The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points.

Exosomes and other classes of extracellular vesicles (EVs) have gained interest due to their role in cell-to-cell communication. Knowledge of the molecular content of EVs may provide important information on features of parental cells and mechanisms of cross-talk between cells. To study functions of EVs it is essential to know their composition, that includes proteins, nucleic acids, and other classes biomolecules. The metabolome, set of molecules the most directly related to the cell phenotype, is the least researched component of EVs. However, the metabolome of EVs circulating in human blood and other bio-fluids is of particular interest because of its potential diagnostic value in cancer and other health conditions. On the other hand, the metabolome of EVs released to culture media in controlled conditions in vitro could shed light on important aspects of communication between cells in model systems. This paper summarizes the most common approaches implemented in EV metabolomics and integrates currently available data on the composition of the metabolome of EVs obtained in different models with particular focus on human body fluids and cancer cells.

Background: In general, the serum metabolome reflects the patient’s body response to both disease state and implemented treatment. Though serum-derived exosomes are an emerging type of liquid biopsy, the metabolite content of these vesicles remains under researched. The aim of this pilot study was to compare the metabolite profiles of the whole serum and serum-derived exosomes in the context of differences between cancer patients and healthy controls as well as patients’ response to radiotherapy (RT). Methods: Serum samples were collected from 10 healthy volunteers and 10 patients with head and neck cancer before and after RT. Metabolites extracted from serum and exosomes were analyzed by the gas chromatography–mass spectrometry (GC–MS). Results: An untargeted GC–MS-based approach identified 182 and 46 metabolites in serum and exosomes, respectively. Metabolites that differentiated cancer and control samples, either serum or exosomes, were associated with energy metabolism. Serum metabolites affected by RT were associated with the metabolism of amino acids, sugars, lipids, and nucleotides. Conclusions: cancer-related features of energy metabolism could be detected in both types of specimens. On the other hand, in contrast to RT-induced changes observed in serum metabolome, this pilot study did not reveal a specific radiation-related pattern of exosome metabolites.

Background Serum metabolome reflects a general patient's body response to both disease state (e.g., cancer) and implemented treatment (e.g., radiotherapy, RT). Though serum-derived exosomes are an emerging type of liquid biopsy, a metabolite content of these vesicles remains under-researched yet. In this pilot study, we aimed to compare metabolite profiles of the whole serum and serum-derived exosomes in the context of differences between cancer patients and healthy controls as well as patients’ response to RT. Methods Serum samples were collected from 10 healthy volunteers and 10 patients with head and neck cancer, in the latter group samples were taken before and after the end of RT. Exosomes were isolated from serum by the size-exclusion chromatography. Metabolites were purified from the complete serum and exosomes using a methanol extraction and analyzed by the gas chromatography-mass spectrometry (GC-MS). Results An untargeted GC-MS-based approach allowed the detection of 182 and 46 metabolites in serum and exosomes, respectively (33 compounds were present in both types of specimens). The unsupervised analyses revealed that metabolite profiles of the whole serum but not serum-derived exosomes enabled the separation of all 3 groups of samples. There were 27 compounds whose serum levels were markedly different (large effect size) between control and cancer samples and 12 compounds whose serum levels were markedly different between cancer pre-RT and post-RT samples. On the other hand, only 4 metabolites present in exosomes showed markedly different levels between cancer and control samples. Noteworthy, metabolites that differentiated cancer and control samples, either serum or exosomes, were associated with energy metabolism pathways. Serum metabolites affected by RT were associated with pathways involved in the metabolism of amino acids, sugars, lipids, and nucleotides. Conclusions Metabolite profile of serum-derived exosomes is less complex than that of the complete serum. However, cancer-specific features of energy metabolism could be detected in both types of specimens. On the other hand, in contrast to RT-induced changes observed in serum metabolome, this pilot study did not reveal a specific radiation-related pattern of exosome metabolites.