title: De novo C16- and C24-ceramide generation contributes to spontaneous neutrophil apoptosis.
abstract: Neutrophils rapidly undergo spontaneous apoptosis following their release from the bone marrow. Although central to leukocyte homeostasis, the mechanisms that regulate neutrophil apoptosis remain poorly understood. We show here that apoptosis of cultured neutrophils is preceded by a substantial increase in the intracellular levels of 16 and 24 carbon atom (C(16)- and C(24))-ceramides, which are lipid second messengers of apoptosis and stress signaling. Treatment of neutrophils with fumonisin B(2), a selective inhibitor of the de novo pathway of ceramide synthesis, prevented accumulation of C(16)- and C(24)-ceramides. Moreover, fumonisin B(2) significantly reduced caspase-3, -8, and -9 activation and apoptosis in these cells. Conversely, 3-O-methylsphingomyelin and fantofarone, which are specific inhibitors of neutral and acid sphingomyelinases, respectively, neither inhibited C(16)- and C(24)-ceramide production nor decreased the apoptosis rate in neutrophils, indicating that in these cells, ceramides are not generated from membrane sphingomyelin. Further experiments showed that increasing endogenous C(16)- and C(24)-ceramide levels by using DL-threo-1-phenyl-2-palmitoylamino-3-morpholino-1-propanol and (1S,2R)-D-erythro-2-(N-myristoylamino)-1-phenyl-1-propanol, two inhibitors of ceramide metabolism, enhances caspase-3, -8, and -9 activity and increases neutrophil apoptosis. Similarly, apoptosis was induced rapidly when synthetic C(16)- and/or C(24)-ceramides were added to neutrophil cultures. Finally, GM-CSF, a cytokine that delays neutrophil apoptosis, abrogated C(16)- and C(24)-ceramide accumulation totally in cultured neutrophils, whereas Fas ligation accelerated apoptosis in these cells without affecting de novo ceramide production. We conclude that de novo generation of C(16)- and C(24)-ceramides contributes to spontaneous neutrophil apoptosis via caspase activation and that GM-CSF exerts its antiapoptotic effects on neutrophils, at least partly through inhibition of ceramide accumulation.
authors.affiliation: Department of Physiology, University of Liège, Boulevard de Colonster, Bâtiment B42, Sart-Tilman, B-4000, Liège, Belgium.
authors.forename: Grégory
authors.initials: G
authors.lastname: Seumois
authors.forename: Marianne
authors.initials: M
authors.lastname: Fillet
authors.forename: Laurent
authors.initials: L
authors.lastname: Gillet
authors.forename: Céline
authors.initials: C
authors.lastname: Faccinetto
authors.forename: Christophe
authors.initials: C
authors.lastname: Desmet
authors.forename: Cédric
authors.initials: C
authors.lastname: François
authors.forename: Benjamin
authors.initials: B
authors.lastname: Dewals
authors.forename: Cécile
authors.initials: C
authors.lastname: Oury
authors.forename: Alain
authors.initials: A
authors.lastname: Vanderplasschen
authors.forename: Pierre
authors.initials: P
authors.lastname: Lekeux
authors.forename: Fabrice
authors.initials: F
authors.lastname: Bureau
mesh_terms: D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D002518:Ceramides / Q000096:biosynthesis*; D037341:Fumonisins / Q000494:pharmacology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D007212:Indolizines / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D009226:Myristates / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D010627:Phenethylamines / Q000494:pharmacology; D011412:Propanolamines / Q000494:pharmacology; D013107:Sphingolipids / Q000494:pharmacology; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism / Q000494:pharmacology
chemical_list: C095823:1-phenyl-2-palmitoylamino-3-morpholino-1-propanol; C508074:3-O-methylsphingomyelin; D002518:Ceramides; D037341:Fumonisins; D007212:Indolizines; D009025:Morpholines; D009226:Myristates; D010627:Phenethylamines; D011412:Propanolamines; D013107:Sphingolipids; D013109:Sphingomyelins; C056934:fumonisin B2; C097760:N-palmitoylsphingosine; C099657:2-(N-myristoylamino)-1-phenyl-1-propanol; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D013108:Sphingomyelin Phosphodiesterase; D020169:Caspases; C059901:fantofarone