μ‐Calpain Activation and Calpain‐Mediated Cytoskeletal Proteolysis Following Traumatic Brain Injury

Abstract

Increasing evidence suggests that excessive activation of the calcium‐activated neutral protease μ‐calpain could play a major role in calcium‐mediated neuronal degeneration after acute brain injuries. To further investigate the changes of the in vivo activity of μ‐calpain after unilateral cortical impact injury in vivo, the ratio of the 76‐kDa activated isoform of μ‐calpain to its 80‐kDa precursor was measured by western blotting. This μ‐calpain activation ratio increased to threefold in the pellet of cortical samples ipsilateral to the injury site at 15 min, 1 h, 3 h, and 6 h after injury and returned to control levels at 24–48 h after injury. We also investigated the effect of μ‐calpain activation on proteolysis of the neuronal cytoskeletal protein α‐spectrin. Immunoreactivity for α‐spectrin breakdown products was detectable within 15 min after injury in cortical samples ipsilateral to the injury site. The levels of α‐spectrin breakdown products increased in a biphasic manner, with a large increase between 15 min and 6 h after injury, followed by a smaller increase between 6 and 24 h after the insult. No further accumulation of α‐spectrin breakdown products was observed between 24 and 48 h after injury. Histopathological examinations using hematoxylin and eosin staining demonstrated dark, shrunken neurons within 15 min after traumatic brain injury. No evidence of μ‐calpain autolysis, calpain‐mediated α‐spectrin degradation, or hematoxylin and eosin neuronal pathology was detected in the contralateral cortex. Although μ‐calpain autolysis and cytoskeletal proteolysis occurred concurrently with early morphological alterations, evidence of calpain‐mediated proteolysis preceded the full expression of evolutionary histopathological changes. Our results indicate that rapid and persistent μ‐calpain activation plays an important role in cortical neuronal degeneration after traumatic brain injury. Our data also suggest that specific inhibitors of calpain could be potential therapeutic agents for the treatment of traumatic brain injury in vivo.

Therapy Development for Diffuse Axonal Injury

“…In particular, activated calpain has been well studied with regard to proteolysis of subaxolemmal spectrin, with the resultant breakdown products detected in damaged axons and in the CSF. 38,41,42,133,134,165,168,[170][171][172] Less well known is the role of calpain on ankyrin proteolysis, which may lead to disordered sodium channel arrangement at the nodes of Ranvier, and its altered binding to neurofascin, which may contribute to axolemmal instability. 173 Several in vivo studies have examined various axonal protective strategies targeting calcium-mediated responses by either direct or indirect means.…”

A Pharmacological Analysis of the Neuroprotective Efficacy of the Brain- and Cell-Permeable Calpain Inhibitor MDL-28170 in the Mouse Controlled Cortical Impact Traumatic Brain Injury Model

“…Additionally, the apoptosis-associated protease caspase-3 can cleave spectrin to produce a unique 150-kDa degradation fragment that is further cleaved to a 120-kDa product (Wang, 2000). a-Spectrin cleavage has been used extensively as a quantified measure of calpain activity in both animal models of traumatic brain injury (TBI; Buki et al, 1999;Kampfl et al, 1996;Kupina et al, 2003aKupina et al, , 2001Kupina et al, , 2002Mbye et al, 2009;Saatman et al, 1996), and more recently in TBI patients (Brophy et al, 2009). Based on these published reports, the neuroprotective potential of the calpain inhibitors may be predicted by their ability to attenuate post-traumatic spectrin degradation.…”

Down-Regulation of Brain-Pancreas Relative Protein in Diabetic Rats and by High Glucose in PC12 Cells: Prevention by Calpain Inhibitors

“…It has been implicated in a large number of physiological processes and pathological conditions, including susceptibility to diabetes (26,27). Increased calpain activity has been associated with neuronal degeneration caused by traumatic events such as excitotoxicity and hypoxia / ischemia (28,29). The calpain inhibitor was shown to correct penile nitrergic nerve deficit in diabetic mice (30).…”