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          Institute: MPI für Herz- und Lungenforschung (W. G. Kerckhoff Institut)     Collection: Publikationen des W. G. Kerckhoff-Instituts     Display Documents

ID: 354761.0, MPI für Herz- und Lungenforschung (W. G. Kerckhoff Institut) / Publikationen des W. G. Kerckhoff-Instituts
Ischemic acidosis causes apoptosis in coronary endothelial cells through activation of caspase-12
Authors:Kumar, S.; Kasseckert, S.; Kostin, S.; Abdallah, Y.; Schafer, C.; Kaminski, A.; Reusch, H. P.; Piper, H. M.; Steinhoff, G.; Ladilov, Y.
Date of Publication (YYYY-MM-DD):2007-01-01
Journal Abbrev.:Cardiovasc Res
Issue / Number:1
Start Page:172
End Page:180
Audience:Not Specified
Abstract / Description: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.
Free Keywords: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
External Publication Status:published
Document Type:Article
Communicated by:N. N.
Affiliations:MPI für physiologische und klinische Forschung
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