Abstract WP325: Peripheral Blood Mononuclear Cells Preconditioned by Oxygen-Glucose Deprivation Regulate Microglial Polarization by Secretome After Cerebral Ischemia

Abstract

Introduction: We previously reported that oxygen-glucose deprivation (OGD) decreased the expression of exosomal miR155-5p from peripheral blood mononuclear cells (PBMCs) and that the administration of PBMCs preconditioned by OGD (OGD-PBMCs) enhanced the VEGF expression from the resident microglia, leading to tissue remodeling in ischemic rats. However, the effect of exosomal miR155-5p downregulation on PBMCs and microglia is still unclear. Hypothesis: OGD-PBMCs polarize PBMCs and microglia towards a tissue-protective phenotype by secretome. Methods: We prepared PBMCs from human blood by centrifugation and cultured PBMCs under OGD conditions for 18 h. We transferred the conditioned media from OGD-PBMCs to PBMCs under normoxic conditions and evaluated protective marker SSEA-3 expression by flow cytometric analysis. Next, we cocultured OGD-PBMCs and microglia under normoxic conditions with or without miR-155-5p antisense oligonucleotide (ASO) inhibitor before performing western blotting to assess the VEGF expression levels in the microglia. Finally, we administered normoxic PBMC treated with miR155-5p ASO inhibitor to rats 7 d after cerebral ischemia and performed microscopic analysis to evaluate VEGF expression. Results: The frequency of stem cell marker SSEA-3-positive cells in PBMCs incubated with conditioned media from OGD-PBMCs was more prominent than that in PBMCs incubated with normal glucose media (P = 0.028). The VEGF expression levels in the microglia cocultured with OGD-PBMCs increased compared to that without OGD-PBMCs (P = 0.025). Moreover, VEGF expression levels in the microglia cocultured with OGD-PBMCs were higher with miR-155-5p ASO than without it (P = 0.049). miR-155-5p ASO-treated PBMC administration increased VEGF expression 21 d after administration in ischemic rats (28 d after cerebral ischemia). VEGF expression was not observed in ischemic rats without administration. Conclusions: OGD-PBMCs may cause phenotype conversion in resident microglia by the secretome, including reduction of exosomal miR-155-5p, after cerebral ischemia.