Revising the structure of a new eicosanoid from human platelets to 8,9-11,12-diepoxy-13-hydroxy-eicosadienoic acid

Andrei Kornilov, Paul D. Kennedy, Maceler Aldrovandi, Andrew Watson, Christine Hinz, Bryan Harless, Joseph Colombo, Kirk M. Maxey, Victoria J. Tyrrell, Matthew Simon, Varinder K. Aggarwal, William E. Boeglin, Alan R. Brash, Robert C. Murphy, Valerie B. O'donnell

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Abstract

Eicosanoids are critical mediators of fever, pain and inflammation generated by immune and tissue cells. We recently described a new bioactive eicosanoid generated by cyclooxygenase-1 (COX-1) turnover during platelet activation that can stimulate human neutrophil integrin expression. On the basis of mass spectrometry (MS/MS and MS3), stable isotope labeling and GC/MS analysis, we previously proposed a structure of 8-hydroxy-9,11-dioxolane eicosatetraenoic acid (DXA3). Here, we achieved enzymatic synthesis and 1H-NMR characterization of this compound with results in conflict with the previously proposed structural assignment. Accordingly, by using LC-MS, we screened autoxidation reactions of 11-HpETE and thereby identified a candidate sharing the precise reverse phase chromatographic and MS characteristics of the platelet product. We optimized these methods to increase yield, allowing full structural analysis by 1H-NMR. The revised assignment is presented here as 8,9-11,12-diepoxy-13-hydroxy-eicosadienoic acid, abbreviated to 8,9-11,12-DiEp-13-HEDE or DiEpHEDE, substituted for the previous name DXA3. We found that in platelets, the lipid likely forms via dioxolane ring opening with rearrangement to the diepoxy moieties, followed by oxygen insertion at C13. We present its enzymatic biosynthetic pathway and MS/MS fragmentation pattern, and using the synthetic compound, demonstrate that it has bioactivity. For the platelet lipid, we estimate 16 isomers based on our current knowledge (and 4 isomers for the synthetic lipid). Determining the exact isomeric structure of the platelet lipid remains to be undertaken.
Original languageEnglish
Pages (from-to)9225-9238
Number of pages14
JournalJournal of Biological Chemistry
Volume294
Issue number23
Early online date6 May 2019
DOIs
Publication statusPublished - 7 Jun 2019

Keywords

  • Immunity
  • leukocyte-regulating lipid
  • 8-hydroxy-9,11- dioxolane eicosatetraenoic acid (DXA3)
  • 8,9-11,12-diepoxy-13-hydroxy-eicosadienoic acid (8,9-11,12-DiEp-13-HEDE)
  • DiEpHEDE
  • eicosanoid
  • lipid metabolism
  • cyclooxygenase (COX)
  • platelet
  • lipid

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