Acetaminophen potentiates staurosporine-induced death in a human neuroblastoma cell line.
OBJECTIVE
Neuroblastoma is the most common solid tumour in infants characterized by a high resistance to apoptosis. Recently, the cyclo-oxygenase pathway has been considered a potential target in the treatment of different kinds of tumours. The aim of the present work was to investigate a possible relationship between cyclo-oxygenase pathway and stauroporine-induced apoptosis in the neuroblastoma cell line SH-SY5Y.


METHODS
Cellular viability was measured by release of LDH. DNA fragmentation was visualized by electrophoresis on agarose gel containing ethidium bromide. Cyclo-oxygenase activity was measured in microsomal fractions obtained from cells by quantification of its final product PGE2 by RIA. Caspase-3 activity was measured fluorimetrically and Western blot analysis was performed to assess cytochrome c expression.


RESULTS
We have found that staurosporine (500 nM) induced cellular death in a time-dependent manner in SH-SY5Y human neuroblastoma cells. Cyclo-oxygenase enzymatic activity was present in SH-SY5Y human neuroblastoma cells under basal conditions and pharmacological experiments using COX inhibitors indicate that cyclo-oxygenase-1 and cyclo-oxygenase-3 are the active isoforms in these cells. Co-incubation of SH-SY5Y cells with staurosporine (500 nM) and acetaminophen for 24 h potentiated staurosporine-mediated cellular death in a concentration-dependent manner. This process is mediated by an increase in cytochrome c release and caspase 3 activation and is prevented by N-acetylcysteine or the superoxide dismutase mimetic, MnTBAP.


CONCLUSIONS
Acetaminophen potentiates staurosporine-mediated neuroblastoma cell death. The mechanism of action of acetaminophen seems to be related to production of reactive oxygen species and decreased intracellular glutathione levels.
Posadas
I
I
Unidad Asociada Neurodeath, UCLM-CSIC, Universidad de Castilla-La Mancha, Avda. de Almansa, Albacete, Spain.
Vellecco
V
V
Santos
P
P
Prieto-Lloret
J
J
Ceña
V
V
D000082:Acetaminophen / Q000494:pharmacology*; D000111:Acetylcysteine / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D051545:Cyclooxygenase 1 / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology*; D065607:Cytochrome P-450 Enzyme Inhibitors; D045304:Cytochromes c / Q000378:metabolism; D015232:Dinoprostone / Q000378:metabolism; D004221:Disulfiram / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D016166:Free Radical Scavengers / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008665:Metalloporphyrins / Q000494:pharmacology; D009447:Neuroblastoma / Q000201:enzymology / Q000503:physiopathology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism; D019311:Staurosporine / Q000494:pharmacology*; D013997:Time Factors
D000970:Antineoplastic Agents; D016861:Cyclooxygenase Inhibitors; D065607:Cytochrome P-450 Enzyme Inhibitors; D016166:Free Radical Scavengers; D008665:Metalloporphyrins; C097284:manganese(III)-tetrakis(4-benzoic acid)porphyrin; D000082:Acetaminophen; D045304:Cytochromes c; D051545:Cyclooxygenase 1; D011451:Prostaglandin-Endoperoxide Synthases; C466421:cyclooxygenase-3; D053148:Caspase 3; D005978:Glutathione; D019311:Staurosporine; D015232:Dinoprostone; D004221:Disulfiram; D000111:Acetylcysteine