Journal of Extracellular Vesicles最新文献

The seminal work by Shi et al. unveils a novel cytoprotective mechanism in intervertebral disc degeneration, whereby USP5-rich apoptotic extracellular vesicles (ApoEVs) stabilize the transcription factor E2F1 to suppress apoptosis and enhance DNA repair in nucleus pulposus cells. While this study elegantly reframes the reparative potential of ApoEVs, its conceptual and translational premises invite critical scrutiny. This letter highlights several overlooked challenges. We argue that the contextual duality of E2F1—a well-established regulator of both cell survival and senescence—poses a considerable therapeutic hazard, as sustained E2F1 stabilization may inadvertently trigger a senescent phenotype. Moreover, the generalizability of this mechanism remains uncertain: ApoEV cargo and functionality are highly dependent on the apoptotic stimulus, and vesicles derived from etoposide-induced apoptosis may not mirror those produced under pathophysiological disc conditions. Crucially, the proposed pathway—from vesicle internalization and endosomal escape of luminal USP5 to its nuclear trafficking and engagement with E2F1—represents a mechanistic black box requiring rigorous validation. Resolving these issues is essential to translating this compelling discovery into a safe and effective therapeutic strategy.

Extracellular vesicles (EVs) are nanosized particles secreted by all cell types. As EVs are naturally occurring carriers of biological cargo, they serve as a promising candidate for drug delivery applications. Potential advantages of EVs as drug delivery systems include biological stability, intrinsic targeting properties and ability to overcome natural barriers. However, limitations such as cumbersome production and isolation procedures, batch-to-batch variability, and challenges related to efficient cargo loading limit their potential for clinical applications. Here, we introduce EV mimetics, prepared by incorporating full-length membrane proteins in the lipid bilayer of liposomes, using cell-free protein synthesis. These structures mimic functional characteristics of EVs, while offering advantages in terms of ease of manufacture, controllability and potential for efficient cargo loading. To demonstrate the feasibility of producing proteoliposomes as EV mimetics, we selected EV-associated CD47, CD39 and N-Cadherin as model proteins. We show successful production and purification of CD47, CD39 and N-Cadherin containing EV mimetics. Additionally, for CD47, we show that reaction conditions can be tailored to enhance EV mimetic yield. Furthermore, proteinase K protection assays and immuno-labelling electron microscopy revealed that correct membrane protein topology is preserved for CD47 and CD39. N-Cadherin EV mimetics show enhanced uptake by N-Cadherin-expressing MDA-MB-231 cells, proving membrane protein functionality is preserved. We demonstrate the versatility of the methodology by producing EV mimetics using a wide variety of liposomal formulations. Finally, we show that two distinct membrane proteins can be inserted in the same EV mimetic, further indicating versatility and broad applicability. This study presents a modular and controllable strategy for cell-free synthesis of functional EV mimetics, which provides a meaningful step toward addressing challenges in EV-inspired drug delivery development.

Current antidepressants face limitations due to the blood–brain barrier (BBB), systemic side effects and delayed onset. Here, we engineered an intranasal thermosensitive hydrogel (EVs@IN) encapsulating Chlorella vulgaris-derived extracellular vesicles (EVs) for sustained nose-to-brain delivery. EVs@IN significantly enhanced nasal mucosal retention and facilitated targeted transport of EVs to the hippocampus via olfactory pathways, while minimizing pulmonary exposure and clearance. In mouse models of depression (LPS-induced and CUMS), intranasal EVs@IN elicited rapid and potent alleviation of depressive- and anxiety-like behaviours. Mechanistically, EVs modulated astrocyte phenotypic transformation, reducing the release of neurotoxic complement C3 and suppressing neuroinflammation. Concurrently, they activated the Nrf2-Pgc-1α pathway, enhanced antioxidant defences (elevated SOD and GSH), mitigated oxidative stress and restored synaptic plasticity and neurogenesis in the hippocampus. Furthermore, we demonstrated the capacity of EVs to serve as efficient drug carriers for brain delivery. EVs@IN exhibited excellent long-term biocompatibility in vivo. Our findings establish plant-derived EVs within a sustained-release intranasal platform as a promising, scalable and BBB-bypassing strategy for the rapid treatment of depression and potentially other neuropsychiatric disorders.

Gastric cancer (GC) persists as one of the most lethal malignancies globally, primarily due to late-stage diagnosis, limited therapeutic targeting options, and inherent resistance to conventional therapies. While molecular profiling has advanced our understanding of GC, the development of effective delivery systems capable of precise tumour targeting and enhanced treatment response remains an unmet need. In this work, we explored targeted therapeutic approaches for GC by leveraging patient-derived organoid models. Firstly, we confirmed claudin-4 (CLDN4) as an overexpressed target in malignant epithelial cells in GC through comprehensive analysis of multiple single-cell RNA sequencing datasets. Capitalising on this discovery, we developed an innovative nano-therapeutic biomaterial, designated NESC (NK-sEV-SpoVM-c-CPE^Q317I), by engineering natural killer cell-derived small extracellular vesicles (NK-sEVs) with a CLDN4-targeting peptide and a membrane-curvature-sensing domain. Multimodal imaging further confirmed tumour-specific accumulation of NESC, underscoring its targeting precision. Proteomic profiling and functional assays revealed that NK-sEVs possessed intrinsic radiosensitising properties, which were significantly augmented upon conjugation with the targeting peptide. The resulting NESC platform demonstrated robust tumour-suppressive activity and enhanced radiosensitisation in both GC organoids and organoid-derived xenograft models. Collectively, by harnessing patient-derived organoids for functional validation, this study not only establishes a versatile framework for developing targeted sEV-based therapeutics but also provides a translational foundation for future clinical applications in GC management.

Extracellular vesicles (EVs) are increasingly recognized as mediators of central nervous system (CNS) function and pathologies, including multiple sclerosis (MS). While plasma-derived EVs have been explored as biomarkers in MS, little is known about EVs in CNS tissue. Here, we characterize EVs from postmortem normal-appearing white matter (NAWM) of MS and control brains. EVs were separated by differential centrifugation followed by size exclusion chromatography and characterized using nanoflow cytometry, single-particle reflectance imaging sensing (SP-IRIS) and transmission electron microscopy. EV size, yield and morphology did not differ significantly between MS and control samples. Despite the small sample size (n = 4 per group), proteomic analyses revealed downregulation of synaptic and mitochondrial proteins and upregulation of complement and inflammatory proteins and pathways in MS NAWM EVs. This suggests that EVs reflect ongoing synaptic pathology, metabolic dysfunction and CNS-compartmentalized inflammation and that they may actively contribute to these pathological processes. Deconvolution analyses suggest a shift in EV cellular origin, with an increased astrocytic and decreased neuronal EV contribution in MS. Several proteomic changes we observed in CNS-derived EVs have also been reported in circulating EVs of people with MS, establishing this CNS tissue EV study as a valuable resource for identifying biomarker candidates for brain-derived plasma EV studies.

Intercellular communication is essential for healthy embryo development, yet the role and dynamics of extracellular space in the maternal-embryonic dialogue remain unclear. Furthermore, little is known of maternal and embryonic metabolic states during early endometrial preparation and after the embryo enters the uterine cavity. Using a human in vitro co-culture model and extracellular vesicle (EV)-specific tools, we dynamically tracked EV secretion, uptake and processing between embryonic and endometrial cells at early stages of cell-to-cell communication. Hormonal stimulation altered endometrial secretory output, producing distinct EV populations. Stimulated EVs (St-EVs) differed from non-stimulated EVs (NSt-EVs) in size, secretion dynamics, uptake efficiency and metabolic cargo, selectively packaging energy-related metabolites and aryl hydrocarbon receptor (AhR) ligands. AhR inhibition increased spheroid attachment, suggesting that AhR signalling regulates implantation by modulating the endometrial environment. Additionally, lipid droplets (LDs) affected by endometrial- and embryo-derived EVs were actively secreted and taken up by embryonic cells, highlighting their role in implantation. EVs were not only exchanged between the embryo and endometrium but were also rapidly internalized, influencing mitochondrial activity, lipid metabolism and extracellular matrix remodelling. Translation of EV-derived mRNA occurred within 1 h of uptake, driving cellular changes and enhancing embryo attachment. These findings suggest EVs, extracellular metabolites and LDs mobilized between the endometrium and embryo coordinate to promote embryo attachment and implantation. This study advances our understanding of embryo-maternal EV-mediated communication and provides a valuable model for investigating EV-mediated simultaneous intercellular bidirectional crosstalk in other biological contexts.

The emergence and worldwide dissemination of antibiotic resistance genes (ARGs) compromise antibiotic therapy and are a major public health crisis. Horizontal gene transfer (HGT) plays a major role in the spread of ARGs among bacterial pathogens. Outer membrane vesicles (OMVs), which are membrane-bound particles and naturally released by Gram-negative bacteria, have been reported to carry a variety of cargos such as DNA, proteins and lipids. However, it remains unknown whether OMVs mediate transfer of ARGs in Campylobacter, an important foodborne pathogen whose resistance to antibiotics poses a serious threat to public health. To close this knowledge gap, we determined the role of OMVs in ARG transfer. Using a non-conjugative plasmid (pRY112), we demonstrated that OMVs successfully transferred the plasmid from Campylobacter coli to Campylobacter jejuni. Additionally, OMVs transferred chromosomally encoded florfenicol resistance from a clinical C. coli isolate (SH89) to C. jejuni. The OMV-mediated transfer is independent of natural transformation as both DNase I treatment (for digestion of external-free DNA) and use of a strain deficient of natural transformation as the recipient strain did not affect OMV-mediated ARG transfer. Transmission electron microscopy revealed direct fusion between OMVs and recipient bacterial membranes, suggesting membrane fusion as the mechanism for OMV-mediated DNA transfer. Furthermore, we showed that OMVs derived from strains expressing a functionally-enhanced CmeB (FE-CmeB) transiently protect florfenicol-susceptible C. jejuni against selection by the antibiotic. Together, these findings indicate that OMVs mediate the transfer of both plasmid- and chromosome-encoded ARGs in Campylobacter and define OMVs as a novel pathway for Campylobacter to acquire antibiotic resistance via HGT.

According to the endosymbiotic theory of mitochondrial origin, an α-proteobacterium entered a prokaryotic cell and, through symbiosis, evolved into the mitochondria—the powerhouse of the cell. Like other bacteria, the α-proteobacteria generate their own extracellular vesicles (EVs), a trait that was passed onto the mitochondria, enabling them to generate mitochondria-derived vesicles (MDVs). MDVs, similar to small EVs (sEVs), are vesicles ranging from 30 to 200 nm in diameter and carry cargo for degradation by lysosomes and peroxisomes. MDVs share several features with sEVs, including targeted cargo degradation, biogenesis, packaging into multivesicular bodies, nucleic acid and protein transportation, induction of immune responses, and surface antigen presentation. MDVs may also be released from the cell in a manner similar to sEVs, potentially influencing intercellular communication and immune responses. Furthermore, the presence of MDVs presents opportunities for early disease detection, including neurodegenerative disorders and cancer. In this review, we explore the differences and similarities between MDVs and EVs, including their roles in immunity.

Adeno-associated virus (AAV)–mediated gene therapies face critical clinical limitations, including immune-mediated neutralization by pre-existing antibodies and dose-dependent hepatotoxicity. Extracellular vesicle-encapsulated AAVs (EV-AAVs) offer a promising solution by shielding AAVs from antibody recognition, yet existing production methods remain inefficient and impractical for clinical application. Here, we developed a cellular nanoporation (CNP) platform that enables scalable, high-yield generation of EV-AAVs, achieving an approximately 11-fold increase in production efficiency compared with conventional methods. In LDLR-deficient murine models with pre-existing neutralizing antibodies (1:200), EV-AAV-LDLR at half the standard AAV dose robustly restored hepatic LDL receptor expression and attenuated atherosclerosis progression. Notably, EV-AAV exhibited superior immune evasion capabilities, maintaining 2.3-fold higher hepatic transduction efficiency than conventional AAV upon secondary dosing due to antibody shielding. Importantly, EV-AAV therapy markedly reduced hepatotoxicity, with serum AST/ALT levels comparable to saline-treated controls, thereby overcoming a critical safety barrier of high-dose AAV treatment. These results demonstrate CNP as a clinically translatable platform for scalable EV-AAV manufacturing, enabling effective multi-dose regimens while overcoming key immunological and toxicity barriers in liver-directed gene therapy for familial hypercholesterolaemia.

Solid tissue-derived extracellular vesicles (ST-EVs) are extracellular vesicles (EVs) separated directly from solid tissues of both vertebrates and invertebrates. ST-EVs provide a physiologically relevant snapshot of tissue-specific molecular dynamics and can be enriched directly in situ, from tissues in their natural state, preserving the native characteristics of ST-EVs. However, their enrichment presents unique technical challenges compared to EVs derived from biofluids or cell culture media. The need for transparent reporting in ST-EV research is crucial to enhance the reproducibility, comparability, and reliability of research findings. The Solid Tissue Task Force, part of the Scientific Reproducibility Subcommittee of International Society for Extracellular Vesicles, aims to recommend reporting parameters and identify outstanding questions related to the pre-analytical and analytical handling of solid tissues, as well as ST-EV separation and characterization. These steps are essential for advancing the understanding of the biological roles of ST-EVs and their potential clinical applications.