title: Expression of LTC4 synthase during the development of eosinophils in vitro from cord blood progenitors.
abstract: The expression of leukotriene C4 synthase (LTC4S) was examined during the development of eosinophils in vitro from cord blood mononuclear cells. At 7 days, the cells contained mRNA and sodium dodecyl sulfate-polyacrylamide gel electrophoresis immunoblot signals for cytosolic phospholipase A2 (cPLA2), 5-lipoxygenase (5-LO), and 5-lipoxygenase-activating protein (FLAP), but lacked LTC4S and did not generate cysteinyl leukotrienes when stimulated with 20 mumol/L calcium ionophore. At 14 days, 94% of the cells were of eosinophil lineage, both LTC4S mRNA transcript and protein were present, and ionophore stimulation resulted in the generation of 23.9 +/- 6.0 pmol cysteinyl leukotrienes/10(6) eosinophil-lineage cells (mean +/- SEM, n = 6). At 28 days, progressive eosinophil maturation was accompanied by further increments in 5-LO, FLAP, and LTC4S proteins, and by the ionophore-induced production of 94.6 +/- 9.0 pmol cysteinyl leukotrienes/10(6) eosinophil-lineage cells (n = 6). Cells selected for CD34 expression lacked detectable 5-LO/LTC4S pathway proteins, and with culture generally expressed immunodetectable cPLA2 and 5-LO proteins by 3 days, FLAP protein by 7 days, and LTC4S protein by 10 days. Thus, during the development of eosinophils in vitro, cPLA2, 5-LO, and FLAP are expressed before LTC4S. Once the lineage is established by morphologic criteria, the eosinophilopoietic cytokines mediate upregulation of FLAP and LTC4S, members of a newly recognized gene family, and of 5-LO, during ongoing cell maturation.
authors.affiliation: Department of Medicine, Harvard Medical School, Boston, MA, USA.
authors.forename: J A
authors.initials: JA
authors.lastname: Boyce
authors.forename: B K
authors.initials: BK
authors.lastname: Lam
authors.forename: J F
authors.initials: JF
authors.lastname: Penrose
authors.forename: D S
authors.initials: DS
authors.lastname: Friend
authors.forename: S
authors.initials: S
authors.lastname: Parsons
authors.forename: W F
authors.initials: WF
authors.lastname: Owen
authors.forename: K F
authors.initials: KF
authors.lastname: Austen
mesh_terms: D058927:5-Lipoxygenase-Activating Proteins; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology; D002352:Carrier Proteins / Q000378:metabolism; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology; D004338:Drug Combinations; D004790:Enzyme Induction; D004804:Eosinophils / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D005312:Fetal Blood / Q000166:cytology*; D005982:Glutathione Transferase / Q000096:biosynthesis* / Q000235:genetics; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D007231:Infant, Newborn; D007377:Interleukin-3 / Q000494:pharmacology; D015848:Interleukin-5 / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007797:Laminin / Q000494:pharmacology; D015289:Leukotrienes / Q000096:biosynthesis; D008084:Lipoxygenase / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D010641:Phenotype; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D011509:Proteoglycans / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000494:pharmacology
chemical_list: D058927:5-Lipoxygenase-Activating Proteins; C548592:ALOX5AP protein, human; D002352:Carrier Proteins; D004338:Drug Combinations; D007377:Interleukin-3; D015848:Interleukin-5; D007476:Ionophores; D007797:Laminin; D015289:Leukotrienes; D008565:Membrane Proteins; D011509:Proteoglycans; D012333:RNA, Messenger; D011994:Recombinant Proteins; C059485:matrigel; D000001:Calcimycin; D003094:Collagen; D008084:Lipoxygenase; D005982:Glutathione Transferase; D010741:Phospholipases A; D054467:Phospholipases A2; C041116:leukotriene-C4 synthase; D002118:Calcium