n-oleoylethanolamine and Reperfusion-Injury

Endocannabinoids alter permeability at various epithelial barriers, and cannabinoid receptors and endocannabinoid levels are elevated by stroke, with potential neuroprotective effects. We therefore explored the role of endocannabinoids in modulating blood-brain barrier (BBB) permeability in normal conditions and in an ischaemia/reperfusion model.. Human brain microvascular endothelial cell and astrocyte co-cultures modelled the BBB. Ischaemia was modelled by oxygen-glucose deprivation (OGD) and permeability was measured by transepithelial electrical resistance. Endocannabinoids or endocannabinoid-like compounds were assessed for their ability to modulate baseline permeability or OGD-induced hyperpermeability. Target sites of action were investigated using receptor antagonists and subsequently identified with real-time PCR.. Anandamide (10 μM) and oleoylethanolamide (OEA, 10 μM) decreased BBB permeability (i.e. increased resistance). This was mediated by cannabinoid CB2 receptors, transient receptor potential vanilloid 1 (TRPV1) channels, calcitonin gene-regulated peptide (CGRP) receptor (anandamide only) and PPARα (OEA only). Application of OEA, palmitoylethanolamide (both PPARα mediated) or virodhamine (all 10 μM) decreased the OGD-induced increase in permeability during reperfusion. 2-Arachidonoyl glycerol, noladin ether and oleamide did not affect BBB permeability in normal or OGD conditions. N-arachidonoyl-dopamine increased permeability through a cytotoxic mechanism. PPARα and γ, CB1 receptors, TRPV1 channels and CGRP receptors were expressed in both cell types, but mRNA for CB2 receptors was only present in astrocytes.. The endocannabinoids may play an important modulatory role in normal BBB physiology, and also afford protection to the BBB during ischaemic stroke, through a number of target sites.

Topics: Arachidonic Acids; Astrocytes; Blood-Brain Barrier; Cells, Cultured; Coculture Techniques; Electric Impedance; Endocannabinoids; Endothelial Cells; Humans; Oleic Acids; Permeability; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB2; Reperfusion Injury; RNA, Messenger

Oleoylethanolamide (OEA) is a high-affinity agonist of peroxisome proliferator-activated receptor α (PPARα) which may act as an endogenous neuroprotective factor. However, it is not clear whether orally administered OEA is effective against ischemic brain injury. In our study, transient focal cerebral ischemia was induced by middle cerebral artery occlusion for 90 min followed by reperfusion. To evaluate its preventive effects, OEA (10, 20 or 40 mg/kg, ig) was administered for 3 days before ischemia. To evaluate its therapeutic effects, OEA (40 mg/kg, ig) was administered at 0.5 or 1h before reperfusion or at 0 or 1h after reperfusion. In some experiments, the PPARα antagonist MK886 (10mg/kg, ig) was administered 0.5h before OEA. Neurological deficit score, infarct volume and brain edema degree were determined at 24h after reperfusion. Blood-brain barrier (BBB) disruption was evaluated by Evans blue (EB) leakage at 6h after reperfusion. Real-time RT-PCR and western blot were performed to detect PPARα mRNA and protein expression. Oral OEA pretreatment improved neurological dysfunction reduced infarct volume and alleviated brain edema in a dose-dependent manner; the most effective dose was 40 mg/kg. The therapeutic time is within 1h after reperfusion. OEA also increased PPARα mRNA and protein expression in the ischemic brain. The PPARα antagonist MK886 abolished the protective effects of OEA. In conclusion, our results indicate that orally administered OEA protects against acute cerebral ischemic injury in mice, at least in part by activating PPARα.

Topics: Administration, Oral; Animals; Blood-Brain Barrier; Brain; Brain Edema; Brain Ischemia; Dose-Response Relationship, Drug; Drug Evaluation; Endocannabinoids; Male; Mice; Oleic Acids; PPAR alpha; Reperfusion Injury

The molecular mechanisms of hepatic ischemia/reperfusion (I/R) damage are incompletely understood. We investigated the role of ceramide in a murine model of warm hepatic I/R injury. This sphingolipid induces cell death and participates in tumor necrosis factor (TNF) signaling. Hepatic ceramide levels transiently increased after the reperfusion phase of the ischemic liver in mice, because of an early activation of acidic sphingomyelinase (ASMase) followed by acid ceramidase stimulation. In vivo administration of an ASMase inhibitor, imipramine, or ASMase knockdown by siRNA decreased ceramide generation during I/R, and attenuated serum ALT levels, hepatocellular necrosis, cytochrome c release, and caspase-3 activation. ASMase-induced ceramide generation activated JNK resulting in BimL phosphorylation and translocation to mitochondria, as the inhibition of ASMase by imipramine prevented these events. In contrast, blockade of ceramide catabolism by N-oleyolethanolamine (NOE), a ceramidase inhibitor, enhanced ceramide levels and potentiated I/R injury compared with vehicle-treated mice. Pentoxifylline treatment prevented TNF upregulation and ASMase activation. Furthermore, 9 of 11 mice treated with imipramine survived 7 days after total liver ischemia, compared with 4 of 12 vehicle-treated mice, whereas 8 of 8 NOE-treated mice died within 2 days of total liver ischemia. In conclusion, ceramide generated from ASMase plays a key role in I/R-induced liver damage, and its modulation may be of therapeutic relevance.

Topics: Animals; Blotting, Western; Disease Models, Animal; Down-Regulation; Endocannabinoids; Enzyme Inhibitors; Ethanolamines; Follow-Up Studies; Galactosylgalactosylglucosylceramidase; Liver; Mice; Mice, Inbred C57BL; Oleic Acids; Polymerase Chain Reaction; Reperfusion Injury; RNA, Messenger; Sphingomyelin Phosphodiesterase