7-Ketocholesterol and staurosporine induce opposite changes in intracellular pH, associated with distinct types of cell death in ECV304 cells.
Incubation of ECV304 cells with 7-ketocholesterol, a lipid component of oxidized low-density lipoproteins, caused a concentration- and time-dependent decrease in the number of viable cells. Other cholesterol oxides, 7 beta-hydroxycholesterol and 25-hydroxycholesterol, but not cholesterol, were only weakly cytotoxic. No evidence for activation of caspase-3 and -8, DNA laddering, or release of cytochrome c from mitochondria into the cytoplasm was obtained in 7-ketocholesterol-treated cells, indicating that cell death was not due to apoptosis. As a positive control for apoptosis, ECV304 cells were treated with staurosporine, which indeed caused significant activation of caspase-3 activity, DNA laddering, and cytochrome c release. Cellular morphology and actin cytoskeletal organization were distinctly different after exposure to the two drugs. Furthermore, staurosporine caused intracellular acidification, whereas 7-ketocholesterol induced a significant alkalinization, which was abolished by 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid. In conclusion, in ECV304 cells 7-ketocholesterol induces some typical hallmarks of necrotic cell death but not of apoptosis.
Ghelli
Anna
A
Dipartimento di Biologia Ev. Sp., Università di Bologna, Via Irnerio 42, 40126 Bologna, Italy.
Porcelli
Anna Maria
AM
Zanna
Claudia
C
Rugolo
Michela
M
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002452:Cell Count; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003574:Cytochrome c Group / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects / Q000648:ultrastructure; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007424:Intracellular Fluid / Q000737:chemistry; D007653:Ketocholesterols / Q000494:pharmacology*; D009336:Necrosis; D019311:Staurosporine / Q000494:pharmacology*; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism
D003574:Cytochrome c Group; D007653:Ketocholesterols; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D019311:Staurosporine; C003001:7-ketocholesterol