Activation of PAR1 contributes to ferroptosis of Schwann cells and inhibits regeneration of myelin sheath after sciatic nerve crush injury in rats via Hippo-YAP/ACSL4 pathway

Peripheral nerve injury (PNI) is characterized by high incidence and sequela rate. Recently, there was increasing evidence that has shown ferroptosis may impede functional recovery. Our objective is to explore the novel mechanism that regulates ferroptosis after PNI. LC-MS/MS proteomics was used to explore the possible differential signals, while PCR array was performed to investigate the differential factors. Besides, we also tried to activate or inhibit the key factors and then observe the level of ferroptosis. Regeneration of myelin sheath was finally examined in vivo via transmission electron microscopy. Proteomics analysis suggested coagulation signal was activated after sciatic nerve crush injury, in which high expression of F2 (encoding thrombin) and F2r (encoding PAR1) were observed. Both thrombin and PAR1-targeted activator TRAP6 can induce ferroptosis in RSC96 cells, which can be rescued by Vorapaxar (PAR1 targeted inhibitor) in vitro. Further PCR array revealed that activation of PAR1 induced ferroptosis in RSC96 cells by increasing expression of YAP and ACSL4. Immunofluorescence of sciatic nerve confirmed that the expression of YAP and ACSL4 were simultaneously reduced after PAR1 inhibition, which may contribute to myelin regeneration after injury in SD rats. Inhibition of PAR1 can relieve ferroptosis after sciatic nerve crush injury in SD rats through Hippo-YAP/ACSL4 pathway, thereby regulating myelin regeneration after injury. In summary, PAR1/Hippo-YAP/ACSL4 pathway may be a promising therapeutic target for promoting functional recovery post-sciatic crush injury. Crush injury triggers the local tissue release thrombin, which cleaves the N-terminal peptide of its receptor PAR1 and activates the transmembrane core of PAR1, and finally lead to “turn-off” the Hippo signaling. The phosphorylation cascade of MST1/2 and LATS1/2 is halted, thus significantly reducing the phosphorylation level of YAP. Unphosphorylated YAP molecules will not be degraded, so more YAP can enter the nucleus and work with other transcriptional co-factors to promote the transcription of downstream molecule ACSL4, finally promoting the ferroptosis in SC after crush injury. [Display omitted] •Thrombin induces ferroptosis by activating PAR1.•PAR1/Hippo-YAP/ACSL4 pathway may be a potential therapeutic target for PNI.