Visualization of cell death in mice with focal cerebral ischemia using fluorescent annexin A5, propidium iodide, and TUNEL staining.
To monitor stroke-induced brain damage and assess neuroprotective therapies, specific imaging of cell death after cerebral ischemia in a noninvasive manner is highly desirable. Annexin A5 has been suggested as a marker for imaging cell death under various disease conditions including stroke. In this study, C57BL6/N mice received middle cerebral artery occlusion (MCAO) and were injected intravenously with either active or inactive Cy5.5-annexin A5 48 hours after reperfusion. Some mice also received propidium iodide (PI), a cell integrity marker. Only in mice receiving active Cy5.5-annexin A5 were fluorescence intensities significantly higher over the hemisphere ipsilateral to MCAO than on the contralateral side. This was detected noninvasively and ex vivo 4 and 8 hours after injection. The majority of cells positive for fluorescent annexin A5 were also positive for PI and fragmented DNA as detected by terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate-biotin nick end labeling (TUNEL) staining. This study demonstrates the high specificity of annexin A5 for visualization of cell death in a mouse model of stroke. To our knowledge, this is the first study to compare the distribution of injected active and inactive annexin A5, PI, and TUNEL staining. It provides important information on the experimental and potential clinical applications of annexin A5-based imaging agents in stroke.
Department of Experimental Neurology, Center for Stroke Research Berlin, Small Animal Imaging Center, Charité-University Medicine Berlin, Charitéplatz 1, Berlin, Germany.
Peyman
P
Bahmani
Eyk
E
Schellenberger
Jan
J
Klohs
Jens
J
Steinbrink
Ryan
R
Cordell
Marietta
M
Zille
Jochen
J
Müller
Denise
D
Harhausen
Leo
L
Hofstra
Chris
C
Reutelingsperger
Tracy Deanne
TD
Farr
Ulrich
U
Dirnagl
Andreas
A
Wunder
D000818:Animals; D017304:Annexin A5; D015415:Biomarkers / Q000032:analysis; D002545:Brain Ischemia / Q000473:pathology*; D002232:Carbocyanines; D016923:Cell Death / Q000502:physiology; D005456:Fluorescent Dyes; D020287:In Situ Nick-End Labeling; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence / Q000379:methods*; D011419:Propidium; D013194:Staining and Labeling / Q000379:methods
D017304:Annexin A5; D015415:Biomarkers; C098793:CY5.5 cyanine dye; D002232:Carbocyanines; D005456:Fluorescent Dyes; D011419:Propidium
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