Platelet miRNA-Based Signature as a Potential Non-Invasive Tool for Discriminating Patients with Secondary Progressive Multiple Sclerosis from Healthy Individuals

Multiple sclerosis (MS) lacks minimally invasive biomarkers that reliably reflect disease-associated pathophysiological alterations, particularly during the progressive stage of the disease. Because blood platelets contribute to thrombo-inflammatory signalling and are the major vascular carriers of regulatory miRNA, we investigated whether platelet miRNA alterations are associated with platelet hyperreactivity in MS and whether these changes originate during thrombopoiesis by examining megakaryocytes. We analyzed platelets and megakaryocytes from MS patients and healthy controls by transmission electron microscopy, flow cytometry, ELISA, miRNA profiling based on microarrays, and Real-Time PCR validation. Logistic regression and bioinformatic analyses were used to evaluate the potential and biological relevance. MS platelets exhibited a hyperreactive phenotype, characterized by enhanced degranulation, microparticle release, P-selectin expression, and collagen-induced aggregation. GPVI levels were significantly elevated in platelets but remained unchanged in megakaryocytes. MiRNA profiling revealed widespread downregulation in platelets, while megakaryocytes showed selective upregulation of miR-223-3p. Following microarray screening, PCR validation of 10 selected miRNAs revealed widespread downregulation in platelets, while only miR-223-3p was significantly upregulated in megakaryocytes from MS patients. A two-miRNA model (let-7a-5p and miR-146a-5p) showed good internal discrimination (AUC = 0.89; sensitivity 89%; specificity 85%), but lower external performance (C-index = 0.66; sensitivity 50%; specificity 73%). MS is associated with a distinct platelet miRNA signature linked to acquired platelet hyperreactivity and vesiculation. Platelet-derived let-7a-5p and miR-146a-5p represent potential non-invasive biomarker candidates, although their diagnostic utility requires further validation in larger independent cohorts.