Human blood-brain barrier spheroid models for the evaluation of drugs targeting brain vascular inflammation.

The blood-brain barrier (BBB), primarily formed by brain microvascular endothelial cells (BMECs), restricts substance entry into the brain. BBB inflammation, involving barrier dysfunction and immune cell recruitment, contributes to neurological diseases and represents a therapeutic target. In this study, we characterized human immortalized cell-based multicellular spheroidal BBB (hiMCS-BBB) models as a tool for evaluating effects of BBB inflammation-targeted drugs. First, we examined the responses of the hiMCS-BBB models to the pro-inflammatory cytokine (TNF-α and IFN-γ) exposure. The cytokine treatment reduced claudin-5 protein expression levels at the BBB, accompanied by a 1.4 ± 0.1-fold increase in lucifer yellow permeability. Consistently, E-selectin protein expression was upregulated, accompanied by a 22.1-fold increase in THP-1 cell adhesion to the BBB. Then, we tested whether natalizumab (a monoclonal antibody used for multiple sclerosis treatment) and JPH203 (an inhibitor of L-type amino acid transporter 1) could modulate the immune cell adhesion. Both agents remarkably suppressed the cytokine-induced THP-1 cell adhesion to 0.2-fold relative to the respective controls. To summarize, our results demonstrate that the hiMCS-BBB models exhibit key features of BBB inflammation in response to inflammation stimuli and recapitulate clinically relevant responses to anti-inflammatory drugs, highlighting their potential to accelerate BBB-targeting drug development.