Abstract P751: Global Analyses of Protein and Microrna Cargo in Neural Stem Cell Derived-Exosomes After Brain Ischemia

Background: Adult neurogenesis facilitates brain remodeling after stroke. Exosomes derived from neural stem cells (NSCs) promote ischemic neurovascular remodeling including angiogenesis and axonal outgrowth, potentially by transferring their cargo proteins and miRNAs to recipient cells. However, cargo profiles of proteins and miRNAs in NSC exosomes have not been investigated. Methods: Exosomes were isolated from supernatants of cultured NSCs harvested from the subventricular zone of rats subjected to 7 day middle cerebral artery occlusion (MCAO) and non-MCAO rats, respectively. Mass spectrometry and miRNA array were utilized to determine the protein and miRNA profiles of NSC-derived exosomes (NSC-Exos). Bioinformatic pathway analyses were performed using Ingenuity Pathway Analysis (IPA). Results: Exosome markers and size distribution (50-200nm) were validated with Western blot, transmission electron microscopy and Nanosight measurements, respectively. Proteomics analysis yielded a total of 1,770 proteins in ischemic NSC-Exos. Bioinformatics analysis identified 24, 23 and 23 proteins that were related to neuronal cell proliferation, migration and differentiation, respectively. Intriguingly, enrichment signaling pathway analysis revealed cargo proteins in ischemic NSC-Exos were highly associated with dysfunction, membrane, and permeability of mitochondrion, indicating a critical role of extracellular mitochondrion in stroke-induced neurogenesis. In addition, 318 miRNAs were detected in ischemic NSC-Exos. Gene ontology analysis demonstrated that differentially expressed miRNAs between ischemic and non-ischemic NSC-Exos were highly related to inflammation, cell proliferation, cell cycle, and differentiation. The top 3 upregulated miRNAs including miR-106b, miR-542,miR-125b were validated in ischemic NSC-Exos using RT-PCR. The functions of these miRNAs are related to the induction of angiogenesis. Summary/Conclusion: Our results for the first time demonstrate that ischemic NSC-Exos contain a robust profile of protein and miRNA effectors, which may provide new insights into the function of NSC-Exos in stroke-induced neurogenesis and potentially lead to new therapeutic targets against stroke.