Bilirubin induces apoptosis via the mitochondrial pathway in developing rat brain neurons.
Increased levels of unconjugated bilirubin, the end-product of heme catabolism, are detrimental to the central nervous system. To examine the role of apoptosis in bilirubin-induced toxicity and to characterize the biochemical pathway of cell death, we exposed developing rat brain neurons to purified unconjugated bilirubin at concentrations below and above saturation of human serum albumin. Isolated neurons treated with bilirubin showed increased levels of apoptosis. Mitochondrial cytochrome c was extensively released and accumulated in cytosol. Consistent with this observation, caspase-3 was activated and the full-length substrate poly(ADP)ribose polymerase (PARP) degraded, even in the presence of very modestly elevated concentrations of bilirubin. In parallel, all events were prevented in cells preincubated with ursodeoxycholate. Further experiments showed that bilirubin diminished mitochondrial transmembrane potential (DeltaPsi(m)) and increased mitochondrial-associated Bax protein levels, while directly disrupting membrane lipid and protein structure. In conclusion, bilirubin induces mitochondrial depolarization and Bax translocation via physical interaction with membranes, mediating the mitochondrial pathway of apoptosis in neurons exposed to bilirubin. These results provide a novel insight into the mechanism of bilirubin-induced toxicity.
Rodrigues
Cecília M P
CM
Centro de Patogénese Molecular, Faculdade de Farmácia, University of Lisbon, Lisbon, Portugal. cmprodrigues@ff.ul.pt
Solá
Susana
S
Brites
Dora
D
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001663:Bilirubin / Q000633:toxicity*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D003574:Cytochrome c Group / Q000378:metabolism; D005260:Female; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D008051:Lipid Bilayers / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009474:Neurons / Q000166:cytology* / Q000201:enzymology*; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D011247:Pregnancy; D011506:Proteins / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D051381:Rats; D017208:Rats, Wistar; D051028:bcl-2-Associated X Protein
C494828:Bax protein, rat; D003574:Cytochrome c Group; D008051:Lipid Bilayers; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C499322:Parp1 protein, rat; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D001663:Bilirubin