APOPTOSIS OF MOTOR-NEURONS WITH INDUCTION OF CASPASES IN THE SPINAL-CORD AFTER ISCHEMIA

Background and Purpose-Some neuronal subpopulations are especially vul nerable to ischemic injury. In the spinal cord, large motor neurons ar e vulnerable to ischemia and are selectively lost after transient isch emia. However, the mechanisms of the neuronal loss have been uncertain . We hypothesized that spinal motor neurons might be lost by apoptosis and investigated a possible mechanism of neuronal death by detection of double-strand breaks in genomic DNA and immunohistochemical analysi s for caspases, ie, interleukin-1 beta converting enzyme (ICE), Nedd-2 , and CPP32, Methods-We used a rabbit spinal cord ischemia model creat ed with a balloon catheter. The spinal cord was removed at 8 hours, 1, 2, or 7 days after 15 minutes of transient ischemia, and histological changes were studied with hematoxylin-eosin staining. To detect doubl e-strand breaks in DNA, a staining with terminal deoxynucleotidyl tran sferase-mediated dUTP-biotin in situ nick end labeling (TUNEL) was per formed. Furthermore, expression of ICE, Nedd-2, and CPP32 was investig ated by Western blotting and immunohistochemical analysis. Results-Mot or neurons were selectively lost at 7 days after transient ischemia. T UNEL study demonstrated that no cells were positively labeled until 1 day after ischemia, but nuclei of some motor neurons were positively l abeled at 2 days. Western blot analysis revealed no immunoreactivity f or ICE and slight immunoreactivities for Nedd-2 and CPP32 in the sham- operated spinal cords. However, immunoreactivity became apparent at 8 hours after transient ischemia, decreased at 1 day, and returned to ba seline level at 2 days. Immunohistochemical analysis demonstrated that motor neurons were responsible for induction of those caspases. Concl usions-Double-strand breaks in genomic DNA and induction of three casp ases were demonstrated. These results indicate that motor neuron death in the spinal cord after transient ischemia is profoundly associated with activation of apoptotic processes.