Abstract TP256: Interactions Between Activated Microglia and Macrophages on Polarization and Chemotaxis Effect After Ischemic Stroke

Abstract

The inflammatory response following acute ischemic stroke is a well-known and widely studied phenomenon, but the mechanism is still unclear. After ischemic stroke, microglia and recruited macrophages play major roles in neuroinflammation after ischemic stroke. To characterize these roles, we explored how these cells affect counterpart’s differentiation, polarization and infiltration or migration. Methods: BV2 (microglia cell line) were treated with lipopolysaccharides (LPS; 10ng/mL) or interleukin-4 (IL-4; 25ng/mL), and the supernatant was collected as M1 or M2 conditioned media of BV2. The supernatant of PMA differentiated THP-1 (monocyte cell line) followed by LPS (20ng/mL) treatment or interleukin-13 (IL-13; 25ng/mL) & IL-4(25ng/mL) co-treatment was collected as M1 or M2 conditioned media of THP-1. After BV2 or THP-1 cultured in conditioned media, the activation and polarization were assessed by ICC and confocal microscopy for CD11b, CD86 and CD206. Transwell inserts of 3 and 8 μm pore membrane were used for THP-1 infiltration and BV2 migration assay. M1/M2 conditioned media of BV2 and THP-1 and the brain tissue of ischemic mouse model were assessed by proteome profiler array (PPA) to find target cytokine and chemokine. Results: THP-1 and BV2 expressed CD206 in M2 conditioned media of BV2 or THP-1. M2 conditioned media of BV2 increased the infiltratory ability of THP-1 while M1 conditioned media of THP-1 enhanced BV2 migration. MIP-2 was significantly expressed in the M1 conditioned media of BV2 and MIP-1a was high in the M2 conditioned media of BV2. MIF and CCL5 were highly expressed in M1 and M2 conditioned media of THP-1, particularly in M1. After ischemic stroke, chemokines were significantly expressed at 3 days than at 7 days in the PPA and several spots were match to the conditioned media of BV2. Conclusion: Our study suggests a new insight into the interaction of microglia and monocyte through their ability to expression of cytokine and chemokine after ischemic stroke. Modulation of microglial and monocyte by key cytokine and chemokine represents huge potential for new therapeutic strategies in acute brain injury.