Biology subjects

Zendrini, A.

Publications and source records attributed to Zendrini, A..

4 recordsLinked to original sources

bioRxiv2023

Addressing heterogeneity in direct analysis of Extracellular Vesicles and analogues using Membrane-Sensing Peptides as Pan-Affinity Probes

Extracellular vesicles (EVs), crucial mediators of cell-to-cell communication, hold immense potential for diagnostic applications due to their ability to enrich protein biomarkers in body fluids. However, challenges in isolating EVs from complex biological specimens hinder their widespread use. In this frame, integrated isolation-and-analysis workflows are the go-to strategy, most of which see the prevalence of immunoaffinity methods. Yet, the high heterogeneity of EVs poses challenges, as proposed ubiquitous markers are less homogenously prevalent than believed, raising concerns about the reliability of downstream biomarker discovery programs. This issue extends to the burgeoning field of engineered EV-mimetics and bio-nanoparticles, where conventional immune-affinity methods may lack applicability. Addressing these challenges, we introduce the use Membrane Sensing Peptides (MSP) as "universal" affinity ligands for both EVs and EV-analogues. Employing a streamlined process integrating on-bead capture and vesicle phenotyping through Single Molecule Array (SiMoA) technology, we showcase the application of MSP ligands in the integrated analysis of circulating EVs in blood derivatives, eliminating the need for prior EV isolation. Demonstrating the possible clinical translation of MSP technology, we directly detect an EV-associated epitope signature in serum and plasma samples, demonstrating its potential for distinguishing patients with myocardial infarction versus stable angina. At last, notably, MSP exhibits a unique capability to enable the analysis of tetraspanin-lacking Red Blood Cell derived EVs (RBC-EVs). Overall, unlike traditional antibody-based methods, MSP probes work agnostically, overcoming limitations associated with surface protein abundance or scarcity. This highlights the potential of MSP in advancing EV analysis for clinical diagnostics and beyond. Of note, this represents also the first-ever peptide-based application in SiMoA technology.

biochemistry↗

bioRxiv2023

A decision-making tool to navigate through Extracellular Vesicle research and product development

Due to their intercellular communication properties and their involvement in a wide range of biological processes, extracellular vesicles (EVs) are increasingly being studied and exploited for different applications. Nevertheless, their complex nature and heterogeneity, as well as the challenges related to their isolation, purification and characterization procedures, require cautious assessment of the quality and quantity parameters to monitor. This translates into a multitude of choices and putative solutions that lie in front of any EV researcher, in both research and translational environments, resembling a labyrinth with multiple paths to cross and, possibly, more than one exit. In this respect, decision-making tools might represent our modern Ariadnes string to follow not to get lost or distracted along the journey, to choose the shorter and best-fit-to-source EV application(s) and vice versa. Here, we present the implementation of a multi-criteria EV decision-making grid (EV-DMG) as a novel, customizable, efficient and easy-to-use tool to support responsible EV research and innovation. By identifying and weighting key assessment criteria for comparing distinct EV-based preparations and/or processes, our EV-DMG may assist any EV community member in making informed, transparent and reproducible decisions regarding the EV sources and/or samples to be managed, as well as the most suitable production and/or analytical pipelines to be adopted for targeting a defined aim or application.

cell biology↗

bioRxiv2022

Compositional profiling of extracellular vesicles/lipoproteins mixtures by AFM nanomechanical imaging

The widely overlapping physicochemical properties of lipoproteins (LPs) and extracellular vesicles (EVs) represents one of the main obstacles for the isolation and characterization of these pervasive biogenic lipid nanoparticles. We herein present the application of an atomic force microscopy (AFM)-based quantitative morphometry assay to the rapid nanomechanical screening of mixed LPs and EVs samples. The method can determine the diameter and the mechanical stiffness of hundreds of individual nanometric objects within few hours. The obtained diameters are in quantitative accord with those measured via cryo-electron microscopy (cryo-EM); the assignment of a specific nanomechanical readout to each object enables the simultaneous discrimination of co-isolated EVs and LPs even if they have overlapping size distributions. EVs and all classes of LPs are shown to be characterized by specific combinations of diameter and stiffness, thus making it possible to estimate their relative abundance in EV/LP mixed samples in terms of stoichiometric ratio, surface area and volume. As a side finding, we show how the mechanical behaviour of specific LP classes is correlated to distinctive structural features revealed by cryo-EM. To the best of our knowledge, these results represent the first systematic single-particle mechanical investigation of lipoproteins. The described approach is label-free, single-step and relatively quick to perform. Importantly, it can be used to analyze samples which prove very challenging to assess with several established techniques due to ensemble-averaging, low sensibility to small particles, or both, thus providing a very useful tool for quickly assessing the purity of EV/LP isolates including plasma- and serum-derived preparations.

biophysics↗

bioRxiv2021

Antithrombin glycoforms from type II antithrombin deficient patients selectively adsorb onto the surface of plasma extracellular vesicles

Antithrombin (AT) is a glycoprotein produced by the liver and a principal antagonist of active clotting proteases. A deficit in AT function leads to AT qualitative deficiency, challenging to diagnose. Here we report that active AT may travel physiosorbed on the surface of plasma extracellular vesicles (EVs), contributing to form the "EV-protein corona". The corona is enriched in specific AT glycoforms, thus suggesting glycosylation to play a key role in AT partitioning between EVs and plasma. Differences in AT glycoform composition of the corona of EVs separated from plasma of healthy and AT qualitative deficiency-affected subjects were also noticed. This suggests deconstructing the plasma into its nanostructured components, as extracellular vesicles, could help to unravel pathophysiological mechanisms otherwise undiscovered.

biochemistry↗