Extra-cellular vesicles (EVs) are bilayer membrane structures enriched with proteins, nucleic acids and other active molecules and have been implicated in many physiological and pathological processes over the past decade. Recently, evidence suggests EVs to play a more dichotomic role in the regulation of the immune system, whereby an immune response may be enhanced or supressed by EVs depending on their cell of origin and its functional state. EVs derived from antigen (Ag)-presenting cells for instance, have been involved in both innate and acquired (or adaptive) immune responses, as Ag carriers or presenters, or as vehicles for delivering active signalling molecules. On the other hand, tumor and stem cell derived EVs have been identified to exert an inhibitory effect on immune responses by carrying immuno-modulatory effectors, such as transcriptional factors, non-coding RNA (Species) and cytokines. In addition, stem cell-derived EVs have also been reported to impair dendritic cell maturation and to regulate the activation, differentiation and proliferation of B cells. They have been shown to control natural killer cell activity and to suppress the innate immune response. Studies reporting the role of EVs on T lymphocyte modulation are controversial. Discrepancy in literature may be due to stem cell culture conditions, methods of EV purification, EV molecular content and functional state of both parental and target cells. However, mesenchymal stem cell-derived EVs were shown to play a more suppressive role by shifting T cells from an activated to a T regulatory phenotype. In this review we will discuss how stem cell-derived EVs may contribute towards the modulation of the immune response. Collectively, stem cell-derived EVs mainly exhibit an inhibitory effect on the immune system.

Extracellular vesicles (EVs) derived from mesenchymal stem cells isolated from both bone marrow (BMSCs) and adipose tissue (ADSCs) show potential therapeutic effects. These vesicles often show a similar beneficial effect on tissue regeneration, but in some contexts, they exert different biological properties. To date, a comparison of their molecular cargo that could explain the different biological effect is not available. Here, we demonstrated that ADSC-EVs, and not BMSC-EVs, promote wound healing on a murine model of diabetic wounds. Besides a general similarity, the bioinformatic analysis of their protein and miRNA cargo highlighted important differences between these two types of EVs. Molecules present exclusively in ADSC-EVs were highly correlated to angiogenesis, whereas those expressed in BMSC-EVs were preferentially involved in cellular proliferation. Finally, in vitro analysis confirmed that both ADSC and BMSC-EVs exploited beneficial effect on cells involved in skin wound healing such as fibroblasts, keratinocytes and endothelial cells, but through different cellular processes. Consistent with the bioinformatic analyses, BMSC-EVs were shown to mainly promote proliferation, whereas ADSC-EVs demonstrated a major effect on angiogenesis. Taken together, these results provide deeper comparative information on the cargo of ADSC-EVs and BMSC-EVs and the impact on regenerative processes essential for diabetic wound healing.

Several studies in the past have investigated the expression of micro RNAs (miRNAs) in saliva as potential biomarkers. Since miRNAs associated with extracellular vesicles (EVs) are known to be protected from enzymatic degradation, we evaluated whether salivary EVs from patients with oral squamous cell carcinoma (OSCC) were enriched with specific subsets of miRNAs. OSCC patients and controls were matched with regards to age, gender and risk factors. Total RNA was extracted from salivary EVs and the differential expression of miRNAs was evaluated by qRT-PCR array and qRT-PCR. The discrimination power of up-regulated miRNAs as biomarkers in OSCC patients versus controls was evaluated by the Receiver Operating Characteristic (ROC) curves. A preliminary qRT-PCR array was performed on samples from 5 OSCC patients and 5 healthy controls whereby a subset of miRNAs were identified that were differentially expressed. On the basis of these results, a cohort of additional 16 patients and 6 controls were analyzed to further confirm the miRNAs that were up-regulated or selectively expressed in the previous pilot study. The following miRNAs: miR-302b-3p and miR-517b-3p were expressed only in EVs from OSCC patients and miR-512-3p and miR-412-3p were up-regulated in salivary EVs from OSCC patients compared to controls with the ROC curve showing a good discrimination power for OSCC diagnosis. The Kyoto Encyclopedia of Gene and Genomes (KEGG) pathway analysis suggested the possible involvement of the miRNAs identified in pathways activated in OSCC. In this work, we suggest that salivary EVs isolated by a simple charge-based precipitation technique can be exploited as a non-invasive source of miRNAs for OSCC diagnosis. Moreover, we have identified a subset of miRNAs selectively enriched in EVs of OSCC patients that could be potential biomarkers.

Evidence indicates that Alix, an accessory protein of the endosomal sorting complex required for transport (ESCRT), is involved in the biogenesis of extracellular vesicles (EVs). EVs contain selected patterns of microRNAs (miRNAs or miRs); however, little is known about the mechanisms of miRNA enrichment in EVs. The aim of the present study was to evaluate whether Alix is involved in the packaging of miRNAs within EVs released by human liver stem‑like cells (HLSCs). EVs released from HLSCs were enriched with miRNAs and expressed Alix and several RNA-binding proteins, including Argonaute 2 (Ago2), a member of the Argonaute family known to be involved in the transport and the processing of miRNAs. Co-immunoprecipitation experiments revealed an association between Alix and Ago2. The results from RT-qPCR indicated that in the Alix/Ago2 immunoprecipitates, miRNAs were detectable. EVs were instrumental in transferring selected miRNAs from HLSCs to human endothelial cells absent in the latter cells. Alix knockdown did not influence the number of EVs released by HLSCs, but it significantly decreased miRNA expression levels in the EVs and consequently their transfer to the endothelium. Our findings indicate that Alix binds to Ago2 and miRNAs, suggesting that it plays a key role in miRNA enrichment during EV biogenesis. These results may represent a novel function of Alix, demonstrating its involvement in the EV-mediated transfer of miRNAs.

Extracellular vesicles (EVs) are considered to be a novel complex mechanism of cell communication within the tumor microenvironment. EVs may act as vehicles for transcription factors and nucleic acids inducing epigenetic changes in recipient cells. Since tumor EVs may be present in patient biological fluids, it is important to investigate their function and molecular mechanisms of action. It has been shown that tumor cells release EVs, which are capable in regulating cell apoptosis, proliferation, invasion, epithelial-mesenchymal transition, as well as, to suppress activity of immune cells, to enhance angiogenesis, and to prepare a favorable microenvironment for metastasis. On the other hand EVs derived from stromal cells, such as mesenchymal stem cells, may influence the phenotype of tumor cells through reciprocal crosstalk greatly influenced by the transcription factors and nucleic acids they carry. In particular, non-coding RNAs (ncRNA), including microRNAs and long ncRNA RNAs, have recently been identified as the main candidates for the phenotypic changes induced in the recipient cells by EVs. Non-coding RNAs, which are important regulators of mRNA and protein expression, can function either as tumor suppressors or as oncogenes, depending on their targets. Herein, we have attempted to revise actual evidence reported in the literature on the role of EVs in tumor biology with particular regards to the crosstalk of ncRNAs between cancer cells and mesenchymal stem cells.