title: Role of antioxidants in the O-hydroxyethyl-D-(Ser)8-cyclosporine A (SDZ IMM125)-induced apoptosis in rat hepatocytes.
abstract: The mechanisms underlying the apoptotic activity of the immunosuppressive drug cyclosporine A and its O-hydroxyethyl-D-(Ser)(8)-derivative SDZ IMM125 in rat hepatocytes are not yet fully understood. It was the purpose of the present study to investigate the role of anti- and pro-oxidants and of caspase-3 and intracellular Ca(2+) in SDZ IMM125-induced apoptosis in rat hepatocytes. SDZ IMM125 induced an increase in chromatin condensation and fragmentation, and the activation of caspase-3. Supplementing the cell cultures with the antioxidants, D,L-alpha-tocopherol-polyethylene-glycol-1000-succinate, ascorbic acid, and the reducing agent, dithiothreitol, significantly inhibited the SDZ IMM125-mediated increase in chromatin condensation and fragmentation, and caspase-3 activity. D,L-alpha-tocopherol-polyethylene-glycol-1000-succinate and dithiothreitol caused significant inhibition on SDZ IMM125-mediated cellular Ca(2+) uptake. The glutathione synthetase inhibitor, buthionine sulfoximine, increased SDZ IMM125-mediated caspase-3 action in parallel to chromatin condensation and fragmentation as well as Ca(2+) influx. Supplementation the culture medium with the intracellular Ca(2+) chelator bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid as well as omission of calcium in the medium reduced SDZ IMM125-induced apoptosis whereas the calcium supplementation of the culture medium elevated SDZ IMM125-induced apoptosis. Calcium antagonists inhibited SDZ IMM125-induced caspase-3 activation. Our data indicate that SDZ IMM125-mediated apoptosis in rat hepatocytes can be inhibited by antioxidants, and that the intracellular redox-state can act as a modulator of cytotoxicity and apoptosis. Further, the results suggest that SDZ IMM125-induced uptake of extracellular calcium is also a redox-sensitive process and that the increased intracellular calcium might directly cause apoptosis by increasing the caspase-3 activity as a central event in the cyclosporine-induced apoptotic mechanism.
authors.affiliation: Novartis Pharma AG, Integrative Compound and Product Profiling, Functional Validation, CH-4002 Basel, Switzerland.
authors.forename: Sibylle
authors.initials: S
authors.lastname: Grub
authors.forename: Wolfgang E
authors.initials: WE
authors.lastname: Trommer
authors.forename: Armin
authors.initials: A
authors.lastname: Wolf
mesh_terms: D000818:Animals; D000963:Antimetabolites / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002843:Chromatin / Q000187:drug effects; D016572:Cyclosporine / Q000494:pharmacology; D003524:Cyclosporins / Q000494:pharmacology*; D053938:DNA Fragmentation / Q000187:drug effects; D004347:Drug Interactions; D022781:Hepatocytes / Q000187:drug effects* / Q000473:pathology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008297:Male; D051381:Rats; D017208:Rats, Wistar
chemical_list: D000963:Antimetabolites; D000975:Antioxidants; D002843:Chromatin; D003524:Cyclosporins; D019328:Buthionine Sulfoximine; D016572:Cyclosporine; D007770:L-Lactate Dehydrogenase; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; C075935:SDZ IMM 125; D002118:Calcium