MST1/Drp1 axis mediates microglia pro-inflammatory activation following cerebral ischemia-reperfusion injury

Abstract The present study investigated the role of the mammalian sterile 20-like kinase 1/dynamin-related protein 1 (MST1/Drp1) axis in regulating microglia pro-inflammatory activation during cerebral ischemia-reperfusion injury (CIRI). An in vivo model of middle cerebral artery occlusion/reperfusion (MCAO/R) in rats and an in vitro oxygen-glucose deprivation/reoxygenation (OGD/R) model in BV-2 microglial cells and primary microglia were established. Inhibitors of MST1 (XMU-MP-1) or/and Drp1 (Mdivi-1), along with genetic approaches including siMST1-mediated knockdown and plasmid-based overexpression, were utilized in the models. The expression and activation of MST1 and Drp1, mitochondrial morphology changes, microglia pro-inflammatory activation makers, pro-inflammatory cytokine release, DNA fragmentation and neurological function were evaluated. The findings indicated that reperfusion or reoxygenation led to a rise in total and phosphorylation levels of MST1 and Drp1. The reperfusion also facilitated the Drp1 translocation toward mitochondria, and resulted in increased mitochondrial morphological changes. MST1 or/and Drp1 inhibitors decreased p-MST1 and p-Drp1(Ser616) levels, attenuated mitochondrial fission, suppressed microglia pro-inflammatory activation, pro-inflammatory factors release (TNF-α, IL-6 and IL-1β). Overall, these effects ultimately mitigated cerebral injury as evidenced by reduced DNA fragmentation, decreased cerebral infarct volumes, and improved neurological function. Combined inhibitors further exerted ameliorative effects on the above-mentioned parameters. In the in vitro experiments, siMST1 knockdown attenuated p-Drp1(Ser616) expression and suppressed microglia pro-inflammatory activation under OGD/R conditions. These protective effects were reversed by Drp1 overexpression. These findings indicate that p-MST1 drives microglia pro-inflammatory activation via promoting the p-Drp1(Ser616)-mediated excessive mitochondrial fission during CIRI.