Induction of endothelial cell apoptosis by TNF alpha: modulation by inhibitors of protein synthesis.
Our objective was to examine the induction of endothelial cell apoptosis by the proinflammatory ligand, TNF alpha. We hypothesized that TNF alpha influences endothelial cell viability by altering the balance of regulatory molecules that either induce or suppress apoptosis. Since the identity of these regulators is unknown, our approach was to examine induction of endothelial cell apoptosis by TNF alpha alone and in the context of an inhibitor of transcription or translation. TNF alpha was able to induce bovine pulmonary artery endothelial cell apoptosis in a dose-dependent fashion. Inhibition of transcription with actinomycin D or translation with cycloheximide also resulted in apoptosis, which reached a maximum value of approximately 35 to 40% of cells after 24 h. TNF alpha induction of apoptosis was either potentiated or abrogated by cycloheximide, depending on the timing of TNF alpha exposure in relation to inhibition of protein synthesis. When cycloheximide was added concomitantly with midrange concentrations of TNF alpha, there was both a dramatic acceleration and a synergistic increase in the observed apoptotic response, with all endothelial cells dying within 24 h. When cycloheximide was added as a function of time after termination of TNF alpha exposure, the synergistic induction of apoptosis was maintained at > 70% of its maximum value for 1 h, declining monotonically in a time-dependent fashion to baseline values after 6 h. In contrast, when TNF alpha was added after protein synthesis was inhibited, no additional increase in apoptosis above that observed with inhibition of protein synthesis alone was observed. Our results are consistent with the concept that endothelial cell viability depends on an interaction of inducers and suppressors of apoptosis, which are susceptible to modulation by TNF alpha.
Polunovsky
V A
VA
Department of Medicine, University of Minnesota Medical School, Minneapolis 55455.
Wendt
C H
CH
Ingbar
D H
DH
Peterson
M S
MS
Bitterman
P B
PB
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002417:Cattle; D002470:Cell Survival; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology*; D004247:DNA / Q000032:analysis; D003609:Dactinomycin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000648:ultrastructure; D008954:Models, Biological; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D011500:Protein Synthesis Inhibitors; D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D004247:DNA; D003513:Cycloheximide