(Stroke. 1997;28:2012-2017.)
© 1997 American Heart Association, Inc.
Articles |
Cell Death Suggestive of Apoptosis After Spinal Cord Ischemia in Rabbits
From the Division of Cardiothoracic Surgery (M.E.M., G.K.K, N.T.K.) and Department of Neurology (Y.W., R.H., J.A.D., M.F.J., C.Y.H.), Washington University School of Medicine, St Louis, Mo.
Correspondence to Nicholas T. Kouchoukos, MD, 3009 N Ballas Rd, Room 266, St Louis, MO 63131.
Background and Purpose After spinal cord ischemia, some neurons remain viable after an ischemic insult but may be at risk of dying during reperfusion. We searched for morphological and biochemical features of apoptosis, which is a mechanism of delayed neuronal death, in a rabbit model of spinal cord ischemia.
Methods The infrarenal aorta of White New Zealand rabbits (n=24) was occluded for 40 minutes using a loop tourniquet. Rabbits were killed after 12, 24, or 48 hours (n=8 per group). The loop was placed but never tightened in sham-operated rabbits (n=6). The lumbar segment of the spinal cord (L5 to L7) was used for morphological studies, including hematoxylin and eosin staining and a modified terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end-labeling (TUNEL) staining method. Electron microscopy was used to examine ultrastructural morphology. In addition, lumbar tissue was used for biochemical investigation of DNA laddering by agarose gel electrophoresis.
Results After ischemia, the affected areas
contained neurons with positive TUNEL staining. Positive neurons were
located in laminae III to IX, although most were concentrated in the
intermediate and ventral areas. Adjacent sections stained with
hematoxylin and eosin exhibited ischemic cell changes (red and
ghost neurons), while apoptotic bodies were also apparent. In
addition, electron microscopy of ischemic tissue samples
exhibited ultrastructural characteristics of apoptosis,
including nuclear condensation and relatively normal organelle
morphology. Finally, isolated DNA revealed a ladder on agarose gel
electrophoresis, indicating DNA fragmentation into 
180 multiples of
base pairs.
Conclusions Spinal cord ischemia in rabbits induces morphological and biochemical changes suggestive of apoptosis. These data raise the possibility that apoptosis contributes to neuronal cell death after spinal cord ischemia.
Key Words: apoptosis • neuronal death • reperfusion • spinal cord • rabbits
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