Effects of M1 microglia-derived exosomes on the function of the blood-brain barrier

Abstract:

Objective To investigate the effects of M1 microglia-derived exosomes（M1-exo） on the function of an in-vitro blood-brain barrier（BBB） model and the expression of tight junction proteins between vascular endothelial cells.Methods Mouse microglia-derived BV2 cells were stimulated with lipopolysaccharide to be polarized towards the M1 phenotype，and exosomes were isolated and extracted after the phenotype was confirmed by flow cytometry. An in-vitro BBB model was constructed using the mouse brain microvascular endothelial cell line b.End3 and primary cultured mouse astrocytes. The BBB model was co-cultured with exosomes of different sources in three groups：normal culture for b.End3 cells（b.End3 group）；b.End3 cells + 25 μg/mL exosomes derived from normal BV-2 cells（b.End3+BV2-exo group）；and b.End3 cells+25 μg/mL M1-exo（b.End3+M1-exo group）. Trans-endothelial electrical resistance（TEER） and Lucifer yellow permeability were examined. The expression of tight junction complex proteins（claudin-1，occludin，zonula occludens-1［ZO-1］，and junctional adhesion molecules） was measured by Western blot.Results Microglia were successfully polarized to the M1 type，and the positivity rate of the M1 marker CD16/32 was significantly increased compared with the control group（P=0.023）. After co-culture with M1-exo，the TEER of the in-vitro BBB model was significantly decreased（P<0.001），and Lucifer yellow permeability was significantly increased（P<0.001）. Compared with the b.End3 group and b.End3+BV2-exo group，the b.End3+M1-exo group showed significantly decreased expression levels of claudin-1，occludin，and ZO-1.Conclusion M1-exo can disrupt the integrity of BBB and interfere with its normal function.