Cerebrospinal fluid-derived extracellular vesicles after spinal cord injury promote vascular regeneration via PI3K/AKT signaling pathway.

Abstract

The cerebrospinal fluid (CSF), which surrounds the brain and spinal cord, is predominantly produced by the choroid plexus of the ventricle. Although CSF-derived extracellular vesicles (CSF-EVs) may be utilized as diagnostic and prognostic indicators for illnesses of the central nervous system (CNS), it is uncertain if CSF-EVs may have an impact on neurological function after spinal cord injury (SCI). Here, we isolated EVs using ultracentrifugation after extracting CSF from Bama miniature pigs. We then combined CSF-EVs with hydrogel and put it on the spinal cord's surface. To determine if CSF-EVs had an impact on mice's neurofunctional recovery, behavioral evaluations were employed. Both in vitro and in vivo, the effect of CSF-EVs on angiogenesis was assessed. We investigated whether CSF-EVs stimulated the PI3K/AKT pathway to alter angiogenesis using the PI3K inhibitor LY294002. CSF-EVs were successfully isolated and identified by transmission electron microscope (TEM), nano-tracking analysis (NTA), and western blot. CSF-EVs could be ingested by vascular endothelial cells as proved by in vivo imaging and immunofluorescence. We demonstrated that CSF-EVs derived from pigs with SCI (SCI-EVs) showed a better effect on promoting vascular regeneration as compared to CSF-EVs isolated from pigs receiving laminectomy (Sham-EVs). Behavioral assessments demonstrated that SCI-EVs could dramatically enhance motor and sensory function in mice with SCI. Western blot analysis suggested that SCI-EVs promote angiogenesis by activating PI3K/AKT signaling pathway, and the pro-angiogenetic effect of SCI-EVs was attenuated by the application of the LY294002 (PI3K inhibitor). Our study revealed that CSF-EVs could enhance vascular regeneration by activating the PI3K/AKT pathway, hence improving motor function recovery after SCI, which may offer potential novel therapeutic options for acute SCI. This study demonstrated the promotion of vascular regeneration and neurological function of CSF-derived exosomes, which may provide a potential therapeutic approach for the treatment of spinal cord injury.