Extracellular vesicles (EVs) have recently come into the spotlight as potential cancer biomarkers. Isolation of pure EVs is complex, so wider use requires reliable and time‑efficient isolation methods. In the present study, galectin‑based magnetic glycan recognition particles, EXÖBead^® were investigated for their practicality as a novel EV isolation technique, exemplified here for squamous cell carcinoma of the head and neck. Analysis of the isolation method showed a high concentration of pure EVs with detection of specific EV markers such as CD9, CD63, CD81 and TSG101. No apolipoprotein A1 was shown in the isolates, indicating low contamination of this isolation technique compared with size exclusion chromatography. In addition, common leukocyte antigen (CD45), three HNSCC [epithelial cell adhesion molecule (EpCAM), pan‑cytokeratin and programmed death‑ligand 1 (PD‑L1)] and PanEV markers (premixed CD9, CD63 and CD81 antibodies) were measured by bead‑based flow cytometry (BFC). BFC revealed that CD45^Neg PanEV⁺, EpCAM⁺ PanEV⁺ and PD‑L1⁺ PanEV⁺ were significantly higher in tumor patients compared with healthy control plasma. CD45^Neg PanEV⁺ and CD45⁺ PanEV⁺ carrying two or three HNSCC biomarkers were also significantly higher in tumor patients compared with healthy controls (BFC). Comparison of the functional immunosuppression effect of eluted tumor patient plasma EVs from EXÖBead^® and commercial polyethylene glycol isolation showed a significant tumor‑dependent increase in concentration of EVs. A peripheral blood mononuclear cell activation assay also showed that the T‑cell functionality of tumor patient plasma EVs isolated with EXÖBead^® was preserved in vitro. In conclusion, isolation using galectin‑based magnetic glycan recognition particles is a novel method for isolating plasma EVs with low lipoprotein contamination. Bead‑based flow cytometry provided an easy way to understand EV subpopulations. EXÖBead^® therefore showed great potential as a new isolation tool with high throughput capacity that could potentially be used in a clinical setting.

Exosomes are nanovesicles involved in intercellular communications. They are released by a variety of cell types; however, their presence in the inner ear has not been described in the literature. The aims of this study were to determine if exosomes are present in the inner ear and, if present, characterize the changes in their protein content in response to ototoxic stress. In this laboratory investigation, inner ear explants of 5-day-old Wistar rats were cultured and treated with either cisplatin or gentamicin. Hair cell damage was assessed by confocal microscopy. Exosomes were isolated using ExoQuick, serial centrifugation, and mini-column methods. Confirmation and characterization of exosomes was carried out using transmission electron microscopy (TEM), ZetaView, BCA protein analysis, and proteomics. Vesicles with a typical size distribution for exosomes were observed using TEM and ZetaView. Proteomic analysis detected typical exosome markers and markers for the organ of Corti. There was a statistically significant reduction in the exosome protein level and number of particles per cubic centimeter when the samples were exposed to ototoxic stress. Proteomic analysis also detected clear differences in protein expression when ototoxic medications were introduced. Significant changes in the proteomes of the exosomes were previously described in the context of hearing loss and ototoxic treatment. This is the first report describing exosomes derived from the inner ear. These findings may present an opportunity to conduct further studies with the hope of using exosomes as a biomarker to monitor inner ear function in the future.

Exosomes are nanovesicles part of a recently described intercellular communication system. Their properties seem promising as a biomarker in cancer research, where more sensitive monitoring and therapeutic applications are desperately needed. In the case of head and neck squamous cell carcinoma (HNSCC), overall survival often remains poor, although huge technological advancements in the treatment of this disease have been made. In the following review, diagnostic and therapeutic properties are highlighted and summarised. Impressive first results have been obtained but more research is needed to implement these innovative techniques into daily clinical routines.

Glioblastomas are among the most aggressive tumors, and with low survival rates. They are characterized by the ability to create a highly immunosuppressive tumor microenvironment. Exosomes, small extracellular vesicles (EVs), mediate intercellular communication in the tumor microenvironment by transporting various biomolecules (RNA, DNA, proteins, and lipids), therefore playing a prominent role in tumor proliferation, differentiation, metastasis, and resistance to chemotherapy or radiation. Exosomes are found in all body fluids and can cross the blood–brain barrier due to their nanoscale size. Recent studies have highlighted the multiple influences of tumor-derived exosomes on immune cells. Owing to their structural and functional properties, exosomes can be an important instrument for gaining a better molecular understanding of tumors. Furthermore, they qualify not only as diagnostic and prognostic markers, but also as tools in therapies specifically targeting aggressive tumor cells, like glioblastomas.

Introduction: Bacteria inhabit the in- and outside of the human body, such as skin, gut or the oral cavity where they play an innoxious, beneficial or even pathogenic role. It is well known that bacteria can secrete membrane vesicles (MVs) like eukaryotic cells with extracellular vesicles (EVs). Several studies indicate that bacterial membrane vesicles (bMVs) play a crucial role in microbiome-host interactions. However, the composition of such bMVs and their functionality under different culture conditions are still largely unknown.Methods: To gain a better insight into bMVs, we investigated the composition and functionality of E. coli (DSM 105380) bMVs from the culture media Lysogeny broth (LB) and RPMI 1640 throughout the different phases of growth (lag-, log- and stationary-phase). bMVs from three time points (8 h, 54 h, and 168 h) and two media (LB and RPMI 1640) were isolated by ultracentrifugation and analyzed using nanoparticle tracking analysis (NTA), cryogenic electron microscopy (Cryo-EM), conventional transmission electron microscopy (TEM) and mass spectrometry-based proteomics (LC–MS/MS). Furthermore, we examined pro-inflammatory cytokines IL-1β and IL-8 in the human monocyte cell line THP-1 upon bMV treatment.Results: Particle numbers increased with inoculation periods. The bMV morphologies in Cryo-EM/TEM were similar at each time point and condition. Using proteomics, we identified 140 proteins, such as the common bMV markers OmpA and GroEL, present in bMVs isolated from both media and at all time points. Additionally, we were able to detect growth-condition-specific proteins. Treatment of THP-1 cells with bMVs of all six groups lead to significantly high IL-1β and IL-8 expressions.Conclusion: Our study showed that the choice of medium and the duration of culturing significantly influence both E. coli bMV numbers and protein composition. Our TEM/Cryo-EM results demonstrated the presence of intact E. coli bMVs. Common E. coli proteins, including OmpA, GroEL, and ribosome proteins, can consistently be identified across all six tested growth conditions. Furthermore, our functional assays imply that bMVs isolated from the six groups retain their function and result in comparable cytokine induction.