NLPR3 inflammasome inhibition alleviates hypoxic endothelial cell death in vitro and protects blood–brain barrier integrity in murine stroke

• Published in: Cell death & disease Vol. 13; no. 1; pp. 20 - 10
• Main Authors: Bellut, Maximilian, Papp, Lena, Bieber, Michael, Kraft, Peter, Stoll, Guido, Schuhmann, Michael K.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 20.12.2021; Springer Nature B.V; Nature Publishing Group
• Subjects: 13/1; 13/31; 13/51; 14/1; 14/28; 14/63; 38/1; 692/308/2778; 692/420/256/2177; 692/699/375/534; Animals; Antibodies; Apoptosis; Biochemistry; Biomedical and Life Sciences; Blood-brain barrier; Blood-Brain Barrier - drug effects; Blood-Brain Barrier - metabolism; Brain; Brain Infarction - drug therapy; Brain Infarction - metabolism; Carotid arteries; Caspase-1; Cell Biology; Cell Culture; Cell death; Cell Hypoxia - drug effects; Cells, Cultured; Cerebral blood flow; Chemokines; Cytokines; Disease Models, Animal; Edema; Endothelial cells; Endothelial Cells - drug effects; Endothelial Cells - metabolism; Experiments; Furans - administration & dosage; Gelatinase B; Glucose; Glucose - metabolism; Hemorrhage; Humidity; Hypoxia; Immunology; Indenes - administration & dosage; Inflammasomes; Inflammasomes - antagonists & inhibitors; Inflammasomes - metabolism; Inflammation; Injections, Intraperitoneal; Ischemia; Ischemic Stroke - drug therapy; Ischemic Stroke - metabolism; Laboratory animals; Life Sciences; Male; Matrix metalloproteinase; Metalloproteinase; Mice; Mice, Inbred C57BL; NLR Family, Pyrin Domain-Containing 3 Protein - antagonists & inhibitors; NLR Family, Pyrin Domain-Containing 3 Protein - metabolism; Protective Agents - administration & dosage; Proteins; Pyrin protein; Pyroptosis; Pyroptosis - drug effects; Reperfusion; Signal Transduction - drug effects; Stroke; Sulfonamides - administration & dosage; Treatment Outcome; Veins & arteries
• ISSN: 2041-4889; 2041-4889
• DOI: 10.1038/s41419-021-04379-z

In ischemic stroke (IS) impairment of the blood–brain barrier (BBB) has an important role in the secondary deterioration of neurological function. BBB disruption is associated with ischemia-induced inflammation, brain edema formation, and hemorrhagic infarct transformation, but the underlying mechanisms are incompletely understood. Dysfunction of endothelial cells (EC) may play a central role in this process. Although neuronal NLR-family pyrin domain-containing protein 3 (NLRP3) inflammasome upregulation is an established trigger of inflammation in IS, the contribution of its expression in EC is unclear. We here used brain EC, exposed them to oxygen and glucose deprivation (OGD) in vitro, and analyzed their survival depending on inflammasome inhibition with the NLRP3-specific drug MCC950. During OGD, EC death could significantly be reduced when targeting NLRP3, concomitant with diminished endothelial NLRP3 expression. Furthermore, MCC950 led to reduced levels of Caspase 1 (p20) and activated Gasdermin D as markers for pyroptosis. Moreover, inflammasome inhibition reduced the secretion of pro-inflammatory chemokines, cytokines, and matrix metalloproteinase-9 (MMP9) in EC. In a translational approach, IS was induced in C57Bl/6 mice by 60 mins transient middle cerebral artery occlusion and 23 hours of reperfusion. Stroke volume, functional outcome, the BBB integrity, and—in good agreement with the in vitro results—MMP9 secretion as well as EC survival improved significantly in MCC950-treated mice. In conclusion, our results establish the NLRP3 inflammasome as a critical pathogenic effector of stroke-induced BBB disruption by activating inflammatory signaling cascades and pyroptosis in brain EC.