Synergistic protection of a general caspase inhibitor and MK-801 in bilirubin-induced cell death in human NT2-N neurons.
Unconjugated bilirubin (UCB) induces both apoptosis and necrosis in neurons. To investigate the role of caspases and excitotoxicity in UCB-induced cell death, we exposed NT2-N neurons to 5 microM UCB (a concentration known to induce apoptosis) or 2 microM staurosporine (positive apoptosis control) and investigated the effects of treatments with the specific caspase-3 inhibitor, zDEVD.FMK (20 and 100 microM), or the general caspase inhibitor, zVAD.FMK (20 and 100 microM), and/or the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (10 microM) during a 24- or 48-h exposure. UCB increased caspase-3 activity 2.3-fold after 6 h. Despite this, treatment with zDEVD.FMK did not prevent cell death. zVAD.FMK enhanced neuronal survival by reducing apoptotic nuclear fragmentation, while MK-801 enhanced survival by reducing apoptotic nuclear condensation; both without affecting the MTT assays. Combined treatment reduced both apoptotic morphologies (without affecting necrosis), and this effect was also reflected in the MTT assays [corrected] We conclude that NMDA receptor-mediated pathways and caspase-mediated pathways are involved in UCB-induced cell death in human NT2-N neurons. Concomitant inhibition of both pathways results in synergistic protection.
Hankø
Erik
E
Department of Pediatric Research, Riskhospitalet-Radiumhospitalet University Hospital, University of Oslo, Norway. erikh@ulrik.uio.no
Hansen
Thor W D
TW
Almaas
Runar
R
Paulsen
Ragnhild
R
Rootwelt
Terje
T
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D001663:Bilirubin / Q000633:toxicity*; D053148:Caspase 3; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D016291:Dizocilpine Maleate / Q000494:pharmacology*; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007647:Kernicterus / Q000201:enzymology / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors*; D014407:Tumor Cells, Cultured
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D016194:Receptors, N-Methyl-D-Aspartate; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D016291:Dizocilpine Maleate; C505407:CASP3 protein, human; D053148:Caspase 3; D001663:Bilirubin