Journal of Extracellular Vesicles最新文献

The discovery that extracellular RNAs (exRNA) can act as endocrine signalling molecules established a novel paradigm in intercellular communication. ExRNAs can be transported, both locally and systemically in virtually all body fluids. In association with an array of carrier vehicles of varying complexity, exRNA can alter target cell phenotype. This highlights the important role secreted exRNAs have in regulating human health and disease. The NIH Common Fund exRNA Communication program was established in 2012 to accelerate and catalyze progress in the exRNA biology field. The program addressed both exRNA and exRNA carriers, and served to generate foundational knowledge for the field from basic exRNA biology to future potential clinical applications as biomarkers and therapeutics. To address scientific challenges, the exRNA Communication program developed novel tools and technologies to isolate exRNA carriers and analyze their cargo. Here, we discuss the outcomes of the NIH Common Fund exRNA Communication program, as well as the evolution of exRNA as a scientific field through the analysis of scientific publications and NIH funding. ExRNA and associated carriers have potential clinical use as biomarkers, diagnostics, and therapeutics. Recent translational applications include exRNA-related technologies repurposed as novel diagnostics in response to the COVID-19 pandemic, the clinical use of extracellular vesicle-based biomarker assays, and exRNA carriers as drug delivery platforms. This comprehensive landscape analysis illustrates how discoveries and innovations in exRNA biology are being translated both into the commercial market and the clinic. Analysis of program outcomes and NIH funding trends demonstrate the impact of this NIH Common Fund program.

As functional derivatives of mesenchymal stem cells (MSCs), small extracellular vesicles (sEVs) have garnered significant attention and application in regenerative medicine. However, the technical limitations for large-scale isolation of sEVs and their heterogeneous nature have added complexity to their applications. It remains unclear if the heterogeneous sEVs represent different aspects of MSCs functions. Here, we provide a method for the large-scale production of sEVs subpopulations derived from human umbilical cord mesenchymal stem cells (HucMSCs), utilizing tangential flow filtration combined with size exclusion chromatography. The resulting subpopulations, S1-sEVs and S2-sEVs, exhibited stable variations in size, membrane-marked proteins, and carrying cargos, thereby displaying distinct functions both in vitro and in animal disease models. S1-sEVs, that highly expressed CD9, HRS and GPC1, demonstrated a greater immunomodulatory impact, while S2-sEVs with enriched expression of CD63 and FLOT1/2 possessed enhanced capacities in promoting cell proliferation and angiogenesis. These discrepancies are attributed to the specific proteins and miRNAs they contain. Further investigation revealed that the two distinct sEVs subpopulations corresponded to different biological processes: the ESCRT pathway (S1-sEVs) and the ESCRT-independent pathway represented by lipid rafts (S2-sEVs). Therefore, we propose the potential for large-scale isolation and purification of sEVs subpopulations from HucMSCs with distinct functions. This approach may provide advantages for targeted therapeutic interventions in various MSC indications.

Japanese encephalitis virus (JEV) is a neurotropic zoonotic pathogen that poses a serious threat to public health. Currently, there is no specific therapeutic agent available for JEV infection, primarily due to the complexity of its infection mechanism and pathogenesis. Extracellular vesicles (EVs) have been known to play an important role in viral infection, but their specific functions in JEV infection remain unknown. Here, ultracentrifugation in combination with density gradient centrifugation was conducted to purify EVs from JEV-infected cells. The purified EVs were found to be infectious, with virions observed inside the EVs. Furthermore, our study showed the formation process of virion-containing EVs both in vitro and in vivo, which involved the fusion of multivesicular bodies with the cell membrane, leading to the release of virion-containing intraluminal vesicles into the extracellular space. Further studies revealed that EVs played a crucial role in JEV propagation by facilitating viral entry and assembly-release. Furthermore, EVs assisted JEV in evading the neutralizing antibodies and promoted viral capability to cross the blood-brain and placental barriers. Moreover, in vivo experiments demonstrated that EVs were beneficial for JEV infection and pathogenicity. Taken together, our findings highlight the significant contribution of EVs in JEV infection and provide valuable insights into JEV pathogenesis.

CprA is a short-chain dehydrogenase/reductase (SDR) that contributes to resistance against colistin and antimicrobial peptides. The cprA gene is conserved across Pseudomonas aeruginosa clades and its expression is directly regulated by the two-component system PmrAB. We have shown that cprA expression leads to the production of outer membrane vesicles (OMVs) that block autophagic flux and have a greater capacity to activate the non-canonical inflammasome pathway. In a murine model of sepsis, a P. aeruginosa strain deleted for cprA was less virulent than the wild-type (WT) strain. These results demonstrate the important role of CprA in the pathogenicity of P. aeruginosa. It is worth noting that CprA is also a functional ortholog of hemolysin F (HlyF), which is encoded by virulence plasmids of Escherichia coli. We have shown that other cryptic SDRs encoded by mammalian and plant pathogens, such as Yersinia pestis and Ralstonia solanacearum are functional orthologs of CprA and HlyF. These SDRs also induce the production of OMVs which block autophagic flux. This study uncovers a new family of virulence determinants in Gram-negative bacteria, offering potential for innovative therapeutic interventions and deeper insights into bacterial pathogenesis.

Extracellular vesicles (EVs) from brain-seeking breast cancer cells (Br-EVs) breach the blood-brain barrier (BBB) via transcytosis and promote brain metastasis. Here, we defined the mechanisms by which Br-EVs modulate brain endothelial cell (BEC) dynamics to facilitate their BBB transcytosis. BEC treated with Br-EVs show significant downregulation of Rab11fip2, known to promote vesicle recycling to the plasma membrane and significant upregulation of Rab11fip3 and Rab11fip5, which support structural stability of the endosomal compartment and facilitate vesicle recycling and transcytosis, respectively. Using machine learning and quantitative global proteomic, we identified novel Br-EV-induced changes in BECs morphology, motility, and proteome that correlate with decreased BEC cytoplasm and cytoskeletal organization and dynamics. These results define early steps leading to breast-to-brain metastasis and identify molecules that could serve as targets for therapeutic strategies for brain metastasis.

Parasitic helminths secrete extracellular vesicles (EVs) into their host tissues to modulate immune responses, but the underlying mechanisms are poorly understood. We demonstrate that Ascaris EVs are efficiently internalised by monocytes in human peripheral blood mononuclear cells and increase the percentage of classical monocytes. Furthermore, EV treatment of monocytes induced a novel anti-inflammatory phenotype characterised by CD14⁺, CD16^-, CC chemokine receptor 2 (CCR2^-) and programmed death-ligand 1 (PD-L1)⁺ cells. In addition, Ascaris EVs induced T cell anergy in a monocyte-dependent mechanism. Targeting professional phagocytes to induce both direct and indirect pathways of immune modulation presents a highly novel and efficient mechanism of EV-mediated host-parasite communication. Intra-peritoneal administration of EVs induced protection against gut inflammation in the dextran sodium sulphate model of colitis in mice. Ascaris EVs were shown to affect circulating immune cells and protect against gut inflammation; this highlights their potential as a subject for further investigation in inflammatory conditions driven by dysregulated immune responses. However, their clinical translation would require further studies and careful consideration of ethical implications.

Emerging evidence indicates that autophagy is tightly connected to the endocytic pathway. Here, we questioned the role of presenilins (PSENs 1 and 2), previously shown to be involved in autophagy regulation, in the secretion of small endocytic-originating extracellular vesicles known as exosomes. Indeed, while wild-type cells responded to stimuli promoting both multivesicular endosome (MVE) formation and secretion of small extracellular vesicles (sEVs) enriched in canonical exosomal proteins, PSEN-deficient cells were almost unaffected to these stimuli. Moreover, in PSEN-deficient cells, the re-expression of either PSEN1 or the functional active PSEN1delta9 mutant led to a rescue of most sEV secretion, while the deletion of PSEN1 alone almost fully phenocopied total PSEN invalidation. We found that the lack of sEV secretion in PSEN-deficient cells was also due to overactivated autophagy promoting MVEs to degradation rather than to plasma membrane fusion. Hence, in these cells, the autophagic blocker bafilomycin A1 (BafA1) not only increased the intracellular levels of the MVE protein CD63, but also turned on sEV secretion by stimulating autophagy-dependent unconventional secretion. In that case, sEVs arised from amphisomes and were enriched in both canonical exosomal proteins and lysosomal-autophagy-associated cargo. Altogether, we here demonstrate that PSENs, and particularly PSEN1, act as hub proteins controlling the balance between endosomal/autophagic degradation and secretion. More generally, our findings strengthen the view of a strong interconnection between the endocytic and autophagic pathways and their complementary roles in sEV secretion.

Despite immense interest in biomarker applications of extracellular vesicles (EVs) from blood, our understanding of circulating EVs under physiological conditions in healthy humans remains limited. Using imaging and multiplex bead-based flow cytometry, we comprehensively quantified circulating EVs with respect to their cellular origin in a large cohort of healthy blood donors. We assessed coefficients of variations to characterize their biological variation and explored demographic, clinical, and lifestyle factors contributing to observed variation. Cell-specific circulating EV subsets show a wide range of concentrations that do not correlate with cell-of-origin concentrations in blood, suggesting steady-state EV subset concentrations are regulated by complex mechanisms, which differ even for EV subsets from the same cell type. Interestingly, tetraspanin+ circulating EVs largely originate from platelets and to a lesser extent from lymphocytes. Principal component analysis (PCA) and association analyses demonstrate high biological inter-individual variation in circulating EVs across healthy humans, which are only partly explained by the influence of sex, menopausal status, age and smoking on specific circulating EV and/or tetraspanin+ circulating EV subsets. No global influence of the explored subject's factors on circulating EVs was detected. Our findings provide the first comprehensive, quantitative data towards the cell-origin atlas of plasma EVs, with important implications in the clinical use of EVs as biomarkers.

B cell maturation is crucial for effective adaptive immunity. It requires a complex signalling network to mediate antibody diversification through mutagenesis. B cells also rely on queues from other cells within the germinal centre. Recently, a novel class of intercellular signals mediated by extracellular vesicles (EVs) has emerged. Studies have shown that B cell EV-mediated signalling is involved in immune response regulation and tumorigenesis. However, the mechanistic role of B cell EVs is not yet established. We herein study the biological properties and physiological function of B cell EVs during B cell maturation. We use emerging technologies to profile B cell EV surface marker signatures at the single particle level, molecular cargo and physiological roles in B cell maturation. EV ncRNA cargo, characterised by RNA-seq, identified an EV-mediated novel non-coding RNA (ncRNA) regulatory network for B cell maturation. We show that a previously uncharacterised micro-RNA (miR-5099) in combination with a set of long ncRNA are carried within B cell EVs and could contribute to antibody diversification. The physiological role of EVs in B cell maturation is investigated using EV blockade assays and complementation studies using diverse EV sources further confirmed the physiological role and mode of action of EVs in B cell maturation.