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  • 1
    Online Resource
    Online Resource
    Efficient Small Extracellular Vesicles (EV) Isolation Method and Evaluation of EV-Associated DNA Role in Cell–Cell Communication in Cancer
    MDPI AG ; 2022
    In:  Cancers Vol. 14, No. 9 ( 2022-04-20), p. 2068-
    Details
    In: Cancers, MDPI AG, Vol. 14, No. 9 ( 2022-04-20), p. 2068-
    Abstract: Small extracellular vesicles (sEVs) play essential roles in intercellular signaling both in normal and pathophysiological conditions. Comprehensive studies of dsDNA associated with sEVs are hampered by a lack of methods, allowing efficient separation of sEVs from free-circulating DNA and apoptotic bodies. In this work, using controlled culture conditions, we enriched the reproducible separation of sEVs from free-circulated components by combining tangential flow filtration, size-exclusion chromatography, and ultrafiltration (TSU). EV-enriched fractions (F2 and F3) obtained using TSU also contained more dsDNA derived from the host genome and mitochondria, predominantly localized inside the vesicles. Three-dimensional reconstruction of high-resolution imaging showed that the recipient cell membrane barrier restricts a portion of EV-DNA. Simultaneously, the remaining EV-DNA overcomes it and enters the cytoplasm and nucleus. In the cytoplasm, EV-DNA associates with dsDNA-inflammatory sensors (cGAS/STING) and endosomal proteins (Rab5/Rab7). Relevant to cancer, we found that EV-DNA isolated from leukemia cell lines communicates with mesenchymal stromal cells (MSCs), a critical component in the BM microenvironment. Furthermore, we illustrated the arrangement of sEVs and EV-DNA at a single vesicle level using super-resolution microscopy. Altogether, employing TSU isolation, we demonstrated EV-DNA distribution and a tool to evaluate the exact EV-DNA role of cell–cell communication in cancer.
    Type of Medium: Online Resource
    ISSN: 2072-6694
    URL: Article
    DOI: 10.3390/cancers14092068
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
    detail.hit.zdb_id: 2527080-1
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  • 2
    Online Resource
    Online Resource
    Tumour Progression Stage-Dependent Secretion of YB-1 Stimulates Melanoma Cell Migration and Invasion
    MDPI AG ; 2020
    In:  Cancers Vol. 12, No. 8 ( 2020-08-18), p. 2328-
    Details
    In: Cancers, MDPI AG, Vol. 12, No. 8 ( 2020-08-18), p. 2328-
    Abstract: Secreted factors play an important role in intercellular communication. Therefore, they are not only indispensable for the regulation of various physiological processes but can also decisively advance the development and progression of tumours. In the context of inflammatory disease, Y-box binding protein 1 (YB-1) is actively secreted and the extracellular protein promotes cell proliferation and migration. In malignant melanoma, intracellular YB-1 expression increases during melanoma progression and represents an unfavourable prognostic marker. Here, we show active secretion of YB-1 from melanoma cells as opposed to benign cells of the skin. Intriguingly, YB-1 secretion correlates with the stage of melanoma progression and depends on a calcium- and ATP-dependent non-classical secretory pathway leading to the occurrence of YB-1 in the extracellular space as a free protein. Along with an elevated YB-1 secretion of melanoma cells in the metastatic growth phase, extracellular YB-1 exerts a stimulating effect on melanoma cell migration, invasion, and tumourigenicity. Collectively, these data suggest that secreted YB-1 plays a functional role in melanoma cell biology, stimulating metastasis, and may serve as a novel biomarker in malignant melanoma that reflects tumour aggressiveness.
    Type of Medium: Online Resource
    ISSN: 2072-6694
    URL: Article
    DOI: 10.3390/cancers12082328
    Language: English
    Publisher: MDPI AG
    Publication Date: 2020
    detail.hit.zdb_id: 2527080-1
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  • 3
    Online Resource
    Online Resource
    Evaluation of Immunoregulatory Biomarkers on Plasma Small Extracellular Vesicles for Disease Progression and Early Therapeutic Response in Head and Neck Cancer
    MDPI AG ; 2022
    In:  Cells Vol. 11, No. 5 ( 2022-03-05), p. 902-
    Details
    In: Cells, MDPI AG, Vol. 11, No. 5 ( 2022-03-05), p. 902-
    Abstract: Head and Neck Cancers (HNCs) have highly immunosuppressive properties. Small extracellular vesicles (sEVs), including exosomes, nanosized mediators of intercellular communication in the blood, carry immunosuppressive proteins and effectively inhibit anti-tumor immune responses in HNCs. This study evaluates immunosuppressive markers on sEVs from 40 HNC patients at different disease stages and 3- and 6-month follow-up after surgery and/or chemoradiotherapy. As controls, sEVs from normal donors (NDs) are examined. Immunoregulatory surface markers on sEVs were detected as relative fluorescence intensity (RFI) using on-bead flow cytometry, and their expression levels were monitored in the early and late stages of HNC and during follow-up. In parallel, the sEV-mediated apoptosis of CD8+ Jurkat cells was assessed. Together with TGF-β1 and PD-L1 abundance, total sEV proteins are elevated with disease progression. In contrast, total sEV protein, including TGF-β1, PD-1 and PD-L1, decrease upon therapy response during follow-up. Overall survival analysis implies that high sEV PD-1/PD-L1 content is an unfavorable prognostic marker in HNC. Consistently, the sEV-mediated induction of apoptosis in CD8+ T cells correlates with the disease activity and therapy response. These findings indicate that a combination of immunoregulatory marker profiles should be preferred over a single marker to monitor disease progression and therapy response in HNC.
    Type of Medium: Online Resource
    ISSN: 2073-4409
    URL: Article
    DOI: 10.3390/cells11050902
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
    detail.hit.zdb_id: 2661518-6
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  • 4
    Online Resource
    Online Resource
    Challenges in the Isolation and Proteomic Analysis of Cancer Exosomes—Implications for Translational Research
    MDPI AG ; 2019
    In:  Proteomes Vol. 7, No. 2 ( 2019-05-15), p. 22-
    Details
    In: Proteomes, MDPI AG, Vol. 7, No. 2 ( 2019-05-15), p. 22-
    Abstract: Exosomes belong to the group of extracellular vesicles (EVs) that derive from various cell populations and mediate intercellular communication in health and disease. Like hormones or cytokines, exosomes released by cells can play a potent role in the communication between the cell of origin and distant cells in the body to maintain homeostatic or pathological processes, including tumorigenesis. The nucleic acids, and lipid and protein cargo present in the exosomes are involved in a myriad of carcinogenic processes, including cell proliferation, tumor angiogenesis, immunomodulation, and metastasis formation. The ability of exosomal proteins to mediate direct functions by interaction with other cells qualifies them as tumor-specific biomarkers and targeted therapeutic approaches. However, the heterogeneity of plasma-derived exosomes consistent of (a) exosomes derived from all kinds of body cells, including cancer cells and (b) contamination of exosome preparation with other extracellular vesicles, such as apoptotic bodies, makes it challenging to obtain solid proteomics data for downstream clinical application. In this manuscript, we review these challenges beginning with the choice of different isolation methods, through the evaluation of obtained exosomes and limitations in the process of proteome analysis of cancer-derived exosomes to identify novel protein targets with functional impact in the context of translational oncology.
    Type of Medium: Online Resource
    ISSN: 2227-7382
    URL: Article
    DOI: 10.3390/proteomes7020022
    Language: English
    Publisher: MDPI AG
    Publication Date: 2019
    detail.hit.zdb_id: 2720995-7
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