51 research outputs found
A Substrate-Mimicking Basement Membrane Drives the Organization of Human Mesenchymal Stromal Cells and Endothelial Cells Into Perivascular Niche-Like Structures
Extracellular matrix-derived products (e.g. Matrigel) are widely used for in vitro cell cultures both as two-dimensional (2D) substrates and as three-dimensional (3D) encapsulation gels because of their ability to control cell phenotypes through biospecific cues. However, batch-to-batch variations, poor stability, cumbersome handling, and the relatively high costs strictly limit their use. Recently, a new substrate known as PhenoDrive-Y has been used as 2D coating of tissue culture plastic showing to direct the bone marrow mesenchymal stromal cells (MSCs) toward the formation of 3D spheroids. When organized into 3D spheroids, the MSCs expressed levels of pluripotency markers and of paracrine angiogenic activity higher than those of the MSCs adhering as fibroblast-like colonies on tissue culture plastic. The formation of the spheroids was attributed to the properties of this biomaterial that resemble the main features of the basement membrane by mimicking the mesh structure of collagen IV and by presenting the cells with orderly spaced laminin bioligands. In this study, PhenoDrive-Y was compared to Matrigel for its ability to drive the formation of perivascular stem cell niche-like structures in 2D co-culture conditions of human endothelial cells and adult bone marrow MSCs. Morphological analyses demonstrated that, when compared to Matrigel, PhenoDrive-Y led endothelial cells to sprout into a more consolidated tubular network and that the MSCs nestled as compact spheroids above the anastomotic areas of this network resemble more closely the histological features of the perivascular stem cell niche. A study of the expressions of relevant markers led to the identification of the pathways linking the PhenoDrive-Y biomimicking properties to the acquired histological features, demonstrating the enhanced levels of stemness, renewal potential, predisposition to migration, and paracrine activities of the MSCs
The Real-Time Validation of the Effectiveness of Third-Generation Hyperbranched Poly(ɛ-lysine) Dendrons-Modified KLVFF Sequences to Bind Amyloid-β1-42 Peptides Using an Optical Waveguide Light-Mode Spectroscopy System
The aggregation of cytotoxic amyloid peptides (Aβ(1-42)) is widely recognised as the cause of brain tissue degeneration in Alzheimer’s disease (AD). Indeed, evidence indicates that the deposition of cytotoxic Aβ(1-42) plaques formed through the gradual aggregation of Aβ(1-42) monomers into fibrils determines the onset of AD. Thus, distinct Aβ(1-42) inhibitors have been developed, and only recently, the use of short linear peptides has shown promising results by either preventing or reversing the process of Aβ(1-42) aggregation. Among them, the KLVFF peptide sequence, which interacts with the hydrophobic region of Aβ(16-20), has received widespread attention due to its ability to inhibit fibril formation of full-length Aβ(1-42). In this study, hyperbranched poly-L-lysine dendrons presenting sixteen KLVFF at their uppermost molecular branches were designed with the aim of providing the KLVFF sequence with a molecular scaffold able to increase its stability and of improving Aβ(1-42) fibril formation inhibitory effect. These high-purity branched KLVFF were used to functionalise the surface of the metal oxide chip of the optical waveguide lightmode spectroscopy sensor showing the more specific, accurate and rapid measurement of Aβ(1-42) than that detected by linear KLVFF peptides
A Comparative In VitroStudy of the Effect of Biospecific Integrin Recognition Processes and Substrate Nanostructure on Stem Cell 3D Spheroid Formation
Development of scaffold-free vascularized pancreatic beta-islets in vitro models by the anchoring of cell lines to a bioligand-functionalized gelatine substrate
Bioengineered pancreatic β-islets have been widely advocated for the research and treatment of diabetes by offering both suitable cell culture models for the study of the pathology and the testing of new drugs and a therapy in those patients no longer responding to insulin administration and as an alternative to the shortage of donors for organ and islet transplantation. Unlike most of the studies published so far where pancreatic islets of pancreatic β-cells are encapsulated in hydrogels, this study demonstrate the formation of bioengineered pancreatic islets through cell anchoring to a gelatine-based biomaterial, PhenoDrive-Y, able to mimic the basement membrane of tissues. Through simple culture conditions, PhenoDrive-Y led human pancreatic β-cell lines and human umbilical endothelial cell lines to form organized structures closely resembling the natural vascularized pancreatic islets. When compared to gelatine, the cultures in presence of PhenoDrive-Y show higher degree of organization in tissue-like structures, a more pronounced endothelial sprouting and higher expression of typical cell markers. Noticeably, when challenged by hyperglycaemic conditions, the cells embedded in the PhenoDrive-Y assembled spheroids responded with higher levels of insulin production. In conclusion, the present work demonstrates the potential of PhenoDrive-Y as substrate for the development of bioengineered vascularized pancreatic islets and to be particularly suitable as a model for in vitro studies and testing of new therapeutics. [Figure: see text
Insights on Lulworthiales Inhabiting the Mediterranean Sea and Description of Three Novel Species of the Genus Paralulworthia
The order Lulworthiales, with its sole family Lulworthiaceae, consists of strictly marine genera found on a wide range of substrates such as seagrasses, seaweeds, and seafoam. Twenty-one unidentified Lulworthiales were isolated in previous surveys aimed at broadening our understanding of the biodiversity hosted in the Mediterranean Sea. Here, these organisms, mostly found in association with Posidonia oceanica and with submerged woods, were examined using thorough multi-locus phylogenetic analyses and morphological observations. Maximum-likelihood and Bayesian phylogeny based on nrITS, nrSSU, nrLSU, and four protein-coding genes led to the introduction of three novel species of the genus Paralulworthia: P. candida, P. elbensis, and P. mediterranea. Once again, the marine environment is a confirmed huge reservoir of novel fungal lineages with an under-investigated biotechnological potential waiting to be explored
L'infrastruttura di ricerca MIRRI (Microbial Resource Reserch Infrastructure): il network delle collezioni di microrganismi per lo sviluppo sostenibile in Europa
CD81 controls sustained T cell activation signaling and defines the maturation stages of cognate immunological synapses
In this study, we investigated the dynamics of the molecular interactions of tetraspanin CD81 in T lymphocytes, and we show that CD81 controls the organization of the immune synapse (IS) and T cell activation. Using quantitative microscopy, including fluorescence recovery after photobleaching (FRAP), phasor fluorescence lifetime imaging microscopy-Föster resonance energy transfer (phasorFLIM-FRET), and total internal reflection fluorescence microscopy (TIRFM), we demonstrate that CD81 interacts with ICAM-1 and CD3 during conjugation between T cells and antigen-presenting cells (APCs). CD81 and ICAM-1 exhibit distinct mobilities in central and peripheral areas of early and late T cell-APC contacts. Moreover, CD81-ICAM-1 and CD81- CD3 dynamic interactions increase over the time course of IS formation, as these molecules redistribute throughout the contact area. Therefore, CD81 associations unexpectedly define novel sequential steps of IS maturation. Our results indicate that CD81 controls the temporal progression of the IS and the permanence of CD3 in the membrane contact area, contributing to sustained T cell receptor (TCR)-CD3-mediated signaling. Accordingly, we find that CD81 is required for proper T cell activation, regulating CD3ζ, ZAP-70, LAT, and extracellular signal-regulated kinase (ERK) phosphorylation; CD69 surface expression; and interleukin- 2 (IL-2) secretion. Our data demonstrate the important role of CD81 in the molecular organization and dynamics of the IS architecture that sets the signaling threshold in T cell activationThis work was supported by SAF2011-25834 from the Spanish Ministry of
Science and Innovation, INDISNET-S2011/BMD-2332 from the Comunidad
de Madrid, Cardiovascular Network RD12-0042-0056 from the Instituto
Salud Carlos III, and ERC-2011-AdG 294340-GENTRI
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