title: 5,6-δ-DHTL, a stable metabolite of arachidonic acid, is a potential EDHF that mediates microvascular dilation.
abstract: Prominent among the endothelium-derived hyperpolarizing factors (EDHFs) are the Cytochrome P450 (CYP) epoxygenase-derived arachidonic acid metabolites-the epoxyeicosatrienoic acids (EETs), that are known as vasodilators in the microcirculation. Among the EET isomers, 5,6-EET undergoes rapid lactonization in aqueous solution to the more stable 5,6-δ DHTL (5,6-dihydroxytrienoic lactone) isomer. It is unclear whether this metabolic transformation maintains its vasodilator potential and what is the mechanism of action. Thus, the aim of this study was to investigate the capacity of the lactone isomer, 5,6- δ DHTL, to induce dilation of arterioles and explore the endothelial Ca2+response mechanism.
  
  
  
  In isolated human microvessels 5,6- δ DHTL induced a dose dependent vasodilation, that was inhibited by mechanical denudation of the endothelial layer. This 5,6- δ DHTL -dependent dilation was partially reduced in the presence of L-NAME (NOS inhibitor) or the NO-scavenger, cPTIO (by 19.7%, which was not statistically significantly). In human endothelial cells, 5,6- δ DHTL induced an increase in intracellular Ca2+([Ca2+]i) in a dose dependent manner. This increase in [Ca2+]i was similar to that induced by the 5,6-EET isomer, and significantly higher than observed by administering the hydrolytic dihydroxy isomer, 5,6-DHET. Further experiments aimed to investigate the mechanism of action revealed, that the 5,6-δ DHTL-mediated ([Ca2+]i elevation was reduced by IP3 and ryanodine antagonists, but not by antagonists to the TRPV4 membrane channel. Similar to their effect on the dilation response in the arteries, NO inhibitors reduced the 5,6-δ DHTL-mediated ([Ca2+]i elevation by 20%. Subsequent 5,6-δ DHTL -dependent K+ion efflux from endothelial cells, was abolished by the inhibition of small and intermediate conductance KCa.
  
  
  
  The present study shows that 5,6-δ DHTL is a potential EDHF, that dilates microvessels through a mechanism that involves endothelial dependent Ca2+entry, requiring endothelial hyperpolarization. These results suggest the existence of additional lactone-containing metabolites that can be derived from the PUFA metabolism and which may function as novel EDHFs.
authors.affiliation: Laboratory of Human Health and Nutrition Sciences, MIGAL-Galilee Research Institute, Ltd., Kiryat Shmona, Israel; Tel-Hai College, Upper Galilee, Israel.
authors.forename: Reut
authors.initials: R
authors.lastname: Levi-Rosenzvig
authors.affiliation: Department of Medicine and Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI, USA; Department of Physiology, Medical College of Wisconsin, Milwaukee, WI, USA.
authors.forename: Andreas M
authors.initials: AM
authors.lastname: Beyer
authors.affiliation: Department of Medicine and Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI, USA.
authors.forename: Joseph
authors.initials: J
authors.lastname: Hockenberry
authors.affiliation: Laboratory of Human Health and Nutrition Sciences, MIGAL-Galilee Research Institute, Ltd., Kiryat Shmona, Israel.
authors.forename: Rotem Shelly
authors.initials: RS
authors.lastname: Ben-Shushan
authors.affiliation: The Baruch Padeh Medical Center, Poriya, Israel.
authors.forename: Dimitry
authors.initials: D
authors.lastname: Chuyun
authors.affiliation: Laboratory of Human Health and Nutrition Sciences, MIGAL-Galilee Research Institute, Ltd., Kiryat Shmona, Israel; Tel-Hai College, Upper Galilee, Israel.
authors.forename: Shahar
authors.initials: S
authors.lastname: Atiya
authors.affiliation: Laboratory of Human Health and Nutrition Sciences, MIGAL-Galilee Research Institute, Ltd., Kiryat Shmona, Israel; Tel-Hai College, Upper Galilee, Israel.
authors.forename: Snait
authors.initials: S
authors.lastname: Tamir
authors.affiliation: Department of Medicine and Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI, USA.
authors.forename: David D
authors.initials: DD
authors.lastname: Gutterman
authors.affiliation: Laboratory of Human Health and Nutrition Sciences, MIGAL-Galilee Research Institute, Ltd., Kiryat Shmona, Israel; Tel-Hai College, Upper Galilee, Israel. Electronic address: andreas@migal.org.il.
authors.forename: Andrea
authors.initials: A
authors.lastname: Szuchman-Sapir
mesh_terms: D001160:Arterioles / Q000187:drug effects; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D004353:Drug Evaluation, Preclinical; D042783:Endothelial Cells / Q000187:drug effects; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000502:physiology; D004789:Enzyme Activation; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D014664:Vasodilation; D014665:Vasodilator Agents / Q000494:pharmacology*
chemical_list: C000625614:5,6-dihydroxytrienoic lactone; D006893:Hydroxyeicosatetraenoic Acids; D014665:Vasodilator Agents; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III
keywords: 5,6-DHTL; Endothelium-dependent hyperpolarization factor; Lactones; Microvessels; Vasodilation