Calcineurin pathway is required for endothelin-1-mediated protection against oxidant stress-induced apoptosis in cardiac myocytes.
Endothelin-1 (ET-1) acts not only as a growth-promoting peptide but also as a potent survival factor against myocardial cell apoptosis. However, the signaling pathways leading to myocardial cell protection by ET-1 are poorly understood. Using a culture system of primary cardiac myocytes derived from neonatal rats, we show in the present study that ET-1 almost completely blocked the hydrogen peroxide-induced increase in the percentage of TdT-mediated dUTP-biotin nick-end labeling-positive myocytes. Apoptosis inhibition by ET-1 was confirmed by cytofluorometric analysis as well as by examination of the ladder formation, morphological features, and caspase-3 cleavage. We have found that ET-1 converts the nuclear factor of activated T lymphocytes (NFATc) in cardiac myocytes into high-mobility forms and translocates cytoplasmic NFATc to the nuclei. In addition, ET-1 stimulates the interaction between NFATc and the cardiac-restricted zinc-finger protein GATA4 in these cells. The immunosuppressants cyclosporin A and FK506, which antagonize calcineurin, negated the inhibitory effect of ET-1 on apoptosis. Calcineurin activation de novo was sufficient to inhibit hydrogen peroxide-induced apoptosis. ET-1 induced the expression of an antiapoptotic protein bcl-2 in cardiac myocytes in a cyclosporin A-dependent manner, but it did not alter the expression of bax. Cyclosporin A also attenuated the ET-1-stimulated transcription of the bcl-2 gene in these cells. These findings demonstrate that the calcineurin pathway is required for the inhibitory effect of ET-1 on oxidant stress-induced apoptosis in cardiac myocytes.
Kakita
T
T
Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
Hasegawa
K
K
Iwai-Kanai
E
E
Adachi
S
S
Morimoto
T
T
Wada
H
H
Kawamura
T
T
Yanazume
T
T
Sasayama
S
S
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D019703:Calcineurin / Q000378:metabolism*; D065095:Calcineurin Inhibitors; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D019332:Endothelin-1 / Q000378:metabolism* / Q000494:pharmacology; D005434:Flow Cytometry; D050991:GATA4 Transcription Factor; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007166:Immunosuppressive Agents / Q000494:pharmacology; D020287:In Situ Nick-End Labeling; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000502:physiology; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D051028:bcl-2-Associated X Protein
C494828:Bax protein, rat; D065095:Calcineurin Inhibitors; D004268:DNA-Binding Proteins; D019332:Endothelin-1; D050991:GATA4 Transcription Factor; D007166:Immunosuppressive Agents; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D016877:Oxidants; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014157:Transcription Factors; D051028:bcl-2-Associated X Protein; D006861:Hydrogen Peroxide; D019703:Calcineurin; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases