Molecular Encoding of Ischemic Stroke and its Resolution after Human Neural Stem Cell Therapy by Extracellular Vesicles

Extracellular vesicles (EVs) can cross the blood-brain barrier and enter the systemic circulation, potentially acting as peripheral biomarkers of stroke neuropathology. Here, we investigated alterations in EV RNA cargoes extracted from rat brain and plasma before and after stroke induction via middle cerebral artery occlusion and subsequent human neural stem cells (hNSCs) transplantation. EV RNA coexpression profiles were assessed, and digital source tracking was used to determine EV origin. The therapeutic effects of intra-arterial delivery of hNSCs on ischemic rat brains were quantified, focusing on functional recovery, resolution of ischemic lesions, and the microenvironment. Stroke induced distinct EV secretion patterns, with a notable increase in EV secretion from non-neuronal cells. hNSCs transplantation caused minimal immune rejection and transplanted cells survived in the brain for over a week. Stem cell-derived EVs were detected in peripheral blood, indicating prolonged systemic distribution after transplantation. Gene regulatory network analyses identified specific EV miRNAs that play crucial roles in neurogenesis, wound healing, angiogenesis, and blood-brain barrier integrity. An integrated analysis of EV RNAs in brain and plasma samples revealed that stroke increased correlations in RNA expression between brain and plasma and that hNSCs transplantation reversed the effect. Brain- and plasma-derived EVs carry similar molecular information after stroke, suggesting that plasma-derived EV RNAs reflect stroke pathophysiology. Intra-arterial transplantation of hNSCs improved outcomes after stroke in rats, by promoting endogenous neurogenesis and maintaining blood-brain barrier integrity. The identified EV miRNAs provided a new mechanism by which hNSCs transplantation regulates neural regeneration through the miR-204-5p/EFNB3 axis.