Ischemic acidosis causes apoptosis in coronary endothelial cells through activation of caspase-12.

Abstract

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 pH(o) 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 pH(o) 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.

• Authors: Sanjeev Kumar, Sascha Kasseckert, Sawa Kostin, Yaser Abdallah, Claudia Schafer, Alexander Kaminski, H Peter Reusch, Hans Michael Piper, Gustav Steinhoff, Yury Ladilov
• Journal: Cardiovascular research (Cardiovasc Res) Vol. 73 Issue 1 Pg. 172-80 (Jan 01 2007) ISSN: 0008-6363 [Print] England
• PMID: 17083921 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Cytochromes c
• Caspase 12
• Caspases
• Calcium

Topics

• Acidosis (enzymology)
• Animals
• Apoptosis
• Blotting, Western (methods)
• Calcium (analysis, metabolism)
• Caspase 12 (analysis, genetics, metabolism)
• Caspases (analysis, metabolism)
• Cells, Cultured
• Coronary Vessels
• Cytochromes c (metabolism)
• Cytosol (chemistry, metabolism)
• Endothelial Cells (enzymology, pathology)
• Enzyme Activation
• Immunohistochemistry
• In Situ Nick-End Labeling
• Male
• Mitochondria, Heart (metabolism)
• Myocardial Ischemia (enzymology, pathology)
• RNA Interference
• Rats
• Rats, Wistar