The Role Of PAQR3 In Neuronal Apoptosis In Cerebral Lschemia-Reperfusion

【Objective】Neuronal apoptosis acts as the pivotal pathogenesis of cerebral ischemia/reperfusion(I/R)injury after ischemic stroke.PAQR3(progestin and adipo Q receptor family member 3)is a crucial player who participates in the regulation of cell death.We aim to explore the specific function and the underlying mechanism of PAQR3 in cerebral I/R induced neuronal injury.【Methods】We established a mouse middle cerebral artery occlusion/reperfusion(MCAO/R)model and rat adrenal pheochromocytoma(PC12)cell oxygen-glucose deprivation/reperfusion(OGD/R)model to simulate ischemia-reperfusion injury in vivo and in vitro and detect the expression of PAQR3 after I/R treatment.Meanwhile,immunofluorescence technique was used to detect the co-localization of PAQR3 with PC12 cells and Golgi specific marker GM130.We used lentivirus to knockdown PAQR3 and then investigated the function of PAQR3 in I/R induced neuronal apoptosis through several methods:(1)Cell viability assessment(CCK-8),lactate dehydrogenase(LDH)release assay,Flow cytometric apoptosis assay(Annexin-V/PE)and TUNEL were used to find out the role of PAQR3 in OGD/R-induced neuronal apoptosis;(2)The expression levels of Bax,Bcl-2,caspase-3,activated caspase-3,cytochrome c and activated PARP were detected by western blot technique to further clarify the effect of PAQR3 on neuronal apoptosis induced by OGD/R;(3)The relationship between PAQR3 and PI3K/AKT signaling pathway after OGD/R treatment was studied by western blotting,and we figure out whether PI3K/AKT signaling suppression with LY294002 can abolish the inhibiting neuronal apoptosis effect induced by silencing PAQR3 after OGD/R treatment.【Results】PAQR3 expression is markedly increased in the ischemic hemisphere of C57BL/6 mice and PC12 cells after I/R stimulation.Knockdown PAQR3 can attenuate neuronal apoptosis induced by I/R in PC12 cells and exerts neuroprotective effects.PAQR3 deficiency can significantly raise cell viability and suppress LDH leakage under I/R treatment.Silencing PAQR3 attenuates neuronal apoptosis remarkably with fewer TUNEL-positive cells and lower apoptosis rate under I/R treatment.Mechanistically,knockdown of PAQR3 can inhibit the apoptosis pathway through inducing anti-apoptotic proteins and inhibiting pro-apoptotic proteins.Besides,PI3K/AKT signaling suppression with LY294002 abolished the neuron protection induced by silencing PAQR3.【Conclusion】Our results elucidate that silencing PAQR3 can protect PC12 from OGD/R injury via activating PI3K/AKT pathway.And therefore,provide a novel therapeutic target for the prevention of cerebral I/R injury.