Lipoxin and resolvin biosynthesis is dependent on 5-lipoxygenase activating protein

Christoph Lehmann, Julia Homann, Ann Katrin Ball, René Blöcher, Thea K. Kleinschmidt, Devaraj Basavarajappa, Carlo Angioni, Nerea Ferreirós, Ann Kathrin Häfner, Olof Rådmark, Ewgenij Proschak, Jesper Z. Haeggström, Gerd Geisslinger, Michael J. Parnham, Dieter Steinhilber, Astrid Stefanie Kahnt

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Resolution of acute inflammation is an active process coordinated by proresolving lipid mediators (SPMs) such as lipoxins (LXs) and resolvins (Rvs), which are formed by the concerted action of 2 lipoxygenases (LOs). Because the exact molecular mechanisms of SPM biosynthesis are not completely understood, we aimed to investigate LX and D-type Rv formation in human leukocytes and HEK293T cells overexpressing leukotriene (LT) pathway enzymes. Activity assays in precursor (15-hydroxyeicosatetraenoic acids, 17-HDoHE)-treated granulocytes [polymorphonuclear leukocytes (PMNLs)] showed a strict dependence of LXA4/RvD1 biosynthesis on cell integrity, and incubation with recombinant human 5-LO did not lead to LX or Rv formation. Pharmacologic inhibition of 5-LO activating protein (FLAP) by MK-886 inhibited LXA4/RvD1 biosynthesis in precursor-treated PMNLs (drug concentration causing 50% inhibition ∼0.3/0.2 μM), as did knockdown of the enzyme in MM6 cells, and precursor-treated HEK293T overexpressing 5-LO produced high amounts of LXA4 only in the presence of FLAP. In addition, inhibition of cytosolic phospholipase A (cPLA) interfered with LXA4/RvD1 formation from exogenous precursors in PMNLs. Furthermore, inhibition of the LT synthases LTA4 hydrolase and LTC4 synthase in PMNL/platelet coincubations augmented LXA4 levels. These findings show that several enzymes known to be involved in the biosynthesis of proinflammatory LTs, such as FLAP and cPLA, also contribute to LX and Rv formation.

LanguageEnglish
Pages5029-5043
Number of pages15
JournalFASEB Journal
Volume29
Issue number12
DOIs
Publication statusPublished - 1 Dec 2015
Externally publishedYes

Fingerprint

5-Lipoxygenase-Activating Proteins
Lipoxins
Biosynthesis
Arachidonate 5-Lipoxygenase
Neutrophils
Phospholipases
Leukotrienes
L 663536
Enzymes
Hydroxyeicosatetraenoic Acids
Lipoxygenase
Platelets
Granulocytes
Assays
Leukocytes
Blood Platelets
lipoxin A4
Inflammation
Lipids
Pharmaceutical Preparations

Keywords

  • Cytosolic phospholipase A2alpha
  • LTA4 hydrolase
  • LTC4 synthase
  • Resolution of inflammation

Cite this

Lehmann, C., Homann, J., Ball, A. K., Blöcher, R., Kleinschmidt, T. K., Basavarajappa, D., ... Kahnt, A. S. (2015). Lipoxin and resolvin biosynthesis is dependent on 5-lipoxygenase activating protein. FASEB Journal, 29(12), 5029-5043. https://doi.org/10.1096/fj.15-275487
Lehmann, Christoph ; Homann, Julia ; Ball, Ann Katrin ; Blöcher, René ; Kleinschmidt, Thea K. ; Basavarajappa, Devaraj ; Angioni, Carlo ; Ferreirós, Nerea ; Häfner, Ann Kathrin ; Rådmark, Olof ; Proschak, Ewgenij ; Haeggström, Jesper Z. ; Geisslinger, Gerd ; Parnham, Michael J. ; Steinhilber, Dieter ; Kahnt, Astrid Stefanie. / Lipoxin and resolvin biosynthesis is dependent on 5-lipoxygenase activating protein. In: FASEB Journal. 2015 ; Vol. 29, No. 12. pp. 5029-5043.
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abstract = "Resolution of acute inflammation is an active process coordinated by proresolving lipid mediators (SPMs) such as lipoxins (LXs) and resolvins (Rvs), which are formed by the concerted action of 2 lipoxygenases (LOs). Because the exact molecular mechanisms of SPM biosynthesis are not completely understood, we aimed to investigate LX and D-type Rv formation in human leukocytes and HEK293T cells overexpressing leukotriene (LT) pathway enzymes. Activity assays in precursor (15-hydroxyeicosatetraenoic acids, 17-HDoHE)-treated granulocytes [polymorphonuclear leukocytes (PMNLs)] showed a strict dependence of LXA4/RvD1 biosynthesis on cell integrity, and incubation with recombinant human 5-LO did not lead to LX or Rv formation. Pharmacologic inhibition of 5-LO activating protein (FLAP) by MK-886 inhibited LXA4/RvD1 biosynthesis in precursor-treated PMNLs (drug concentration causing 50{\%} inhibition ∼0.3/0.2 μM), as did knockdown of the enzyme in MM6 cells, and precursor-treated HEK293T overexpressing 5-LO produced high amounts of LXA4 only in the presence of FLAP. In addition, inhibition of cytosolic phospholipase A2α (cPLA2α) interfered with LXA4/RvD1 formation from exogenous precursors in PMNLs. Furthermore, inhibition of the LT synthases LTA4 hydrolase and LTC4 synthase in PMNL/platelet coincubations augmented LXA4 levels. These findings show that several enzymes known to be involved in the biosynthesis of proinflammatory LTs, such as FLAP and cPLA2α, also contribute to LX and Rv formation.",
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Lehmann, C, Homann, J, Ball, AK, Blöcher, R, Kleinschmidt, TK, Basavarajappa, D, Angioni, C, Ferreirós, N, Häfner, AK, Rådmark, O, Proschak, E, Haeggström, JZ, Geisslinger, G, Parnham, MJ, Steinhilber, D & Kahnt, AS 2015, 'Lipoxin and resolvin biosynthesis is dependent on 5-lipoxygenase activating protein', FASEB Journal, vol. 29, no. 12, pp. 5029-5043. https://doi.org/10.1096/fj.15-275487

Lipoxin and resolvin biosynthesis is dependent on 5-lipoxygenase activating protein. / Lehmann, Christoph; Homann, Julia; Ball, Ann Katrin; Blöcher, René; Kleinschmidt, Thea K.; Basavarajappa, Devaraj; Angioni, Carlo; Ferreirós, Nerea; Häfner, Ann Kathrin; Rådmark, Olof; Proschak, Ewgenij; Haeggström, Jesper Z.; Geisslinger, Gerd; Parnham, Michael J.; Steinhilber, Dieter; Kahnt, Astrid Stefanie.

In: FASEB Journal, Vol. 29, No. 12, 01.12.2015, p. 5029-5043.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Lehmann, Christoph

AU - Homann, Julia

AU - Ball, Ann Katrin

AU - Blöcher, René

AU - Kleinschmidt, Thea K.

AU - Basavarajappa, Devaraj

AU - Angioni, Carlo

AU - Ferreirós, Nerea

AU - Häfner, Ann Kathrin

AU - Rådmark, Olof

AU - Proschak, Ewgenij

AU - Haeggström, Jesper Z.

AU - Geisslinger, Gerd

AU - Parnham, Michael J.

AU - Steinhilber, Dieter

AU - Kahnt, Astrid Stefanie

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N2 - Resolution of acute inflammation is an active process coordinated by proresolving lipid mediators (SPMs) such as lipoxins (LXs) and resolvins (Rvs), which are formed by the concerted action of 2 lipoxygenases (LOs). Because the exact molecular mechanisms of SPM biosynthesis are not completely understood, we aimed to investigate LX and D-type Rv formation in human leukocytes and HEK293T cells overexpressing leukotriene (LT) pathway enzymes. Activity assays in precursor (15-hydroxyeicosatetraenoic acids, 17-HDoHE)-treated granulocytes [polymorphonuclear leukocytes (PMNLs)] showed a strict dependence of LXA4/RvD1 biosynthesis on cell integrity, and incubation with recombinant human 5-LO did not lead to LX or Rv formation. Pharmacologic inhibition of 5-LO activating protein (FLAP) by MK-886 inhibited LXA4/RvD1 biosynthesis in precursor-treated PMNLs (drug concentration causing 50% inhibition ∼0.3/0.2 μM), as did knockdown of the enzyme in MM6 cells, and precursor-treated HEK293T overexpressing 5-LO produced high amounts of LXA4 only in the presence of FLAP. In addition, inhibition of cytosolic phospholipase A2α (cPLA2α) interfered with LXA4/RvD1 formation from exogenous precursors in PMNLs. Furthermore, inhibition of the LT synthases LTA4 hydrolase and LTC4 synthase in PMNL/platelet coincubations augmented LXA4 levels. These findings show that several enzymes known to be involved in the biosynthesis of proinflammatory LTs, such as FLAP and cPLA2α, also contribute to LX and Rv formation.

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KW - Cytosolic phospholipase A2alpha

KW - LTA4 hydrolase

KW - LTC4 synthase

KW - Resolution of inflammation

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