Heparan sulfates facilitate harmless amyloidogenic fibril formation interacting with elastin-like peptides

Heparan sulfates (HSs) modulate tissue elasticity in physiopathological conditions by interacting with various matrix constituents as tropoelastin and elastin-derived peptides. HSs bind also to protein moieties accelerating amyloid formation and influencing cytotoxic properties of insoluble fibrils. Interestingly, amyloidogenic polypeptides, despite their supposed pathogenic role, have been recently explored as promising bio-nanomaterials due to their unique and interesting properties. Therefore, we investigated the interactions of HSs, obtained from different sources and exhibiting various degree of sulfation, with synthetic amyloidogenic elastin-like peptides (ELPs), also looking at the effects of these interactions on cell viability and cell behavior using in vitro cultured fibroblasts, as a prototype of mesenchymal cells known to modulate the soft connective tissue environment. Results demonstrate, for the first time, that HSs, with differences depending on their sulfation pattern and chain length, interact with ELPs accelerating aggregation kinetics and amyloid-like fibril formation as well as self-association. Furthermore, these fibrils do not negatively affect fibroblasts' cell growth and parameters of redox balance, and influence cellular adhesion properties. Data provide information for a better understanding of the interactions altering the elastic component in aging and in pathologic conditions and may pave the way for the development of composite matrix-based biomaterials

ECMO for COVID-19 patients in Europe and Israel

Since March 15th, 2020, 177 centres from Europe and Israel have joined the study, routinely reporting on the ECMO support they provide to COVID-19 patients. The mean annual number of cases treated with ECMO in the participating centres before the pandemic (2019) was 55. The number of COVID-19 patients has increased rapidly each week reaching 1531 treated patients as of September 14th. The greatest number of cases has been reported from France (n = 385), UK (n = 193), Germany (n = 176), Spain (n = 166), and Italy (n = 136) .The mean age of treated patients was 52.6 years (range 16–80), 79% were male. The ECMO configuration used was VV in 91% of cases, VA in 5% and other in 4%. The mean PaO2 before ECMO implantation was 65 mmHg. The mean duration of ECMO support thus far has been 18 days and the mean ICU length of stay of these patients was 33 days. As of the 14th September, overall 841 patients have been weaned from ECMO support, 601 died during ECMO support, 71 died after withdrawal of ECMO, 79 are still receiving ECMO support and for 10 patients status n.a. . Our preliminary data suggest that patients placed on ECMO with severe refractory respiratory or cardiac failure secondary to COVID-19 have a reasonable (55%) chance of survival. Further extensive data analysis is expected to provide invaluable information on the demographics, severity of illness, indications and different ECMO management strategies in these patients

Combined effects of solvation and aggregation propensity on the final supramolecular structures adopted by hydrophobic- glycine-rich, elastin-like polypeptides

Previous work on elastin-like polypeptides (ELPs) made of hydrophobic amino acids of the type XxxGlyGlyZzzGly (Xxx, Zzz = Val, Leu) has consistently shown that differing dominant supramolecular structures were formed when the suspending media were varied: helical, amyloid-like fibers when suspended in water and globules evolving into ‘string of bead’ structures, poly(ValGlyGlyValGly), or cigar-like bundles, poly (ValGlyGlyLeuGly), when suspended in methyl alcohol. Comparative experiments with poly (LeuGlyGlyValGly) have further indicated that the interface energy plays a significant role and that solvation effects act in concomitance with the intrinsic aggregation propensity of the repeat sequence. Continuing our investigation on ELPs using surface (XPS, AFM) and bulk (CD, FTIR) techniques for their characterization, here we have compared the effect of suspending solvents (H2O, DMSO, EG, MeOH) on poly(ValGlyGlyValGly), the polypeptide most inclined to form long and well refined helical fibers in water, searching for the signature of intermolecular interactions occurring between the polypeptide chains in the given suspension. The influence of sequence specificities has been studied by comparing poly(ValGlyGlyValGly) and poly(LeuGlyGlyValGly) with a similar degree of polymerization. Deposits on substrates of the polypeptides were characterized taking into account the differing evaporation rate of solvents, and tests on their stability in ultra high vacuum (UHV) were performed. Finally, combining experimental and computational studies, we have revaluated the 3D modelling previously proposed for the supramolecular assembly in water of poly(ValGlyGlyValGly). The results were discussed and rationalized also on the light of published data

Characterisation of helical structure in AFM micrographs of a trimer of the peptide sequence (ValGlyGlyValGly)

Studies by atomic force microscopy of a set of helical fibres formed within the same period of time from the amyloidogenic peptide, (VGGVG)3 show that there is a characteristic increment in diameter, suggesting that the helices are formed from multiple layers. Measurement of the height to pitch ratio suggests that each succeeding layer forms on top of the previous winding and does not fit into the gap between the turns. Dimensions are given for the increment per layer and for the pitch of the helices

Heparan Sulfate Affects Elastin Deposition in Fibroblasts Cultured from Donors of Different Ages

Heparan sulfate (HS), due to its presence on the cell surface and in the extracellular milieu and its ability to modulate cell signaling, has a fundamental role in both physiological and pathological conditions. For decades we have demonstrated the occurrence of interactions between glycosaminoglycans (GAGs) and elastic fibers. In particular, we have recently shown that HS is present inside elastic fibers and plays a role in the assembly and stability of elastin coacervates. Elastin represents, within the extracellular matrix, the component most severely affected during aging, and changes in the synthesis and posttranslational modifications of HS have been described, possibly influencing cellular behavior and protein interactions. Thus, the present study has investigated, in two different in vitro experimental models, the role of HS on elastin deposition and assembly. Results demonstrate that: (1) Biological effects of HS are partly dependent on the physicochemical characteristics of the GAGs(2) HS does not affect attachment, viability, and growth of human dermal fibroblasts(3) HS does not modify elastin gene expression nor elastin synthesis, but favors a-elastin aggregation and, independently from the age of donors, elastin assembly(4) HS significantly increases the expression of fibulin 5, and these effects are especially evident in fibroblasts isolated from aging donors. These data provide a better understanding of the biological role of HS and offer new perspectives regarding the possibility of restoring and/or preserving the elastic component with aging

Cardiopulmonary resuscitation in prone position: A scoping review

Introduction The ongoing pandemic of COVID-19 brought to the fore prone positioning as treatment for patients with acute respiratory failure. With the increasing number of patients in prone position, both spontaneously breathing and mechanically ventilated, cardiac arrest in this position is more likely to occur. This scoping review aimed to summarize the available evidence on cardiopulmonary resuscitation in prone position (‘reverse CPR’) and knowledge or research gaps to be further evaluated. The protocol of this scoping review was prospectively registered on 10th May 2020 in Open Science Framework (https://osf.io/nfuh9). Methods We searched PubMed, EMBASE, MEDLINE and pre-print repositories (bioRxiv and medRxiv) for simulation, pre-clinical and clinical studies on reverse CPR until 31st May 2020. Results We included 1 study on manikins, 31 case reports (29 during surgery requiring prone position) and 2 nonrandomized studies describing reverse CPR. No studies were found regarding reverse CPR in patients with COVID-19. Conclusions Even if the algorithms provided by the guidelines on basic and advanced life support remain valid in cardiac arrest in prone position, differences exist in the methods of performing CPR. There is no clear evidence of superiority in terms of effectiveness of reverse compared to supine CPR in patients with cardiac arrest occurring in prone position. The quality of evidence is low and knowledge gaps (e.g. protocols, training of healthcare personnel, devices for skill acquisition) should be fulfilled by further research. Meanwhile, a case-by-case evaluation of patient and setting characteristics should guide the decision on how to start CPR in such cases

Exome sequencing and bioinformatic approaches reveals rare sequence variants involved in cell signalling and elastic fibre homeostasis: new evidence in the development of ectopic calcification

Elastic fibres undergo aberrant mineralization in genetic as well as in acquired pathologic conditions causing severe impairment of tissue mechanical properties. Despite the number of investigations performed so far, the pathogenesis of these alterations is still elusive, due to both the complexity of the elastin network and the involvement of many genes and/or pro-osteogenic signalling pathways. Whole Exome Sequencing (WES) was performed on DNA from three patients affected by beta-thalassemia exhibiting soft connective tissue calcification. WES data were analysed with a bioinformatic approach, allowing to screen and to select genes carrying rare sequence variants. These genes were matched with those present in Extracellular Matrix DB. This approach enables to shed light on the involvement of the extracellular matrix in the occurrence of ectopic calcification. Results revealed a number of rare sequence variants in genes related to elastic fibre assembly and integrity. For instance, the involvement of fibrillins and collagen type VI in the formation of a modified microfibrillar scaffold may lead to elastic fibres less resilient and more prone to hydroxyapatite deposition. Moreover, data reveal that changes in mitochondrial metabolic pathways are sustained by a genetic background and emphasize that a persistent chronic oxidative stress can further influence extracellular matrix homeostasis and cell signalling through the TGFβ-BMP axis. Eventually, the presence of multiple rare sequence variants in the Solute Carrier Family 25 Member 5 (SLC25A5) gene is suggestive of the role of this gene as a key factor linking mitochondria metabolism, ADP/ATP ratio and oxidative stress thus affecting extracellular matrix homeostasis and activation of pro-osteogenic factors

Interactions between elastin-like peptides and an insulating poly(ortho-aminophenol) membrane investigated by AFM and XPS

This investigation was undertaken to explore the mutual recognition of the pentapeptide (ValGlyGlyValGly)n, a hydrophobic elastin-like peptide (ELP), suspended in deionized water in monomer (n = 1) and trimer (n = 3) forms and the outer surface of a very thin, insulating polymer, poly(ortho-aminophenol) (PoAP), electrochemically grown on a platinum foil by cyclic voltammetry in a neutral medium (phosphate-buffered saline, I = 0.1M) immersed in the suspension. As a prior task, the proved propensity of the ValGlyGlyValGly sequence, at the given minimal length (three or more repeats), to self-assemble into amyloid-like fibrils when solubilized in an aqueous environment was considered within the framework of testing PoAP surfaces for the specific detection of amyloid precursors. From our knowledge of the chemical structure and physical properties of both biomacromolecule families obtained in previous studies, we focused on the efficacy of the binding sites offered to ELP fibrils by PoAP in its as-prepared form or properly modified either by postsynthesis oxidation or by adsorption/entrapping of ELP monomer(s) with or without protecting terminal groups. Consistent with all methods of preparation, the best surfaces, recognizable by the trimer fibrils, are those modified to carry a larger number of carbonyls, particularly by entrapment of ELP monomer(s) during PoAP electrosynthesis using an imprinting-inspired method. The degree of attachment of fibrillar aggregates, detected by atomic force microscopy and X-ray photoelectron spectroscopy, provides unequivocal evidence of the cooperative forces involving PoAP–ELP interactions. The results obtained suggest the prospect of using the proposed Pt/PoAP/ELP systems as biodetectors in Alzheimer disease