(Circulation. 2000;102:1564.)
© 2000 American Heart Association, Inc.
Basic Science Reports |
Cardiomyocyte Death Induced by Myocardial Ischemia and Reperfusion
Measurement With Recombinant Human Annexin-V in a Mouse Model
Presented in part at the Keystone Meeting (Apoptosis and Programmed Cell Death), Breckenridge, Colo, April 6–11, 1999, and at The Lancet Conference (The Challenge of Acute Coronary Syndromes), Copenhagen, Denmark, June 9–11, 1999.
From the Departments of Cardiology (E.A.W.J.D., L.H., P.A.F.D., E.D, H.J.J.W.), Biochemistry (W.L.v.H., S.V.D.E., C.P.M.R), Surgery (M.A.R.C.D.), Pharmacology (J.F.M.S.), Electron Microscopy (P.F.), and Pathology (M.J.A.P.D.), Cardiovascular Research Institute, Maastricht, the Netherlands.
Correspondence to L. Hofstra, MD, PhD, Department of Cardiology, University Hospital Maastricht, PO Box 5800, 6202 AZ Maastricht, Netherlands. E-mail L.Hofstra{at}cardio.azm.nl
Introduction—Phosphatidylserine (PS) externalization is regarded as one of the earliest hallmarks of cells undergoing programmed cell death. We studied the use of labeled human recombinant annexin-V, a protein selectively binding to PS, to detect cardiomyocyte death in an in vivo mouse model of cardiac ischemia and reperfusion (I/R).
Methods and Results—I/R was induced in mouse hearts by ligation and subsequent release of a suture around the left anterior descending coronary artery. Annexin-V (25 mg/kg) fused to a marker molecule was injected intra-arterially 30 minutes before euthanasia. After 15 minutes of ischemia followed by 30 minutes of reperfusion, 1.4±1.2% (mean±SD) of the cardiomyocytes in the area at risk were annexin-V positive (n=6). This increased to 11.4±1.9% after 15 minutes of ischemia followed by 90 minutes of reperfusion (n=7) and to 20.2±3.3% after 30 minutes of ischemia followed by 90 minutes of reperfusion (n=7). In control mice, including those injected with annexin-V at the binding site of PS, no annexin-V–positive cells were observed. DNA gel electrophoresis showed typical laddering starting after 15 minutes of ischemia followed by 30 minutes of reperfusion, suggesting activation of the cell death program. Intervention in the cell death program by pretreatment with a novel Na+-H+ exchange inhibitor substantially decreased annexin-V–positive cardiomyocytes from 20.2% to 2.2% in mice after 30 minutes of ischemia followed by 90 minutes of reperfusion.
Conclusions—These data suggest that labeled annexin-V is useful for in situ detection of cell death in an in vivo model of I/R in the heart and for the evaluation of cell death–blocking strategies.
Key Words: apoptosis • cells • myocardial infarction • reperfusion • proteins
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