Background <p>Neuromyelitis optica spectrum disorder (NMOSD) and multiple sclerosis (MS) are central demyelinating diseases with overlapping features but differing immunopathogenesis. Extracellular vesicle (EV)-associated microRNAs (miRNAs) have emerged as potential biomarkers, but their roles in NMOSD remain underexplored. This study aimed to assess EV-miRNA expression changes across NMOSD stages compared to relapsing–remitting MS (RRMS) and healthy controls (HC).</p> Methods <p>We enrolled 43 participants, including HC (<i>n</i> = 10), RRMS (<i>n</i> = 10), NMOSD in remission (NMOSD-r, <i>n</i> = 13), and NMOSD exacerbation (NMOSD-ex, <i>n</i> = 10). Plasma-derived EVs from all participants were isolated, and their morphology, size, and concentration were assessed. The expression of let-7i-5p, miR-21-5p, and miR-30b-5p in EV was quantified. EVs of NMOSD-ex patients were re-evaluated at one month after corticosteroid treatment (NMOSD-fu). The correlation between alterations of EV miRNA levels and clinical outcomes was analyzed.</p> Results <p>EV concentrations isolated from patients with RRMS, NMOSD-r and NMOSD-ex were significantly lower compared to those of HC, and the smallest of EV sizes was observed in NMOSD-ex group. MiR-30b-5p expression was markedly reduced in NMOSD and RRMS compared to HC, whereas let-7i-5p and miR-21-5p exhibited nonsignificant downward trends. In NMOSD-ex group, miR-30b-5p levels positively correlated with higher glial fibrillary acidic protein (GFAP) and greater disability. Following corticosteroid treatment, increased expression of miR-30b-5p was individually associated with motor function recovery, although the degree of visual improvement was comparatively limited.</p> Conclusion <p>Plasma-derived EV-associated miR-30b-5p may reflect a protective role in response to astrocytic injury and serve as a valuable biomarker for disease activity and treatment response in NMOSD.</p>