12-hydroxy-5-8-10-14-eicosatetraenoic-acid and Subarachnoid-Hemorrhage

The monohydroxylated metabolite of arachidonic acid, 20-hydroxyeicosatetraenoic acid (20-HETE), is a potent vasoconstrictor of cerebral microvessels. 20-HETE formation is substantially elevated in the cerebral spinal fluid (CSF) in the rat subarachnoid hemorrhage (SAH) model. The presence of 20-HETE in human CSF has not been demonstrated. Therefore, it was the purpose of this study to determine if HETE metabolites are present in human CSF after SAH.. CSF samples were collected daily from four SAH patients over 15 days. HETE metabolites were separated by HPLC with identification by ion-trap MS/MS and quantification via single quadrupole MS operating in negative single ion monitoring mode.. Two major metabolites were identified as 12-HETE and 20-HETE. 20-HETE maximal concentrations were 2.9 and 0.7 ng/ml at approximately 70 h in the two patients with symptomatic cerebral vasospasm (SV) after SAH. Concentrations of 12-HETE in these patients peaked at 21.9 ng/ml and 2.8 ng/ml. Concentrations of 20-HETE and 12-HETE were non-detectible in the majority of the samples obtained from two matched SAH patients without SV.. This study is the first to demonstrate that 20-HETE and 12-HETE are present in the CSF of SAH patients at physiologically relevant concentrations. Based on this information future prospective studies will allow for the delineation of the role of these metabolites in the pathogenesis of SAH.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Adult; Aged; Arachidonic Acid; Brain Ischemia; Cerebral Arteries; Cerebrospinal Fluid; Chromatography, High Pressure Liquid; Humans; Hydroxyeicosatetraenoic Acids; Mass Spectrometry; Middle Aged; Neurochemistry; Subarachnoid Hemorrhage; Time Factors; Vasospasm, Intracranial

A possible mechanism for the induction of protein kinase C (PKC)-dependent vascular contraction independent to the increase of intracellular Ca++ was investigated in the pathogenesis of cerebral vasospasm in the double subarachnoid haemorrhage (SAH) model. The level of 1,2-diacylglycerol (DAG), which is an intrinsic PKC activator, significantly increased from days 4 to 7 in the basilar artery after the initial SAH, and the continuous administration of 1,2-bis(nicotinamido)-propane (AVS), a novel free radical scavenger, not only lowered the concentration of lipid peroxides in the CSF but also successfully suppressed the basilar arterial wall in the same model. It was suggested that lipid peroxides generated in the subarachnoid clot affect the DAG content of the cerebral artery. Analysis of hydroxy-eicosatetraenoic acids (HETEs) with high performance liquid chromatography (HPLC) revealed the production of relatively large amount of 12-HETE in the subarachnoid clot. To examine the potential effect of exogenous 12-HETE on the DAG content of the cerebral artery, the basilar artery was incubated with 12-HETE in vitro. 12-HETE induced a concentration-dependent slow increase in DAG content in the arterial wall after 6 hours of incubation. Under conditions in which DAG formation was facilitated by the Ca(++)-ionophore, DAG accumulation in the basilar artery was enhanced in the presence of 12-HETE. It was suggested that 12-HETE generated in the subarachnoid clot, induced DAG accumulation in the arterial wall by inhibition of DAG metabolism, resulting in the induction of prolonged PKC-dependent smooth muscle contraction in the pathogenesis of cerebral vasospasm.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Basilar Artery; Diglycerides; Dogs; Female; Hydroxyeicosatetraenoic Acids; Ischemic Attack, Transient; Lipid Peroxidation; Male; Muscle, Smooth, Vascular; Protein Kinase C; Subarachnoid Hemorrhage; Vasoconstriction

To examine the possible involvement of lipoxygenase products from arachidonic acid in the pathogenesis of delayed vasospasm after subarachnoid hemorrhage (SAH), we measured the contents of hydroxyeicosatetraenoic acids (HETEs) in the subarachnoid clot, the cerebrospinal fluid, and the basilar artery, using the canine "two-hemorrhage" model. Lipoxygenase activity in the subarachnoid clot and the basilar artery was measured, ex vivo, using samples obtained 7 days after SAH. For a quantitative analysis of HETEs, each sample was homogenized with either ice-cold saline or methanol. The lipid extract was then submitted to reverse-phase HPLC. The identity of each HETE was further confirmed using straight-phase HPLC and gas chromatography-mass spectrometry. When the basilar artery was homogenized with ice-cold saline, a significant increase in the 5-HETE content was observed on SAH day 8. However, when the artery was homogenized with methanol, HETEs were not detected. In the case of incubation in the presence of arachidonic acid and calcium ionophore A23187, the 5-lipoxygenase activity was remarkably increased in the basilar artery exposed to SAH, compared to that of normal dogs. The subarachnoid clot contained a significant amount of 12-HETE (average 1.8 nmol/g wet weight) from day 2 to day 8. The administration of 1,2-bis(nicotinamido)propane significantly ameliorated vasospasm in the two-hemorrhage model, simultaneously inhibiting the 5-lipoxygenase activity of the basilar artery. Our observations show that the activities of 12- and 5-lipoxygenases are significantly increased after SAH in the subarachnoid clot and the basilar artery, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arachidonic Acid; Arachidonic Acids; Basilar Artery; Chromatography, High Pressure Liquid; Dogs; Gas Chromatography-Mass Spectrometry; Hydroxyeicosatetraenoic Acids; Ischemic Attack, Transient; Lipoxygenase; Subarachnoid Hemorrhage