Is digitalis compound-induced cardiotoxicity, mediated through guinea-pig cardiomyocytes apoptosis?
Our aim in performing this study was to analyze in vivo the cell death mechanism induced by toxic doses of digitalis compounds on guinea-pig cardiomyocytes. We analyzed three study groups of five male guinea pigs each. Guinea pigs were intoxicated under anesthesia with ouabain or digoxin (at a 50-60% lethal dose); the control group did not receive digitalis. A 5-hours period elapsed before guinea pig hearts were extracted to obtain left ventricle tissue. We carried out isolation of mitochondria and cytosol, cytochrome c and caspase-3 and -9 determination, and electrophoretic analysis of nuclear DNA. TdT-mediated DUTP-X nick end labeling (TUNEL) reaction was performed in histologic preparations to identify in situ apoptotic cell death. Ultrastructural analysis was performed by electron microscopy. Electrophoretic analysis of DNA showed degradation into fragments of 200-400 base pairs in digitalis-treated groups. TUNEL reaction demonstrated the following: in the control group, <10 positive nuclei per field; in the digoxin-treated group, 2-14 positive nuclei per field, while in the ouabain-treated group counts ranged from 9-30 positive nuclei per field. Extracts from ouabain-treated hearts had an elevation of cytochrome c in cytosol and a corresponding decrease in mitochondria; this release of cytochrome c provoked activation of caspase-9 and -3. Electron microscopy revealed presence of autophagic vesicles in cytoplasm of treated hearts. Toxic dosages of digitalis at 50-60% of the lethal dose are capable of inducing cytochrome c release from mitochondria, processing of procaspase-9 and -3, and DNA fragmentation; these observations are mainly indicative of apoptosis, although a mixed mechanism of cell death cannot be ruled out.
Ramirez-Ortega
Margarita
M
Departamento de Farmacologia, Instituto Nacional de Cardiologia Ignacio Chavez, Juan Badiano 1, Col. Seccion XVI, 14080 Mexico, D. F., Mexico. margarita0022@yahoo.com
Zarco
Gabriela
G
Maldonado
Vilma
V
Carrillo
Jose F
JF
Ramos
Pilar
P
Ceballos
Guillermo
G
Melendez-Zajgla
Jorge
J
Garcia
Noemí
N
Zazueta
Cecilia
C
Chanona
Jose
J
Suarez
Jorge
J
Pastelin
Gustavo
G
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002316:Cardiotonic Agents / Q000633:toxicity*; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D053938:DNA Fragmentation; D004071:Digitalis Glycosides / Q000633:toxicity*; D006168:Guinea Pigs; D020287:In Situ Nick-End Labeling; D008297:Male; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D009206:Myocardium / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D010042:Ouabain / Q000633:toxicity
D002316:Cardiotonic Agents; D002843:Chromatin; D004071:Digitalis Glycosides; D010042:Ouabain; D045304:Cytochromes c; D053148:Caspase 3; D053453:Caspase 9