title: Ceramide signaling in fenretinide-induced endothelial cell apoptosis.
abstract: Stress stimuli can mediate apoptosis by generation of the lipid second messenger, ceramide. Herein we investigate the molecular mechanism of ceramide signaling in endothelial apoptosis induced by fenretinide (N-(4-hydroxyphenyl)retinamide (4-HPR)). 4-HPR, a synthetic derivative of retinoic acid that induces ceramide in tumor cell lines, has been shown to have antiangiogenic effects, but the molecular mechanism of these is largely unknown. We report that 4-HPR was cytotoxic to endothelial cells (50% cytotoxicity at 2.4 microm, 90% at 5.36 microm) and induced a caspase-dependent endothelial apoptosis. 4-HPR (5 microm) increased ceramide levels in endothelial cells 5.3-fold, and the increase in ceramide was required to achieve the apoptotic effect of 4-HPR. The 4-HPR-induced increase in ceramide was suppressed by inhibitors of ceramide synthesis, fumonisin B(1), myriocin, and l-cycloserine, and 4-HPR transiently activated serine palmitoyltransferase, demonstrating that 4-HPR induced de novo ceramide synthesis. Sphingomyelin levels were not altered by 4-HPR, and desipramine had no effect on ceramide level, suggesting that sphingomyelinase did not contribute to the 4-HPR-induced ceramide increase. Finally, the pancaspase inhibitor, t-butyloxycarbonyl-aspartyl[O-methyl]-fluoromethyl ketone, suppressed 4-HPR-mediated apoptosis but not ceramide accumulation, suggesting that ceramide is upstream of caspases. Our results provide the first evidence that increased ceramide biosynthesis is required for 4-HPR-induced endothelial apoptosis and present a molecular mechanism for its antiangiogenic effects.
authors.affiliation: Division of Hematology-Oncology, Department of Pediatrics, Childrens Hospital Los Angeles, Keck School of Medicine, University of Southern California, 4650 Sunset Boulevard, Los Angeles, CA 90027, USA. epstein@usc.edu
authors.forename: Anat
authors.initials: A
authors.lastname: Erdreich-Epstein
authors.forename: Linda B
authors.initials: LB
authors.lastname: Tran
authors.forename: Nina N
authors.initials: NN
authors.lastname: Bowman
authors.forename: Hongtao
authors.initials: H
authors.lastname: Wang
authors.forename: Myles C
authors.initials: MC
authors.lastname: Cabot
authors.forename: Donald L
authors.initials: DL
authors.lastname: Durden
authors.forename: Jitka
authors.initials: J
authors.lastname: Vlckova
authors.forename: C Patrick
authors.initials: CP
authors.lastname: Reynolds
authors.forename: Monique F
authors.initials: MF
authors.lastname: Stins
authors.forename: Susan
authors.initials: S
authors.lastname: Groshen
authors.forename: Melissa
authors.initials: M
authors.lastname: Millard
mesh_terms: D000217:Acyltransferases / Q000378:metabolism; D020533:Angiogenesis Inhibitors / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D002518:Ceramides / Q000378:metabolism / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D003523:Cycloserine / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism / Q000473:pathology*; D004789:Enzyme Activation; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology; D017313:Fenretinide / Q000494:pharmacology*; D005434:Flow Cytometry; D016193:G1 Phase; D005963:Glucosylceramides / Q000378:metabolism; D006801:Humans; D008861:Microsomes; D017382:Reactive Oxygen Species; D016192:Resting Phase, Cell Cycle; D051102:Serine C-Palmitoyltransferase; D015398:Signal Transduction; D013109:Sphingomyelins / Q000378:metabolism; D013997:Time Factors; D014212:Tretinoin / Q000494:pharmacology
chemical_list: D020533:Angiogenesis Inhibitors; D000970:Antineoplastic Agents; D002518:Ceramides; D005229:Fatty Acids, Monounsaturated; D005963:Glucosylceramides; D017382:Reactive Oxygen Species; D013109:Sphingomyelins; D017313:Fenretinide; D014212:Tretinoin; D003523:Cycloserine; D000217:Acyltransferases; D051102:Serine C-Palmitoyltransferase; D020169:Caspases; C001996:thermozymocidin