Clinical Values of Serum Extracellular Vesicle MicroRNA Profiles as Molecular Biomarkers for Ischemic Stroke and Transient Ischemic Attack.

Initial sequencing identified 134 differentially expressed EV miRNAs in ischemic stroke patients compared to controls.

• Single-molecule sequencing was performed on pooled sera from 50 IS patients and 50 controls
• Of the 134 differentially expressed EV miRNAs, 36 were upregulated and 98 were downregulated in IS
• Altered miRNAs were subsequently validated via individual qRT-PCR in the same cohort of 50 IS patients and 50 controls

EV let-7f-5p was significantly decreased in both ischemic stroke and transient ischemic attack patients.

• Validation was performed via individual qRT-PCR in the initial cohort and confirmed in expanded internal validation cohorts (100 IS, 40 TIA, 100 controls)
• Results were further tested in an external validation cohort (32 IS, 8 TIA, 32 controls)
• Reduced let-7f-5p was independently associated with both IS and TIA in multivariable logistic regression analysis
• let-7f-5p reduction was observed across both internal and external validation cohorts

EV miR-486-5p and let-7b-5p were elevated in ischemic stroke, with let-7b-5p showing higher levels in IS compared to TIA.

• Both miR-486-5p and let-7b-5p were validated as upregulated specifically in IS patients
• let-7b-5p levels were higher in IS than in TIA, suggesting potential utility for distinguishing between the two conditions
• Increased EV let-7b-5p was independently associated with IS in multivariable logistic regression analysis
• Findings were confirmed in expanded internal (100 IS, 40 TIA, 100 controls) and external (32 IS, 8 TIA, 32 controls) validation cohorts

A combined EV miRNA panel may effectively distinguish IS and TIA from controls and stratify IS severity and TIA subsequent stroke risk.

• Diagnostic performance was assessed via ROC and logistic regression analyses
• The panel consists of downregulated let-7f-5p, upregulated miR-486-5p, and upregulated let-7b-5p
• The panel was evaluated for capacity to stratify IS severity
• The panel was also evaluated for its ability to stratify TIA subsequent stroke risk

Bioinformatic analysis predicted FOXO1 and BDNF as key targets of the identified EV miRNAs.

• Bioinformatics analysis was employed to explore mechanisms downstream of the identified miRNAs
• Decreased FOXO1 and increased BDNF were observed in IS serum and serum EVs
• FOXO1 downregulation was replicated in an in vitro oxygen-glucose deprivation/reperfusion (OGD/R) cellular model
• These findings suggest FOXO1 and BDNF dysregulation may contribute to ischemic injury pathology

Serum extracellular vesicle miRNA profiling was evaluated as a non-invasive diagnostic approach for ischemic stroke and transient ischemic attack.

• The study used a multi-stage design: initial screening with single-molecule sequencing, followed by individual qRT-PCR validation, and testing in expanded internal and external cohorts
• Total cohorts included up to 100 IS patients, 40 TIA patients, and 100 controls in the internal validation phase
• An external validation cohort of 32 IS, 8 TIA, and 32 controls was also used
• Both ROC analysis and logistic regression were used to assess diagnostic performance