Breakdown of blood–brain barrier, astrocytic endfeet swelling, and abnormal behaviors by blockade of TROY signaling in TROY-Fc transgenic mice

Abstract

An orphan receptor of tumor necrosis factor receptor superfamily (TNFRSF), TROY (TNFRSF19), forms the receptor complex with Nogo-66 receptor/LINGO-1 and DR6 in neurons and cerebral endothelial cells, respectively. Although TROY is expressed in astrocytes of the brain under normal conditions, its function is still unknown. Here, we demonstrated that TROY was strongly expressed in astrocytes rather than in neurons and endothelial cells in the adult mouse brain under normal conditions. Recombinant soluble form of the extracellular domain (ECD) of mouse TROY (sTROY) bound to megalencephalic leukoencephalopathy with subcortical cysts 1 (Mlc1)-positive astrocytes, endfeet of which were surrounding large vessels, in the adult mouse brain. These findings suggest the presence of an unidentified ligand that directly binds to TROY in such astrocytes. To antagonize TROY ligand(s) in vivo, we generated transgenic (Tg) mice overexpressing a chimeric protein consisting of the ECD of mouse TROY and the Fc domain of human immunoglobulin G₁ (TROY-Fc). Adult TROY-Fc Tg mice showed the enhanced permeability of the blood–brain barrier (BBB), which is consistent with TROY-deficient mice, and swollen astrocytic endfeet. The expressions of tight junction proteins (claudin-5, claudin-12, occludin, and zonula occludens-1) and src-suppressed C-kinase substrate (SSeCKS) were reduced in adult Tg mice, whereas aquaporin-4 (AQP4) expression was increased. In postnatal Tg mice, the expressions of vascular endothelial growth factor and AQP4 were increased in TROY-Fc Tg mice, whereas angiopoietin-1 and SSeCKS were reduced. There was no significant change in the expression of endogenous TROY in postnatal and adult Tg mice. Behavioral tests revealed that TROY-Fc Tg mice exhibited a reduction in exploratory motivation and an enhancement of responses to stress. Thus, the signaling mediated by unidentified TROY ligand(s)-TROY interaction may be essential for proper neuronal function through the maintenance of intact BBB.