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Extracellular vesicle properties and functions are defined by cell fitness status

ABSTRACT: Recent research has focused on understanding the influence of dying cells on immune function and the surrounding microenvironment. Apoptotic cells release a variety of extracellular vesicles (EVs), such as exosomes and microvesicles, and form apoptotic bodies through membrane blebbing. Despite increasing interest in EVs, the effects of apoptotic cell- versus live cell-derived EVs, have been poorly studied. Here, via transmission electron microscopy, nanoparticle tracking analysis, proteomics, and flow cytometry, we characterise the distinct features of EVs from live versus apoptotic cells. Using a model of Schistosoma mansoni infection, characterised by the presence of dying cells due to parasite egg accumulation, we demonstrated that adoptive transfer of apoptotic T cell-derived EVs attenuates liver pathology, in contrast to EVs from live T cells. Analysis of the transcriptomic profile of target macrophages and in vitro functional assays revealed that apoptotic cell-derived EVs activate NO-related pathways, which modulate macrophage function. Furthermore, these EVs specifically promoted fibroblast-mediated wound healing in vitro. Collectively, our findings highlight that cell fitness influences EV properties and their immune regulatory function. Investigating the differences between EVs from living and apoptotic cells is essential for advancing our understanding of immune regulation and optimising the development of EV-based therapeutic strategies.

INSTRUMENT(S):

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Antonia Elisabeth Gocke

LAB HEAD: Hartmut Schlüter

PROVIDER: PXD063388 | Pride | 2026-03-02

REPOSITORIES: Pride

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			Action	DRS
	240313_KM_EVTest_aT_EVs_1.raw	Raw
	240313_KM_EVTest_aT_EVs_2.raw	Raw
	240313_KM_EVTest_aT_EVs_3.raw	Raw
	240313_KM_EVTest_liveT_EVs_1.raw	Raw
	240313_KM_EVTest_liveT_EVs_2.raw	Raw

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Project description:Glioblastoma multiforme (GBM) is a highly aggressive and heterogeneous form of primary brain tumors, driven by a complex repertoire of oncogenic alterations, including the constitutively active epidermal growth factor receptor (EGFRvIII). EGFRvIII impacts both cell-intrinsic and non-cell autonomous aspects of GBM progression, including cell invasion, angiogenesis and modulation of the tumor microenvoiront, this is, at least in part, attributable to the release and intercellular trafficking of extracellular vesicles (EVs), heterogeneous membrane structures containing multiple bioactive macromolecules. Here we analyzed the impact of EGFRvIII on the profile of glioma EVs using a series of isogenic tumor cell lines, in which this oncogene exhibits a strong transforming activity. Thus, we observed that EGFRvIII expression alters several properties of glioma EVs, including their output and global protein composition. Using mass spectrometry, quantitative proteomic analysis and Gene Ontology terms filters, we observed that EVs released by EGFRVIII-transformed cells were enriched for extracellular exosome and focal adhesion related proteins. Among them, we validated the association of pro-invasive proteins (CD44, BSG, CD151) with EVs of EGFRvIII expressing cells, and down-regulation of exosomal markers (CD81 and CD82) in EVs of their indolent counterparts. Nano-flow cytometry revealed that the EV output from individual glioma cell lines was highly heterogeneous, such that only a fraction of EVs contained specific proteins such as EGFR, CD9, or others. Notably, cells expressing EGFRvIII released ample EVs double positive for CD44/BSG, and these proteins also co-localized in cellular filopodia. We also detected the expression of homophilic adhesion molecules and increased homologous EV uptake by EGFRvIII-positive glioma cells. These results suggest that oncogenic EGFRvIII reprograms the proteome of GBM-related EVs, a notion with considerable implications for their biological activity and properties relevant for development of EV-based cancer biomarkers.

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Extracellular vesicle properties and functions are defined by cell fitness status

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