EXPRESSION OF LTC(4) SYNTHASE DURING THE DEVELOPMENT OF EOSINOPHILS IN-VITRO FROM CORD-BLOOD PROGENITORS

The expression of leukotriene C-4 synthase (LTC(4)S) was examined duri ng 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 p hospholipase A(2) (cPLA(2)), 5-lipoxygenase (5-LO), and 5-lipoxygenase -activating protein (FLAP), but lacked LTC(4)S and did not generate cy steinyl leukotrienes when stimulated with 20 mu mol/L calcium ionophor e. At 14 days, 94% of the cells were of eosinophil lineage, both LTC(4 )S mRNA transcript and protein were present, and ionophore stimulation resulted in the generation of 28.9 +/- 6.0 pmol cysteinyl leukotriene s/10(6) eosinophil lineage cells (mean +/- SEM, n = 6). At 28 days, pr ogressive eosinophil maturation was accompanied by further increments in 5-LO, FLAP, and LTC(4)S proteins, and by the ionophore-induced prod uction of 94.6 +/- 9.0 pmol cysteinyl leukotrienes/10(6) eosinophil-li neage cells (n = 6). Cells selected for CD34 expression lacked detecta ble 5-LO/LTC(4)S pathway proteins, and with culture generally expresse d immunodetectable cPLA(2) and 5-LO proteins by 3 days, FLAP protein b y 7 days, and LTC(4)S protein by 10 days. Thus, during the development of eosinophils in vitro, cPLA(2), 5-LO, and FLAP are expressed before LTC(4)S. Once the lineage is established by morphologic criteria, the eosinophilopoietic cytokines mediate upregulation of FLAP and LTC(4)S , members of a newly recognized gene family, and of 5-LO, during ongoi ng cell maturation. (C) 1996 by The American Society of Hematology.