Copyright © 2006, European Society of Cardiology
Ischemic acidosis causes apoptosis in coronary endothelial cells through activation of caspase-12
aDepartment of Clinical Pharmacology, Ruhr-Universität Bochum, Germany
bDepartment of Cardiac Surgery, University of Rostock, Germany
cInstitute of Physiology, University of Giessen, Germany
dMax-Planck-Institute for Heart and Lung Research, Bad Nauheim, Germany
* Corresponding author. Department of Clinical Pharmacology, Universitätsstrasse 150, D-44801 Bochum, Germany. Tel.: +49 0234/32 27639; fax: +49 234/32 14904. Email address: yury.ladilov{at}ruhr-uni-bochum.de
Objective: Myocardial ischemia has been shown to induce apoptosis of endothelial cells (EC). However, the mechanism of this endothelial injury is still poorly understood. To analyse the signaling pathway of ischemia-induced EC apoptosis was the aim of the present study.
Methods: The primary culture of rat coronary EC was exposed to simulated ischemia (glucose-free anoxia at pHo 6.4). Apoptosis was defined by staining of nuclei with Hoechst-33342 and TUNEL. Cytosolic Ca2+ and pH were measured with Fura-2 and BCECF, respectively.
Results: Apoptosis (29.2±1.7% of cells) induced by exposure to simulated ischemia for 2 h was accompanied by cytosolic Ca2+ overload (1090±52 nmol/l) and acidosis (pHi=6.52±0.13). Simulated ischemia had no significant effect on caspase-8 cleavage, but induced cleavage of caspase-3 and caspase-12 and led to a slight release of cytochrome C. Prevention of cytosolic acidosis (anoxia at pHo 7.4) had no effect on cytochrome C release, but significantly reduced apoptosis, attenuated cytosolic Ca2+ overload, and prevented cleavage of caspase-12. A similar effect was achieved by inhibition of Ca2+ release channels in the endoplasmic reticulum with ryanodine and xestospongin C. Knock-down of caspase-12 with small interfering RNA suppressed caspase-3 activation and reduced apoptotic cell number by about 70%.
Conclusion: Acidosis, rather than anoxia, is an important trigger of apoptosis in EC under simulated ischemia. The main pathway of the simulated ischemia-induced apoptosis consists of the Ca2+ leak from the ER followed by activation of caspase-12 and caspase-3.
KEYWORDS Coronary endothelial cell; Apoptosis; Acidosis; Caspase-12
Ajay Shah (King's College London) served as Guest Editor for this article.
Time for primary review 34 days
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