concentration from 25mg/ml to 50mg/ml and approached a plateau above this concentration. The presence of elastin within the pores of PCL improved the water uptake properties of PCL scaffolds the water uptake ratio of PCL was enhanced 100-fold from 0.030 ± 0.005g liquid/g polymer to 11.80 ± 0.01g liquid/g polymer, when the elastin solution concentration was 50mg/ml. These composites exhibited lower compressive modulus and energy loss compared to pure PCL scaffolds due to their higher water content and elasticity. In vitro studies show that these composites can support primary articular cartilage chondrocyte adhesion and proliferation within the 3D structures. These results demonstrate the potential of using PCL/elastin composites for cartilage repair.

A step closer to elastogenesis on demand; Inducing mature elastic fibre deposition in a natural biomaterial scaffold

Publisher: Elsevier BV

Date: 2021

DOI: 10.1016/J.MSEC.2020.111788

Increasing the Pore Size of Electrospun Scaffolds

Publisher: Mary Ann Liebert Inc

Date: 10-2011

DOI: 10.1089/TEN.TEB.2011.0235

Abstract: Electrospinning has gained much attention in the past decade as an effective means of generating nano- to micro-scale polymer fibers that resemble native extracellular matrix. High porosity, pore interconnectivity, and large surface area to volume ratio of electrospun scaffolds make them highly conducive to cellular adhesion and growth. However, inherently small pores of electrospun scaffolds do not promote adequate cellular infiltration and tissue ingrowth. Cellular infiltration into the scaffold is essential for a range of tissue engineering applications and is particularly important in skin and musculoskeletal engineering. Pore size, porosity, and pore interconnectivity dictate the extent of cellular infiltration and tissue ingrowth into the scaffold influence a range of cellular processes and are crucial for diffusion of nutrients, metabolites, and waste products. A number of electrospinning techniques and postelectrospinning modifications have, therefore, been developed in order to increase the pore size of electrospun scaffolds. Diverse techniques ranging from simple variations in the electrospinning parameters to complex methodologies requiring highly specialized equipment have been explored and are described in this article.

Fuzzy binding model of molecular interactions between tropoelastin and integrin alphaVbeta3

Publisher: Elsevier BV

Date: 08-2021

DOI: 10.1016/J.BPJ.2021.04.037

Engineering a highly elastic human protein–based sealant for surgical applications

Publisher: American Association for the Advancement of Science (AAAS)

Date: 04-10-2017

DOI: 10.1126/SCITRANSLMED.AAI7466

Abstract: A highly elastic and adhesive photocrosslinkable surgical sealant using a modified human protein controls liquid leakages without the need for suturing.

Elastin architecture.

Publisher: Elsevier BV

Date: 11-2019

DOI: 10.1016/J.MATBIO.2019.07.005

Abstract: Elastic fibers are an essential component of the extracellular matrix where they provide structural integrity and elastic recoil in a number of important tissues. A major constituent of these fibers is elastin, an insoluble metabolically stable polymer formed via extensive crosslinking of the monomeric precursor tropoelastin. Research over the past few decades has shown that tropoelastin possesses unique structural features that differ from both intrinsically disordered and globular proteins. This review details the advances in our understanding of tropoelastin's structural properties and illustrates how these dictate its biological function.

Recombinant Human Elastin (rhELN)) Enhances Endothelial Progenitor Cell (EPC)-mediated Endothelialisation

Publisher: Elsevier BV

Date: 2011

DOI: 10.1016/J.HLC.2011.05.103

Transient tropoelastin nanoparticles are early-stage intermediates in the coacervation of human tropoelastin whose aggregation is facilitated by heparan sulfate and heparin decasaccharides

Publisher: Elsevier BV

Date: 03-2010

DOI: 10.1016/J.MATBIO.2009.10.003

Abstract: Tropoelastin assembly is a key step in the formation of elastin. We consider how nanoscale intracellular assemblies of tropoelastin can congregate in an extracellular environment to give microscale aggregates. We describe novel 200-300 nm spherical particles that serve as intermediates in the formation of the coacervate. Their aggregation gives 800 nm to 1 microm species. This process is facilitated by heparan sulfate and dermatan sulfate interactions which effectively lower the critical concentration to facilitate this transition. This coacervation process was examined using a panel of heparin chains of various lengths and showed greatest efficacy for the decasaccharide, followed by the octasaccharide, while the hexasaccharide displayed the shortest efficacious length. We propose that these oligosaccharide interactions enable the charge-mediated aggregation of positively charged tropoelastin. This biochemistry models glycosaminoglycan interactions on the cell surface during elastogenesis which is characterized by the clustering of nascent tropoelastin aggregates to form micron-sized spherules.

Synthesis of highly porous crosslinked elastin hydrogels and their interaction with fibroblasts in vitro

Publisher: Elsevier BV

Date: 09-2009

DOI: 10.1016/J.BIOMATERIALS.2009.05.014

Abstract: In this study the feasibility of using high pressure CO2 to produce porous alpha-elastin hydrogels was investigated. Alpha-elastin was chemically crosslinked with hexamethylene diisocyanate that can react with various functional groups in elastin such as lysine, cysteine, and histidine. High pressure CO2 substantially affected the characteristics of the fabricated hydrogels. The pore size of the hydrogels was enhanced 20-fold when the pressure was increased from 1 bar to 60 bar. The swelling ratio of the s les fabricated by high pressure CO2 was also higher than the gels produced under atmospheric pressure. The compression modulus of alpha-elastin hydrogels was increased as the applied strain magnitude was modified from 40% to 80%. The compression modulus of hydrogels produced under high pressure CO2 was 3-fold lower than the gels formed at atmospheric conditions due to the increased porosity of the gels produced by high pressure CO2. The fabrication of large pores within the 3D structures of these hydrogels substantially promoted cellular penetration and growth throughout the matrices. The highly porous alpha-elastin hydrogel structures fabricated in this study have potential for applications in tissue engineering.

Tropoelastin Switch and Modulated Endothelial Cell Binding to PTFE

Publisher: Springer Science and Business Media LLC

Date: 27-10-2011

DOI: 10.1007/S12668-011-0018-1

Injectable and Magnetic Responsive Hydrogels with Bioinspired Ordered Structures

Publisher: American Chemical Society (ACS)

Date: 06-02-2019

DOI: 10.1021/ACSBIOMATERIALS.8B01179

Towards development of a novel bio-engineered vascular bypass conduit

Publisher: Elsevier BV

Date: 02-2009

DOI: 10.1016/J.HLC.2008.11.038

Abnormal levels of serum antielastin antibodies in children with diabetes mellitus type 1

Publisher: SAGE Publications

Date: 12-2006

DOI: 10.2310/6650.2006.06009

Abstract: Antibodies to α-elastin (elastin breakdown product) and elastin sequences devoid of cross-linked regions (linear elastin) are found in the serum of all human subjects and correlate with their respective serum peptide levels. The aim of this study was to determine if the serum level of antielastin antibodies (AEAbs) differs between type 1 diabetic children and nondiabetic children. Enzyme-linked immunosorbent assay was used to measure the levels of immunoglobulin (Ig)G and IgM AEAbs in the sera of 45 diabetic children (mean age 12.8 ± 3.2 years, diabetes duration 5.3 ± 3.6 years). Twenty-two children presented with vascular complications (group 1), whereas 23 displayed no vascular complications (group 2). The controls were 18 healthy children (mean age 11.9 ± 2.3 years). Diabetic patients showed statistically significant higher levels of IgM α-AEAbs (0.82 ± 0.26 vs 0.61 ± 0.14, p = .0013) than the control group. In group 1, α-AEAbs showed statistically significant higher level than controls: IgG (0.86 ± 0.42 vs 0.59 ± 0.12 p = .0109) and IgM (0.88 ± 0.24 vs 0.61 ± 0.14 p = .0001). IgM antilinear elastin antibodies (ALEAbs) in group 1 were significantly lower than in controls (0.462 ± 0.191 vs 0.652 ± 0.127 p= .0009). IgG α-AEAbs showed correlation with microalbuminuria ( r = −.26 p = .05) and IgM ALEAbs correlated with microalbuminuria ( r = −.32 p = .035). IgG α-AEAbs correlated with neuropathy ( r = −.32 p = .035). Group 1 patients displayed a correlation between IgG ALEAbs and retinopathy ( r = −.48 p = .023) and IgM ALEAbs and microalbuminuria ( r = .52 p = .014). Levels of AEAbs and ALEAbs can serve as immunologic markers of the extent of elastin degradation. These markers may provide a tool to study elastin metabolism and a potential clinical role for AEAbs in the pathogenesis and development of vascular complications in diabetic children.

Synthetic Recombinant Human Elastin-based Vascular Grafts with Biological and Mechanical Properties Matching the Human Vasculature

Publisher: Elsevier BV

Date: 2010

DOI: 10.1016/J.HLC.2010.06.581

Human‐Recombinant‐Elastin‐Based Bioinks for 3D Bioprinting of Vascularized Soft Tissues

Publisher: Wiley

Date: 10-2020

DOI: 10.1002/ADMA.202003915

Elastin sequences trigger transient proinflammatory responses by human dermal fibroblasts

Publisher: Wiley

Date: 13-05-2013

DOI: 10.1096/FJ.13-231787

Action of tropoelastin and synthetic elastin sequences on vascular tone and on free Ca2+ level in human vascular endothelial cells.

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 23-02-1998

DOI: 10.1161/01.RES.82.3.328

Abstract: Abstract —The elastic properties of extensible tissues such as arteries and skin are mainly due to the presence of elastic fibers whose major component is the extracellular matrix protein elastin. Pathophysiological degradation of this protein leads to the generation of elastin peptides that have been identified in the circulation in the ng/mL to μg/mL range. Similar concentrations of an elastin peptide preparation (κ-elastin) were previously demonstrated to induce, among other biological actions, a dose- and endothelium-dependent vasorelaxation mediated by the elastin/laminin receptor and by endothelial NO production. To determine the elastin sequence(s) responsible for vasomotor activity and to learn more about possible signaling pathways, we have compared the action of different concentrations (10 −13 to 10 −7 mol/L) of recombinant human tropoelastin, eight synthetic elastin peptides, and a control peptide (VPVGGA) on both rat aortic ring tension and [Ca 2+ ] i of cultured human umbilical vein endothelial cells. No vasoactivity could be detected for VPVGGA and for the elastin-related sequences VGVGVA, PGVGVA, and GVGVA. Tropoelastin, VGV, PGV, and VGVAPG were found to induce an endothelium- and dose-dependent vasorelaxation and to increase endothelial [Ca 2+ ] i , whereas PVGV and VGVA produced these effects only at low concentration (10 −11 mol/L). A likely candidate for mediating the elastin peptide–related effects is the elastin/laminin receptor, since the presence of lactose strongly inhibited the vasoactivity associated with these compounds. Our results show that although the flanking amino acids modulate its activity, VGV seems to be the core sequence recognized by the elastin receptor.

Stages in tropoelastin coalescence during synthetic elastin hydrogel formation

Publisher: Elsevier BV

Date: 04-2010

DOI: 10.1016/J.MICRON.2009.11.003

Abstract: Synthetic human tropoelastin was chemically cross-linked to form elastic hydrogel-like structures in vitro. Discrete stages were identified during elastic hydrogel formation by cross-linking tropoelastin with bis(sulfosuccinimidyl) suberate at a range of protein concentrations during this process. In the early stages of this process, particles with the same dimensions as tropoelastin were seen. As hydrogel formation progressed, monomer width fibres were also observed. Overall, four distinct stages were identified: (1) tropoelastin monomers form discrete particles in the order of 200 nm diameter, (2) these particles merge to form larger spheres, (3) spheres coalesce into open linked networks, (4) coalesced spheres consolidate to form a porous structure to give synthetic elastin hydrogels.

Tropoelastin — A multifaceted naturally smart material

Publisher: Elsevier BV

Date: 04-2013

DOI: 10.1016/J.ADDR.2012.06.009

Abstract: Tropoelastin dominates the physical performance of human elastic tissue as it is assembled to make elastin. Tropoelastin is increasingly appreciated as a protein monomer with a defined solution shape comprising modular, bridged regions that specialize in elasticity and cell attachment, which collectively participate in macromolecular assembly. This modular, multifaceted molecule is being exploited to enhance the physical performance and biological presentation of engineered constructs to augment and repair human tissues. These tissues include skin and vasculature, and emphasize how growing knowledge of tropoelastin can be powerfully adapted to add value to pre-existing devices like stents and novel, multi-featured biological implants.

Elastin in asthma

Publisher: Elsevier BV

Date: 04-2012

DOI: 10.1016/J.PUPT.2012.02.001

Abstract: Extracellular matrix is generally increased in asthma, causing thickening of the airways which may either increase or decrease airway responsiveness, depending on the mechanical requirements of the deposited matrix. However, in vitro studies have shown that the altered extracellular matrix produced by asthmatic airway smooth muscle cells is able to induce increased proliferation of non-asthmatic smooth muscle cells, which is a process believed to contribute to airway hyper-responsiveness in asthma. Elastin is an extracellular matrix protein that is altered in asthmatic airways, but there has been no systematic investigation of the functional effect of these changes. This review reveals ergent reports of the state of elastin in the airway wall in asthma. In some layers of the airway it has been described as increased, decreased and/or fragmented, or unchanged. There is also considerable evidence for an imbalance of matrix metalloproteinases, which degrade elastin, and their respective inhibitors the tissue inhibitors of metalloproteinases, which collectively help to explain observations of both increased elastin and elastin fragments. A loss of lung elastic recoil in asthma suggests a mechanical role for disordered elastin in the aetiology of the disease, but extensive studies of elastin in other tissues show that elastin fragments elicit cellular effects such as increased proliferation and inflammation. This review summarises the current understanding of the role of elastin in the asthmatic airway.

Synthetic Vascular Conduits for Coronary Artery Bypass Surgery. Synopsis and State of the Art

Publisher: Elsevier BV

Date: 04-2011

DOI: 10.1016/J.HLC.2010.11.020

Asn78 and His81 form a destabilizing locus within the Max HLH-LZ homodimer

Publisher: Wiley

Date: 27-11-2001

DOI: 10.1016/S0014-5793(01)03166-0

Abstract: The human Max protein lies at the center of the Myc/Max/Mad family of transcription factors. Its role at the center of this regulatory network is dependent on the helix-loop-helix leucine zipper (HLH-LZ) dimerization domain. The Max LZ contains three residues that deviate from the pattern of hydrophobic amino acids normally present at the interface of LZ dimers: Asn(78), His(81) and Asn(92). In contrast to interfacial Asn residues in other LZ proteins, we have shown that Asn(92) does not act to destabilize the homodimer. Here we describe thermal denaturation experiments performed on Asn(78) and His(81) mutants demonstrating that these residues are involved in actively destabilizing the Max homodimer.

Tailoring the porosity and pore size of electrospun synthetic human elastin scaffolds for dermal tissue engineering

Publisher: Elsevier BV

Date: 10-2011

DOI: 10.1016/J.BIOMATERIALS.2011.05.065

Abstract: We obtained low and high porosity synthetic human elastin scaffolds by adapting low (1 mL/h) and high (3 mL/h) flow rates respectively during electrospinning. Physical, mechanical and biological properties of these scaffolds were screened to identify the best candidates for the bioengineering of dermal tissue. SHE scaffolds that were electrospun at the higher flow rate presented increased fiber diameter and greater average pore size and over doubling of overall scaffold porosity. Both types of scaffold displayed Young's moduli comparable to that of native elastin, but the high porosity scaffolds possessed higher tensile strength. Low and high porosity scaffolds supported early attachment, spreading and proliferation of primary dermal fibroblasts, but only high porosity scaffolds supported active cell migration and infiltration into the scaffold. High porosity SHE scaffolds promoted cell persistence and scaffold remodeling in vitro with only moderate scaffold contraction. The scaffolds persisted for at least 6 weeks in a mouse subcutaneous implantation study with fibroblasts on the exterior and infiltrating, evidence of scaffold remodeling including de novo collagen synthesis and early stage angiogenesis.

Plasma ion activated ePTFE vascular grafts with a covalently immobilized recombinant human tropoelastin coating reduce neointimal hyperplasia.

Publisher: American Chemical Society (ACS)

Date: 12-07-2016

DOI: 10.1021/ACSBIOMATERIALS.6B00208

Hierarchical assembly of elastin materials

Publisher: Elsevier BV

Date: 06-2019

DOI: 10.1016/J.COCHE.2019.01.004

Coarse-grained model of tropoelastin self-assembly into nascent fibrils

Publisher: Elsevier BV

Date: 06-2019

DOI: 10.1016/J.MTBIO.2019.100016

Molecular Orientation of Tropoelastin is Determined by Surface Hydrophobicity

Publisher: American Chemical Society (ACS)

Date: 06-01-2012

DOI: 10.1021/BM201404X

Abstract: Tropoelastin is the precursor of the extracellular protein elastin and is utilized in tissue engineering and implant technology by adapting the interface presented by surface-bound tropoelastin. The preferred orientation of the surface bound protein is relevant to biointerface interactions, as the C-terminus of tropoelastin is known to be a binding target for cells. Using recombinant human tropoelastin we monitored the binding of tropoelastin on hydrophilic silica and on silica made hydrophobic by depositing a self-assembled monolayer of octadecyl trichlorosilane. The layered organization of deposited tropoelastin was probed using neutron and X-ray reflectometry under aqueous and dried conditions. In a wet environment, tropoelastin retained a solution-like structure when adsorbed on silica but adopted a brush-like structure when on hydrophobized silica. The orientation of the surface-bound tropoelastin was investigated using cell binding assays and it was found that the C-terminus of tropoelastin faced the bulk solvent when bound to the hydrophobic surface, but a mixture of orientations was adopted when tropoelastin was bound to the hydrophilic surface. Drying the tropoelastin-coated surfaces irreversibly altered these protein structures for both hydrophilic and hydrophobic surfaces.

Directed cell attachment by tropoelastin on masked plasma immersion ion implantation treated PTFE

Publisher: Elsevier BV

Date: 10-2011

DOI: 10.1016/J.BIOMATERIALS.2011.05.060

Abstract: The ability to generate cell patterns on polymer surfaces is critical for the detailed study of cellular biology, the fabrication of cell-based biosensors, cell separation techniques and for tissue engineering. In this study contact tape masking and steel shadow masks were used to exclude plasma immersion ion implantation (PIII) treatment from defined areas of polytetrafluoroethylene (PTFE) surfaces. This process enabled patterned covalent binding of the cell adhesive protein, tropoelastin, without employing chemical linking molecules. Tropoelastin coating rendered the untreated regions cell adhesive and the PIII-treated area non-adhesive, allowing very fine patterning of cell adhesion to PTFE surfaces. A blocking step, such as with BSA or PEG, was not required to prevent cell binding to the underlying PIII-treated regions as tropoelastin coating alone performed this blocking function. Although tropoelastin coated the entire PTFE surface, the cell binding C-terminus of tropoelastin was markedly less solvent exposed on the PIII-treated, hydrophilic regions. The differential exposure of the C-terminus correlated with the patterned distribution of tropoelastin-mediated cell adhesion. This new methodology specifically enables directed cell behavior on a polymer surface using a simple one-step treatment process, by modulating the adhesive activity of a single extracellular matrix protein.

Silk–ionomer and silk–tropoelastin hydrogels as charged three-dimensional culture platforms for the regulation of hMSC response

Publisher: Hindawi Limited

Date: 06-04-2017

DOI: 10.1002/TERM.2152

Tropoelastin as a thermodynamically unfolded premolten globule protein: The effect of trimethylamine N-oxide on structure and coacervation

Publisher: Elsevier BV

Date: 07-2009

DOI: 10.1016/J.ABB.2009.06.001

Abstract: Tropoelastin is the monomer building block of the biopolymer elastin, which is responsible for elasticity in arteries, lung and skin. Previous studies have shown that, in contrast to predictions made based on primary sequence, tropoelastin has little regular secondary structure in aqueous solution and displays considerable flexibility. This investigation defines the level of residual structure present in tropoelastin and uses the naturally-occurring structure-inducing osmolyte trimethylamine N-oxide to examine the potential for regular structure in tropoelastin. Tropoelastin is defined as a thermodynamically unfolded premolten globule, which can account for its ability to elastically deform.

mle-1, a mariner-like transposable element in the nematode Trichostrongylus colubriformis.

Publisher: Elsevier BV

Date: 04-1997

DOI: 10.1016/S0378-1119(96)00816-5

Abstract: A mariner-like element termed mle-1 was discovered in the parasitic nematode Trichostrongylus colubriformis. The mle-1 has features which support its assignment as a mariner-like transposable element. Cloned mle-1 was derived from an intron of the tar-1 gene. It comprises 893 bp, includes two 27 bp flanking perfect inverted repeats and is present at approximately 50 copies in the genome. The element contains a coding region which displays homology to transposases, with the greatest amino acid similarity to a Caenorhabditis elegans mariner-like transposase. The coding region contains two 12 bp repeats these repeats flank an 11 bp segment which accounts for a frameshift in this region. As a candidate transposon, mle-1 provides potential for genetic manipulation of this and related organisms.

Complementary mass spectrometric techniques to achieve complete sequence coverage of recombinant human tropoelastin

Publisher: Wiley

Date: 22-09-2005

DOI: 10.1002/RCM.2164

Specificity in the coacervation of tropoelastin: solvent exposed lysines

Publisher: Elsevier BV

Date: 03-2005

DOI: 10.1016/J.JSB.2004.11.006

Abstract: Tropoelastin protein monomers associate by coacervation and are cross-linked in vivo to form elastin macro-assemblies. We provide evidence for specific protein domain contact points between tropoelastin monomers during association by coacervation. The homobifunctional cross-linker bis(sulfosuccinimidyl) suberate served as a rapid reporter of adjacent lysines and preferentially exposed domains. Intact cross-linked peptide pairs were identified after protease digestion and high-resolution electrospray mass spectrometry followed by MS/MS sequencing. Mapping of the assigned sequences indicated that the region in the monomer spanning domains 19-25 was readily accessible to solvent and enriched in cross-linking. Domains 12 and 36 were also prevalent, where these two regions were not previously thought to play a major role in the formation of mature elastin. A specificity for particular lysines allowed for the construction of a model for the first close contacts between domains and the first detailed study of the cross-linking of tropoelastin.

Haemonchus contortus: Sequence Heterogeneity of Internucleotide Binding Domains from P-Glycoproteinsand an Association with Avermectin/Milbemycin Resistance

Publisher: Elsevier BV

Date: 03-1999

DOI: 10.1006/EXPR.1998.4373

Abstract: P-Glycoproteins are transmembrane proteins associated with acquired multidrug resistance in mammalian cells and some protozoan parasites by a process of active drug export. P-glycoproteins contain two nucleotide binding domains which couple ATP to the drug transport process. The region between the nucleotide binding domains of P-glycoproteins, termed the internucleotide binding domain (IBD), was PCR- lified from adult and larval cDNA libraries using degenerate primers. The 11 clones isolated by this method fall into several distinct groups, with one group of alleles displaying between 82 and 99% identity at the nucleotide level. This sets a baseline for sequence variation of transcribed alleles from a parasitic nematode. Northern blotting showed that P-glycoprotein genes are transcribed in a developmentally regulated fashion in Haemonchus contortus. Southern blots of H. contortus drug-resistant isolates with an IBD probe revealed a pattern consistent with the involvement of P-glycoprotein in resistance to avermectin/milbemycin anthelmintics.

A model two-component system for studying the architecture of elastin assembly in vitro

Publisher: Elsevier BV

Date: 03-2005

DOI: 10.1016/J.JSB.2004.11.005

Abstract: Tropoelastin is encoded by a single human gene that spans 36 exons and is oxidized in vivo by mammalian lysyl oxidase at the epsilon amino group of available lysines to give the adipic semialdehyde, which then facilitates covalent cross-link formation in an enzyme-free process involving tropoelastin association. We demonstrate here that this process is effectively modeled by a two protein component system using purified lysyl oxidase from the yeast Pichia pastoris to facilitate the oxidation and subsequent cross-linking of recombinant human tropoelastin. The oxidized human tropoelastin forms an elastin-like polymer (EL) that is elastic, shows hydrogel behavior and contains typical elastin cross-links including lysinonorleucine, allysine aldol, and desmosine. Protease digestion and subsequent mass-spectrometry analysis of multiple ELs allowed for the identification of specific intra- and inter-molecular cross-links, leading to a model of the molecular architecture of elastin assembly in vitro. Specific intra-molecular cross-links were confined to the region of tropoelastin encoded by exons 6-15. Inter-molecular cross-links were prevalent between the regions encoded by exons 19-25. We find that assembly of tropoelastin molecules in ELs are highly enriched for a defined subset of cross-links.

Computational smart polymer design based on elastin protein mutability.

Publisher: Elsevier BV

Date: 05-2017

DOI: 10.1016/J.BIOMATERIALS.2017.01.041

The solution structure of the leucine zipper motif of the Jun oncoprotein homodimer.

Publisher: Wiley

Date: 06-1993

DOI: 10.1111/J.1432-1033.1993.TB17937.X

Abstract: Proton NMR studies have been performed on a 9.8-kDa synthetic fragment comprising the homodimeric leucine zipper domain of the human oncoprotein Jun to ascertain its conformation in aqueous solution. Analysis of two-dimensional scalar and dipolar-coupling experiments enabled almost all proton resonances to be sequence-specifically assigned and further revealed that the Jun leucine zipper forms a completely symmetric dimer in solution, consistent with the formation of a coiled-coil arrangement of parallel alpha-helical strands. The rates of exchange of in idual amide protons with solvent, as well as hydrogen-bond lengths predicted from amide proton chemical shifts, are shown to correlate with residue position in the coiled-coil. A subset of 209 unambiguous distance constraints was compiled using rules recently formulated for interpreting the NOESY spectra of symmetric coiled-coils, and these were used in combination with experimentally determined hydrogen bond and dihedral angle constraints to compute a solution structure for the Jun leucine zipper domain.

The hydrophobic domain 26 of human tropoelastin is unstructured in solution

Publisher: Elsevier BV

Date: 05-2005

DOI: 10.1016/J.JSB.2005.02.005

Abstract: Elastin is the protein responsible for the elastic properties of vertebrate tissue. Very little is currently known about the structure of elastin or of its soluble precursor tropoelastin. We have used high-resolution solution NMR methods to probe the conformational preferences of a conserved hydrophobic region in tropoelastin, domain 26 (D26). Using a combination of homonuclear, 15N-separated and triple resonance experiments, we have obtained essentially full chemical shift assignments for D26 at 278K. An analysis of secondary chemical shift changes, as well as NOE and 15N relaxation data, leads us to conclude that this domain is essentially unstructured in solution and does not interact with intact tropoelastin. D26 does not display exposed hydrophobic clusters, as expected for a fully unfolded protein and commensurate with an absence of flexible structural motifs, as identified by lack of binding of the fluorescent probe 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid. Sedimentation equilibrium data establish that this domain is strictly monomeric in solution. NMR spectra recorded at 278 and 308K indicate that no significant structural changes occur for this domain over the temperature range 278-308K, in contrast to the characteristic coacervation behavior that is observed for the full-length protein.

Tropoelastin: A versatile, bioactive assembly module

Publisher: Elsevier BV

Date: 04-2014

DOI: 10.1016/J.ACTBIO.2013.08.003

High resolution NMR solution structure of the leucine zipper domain of the c-Jun homodimer

Publisher: Elsevier BV

Date: 06-1996

DOI: 10.1074/JBC.271.23.13663

Expression of recombinant human tropoelastin in Saccharomyces cerevisiae containing a synthetic gene with a high codon adaptation index coupled to the SUC2 invertase signal sequence

Publisher: Wiley

Date: 05-2001

DOI: 10.1002/1521-3846(200105)21:2<189::AID-ABIO189>3.0.CO;2-Z

Tropoelastin enhances nitric oxide production by endothelial cells.

Publisher: Future Medicine Ltd

Date: 06-2016

DOI: 10.2217/NNM-2016-0052

Abstract: Aims: This study aimed to characterize the role of tropoelastin in eliciting a nitric oxide response in endothelial cells. Materials and methods: Nitric oxide production in cells was quantified following the addition of known nitric oxide synthase pathway inhibitors such as LNAME and 1400W. The effect of eNOS siRNA knockdowns was studied using western blotting and assessed in the presence of PI3K-inhibitor, wortmannin. Results: Tropoelastin-induced nitric oxide production was LNAME and wortmannin sensitive, while being unaffected by treatment with 1400W. Conclusion: Tropoelastin acts through a PI3K-specific pathway that leads to the phosphorylation of eNOS to enhance nitric oxide production in endothelial cells. This result points to the benefit of the use of tropoelastin in vascular applications, where NO production is a characteristic marker of vascular health.

Mechanistic insight into the elastin degradation process by the metalloprotease myroilysin from the deep-sea bacterium Myroides profundi D25.

Publisher: MDPI AG

Date: 18-03-2015

DOI: 10.3390/MD13031481

Degradation of tropoelastin by matrix metalloproteinases – cleavage site specificities and release of matrikines

Publisher: Wiley

Date: 30-03-2010

DOI: 10.1111/J.1742-4658.2010.07616.X

Abstract: To provide a basis for the development of approaches to treat elastin-degrading diseases, the aim of this study was to investigate the degradation of the natural substrate tropoelastin by the elastinolytic matrix metalloproteinases MMP-7, MMP-9, and MMP-12 and to compare the cleavage site specificities of the enzymes using complementary MS techniques and molecular modeling. Furthermore, the ability of the three proteases to release bioactive peptides was studied. Tropoelastin was readily degraded by all three MMPs. Eighty-nine cleavage sites in tropoelastin were identified for MMP-12, whereas MMP-7 and MMP-9 were found to cleave at only 58 and 63 sites, respectively. Cleavages occurred predominantly in the N-terminal and C-terminal regions of tropoelastin. With respect to the cleavage site specificities, the study revealed that all three MMPs similarly tolerate hydrophobic and/or aliphatic amino acids, including Pro, Gly, Ile, and Val, at P(1)'. MMP-7 shows a strong preference for Leu at P(1)', which is also well accepted by MMP-9 and MMP-12. Of all three MMPs, MMP-12 best tolerates bulky charged and aromatic amino acids at P(1)'. All three MMPs showed a clear preference for Pro at P(3) that could be structurally explained by molecular modeling. Analysis of the generated peptides revealed that all three MMPs show a similar ability to release bioactive sequences, with MMP-12 producing the highest number of these peptides. Furthermore, the generated peptides YTTGKLPYGYGPGG, YGARPGVGVGGIP, and PGFGAVPGA, containing GxxPG motifs that have not yet been proven to be bioactive, were identified as new matrikines upon biological activity testing.

A sterilizable, biocompatible, tropoelastin surface coating immobilized by energetic ion activation

Publisher: The Royal Society

Date: 02-2017

DOI: 10.1098/RSIF.2016.0837

Abstract: Biomimetic materials which integrate with surrounding tissues and regulate new tissue formation are attractive for tissue engineering and regenerative medicine. Plasma immersion ion-implanted (PIII) polyethersulfone (PES) provides an excellent platform for the irreversible immobilization of bioactive proteins and peptides. PIII treatment significantly improves PES wettability and results in the formation of acidic groups on the PES surface, with the highest concentration observed at 40–80 s of PIII treatment. The elastomeric protein tropoelastin can be stably adhered to PIII-treated PES in a cell-interactive conformation by tailoring the pH and salt levels of the protein–surface association conditions. Tropoelastin-coated PIII-treated PES surfaces are resistant to molecular fouling, and actively promote high levels of fibroblast adhesion and proliferation while maintaining cell morphology. Tropoelastin, unlike other extracellular matrix proteins such as fibronectin, uniquely retains full bioactivity even after medical-grade ethylene oxide sterilization. This dual approach of PIII treatment and tropoelastin cloaking allows for the stable, robust functionalization of clinically used polymer materials for directed cellular interactions.

A novel cell adhesion region in tropoelastin that mediates attachment to integrin alphaVbeta5.

Publisher: Elsevier BV

Date: 2013

DOI: 10.1074/JBC.M113.518381

Nuclear Magnetic Resonance Characterization of the Jun Leucine Zipper Domain: Unusual Properties of Coiled-Coil Interfacial Polar Residues

Publisher: American Chemical Society (ACS)

Date: 09-05-1995

DOI: 10.1021/BI00018A020

Abstract: Leucine zippers constitute a widely observed structural motif which serves to promote both homo- and heterodimerization in a number of DNA-binding proteins. As part of our ongoing efforts to characterize both the structure and the dynamical properties of this dimerization domain as they relate to biological function, we report here the secondary structure in solution of a recombinant dimeric peptide (rJunLZ) comprising residues Arg276-Asn314 of the leucine zipper domain of c-Jun. Two- and three-dimensional homo- and heteronuclear NMR experiments have allowed definition of the secondary structure of rJunLZ and have provided a total of approximately 1500 interproton distance and 62 phi dihedral angle constraints for tertiary structure calculations. Amide proton protection factors, calculated from hydrogen-deuterium exchange experiments, have identified 62 hydrogen bonds in the rJunLZ dimer. We have also examined the role of Asn22, the only polar residue situated at the hydrophobic dimer interface. Virtually all leucine zipper sequences contain such a polar residue (usually Asn) near the center of the motif. X-ray crystallographic studies showed that, in the case of the GCN4 homodimer, the polar residue (Asn) adopts an asymmetric conformation in an otherwise essentially symmetric structure. In contrast, all NMR studies of leucine zipper homodimers to date have suggested that the dimers are completely symmetric in solution. We present evidence that the side-chain amide protons of Asn22 are hydrogen-bonded in solution and that this side chain exchanges rapidly between two distinct conformations. On the basis of these observations, we propose a dynamic model which can explain the apparent differences in symmetry observed in NMR and X-ray crystallographic studies of leucine zipper homodimers. We show that mutation of Asn22 to a hydrophobic Leu residue markedly increases the thermal stability of the rJunLZ homodimer, consistent with a destabilizing role for this residue. However, at temperatures below 30 degrees C, the Asn22-->Leu mutant rearranges to form oligomers larger than the dimer, as was previously observed for the corresponding Asn-->Val mutation in the GCN4 leucine zipper. These results are consistent with the hypothesis that the polar Asn residue commonly observed at the interface of leucine zippers imposes specificity for the dimer structure at the expense of stability [Harbury, P.B., Zhang, T., Kim, P.S., & Alber, T. (1993) Science 262, 1401-1407].

Optically robust, highly permeable and elastic protein films that support dual cornea cell types

Publisher: Elsevier BV

Date: 2019

DOI: 10.1016/J.BIOMATERIALS.2018.10.006

Abstract: Damaged corneas can lead to blindness. Due to the worldwide shortage of donor corneas there is a tremendous unmet demand for a robust corneal replacement that supports growth of the major corneal cell types. Commercial artificial corneas comprise plastic polymers that do not adequately support erse cell growth. We present a new class of protein elastomer-dominated synthetic corneas with attractive performance that intimately couple biologically active tropoelastin to mechanically robust and durable protein silk. Fabricated films substantially replicate the natural cornea physically and by interacting with both key cells types used in cornea repair. Performance encompasses optical clarity at high transmittance, compatible refractive index, substantial glucose permeability, compliant mechanical properties, and support of both growth and function of corneal epithelial and endothelial cells.

Homology models for domains 21-23 of human tropoelastin shed light on lysine crosslinking.

Publisher: Elsevier BV

Date: 06-2010

DOI: 10.1016/J.BBRC.2010.05.013

Abstract: The contiguous crosslinking domain at the center of human tropoelastin encoded by exons 21-23 contains an unusual 'hinge' region thought to adopt both open and closed conformations. Key lysines 425 and 437 have been implicated in both artificial and lysyl oxidase mediated crosslinks. We have examined the importance of hinge conformation to the proximity of these lysines and their ability to undergo intramolecular and intermolecular crosslinks using homology models. The results, counter intuitively, indicate that the more open hinge conformations favor intramolecular crosslinking, while the more closed conformations favor intermolecular crosslinking. We also present evidence that the sidechains of lysines 425 and 437 are able to make direct contact enabling an intramolecular lysyl oxidase mediated crosslink.

Thermodynamic and Hydrodynamic Properties of Human Tropoelastin

Publisher: Elsevier BV

Date: 07-2001

DOI: 10.1074/JBC.M103391200

A turning point in elastin structure

Publisher: Elsevier BV

Date: 11-2006

DOI: 10.1016/J.MATBIO.2006.08.049

The S-star trial bioinformatics course: An on-line learning success

Publisher: Wiley

Date: 2003

DOI: 10.1002/BMB.2003.494031010160

Cloning and structural analysis of partial acetylcholine receptor subunit genes from the parasitic nematode Teladorsagia circumcincta

Publisher: Elsevier BV

Date: 06-2001

DOI: 10.1016/S0304-4017(01)00416-2

Abstract: Nematode nicotinic acetylcholine receptors (nAChRs) are the sites of action for the anthelmintic drug levamisole. Recent findings indicate that the molecular mechanism of levamisole resistance may involve changes in the number and/or functions of target nAChRs. Accordingly, we have used an RT-PCR approach to isolate and characterise partial cDNA clones (tca-1 and tca-2) encoding putative nAChR subunits from the economically important trichostrongyloid, Teladorsagia circumcincta. The predicted tca-1 gene product is a 248 aa fragment (TCA-1) which contains structural motifs typical of ligand-binding (alpha-) subunits, and which shows very high sequence similarities (98.8 and 97.2% amino acid identities) to the alpha-subunits encoded by tar-1 and hca-1 from Trichostrongylus colubriformis and Haemonchus contortus, respectively. Sequence analyses of partial tca-1 cDNAs from one levamisole-resistant and two susceptible populations of T. circumcincta revealed polymorphism at the predicted amino acid level, but there was no apparent association of any particular tca-1 allele with resistance. tca-2 encodes a 67 aa fragment (TCA-2) containing the TM4 transmembrane domain and carboxyl terminus of a putative nAChR structural (non-alpha) subunit. The deduced amino acid sequence of TCA-2 shows highest similarity (75% amino acid identity) to ACR-2, a structural subunit involved in forming levamisole-gated ion channels in Caenorhabditis elegans, but low similarity (43% identity) to the corresponding regions of TAR-1 and HCA-1. tca-2 is the first nAChR subunit gene of this type to be isolated from parasitic nematodes, and it provides a basis for further characterisation of structural subunits in trichostrongyloids.

Cell adhesion to tropoelastin is mediated via the C-terminal GRKRK motif and integrin alphaVbeta3.

Publisher: Elsevier BV

Date: 10-2009

DOI: 10.1074/JBC.M109.017525

Fibulin-5 interacts with fibrillin-1 molecules and microfibrils

Publisher: Portland Press Ltd.

Date: 10-05-2005

DOI: 10.1042/BJ20050368

Abstract: Fibulin-5 plays an important role in elastic fibre formation in vivo. We have investigated the molecular interactions between fibulin-5 and components of fibrillin-rich microfibrils which form a template for elastin. Fibulin-5 interacted in a dose-dependent manner with a fibrillin-1 N-terminal sequence and with tropoelastin, but not with MAGP-1 (microfibril-associated glycoprotein-1) or decorin. Fibulin-5 did not inhibit interactions between fibrillin-1 N- and C-terminal fragments, or fibrillin-1 interactions with tropoelastin. Fibulin-5 may provide a link between tropoelastin and microfibrils in the pericellular space during elastic fibre assembly.

Glycosaminoglycans mediate the coacervation of human tropoelastin through dominant charge interactions involving lysine side chains

Publisher: Elsevier BV

Date: 07-1999

DOI: 10.1074/JBC.274.31.21719

Extracellular Vesicles: Interplay with the Extracellular Matrix and Modulated Cell Responses

Publisher: MDPI AG

Date: 21-03-2022

DOI: 10.3390/IJMS23063389

Abstract: The discovery that cells secrete extracellular vesicles (EVs), which carry a variety of regulatory proteins, nucleic acids, and lipids, has shed light on the sophisticated manner by which cells can communicate and accordingly function. The bioactivity of EVs is not only defined by their internal content, but also through their surface associated molecules, and the linked downstream signaling effects they elicit in target cells. The extracellular matrix (ECM) contains signaling and structural molecules that are central to tissue maintenance and repair. Recently, a subset of EVs residing within the extracellular matrix has been identified. Although some roles have been proposed for matrix-bound vesicles, their role as signaling molecules within the ECM is yet to be explored. Given the close association of EVs and the ECM, it is not surprising that EVs partly mediate repair and regeneration by modulating matrix deposition and degradation through their cellular targets. This review addresses unique EV features that allow them to interact with and navigate through the ECM, describes how their release and content is influenced by the ECM, and emphasizes the emerging role of stem-cell derived EVs in tissue repair and regeneration through their matrix-modulating properties.

Fabricating Organized Elastin in Vascular Grafts

Publisher: Elsevier BV

Date: 05-2020

DOI: 10.1016/J.TIBTECH.2020.09.003

Yeast artificial chromosomes: Rapid extraction for high resolution analysis

Publisher: Oxford University Press (OUP)

Date: 1990

DOI: 10.1093/NAR/18.8.2193

Abstract: Evidence for an RNA gain-of-function toxicity has now been provided for an increasing number of human pathologies. Myotonic dystrophies (DM) belong to a class of RNA-dominant diseases that result from RNA repeat expansion toxicity. Specifically, DM of type 1 (DM1), is caused by an expansion of CUG repeats in the 3'UTR of the DMPK protein kinase mRNA, while DM of type 2 (DM2) is linked to an expansion of CCUG repeats in an intron of the ZNF9 transcript (ZNF9 encodes a zinc finger protein). In both pathologies the mutant RNA forms nuclear foci. The mechanisms that underlie the RNA pathogenicity seem to be rather complex and not yet completely understood. Here, we describe Drosophila models that might help unravelling the molecular mechanisms of DM1-associated CUG expansion toxicity. We generated transgenic flies that express inducible repeats of different type (CUG or CAG) and length (16, 240, 480 repeats) and then analyzed transgene localization, RNA expression and toxicity as assessed by induced lethality and eye neurodegeneration. The only line that expressed a toxic RNA has a (CTG)(240) insertion. Moreover our analysis shows that its level of expression cannot account for its toxicity. In this line, (CTG)(240.4), the expansion inserted in the first intron of CG9650, a zinc finger protein encoding gene. Interestingly, CG9650 and (CUG)(240.4) expansion RNAs were found in the same nuclear foci. In conclusion, we suggest that the insertion context is the primary determinant for expansion toxicity in Drosophila models. This finding should contribute to the still open debate on the role of the expansions per se in Drosophila and in human pathogenesis of RNA-dominant diseases.

Promoting Tropoelastin Expression in Arterial and Venous Vascular Smooth Muscle Cells and Fibroblasts for Vascular Tissue Engineering

Publisher: Mary Ann Liebert Inc

Date: 10-2016

DOI: 10.1089/TEN.TEC.2016.0173

Depth-Resolved Structural and Compositional Characterization of Ion-Implanted Polystyrene that Enables Direct Covalent Immobilization of Biomolecules

Publisher: American Chemical Society (ACS)

Date: 15-07-2015

DOI: 10.1021/ACS.JPCC.5B05164

Elastin

Publisher: Elsevier

Date: 2005

DOI: 10.1016/S0065-3233(05)70013-9

Proteomic Analysis of the Genetic Premature Aging Disease Hutchinson Gilford Progeria Syndrome Reveals Differential Protein Expression and Glycosylation

Publisher: American Chemical Society (ACS)

Date: 28-08-2003

DOI: 10.1021/PR034035K

Abstract: Proteomics has revealed differential protein expression and glycosylation in membrane proteins from premature aging Hutchinson-Gilford progeria syndrome fibroblasts (progeria). Progeria is a rare autosomal dominant genetic disorder of premature aging characterized by marked growth retardation and specific, progressive, premature senescent changes of the skin and other tissues. Affected children live to an average age of 13 years. The 1q20-24 region of chromosome 1 which codes for one of these proteins, lamin A/C, has previously been implicated by Brown et al. (1990) who described identical twins with progeria, where cytogenetic analysis showed an inverted insertion in the long arm of the chromosome in 70% of cells. Luengo et al. (2002) similarly reported an interstitial deletion of chromosome 1q23, in a 9-year-old patient with a classic clinical picture of progeria.

Lamin A expression levels are unperturbed at the normal and mutant alleles but display partial splice site selection in Hutchinson-Gilford progeria syndrome

Publisher: BMJ

Date: 09-2004

DOI: 10.1136/JMG.2004.019323

Elastin-coated ePTFE Vascular Conduit

Publisher: Elsevier BV

Date: 2008

DOI: 10.1016/J.HLC.2008.05.043

An immobilized fork as a termination of replication intermediate in Bacillus subtilis

Publisher: Elsevier BV

Date: 03-1986

DOI: 10.1016/0022-2836(86)90304-9

Abstract: The structure of a DNA intermediate associated with termination of chromosome replication in Bacillus subtilis and derived from a unique BamHI 24.8 X 10(3) base-pair (bp) region of the chromosome has been investigated. The intermediate has properties expected for a forked structure. Gel electrophoresis followed by Southern transfer and hybridization to cloned DNA has shown it to comprise single strands of 15.4 X 10(3) bp and 24.8 X 10(3) bp, in approximately equimolar amounts. After purification away from the bulk of chromosomal DNA, electron microscopy of the intermediate established that 15% of the DNA was present as branched molecules and a significant proportion (11 of 31) of these contained two arms of matching length. The average dimensions (best estimates) of this unique class of Y-shaped molecule were 9.5(+/- 0.3) X 10(3), 15.1(+/- 0.4) X 10(3) and 24.6 24.6(+/- 0.6) X 10(3) bp for the stem, arms and end-to-end length, respectively. These values are consistent with the single strand composition of the intermediate as found. Furthermore, hybridization of the single strands to DNA from known locations within the BamHI 24.8 X 10(3) bp region has established the orientation of the forked intermediate relative to the genetic map. The intermediate presumably reflects the immobilization of the clockwise replication fork within the 24.8 X 10(3) bp region, at a location approximately 15.4 X 10(3) bp from the right end.

Electrospun Elastin-based Vascular Grafts

Publisher: Elsevier BV

Date: 2008

DOI: 10.1016/J.HLC.2008.05.042

In vivo biocompatibility of a plasma-activated, coronary stent coating

Publisher: Elsevier BV

Date: 11-2012

DOI: 10.1016/J.BIOMATERIALS.2012.07.059

Abstract: Bare metal and drug-eluting coronary stents suffer an inherent lack of vascular cell and blood compatibility resulting in adverse patient responses. We have developed a plasma-activated coating (PAC) for metallic coronary stents that is durable, withstands crimping and expansion, has low thrombogenicity and can covalently bind proteins, linker-free. This has been shown to enhance endothelial cell interactions in vitro and has the potential to promote biointegration of stents. Using the rabbit denuded iliac artery model, we show for the first time that PAC is a feasible coating for coronary stents in vivo. The coating integrity of PAC was maintained following implantation and expansion. The rate of endothelialization, strut coverage, neointimal response and the initial immune response were equivalent to bare metal stents. Furthermore, the initial thrombogenicity caused by the PAC stents showed a reduced trend compared to bare metal stents. This work demonstrates a robust, durable, non-cytotoxic plasma-based coating technology that has the ability to covalently immobilize bioactive molecules for surface modification of coronary stents. Improvements in the clinical performance of implantable cardiovascular devices could be achieved by the immobilization of proteins or peptides that trigger desirable cellular responses.

Biomechanics of Synthetic Elastin: Insights from Magnetic Resonance Microimaging

Publisher: Trans Tech Publications, Ltd.

Date: 05-2013

DOI: 10.4028/WWW.SCIENTIFIC.NET/AMR.699.457

Abstract: We used Magnetic Resonance microimaging (MRI) to study the compressive behaviour of synthetic elastin. Compression-induced changes in the elastin s le were quantified using longitudinal and transverse spin relaxation rates (R1 and R2, respectively). Spatially-resolved maps of each spin relaxation rate were obtained, allowing the heterogeneous texture of the s le to be observed with and without compression. Compression resulted in an increase of both the mean R1 and the mean R2, but most of this increase was due to sub-locations that exhibited relatively low R1 and R2 in the uncompressed state. This behaviour can be described by differential compression, where local domains in the hydrogel with a relatively low biopolymer content compress more than those with a relatively high biopolymer content.

Multifunctional SilkTropoelastin Biomaterial Systems

Publisher: Wiley

Date: 10-2013

DOI: 10.1002/IJCH.201300082

Abstract: New multifunctional, degradable, polymeric biomaterial systems would provide versatile platforms to address cell and tissue needs in both in vitro and in vivo environments. While protein‐based composites or alloys are the building blocks of biological organisms, similar systems have not been largely exploited, to date, to generate ad hoc biomaterials able to control and direct biological functions by recapitulating their inherent structural and mechanical complexities. Therefore, we have recently proposed silktropoelastin material platforms that are able to conjugate a mechanically robust and durable protein, silk, to a highly flexible and biologically active protein, tropoelastin. This review focuses on the elucidation of the interactions between silk and tropoelastin to control the structure of the material its properties, and ultimately functions. In addition, an approach is provided for novel material designs to provide tools to control biological outcomes through surface roughness, elasticity, and net charge for neuronal and mesenchymal stem‐cell‐based tissue engineering.

Does Progeria Provide the Best Model of Accelerated Ageing in Humans?

Publisher: S. Karger AG

Date: 1990

DOI: 10.1159/000213181

Abstract: Studies of the causes of ageing are often obscured by the complexity of this phenomenon, hindering investigations in this area. In particular, the variable characteristics of ageing complicate analysis at a molecular level. It is proposed that to gain insight into ageing, the problem must first be simplified by restricting studies to an aspect of the ageing processes. It is suggested that progeria, which presents a less complicated etiology and phenotype than other accelerated ageing diseases, allows research to focus on a regulation site involved in development and ageing.

Emerging concepts in bone repair and the premise of soft materials

Publisher: Elsevier BV

Date: 04-2022

DOI: 10.1016/J.COPBIO.2021.12.004

Abstract: Human bone has a strong regenerative capacity that allows for restoration of its function and structure after damage. For degenerative bone diseases or large defects, bone regeneration requirements exceed the natural potential for self-healing, so bone grafts or bone substitute materials are required to support the regeneration of bone tissue. Compared to the plethora of endogenous bioactive molecules and cells in native bone grafts, the regenerative capacity of tissue-engineered materials is limited. The modest clinical impact of tissue-engineered strategies in this domain can be attributed to a failure to fully recognize key physical and biological events during bone healing, and to recapitulate the structure and composition of the target tissue to generate truly biomimetic grafts. This limitation has motivated the emergence of new strategies such as immunomodulation, endochondral ossification routes, engineered microtissues and hematoma regulation, and the development of advanced biomaterials including gene-activated matrices, soft microgels and hierarchically designed materials.

Covalent immobilisation of tropoelastin on a plasma deposited interface for enhancement of endothelialisation on metal surfaces

Publisher: Elsevier BV

Date: 03-2009

DOI: 10.1016/J.BIOMATERIALS.2008.11.009

Abstract: Currently available endovascular metallic implants such as stents exhibit suboptimal biocompatibility in that they re-endothelialise poorly leaving them susceptible to thrombosis. To improve the interaction of these implants with endothelial cells we developed a surface coating technology, enabling the covalent attachment of biomolecules to previously inert metal surfaces. Using horseradish peroxidase as a probe, we demonstrate that the polymerised surface can retain the presentation and activity of an immobilised protein. We further demonstrated the attachment of tropoelastin, an extracellular matrix protein critical to the correct arrangement and function of vasculature. Not only it is structurally important, but it plays a major role in supporting endothelial cell growth, while modulating smooth muscle cell infiltration. Tropoelastin was shown to bind to the surface in a covalent monolayer, supplemented with additional physisorbed multilayers on extended incubation. The physisorbed tropoelastin layers can be washed away in buffer or SDS while the first layer of tropoelastin remains tightly bound. The plasma coated stainless steel surface with immobilised tropoelastin was subsequently found to have improved biocompatibility by promoting endothelial cell attachment and proliferation relative to uncoated stainless steel controls. Tropoelastin coatings applied to otherwise inert substrates using this technology could thus have broad applications to a range of non-polymeric vascular devices.

Plasma ion implantation enabled bio-functionalization of PEEK improves osteoblastic activity

Publisher: AIP Publishing

Date: 06-2018

DOI: 10.1063/1.5010346

Abstract: Slow appositional growth of bone in vivo is a major problem associated with polyether ether ketone (PEEK) based orthopaedic implants. Early stage promotion of osteoblast activity, particularly bone nodule formation, would help to improve contact between PEEK implantable materials and the surrounding bone tissue. To improve interactions with bone cells, we explored here the use of plasma immersion ion implantation (PIII) treatment of PEEK to covalently immobilize biomolecules to the surface. In this study, a single step process was used to covalently immobilize tropoelastin on the surface of PIII modified PEEK through reactions with radicals generated by the treatment. Improved bioactivity was observed using the human osteoblast-like cell line, SAOS-2. Cells on surfaces that were PIII-treated or tropoelastin-coated exhibited improved attachment, spreading, proliferation, and bone nodule formation compared to cells on untreated s les. Surfaces that were both PIII-treated and tropoelastin-coated triggered the most favorable osteoblast-like responses. Surface treatment or tropoelastin coating did not alter alkaline phosphatase gene expression and activity of bound cells but did influence the expression of other bone markers including osteocalcin, osteonectin, and collagen I. We conclude that the surface modification of PEEK improves osteoblast interactions, particularly with respect to bone apposition, and enhances the orthopedic utility of PEEK.

Electrospun synthetic human elastin:collagen composite scaffolds for dermal tissue engineering

Publisher: Elsevier BV

Date: 10-2012

DOI: 10.1016/J.ACTBIO.2012.06.032

Abstract: We present an electrospun synthetic human elastin:collagen composite scaffold aimed at dermal tissue engineering. The panel of electrospun human tropoelastin and ovine type I collagen blends comprised 80% tropoelastin+20% collagen, 60% tropoelastin+40% collagen and 50% tropoelastin+50% collagen. Electrospinning efficiency decreased with increasing collagen content under the conditions used. Physical and mechanical characterization encompassed fiber morphology, porosity, pore size and modulus, which were prioritized to identify the optimal candidate for dermal tissue regeneration. Scaffolds containing 80% tropoelastin and 20% collagen (80T20C) were selected on this basis for further cell interaction and animal implantation studies. 80T20C enhanced proliferation and migration rates of dermal fibroblasts in vitro and were well tolerated in a mouse subcutaneous implantation study where they persisted over 6 weeks. The 80T20C scaffolds supported fibroblast infiltration, de novo collagen deposition and new capillary formation.

Elastin-based materials

Publisher: Royal Society of Chemistry (RSC)

Date: 2010

DOI: 10.1039/B919452P

Abstract: Elastin is a versatile elastic protein that dominates flexible tissues capable of recoil, and facilitates commensurate cell interactions in these tissues in all higher vertebrates. Elastin's persistence and insolubility h ered early efforts to construct versatile biomaterials. Subsequently the field has progressed substantially through the adapted use of solubilized elastin, elastin-based peptides and the increasing availability of recombinant forms of the natural soluble elastin precursor, tropoelastin. These interactions allow for the formation of a sophisticated range of biomaterial constructs and composites that benefit from elastin's physical properties of innate assembly and elasticity, and cell interactive properties as discussed in this tutorial review.

Subtle balance of tropoelastin molecular shape and flexibility regulates dynamics and hierarchical assembly

Publisher: American Association for the Advancement of Science (AAAS)

Date: 05-02-2016

DOI: 10.1126/SCIADV.1501145

Abstract: Tropoelastin’s local and global structures dictate molecular dynamics and are essential for efficient assembly into elastin.

Linker-free covalent attachment of the extracellular matrix protein tropoelastin to a polymer surface for directed cell spreading

Publisher: Elsevier BV

Date: 11-2009

DOI: 10.1016/J.ACTBIO.2009.05.016

Abstract: Polymers are used for the fabrication of many prosthetic implants. It is desirable for these polymers to promote biological function by promoting the adhesion, differentiation and viability of cells. Here we have used plasma immersion ion implantation (PIII) treatment of polystyrene to modify the polymer surface, and so modulate the binding of the extracellular matrix protein tropoelastin. PIII treated, but not untreated polystyrene, bound tropoelastin in a sodium dodecyl sulfate (SDS)-resistant manner, consistent with previous enzyme-binding data that demonstrated the capability of these surfaces to covalently attach proteins without employing chemical linking molecules. Furthermore sulfo-NHS acetate (SNA) blocking of tropoelastin lysine side chains eliminated the SDS-resistant binding of tropoelastin to PIII-treated polystyrene. This implies tropoelastin is covalently attached to the PIII-treated surface via its lysine side chains. Cell spreading was only observed on tropoelastin coated, PIII-treated polystyrene surfaces, indicating that tropoelastin was more biologically active on the PIII-treated surface compared to the untreated surface. A contact mask was used to pattern the PIII treatment. Following tropoelastin attachment, cells spread preferentially on the PIII-treated sections of the polystyrene surface. This demonstrates that PIII treatment of polystyrene improves the polymer's tropoelastin binding properties, with advantages for tissue engineering and prosthetic design.

Elastic proteins and elastomeric protein alloys

Publisher: Elsevier BV

Date: 06-2016

DOI: 10.1016/J.COPBIO.2015.12.020

Southern transfer and hybridization: A class experiment

Publisher: Elsevier BV

Date: 10-1992

DOI: 10.1016/0307-4412(92)90202-W

The linker-free covalent attachment of collagen to plasma immersion ion implantation treated polytetrafluoroethylene and subsequent cell-binding activity

Publisher: Elsevier BV

Date: 03-2010

DOI: 10.1016/J.BIOMATERIALS.2009.12.009

Abstract: It is desirable that polymers used for the fabrication of prosthetic implants promote biological functions such as cellular adhesion, differentiation and viability. In this study, we have used plasma immersion ion implantation (PIII) to modify the surface of polytetrafluoroethylene (PTFE), thereby modulating the binding mechanism of collagen. The amount of collagen bound to the polymer surface following PIII-treatment was similar to that bound by non-covalent physisorption. In a manner consistent with previous enzyme and tropoelastin binding data, the collagen bound to the PIII-treated PTFE surface was resistant to sodium dodecyl sulfate (SDS) elution whilst collagen bound to the untreated surface was fully removed. This demonstrates the capability of PIII-treated surfaces to covalently attach collagen without employing chemical linking molecules. Only the collagen bound to the PIII-treated PTFE surface supported human dermal fibroblast attachment and spreading. This indicates that collagen on the PIII-treated surface possesses increased adhesive activity as compared to that on the untreated surface. Cell adhesion was inhibited by EDTA when the collagen was bound to PIII-treated PTFE, as expected for integrin involvement. Additionally this adhesion was sensitive to the conformation of the bound collagen. Increased actin cytoskeletal assembly was observed on cells spreading onto collagen-coated PIII-treated PTFE compared to the collagen-coated untreated PTFE. These data demonstrate the retention of collagen's biological properties following its attachment to PIII-treated PTFE, suggesting advantages for tissue engineering and prosthetic design.

Protein Interaction Studies of MAGP-1 with Tropoelastin and Fibrillin-1

Publisher: Elsevier BV

Date: 10-2001

DOI: 10.1074/JBC.M104533200

A Scal RFLP at the E-selectin (SELE) locus in a progeria family

Publisher: Oxford University Press (OUP)

Date: 1993

DOI: 10.1093/HMG/2.6.825

Biochemistry of tropoelastin.

Publisher: Wiley

Date: 15-11-1998

DOI: 10.1046/J.1432-1327.1998.2580001.X

In vitro degradation of human tropoelastin by MMP-12 and the generation of matrikines from domain 24

Publisher: Elsevier BV

Date: 03-2009

DOI: 10.1016/J.MATBIO.2008.12.002

Abstract: Degradation of elastic fibers in tissues can result in the development of disorders that include aneurysms, atherosclerosis, and loss of skin elasticity. Tropoelastin is the precursor of the cross-linked elastin and its expression is triggered by elastin-degrading factors as a response to damage. Factors like UV radiation not only increase the expression of tropoelastin but also potent metalloelastases such as macrophage elastase (MMP-12). The development of elastin-degrading diseases, moreover, is a chronic process during which elastin and tropoelastin are repeatedly exposed to attacks by MMP-12. Hence, in this work we report the in vitro susceptibility of tropoelastin and the potential of MMP-12 to generate matrikines. This work provides evidence that tropoelastin is substantially and rapidly degraded by MMP-12 even at very dilute enzyme concentrations. MMP-12 cleaves at least 86 sites in tropoelastin. Analysis of the generated peptides revealed that some small peptides contained the motif GXXPG that may enable them to bind with the elastin binding protein (EBP). Furthermore, using synthesized peptides it was confirmed that several sites in the sequence encoded by exon 24 which contains repetitive units of biologically active VGVAPG domains are susceptible to attack by MMP-12, provided that the active subsites in MMP-12 (S(4) to S(4)') are occupied. Such cleavage events have lead to the generation of ligands that may bind to EBP.

Tropoelastin coated PLLA-PLGA scaffolds promote vascular network formation

Publisher: Elsevier BV

Date: 04-2017

DOI: 10.1016/J.BIOMATERIALS.2017.01.015

Abstract: The robust repair of large wounds and tissue defects relies on blood flow. This vascularization is the major challenge faced by tissue engineering on the path to forming thick, implantable tissue constructs. Without this vasculature, oxygen and nutrients cannot reach the cells located far from host blood vessels. To make viable constructs, tissue engineering takes advantage of the mechanical properties of synthetic materials, while combining them with ECM proteins to create a natural environment for the tissue-specific cells. Tropoelastin, the precursor of the elastin, is the ECM protein responsible for elasticity in erse tissues, including robust blood vessels. Here, we seeded endothelial cells with supporting cells on PLLA/PLGA scaffolds treated with tropoelastin, and examined the morphology, expansion and maturity of the newly formed vessels. Our results demonstrate that the treated scaffolds elicit a more expanded, complex and developed vascularization in comparison to the untreated group. Implantation of tropoelastin-treated scaffolds into mouse abdominal muscle resulted in enhanced perfusion of the penetrating vasculature and improved integration. This study points to the great potential of these combined materials in promoting the vascularization of implanted engineered constructs, which can be further exploited in the fabrication of clinically relevant engineered tissues.

Rational design of tropoelastin peptide-based inhibitors of metalloproteinases

Publisher: Elsevier BV

Date: 2003

DOI: 10.1016/S0003-9861(02)00612-4

Abstract: Abnormal production of matrix metalloproteinases (MMPs) has been observed in a variety of diseases, such as emphysema, atherosclerosis, and cancer metastasis. Destruction of connective tissue ensues and elastin is often a key target. Three of the main elastolytic MMPs are the gelatinases MMP-2 and MMP-9 and the metalloelastase MMP-12. To investigate the possibility of using peptides to inhibit the elastolytic activity of these enzymes, we mapped the sites within tropoelastin recognized by MMP-9 and MMP-12. Peptides that correspond to regions overlapping these sites were then tested for their ability to inhibit these MMPs. These included an unmodified peptide directed against MMP-9 (peptide PP), cysteine-containing peptides that mimicked either the MMP-9 (peptide NCP) or the MMP-12 (peptide lin24) cleavage sites in tropoelastin and their cyclized forms (CP and cyc24, respectively), and a peptide containing a zinc-chelating hydroxamate group directed against MMP-9 (HP). The presence of a free sulfhydryl or hydroxamate group capable of chelating the zinc ion in the active site of the MMPs was generally found to increase the inhibitory activity of the peptides. The specificity of the inhibitors varied, with some of the inhibitors showing activity against all of the MMPs examined. None of the inhibitors had any significant effect on the activity of the unrelated serine protease, plasmin. K(i) values for the inhibitors were in the micromolar range. Our results suggest ways of developing other MMP inhibitors based on substrate recognition sites that may provide greater levels of inhibition.

Elastin-based biomaterials and mesenchymal stem cells

Publisher: Royal Society of Chemistry (RSC)

Date: 2015

DOI: 10.1039/C5BM00038F

Abstract: Elastin-based biomaterials can direct mesenchymal stem cell differentiation.

Stability of a Therapeutic Layer of Immobilized Recombinant Human Tropoelastin on a Plasma-Activated Coated Surface

Publisher: Springer Science and Business Media LLC

Date: 20-11-2010

DOI: 10.1007/S11095-010-0327-Z

Abstract: To modify blood-contacting stainless surfaces by covalently coating them with a serum-protease resistant form of tropoelastin (TE). To demonstrate that the modified TE retains an exposed, cell-adhesive C-terminus that persists in the presence of blood plasma proteases. Recombinant human TE and a point mutant variant (R515A) of TE were labeled with (125)Iodine and immobilized on plasma-activated stainless steel (PAC) surfaces. Covalent attachment was confirmed using rigorous detergent washing. As kallikrein and thrombin dominate the serum degradation of tropoelastin, supraphysiological levels of these proteases were incubated with covalently bound TE and R515A, then assayed for protein levels by radioactivity detection. Persistence of the C-terminus was assessed by ELISA. TE was significantly retained covalently on PAC surfaces at 88 ± 5% and 71 ± 5% after treatment with kallikrein and thrombin, respectively. Retention of R515A was 100 ± 1.3% and 87 ± 2.3% after treatment with kallikrein and thrombin, respectively, representing significant improvements over TE. The functionally important C-terminus was cleaved in wild-type TE but retained by R515A. Protein persists in the presence of human kallikrein and thrombin when covalently immobilized on metal substrata. R515A displays enhanced protease resistance and retains the C-terminus presenting a protein interface that is viable for blood-contacting applications.

Analysis of the terminus region of the Bacillus subtilis chromosome.

Publisher: Springer Science and Business Media LLC

Date: 10-1981

DOI: 10.1038/293673A0

Abstract: Alzheimer's disease (AD) is characterized by irreversible and progressive loss of memory and cognition and profound neuronal loss. Current therapeutic strategies for the treatment of AD have been directed to a variety of targets with the aim of reversing or preventing the disease but, unfortunately, the available treatments often produce no significant clinical benefits. During the last decades compounds that inhibit or modulate γ-secretase, reducing β amyloid (Aβ) levels, have been considered as potential therapeutics for AD. Among these the (R)-enantiomer of flurbiprofen (FLU) seems to be very promising, but it shows low brain penetration. In this study, in order to improve the properties of FLU against Alzheimer's pathogenesis we synthesized some novel FLU lipophilic analogues. Lipophilicity of the new molecules has been characterized in terms of clogP, log K(C18/W) and log K(IAM/W) values. Permeability has been determined in both gastrointestinal PAMPA (PAMPA-GI) at different pH values and in brain blood barrier PAMPA (PAMPA-BBB) models. They were also tested for their ability to inhibit in vitro γ-secretase activity using rat CTXTNA2 astrocytes. Interestingly, the investigated molecules demonstrated to reduce Aβ 42 levels without affecting the amyloid precursor protein APP level in a clear concentrations-dependent manner.

A multilayered synthetic human elastin/polycaprolactone hybrid vascular graft with tailored mechanical properties

Publisher: Elsevier BV

Date: 2011

DOI: 10.1016/J.ACTBIO.2010.07.022

Abstract: Small-diameter synthetic vascular graft materials fail to match the patency of human tissue conduits used in vascular bypass surgery. The foreign surface retards endothelialization and is highly thrombogenic, while the mismatch in mechanical properties induces intimal hyperplasia. Using recombinant human tropoelastin, we have developed a synthetic vascular conduit for small-diameter applications. We show that tropoelastin enhances endothelial cell attachment (threefold vs. control) and proliferation by 54.7 ± 1.1% (3 days vs. control). Tropoelastin, when presented as a monomer and when cross-linked into synthetic elastin for biomaterials applications, had low thrombogenicity. Activation of the intrinsic pathway of coagulation, measured by plasma clotting time, was reduced for tropoelastin (60.4 ± 8.2% vs. control). Platelet attachment was also reduced compared to collagen. Reductions in platelet interactions were mirrored on cross-linked synthetic elastin scaffolds. Tropoelastin was subsequently incorporated into a synthetic elastin olycaprolactone conduit with mechanical properties optimized to mimic the human internal mammary artery, including permeability, compliance, elastic modulus and burst pressure. Further, this multilayered conduit presented a synthetic elastin internal lamina to circulating blood and demonstrated suturability and mechanical durability in a small scale rabbit carotid interposition model.

Transglutaminase-Mediated Cross-Linking of Tropoelastin to Fibrillin Stabilises the Elastin Precursor Prior to Elastic Fibre Assembly

Publisher: Elsevier BV

Date: 10-2020

DOI: 10.1016/J.JMB.2020.08.023

Rapid method for preparation and cleavage of bacterial DNA for pulsed-field gel electrophoresis

Publisher: Oxford University Press (OUP)

Date: 1989

DOI: 10.1093/NAR/17.2.814

Structure and Activity of Aspergillus nidulans Copper Amine Oxidase

Publisher: American Chemical Society (ACS)

Date: 03-06-2011

DOI: 10.1021/BI200555C

Abstract: Aspergillus nidulans amine oxidase (ANAO) has the unusual ability among the family of copper and trihydroxyphenylalanine quinone-containing amine oxidases of being able to oxidize the amine side chains of lysine residues in large peptides and proteins. We show here that in common with the related enzyme from the yeast Pichia pastoris, ANAO can promote the cross-linking of tropoelastin and oxidize the lysine residues in α-casein proteins and tropoelastin. The crystal structure of ANAO, the first for a fungal enzyme in this family, has been determined to a resolution of 2.4 Å. The enzyme is a dimer with the archetypal fold of a copper-containing amine oxidase. The active site is the most open of any of those of the structurally characterized enzymes in the family and provides a ready explanation for its lysine oxidase-like activity.

The nicotinic acetylcholine α-subunit gene tar-1 is located on the X chromosome but its coding sequence is not involved in levamisole resistance in an isolate of Trichostrongylus colubriformis

Publisher: Elsevier BV

Date: 12-1997

DOI: 10.1016/S0166-6851(97)00179-5

Abstract: The polymerase chain reaction was used to lify fragments comprising the known reading frame of the nematode nicotinic acetylcholine alpha-subunit gene tar-1. Sequences were derived from DNA prepared from bulk collections of worms and from in idual male and female Trichostrongylus colubriformis. In each case a levamisole-resistant (BCk) and a drug susceptible population were examined. Although several nucleotide transitions were detected no amino acid sequence variations were found between the isolates and between in idual worms, indicating that the coding sequence of this gene is not responsible for levamisole-resistance in the isolate tested. However, an intronic allelic T/C variation at position 4955 was observed in both populations. It has been reported that levamisole-resistance in the BCk isolate of T. colubriformis is due to a sex-linked recessive gene or gene complex. A restriction fragment length polymorphism formed by the allelic variation was found and was detectable by digestion with the restriction endonuclease NlaIII. Statistical comparison of allele frequencies from in idual male and female worms was consistent with sex-linkage of tar-1 (P < 0.05) but showed no correlation with levamisole resistance status. The polymorphism described will provide a useful X-chromosome marker and represents the first mapped genetic locus in this species.

Tropoelastin Implants That Accelerate Wound Repair

Publisher: Wiley

Date: 16-02-2018

DOI: 10.1002/ADHM.201701206

Abstract: A novel, pure, synthetic material is presented that promotes the repair of full-thickness skin wounds. The active component is tropoelastin and leverages its ability to promote new blood vessel formation and its cell recruiting properties to accelerate wound repair. Key to the technology is the use of a novel heat-based, stabilized form of human tropoelastin which allows for tunable resorption. This implantable material contributes a tailored insert that can be shaped to the wound bed, where it hydrates to form a conformable protein hydrogel. Significant benefits in the extent of wound healing, dermal repair, and regeneration of mature epithelium in healthy pigs are demonstrated. The implant is compatible with initial co-treatment with full- and split-thickness skin grafts. The implant's superiority to sterile bandaging, commercial hydrogel and dermal regeneration template products is shown. On this basis, a new concept for a prefabricated tissue repair material for point-of-care treatment of open wounds is provided.

Synthesis of functionalized-thermo responsive-water soluble co-polymer for conjugation to protein for biomedical applications

Publisher: Springer Science and Business Media LLC

Date: 2013

DOI: 10.1557/OPL.2013.86

Abstract: The aim of this study was to develop a thermo-responsive and bioactive polymer with suitable mechanical properties for musculoskeletal tissue engineering applications. A copolymer was synthesized that comprised of hydrophilic polyethylene glycol, thermo responsive N -isopropylacrylamide (NIPAAm), 2-hydroxyethyl methacrylate-poly(lactide) (HEMA-PLA) to enhance mechanical strength and an active N- acryloxysuccinimide (NAS) group for conjugation to proteins to enhance biological properties. A model protein such as elastin was used to assess the feasibility of conjugating this polymer to protein. The results of 1 HNMR analyses confirmed that random polymerization was viable technique for synthesis of this copolymer. The co-polymers synthesized with PEG content of 3 mol% were water soluble. A hydrogel was created by dissolving the copolymer and elastin below room temperature in aqueous media, followed by rapid gelation at 37°C. The results of Fourier transform infrared analyses confirmed the conjugation of protein to copolymer due to significant reduction of ester group absorption (1735 cm −1 ). This data confirmed molecular interaction between protein and the temperature responsive co-polymer. Our preliminary results demonstrated that it is viable to tune different properties of this hydrogel by changing the composition of co-polymer.

Elastin biology and tissue engineering with adult cells

Publisher: Walter de Gruyter GmbH

Date: 04-2013

DOI: 10.1515/BMC-2012-0040

Abstract: The inability of adult cells to produce well-organized, robust elastic fibers has long been a barrier to the successful engineering of certain tissues. In this review, we focus primarily on elastin with respect to tissue-engineered vascular substitutes. To understand elastin regulation during normal development, we describe the role of various elastic fiber accessory proteins. Biochemical pathways regulating expression of the elastin gene are addressed, with particular focus on tissue-engineering research using adult-derived cells.

Tropoelastin bridge region positions the cell-interactive C terminus and contributes to elastic fiber assembly

Publisher: Proceedings of the National Academy of Sciences

Date: 10-02-2012

DOI: 10.1073/PNAS.1111615108

Abstract: The tropoelastin monomer undergoes stages of association by coacervation, deposition onto microfibrils, and cross-linking to form elastic fibers. Tropoelastin consists of an elastic N-terminal coil region and a cell-interactive C-terminal foot region linked together by a highly exposed bridge region. The bridge region is conveniently positioned to modulate elastic fiber assembly through association by coacervation and its proximity to dominant cross-linking domains. Tropoelastin constructs that either modify or remove the entire bridge and downstream regions were assessed for elastogenesis. These constructs focused on a single alanine substitution (R515A) and a truncation (M155n) at the highly conserved arginine 515 site that borders the bridge. Each form displayed less efficient coacervation, impaired hydrogel formation, and decreased dermal fibroblast attachment compared to wild-type tropoelastin. The R515A mutant protein additionally showed reduced elastic fiber formation upon addition to human retinal pigmented epithelium cells and dermal fibroblasts. The small-angle X-ray scattering nanostructure of the R515A mutant protein revealed greater conformational flexibility around the bridge and C-terminal regions. This increased flexibility of the R515A mutant suggests that the tropoelastin R515 residue stabilizes the structure of the bridge region, which is critical for elastic fiber assembly.

Development of tropoelastin-functionalized anisotropic PCL scaffolds for musculoskeletal tissue engineering

Publisher: Oxford University Press (OUP)

Date: 27-10-2022

DOI: 10.1093/RB/RBAC087

Abstract: The highly organized extracellular matrix (ECM) of musculoskeletal tissues, encompassing tendons, ligaments and muscles, is structurally anisotropic, hierarchical and multi-compartmental. These features collectively contribute to their unique function. Previous studies have investigated the effect of tissue-engineered scaffold anisotropy on cell morphology and organization for musculoskeletal tissue repair and regeneration, but the hierarchical arrangement of ECM and compartmentalization are not typically replicated. Here, we present a method for multi-compartmental scaffold design that allows for physical mimicry of the spatial architecture of musculoskeletal tissue in regenerative medicine. This design is based on an ECM-inspired macromolecule scaffold. Polycaprolactone (PCL) scaffolds were fabricated with aligned fibers by electrospinning and mechanical stretching, and then surface-functionalized with the cell-supporting ECM protein molecule, tropoelastin (TE). TE was attached using two alternative methods that allowed for either physisorption or covalent attachment, where the latter was achieved by plasma ion immersion implantation (PIII). Aligned fibers stimulated cell elongation and improved cell alignment, in contrast to randomly oriented fibers. TE coatings bound by physisorption or covalently following 200 s PIII treatment promoted fibroblast proliferation. This represents the first cytocompatibility assessment of novel PIII-treated TE-coated PCL scaffolds. To demonstrate their versatility, these 2D anisotropic PCL scaffolds were assembled into 3D hierarchical constructs with an internally compartmentalized structure to mimic the structure of musculoskeletal tissue.

Ion implanted, radical-rich surfaces for the rapid covalent immobilization of active biomolecules

Publisher: AIP

Date: 2013

DOI: 10.1063/1.4802352

Fabricated tropoelastin-silk yarns and woven textiles for diverse tissue engineering applications

Publisher: Elsevier BV

Date: 06-2019

DOI: 10.1016/J.ACTBIO.2019.04.029

Abstract: Electrospun yarns offer substantial opportunities for the fabrication of elastic scaffolds for flexible tissue engineering applications. Currently available yarns are predominantly made of synthetic elastic materials. Thus scaffolds made from these yarns typically lack cell signaling cues. This can result in poor integration or even rejection on implantation, which drive demands for a new generation of yarns made from natural biologically compatible materials. Here, we present a new type of cell-attractive, highly twisted protein-based yarns made from blended tropoelastin and silk fibroin. These yarns combine physical and biological benefits by being rendered elastic and bioactive through the incorporation of tropoelastin and strengthened through the presence of silk fibroin. Remarkably, the process delivered multi-meter long yarns of tropoelastin-silk mixture that were conducive to fabrication of meshes on hand-made frames. The resulting hydrated meshes are elastic and cell interactive. Furthermore, subcutaneous implantation of the meshes in mice demonstrates their tolerance and persistence over 8 weeks. This combination of mechanical properties, biocompatibility and processability into erse shapes and patterns underscores the value of these materials and platform technology for tissue engineering applications. STATEMENT OF SIGNIFICANCE: Synthetic yarns are used to fabricate textile materials for various applications such as surgical meshes for hernia repair and pelvic organ prolapse. However, synthetic materials lack the attractive biological and physical cues characteristic of extracellular matrix and there is a demand for materials that can minimize postoperative complications. To address this need, we made yarns from a combination of recombinant human tropoelastin and silk fibroin using a modified electrospinning approach that blended these proteins into functional yarns. Prior to this study, no protein-based yarns using tropoelastin were available for the fabrication of functional textile materials. Multimeter-long, uniform and highly twisted yarns based on these proteins were elastic and cell interactive and demonstrated processing to yield textile fabrics. By using these yarns to weave fabrics, we demonstrate that an elastic human matrix protein blend can deliver a versatile platform technology to make textiles that can be explored for efficacy in tissue repair.

Hutchinson-Gilford progeria fibroblasts exhibit metabolically normal uridine uptake and RNA synthetic rates

Publisher: Elsevier BV

Date: 05-1995

DOI: 10.1006/BBRC.1995.1650

Abstract: The accelerated aging disease Hutchinson-Gilford progeria syndrome displays altered messenger RNA levels in cultured fibroblasts, yet little is known of effects on transcription by RNA polymerases other than RNA polymerase II. Total RNA metabolism was examined by incubation of Hutchinson-Gilford progeria fibroblasts with [5-3H]uridine in asynchronous culture. Uptake of radiolabel was quantitative and was incorporated preferentially (99 +/- 0.3%) into newly synthesized RNA. Progeria and control cultures showed comparable rates of uptake of radiolabel, time courses of RNA synthesis, and relative intensity profiles of newly synthesized ribosomal RNA. These characteristics held over a greater than ten-fold range of cell densities (5 x 10(3) to 8 x 10(4) cells/cm2). Progeria and control fibroblasts thus have comparable metabolic capacities for uridine uptake and net RNA production, emphasizing the relative specificity of transcriptional changes previously identified in progeria cells.

A Synthetic Internal Mammary Artery

Publisher: Elsevier BV

Date: 2011

DOI: 10.1016/J.HLC.2010.10.034

Universal Biomolecule Binding Interlayers Created by Energetic Ion Bombardment

Publisher: Springer Science and Business Media LLC

Date: 2011

DOI: 10.1557/OPL.2011.1382

Abstract: The ability to strongly attach biomolecules such as enzymes and antibodies to surfaces underpins a host of technologies that are rapidly growing in utility and importance. Such technologies include biosensors for medical and environmental applications and protein or antibody diagnostic arrays for early disease detection. Emerging new applications include continuous flow reactors for enzymatic chemical, textile or biofuels processing and implantable biomaterials that interact with their host via an interfacial layer of active biomolecules. In many of these applications it is desirable to maintain physical properties of an underlying material whilst engineering a surface suitable for attachment of proteins or peptide constructs. Nanoscale polymeric interlayers are attractive for this purpose. We have developed interlayers[1] that form the basis of a new biomolecule binding technology with significant advantages over other currently available methods. The interlayers, created by the ion implantation of polymer like surfaces, achieve covalent immobilization on immersion of the surface in protein solution. The interlayers can be created on any underlying material and ion stitched into its surface. The covalent immobilization of biomolecules from solution is achieved through the action of highly reactive free radicals in the interlayer. In this paper, we present characterisation of the structure and properties of the interlayers and describe a detailed kinetic model for the covalent attachment of protein molecules directly from solution.

Rapid Regeneration of a Neoartery with Elastic Lamellae

Publisher: Wiley

Date: 17-10-2022

DOI: 10.1002/ADMA.202205614

Abstract: Native arteries contain a distinctive intima‐media composed of organized elastin and an adventitia containing mature collagen fibrils. In contrast, implanted biodegradable small‐diameter vascular grafts do not present spatially regenerated, organized elastin. The elastin‐containing structures within the intima‐media region encompass the elastic lamellae (EL) and internal elastic lamina (IEL) and are crucial for normal arterial function. Here, the development of a novel electrospun small‐diameter vascular graft that facilitates de novo formation of a structurally appropriate elastin‐containing intima‐media region following implantation is described. The graft comprises a non‐porous microstructure characterized by tropoelastin fibers that are embedded in a PGS matrix. After implantation in mouse abdominal aorta, the graft develops distinct cell and extracellular matrix profiles that approximate the native adventitia and intima‐media by 8 weeks. Within the newly formed intima‐media region there are circumferentially aligned smooth muscle cell layers that alternate with multiple EL similar to that found in the arterial wall. By 8 months, the developed adventitia region contains mature collagen fibrils and the neoartery presents a distinct IEL with thickness comparable to that in mouse abdominal aorta. It is proposed that this new class of material can generate the critically required, organized elastin needed for arterial regeneration.

ChemInform Abstract: Elastin‐Based Materials

Publisher: Wiley

Date: 26-11-2010

DOI: 10.1002/CHIN.201051252

Cloning DNA from the replication terminus region of the Bacillus subtilis chromosome.

Publisher: Elsevier BV

Date: 09-1983

DOI: 10.1016/0378-1119(83)90133-6

Abstract: DNA from the Bacillus subtilis 168 prototroph, SB19, was partially cleaved with MboI and cloned into the BamHI site of the Escherichia coli cosmid vector, pHC79. [3H]thymine-labelled DNA from the replication terminus region of the B. subtilis dna-1 chromosome was used to identify, by hybridization, clones harboring recombinant cosmids carrying inserts from the terminus region. Restriction maps have been constructed for two such cosmids carrying overlapping DNA inserts that cover or extend into four of the previously identified five SalI fragments which are replicated last. The composite map of the cloned region, together with the available data on the replication order of fragments within it, is consistent with its replication being achieved by the unidirectional movement of a fork through it and towards the late replicating 16.2-kb SalI fragment. Most, if not all, of the terminus sequences in at least one of the recombinant cosmids are missing from a viable strain of B. subtilis that carries a deletion in the SP beta-gltA region of the chromosome.

“Setting paint” analogy for the hydrophobic self-association of tropoelastin into elastin-like hydrogel

Publisher: Wiley

Date: 05-2009

DOI: 10.1002/BIP.21133

Abstract: Alkaline tropoelastin solutions (pH 11) were optically clear at low temperatures, but a firm gel formed when the temperature was raised to 37 degrees C. Reversion to a clear solution took place if the temperature was lowered to below 20 degrees C within less than 2 h, but not if 37 degrees C was maintained for several hours. The precipitated elastin-like hydrogel thus formed did not visually redissolve at low temperatures. Tropoelastin hydrogel was stable to subsequent washings with alkaline solution at 37 degrees C, but at 4 degrees C some hydrogel redissolved showing that association is at least partly reversible. Washing the hydrogel with neutral 8M urea solution at 4 degrees C dissolved less than 10% of tropoelastin in 24 h. We characterized this phenomenon by combining temperature-controlled light microscopy analysis, 1H NMR spectroscopy (temperature, diffusion, and relaxation time studies), and UV-absorption-based concentration measurements. The self-association of tropoelastin at pH 11 is due to hydrophobic interactions in an emulsion-like system in which the spherules coalesce in a manner like a water-based latex paint that forms a durable hydrophobic sheet as water and the organic solvent evaporate. In the present case, the sedimentation and entanglement of the tropoelastin porous sheets means that reverse dissolution is a kinetically slow process.

Tropoelastin-Coated Tendon Biomimetic Scaffolds Promote Stem Cell Tenogenic Commitment and Deposition of Elastin-Rich Matrix

Publisher: American Chemical Society (ACS)

Date: 15-05-2019

DOI: 10.1021/ACSAMI.9B04616

Abstract: Tendon tissue engineering strategies that recreate the biophysical and biochemical native microenvironment have a greater potential to achieve regeneration. Here, we developed tendon biomimetic scaffolds using mechanically competent yarns of poly-ε-caprolactone, chitosan, and cellulose nanocrystals to recreate the inherent tendon hierarchy from a nano-to-macro scale. These were then coated with tropoelastin (TROPO) through polydopamine (PDA) linking, to mimic the native extracellular matrix (ECM) composition and elasticity. Both PDA and TROPO coatings decreased surface stiffness without masking the underlying substrate. We found that human adipose-derived stem cells (hASCs) seeded onto these TROPO biomimetic scaffolds more rapidly acquired their spindle-shape morphology and high aspect ratio characteristic of tenocytes. Immunocytochemistry shows that the PDA and TROPO-coated surfaces boosted differentiation of hASCs toward the tenogenic lineage, with sustained expression of the tendon-related markers scleraxis and tenomodulin up to 21 days of culture. Furthermore, these surfaces enabled the deposition of a tendon-like ECM, supported by the expression of collagens type I and III, tenascin, and decorin. Gene expression analysis revealed a downregulation of osteogenic and fibrosis markers in the presence of TROPO when compared with the control groups, suggesting proper ECM deposition. Remarkably, differentiated cells exposed to TROPO acquired an elastogenic profile due to the evident elastin synthesis and deposition, contributing to the formation of a more mimetic matrix in comparison with the PDA-coated and uncoated conditions. In summary, our biomimetic substrates combining biophysical and biological cues modulate stem cell behavior potentiating their long-term tenogenic commitment and the production of an elastin-rich ECM.

Plasma-Activated Substrate with a Tropoelastin Anchor for the Maintenance and Delivery of Multipotent Adult Progenitor Cells

Publisher: Wiley

Date: 25-09-2018

DOI: 10.1002/MABI.201800233

Abstract: Conventional wound therapy utilizes wound coverage to prevent infection, trauma, and fluid and thermal loss. However, this approach is often inadequate for large and/or chronic wounds, which require active intervention via therapeutic cells to promote healing. To address this need, a patch which delivers multipotent adult progenitor cells (MAPCs) is developed. Medical-grade polyurethane (PU) films are modified using plasma immersion ion implantation (PIII), which creates a radical-rich layer capable of rapidly and covalently attaching biomolecules. It is demonstrated that a short treatment duration of 400 s maximizes surface activation and wettability, minimizes reduction in gas permeability, and preserves the hydrolytic resistance of the PU film. The reactivity of PIII-treated PU is utilized to immobilize the extracellular matrix protein tropoelastin in a functional conformation that stably withstands medical-grade ethylene oxide sterilization. The PIII-treated tropoelastin-functionalized patch significantly promotes MAPC adhesion and proliferation over standard PU, while fully maintaining cell phenotype. Topical application of the MAPC-seeded patch transfers cells to a human skin model, while undelivered MAPCs repopulate the patch surface for subsequent cell transfer. The potential of this new wound patch as a reservoir for the sustained delivery of therapeutic MAPCs and cell-secreted factors for large and/or non-healing wounds is indicated in the findings.

The N-Terminal A Domain of Staphylococcus aureus Fibronectin-Binding Protein A Binds to Tropoelastin †

Publisher: American Chemical Society (ACS)

Date: 22-05-2007

DOI: 10.1021/BI700454X

Abstract: Staphylococcus aureus is an important human pathogen. Its virulence factors include a variety of MSCRAMMs (microbial surface component recognizing adhesive matrix molecules), each capable of binding specifically to the host extracellular matrix. The fibronectin-binding protein, FnBPA, has been shown previously to bind immobilized fibronectin, fibrinogen, and alpha-elastin peptides. Here we show that region A of FnBPA (rAFnBPA) binds to recombinant human tropoelastin. Binding occurs to three separate truncates of tropoelastin, encompassing domains 2-18, 17-27, and 27-36, signifying that the interaction occurs at multiple sites. The greatest affinity was for the N-terminal truncate. We observed a pH dependency for the rAFnBPA-tropoelastin interaction with strong, nonsaturable binding at low pH. The interaction ceased at higher pH. These data support a model of surface-surface interactions between the negative charges present on rAFnBPA and the positive lysines of tropoelastin. A protein lacking the negatively charged C-terminal fibronectin-binding motif of the A domain of FnBPA and another construct lacking subdomain N1 were both capable of binding immobilized tropoelastin with a lower affinity. The binding properties of five site-directed mutants of rAFnBPA were compared with wild-type rAFnBPA. There was no decreased affinity for immobilized tropoelastin, in contrast to the defective binding of these mutants to alpha-elastin and fibrinogen. The data indicate novel interactions between tropoelastin and FnBPA that include the use of surface charges. These results demonstrate that FnBPA is capable of directly binding tropoelastin prior to its incorporation into elastin.

Glycosaminoglycan-mediated coacervation of tropoelastin abolishes the critical concentration, accelerates coacervate formation, and facilitates spherule fusion: implications for tropoelastin microassembly.

Publisher: American Chemical Society (ACS)

Date: 12-06-2008

DOI: 10.1021/BM7013153

Abstract: Elastogenesis and elastin repair depend on the secretion of tropoelastin from the cell, yet cellular production is low in the many biological systems that have been studied. To address the apparent paradox of a paucity of tropoelastin for cell surface microassembly, we examined the effects of the glycosaminoglycans heparin, heparan sulfate, and chondroitin sulfate B, on tropoelastin aggregate formation through coacervation. We found a significant effect, particularly of heparin, on the minimum or critical concentration of tropoelastin, which was required for microassembly, lowering critical concentration to a point that it was no longer detectable. The assemblies resulted in protein droplet formation that was visually indistinguishable from the spherules that typify coacervation. The spherules readily coalesced in the presence of heparin and higher concentrations of tropoelastin, resulting in an almost continuous layer of coacervated tropoelastin. Four stages of droplet behavior were observed: early droplet formation, approximately 6 mum droplet formation, and fusion of droplets followed by the formation of a coalesced layer. We conclude that glycosaminoglycans in the extracellular matrix have the capacity to promote coacervation at low concentrations of tropoelastin.

A novel tropoelastin-based resorbable surgical mesh for pelvic organ prolapse repair

Publisher: Elsevier BV

Date: 09-2020

DOI: 10.1016/J.MTBIO.2020.100081

Cloning and sequence analysis of the candidate nicotinic acetylcholine receptor alpha subunit gene tar-1 from Trichostrongylus colubriformis

Publisher: Elsevier BV

Date: 12-1996

DOI: 10.1016/S0378-1119(96)00520-3

Abstract: A T. colubriformis genomic library in lambda EMBL3 was screened for sequences homologous to the Caenorhabditis elegans unc-38 nicotinic acetylcholine receptor (nAChR) alpha-subunit gene. The candidate gene tar-1 (for Trichostrongylus acetylcholine receptor subunit gene 1) comprising 13704 base pairs was thus identified. BLAST comparison of the sequenced clone with GenBank, followed by comparison of translated regions in six reading frames with protein databases, identified clearly defined tracts corresponding to 12 putative exons sharing high sequence homology to other nAChR genes and able to code for sequential regions of a putative nAChR alpha-subunit protein (tar-1). Tar-1 shares sequence similarities with over 40 nAChR subunit proteins. The highest similarity (91.6%) is with unc-38, suggesting that nAChR sequences from nematodes are closely related. The sequence includes motifs typical of these molecules including adjacent cysteine residues at the ACh binding site and four transmembrane regions. The DNA sequence presents the longest genomic tract described for this organism and should prove useful as a probe source in the search for nAChR genes from this and other nematodes and for studying the molecular mechanism of resistance to levamisole, a drug which is known to act on nAChRs of worms and which is widely used for parasite control.

Engineering a sprayable and elastic hydrogel adhesive with antimicrobial properties for wound healing

Publisher: Elsevier BV

Date: 09-2017

DOI: 10.1016/J.BIOMATERIALS.2017.05.011

Abstract: Hydrogel-based bioadhesives have emerged as alternatives for sutureless wound closure, since they can mimic the composition and physicochemical properties of the extracellular matrix. However, they are often associated with poor mechanical properties, low adhesion to native tissues, and lack of antimicrobial properties. Herein, a new sprayable, elastic, and biocompatible composite hydrogel, with broad-spectrum antimicrobial activity, for the treatment of chronic wounds is reported. The composite hydrogels were engineered using two ECM-derived biopolymers, gelatin methacryloyl (GelMA) and methacryloyl-substituted recombinant human tropoelastin (MeTro). MeTro/GelMA composite hydrogel adhesives were formed via visible light-induced crosslinking. Additionally, the antimicrobial peptide Tet213 was conjugated to the hydrogels, instilling antimicrobial activity against Gram (+) and (-) bacteria. The physical properties (e.g. porosity, degradability, swellability, mechanical, and adhesive properties) of the engineered hydrogel could be fine-tuned by varying the ratio of MeTro/GelMA and the final polymer concentration. The hydrogels supported in vitro mammalian cellular growth in both two-dimensional and three dimensional cultures. The subcutaneous implantation of the hydrogels in rats confirmed their biocompatibility and biodegradation in vivo. The engineered MeTro/GelMA-Tet213 hydrogels can be used for sutureless wound closure strategies to prevent infection and promote healing of chronic wounds.

Elastomers in vascular tissue engineering

Publisher: Elsevier BV

Date: 08-2016

DOI: 10.1016/J.COPBIO.2016.04.008

Charge‐Tunable Autoclaved Silk‐Tropoelastin Protein Alloys That Control Neuron Cell Responses

Publisher: Wiley

Date: 08-03-2013

DOI: 10.1002/ADFM.201202685

A unique DNA intermediate associated with termination of chromosome replication in Bacillus subtilis.

Publisher: Elsevier BV

Date: 12-1984

DOI: 10.1016/0092-8674(84)90475-6

Abstract: A unique DNA structural intermediate associated with termination of replication in Bacillus subtilis has been identified. This was achieved by probing for the Bam HI DNA fragment within which the terminus of the chromosome (terC) resides. The intermediate migrates more slowly than the bulk of digested chromosomal DNA upon electrophoresis. It is more readily detected in DNA from a merodiploid strain in which the clockwise fork should be immobilized at terC for longer than usual, and it is destroyed by S1 nuclease. The intermediate may be a forked or related structure. These findings provide evidence for the existence of a specific, sequence-based termination site that blocks or severely impedes the movement of at least the clockwise fork, which is the first to encounter it.

Flexibility in the solution structure of human tropoelastin

Publisher: American Chemical Society (ACS)

Date: 14-06-2007

DOI: 10.1021/BI700139K

Abstract: We investigated the flexibility of full-length tropoelastin in solution by using far- and near-ultraviolet circular dichroism (UV CD) and fluorescence spectroscopy to probe for structural flexibility and residue mobility within secondary and tertiary features of the monomer. Fluorescence spectroscopy revealed the presence of exposed hydrophobicity through the binding of the hydrophobic probe 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonate (bis-ANS), which demonstrates that hydrophobic regions form clusters and are not confined to a molecular core. Near-UV CD indicated substantial mobility of aromatic residues. Structural prediction programs (PONDR, DisEMBL, and Globplot version 2.0) estimated 75 +/- 2% disorder in the tertiary structure of tropoelastin on the basis of primary sequence information. A single-site substitution of Trp for Gln (Q513W) at the tropoelastin domain 25-26 interface facilitated fluorescence spectroscopy for revealing that this region is exposed to solvent. Polarization anisotropy demonstrated substantial flexibility of W513 and little change upon denaturation of the monomer with guanidine hydrochloride. Comparable movement was found for native sequence aromatic residues in the presence of glycosaminoglycans and trifluoroethanol. These data prove the intrinsic flexibility of specific residues and adjacent sequences in any native conformation(s) they may take. This study is the first characterization of the level of mobility in defined regions of the full-length tropoelastin monomer and provides direct evidence for regions of flexible structure in tropoelastin.

In situ polymerization of tropoelastin in the absence of chemical cross-linking

Publisher: Elsevier BV

Date: 02-2009

DOI: 10.1016/J.BIOMATERIALS.2008.10.018

Abstract: Tropoelastin, the polypeptide monomer precursor of elastin, is covalently cross-linked to give stable elastic structures. We show here that elastic biomaterials can be generated from tropoelastin in the absence of the classically accepted cross-linking pathway. Under alkaline conditions tropoelastin proceeds through a sol-gel transition leading to the formation of an irreversible hydrogel. This does not occur at neutral pH. The resulting biomaterial is stable, elastic and flexible. Scanning electron microscopy revealed that the hydrogel forms through the coalescence of approximately 1 microm quantized protein spheres. These spheres resemble the tropoelastin-rich globules that accumulate on cultured cell surfaces during elastin formation. In vitro cell culture studies demonstrate that the hydrogel can support human skin fibroblast proliferation. In vivo studies demonstrate that following injection, the tropoelastin solution undergoes rapid localized gelation to form a persistent mass. These subcutaneous rodent injection data establish the material's potential as a novel cell-compatible elastic scaffold that can be formed in situ.

Synthetic human elastin microfibers: Stable cross-linked tropoelastin and cell interactive constructs for tissue engineering applications

Publisher: Elsevier BV

Date: 02-2010

DOI: 10.1016/J.ACTBIO.2009.08.011

Abstract: Elastin is a key extracellular matrix protein in a range of tissues and a viable candidate for elastic tissue engineering. Elastin is not typically incorporated into engineered scaffolds because of lack of access to pure, homogeneous human elastin. Recombinant human tropoelastin, the monomer precursor of elastin, can be chemically cross-linked to form a polymer and used to generate biomaterials with physical properties similar to native elastin. In this study, we use electrospinning to generate versatile tropoelastin microfibers. Tropoelastin retained structural and biological properties, including secondary structure and coacervation temperature, after fiber formation but was solubilized on exposure to an aqueous environment prior to cross-linking. Two cross-linking methods were utilized to generate synthetic elastin microfibers that were stable in aqueous environments. Microfibers stably persisted for up to 180 days at 37 degrees C. Three primary human cell types derived from elastic tissues were assessed and found to attach and proliferate on both types of microfibers.

Substrate elasticity provides mechanical signals for the expansion of hemopoietic stem and progenitor cells

Publisher: Springer Science and Business Media LLC

Date: 10-2010

DOI: 10.1038/NBT.1687

Abstract: Surprisingly little is known about the effects of the physical microenvironment on hemopoietic stem and progenitor cells. To explore the physical effects of matrix elasticity on well-characterized primitive hemopoietic cells, we made use of a uniquely elastic biomaterial, tropoelastin. Culturing mouse or human hemopoietic cells on a tropoelastin substrate led to a two- to threefold expansion of undifferentiated cells, including progenitors and mouse stem cells. Treatment with cytokines in the presence of tropoelastin had an additive effect on this expansion. These biological effects required substrate elasticity, as neither truncated nor cross-linked tropoelastin reproduced the phenomenon, and inhibition of mechanotransduction abrogated the effects. Our data suggest that substrate elasticity and tensegrity are important mechanisms influencing hemopoietic stem and progenitor cell subsets and could be exploited to facilitate cell culture.

Structural changes and facilitated association of tropoelastin

Publisher: Elsevier BV

Date: 02-2003

DOI: 10.1016/S0003-9861(02)00719-1

Abstract: Circular dichroism studies of tropoelastin secondary structure show 4+/-1% alpha-helix in aqueous solutions. This is in contrast to the substantially higher amounts (up to 23+/-7%) of alpha-helix predicted by computer algorithms, which propose that regions of alpha-helix are limited to the alanine-rich cross-linking domains. Through the addition of trifluoroethanol, the amount of alpha-helix increased to 17+/-1%, equivalent to that expected on the basis of primary structure. The physiological ability of the protein to coacervate and the critical concentration of monomer required for coacervation were unaffected by levels of alpha-helix. However, the temperature required for coacervation decreased linearly with increasing alpha-helical structure, which correlates with the participation of alpha-helices in association. We propose that the alanine-rich cross-linking domains exist as nascent helices in tropoelastin in aqueous solution. We further suggest a novel mechanism for coacervation whereby formation of alpha-helices and subsequent helical side chain interactions limit the conformational flexibility of the polypeptide, to facilitate associations between hydrophobic domains during elastogenesis.

Cellular interactions with elastin

Publisher: Elsevier BV

Date: 09-2005

DOI: 10.1016/J.PATBIO.2004.12.022

Abstract: Elastin is a key structural component of the extracellular matrix. Tropoelastin is the soluble precursor of elastin. In addition to providing elastic recoil to various tissues such as the aorta and lung, elastin, tropoelastin and elastin degradation products are able to influence cell function and promote cellular responses. These responses include chemotaxis, proliferation and cell adhesion. The interaction of elastin products with cells has been attributed to the elastin receptor. However, additional cell-surface receptors have also been identified. These include G protein-coupled receptors and integrins. The potential roles of these receptors in cell-elastin interactions, with particular focus on elastin formation are discussed.

Freestanding hierarchical vascular structures engineered from ice

Publisher: Elsevier BV

Date: 02-2019

DOI: 10.1016/J.BIOMATERIALS.2018.11.011

Abstract: The ability to engineer a synthetic hierarchical vascular network is one of the most demanding and unaddressed challenges in tissue engineering and regenerative medicine. A material that is both structurally rigid and biocompatible is needed to fabricate freestanding hierarchical vascular structures with defined dimensions and geometry. This is particularly important for creating commercially viable and easily suturable synthetic vasculature. Here, we present the surprising discovery that ice is a versatile material which satisfies these requirements. We demonstrate utilizing ice as a sacrificial scaffold, onto which a erse range of materials were coated, including tropoelastin, polycaprolactone (PCL), silk, and polydimethylsiloxane (PDMS). We present ice facilitating the fabrication of freestanding hierarchical vascular structures with variable lumen dimensions, and validate the vascular application of these vessels by demonstrating their mechanical tunability, biocompatibility, and permeability to nutrient diffusion. This adaptable technology delivers engineered synthetic vasculature and has potential wider applications encompassing tissue engineering bespoke structures.

Immobilisation of a fibrillin-1 fragment enhances the biocompatibility of PTFE

Publisher: Elsevier BV

Date: 04-2014

DOI: 10.1016/J.COLSURFB.2014.01.042

Abstract: Current vascular biomaterials exhibit poor biocompatibility characterised by failure to promote endothelialisation, predisposition to neoinitmal hyperplasia and excessive thrombogenicity. Fibrillin-1, a major constituent of microfibrils is associated with elastic fibres in the arterial wall. Fibrillin-1 binds to endothelial cells through an RGD cell adhesion motif in the fourth TB module. The RGD motif is present in PF8, a recombinant fibrillin-1 fragment. We investigated the potential of PF8 to improve the biocompatibility of PTFE. PF8 enhanced endothelial cell attachment and cell proliferation to a greater extent than fibronectin (p<0.01). PF8 immobilised on PTFE using plasma immersion ion implantation (PIII), retained these favourable cell interactive properties, again promoting endothelial cell attachment and proliferation. The thrombogenicity of covalently bound PF8 on PTFE was assessed in both static and dynamic conditions. In static conditions, uncoated PIII treated PTFE was more thrombogenic than untreated PTFE, while PF8 coating reduced thrombogenicity. Under flow, there was no difference in the thrombogenicity of PF8 coated PTFE and untreated PTFE. Immobilised PF8 shows a striking ability to promote attachment and growth of endothelial cells on PTFE, while providing a non-thrombogenic surface. These features make PF8 a promising candidate to improve the biocompatibility of current synthetic vascular grafts.

Severe Burn Injuries and the Role of Elastin in the Design of Dermal Substitutes

Publisher: Mary Ann Liebert Inc

Date: 04-2011

DOI: 10.1089/TEN.TEB.2010.0452

Autosomal Recessive Cutis Laxa 1C Mutations Disrupt the Structure and Interactions of Latent TGFβ Binding Protein-4

Publisher: Frontiers Media SA

Date: 03-09-2021

DOI: 10.3389/FGENE.2021.706662

Abstract: Latent TGFβ binding protein-4 (LTBP4) is a multi-domain glycoprotein, essential for regulating the extracellular bioavailability of TGFβ and assembly of elastic fibre proteins, fibrillin-1 and tropoelastin. LTBP4 mutations are linked to autosomal recessive cutis laxa type 1C (ARCL1C), a rare congenital disease characterised by high mortality and severely disrupted connective tissues. Despite the importance of LTBP4, the structure and molecular consequences of disease mutations are unknown. Therefore, we analysed the structural and functional consequences of three ARCL1C causing point mutations which effect highly conserved cysteine residues. Our structural and biophysical data show that the LTBP4 N- and C-terminal regions are monomeric in solution and adopt extended conformations with the mutations resulting in subtle changes to their conformation. Similar to LTBP1, the N-terminal region is relatively inflexible, whereas the C-terminal region is flexible. Interaction studies show that one C-terminal mutation slightly decreases binding to fibrillin-1. We also found that the LTBP4 C-terminal region directly interacts with tropoelastin which is perturbed by both C-terminal ARCL1C mutations, whereas an N-terminal mutation increased binding to fibulin-4 but did not affect the interaction with heparan sulphate. Our results suggest that LTBP4 mutations contribute to ARCL1C by disrupting the structure and interactions of LTBP4 which are essential for elastogenesis in a range of mammalian connective tissues.

Protein-based composite materials

Publisher: Elsevier BV

Date: 05-2012

DOI: 10.1016/S1369-7021(12)70091-3

Fabrication Techniques for Vascular and Vascularized Tissue Engineering

Publisher: Wiley

Date: 12-08-2019

DOI: 10.1002/ADHM.201900742

Abstract: Impaired or damaged blood vessels can occur at all levels in the hierarchy of vascular systems from large vasculatures such as arteries and veins to meso- and microvasculatures such as arterioles, venules, and capillary networks. Vascular tissue engineering has become a promising approach for fabricating small-diameter vascular grafts for occlusive arteries. Vascularized tissue engineering aims to fabricate meso- and microvasculatures for the prevascularization of engineered tissues and organs. The ideal small-diameter vascular graft is biocompatible, bridgeable, and mechanically robust to maintain patency while promoting tissue remodeling. The desirable fabricated meso- and microvasculatures should rapidly integrate with the host blood vessels and allow nutrient and waste exchange throughout the construct after implantation. A number of techniques used, including engineering-based and cell-based approaches, to fabricate these synthetic vasculatures are herein explored, as well as the techniques developed to fabricate hierarchical structures that comprise multiple levels of vasculature.

Highly Elastic Micropatterned Hydrogel for Engineering Functional Cardiac Tissue

Publisher: Wiley

Date: 26-04-2013

DOI: 10.1002/ADFM.201300570

The fabrication of elastin-based hydrogels using high pressure CO2

Publisher: Elsevier BV

Date: 2009

DOI: 10.1016/J.BIOMATERIALS.2008.09.031

Abstract: The aim of this study was to investigate the effect of high pressure CO(2) on the crosslinking of elastin-based polymers and the characteristics of the fabricated hydrogels. A hydrogel was fabricated by chemically crosslinking alpha-elastin with glutaraldehyde at high pressure CO(2). The effects of pressure, reaction time, and crosslinker concentration on the characteristics of the fabricated hydrogels were determined. The reaction time had negligible effect on either the swelling ratio or the pore size of the fabricated hydrogels. Increasing the processing pressure from 30bar to 150bar resulted in a 60% increase in the hydrogel swelling ratio. The crosslinked hydrogels displayed stimuli-responsive characteristics towards temperature and salt concentration. The dense gas process facilitated coacervation, expedited the crosslinking reaction, and dramatically affected the micro- and macrostructures of pores within the s le. The results of micro-CT scan and SEM images demonstrated that pore interconnectivity was substantially enhanced for alpha-elastin hydrogels fabricated using high pressure CO(2). Dense gas CO(2) reduced the wall thickness and size of the pores and importantly induced channels within the structure of the alpha-elastin hydrogels. In vitro cell culture studies demonstrated that the channels facilitated fibroblast penetration and proliferation within alpha-elastin structures.

The Role of Elastin in Wound Healing and Dermal Substitute Design

Publisher: Springer Vienna

Date: 2013

DOI: 10.1007/978-3-7091-1586-2_5

Total synthesis and expression in Escherichia coli of a gene encoding human tropoelastin.

Publisher: Elsevier BV

Date: 1995

DOI: 10.1016/0378-1119(94)00848-M

Abstract: To elucidate the structural features and interactions of tropoelastin (TEL) molecules which assist in giving the elastic fibre its physical properties, a 2210-bp synthetic human TEL-encoding gene (SHEL) was constructed for expression in Escherichia coli. To this end, a model of codon adjustment was tested which better suits the polypeptide biosynthetic needs of E. coli than the human sequence, where over one-third of this natural sequence contains expression-limiting rare codons and 4 amino acids alone account for 75% of the resulting polypeptide. This large synthetic TEL gene was expressed at a high level as the recombinant counterpart of human TEL and as a C-terminal fusion with glutathione S-transferase. This demonstrates that a synthetic approach based upon matching codon usage to that of the host organism can support significant expression of recombinant sequences. The synthetic gene incorporates the facility for simple cassette replacement in future insertion, deletion and mutagenesis experiments, including the introduction and removal of exon homologues. The resulting soluble polypeptide is easily purified and displays properties expected for this protein.

Colocalization in vivo and association in vitro of perlecan and elastin

Publisher: Springer Science and Business Media LLC

Date: 28-08-2011

DOI: 10.1007/S00418-011-0854-7

Abstract: We have colocalized elastin and fibrillin-1 with perlecan in extracellular matrix of tensional and weight-bearing connective tissues. Elastin and fibrillin-1 were identified as prominent components of paraspinal blood vessels, and posterior longitudinal ligament in the human fetal spine and outer annulus fibrosus of the fetal intervertebral disc. We also colocalized perlecan with a synovial elastic basal lamina, where the attached synovial cells were observed to produce perlecan. Elastin, fibrillin-1 and perlecan were co-localized in the intima and media of small blood vessels in the synovium and in human fetal paraspinal blood vessels. Elastic fibers were observed at the insertion point of the anterior cruciate ligament to bone in the ovine stifle joint where they colocalized with perlecan. Elastin has not previously been reported to be spatially associated with perlecan in these tissues. Interactions between the tropoelastin and perlecan heparan sulfate chains were demonstrated using quartz crystal microbalance with dissipation solid phase binding studies. Electrostatic interactions through the heparan sulfate chains of perlecan and core protein mediated the interactions with tropoelastin, and were both important in the coacervation of tropoelastin and deposition of elastin onto perlecan immobilized on the chip surface. This may help us to understand the interactions which are expected to occur in vivo between the tropoelastin and perlecan to facilitate the deposition of elastin and formation of elastic microfibrils in situ and would be consistent with the observed distributions of these components in a number of connective tissues.

Tropoelastin Improves Post-Infarct Cardiac Function

Publisher: Ovid Technologies (Wolters Kluwer Health)

Date: 06-01-2023

DOI: 10.1161/CIRCRESAHA.122.321123

Abstract: Myocardial infarction (MI) is among the leading causes of death worldwide. Following MI, necrotic cardiomyocytes are replaced by a stiff collagen-rich scar. Compared to collagen, the extracellular matrix protein elastin has high elasticity and may have more favorable properties within the cardiac scar. We sought to improve post-MI healing by introducing tropoelastin, the soluble subunit of elastin, to alter scar mechanics early after MI. We developed an ultrasound-guided direct intramyocardial injection method to administer tropoelastin directly into the left ventricular anterior wall of rats subjected to induced MI. Experimental groups included shams and infarcted rats injected with either PBS vehicle control or tropoelastin. Compared to vehicle treated controls, echocardiography assessments showed tropoelastin significantly improved left ventricular ejection fraction (64.7±4.4% versus 46.0±3.1% control) and reduced left ventricular dyssynchrony (11.4±3.5 ms versus 31.1±5.8 ms control) 28 days post-MI. Additionally, tropoelastin reduced post-MI scar size (8.9±1.5% versus 20.9±2.7% control) and increased scar elastin (22±5.8% versus 6.2±1.5% control) as determined by histological assessments. RNA sequencing (RNAseq) analyses of rat infarcts showed that tropoelastin injection increased genes associated with elastic fiber formation 7 days post-MI and reduced genes associated with immune response 11 days post-MI. To show translational relevance, we performed immunohistochemical analyses on human ischemic heart disease cardiac s les and showed an increase in tropoelastin within fibrotic areas. Using RNA-seq we also demonstrated the tropoelastin gene ELN is upregulated in human ischemic heart disease and during human cardiac fibroblast-myofibroblast differentiation. Furthermore, we showed by immunocytochemistry that human cardiac fibroblast synthesize increased elastin in direct response to tropoelastin treatment. We demonstrate for the first time that purified human tropoelastin can significantly repair the infarcted heart in a rodent model of MI and that human cardiac fibroblast synthesize elastin. Since human cardiac fibroblasts are primarily responsible for post-MI scar synthesis, our findings suggest exciting future clinical translation options designed to therapeutically manipulate this synthesis.

Tropoelastin is a Flexible Molecule that Retains its Canonical Shape

Publisher: Wiley

Date: 08-10-2018

DOI: 10.1002/MABI.201800250

Abstract: Tropoelastin is the dominant building block of elastic fibers, which form a major component of the extracellular matrix, providing structural support to tissues and imbuing them with elasticity and resilience. Recently, the atomistic structure of human tropoelastin is described, obtained through accelerated s ling via replica exchange molecular dynamics simulations. Here, principal component analysis is used to consider the ensemble of structures accessible to tropoelastin at body temperature (37 °C) at which tropoelastin naturally self-assembles into aggregated coacervates. These coacervates are relevant because they are an essential intermediate assembly stage, where tropoelastin molecules are then cross-linked at lysine residues and integrated into growing elastic fibers. It is found that the ensemble preserves the canonical tropoelastin structure with an extended molecular body flanked by two protruding legs, and identifies variations in specific domain positioning within this global shape. Furthermore, it is found that lysine residues show a large variation in their location on the tropoelastin molecule compared with other residues. It is hypothesized that this perturbation of the lysines increases their accessibility and enhances cross-linking. Finally, the principal component modes are extracted to describe the range of tropoelastin's conformational fluctuation to validate tropoelastin's scissor-twist motion that was predicted earlier.

The elastin matrix in tissue engineering and regeneration

Publisher: Elsevier BV

Date: 06-2018

DOI: 10.1016/J.COBME.2018.02.007

Domain 26 of tropoelastin plays a dominant role in association by coacervation.

Publisher: Elsevier BV

Date: 09-2000

DOI: 10.1074/JBC.M004265200

Changes in elastin structure and extensibility induced by hypercalcemia and hyperglycemia

Publisher: Elsevier BV

Date: 06-2023

DOI: 10.1016/J.ACTBIO.2022.03.041

Abstract: Elastin is a key elastomeric protein responsible for the elasticity of many organs, including heart, skin, and blood vessels. Due to its intrinsic long life and low turnover rate, damage in elastin induced by pathophysiological conditions, such as hypercalcemia and hyperglycemia, accumulates during biological aging and in aging-associated diseases, such as diabetes mellitus and atherosclerosis. Prior studies have shown that calcification induced by hypercalcemia deteriorates the function of aortic tissues. Glycation of elastin is triggered by hyperglycemia and associated with elastic tissue damage and loss of mechanical functions via the accumulation of advanced glycation end products. To evaluate the effects on elastin's structural conformations and elasticity by hypercalcemia and hyperglycemia at the molecular scale, we perform classical atomistic and steered molecular dynamics simulations on tropoelastin, the soluble precursor of elastin, under different conditions. We characterize the interaction sites of glucose and calcium and associated structural conformational changes. Additionally, we find that elevated levels of calcium ions and glucose hinder the extensibility of tropoelastin by rearranging structural domains and altering hydrogen bonding patterns, respectively. Overall, our investigation helps to reveal the behavior of tropoelastin and the biomechanics of elastin biomaterials in these physiological environments. STATEMENT OF SIGNIFICANCE: Elastin is a key component of elastic fibers which endow many important tissues and organs, from arteries and veins, to skin and heart, with strength and elasticity. During aging and aging-associated diseases, such as diabetes mellitus and atherosclerosis, physicochemical stressors, including hypercalcemia and hyperglycemia, induce accumulated irreversible damage in elastin, and consequently alter mechanical function. Yet, molecular mechanisms associated with these processes are still poorly understood. Here, we present the first study on how these changes in elastin structure and extensibility are induced by hypercalcemia and hyperglycemia at the molecular scale, revealing the essential roles that calcium and glucose play in triggering structural alterations and mechanical stiffness. Our findings yield critical insights into the first steps of hypercalcemia- and hyperglycemia-mediated aging.

Mapping of macrophage elastase cleavage sites in insoluble human skin elastin

Publisher: Elsevier BV

Date: 06-2008

DOI: 10.1016/J.MATBIO.2008.02.001

Abstract: Macrophage elastase (MMP-12) is a member of the family of matrix metalloproteinases (MMPs) and is active against multiple extracellular protein substrates such as elastin. Its effect on elastin is central to emphysema in the lung and photoaging of skin. Its expression in the skin increases on photodamaged skin and upon aging. Detecting and characterizing peptides cleaved in elastin, therefore, helps to understand such degradative disease processes in the skin and is also needed to assist in the rational design of agents that specifically inhibit the degradation. In this study, cleavage sites of MMP-12 in human skin elastin were extensively investigated. The peptides formed as a result of cleavages by this enzyme in the human skin elastin were characterized using mass spectrometry. A total of 41 peptides ranging from 4 to 41 amino acids were identified and 36 cleavage sites were determined. Amino acids encoded by exons 5, 6, 26, 28-31 were particularly susceptible to cleavages by MMP-12 and none or very few cleavages were detected from domains encoded by the remaining exons. The amino acid preferences of the different subsites on the catalytic domain of MMP-12 were analyzed.

A cell adhesive peptide from tropoelastin promotes sequential cell attachment and spreading via distinct receptors

Publisher: Wiley

Date: 15-06-2017

DOI: 10.1111/FEBS.14114

Abstract: Tropoelastin is the dominant monomer that assembles to form elastin, which confers elasticity to vertebrate elastic tissues including skin, arteries, and lungs. Tropoelastin interacts with cells through cell surface receptors including integrins and glycosaminoglycans (GAGs). As the region 17-18 is recognized as a key region in cell attachment and spreading, we utilized C-terminal truncated tropoelastin constructs containing dissected sections of domain 18. We mapped a cell-interactive sequence of tropoelastin to domain 17 and the first six amino acids (aa) of domain 18. Further delineation identified a 21-residue sequence (Peptide 302-322) which promoted cell attachment and spreading indistinguishable from that to N18, a construct encompassing the full domains 17-18. Alanine substitution of the lysines at positions 11 and 14 in Peptide 302-322 effectively abolished cell binding. This reliance on lysines pointed to a role for GAGs, which was assessed by heparan sulfate inhibition, leading to 85.9 ± 4.2% decreased cell binding, while inhibition of integrins using ethylenediaminetetraacetic acid did not affect attachment. In contrast, selective antibody blocking of the integrin α

Microfibril-associated glycoprotein-1 binding to tropoelastin

Publisher: Wiley

Date: 29-06-2004

DOI: 10.1111/J.1432-1033.2004.04246.X

Fabrication of porous PCL/elastin composite scaffolds for tissue engineering applications

Publisher: Elsevier BV

Date: 11-2011

DOI: 10.1016/J.SUPFLU.2011.06.010

Biomaterials derived from silk–tropoelastin protein systems

Publisher: Elsevier BV

Date: 11-2010

DOI: 10.1016/J.BIOMATERIALS.2010.07.044

Molecular-level characterization of elastin-like constructs and human aortic elastin

Publisher: Elsevier BV

Date: 09-2014

DOI: 10.1016/J.MATBIO.2014.07.006

Abstract: This study aimed to characterize the structures of two elastin-like constructs, one composed of a cross-linked elastin-like polypeptide and the other one of cross-linked tropoelastin, and native aortic elastin. The structures of the insoluble materials and human aortic elastin were investigated using scanning electron microscopy. Additionally, all s les were digested with enzymes of different specificities, and the resultant peptide mixtures were characterized by ESI mass spectrometry and MALDI mass spectrometry. The MS(2) data was used to sequence linear peptides, and cross-linked species were analyzed with the recently developed software PolyLinX. This enabled the identification of two intramolecularly cross-linked peptides containing allysine aldols in the two constructs. The presence of the tetrafunctional cross-link desmosine was shown for all analyzed materials and its quantification revealed that the cross-linking degree of the two in vitro cross-linked materials was significantly lower than that of native elastin. Molecular dynamics simulations were performed, based on molecular species identified in the s les, to follow the formation of elastin cross-links. The results provide evidence for the significance of the GVGTP hinge region of domain 23 for the formation of elastin cross-links. Overall, this work provides important insight into structural similarities and differences between elastin-like constructs and native elastin. Furthermore, it represents a step toward the elucidation of the complex cross-linking pattern of mature elastin.

Elastomeric recombinant protein-based biomaterials

Publisher: Elsevier BV

Date: 08-2013

DOI: 10.1016/J.BEJ.2013.05.006

A novel elastin gene mutation (1281delC) in a family with supravalvular aortic stenosis: A mutation cluster within exon 20

Publisher: Hindawi Limited

Date: 2001

DOI: 10.1002/1098-1004(2001)17:1<81::AID-HUMU30>3.0.CO;2-X

Novel Recombinant Tropoelastin Implants Restore Skin Extracellular Matrix

Publisher: SanovaWorks

Date: 11-2020

DOI: 10.36849/JDD.2020.5375

Role for Cela1 in Postnatal Lung Remodeling and Alpha-1 Antitrypsin–Deficient Emphysema

Publisher: American Thoracic Society

Date: 08-2018

DOI: 10.1165/RCMB.2017-0361OC

Binding of the cell adhesive protein tropoelastin to PTFE through plasma immersion ion implantation treatment

Publisher: Elsevier BV

Date: 08-2011

DOI: 10.1016/J.BIOMATERIALS.2011.03.079

Abstract: The interaction of proteins and cells with polymers is critical to their use in scientific and medical applications. In this study, plasma immersion ion implantation (PIII) was used to modify the surface of polytetrafluorethylene (PTFE), enabling the covalent binding of a cell adhesive protein, tropoelastin, without employing chemical linking molecules. Tropoelastin coating of untreated or PIII treated PFTE simultaneously promoted and blocked cell interactions respectively, i.e. PIII treatment of the PTFE surface completely inverses the cell interactive properties of bound tropoelastin. This activity persisted over long term storage of the PIII treated surfaces. The integrin binding C-terminus of tropoelastin was markedly less solvent exposed when bound to PIII treated PTFE than untreated PTFE, accounting for the modulation of cell adhesive activity. This presents a new methodology to specifically modulate cell behavior on a polymer surface using a simple one step treatment process, by adjusting the adhesive activity of a single extracellular matrix protein.

Characterization of Endothelial Progenitor Cell Interactions with Human Tropoelastin

Publisher: Public Library of Science (PLoS)

Date: 26-06-2015

DOI: 10.1371/JOURNAL.PONE.0131101

Fabricated Elastin

Publisher: Wiley

Date: 13-03-2015

DOI: 10.1002/ADHM.201400781

Deficient coacervation of two forms of human tropoelastin associated with supravalvular aortic stenosis

Publisher: Wiley

Date: 15-11-1999

DOI: 10.1046/J.1432-1327.1999.00891.X

Abstract: Human tropoelastin associates by coacervation and is subsequently cross-linked to make elastin. In Williams syndrome, defective elastin deposition is associated with hemizygous deletion of the tropoelastin gene in supravalvular aortic stenosis (SVAS). Remarkably, point-mutation forms of SVAS correspond to incomplete forms of tropoelastin which include in-frame termination by nonsense mutations, yet the resulting phenotype of these disorders is not explained because expression variably occurs from both normal and mutant alleles. Proteins corresponding to two truncated tropoelastin mutants were expressed and purified to homogeneity. Coacervation of these proteins occurred as expected with increasing temperature, but substantially contrasted with that of the performance of a normal tropoelastin. Significantly, association by coacervation of the truncated SVAS tropoelastin molecules was negligible at 37 degrees C, which contrasted with the substantial coacervation seen for normal tropoelastin. Furthermore their midpoints of coacervation increased and correlated with the extent of deletion, in accord with the loss of hydrophobic regions required for tropoelastin association. Their secondary structures are similar, as evidenced by CD studies. We propose a model for point-mutation SVAS in which aberrant tropoelastin molecules are incompetent and are mainly excluded from participation in coacervation and consequently in elastogenesis. These forms of SVAS may consequently be considered functionally similar to a hemizygous deletion, and mark point-mutation SVAS as a disorder of defective coacervation.

In vitro studies of cells grown on the superconductor PrOxFeAs

Publisher: Elsevier BV

Date: 06-2009

DOI: 10.1016/J.MICRON.2008.12.005

Abstract: The recent discovery of arsenic-based high temperature superconductors has reignited interest in the study of superconductor: biological interfaces. However, the new superconductor materials involve the chemistry of arsenic and their toxicity remains unclear [Hand, E., 2008. Nature 452 (24), 922]. In this study the possible adverse effects of this new family of superconductors on cells have been examined. Cell culture studies in conjunction with microscopy and viability assays were employed to examine the influence of arsenic-based superconductor PrO(x)FeAs (x=0.75) material in vitro. Imaging data revealed that cells were well adhered and spread on the surface of the superconductor. Furthermore, cytotoxicity studies showed that cells were unaffected during the time-course of the experiments, providing support for the biocompatibility aspects of PrO(x)FeAs-based superconductor material.

Tuneable cellulose nanocrystal and tropoelastin-laden hyaluronic acid hydrogels

Publisher: SAGE Publications

Date: 08-07-2019

DOI: 10.1177/0885328219859830

Plasma-based biofunctionalization of vascular implants

Publisher: Future Medicine Ltd

Date: 12-2012

DOI: 10.2217/NNM.12.161

Abstract: Polymeric and metallic materials are used extensively in permanently implanted cardiovascular devices and devices that make temporary but often prolonged contact with body fluids and tissues. Foreign body responses are typically triggered by host interactions at the implant surface, making surface modifications to increase biointegration desirable. Plasma-based treatments are extensively used to modify erse substrates modulating surface chemistry, wettability and surface roughness, as well as facilitating covalent biomolecule binding. Each aspect impacts on facets of vascular compatibility including endothelialization and blood contact. These modifications can be readily applied to polymers such as Dacron ® and expanded polytetrafluoroethylene, which are widely used in bypass grafting and the metallic substrates of stents, valves and pacemaker components. Plasma modification of metals is more challenging given the need for coating deposition in addition to surface activation, adding the necessity for robust interface adhesion. This review examines the evolving plasma treatment technology facilitating the biofunctionalization of polymeric and metallic implantable cardiovascular materials.

Tailoring the biofunctionality of collagen biomaterials via tropoelastin incorporation and EDC-crosslinking

Publisher: Elsevier BV

Date: 11-2021

DOI: 10.1016/J.ACTBIO.2021.08.027

Abstract: Recreating the cell niche of virtually all tissues requires composite materials fabricated from multiple extracellular matrix (ECM) macromolecules. Due to their wide tissue distribution, physical attributes and purity, collagen, and more recently, tropoelastin, represent two appealing ECM components for biomaterials development. Here we blend tropoelastin and collagen, harnessing the cell-modulatory properties of each biomolecule. Tropoelastin was stably co-blended into collagen biomaterials and was retained after EDC-crosslinking. We found that human dermal fibroblasts (HDF), rat glial cells (Rugli) and HT1080 fibrosarcoma cells ligate to tropoelastin via EDTA-sensitive and EDTA-insensitive receptors or do not ligate with tropoelastin, respectively. These differing elastin-binding properties allowed us to probe the cellular response to the tropoelastin-collagen composites assigning specific bioactivity to the collagen and tropoelastin component of the composite material. Tropoelastin addition to collagen increased total Rugli cell adhesion, spreading and proliferation. This persisted with EDC-crosslinking of the tropoelastin-collagen composite. Tropoelastin addition did not affect total HDF and HT1080 cell adhesion however, it increased the contribution of cation-independent adhesion, without affecting the cell morphology or, for HT1080 cells, proliferation. Instead, EDC-crosslinking dictated the HDF and HT1080 cellular response. These data show that a tropoelastin component dominates the response of cells that possess non-integrin based tropoelastin receptors. EDC modification of the collagen component directs cell function when non-integrin tropoelastin receptors are not crucial for cell activity. Using this approach, we have assigned the biological contribution of each component of tropoelastin-collagen composites, allowing informed biomaterial design for directed cell function via more physiologically relevant mechanisms. STATEMENT OF SIGNIFICANCE: Biomaterials fabricated from multiple extracellular matrix (ECM) macromolecules are required to fully recreate the native tissue niche where each ECM macromolecule engages with a specific repertoire of cell-surface receptors. Here we investigate combining tropoelastin with collagen as they interact with cells via different receptors. We identified specific cell lines, which associate with tropoelastin via distinct classes of cell-surface receptor. These showed that tropoelastin, when combined with collagen, altered the cell behaviour in a receptor-usage dependent manner. Integrin-mediated tropoelastin interactions influenced cell proliferation and non-integrin receptors influenced cell spreading and proliferation. These data shed light on the interplay between biomaterial macromolecular composition, cell surface receptors and cell behaviour, advancing bespoke materials design and providing functionality to specific cell populations.

Clinical Relevance of Elastin in the Structure and Function of Skin

Publisher: Oxford University Press (OUP)

Date: 14-05-2021

DOI: 10.1093/ASJOF/OJAB019

Abstract: Elastin is the main component of elastic fibers, which provide stretch, recoil, and elasticity to the skin. Normal levels of elastic fiber production, organization, and integration with other cutaneous extracellular matrix proteins, proteoglycans, and glycosaminoglycans are integral to maintaining healthy skin structure, function, and youthful appearance. Although elastin has very low turnover, its production decreases after in iduals reach maturity and it is susceptible to damage from many factors. With advancing age and exposure to environmental insults, elastic fibers degrade. This degradation contributes to the loss of the skin’s structural integrity combined with subcutaneous fat loss, this results in looser, sagging skin, causing undesirable changes in appearance. The most dramatic changes occur in chronically sun-exposed skin, which displays sharply altered amounts and arrangements of cutaneous elastic fibers, decreased fine elastic fibers in the superficial dermis connecting to the epidermis, and replacement of the normal collagen-rich superficial dermis with abnormal clumps of solar elastosis material. Disruption of elastic fiber networks also leads to undesirable characteristics in wound healing, and the worsening structure and appearance of scars and stretch marks. Identifying ways to replenish elastin and elastic fibers should improve the skin’s appearance, texture, resiliency, and wound-healing capabilities. However, few therapies are capable of repairing elastic fibers or substantially reorganizing the elastin/microfibril network. This review describes the clinical relevance of elastin in the context of the structure and function of healthy and aging skin, wound healing, and scars and introduces new approaches being developed to target elastin production and elastic fiber formation.

Elastin as a Nonthrombogenic Biomaterial

Publisher: Mary Ann Liebert Inc

Date: 04-2011

DOI: 10.1089/TEN.TEB.2010.0432

Erratum to large-scale investigation of leishmania interaction networks with host extracellular matrix by surface plasmon resonance imaging [Infection and Immunity 82, 2, (2014) 594-606]

Publisher: American Society for Microbiology

Date: 04-2014

DOI: 10.1128/IAI.01561-14

Tropoelastin Promotes the Formation of Dense, Interconnected Endothelial Networks

Publisher: MDPI AG

Date: 06-09-2021

DOI: 10.3390/BIOM11091318

Abstract: Tropoelastin, the soluble precursor of elastin, has been used for regenerative and wound healing purposes and noted for its ability to accelerate wound repair by enhancing vascularization at the site of implantation. However, it is not clear whether these effects are directly due to the interaction of tropoelastin with endothelial cells or communicated to endothelial cells following interactions between tropoelastin and neighboring cells, such as mesenchymal stem cells (MSCs). We adapted an endothelial tube formation assay to model in vivo vascularization with the goal of exploring the stimulatory mechanism of tropoelastin. In the presence of tropoelastin, endothelial cells formed less tubes, with reduced spreading into capillary-like networks. In contrast, conditioned media from MSCs that had been cultured on tropoelastin enhanced the formation of more dense, complex, and interconnected endothelial tube networks. This pro-angiogenic effect of tropoelastin is mediated indirectly through the action of tropoelastin on co-cultured cells. We conclude that tropoelastin inhibits endothelial tube formation, and that this effect is reversed by pro-angiogenic crosstalk from tropoelastin-treated MSCs. Furthermore, we find that the known in vivo pro-angiogenic effects of tropoelastin can be modeled in vitro, highlighting the value of tropoelastin as an indirect mediator of angiogenesis.

Plasma-Activated Tropoelastin Functionalization of Zirconium for Improved Bone Cell Response

Publisher: American Chemical Society (ACS)

Date: 08-03-2016

DOI: 10.1021/ACSBIOMATERIALS.6B00049

Effect of Dense Gas CO₂ on the Coacervation of Elastin

Publisher: American Chemical Society (ACS)

Date: 26-03-2008

DOI: 10.1021/BM700891B

Abstract: In this study for the first time the effect of high-pressure CO2 on the coacervation of alpha-elastin was investigated using analytical techniques including light spectroscopy and attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopic imaging and circular dichroism (CD) spectroscopy. The coacervation behavior of alpha-elastin, a protein biopolymer, was determined at temperatures below 40 degrees C and pressures lower than 180 bar. At these conditions elevated pressures did not disrupt the ability of alpha-elastin to coacervate. It was feasible to monitor the presence of amide I, II, and III bands for alpha-elastin at high-pressure CO2 using ATR-FTIR imaging. At a constant temperature the peak absorption was substantially enhanced by increasing the pressure of the system. CD analysis demonstrated the preservation of secondary structure attributes of alpha-elastin exposed to dense gas CO2 at the pressure range investigated in this study. The lower critical solution temperature of alpha-elastin was dramatically decreased from 37 to 16 degrees C when the CO2 pressure increased from 1 to 50 bar, without a significant change after that. Carbon dioxide at high pressures also impeded the reversible coacervation of alpha-elastin solution. These effects were predominantly associated with the lowered pH of the aqueous solution and maybe the interaction between CO2 and hydrophobic domains of alpha-elastin.

Photocrosslinkable Gelatin/Tropoelastin Hydrogel Adhesives for Peripheral Nerve Repair

Publisher: Mary Ann Liebert Inc

Date: 09-2018

DOI: 10.1089/TEN.TEA.2017.0502

A radioassay for synaptic core complex assembly: Screening of herbal extracts for effectors

Publisher: Elsevier BV

Date: 10-2006

DOI: 10.1016/J.AB.2006.05.036

Abstract: Synaptic core complex formation between syntaxin and synaptosome-associated protein of 25 kDa (SNAP25) on the plasma membrane and synaptobrevin on the vesicle membrane is responsible for membrane fusion and neurotransmitter release. A radiolabeled protein binding assay for synaptic core complex formation was developed. The components of this assay included recombinant glutathione S-transferase (GST)-syntaxin immobilized on glutathione agarose beads, SNAP25 and (125)I-labeled synaptobrevin. Reactions were performed in tubes containing filter inserts to facilitate removal of unbound protein. The radiolabeled protein bound was then quantified by gamma counter. A K(d) of 1.6 microM was determined for the GST-syntaxin/SNAP25/synaptobrevin complex, and a K(d) of 12 microM was determined for the GST-syntaxin/synaptobrevin complex. The assay was used to screen 14 herbal extracts for effectors of core complex formation. Herbs traditionally used to treat neurological conditions such as depression, anxiety, and stress were chosen. A Hypericum perforatum extract was found to have a nonspecific effect via protein complexation, whereas an Albizzia julibrissin extract was found to reduce the level of core complex formation. The assay was used to further investigate the effect of the A. julibrissin extract. The discovery of an inhibitor of core complex formation demonstrates the efficacy of the assay in screening natural products for substances that affect core complex formation.

Stress-induced senescence in mesenchymal stem cells: Triggers, hallmarks, and current rejuvenation approaches

Publisher: Elsevier BV

Date: 06-2023

DOI: 10.1016/J.EJCB.2023.151331

Tropoelastin Incorporation into a Dermal Regeneration Template Promotes Wound Angiogenesis

Publisher: Wiley

Date: 02-12-2014

DOI: 10.1002/ADHM.201400571

Abstract: Severe burn injury results in substantial skin loss and cannot be treated by autografts. The Integra Dermal Regeneration Template (IDRT) is the leading synthetic skin substitute because it allows for wound bed regeneration and wound healing. However, all substitutes suffer from slow blood vessel ingrowth and would benefit considerably from enhanced vascularization to nurture tissue repair. It is shown here that by incorporating the human elastic protein tropoelastin into a dermal regeneration template (TDRT) we can promote angiogenesis in wound healing. In small and large animal models comprising mice and pigs, the hybrid TDRT biomaterial and IDRT show similar contraction to autografts and decrease wound contraction compared to open wounds. In mice, TDRT accelerates early stage angiogenesis by 2 weeks, as evidenced by increased angiogenesis fluorescent radiant efficiency in live animal imaging and the expression of endothelial cell adhesion marker CD146. In the pig, a full thickness wound repair model confirms increased numbers of blood vessels in the regenerating areas of the dermis closest to the hypodermis and immediately below the epidermis at 2 weeks post-surgery. It is concluded that including tropoelastin in a dermal regeneration template has the potential to promote wound repair through enhanced vascularization.

Elevated levels of glycoprotein gp200 in progeria fibroblasts

Publisher: Springer Science and Business Media LLC

Date: 1993

DOI: 10.1007/BF00925984

Conservation of the 168 divIB gene in Bacillus subtilis W23 and B. licheniformis, and evidence for homology to ftsQ of Escherichia coli

Publisher: Elsevier BV

Date: 09-1994

DOI: 10.1016/0378-1119(94)90043-4

Abstract: The chromosomal regions of Bacillus subtilis (Bs) W23 and Bacillus licheniformis (Bl), which span the sequence encoding the homolog of the ision initiation gene, IB, of Bs168 were cloned and sequenced. The high level of conservation of the amino acid (aa) sequence of the DivIB protein (99 and 68% identity for BsW23 and Bl, respectively) was consistent with a significant role for this protein in the cell cycle of the two species. The hydropathy profile for DivIB of Bl was almost identical to that of Bs168 and consistent with a membrane location, as previously established for the latter. The higher than average level of identity (87%) of the 31-aa N-terminal cytoplasmic domain of DivIB between Bs168 and Bl raised the possibility of a special role for this domain. Database analyses using the Bl DivIB sequence and similarity analyses also strongly suggested that DivIB, of Bl and Bs, is a homolog of FtsQ of Escherichia coli. The flanking sequences extending into the unidentified orfs both upstream and downstream from IB were highly conserved between Bs168 and Bl at both the nucleotide and aa levels. It was confirmed that orf4 of Bs168 is dispensable.

The immobilization of recombinant human tropoelastin on metals using a plasma-activated coating to improve the biocompatibility of coronary stents

Publisher: Elsevier BV

Date: 11-2010

DOI: 10.1016/J.BIOMATERIALS.2010.07.062

Abstract: Current endovascular stents have sub-optimal biocompatibility reducing their clinical efficacy. We previously demonstrated a plasma-activated coating (PAC) that covalently bound recombinant human tropoelastin (TE), a major regulator of vascular cells in vivo, to enhance endothelial cell interactions. We sought to develop this coating to enhance its mechanical properties and hemocompatibility for application onto coronary stents. The plasma vapor composition was altered by incorporating argon, nitrogen, hydrogen or oxygen to modulate coating properties. Coatings were characterized for 1) surface properties, 2) mechanical durability, 3) covalent protein binding, 4) endothelial cell interactions and 5) thrombogenicity. The N(2)/Ar PAC had optimal mechanical properties and did not delaminate after stent expansion. The N(2)/Ar PAC was mildly hydrophilic and covalently bound the highest proportion of TE, which enhanced endothelial cell proliferation. Acute thrombogenicity was assessed in a modified Chandler loop using human blood. Strikingly, the N(2)/Ar PAC alone reduced thrombus weight by ten-fold compared to 316L SS, a finding unaltered with immobilized TE. Serum soluble P-selectin was reduced on N(2)/Ar PAC and N(2)/Ar PAC + TE (p < 0.05), consistent with reduced platelet activation. We have demonstrated a coating for metal alloys with multifaceted biocompatibility that resists delamination and is non-thrombogenic, with implications for improving coronary stent efficacy.

Tropoelastin inhibits intimal hyperplasia of mouse bioresorbable arterial vascular grafts.

Publisher: Elsevier BV

Date: 04-2017

DOI: 10.1016/J.ACTBIO.2016.12.044

Endothelial Progenitor Cell (EPC) Interaction with Recombinant Human Elastin (rhELN) is Through Integrin αVβ3

Publisher: Elsevier BV

Date: 2012

DOI: 10.1016/J.HLC.2012.05.043

HiPIMS carbon coatings show covalent protein binding that imparts enhanced hemocompatibility

Publisher: Elsevier BV

Date: 11-2018

DOI: 10.1016/J.CARBON.2018.06.024

Allysine modifications perturb tropoelastin structure and mobility on a local and global scale

Publisher: Elsevier BV

Date: 05-2019

DOI: 10.1016/J.MBPLUS.2019.03.001

Building Elastin

Publisher: American Thoracic Society

Date: 06-2001

DOI: 10.1165/AJRCMB.24.6.4304

Abstract: The purpose of this study was to assess the feasibility of crosslinking exogenously produced tropoelastin, the precursor of insoluble elastin, into existing elastin. Tritiated recombinant human tropoelastin (rhTE) was added to neonatal rat aorta smooth-muscle cell (NNRSMC) cultures. As much as 12% of the added rhTE was incorporated into the NNRSMC-derived insoluble elastin with the formation of the elastin crosslinks desmosine (DES) and isodesmosine (IDES) in a time-dependent fashion. The ratio of radioactivity found in DES and IDES crosslinks to that found in lysyl residues increased from 0.18 immediately after incubation with rhTE to 0.76 after 14 d. Crosslinking of rhTE into elastin and the accompanying formation of tritiated water was inhibited by beta-aminoproprionitrile, a potent inhibitor of lysyl oxidase, an enzyme critical for the post-translational processing of elastin and collagen. Acellular NNRSMC matrices were produced and replated with Rat-1 fibroblasts, cells that were found to express lysyl oxidase but not tropoelastin. At 14 d after incubation with rhTE, the ratio of DES and IDES radioactivity to that of lysine in the insoluble elastin was 0.38. We show for the first time that cells expressing lysyl oxidase, but not elastin, as well as elastogenic cells can incorporate rhTE into insoluble elastin with the formation of elastin crosslinks. This novel approach might be used to augment elastin repair in certain pathologic states.

Plasma processing of PDMS based spinal implants for covalent protein immobilization, cell attachment and spreading

Publisher: Springer Science and Business Media LLC

Date: 30-11-2018

DOI: 10.1007/S10856-018-6181-Y

Abstract: PDMS is widely used for prosthetic device manufacture. Conventional ion implantation is not a suitable treatment to enhance the biocompatibility of poly dimethyl siloxane (PDMS) due to its propensity to generate a brittle silicon oxide surface layer which cracks and delaminates. To overcome this limitation, we have developed new plasma based processes to balance the etching of carbon with implantation of carbon from the plasma source. When this carbon was implanted from the plasma phase it resulted in a surface that was structurally similar and intermixed with the underlying PDMS material and not susceptible to delamination. The enrichment in surface carbon allowed the formation of carbon based radicals that are not present in conventional plasma ion immersion implantation (PIII) treated PDMS. This imparts the PDMS surfaces with covalent protein binding capacity that is not observed on PIII treated PDMS. The change in surface energy preserved the function of bound biomolecules and enhanced the attachment of MG63 osteosarcoma cells compared to the native surface. The attached cells, an osteoblast interaction model, showed increased spreading on the treated over untreated surfaces. The carbon-dependency for these beneficial covalent protein and cell linkage properties was tested by incorporating carbon from a different source. To this end, a second surface was produced where carbon etching was balanced against implantation from a thin carbon-based polymer coating. This had similar protein and cell-binding properties to the surfaces generated with carbon inclusion in the plasma phase, thus highlighting the importance of balancing carbon etching and deposition. Additionally, the two effects of protein linkage and bioactivity could be combined where the cell response was further enhanced by covalently tethering a biomolecule coating, as exemplified here with the cell adhesive protein tropoelastin. Providing a balanced carbon source in the plasma phase is applicable to prosthetic device fabrication as illustrated using a 3-dimensional PDMS balloon prosthesis for spinal implant applications. Consequently, this study lays the groundwork for effective treatments of PDMS to selectively recruit cells to implantable PDMS fabricated biodevices.

Effects of Surface Attachment of Recombinant Human Elastin on the Thrombogenicity of Synthetic Vascular Conduits

Publisher: Elsevier BV

Date: 2009

DOI: 10.1016/J.HLC.2009.05.147

Molecular model of human tropoelastin and implications of associated mutations

Publisher: Proceedings of the National Academy of Sciences

Date: 26-06-2018

DOI: 10.1073/PNAS.1801205115

Abstract: Tropoelastin, the precursor molecule to elastic fibers, is a large, flexible elastic protein whose structure has been the subject of investigation and debate over several decades. Here, we present the fully atomistic structure of human tropoelastin, based on molecular dynamics simulations, and validate it with experiments. We explore the functional role of two key residues by inserting alanine substitutions and explain conformational changes and variations in hierarchical assembly. We also predict essential dynamics of the molecule by building elastin network models, to explain experimentally observed differences in assembly. Finally, we study the structural and dynamic molecular changes associated with the acquired cutis laxa disorder. The approach developed here is applicable for studying structure and function of other highly disordered proteins.

Structural disorder and dynamics of elastin.

Publisher: Canadian Science Publishing

Date: 04-2010

DOI: 10.1139/O09-161

Abstract: Elastin is a self-assembling, extracellular-matrix protein that is the major provider of tissue elasticity. Here we review structural studies of elastin from over four decades, and draw together evidence for solution flexibility and conformational disorder that is inherent in all levels of structural organization. The characterization of disorder is consistent with an entropy-driven mechanism of elastic recoil. We conclude that conformational disorder is a constitutive feature of elastin structure and function.

Surface plasma modification and tropoelastin coating of a polyurethane co-polymer for enhanced cell attachment and reduced thrombogenicity

Publisher: Elsevier BV

Date: 08-2014

DOI: 10.1016/J.BIOMATERIALS.2014.04.082

Abstract: Polymers currently utilized for dermal and vascular applications possess sub-optimal biocompatibility which reduces their efficacy. Improving the cell-binding and blood-contacting properties of these polymers would substantially improve their clinical utility. Tropoelastin is a highly extensible extracellular matrix protein with beneficial cell interactive and low thrombogenic properties. We transferred these benefits to the polyurethane block copolymer Elast-Eon E2A through a specific combination of surface plasma modifications and coating with human tropoelastin. The cell-binding activity of bound tropoelastin was modulated by ion implantation of the underlying polymer, and correlated with surface hydrophobicity, carbon and oxygen content. This combined treatment enhanced human dermal fibroblast (HDF) and human umbilical vein endothelial cell (HUVEC) attachment, cytoskeletal assembly and viability, combined with elevated PECAM-1 staining of HUVEC cell junctions. The thrombogenicity of the polymer was ameliorated by tropoelastin coating. We propose that a combination of metered plasma treatment and tropoelastin coating of Elast-Eon can serve to improve the biological performance of implantable devices such as vascular conduits.

The influence of elasticity and surface roughness on myogenic and osteogenic-differentiation of cells on silk-elastin biomaterials

Publisher: Elsevier BV

Date: 12-2011

DOI: 10.1016/J.BIOMATERIALS.2011.08.037

Elastin in Vascular Grafts

Publisher: Springer International Publishing

Date: 2020

DOI: 10.1007/978-3-030-05336-9_13

Coacervation Characteristics of Recombinant Human Tropoelastin

Publisher: Wiley

Date: 11-1997

DOI: 10.1111/J.1432-1033.1997.00092.X

Abstract: Coacervation of soluble tropoelastin molecules is characterized by thermodynamically reversible association as temperature is increased under appropriately juxtaposed ionic conditions, protein concentration and pH. Coacervation plays a critical role in the assembly of these elastin precursors in elastic fiber formation. To examine the effect of physiological parameters on the ability of tropoelastin molecules to associate, solutions of recombinant human tropoelastin were monitored spectrophotometrically by light scattering over a broad range of temperatures. Coacervation of recombinant human tropoelastin is strongly influenced by the concentration of protein and NaCl and to a lesser extent on pH. Trends towards maximal association are apparent when each of these parameters is varied. Remarkably, optimal coacervation is found at 37 degrees C, 150 mM NaCl and pH 7-8. Using the data generated by time courses, estimates of thermodynamic parameters were made. These estimates confirm that coacervation is endothermic and is marked by a strong entropic contribution. Circular dichroism of recombinant human tropoelastin revealed that, rather than being random, the structure is compatible with being largely that, of an all-beta protein (with secondary structure estimated to be 3% alpha-helix, 41% beta-sheet, 21% beta-turn and 33% other), exhibiting a spectrum as previously seen for tropoelastin populations and soluble elastin from naturally-derived sources.

Coacervation is promoted by molecular interactions between the PF2 segment of fibrillin-1 and the domain 4 region of tropoelastin

Publisher: American Chemical Society (ACS)

Date: 09-07-2005

DOI: 10.1021/BI050530D

Abstract: In forming elastic fibers, microfibrils act as the scaffold sites for depositing the elastin precursor tropoelastin. We examined key binding interactions that promote massive tropoelastin association through coacervation. Using a segment of the microfibril protein fibrillin-1, PF2, known to bind full-length tropoelastin, we mapped its interaction site to the N-terminal region of tropoelastin bounded by domains 2 and 18. Precise contact residues between domain 4 of tropoelastin and domain 16 of fibrillin-1 were discovered through a novel combination of transglutaminase cross-linking and mass spectroscopy, with contact sites at residues K38 of tropoelastin and Q669 of fibrillin-1. This is the first report of a role for this region of tropoelastin in microfibril interactions. The addition of PF2 thermodynamically facilitated the coacervation of tropoelastin, resulting in smaller changes in entropy and enthalpy values for the coacervating system. A novel multicomponent in vitro tropoelastin assembly reaction system demonstrated that amassed tropoelastin was spatially and preferentially directed to surfaces coated with PF2 as expected for organized three-dimensional distribution during tissue elastogenesis. This study underscores the role of this part of fibrillin-1 as an anchor point for tropoelastin at the microfibril-elastin junction during the initial stages of elastic fiber assembly.

Plasma mediated protein immobilisation enhances the vascular compatibility of polyurethane with tissue matched mechanical properties

Publisher: IOP Publishing

Date: 04-07-2017

DOI: 10.1088/1748-605X/AA6EB6

Multimer formation as a consequence of separate homodimerization domains: the human c-Jun leucine zipper is a transplantable dimerization module.

Publisher: Oxford University Press (OUP)

Date: 1996

DOI: 10.1093/PROTEIN/9.2.223

Abstract: Human c-Jun and c-Fos leucine zipper domains were examined for their ability to serve as autonomous dimerization domains as part of a heterologous protein construct. Schistosoma japonicum glutathione S-transferase (GST) was fused to recombinant Jun leucine zipper (rJunLZ) and Fos leucine zipper (rFosLZ) domains. SDS-PAGE 'snapshot' analyses based on disulphide linkage of monomers demonstrated the ability of rJunLZ to function as a dimerization motif in a foreign protein environment. Steric hindrance prevented formation of rJunLZ-GST::rFosLZ-GST heterodimers whereas rJunLZ-GST::rFosLZ and rJunLZ:: rFosLZGST formed readily. Furthermore, rJunLZGST generated homodimers suggesting fusion protein heterodimers interact differently to homodimers. Gel filtration chromatography confirmed that GST is a dimer in solution and that attachment of a leucine zipper domain allows further interactions to take place. Sedimentation equilibrium analyses showed that GST is a stable dimer (K(a) > 10(6) M(-1)) with no higher multimeric forms. rFosLZ-GST weakly associates beyond a dimer (K(a) approximately 4 x 10(4) M(-1)) and rJunLZ-GST associates indefinitely (K(a) approximately 4 x 10(5) M(-1)) [corrected], consistent with an isodesmic model of association. The interaction of these leucine zippers independently of GST association demonstrates their utility in the modification of proteins when multimer formation is desired.

Hutchinson-Gilford progeria types defined by differential binding of lectin DSA.

Publisher: Elsevier BV

Date: 04-1995

DOI: 10.1016/0925-4439(94)00081-Z

Abstract: Hutchinson-Gilford Progeria Syndrome (progeria) is an extremely rare childhood disorder characterized by precocious senility which presents features similar to those seen in human aging. We have previously described a consistent increase of the glycoprotein gp200 in progeria skin fibroblasts in vitro. Here we extend these glycosylation studies and present evidence for the existence of two types of progeria skin fibroblasts. These two forms, called D- and D+, are distinguished by their response to the lectin DSA. In the D- group, DSA bound glycoproteins from progeria fibroblast strains AG03513B and AG10750 with markedly lower affinities compared with glycoproteins from three control fibroblast strains. In the D+ group, DSA binding to glycoproteins from four other progeria strains AG01972A, AG06297A, AG06917 and AG03198, was comparable to controls. Discrimination by DSA is the most distinctive feature of the D- and D+ groups, in contrast to binding of lectins Con A, GNA, PHA-L, RCA120, AAA and PNA which show no such selectivity. The data are consistent with a model of altered glycosylation in the D- type of progeria fibroblasts.

Electrospun protein fibers as matrices for tissue engineering

Publisher: Elsevier BV

Date: 10-2005

DOI: 10.1016/J.BIOMATERIALS.2005.03.030

Abstract: Electrospinning has recently emerged as a leading technique for generating biomimetic scaffolds made of synthetic and natural polymers for tissue engineering applications. In this study, we compared collagen, gelatin (denatured collagen), solubilized alpha-elastin, and, as a first, recombinant human tropoelastin as biopolymeric materials for fabricating tissue engineered scaffolds by electrospinning. In extending previous studies, we optimized the shape and size (diameter or width) of the ensuing electrospun fibers by varying important parameters of the electrospinning process, such as solute concentration and delivery rate of the polymers. Our results indicate that the average diameter of gelatin and collagen fibers could be scaled down to 200-500 nm without any beads, while the alpha-elastin and tropoelastin fibers were several microns in width. Importantly, and contrary to any hitherto reported structures of electrospun polymers, fibers composed of alpha-elastin, especially tropoelastin, exhibited "quasi-elastic" wave-like patterns at increased solution delivery rates. The periodicity of these wave-like tropoelastin fibers was partly affected by the delivery rate. Atomic force microscopy was utilized to profile the topography of in idual electrospun fibers and microtensile testing was performed to measure their mechanical properties. Cell culture studies confirmed that the electrospun engineered protein scaffolds support attachment and growth of human embryonic palatal mesenchymal (HEPM) cells.

Integrin alpha(v)beta(3) binds a unique non-RGD site near the C-terminus of human tropoelastin

Publisher: Elsevier BV

Date: 03-2004

DOI: 10.1016/J.BIOCHI.2004.03.002

A Novel Elastin-coated e-PTFE Vascular Conduit

Publisher: Elsevier BV

Date: 08-2010

DOI: 10.1016/J.HLC.2010.04.104

Primary human dermal fibroblast interactions with open weave three-dimensional scaffolds prepared from synthetic human elastin

Publisher: Elsevier BV

Date: 11-2009

DOI: 10.1016/J.BIOMATERIALS.2009.08.017

Abstract: We present an elastic, fibrous human protein-based and cell-interactive dermal substitute scaffold based on synthetic human elastin. Recombinant human tropoelastin promoted primary human dermal fibroblast attachment, spreading and proliferation. Tropoelastin was cross-linked to form a synthetic elastin (SE) hydrogel matrix and electrospun into fibrous SE scaffolds. Fibroblasts attached to and proliferated across SE hydrogel scaffold surfaces for at least 14 days and deposited the extracellular matrix proteins fibronectin and collagen type I. To allow for the benefit of greater cell infiltration, SE was electrospun into open weave, fibrous scaffolds that closely mimic the fibrous nature of the skin dermis. 3D SE scaffolds were robust and consisted of flat, ribbon-like fibers with widths that are similar to native dermal elastic fibers. The scaffolds displayed elasticity close to that of natural elastin. 3D SE retained the ability to interact with primary human dermal fibroblasts, which consistently attached and proliferated to form monolayers spanning the entire scaffold surface. The open weave design, with larger spaces between in idual fibers and greater fiber diameters beneficially allowed for substantial cell infiltration throughout the scaffolds.

Sex determination using the polymerase chain reaction

Publisher: Elsevier BV

Date: 10-1999

DOI: 10.1016/S0307-4412(99)00044-8

Bioengineering a Human Elastin-derived Small-diameter Conduit with Biomechanical and Biological Properties of a Native Human Artery

Publisher: Elsevier BV

Date: 2009

DOI: 10.1016/J.HLC.2009.05.139

A cost-effective and enhanced mesenchymal stem cell expansion platform with internal plasma-activated biofunctional interfaces

Publisher: Elsevier BV

Date: 10-2023

DOI: 10.1016/J.MTBIO.2023.100727

Domains 17-27 of tropoelastin contain key regions of contact for coacervation and contain an unusual tum-containing crosslinking domain

Publisher: Elsevier BV

Date: 03-2007

DOI: 10.1016/J.MATBIO.2006.10.002

Abstract: The central region of tropoelastin including domains 19-25 of human tropoelastin forms a hot-spot for contacts during the inter-molecular association of tropoelastin by coacervation [Wise, S.G., Mithieux, S.M., Raftery, M.J. and Weiss, A.S (2005). "Specificity in the coacervation of tropoelastin: solvent exposed lysines." Journal of Structural Biology 149: 273-81.]. We explored the physical properties of this central region using a sub-fragment bordered by domains 17-27 of human tropoelastin (SHEL 17-27) and identified the intra- and inter-molecular contacts it forms during coacervation. A homobifunctional amine reactive crosslinker (with a maximum reach of 11 A, corresponding to approximately 7 residues in an extended polypeptide chain) was used to capture these contacts and crosslinked regions were identified after protease cleavage and mass spectrometry (MS) with MS/MS verification. An intermolecular crosslink formed between the lysines at positions 353 of each strand of tropoelastin at the lowest of crosslinker concentrations and was observed in all s les tested, suggesting that this residue forms an important initial contact during coacervation. At higher crosslinker concentrations, residues K425 and K437 showed the highest levels of involvement in crosslinks. An intramolecular crosslink between these K425 and K437, separated by 11 residues, indicated that a structural bend must serve to bring these residues into close proximity. These studies were complemented by small angle X-ray scattering studies that confirmed a bend in this important subfragment of the tropoelastin molecule.

Hutchinson-Gilford progeria: faithful DNA maintenance, inheritance and allelic transcription of beta(1-4) galactosyltransferase.

Publisher: Elsevier BV

Date: 03-1998

DOI: 10.1016/S0047-6374(97)00163-2

Abstract: Hutchinson-Gilford progeria syndrome (HGPS) is a fatal segmental aging disorder affecting children. There is a paucity of prior data at the nucleotide level on DNA maintenance in HGPS. We have examined the specific nucleotide sequences and production of allelic transcripts from the locus GGTB2 encoding beta(1-4) galactosyltransferase. Quantitative Northern blots of mRNA from HGPS and control fibroblasts indicated identical mature beta(1-4) galactosyltransferase transcript sizes and amounts, regardless of their altered glycosylation status. DNA sequencing of cDNA derived from HGPS beta(1-4) galactosyltransferase mRNA populations confirmed the encoded amino acid sequence was unaffected. Population studies of 41 unrelated in iduals provided allelic frequency estimates for a novel FokI polymorphism, which was identified in two of six progeria cell strains. The polymorphism was faithfully inherited in a progeria pedigree in a Mendelian manner. Furthermore, the polymorphism provided direct evidence through sequencing of reverse transcription polymerase chain reaction products that both alleles were transcribed and generated mature mRNA. Any defects in transcripts were below detectable levels over the lengths of coding sequences examined, despite multiple replication events from conception leading to the production and maintenance of patient-derived cells. These results indicate faithful transcription in HGPS.

Perspectives on the molecular and biological implications of tropoelastin in human tissue elasticity

Publisher: CSIRO Publishing

Date: 2016

DOI: 10.1071/CH16452

Abstract: The elasticity of a range of vertebrate and particularly human tissues depends on the dynamic and persistent protein elastin. This elasticity is erse, and comprises skin, blood vessels, and lung, and is essential for tissue viability. Elastin is predominantly made by assembling tropoelastin, which is an asymmetric 20-nm-long protein molecule. This overview considers tropoelastin’s molecular features and biological interactions in the context of its value in tissue repair.

Large-Scale Investigation of Leishmania Interaction Networks with Host Extracellular Matrix by Surface Plasmon Resonance Imaging

Publisher: American Society for Microbiology

Date: 02-2014

DOI: 10.1128/IAI.01146-13

Abstract: We have set up an assay to study the interactions of live pathogens with their hosts by using protein and glycosaminoglycan arrays probed by surface plasmon resonance imaging. We have used this assay to characterize the interactions of Leishmania promastigotes with ∼70 mammalian host biomolecules (extracellular proteins, glycosaminoglycans, growth factors, cell surface receptors). We have identified, in total, 27 new partners (23 proteins, 4 glycosaminoglycans) of procyclic promastigotes of six Leishmania species and 18 partners (15 proteins, 3 glycosaminoglycans) of three species of stationary-phase promastigotes for all the strains tested. The ersity of the interaction repertoires of Leishmania parasites reflects their dynamic and complex interplay with their mammalian hosts, which depends mostly on the species and strains of Leishmania . Stationary-phase Leishmania parasites target extracellular matrix proteins and glycosaminoglycans, which are highly connected in the extracellular interaction network. Heparin and heparan sulfate bind to most Leishmania strains tested, and 6- O -sulfate groups play a crucial role in these interactions. Numerous Leishmania strains bind to tropoelastin, and some strains are even able to degrade it. Several strains interact with collagen VI, which is expressed by macrophages. Most Leishmania promastigotes interact with several regulators of angiogenesis, including antiangiogenic factors (endostatin, anastellin) and proangiogenic factors (ECM-1, VEGF, and TEM8 [also known as anthrax toxin receptor 1]), which are regulated by hypoxia. Since hypoxia modulates the infection of macrophages by the parasites, these interactions might influence the infection of host cells by Leishmania .

Cell patterning via linker-free protein functionalization of an organic conducting polymer (polypyrrole) electrode

Publisher: Elsevier BV

Date: 07-2012

DOI: 10.1016/J.ACTBIO.2012.03.023

Abstract: The interaction of proteins and cells with polymers is critical to their use in scientific and medical applications. In this study, plasma immersion ion implantation (PIII) was used to modify the surface of the conducting polymer, polypyrrole, which possesses electrical properties. PIII treatment enabled persistent, covalent binding of the cell adhesive protein, tropoelastin, without employing chemical linking molecules. In contrast tropoelastin was readily eluted from the untreated surface. Through this differential persistence of binding, surface bound tropoelastin supported cell adhesion and spreading on the PIII treated but not the untreated polypyrrole surface. The application of a steel shadow mask during PIII treatment allowed for spatial definition of tropoelastin exclusively to PIII treated regions. The general applicability of this approach to other extracellular matrix proteins was illustrated using collagen I, which displayed similar results to tropoelastin but required extended washing conditions. This approach allowed fine patterning of cell adhesion and spreading to tropoelastin and collagen, specifically on PIII treated polypyrrole regions. We therefore present a methodology to alter the functionality of polypyrrole surfaces, generating surfaces that can spatially control cellular interactions through protein functionalization with the potential for electrical stimulation.

Polyglycerol sebacate‐based elastomeric materials for arterial regeneration

Publisher: Wiley

Date: 22-06-2023

DOI: 10.1002/JBM.A.37583

Abstract: Synthetic vascular grafts are commonly used in patients with severe occlusive arterial disease when autologous grafts are not an option. Commercially available synthetic grafts are confronted with challenging outcomes: they have a lower patency rate than autologous grafts and are currently unable to promote arterial regeneration. Polyglycerol sebacate (PGS), a non‐toxic polymer with a tunable degradation profile, has shown promising results as a small‐diameter vascular graft component that can support the formation of neoarteries. In this review, we first present an overview of the synthesis and modification of PGS followed by an examination of its mechanical properties. We then report on the performance, degradation, regeneration, and remodeling of PGS‐based small‐diameter vascular grafts, with a focus on efforts to reduce thrombosis, prevent dilation, and promote cellular residency and extracellular matrix regeneration that resembles the native artery in spatial distribution and organization. We also highlight recent advances in the incorporation of novel in situ cell sources for arterial regeneration and their potential application in PGS‐based vascular grafts. Finally, we compare vascular grafts fabricated using PGS‐based materials with other elastomeric alternatives.

Direct covalent coupling of proteins to nanostructured plasma polymers: a route to tunable cell adhesion

Publisher: Elsevier BV

Date: 10-2015

DOI: 10.1016/J.APSUSC.2015.05.162

Cross-linked open-pore elastic hydrogels based on tropoelastin, elastin and high pressure CO2

Publisher: Elsevier BV

Date: 03-2010

DOI: 10.1016/J.BIOMATERIALS.2009.11.051

Abstract: In this study the effect of high pressure CO(2) on the synthesis and characteristics of elastin-based hybrid hydrogels was investigated. Tropoelastin/alpha-elastin hybrid hydrogels were fabricated by chemically cross-linking tropoelastin/alpha-elastin solutions with glutaraldehyde at high pressure CO(2). Dense gas CO(2) had a significant impact on the characteristics of the fabricated hydrogels including porosity, swelling ratio, compressive properties, and modulus of elasticity. Compared to fabrication at atmospheric pressure high pressure CO(2) based construction eliminated the skin-like formation on the top surfaces of hydrogels and generated larger pores with an average pore size of 78 +/- 17 microm. The swelling ratios of composite hydrogels fabricated at high pressure CO(2) were lower than the gels produced at atmospheric pressure as a result of a higher degree of cross-linking. Dense gas CO(2) substantially increased the mechanical properties of fabricated hydrogels. The compressive and tensile modulus of 50/50 weight ratio tropoelastin/alpha-elastin composite hydrogels were enhanced 2 and 2.5 fold, respectively, when the pressure was increased from 1 to 60 bar. In vitro studies show that the presence of large pores throughout the hydrogel matrix fabricated at high pressure CO(2) enabled the migration of human skin fibroblast cells 300 microm into the construct.

Heparan sulphate interacts with tropoelastin, with some tropoelastin peptides and is present in human dermis elastic fibers

Publisher: Elsevier BV

Date: 02-2005

DOI: 10.1016/J.MATBIO.2004.12.001

Abstract: A number of reports point to the presence of proteoglycans and/or glycosaminoglycans within elastic fibers in normal and in pathological conditions. We present data that heparan sulphate (HS)-containing proteoglycans are associated with normal elastic fibers in human dermis and that isolated HS chains interact in vitro with recombinant tropoelastin and with peptides encoded by distinct exons of the human tropoelastin gene (EDPs). By immunocytochemistry, HS chains were identified as associated with the amorphous elastin component in the human dermis and remained associated with the residual elastin in the partially degenerated fibers of old subjects. HS appeared particularly concentrated in the mineralization front of elastic fibers in the dermis of patients affected by pseudoxanthoma elasticum (PXE). In in vitro experiments, HS induced substantial changes in the coacervation temperature and in the aggregation properties of recombinant tropoelastin and of synthetic peptides (EDPs) corresponding to sequences encoded by exons 18, 20, 24 and 30 of the human tropoelastin gene. In particular, HS modified the coacervation temperature and favoured the aggregation into ordered structures of tropoelastin molecules and of EDPs 18, 20 and 24, but not of EDP30. These data strongly indicate that HS-elastin interactions may play a role in tissue elastin fibrogenesis as well as modulating elastin stability with time and in diseases.

Covalently bound biomimetic layers on plasma polymers with graded metallic interfaces for in vivo implants

Publisher: Wiley

Date: 13-10-2009

DOI: 10.1002/PPAP.200900045

Cloning, expression, and spectroscopic studies of the Jun leucine zipper domain.

Publisher: Wiley

Date: 02-1994

DOI: 10.1111/J.1432-1033.1994.TB18569.X

Abstract: Association of the human c-Jun and c-Fos proteins depends upon interactions involving their leucine zipper domains. We are interested in elucidating the tertiary structure of the Jun and Fos leucine zipper domains with a view to understanding the precise intermolecular interactions which govern the affinity and specificity of interaction in these proteins, which have the unusual capacity to form either homodimeric or heterodimeric zipper pairs. With this goal in mind, we have developed a bacterial expression system for the efficient production of both unlabelled and isotopically labelled c-Jun leucine zipper domain. A synthetic junLZ gene was created by annealing, ligation, and polymerase-chain-reaction lification of overlapping synthetic oligonucleotides which comprised 132 bp of coding sequence encompassing residues Arg276-Asn314 of c-Jun plus a total of five engineered non-native residues at the N- and C-termini. The junLZ gene was cloned into the pGEX-2T vector from which recombinant c-Jun leucine zipper domain (rJunLZ 46 residues, 5.1 kDa) was overexpressed (approximately 15% total cell protein) in Escherichia coli as a fusion protein of 31.4 kDa, consisting of rJunLZ fused to the carboxy-terminal portion of Schistosoma japonicum glutathione S-transferase. Two markedly different expression strategies have been devised which allow purification of rJunLZ from the soluble or inclusion-body fraction of induced cells. We have used these strategies to produce unlabelled and uniformly 15N-labelled rJunLZ for NMR studies which, in combination with circular dichroic measurements, reveal that rJunLZ most likely forms a symmetric coiled-coil of parallel alpha-helices. We also present 15N-NMR chemical shift assignments for the backbone and sidechain amide nitrogens of rJunLZ, which should assist in determination of a high-resolution structure of the homodimeric Jun leucine zipper using heteronuclear three-dimensional NMR spectroscopy.

Coacervation of tropoelastin

Publisher: Elsevier BV

Date: 09-2011

DOI: 10.1016/J.CIS.2010.10.003

Abstract: The coacervation of tropoelastin represents the first major stage of elastic fiber assembly. The process has been modeled in vitro by numerous studies, initially with mixtures of solubilized elastin, and subsequently with synthetic elastin peptides that represent hydrophobic repeat units, isolated hydrophobic domains, segments of alternating hydrophobic and cross-linking domains, or the full-length monomer. Tropoelastin coacervation in vitro is characterized by two stages: an initial phase separation, which involves a reversible inverse temperature transition of monomer to n-mer and maturation, which is defined by the irreversible coalescence of coacervates into large species with fibrillar structures. Coacervation is an intrinsic ability of tropoelastin. It is primarily influenced by the number, sequence, and contextual arrangement of hydrophobic domains, although hydrophilic sequences can also affect the behavior of the hydrophobic domains and thus affect coacervation. External conditions including ionic strength, pH, and temperature also directly influence the propensity of tropoelastin to self-associate. Coacervation is an endothermic, entropically-driven process driven by the cooperative interactions of hydrophobic domains following destabilization of the clathrate-like water shielding these regions. The formation of such assemblies is believed to follow a helical nucleation model of polymerization. Coacervation is closely associated with conformational transitions of the monomer, such as increased β-structures in hydrophobic domains and α-helices in cross-linking domains. Tropoelastin coacervation in vivo is thought to mainly involve the central hydrophobic domains. In addition, cell-surface glycosaminoglycans and microfibrillar proteins may regulate the process. Coacervation is essential for progression to downstream elastogenic stages, and impairment of the process can result in elastin haploinsufficiency disorders such as supravalvular aortic stenosis.

Bio-functionalisation of polyether ether ketone using plasma immersion ion implantation

Publisher: SPIE

Date: 22-12-2015

DOI: 10.1117/12.2202506

The use of plasma-activated covalent attachment of early domains of tropoelastin to enhance vascular compatibility of surfaces

Publisher: Elsevier BV

Date: 10-2013

DOI: 10.1016/J.BIOMATERIALS.2013.06.036

Engineering magnetically responsive tropoelastin spongy-like hydrogels for soft tissue regeneration

Publisher: Royal Society of Chemistry (RSC)

Date: 2018

DOI: 10.1039/C7TB02035J

Abstract: Magnetic biomaterials are a key focus in medical research.

Tubular Fibrous Scaffolds Functionalized with Tropoelastin as a Small-Diameter Vascular Graft

Publisher: American Chemical Society (ACS)

Date: 17-07-2020

DOI: 10.1021/ACS.BIOMAC.0C00599

Biocompatibility of silk-tropoelastin protein polymers

Publisher: Elsevier BV

Date: 06-2014

DOI: 10.1016/J.BIOMATERIALS.2014.03.024

Abstract: Blended polymers are used extensively in many critical medical conditions as components of permanently implanted devices. Hybrid protein polymers containing recombinant human tropoelastin and silk fibroin have favorable characteristics as implantable scaffolds in terms of mechanical and biological properties. A firefly luciferase transgenic mouse model was used to monitor real-time IL-1β production localized to the site of biomaterial implantation, to observe the acute immune response (up to 5 days) to these materials. Significantly reduced levels of IL-1β were observed in silk/tropoelastin implants compared to control silk only implants at 1, 2 and 3 days post-surgery. Subsequently, mice (n = 9) were euthanized at 10 days (10D) and 3 weeks (3W) post-surgery to assess inflammatory cell infiltration and collagen deposition, using histopathology and immunohistochemistry. Compared to control silk only implants, fewer total inflammatory cells were found in silk/tropoelastin (∼29% at 10D and ∼47% at 3W). Also fewer ingrowth cells (∼42% at 10D and ∼63% at 3W) were observed within the silk/tropoelastin implants compared to silk only. Lower IL-6 (∼52%) and MMP-2 (∼84%) (pro-inflammatory) were also detected for silk/tropoelastin at 10 days. After 3 weeks implantation, reduced neovascularization (vWF ∼43%), fewer proliferating cells (Ki67 ∼58% and PCNA ∼41%), macrophages (F4/80 ∼64%), lower IL-10 (∼47%) and MMP-9 (∼55%) were also observed in silk/tropoelastin materials compared to silk only. Together, these results suggest that incorporation of tropoelastin improves on the established biocompatibility of silk fibroin, uniquely measured here as a reduced foreign body inflammatory response.

Elastin based cell-laden injectable hydrogels with tunable gelation, mechanical and biodegradation properties

Publisher: Elsevier BV

Date: 07-2014

DOI: 10.1016/J.BIOMATERIALS.2014.03.026

Tropoelastin and Elastin Assembly

Publisher: Frontiers Media SA

Date: 25-02-2021

DOI: 10.3389/FBIOE.2021.643110

Abstract: Elastic fibers are an important component of the extracellular matrix, providing stretch, resilience, and cell interactivity to a broad range of elastic tissues. Elastin makes up the majority of elastic fibers and is formed by the hierarchical assembly of its monomer, tropoelastin. Our understanding of key aspects of the assembly process have been unclear due to the intrinsic properties of elastin and tropoelastin that render them difficult to study. This review focuses on recent developments that have shaped our current knowledge of elastin assembly through understanding the relationship between tropoelastin’s structure and function.

Elastin Biopolymers

Publisher: Elsevier

Date: 2011

DOI: 10.1016/B978-0-08-055294-1.00071-4

Blended Polyurethane and Tropoelastin as a Novel Class of Biologically Interactive Elastomer

Publisher: Mary Ann Liebert Inc

Date: 03-2016

DOI: 10.1089/TEN.TEA.2015.0409

Shape of tropoelastin, the highly extensible protein that controls human tissue elasticity

Publisher: Proceedings of the National Academy of Sciences

Date: 28-02-2011

DOI: 10.1073/PNAS.1014280108

Abstract: Elastin enables the reversible deformation of elastic tissues and can withstand decades of repetitive forces. Tropoelastin is the soluble precursor to elastin, the main elastic protein found in mammals. Little is known of the shape and mechanism of assembly of tropoelastin as its unique composition and propensity to self-associate has h ered structural studies. In this study, we solve the nanostructure of full-length and corresponding overlapping fragments of tropoelastin using small angle X-ray and neutron scattering, allowing us to identify discrete regions of the molecule. Tropoelastin is an asymmetric coil, with a protruding foot that encompasses the C-terminal cell interaction motif. We show that in idual tropoelastin molecules are highly extensible yet elastic without hysteresis to perform as highly efficient molecular nanosprings. Our findings shed light on how biology uses this single protein to build durable elastic structures that allow for cell attachment to an appended foot. We present a unique model for head-to-tail assembly which allows for the propagation of the molecule’s asymmetric coil through a stacked spring design.

Elastin Based Constructs

Publisher: InTech

Date: 29-08-2011

DOI: 10.5772/23660

Tropoelastin

Publisher: Elsevier BV

Date: 03-2009

DOI: 10.1016/J.BIOCEL.2008.03.017

Abstract: Tropoelastin is a 60-72 kDa alternatively spliced extracellular matrix protein and a key component of elastic fibres. It is found in all vertebrates except for cyclostomes. Secreted tropoelastin is tethered to the cell surface, where it aggregates into organised spheres for cross-linking and incorporation into growing elastic fibres. Tropoelastin is characterised by alternating hydrophobic and hydrophilic domains and is highly flexible. The conserved C-terminus is an area of the molecule of particular biological importance in that it is required for both incorporation into elastin and for cellular interactions. Mature cross-linked tropoelastin gives elastin, which confers resilience and elasticity on a erse range of tissues. Elastin gene disruptions in disease states and knockout mice emphasise the importance of proper tropoelastin production and assembly, particularly in vascular tissue. Tropoelastin constructs hold promise as biomaterials as they mimic many of elastin's physical and biological properties with the capacity to replace damaged elastin-rich tissue.

Hyaluronic acid in progeria and the aged phenotype?

Publisher: S. Karger AG

Date: 1992

DOI: 10.1159/000213321

Abstract: Hyaluronic acid (HA) is implicated in functions such as vascularity, morphogenesis, repair, and the general integrity of the extracellular matrix. Hence, it is considered possible that HA is involved in the most conspicuous features of the progeroid phenotype. However, it is not known whether the increase in HA excretion seen in progeria patients is due to a primary genetic defect or is a secondary effect due to some deeper problem. The phenomenon of 'normal' aging is suggested to have a more complex etiology and phenotype than progeria and the role of HA levels is less well-defined.

Soluble matrix protein is a potent modulator of mesenchymal stem cell performance

Publisher: Proceedings of the National Academy of Sciences

Date: 18-01-2019

DOI: 10.1073/PNAS.1812951116

Abstract: Extracellular matrix proteins have primarily been designated as supporting scaffolds for cells. This work presents the soluble extracellular matrix component tropoelastin as a powerful proproliferative and cell-attractive molecule that surpasses the potency of conventional growth factors and matrix proteins used in a mesenchymal stem cell (MSC) culture. Tropoelastin is also demonstrated to modulate MSCs both as a substrate coating and as a soluble additive in media, which significantly deviates from the classical dogma of cell anchorage-dependent structural roles of the matrix. We show that these activities of tropoelastin can be harnessed and establish a path to boosting the efficacy of and simplifying processes for clinical MSC expansion and therapeutic MSC recruitment.

Tropoelastin Massively Associates during Coacervation To Form Quantized Protein Spheres

Publisher: American Chemical Society (ACS)

Date: 28-07-2006

DOI: 10.1021/BI0610092

Abstract: Tropoelastin, the precursor of elastin, undergoes a rapid monomer to multimer association in an inverse temperature transition. This association culminates in the rapid formation of stable, optically distinct droplets of tropoelastin. Light scattering and microscope measurements reveal that these droplets are 2-6 microm in diameter. Scanning electron microscopy confirms that the droplets are spherical. Three-dimensional confocal image stacks based on the autofluorescence of tropoelastin reveal that droplets are loaded with hydrated tropoelastin. Droplets are viable intermediates in synthetic elastin macroassembly. Dense clusters of aggregated droplets and partially formed fibers develop when droplets are incubated in the presence of a lysyl oxidase. Lysine-reacting chemical and enzyme-assisted cross-linking conditions generate cross-linked beads due to interactions between multiple, surface-exposed lysine epsilon-amino groups. Droplets represent an efficient mechanism for the bolus delivery during elastogenesis of quantized packages of preaccreted tropoelastin.

Tandem integration of multiple ILV5 copies and elevated transcription in polyploid yeast

Publisher: Wiley

Date: 15-04-1995

DOI: 10.1002/YEA.320110403

Abstract: An industrial yeast strain was modified by introducing DNA into brewing yeast such that the derived cells contain only yeast DNA. Thus selectable markers and bacterial sequences are not present in the final strain, making this procedure attractive for the development of generally acceptable brewing yeast. Linear DNA containing the cloned ILV5 gene was introduced into lager yeast along with an unlinked circular bifunctional plasmid containing a dominant resistance marker. Resistant colonies were screened for site-directed integration of the ILV5 DNA. Candidates were examined by several methods including Southern transfer and polymerase chain reaction. In this way, a strain WM56 was identified containing three tandem copies of ILV5. The lified ILV5 region is stable during repeated subculturing in the absence of selective pressure. Correspondingly elevated levels of ILV5 transcript in strain WM56 compared to the control (i.e. non-tandem) parental strain led to increased amounts of encoded acetohydroxyacid reductoisomerase as evidenced by significantly lower diacetyl production. WM56 appears to be identical to the parental strain judged by CHEF, total restriction digestion patterns, and probing, but differs in the ILV5 region of the chromosome. The method is generally applicable to other yeast strains, and if desired, is amenable to iterated cycles of integration to increase the number of copies.

Interfacial asparagine residues within an amide tetrad contribute to Max helix-loop-helix leucine zipper homodimer stability

Publisher: Elsevier BV

Date: 12-2000

DOI: 10.1074/JBC.M004264200

Plasma Treatment of ePTFE for Covalent Attachment of Human Elastin, and its Effects on Endothelialisation

Publisher: Elsevier BV

Date: 2009

DOI: 10.1016/J.HLC.2009.05.155

Elastolytic Mechanism of a Novel M23 Metalloprotease Pseudoalterin from Deep-sea Pseudoalteromonas sp. CF6-2

Publisher: Elsevier BV

Date: 11-2012

DOI: 10.1074/JBC.M112.405076

Engineered cell-laden human protein-based elastomer

Publisher: Elsevier BV

Date: 07-2013

DOI: 10.1016/J.BIOMATERIALS.2013.03.076

Electrodeposited gels prepared from protein alloys.

Publisher: Future Medicine Ltd

Date: 03-2015

DOI: 10.2217/NNM.14.230

Abstract: Aim: Silk-tropoelastin alloys, composed of recombinant human tropoelastin and regenerated Bombyx mori silk fibroin, are an emerging, versatile class of biomaterials endowed with tunable combinations of physical and biological properties. Electrodeposition of these alloys provides a programmable means to assemble functional gels with both spatial and temporal controllability. Materials & methods: Tropoelastin-modified silk was prepared by enzymatic coupling between tyrosine residues. Hydrogel coatings were electrodeposited using two wire electrodes. Results & discussion: Mechanical characterization and in vitro cell culture revealed enhanced adhesive capability and cellular response of these alloy gels as compared with electrogelled silk alone. Conclusion: These electro-depositable silk-tropoelastin alloys constitute a suitable coating material for nanoparticle-based drug carriers and offer a novel opportunity for on-demand encapsulation/release of nanomedicine.

Elastin in healthy and diseased lung.

Publisher: Elsevier BV

Date: 04-2022

DOI: 10.1016/J.COPBIO.2021.10.025

Abstract: Elastic fibers are an essential part of the pulmonary extracellular matrix (ECM). Intact elastin is required for normal function and its damage contributes profoundly to the etiology and pathology of lung disease. This highlights the need for novel lung-specific imaging methodology that enables high-resolution 3D visualization of the ECM. We consider elastin's involvement in chronic respiratory disease and examine recent methods for imaging and modeling of the lung in the context of advances in lung tissue engineering for research and clinical application.

University Of Sydney

Location: Australia

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Linkage Infrastructure, Equipment And Facilities - Grant ID: LE230100157

Start Date: 2023

End Date: 12-2023

Amount: $1,009,078.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0775598

Start Date: 10-2008

End Date: 11-2008

Amount: $1,000,000.00

Funder: Australian Research Council

Linkage - International - Grant ID: LX0561916

Start Date: 10-2005

End Date: 06-2007

Amount: $24,520.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0344773

Start Date: 02-2003

End Date: 01-2006

Amount: $300,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP110200316

Start Date: 07-2012

End Date: 12-2016

Amount: $540,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP120103911

Start Date: 02-2012

End Date: 12-2014

Amount: $280,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0775720

Start Date: 06-2007

End Date: 06-2008

Amount: $100,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP130103693

Start Date: 04-2013

End Date: 10-2016

Amount: $385,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP210103160

Start Date: 03-2022

End Date: 03-2025

Amount: $539,765.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE100100010

Start Date: 12-2010

End Date: 12-2010

Amount: $720,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0883030

Start Date: 09-2008

End Date: 01-2011

Amount: $450,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP0882243

Start Date: 03-2008

End Date: 03-2012

Amount: $795,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE100100096

Start Date: 2010

End Date: 06-2011

Amount: $450,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP1096608

Start Date: 01-2010

End Date: 01-2013

Amount: $450,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0774289

Start Date: 02-2007

End Date: 02-2010

Amount: $295,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP220101644

Start Date: 07-2022

End Date: 06-2025

Amount: $570,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0882855

Start Date: 2008

End Date: 12-2009

Amount: $900,000.00

Funder: Australian Research Council