COX-1 and COX-2 Contributions to Basal and IL-1β-Stimulated Prostanoid Synthesis in Human Neonatal Cerebral Microvascular Endothelial Cells

Abstract

Mechanisms of endothelium-dependent regulation of cerebral circulation in human neonates are poorly understood owing to the lack of experimental data. Prostanoids, the products of the cyclooxygenase (COX) pathway, appear to be important regulators of blood flow in neonates. COX activity in cultured endothelial cells from small (60 -300 m) and large (Ͼ300 m) microvessels from the autopsy specimens of neonatal human cerebral cortex and cerebellum (22-26 wk gestational age) was detected as production of vasodilator prostanoids, prostacyclin [as 6-keto-prostaglandin (PG) F 1␣ ] and PGE 2 from arachidonic acid. Treatment of neonatal human cerebral microvascular endothelial cells (hCMVEC) with IL-1␤ (50 ng/mL, 17 h) stimulated COX activity 5-to 20-fold. Basal and IL-1␤-stimulated COX activities were inhibited by NS-398, indicating substantial COX-2 contribution to endothelial prostanoid synthesis in neonatal human brain cortex and cerebellum at rest and when mimicking the inflammatory conditions. Increased COX-2-linked activity in response to IL-1␤ was observed in hCMVEC from both cerebrum and cerebellum (5-to 20-fold), while under the same conditions elevated COX-1-linked activity was detected only in hCMVEC from cerebellum (5-to 10-fold). In IL-1␤-treated hCMVEC, a shift toward PGE 2 as the major vasodilator product of the COX pathway was observed. Acute treatment with the protein tyrosine kinase inhibitor, tyrphostin 25, inhibited basal and IL-1␤-induced COX activities, suggesting the importance of posttranslational modifications in endothelial COX-2 activation in human brain. Altogether, these data indicate that both COX-1 and COX-2 contribute to endothelial prostanoid synthesis in the neonatal human brain under basal conditions and in response to proinflammatory cytokine IL-1␤. Cerebral circulatory disorders produce serious complications in human neonates, such as cerebral hemorrhages and hypoxicischemic injuries, that can result in life-long neurologic dysfunction in survivors including cerebral palsy, epilepsy, and mental retardation. Mechanisms of the regulation of cerebral circulation in neonates are understood incompletely, and experimental data are very limited. Vascular endothelium-derived dilator prostanoids, NO, and carbon monoxide are vasorelaxant factors essential in the regulation of cerebral blood flow. There are speciesspecific and age-dependent differences in physiologically relevant endothelial vasorelaxants in the cerebral circulation (1). In human newborns, prostanoids appear to be major factors that contribute to regulation of cerebral blood flow in response to hypercapnia, asphyxia, ischemia, and cerebral hemorrhage (1-3), and indomethacin is widely used in neonatal practice (2,4,5). Similarly, in newborn pigs, prostanoids contribute to the regulation of cerebral blood flow under basal conditions and in response to physiologically important stimuli, such as CO 2 , oxygen, and changes in blood pressure (1). Prostanoids are also important in the pathogenesis of inflammation because they alter in...

Dexamethasone Alters Vascular Reactivity by Enhancing COX-Related Vasodilatation in Fetal Ovine Carotids

“…We therefore performed Western blotting analyses to quantify the effect of DEX treatment on COX-1 or -2 protein abundance. As shown in fi gure 4 , we detected both COX-1 and -2 in our control samples, suggesting that both enzymes are constitutively expressed in near-term lamb carotids, similar to observations in human newborns and neonatal pigs [26,27] . However, we found that DEX treatment had no significant effects on COX-1 or -2 protein expression in our samples ( fi g. 4 ).…”

“…This data suggests that DEX may alter the balance between the PLC and phospholipase A 2 pathway, stimulating an increase in vasodilator prostanoids while inhibiting the signal transduction pathway, leading to contraction. Additional mechanisms by which DEX can result in vasodilation are the possible effects of DEX on COX isoform and subcellular localization [34] , post-translational regulation [25], and other pathways downstream from COX, such as the conversion of arachidonate to prostaglandin E 2 [27] and the activities of other arachidonic metabolites such as epoxyeicosatrienoic [35] or leukotrienes [36] .…”

Selective Inhibitors Differentially Affect Cyclooxygenase-Dependent Pial Arteriolar Responses in Newborn Pigs

“…The assessed COX-dependent responses mainly require COXproducts from endothelial cells, since selective endothelial injury diminishes the responsiveness to these stimuli (14,15). Indeed, freshly isolated or cultured endothelial cells from piglet brains or autopsy specimens of neonatal human cerebral cortex have also been found to produce prostanoids mainly via the activity of COX-2 (16,17).…”

Cerebrovascular Responses in the Fetal Sheep Brain to Low-Dose Endotoxin

“…This is an important observation because it suggests that macrophage penetration into white matter occurs at lower doses of LPS required to provoke the response elsewhere in the brain. Macrophages respond to a wide range of microbial products such as LPS using a toll-like receptor on their surface (33), initiating host-defense mechanisms by release of cytokines, particularly IL-1 and TNF-␣. Macrophage infiltration is a well-defined response to systemic and CNS infections in the adult brain (26,42), and the presence of activated monocytes of peripheral and in situ origin (i.e.…”