Abstract
Purposes Spinal cord ischemia-reperfusion injury (SCII) is initiated following the occlusion of supporting blood vessels, leading to the loss of neurological function. Here, we aimed to study the regenerative properties of tourniquet-induced hindlimb ischemia exosomes (Exos) in SCII Wistar rats. Methods Exos were isolated from rats following tourniquet-induced hindlimb ischemia. CellTracker™ CM-DiI-labeled Exos were injected systematically into SCII rats subjected to 60 min of abdominal aorta occlusion. The distribution of Exos was monitored using an immunofluorescence assay. Using histological examination and real-time PCR analysis, glial cell number, pyknotic and swollen neurons, and expression of apoptosis genes were studied. Oxidative stress was examined by measuring the SOD, GPx activity, MDA, and TAC levels. The neurological assessments were also performed 72 hours after the Exo injection. Results Data revealed cup-shaped spherical Exos with average size and zeta potential of 279.3 nm and 15.6 mV, respectively. The isolated particles were CD9+, CD63+, and CD81+, indicating the existence of typical Exo biomarkers. Histological analysis showed reduced gliosis, pyknotic, and swollen neurons compared to SCII rats after Exos injection (p<0.05). Data indicated the existence of Exos at the site of injury 24 hours after systemic injection. The injection of hypoxic Exos led to inhibition of apoptosis [Bax (~0.6-fold↓), and Bcl-2 (~3.97-fold↑)] and reduction of oxidative stress [MDA (~58%↓), SOD (~310%↑), GPx (~260%↑), and TAC (~300%↑)] compared to SCII rats (p<0.05). Neurological assessments revealed the reduction of withdrawal response and motor deficit index in SCII rats after injection of hypoxic Exos. Conclusion Hypoxic Exos are valid regenerative tools for the alleviation of spinal cord injury following ischemic/reperfusion.