Involvement of de novo ceramide biosynthesis in tumor necrosis factor-alpha/cycloheximide-induced cerebral endothelial cell death.
Cytokines, including tumor necrosis factor-alpha (TNF-alpha), may elicit cytotoxic response through the sphingomyelin-ceramide signal transduction pathway by activation of sphingomyelinases and the subsequent release of ceramide: the universal lipid second messenger. Treatment of bovine cerebral endothelial cells (BCECs) with TNF-alpha for 16 h followed by cycloheximide (CHX) for 6 h resulted in an increase in ceramide accumulation, DNA fragmentation, and cell death. Application of a cell permeable ceramide analogue C2 ceramide, but not the biologically inactive C2 dihydroceramide, also induced DNA laddering and BCEC death in a concentration- and time-dependent manner. TNF-alpha/CHX-mediated ceramide production apparently is not a result of sphingomyelin hydrolysis because sphingomyelin content does not decrease in this death paradigm. In addition, an acidic sphingomyelinase inhibitor, desipramine, had no effect on TNF-alpha/CHX-induced cell death. However, addition of fumonisin B1, a selective ceramide synthase inhibitor, attenuated TNF-alpha/CHX-induced intracellular ceramide elevation and BCEC death. Together, these findings suggest that ceramide plays at least a partial role in this paradigm of BCEC death. Our results show, for the first time, that ceramide derived from de novo synthesis is an alternative mechanism to sphingomyelin hydrolysis in the BCEC death process initiated by TNF-alpha/CHX.
Xu
J
J
Center for the Study of Nervous System Injury and Department of Neurology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Yeh
C H
CH
Chen
S
S
He
L
L
Sensi
S L
SL
Canzoniero
L M
LM
Choi
D W
DW
Hsu
C Y
CY
D000818:Animals; D002417:Cattle; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D002518:Ceramides / Q000096:biosynthesis*; D002560:Cerebrovascular Circulation; D003513:Cycloheximide / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000187:drug effects*; D010088:Oxidoreductases / Q000037:antagonists & inhibitors; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D002518:Ceramides; D011500:Protein Synthesis Inhibitors; D014409:Tumor Necrosis Factor-alpha; D003513:Cycloheximide; D010088:Oxidoreductases; C108138:dihydroceramide desaturase