E-64-d prevents both calpain upregulation and apoptosis in the lesion and penumbra following spinal cord injury in rats

Calpain, a Ca2+-dependent cysteine protease, has been implicated in cytoske letal protein degradation and neurodegeneration in the lesion and adjacent areas following spinal cord injury (SCI). To attenuate apoptosis or Program med cell death (PCD) in SCI, we treated injured rats with E-64-d, a cell pe rmeable and selective inhibitor of calpain. SCI was induced on T12 by the w eight-drop (40 g-cm force) method. Within 15 min, E-64-d(1 mg/kg) in 1.5% D MSO was administered i.v. to the SCI rats. Following 24 h treatment, a 5-cm long spinal cord section with the lesion in the center was collected. The spinal cord section was divided equally into live 1-cm segments (S1: distan t rostral, S2: near rostral, S3: lesion or injury, S4: near caudal and S5: distant caudal) for analysis. Determination of mRNA levels by reverse trans criptase-polymerase chain reaction (RT-PCR) indicated that ratios of bax/bc l-2 and calpoin/calpastatin were increased in spinal cord segments from inj ured rats compared to controls. Degradation of the 68-kD neurofilament prot ein and internucleosomal DNA fragmentation were also increased. All of thes e changes were maximally increased in the lesion and gradually decreased in the adjacent areas of SCI rats, while largely undetectable in E-64-d treat ed rats and absent in sham controls. The results indicate that apoptosis in rat SCI appears to be associated with calpain activity which can he attenu ated by the calpain inhibitor E-64-d. (C) 2000 Elsevier Science B.V. All ri ghts reserved.