Peroxynitrite‑mediated mitophagy could be a crucial therapeutic target for reducing cerebral ischemia–reperfusion injury

Abstract

Basic autophagy/mitophagy is essential for cell survival whereas excessive autophagy/mitophagy is detrimental during cerebral ischemia–reperfusion (I/R) injury. Peroxynitrite (ONOO−), a representative of reactive nitrogen species, is a critical neurotoxic factor in mediating cerebral I/R injury, but its roles in autophagy/mitophagy remain unclear. Herein, we hypothesized that ONOO− could induce PINK1/Parkin-mediated mitophagy activation via triggering dynamin-related protein 1 (Drp1) recruitment to damaged mitochondria, contributing to cerebral I/R injury. The major discoveries revealed that: (1) PINK1/Parkin-mediated mitophagy activation was predominant among general autophagy, leading to rat brain injury at the reperfusion phase after cerebral ischemia; (2) increased nitrotyrosine was found in the plasma of ischemic stroke patients and ischemia–reperfused rat brains, indicating the generation of ONOO− in ischemic stroke; (3) ONOO− was dramatically increased in accompanied with mitochondrial recruitment of Drp1, PINK1/Parkin-mediated mitophagy activation, and progressive infarct size in rat ischemic brains at the reperfusion phase; (4) FeTMPyP, a peroxynitrite decomposition catalyst, remarkably reversed mitochondrial recruitment of Drp1, mitophagy activation and brain injury; (5) ONOO− induced tyrosine nitration of Drp1 peptide and mitochondrial recruitment of Drp1 for mitophagy activation. Those results suggest that ONOO−-induced mitophagy activation aggravates cerebral I/R injury via recruiting Drp1 to damaged mitochondria. Furthermore, we investigated ONOO−-induced mitophagy as a therapeutic target for attenuating cerebral I/R injury by using a natural antioxidant naringin as an example. Naringin possessed strong ONOO− scavenging capability and inhibited the production of superoxide and nitric oxide in SH-SY5Y cells under 10 h oxygen-glucose-deprivation plus 14 h of reoxygenation or ONOO− donor 3-morpholinosydnonimine conditions. Naringin also inhibited NADPH oxidases and iNOS in rat brains with 2 h ischemia plus 22 h reperfusion. Naringin was able to cross the blood–brain barrier, decreased neurological deficit score and infarct size, and attenuated apoptotic cell death. Naringin reduced 3-nitrotyrosine formation, decreased the ratio of LC3-II to LC3-I in mitochondrial fraction, and inhibited the translocation of Parkin to the mitochondria. Taken together, peroxynitrite-mediated mitophagy activation could be a potential therapeutic target for cerebral I/R injury.