Abstract
Introduction: Piezo1 is a calcium-permeable mechanosensitive ion channel that is opened by membrane stretch or increased shear force. In peripheral vasculature, activation of endothelial Piezo1 promotes vasodilation. Whether endothelial Piezo1 regulates cerebral blood flow and the consequences of endothelial cell (EC) Piezo1 loss-of-function in the brain are unknown. The present study was designed to test the overall hypothesis that loss of EC Piezo1 results in reduced cerebral blood flow and subsequent brain pathology consistent with chronic hypoperfusion. Methods: EC specific Piezo1 knockout mice were generated by crossing Piezo1 fl/fl mice with inducible EC-specific Cre mice (Cdh5(Pac)CreERT2). Tamoxifen (75 mg/kg) was administrated to generate EC Piezo1 KO or as control in Piezo1 “floxed only” mice. At 1 month after tamoxifen injection, three cohorts were evaluated: (cohort 1) measure CBF by MRI ASL, (cohort 2) gait analysis (Digigait) and subsequent immunofluorescence (IF), (cohort 3) scRNAseq analysis. Gait analysis was chosen as a behavioral measure sensitive to white matter injury. Results: EC Piezo1 KO resulted in >8% reduction in resting CBF in striatum (n=5; p<0.05, females). Gait analysis showed significant change in stride time, stride length, and cadence (n = 9-10 males, 6-9 females; p< 0.01). IF showed gliosis in the corpus callosum of EC Piezo1 KO mice, in the form of astrogliosis (Gfap) in males and microgliosis (Iba1) in females (n=5-7 in each group; p<0.05). From scRNAseq of EC Piezo1 KO and control brains (males), GO analysis from all clusters revealed GO terms including: inflammatory response, synapse pruning, gliogenesis, learning or memory, and locomotory behavior. GO analysis from EC clusters revealed GO terms including: gliogenesis, myelination, and vascular morphogenesis. Among the most highly upregulated genes in the EC cluster, were Ctss and C1q family genes. In choroid plexus cluster, EC Piezo1 KO showed increased expression of Hif1a target genes. Conclusions: We conclude that EC Piezo1 loss-of-function promotes hypoperfusion sufficient to induce induction of inflammatory signaling and hypoxia-dependent transcription. White matter injury was evident as gliosis in the corpus callosum and by alterations in gait.
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