TIMP1 protects against blood-brain barrier disruption after subarachnoid haemorrhage by inhibiting ubiquitination of astrocytic β1-integrin

Abstract

BackgroundAstrocytes regulate blood-brain barrier (BBB) integrity, whereas subarachnoid haemorrhage (SAH) results in astrocyte dysregulation and BBB disruption. Here, we explored the involvement of tissue inhibitor of matrix metalloprotease-1 (TIMP1) in astrocyte-mediated BBB protection during SAH, along with its underlying mechanisms.MethodsC57BL/6J mice were used to establish a model of SAH. The effects of TIMP1 on SAH outcomes were analysed by intraperitoneal injection of recombinant mouse TIMP1 protein (rm-TIMP1; 250 µg/kg). The roles of TIMP1 and its effector β1-integrin on astrocytes were observed by in vivo transduction with astrocyte-targeted adeno-associated virus carrying TIMP1 overexpression plasmid or β1-integrin RNAi. The molecular mechanisms underlying TIMP1 and β1-integrin interactions were explored in primary cultured astrocytes stimulated with red blood cells (RBCs).ResultsTIMP1 was upregulated after SAH. Administration of rm-TIMP1 mitigated SAH-induced early brain injury (EBI) in male and female mice. TIMP1 was primarily expressed in astrocytes; its overexpression in astrocytes led to increased β1-integrin expression in astrocytes, along with the preservation of astrocytic endfoot attachment to the endothelium and subsequent recovery of endothelial tight junctions. All of these effects were reversed by the knockdown of β1-integrin in astrocytes. Molecular analysis showed that TIMP1 overexpression decreased the RBC-induced ubiquitination of β1-integrin; this effect was partially achieved by inhibiting the interaction between β1-integrin and the E3 ubiquitin ligase Trim21.ConclusionTIMP1 inhibits the interaction between β1-integrin and Trim21 in astrocytes, thereby rescuing the ubiquitination of astrocytic β1-integrin. It subsequently restores interactions between astrocytic endfeet and the endothelium, as well as BBB integrity, eventually mitigating SAH-induced EBI.