Comprehensive proteomics and microRNA analyses of adult neural stem cell derived exosomes after stroke

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Conference Proceeding

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J Extracell Vesicles


Introduction: Neural stem cells (NSC) are known to facilitate healing of ischemic brain tissues. Recent studies show that NSC derived exosomes function as paracrine effectors to promote neurovascular remodelling including angiogenesis and axonal outgrowth after stroke; nevertheless, the contents of the non-stroke and post stroke NSC exosome proteome and miRNA cargo have not been determined. Methods: NSC derived exosomes were purified from conditioned media of cultured NSCs harvested from the subventricular zone of non-ischemic and ischemic rats, respectively. Liquid chromatography mass spectrometry (LCMS) and miRNA array were employed to comprehensively characterize the protein and miRNA contents of NSCs and their derived exosomes after stroke. Bioinformatic analyses were performed using Ingenuity Pathway Analysis (IPA). Results: Exosome markers including CD63, CD9, Alix and size distribution (50-200nm) were verified with Western blot, transmission electron microscopy (TEM) and Nanosight, respectively. In total, proteomics analysis yielded 2409 and 1770 proteins identified in ischemic NSC and NSC derived exosomes, respectively. Bioinformatics analysis identified that 52, 39 and 31 proteins in the NSCs-derived exosomes were related to regulating neuronal cell proliferation, migration and differentiation, respectively. In addition, 318 miRNAs were identified in ischemic NSCs with 26% of miRNAs (84 miRNAs) overlapped with parent NSCs. Gene ontology analysis showed that up- and down-regulated miRNAs with the fold change above 1.5 were highly related to inflammation, invasion, cell proliferation, cell cycle, cell death, differentiation, etc. The top three upregulated miRNAs were validated in ischemic NSCexosomes using real-time RT-PCR. Summary/Conclusion: Collectively, the results of our proteomic and miRNA analysis, to our knowledge, demonstrate for the first time that NSC derived exosomes contain a robust profile of protein and miRNA effectors. These data provide a platform for beginning to understand the mechanism by which NSCs are activated after cerebral ischemia, and may lead to a deeper mechanistic understanding of their role in tissue repair after neural injury.



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