Secretion of interleukin‐1β by astrocytes mediates endothelin‐1 and tumour necrosis factor‐α effects on human brain microvascular endothelial cell permeability

Evidence suggests that endothelin‐1 (ET‐1) plays an essential role in brain inflammation. However, whether ET‐1 contributes directly to blood–brain barrier (BBB) breakdown remains to be elucidated. Using an in vitro BBB model consisting of co‐cultures of human primary astrocytes and brain microvascular endothelial cells (BMVECs), we first investigated the expression of ET‐1 by BMVECs upon stimulation with tumour necrosis factor (TNF)‐α, which plays an essential role in the induction and synthesis of ET‐1 during systemic inflammatory responses. Increased ET‐1 mRNA was detected in the human BMVECs 24 h after TNF‐α treatment. This was correlated with an increase in ET‐1 levels in the culture medium, as determined by sandwich immunoassay. Both TNF‐α and ET‐1 increased the permeability of human BMVECs to a paracellular tracer, sucrose, but only in the presence of astrocytes. The increase in BMVEC permeability by TNF‐α was partially prevented by antibody neutralization of ET‐1 and completely by monoclonal antibody against IL‐1β. Concomitantly, TNF‐α induced IL‐1β mRNA expression by astrocytes in co‐culture and this effect was partially prevented by ET‐1 antibody neutralization. In parallel experiments, treatment of human primary astrocytes in single cultures with ET‐1 for 24 h induced IL‐1β mRNA synthesis and IL‐1β protein secretion in the cell culture supernatant. Taken together, these results provide evidence for paracrine actions involving ET‐1, TNF‐α and IL‐1β between human astrocytes and BMVECs, which may play a central role in BBB breakdown during CNS inflammation.