14,15-EET Reduced Brain Injury from Cerebral Ischemia and Reperfusion via Suppressing Neuronal Parthanatos.

To investigate the effect of 14,15-EET on the parthanatos in neurons induced by cerebral ischemia and reperfusion, middle cerebral artery occlusion and reperfusion (MCAO/R) and oxygen glucose deprivation/reoxygenation (OGD/R) were used to simulate cerebral ischemia reperfusion in vivo and in vitro, respectively. TTC staining and the Tunel method were used to detect cerebral infarct volume and neuronal apoptosis. Western blot and immunofluorescence were used to detect poly (ADP-ribose) polymerase-1 (PARP-1) activation and AIF nuclear translocation. The production of reactive oxygen species (ROS) and the expression of antioxidant genes were detected by Mito SOX, DCFH-DA and qPCR methods. MCAO/R increased cerebral infarct volume and neuronal apoptosis in mice, while 14,15-EET pretreatment increased cerebral infarct volume and neuronal apoptosis. OGD/R induced reactive oxygen species generation, PARP-1 cleavage, and AIF nuclear translocation in cortical neurons. 14,15-EET pretreatment could enhance the antioxidant gene expression of glutathione peroxidase (GSH-Px), heme oxygenase-1 (HO-1) and superoxide dismutase (SOD) in cortical neurons after ischemia and reperfusion. 14,15-EET inhibits the neuronal parthanatos induced by MCAO/R through upregulation of the expression of antioxidant genes and by reducing the generation of reactive oxygen species. This study advances the EET neuroprotection theory and provides a scientific basis for targeted clinical drugs that reduce neuronal parthanatos following cerebral ischemia and reperfusion.

Effect of 14,15-EET on Release of BDNF from Astrocytes and Neuronal Apoptosis in a Co-culture System after Oxygen-glucose Deprivation and Reperfusion / 华中科技大学学报(医学版)

Objective@#To investigate whether post-treatment of cultured astrocytes with 14,15-epoxyeicosatrienoic acids（14,15-EET）would increase the brain derived neurotrophic factor（BDNF）secretion after oxygen-glucose deprivation and reperfusion（OGD/R）, and if this effect would subsequently protect neurons during reperfusion after OGD in the co-cultured system.@*Methods@#Astrocytes and neurons were subjected to OGD/R. Exogenous 14,15-EET were applied to astrocytes in the reperfusion period and ELISA was then performed to measure BDNF secretion from astrocytes at different time points following OGD/R. After that，the OGD neurons were co-cultured with the astrocytes that were previously incubated with DMSO or 14,15-EET.Tunnel staining was used to detect neuronal apoptosis.@*Results@#BDNF secretion was significantly promoted by application of 14,15-EET on astrocytes in the reperfusion stage after OGD. Exposure of OGD neurons to astrocyte media previously conditioned with 14,15-EET reduced the neuronal apoptosis，but the pro-survival effect could be partly reversed by TrkB inhibitor k252a.@*Conclusion@#Exogenous administration of 14,15-EET augments BDNF secretion from astrocytes，which increases TrkB receptor occupancy on neurons and promotes neuronal survival after OGD/R.

Effect of 14,15-EET on Inflammatory Responses of BV2 Cells After Oxygen and Glucose Depriviation/Reoxygenation / 医药导报

Objective To explore the effect of 14,15-epoxyeicosatrienoic acids (14,15-EET) on the inflammatory response of BV2 cells under oxygen and glucose depriviation/reoxygenation (OGD/R) conditions.Methods BV2 cells were randomly divided into three groups,blank control group,vehicle control group,and 14,15-EET group.Under treatment of 14,15-EET,the concentration of inflammatory factor in BV2 cell culture media was detected by ELISA at different time points (reoxygenation for 0,3,6,12,24 h) after OGD1h.The viability of BV2 cells was detected by MTT assay at different time points.At the same conditions,using Transwell migration experiment,migration ability of BV2 cells was observed.Results The 14,15-EET group had the lower levels of inflammatory factor secretion,lower viability and weaker ability of migration than the vehicle control group.The above results were most statistically significant at OGD1h/R12h.Conclusion 14,15-EET can inhibit the inflammation of BV2 cells induced by the injury of OGD reperfusion.

14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis.

14,15-Epoxyeicosatrienoic acid (14,15-EET), a metabolite of arachidonic acid, is enriched in the brain cortex and exerts protective effect against neuronal apoptosis induced by ischemia/reperfusion. Although apoptosis has been well recognized to be closely associated with mitochondrial biogenesis and function, it is still unclear whether the neuroprotective effect of 14,15-EET is mediated by promotion of mitochondrial biogenesis and function in cortical neurons under the condition of oxygen-glucose deprivation (OGD). In this study, we found that 14,15-EET improved cell viability and inhibited apoptosis of cortical neurons. 14,15-EET significantly increased the mitochondrial mass and the ratio of mitochondrial DNA to nuclear DNA. Key makers of mitochondrial biogenesis, peroxisome proliferator activator receptor gamma-coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM), were elevated at both mRNA and protein levels in the cortical neurons treated with 14,15-EET. Moreover, 14,15-EET markedly attenuated the decline of mitochondrial membrane potential, reduced ROS, while increased ATP synthesis. Knockdown of cAMP-response element binding protein (CREB) by siRNA blunted the up-regulation of PGC-1α and NRF-1 stimulated by 14,15-EET, and consequently abolished the neuroprotective effect of 14,15-EET. Our results indicate that 14,15-EET protects neurons from OGD-induced apoptosis by promoting mitochondrial biogenesis and function through CREB mediated activation of PGC-1α and NRF-1.