Journal of extracellular biology最新文献

Vitronectin (VTN) is a potential non-invasive biomarker for renal fibrosis, originally described in urinary extracellular vesicles (uEV) from kidney transplant patients (KTx). However, VTN's specific role in renal fibrosis is unclear, as it is involved in various physiological processes. This study aims to identify other uEV-associated proteins linked to renal fibrosis to clarify which pathways involve VTN. uEV were isolated from 33 KTx patients and five healthy controls. uEV proteins were analysed using proximity extension assay (PEA), and data were normalized and compared using Welch's two-sided t-test to identify differentially expressed proteins between fibrotic (n = 31) and non-fibrotic patients (n = 7). Urinary VTN levels and monocyte chemoattractant protein-1 (MCP-1) were measured by ELISA. PEA analysis identified 33 proteins overexpressed in the fibrotic group. These proteins clustered in STRING analysis, primarily associating with coagulation, fibrinolysis and TNF-inflammation involving macrophages. ELISA detection of MCP-1 further validated the results. High levels of VTN in the fibrotic group were accompanied by the upregulation of fibrinolytic pathway components (PAI-1, tPA and uPAR), which are well-known to interact with VTN. This study highlights TNF-induced inflammation involving macrophages and fibrinolysis as key mechanisms underlying renal fibrosis with direct implications of VTN, which support VTN's potential as a biomarker for this pathological process.

Fasciolosis, caused by Fasciola hepatica, is a parasitic zoonosis that induces liver fibrosis in infected hosts, including ruminants and humans. Extracellular vesicles secreted by F. hepatica (FhEVs) play a crucial role in modulating host immune responses and promoting tissue re-modelling. This work explores the effects of two proteins found in FhEVs, enolase (Fhenolase), enriched in the vesicular lumen, as well as the 16.5-kDa tegument-associated protein (Fh16.5TP), highly abundant in the EV membrane, on hepatic and liver-associated immune cells. Recombinant proteins (r-Fhenolase and r-Fh16.5TP) were produced to evaluate their impact on cell viability, inflammatory responses, proteomic profiles and EV secretion in THP1-XBlue CD14 macrophages, HepG2 hepatocytes and LX-2 hepatic stellate cells (HSCs). Interestingly, r-Fhenolase, but not r-Fh16.5TP, showed anti-inflammatory properties in lipopolysaccharide (LPS)–activated macrophages, by reducing NF-κB activation and inducing significant changes in the protein cargo of macrophage-derived EVs, which contained lower levels of the pro-inflammatory cytokines IL-1β, TNF-α and IL-6. Proteomic analysis of cells treated with r-Fhenolase revealed distinct alterations in proteins related to fibrotic and inflammatory pathways, including a reduction in extracellular matrix (ECM) proteins and suggesting a potential role in mitigating liver fibrosis. Furthermore, r-Fhenolase reduced EV production and fibrotic markers in hepatic cells, but not in macrophages. In contrast, r-Fh16.5TP increased pro-fibrotic proteins in both, cells and EVs, and increased EV production specifically in LX-2 cells, indicating its possible contribution to fibrosis progression in fasciolosis. These findings represent a first approach to analyse EV-associated proteins and study their potential role in the molecular mechanisms of F. hepatica–host interactions.

Cancer continues to be the foremost cause of mortality in humans. Persistent challenges in cancer treatment include inadequate drug targeting, severe toxicological side effects and uncontrolled drug distribution. The bioinspired membrane vesicle drug delivery systems have been emerging as promising therapeutic strategies. This study characterises unique cell-bound membrane vesicles (CBMVs), which are impervious to standard cleaning agents and effectively loaded with doxorubicin (DOX). For the first time, we used iRGD peptide to modify the CBMVs to enhance the CBMVs' targeting capabilities for cancer cells. Laser confocal microscopy and ¹H Nuclear Magnetic Resonance Spectra (¹H NMR) have confirmed the CBMVs' iRGD modification and effective encapsulation with DOX (iRGD-CBMVs-DOX). Then, we used the iRGD-CBMVs-DOX to treat tumour cell lines and tumour-bearing mouse models. Our research identified that iRGD-CBMVs-DOX proves effective in inhibiting cell growth and migration for tumour cell lines, significant anti-tumour ability, reduced organ toxicity and continuous drug administration were revealed in tumour-bearing mouse models. Additionally, the iRGD-CBMVs-DOX demonstrated sustained drug release, indicating their potential for prolonged circulation. These findings are pivotal in enhancing cancer treatment through novel nanomedicine strategies, and highlight the potential of iRGD-modified vesicles (e.g., iRGD-CBMVs) as efficient drug carriers, contributing to targeted and biocompatible drug delivery advancements for cancer treatment.

R. Franko, and M. de Almeida Monteiro Melo Ferraz, “Exploring the Potential of In vitro Extracellular Vesicle Generation in Reproductive Biology,” Journal of Extracellular Biology 3 (2024): e70007. https://doi.org/10.1002/jex2.70007

In the originally published article, the in-text citation for the reference ‘Montecalvo et al., 2008’ was given incorrectly as ‘Montecalvo et al., 1950’. This has been corrected in the online version of the article.

We apologise for this error.

M. L. Zenner, B. Kirkpatrick, T. R. Leonardo, et al., “Prostate-Derived Circulating microRNAs Add Prognostic Value to Prostate Cancer Risk Calculators,” Journal of Extracellular Biology 2 (2023): e122, https://doi.org/10.1002/jex2.122.

In the originally published article, the in-text citation for the reference ‘Cancer Statistics, 2023 (2023)’ was given incorrectly as ‘Cancer Facts & Figures 2023, 1930’. This has been corrected in the online version of the article.

We apologise for this error.

As extracellular vesicles (EVs) are increasingly recognized for their superior functions for therapeutics, the need for large-scale EV isolation technology is becoming more critical for clinical and industrial applications. Most existing EV isolation methods are optimized for small-scale laboratory samples, limiting their efficiency and scalability for large-scale production. Here, an electrokinetic-assisted filtration system (ExoFilter), which introduces charge interaction into physical mesh flow filtration, is proposed as a new candidate to address the challenges of scalable EV isolation. The hybrid filtration system demonstrates outstanding high-throughput EV isolation performance (a flux of ∼750 mL/min) using only a coarse physical filter by electrokinetically arresting EVs flowing through the filter lattice. Furthermore, the recovery efficiency of ExoFilter, analysed based on the ELISA results, was found to be approximately 98%, demonstrating the filter's exceptional efficiency in EV isolation. Additionally, ExoFilter enables the rapid isolation of EVs from small samples as little as 200 µL, facilitating quick and easy blood-based EV research. Furthermore, low-molecular-weight albumin from plasma samples was effectively removed. The high-throughput and high-efficiency characteristics of ExoFilter make it well-suited for scalable EV production, offering greater convenience for various clinical applications.

Despite biomolecule delivery is a natural function of extracellular vesicles (EVs), low loading of exogenous macromolecules such as proteins into EVs limits their interest as convincing protein delivery systems for health applications. In this context, lipid-anchorage of exogenous cargo into EV membrane recently emerged as a promising option to enable their vectorisation into cells. Nevertheless, this option was not explored for protein intracellular delivery, and further characterisation of critical parameters governing the association of a lipid-anchored cargo protein to EVs is still needed to confirm the relevance of this anchorage strategy. Therefore, we sought to identify these parameters in a precise and quantitative manner, using bulk and single nanoparticle analysis methods to identify protein loading capacity and subsequent intracellular delivery. We identified incubation temperature, cargo concentration, lipid anchor (LA) structure (lipid nature, linker) and EV origin as critical factors influencing maximal EV loading capacity. Precise control of these parameters enabled to load cargo protein close to EV saturation without hindering cellular delivery. The structural properties of LA influenced not only cargo protein/EV association but also intracellular delivery into different carcinoma cell lines. By thoroughly characterising Lipid-PEG-protein anchorage, this study evidences the interest of this tunable and controllable approach for efficient EV protein delivery.

Nanoflow cytometry (nanoFCM) is an increasingly important analytical procedure in every aspect of extracellular vesicle (EV) research, particularly in the development of EV-based therapeutics. The main objective of this study was to evaluate and optimise the key determinant factors of nanoFCM in the quantification analysis of EVs to ensure its consistency and reliability in the development of EV therapeutic drugs, thereby serving as a potential quality control measure. Our investigation followed the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) Q14 guideline. We revisited the day-to-day practice of nanoFCM measurement for HEK293 cell-derived and milk-derived EVs (mEVs), focusing on optimising particle quantification and identifying risk factors. Initial evaluation of the procedure revealed a considerable lack of consistency and reliability, which was then subjected to extensive optimisation. The key outcomes of this study include: (1) an optimised analytic procedure incorporating Tween-20, which significantly enhanced the precision and accuracy of the nanoFCM measurement and expanded the reportable range; (2) an analytical target profile (ATP) which provides a preliminary standard for future validation of nanoFCM procedures. Overall, this study serves as a foundation for future efforts towards the standardisation of analytical procedures for EV therapeutics.