Journal of Extracellular Vesicles最新文献

Extracellular vesicles (EVs) are key mediators of intercellular communication, carrying diverse molecular cargo that reflects the dynamic physiological and pathological state of their source cell. While analyses of the entire vesicular population (bulk EV) have advanced our understanding of their roles in health and disease, these approaches often obscure the heterogeneity inherent in EV populations. Emerging single-vesicle analysis technologies offer unprecedented resolution, enabling the identification of individual EV subpopulations and their distinct molecular signatures. Such approaches, combined with digital platforms, can now analyze individual molecules from single EVs, including single-molecule features such as protein, mRNA, double-stranded DNA and single-stranded DNA. This perspective explores the transformative potential of single EV technologies in clinical diagnostics and therapeutic applications. We highlight key advancements including microfluidic platforms, super-resolution microscopy and AI-driven data analyses, that are shaping and advancing the field and its applications. With the development and advancement of clinically viable single EV technologies, we are beginning to appreciate the complexity and abundance of cell type and specific EVs. We further discussed the challenges of sensitivity, specificity, standardization and scalability hindering these technologies' broad acceptance and feasibility in clinical translation. This perspective paper originates from discussions at the Chinese Society of Extracellular Vesicles (CSEV) annual meeting, held in Guangzhou, China, on 16 November 2024. At this meeting, researchers from various fields of EV research, with a particular emphasis on single EV digital, analytical and quantitative technological platforms, discussed the opportunities and challenges of this emerging single-EV-focused technology. The paper aims to provide a roadmap for integrating single EV technologies into routine EV-research and even clinical practice, paving the way for novel scientific and diagnostic tools, personalized therapies, and a deeper understanding of EV heterogeneity and EV biology.

Reliable non-invasive biomarkers for early detection of diabetic nephropathy (DN), a leading cause of chronic kidney disease, remain limited. In this study, we isolate urinary extracellular vesicles (uEVs) using wheat germ agglutinin (WGA)-conjugated magnetic beads and identify cytoskeleton-associated protein 4 (CKAP4) as a potential diagnostic biomarker for DN. Proteomic profiling and flow cytometry show that CKAP4 levels are significantly higher in uEVs from DN patients than in those from diabetic, non-diabetic renal disease (NDRD) and healthy control groups. Receiver operating characteristic (ROC) analysis demonstrates excellent diagnostic performance, with area under the curve (AUC) values of 0.9998 (sensitivity = 98.77%, specificity = 100%) for DN versus controls, and 0.9859 (sensitivity = 95.72%, specificity = 99.24%) for DN versus diabetes mellitus. CKAP4 levels, elevated even at early-stage DN, positively correlate with glomerulosclerosis, increasing with the severity of interstitial fibrosis and tubular atrophy (IFTA). Mechanistically, CKAP4-containing EVs derived from high glucose-treated podocytes promote vascular calcification in vascular smooth muscle cells via YAP signalling. These findings identify CKAP4 in podocyte-derived uEVs as a robust non-invasive biomarker for early DN detection and provide new insights into the vascular pathology associated with the disease.

Subunit vaccines are promising for disease prevention because of their safety and cost-effectiveness. However, their efficacy is limited by low immunogenicity and gastrointestinal degradation after oral administration. To address this issue, low-endotoxin Salmonella choleraesuis strain SC-L3 was engineered via lipid A modification to generate bacterial biomimetic vesicles (BBVs) with reduced endotoxin activity. BBVs were functionalized using ClyA-embedded SpyCatcher and Streptococcus protein G for dual antigen coupling, and further coated with chitosan oligosaccharides (COS) to enhance mucosal penetration and gastrointestinal stability. Using mCherry as a model antigen, we obtained optimized mCherry-CSS-BBV@COS that showed high antigen protection rates (83% and 63% in simulated gastric and intestinal fluids, respectively), capacity for lysosomal escape and effective stimulation of M1 macrophage polarization in vitro. Oral administration of mCherry-CSS-BBV@COS elicited robust systemic IgG and mucosal sIgA responses in mice. Furthermore, dual-antigen BBV conjugates (GDH-gD-Fc-CSS-BBV@COS) co-delivering Streptococcus suis glutamate dehydrogenase and pseudorabies virus gD-Fc induced antigen-specific humoral, mucosal and cellular immunity, conferring complete protection against lethal challenges with the respective pathogens. In summary, we generated a versatile, low-endotoxin BBV platform for oral combination subunit vaccines, offering a novel strategy for protection against viral and bacterial infections.

Cannabis sativa is a medicinal plant that produces a diverse array of pharmacologically active metabolites, making it a valuable resource for pharmaceutical applications. In this study, an adventitious root (AR) culture system was established from C. sativa using two representative plant growth regulators—naphthaleneacetic acid (NAA; hereafter referred to as N-ARs) and indole-3-butyric acid (IBA; hereafter referred to as I-ARs) —from which plant-derived nanovesicles (PDNVs) were subsequently isolated (hereafter N-PDNVs and I-PDNVs, respectively). The resulting N-PDNVs and I-PDNVs exhibited average diameters of 128 ± 2 and 124 ± 4 nm, respectively, with zeta potentials of −12.9 and −15.7 mV. Both PDNV types maintained structural integrity and colloidal stability under diverse external stress conditions, underscoring their physicochemical robustness. Metabolite profiling of PDNVs revealed 25 distinct metabolites. Functionally, I-PDNVs markedly enhanced dendritic cell maturation through Toll-like receptor 2 (TLR2)- and TLR4-dependent pathways, promoted T cell proliferation and activation (notably IFN-γ- and IL-17A-producing subsets), and increased natural killer (NK) cell activity compared with N-PDNVs. In immunosuppressed and tumour-bearing mouse models, I-PDNVs further augmented NK cell, Th1 and cytotoxic T lymphocyte (CTL) responses, thereby confirming their superior potential as immunotherapeutic agents. Moreover, in immunized mouse models, OVA_257-264-encapsulated I-PDNVs demonstrated a clear advantage as a vaccine delivery platform by eliciting a potent OVA_257-264-specific CTL response. When applied as a prophylactic cancer vaccine, they not only delayed tumour growth but also reshaped the antitumour immune landscape, characterized by enhanced CTL responses, reduced regulatory T cell frequencies and diminished exhausted CD8⁺ T cell populations. Collectively, these findings highlight the potential of I-PDNVs as dual-function PDNVs, serving both as immunotherapeutic agents and as vaccine delivery platforms for applications requiring reinforced Th1, CTL and NK cell responses.

Metastasis is the leading cause of death related to breast cancer. Premetastatic niches (PMNs), which are remodelled by the primary tumours in distant organs, are essential for the colonisation of disseminated cancer cells. The vascular niche is among the most pivotal PMNs in breast cancer lung metastasis, and the underlying mechanism remains unclear. Here, we report that breast cancer cells secrete dipeptidyl peptidase 3 (DPP3) via small extracellular vesicles (sEVs) to promote lung metastasis. Mechanistically, circulating DPP3 upregulates RAPGEF4 to activate the Rap1 signalling pathway in the lung endothelial cells through the DPP3–PFKP–YBX1 axis and promotes angiogenesis to remodel the vascular niche, thereby increasing lung metastasis. In addition, ARF4 recognises ISGylated DPP3, which facilitates its packaging into sEVs in breast cancer cells. Finally, treatment with losartan pharmacologically inhibits the ISGylation of DPP3, preventing its secretion via sEVs. In summary, our findings demonstrate that DPP3, which is encapsulated in sEVs and secreted by breast cancer cells, regulates angiogenesis in the lung and remodels vascular niches to promote breast cancer lung metastasis, making it a potential target for the diagnosis and treatment of breast cancer metastasis.

During fibrogenesis, certain negative feedback loops are elicited to restrain persistent and hyperactive fibrotic responses. Activated fibroblasts have been found to acquire anti-fibrotic phenotypes. However, the specific inhibitory modulators remain largely enigmatic. Thus, the present study aimed to examine the intrinsic autoregulatory mechanisms of fibroblasts. Here, we demonstrated that angiotensin II (AngII)-primed cardiac myofibroblast moderated subsequent profibrotic activation. More importantly, this suppressive action was dependent on small extracellular vesicles (sEVs). Strikingly, small RNA sequencing identified an abundant presence of Piwi-interacting RNAs (piRNAs) in sEVs. In cultured primary cardiac fibroblasts, piRNA-62788 was induced by AngII receptor type 2 (AT₂R) stimulation and encapsulated into sEVs. Furthermore, fibrogenic responses were attenuated by piRNA-62788 overexpression, whereas aggravated by piRNA-62788 knockdown. In a mouse model of transverse aortic constriction, either piRNA-62788 agomir or circulating sEVs of patients with heart failure (HF) mitigated adverse cardiac remodelling, while piRNA-62788 inhibitor-containing sEVs accentuated myocardial fibrosis. Mechanistically, piRNA-62788 formed a functional complex with PIWI-like protein 2 (PIWIL2) and bound to the 3’ untranslated region (UTR) region of serum response factor (Srf) mRNA transcripts, leading to inhibition of the SRF signalling. Additionally, plasma sEV-derived piRNA-62788 was significantly upregulated in HF patients and negatively correlated with left ventricular ejection fraction. Collectively, we uncovered a protective negative feedback circuit controlled by AngII/AT₂R/sEVs axis. Understanding this endogenous anti-fibrotic pathway may hold therapeutic promise in HF.

Stem cell niches are complex multi-signalling networks comprised of molecular cues and physical interactions, orchestrated by niche-resident cells and the extracellular factors they produce. The bone niche specifically houses haematopoietic stem cells (HSCs), a critical cell type responsible for producing all blood and immune cells throughout life. Currently, how niches facilitate an ideal environment with simultaneously coordinating both intrinsic and extrinsic cellular signals is unknown. Studies presented here identify the existence of unique extracellular vesicle (EV)-defined niches within the haematopoietic system of human individuals. Bridging studies using proteomic signatures, nanoparticle characterization at single-vesicle resolution and machine learning-based techniques reveal that EVs can be grouped by blood, bone marrow and trabeculae within a human individual. Stem cell assays demonstrate that these niche-defined EVs impart functional effects on stem cells/progenitors based on location within the haematopoietic system. Finally, using single-cell transcriptomic analyses, results identify for the first time how niche-sourced EVs differentially affect the most primitive human HSCs and progenitors. This study highlights the significance of nanoparticles on human immunity and blood production and provides evidence for a new role for EVs, namely the demarcation of distinct nano-niches within biological systems.

Extracellular Vesicles (EVs), nanoscale membrane-bound cell-released structures, are vital for intercellular communication and material transport. Their role in musculoskeletal health and diseases has recently drawn significant attention. This review focuses on the latest EV research in musculoskeletal diseases, including their roles in disease progression and potential as biomarkers and therapies. Musculoskeletal disorders are the third-leading cause of global disability-adjusted life-years among adolescents and young adults. Current treatments face issues like limited tissue regeneration and poor drug targeting. With their natural messenger function and low immunogenicity, EVs have become a research focus. However, their action mechanisms in the musculoskeletal system remain un-systematically understood. This paper reviews EVs’ role in musculoskeletal diseases. It covers classification, biogenesis, release, internalization, cargo and their involvement in muscle cell processes, joint diseases, bone metabolism and disc degeneration. It also explores EVs’ role in musculoskeletal crosstalk and their potential as therapeutic agents and drug carriers through engineering with biomaterials. Future research should delve deeper into EV action mechanisms for better treatments. Overall, while EVs offer new treatment strategies for musculoskeletal diseases, more research is needed to overcome technical and clinical barriers.

For the past decade, the field of extracellular vesicle (EV) research has vastly expanded (Bazzan et al. 2021). Our knowledge of EV biogenesis, their various roles in intercellular communication, and significant contributions to disease initiation and regulation has quickly developed (Möller and Lobb 2020, Kalluri and LeBleu 2020). The burgeoning of EV research is notable in the rapid parallel advancement of appropriate, cutting-edge isolation and analysis techniques (Qiu et al. 2023, De Sousa et al. 2023). Whilst there is a consensus amongst the field on certain aspects of EV function and biology, one crucial, trending topic of controversy is the significance of EV nomenclature.

Accurate and appropriate naming is paramount within and outside the scientific community for several reasons. Firstly, researchers must be able to refer to the same entity properly to ensure that the same phenomenon is evaluated in their specific fields, as well as across fields, to allow legitimate comparisons and improvement. Iterative publications of MISEV guidelines and similar initiatives over the years have enabled significant progress (Welsh et al. 2024). Within the past decade, researchers used over 25 names for EVs, often describing the same entity across different diseases or source materials, with different labels (Choi et al. 2015). Precise designation of EVs is not only a matter of scientific uniformity, but also of scientific integrity. If research groups isolate the same EV but use different terms for it, determining whether the findings are providing deeper insights into a certain topic or contradicting each other can be difficult. In 2023, an average of ∼170 publications per week were deposited with PubMed in the EV field, increasing to about 195 in 2024, a trend that is likely to continue. Thus, it is possible that repeated work has been published without cross-referencing, and that this happens not by ignorance or deceit, but by a difficulty of finding related studies due to discrepancies in EV naming. The nomenclature of EVs was clearly and unambiguously defined in MISEV2018 (Théry et al. 2018) and more recently in MISEV2023 (Welsh et al. 2024). The MISEV guidelines, not only for researchers in the EV field but across disciplines, provide a unified and comparable platform.

Using accurate and specific EV terms is also important for early-career researchers and those new to the field. As numerous publications show, EVs obtained by current experimental methods are not EVs generated by a single mechanism, but rather a mixture of EVs secreted by multiple mechanisms (Di Bella 2022). Hence, a precise understanding of the EV source material used helps to design appropriate experiments and conduct accurate research. MISEV2023 provides an appropriate framework by defining the terms for EV subpopulations and the require