Extracellular vesicles (EVs) are membrane-bound particles secreted by almost all cell types. They are classified depending on their biogenesis and size into exosomes and microvesicles or according to their cell origin. EVs play a role in cell-to-cell communication, including contact-free cell synapsis, carrying active membrane proteins, lipids, and genetic material both inside the particle and on their surface. They have been related to several physiological and pathological conditions. In particular, increasing concentrations of EVs have been found in many autoimmune diseases including multiple sclerosis (MS). MS is a central nervous system demyelinating disease characterized by relapsing of symptoms followed by periods of remission. Close interaction between endothelial cells, leukocytes, monocytes and cells from central nervous system is crucial for the development of MS. This review summarizes the pathological role of EVs in MS and the relationship of EVs with clinical characteristics, therapy and biomarkers of the disease.

The research in extracellular vesicles (EVs) has been rising during the last decade. However, there is no clear consensus on the most accurate protocol to isolate and analyze them. Besides, most of the current protocols are difficult to implement in a hospital setting due to being very time consuming or to requirements of specific infrastructure. Thus, our aim is to compare five different protocols (comprising two different medium-speed differential centrifugation protocols; commercially polymeric precipitation -exoquick-; acid precipitation; and ultracentrifugation) for blood and urine samples to determine the most suitable one for the isolation of EVs. Nanoparticle tracking analysis, flow cytometry, western blot, electronic microscopy and spectrophotometry were used to characterize basic aspects of EVs such us concentration, size distribution, cell-origin and transmembrane markers and RNA concentration. The highest EV concentrations were obtained using the exoquick protocol, followed by both differential centrifugation protocols, while the ultracentrifugation and acid-precipitation protocols yielded considerably lower EV concentrations. The five protocols isolated EVs of similar characteristics regarding markers and RNA concentration however standard protocol recovered only small EVs. EV isolated with exoquick presented difficult to be analyzed with western blot. The RNA concentrations obtained from urine-derived EVs were similar to those obtained from blood-derived ones, despite the urine EV concentration being 10 to 20 times lower. We consider that a medium-speed differential centrifugation could be suitable to be applied in a hospital setting due to require the simplest infrastructure and recover higher concentration of EV than standard protocol. A workflow from sampling to characterization of EVs is proposed.

Aim: To evaluate whether circulating microparticles (MPs) derived from three cell subtypes (platelets, total leukocytes or monocytes) obtained from multiple sclerosis (MS) patients were modulated depending on the clinical status and to investigate the effect of treatments on MP levels. Patients & methods: The MP counts were assessed with flow cytometry. Results: The platelet-derived MP level was higher in untreated MS patients than controls. Relapsing–remitting patients showed the highest levels in the three subtypes of MP while secondary progressive patients presented similar levels to those of healthy controls. Treatments had significant effects increasing the three subtypes of MP counts. Conclusion: We suggest that MPs play a role in MS pathogenesis, reflecting disease status with an increment of their shedding during inflammatory periods and turning to baseline during chronic progressive degeneration.

Remyelination is a key aspect in multiple sclerosis pathology and a special effort is being made to promote it. However, there is still no available treatment to regenerate myelin and several strategies are being scrutinized. Myelination is naturally performed by oligodendrocytes and microRNAs have been postulated as a promising tool to induce oligodendrocyte precursor cell differentiation and therefore remyelination. Herein, DSPC liposomes and PLGA nanoparticles were studied for miR-219a-5p encapsulation, release and remyelination promotion. In parallel, they were compared with biologically engineered extracellular vesicles overexpressing miR-219a-5p. Interestingly, extracellular vesicles showed the highest oligodendrocyte precursor cell differentiation levels and were more effective than liposomes and polymeric nanoparticles crossing the blood–brain barrier. Finally, extracellular vesicles were able to improve EAE animal model clinical evolution. Our results indicate that the use of extracellular vesicles as miR-219a-5p delivery system can be a feasible and promising strategy to induce remyelination in multiple sclerosis patients.

Extracellular vesicles (EVs) are membrane-bound particles involved in intercellular communication. They carry proteins, lipids, and nucleotides such as microRNAs (miRNAs) from the secreting cell that can modulate target cells. We and others have previously described the presence of EVs in peripheral blood of multiple sclerosis (MS) patients and postulated them as novel biomarkers. However, their immune function in MS pathogenesis and the effect during the onset of new immunomodulatory therapies on EVs remain elusive. Here, we isolated plasma EVs from fingolimod-treated MS patients in order to assess whether EVs are affected by the first dose of the treatment. We quantified EVs, analyzed their miRNA cargo, and checked their immune regulatory function. Results showed an elevated EV concentration with a dramatic change in their miRNA cargo 5 h after the first dose of fingolimod. Besides, EVs obtained prior to fingolimod treatment showed an increased immune regulatory activity compared to EVs obtained 5 h post-treatment. This work suggests that EVs are implicated in the mechanism of action of immunomodulatory treatments from the initial hours and opens a new avenue to explore a potential use of EVs for early treatment monitoring.

Multiple sclerosis (MS) is a common inflammatory and degenerative disease that causes neurological disability. It affects young adults and its prevalence is higher in women. The most common form is manifested as a series of acute episodes of neurological disability (relapses) followed by a recovery phase (remission). Recently, non-coding RNAs have emerged as new players in transcriptome regulation and in turn, they could have a significant role in MS pathogenesis. In this context, our aim was to investigate the involvement of microRNAs and snoRNAs in the relapse-remission dynamics of MS in peripheral blood leucocytes, to shed light on the molecular and regulatory mechanisms that underlie this complex process. With this approach, we found that a subset of small non-coding RNAs (sncRNA) is altered in relapse and remission, revealing unexpected opposite changes that are sex dependent. Furthermore, we found that a relapse-related miRNA signature regulated general metabolism processes in leucocytes and miRNA altered in remission are involved in the regulation of innate immunity. We observed that sncRNA dysregulation is different in relapse and remission leading to differences in transcriptome regulation and that this process is sex dependent. In conclusion, relapse and remission have a different molecular background in men and women.