Exploring extracellular vesicle surface markers and microRNA regulation following remote ischemic conditioning in patients with stroke; a randomized-controlled pilot study.

Remote ischemic conditioning (RIC) has demonstrated neuroprotective effects, yet translating these effects into clinical efficacy has proven challenging. This study aims to uncover the molecular mechanisms underlying RIC’s protective effects, which could potentially identify new therapeutic targets. We conducted the ENOS pilot study, a patient-assessor blinded, Sham-controlled clinical trial, to investigate the molecular changes in patients diagnosed with acute ischemic stroke (AIS). Patients were assigned to undergo either RIC (n = 9), involving five cycles of transient ischemia and reperfusion of the arm, or a Sham treatment (n = 9), using a similar device with less pressure to avoid creating ischemia, within 48 h of symptom onset. We collected plasma samples at three time points: at inclusion (pre-RIC), two hours post-initial RIC, and after seven days, with RIC administered twice daily. Analysis focused on brain biomarkers, specifically extracellular vesicle (EV) surface markers and circulating microRNAs (miRNAs). We identified significant differences in the regulation of CD62 on EVs and five specific miRNAs (miR-374a-5p, miR-20a-5p + miR-20b-5p, miR-19b-3p, miR-24-3p, miR-30d-5p) between the RIC and Sham groups. Notably, miR-374a-5p, known for its neuroprotective properties, was significantly increased in patients treated with RIC (p = 0.015). Furthermore, the changes of several of these miRNAs were correlated with improvements in red blood cell (RBC) deformability and aggregation during shear stress. In addition, we observed significant increases in CD62 on EVs at both the two-hour and 7-day follow-ups in the RIC group. These findings suggest that RIC may induce specific changes in EV surface markers and circulating miRNAs which could serve as future biomarkers for the RIC response. The trial was registered on clinicaltrails.gov (NCT04266639) on February 12, 2020 before trial initation on July 29, 2020. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12868-025-00993-1.