Deactivation of ceramide de novo synthesis induces cerebral angiogenesis and microvascular remodeling post-ischemia/reperfusion in mice via mechanisms not predominated by extracellular vesicles

Ceramide is a major constituent of membrane microdomains and controls cell signaling. Previous studies showed that ceramide exacerbates cerebral ischemia/reperfusion injury and that inhibition of acid sphingomyelinase (ASM), which converts sphingomyelin to ceramide, promotes microvascular remodeling via induction of extracellular vesicles (EVs) release. ASM inhibitors have so far not translated into clinical stroke therapies. Of note, ceramide levels can also be lowered by inhibition of serine palmitoyltransferase (SPT), which is the rate-limiting enzyme of ceramide de novo synthesis. To evaluate this pathway as a potential therapeutic target for stroke, we exposed cerebral microvascular endothelial cells to the SPT inhibitor myriocin or induced SPT knockdown by siRNA. Deactivation of de novo ceramide synthesis increased endothelial tube formation, migration, VEGF secretion, and EV release. These EVs did not induce angiogenesis in tube formation assays. Using light sheet microscopy, we demonstrate that myriocin-mediated SPT inhibition increases the length and branch density of thin microvessels (<4 µm diameter), which are drivers of angiogenesis, in the previously ischemic brain tissue of mice exposed to middle cerebral artery occlusion. These data highlight the importance of ceramide de novo synthesis for post-ischemic microvascular remodeling and angiogenesis, while its mode of action differs from ASM inhibitors.