NOTCH3-R544C mutation drives endothelial dysfunction through inflammation, migration impairment, and lipid...

BackgroundCerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is caused by mutations in the NOTCH3 gene. Previous research has predominantly focused on vascular smooth muscle cell pathology, whereas the role of brain microvascular endothelial cells (BMECs) in the disease remains unclear.ObjectiveTo investigate the impact of the NOTCH3-R544C mutation on BMECs function and to elucidate the underlying mechanisms of endothelial dysfunction in CADASIL.MethodsUsing CADASIL transgenic mice and endothelial cell (EC) models stably expressing NOTCH3-R544C, the impact of the mutation on endothelial function was assessed through immunofluorescence staining, RNA-seq analysis, protein-protein interaction (PPI) network mapping, and lipid and cellular function assays.ResultsSignificant NOTCH3 extracellular domain (NOTCH3ECD) deposition and reduced microvascular density are observed in CADASIL mice. R544C mutant cells exhibit abnormal NOTCH3ECD accumulation alongside pronounced gene expression dysregulation, predominantly enriched in pathways related to inflammation, cell migration. The PPI network centers on CXCL10 as a pivotal hub, forming a core "inflammation-migration" pathological axis. R544C cells exhibit heightened inflammatory responses, cholesterol accumulation, reduced cell viability, and increased sensitivity to inflammatory stimuli.ConclusionsThe NOTCH3-R544C mutation disrupts inflammatory regulation, migratory capacity, and lipid metabolic homeostasis in BMECs, leading to endothelial dysfunction and revealing a key mechanism of endothelial injury in CADASIL.