Accumulation of Calpain and Caspase-3 Proteolytic Fragments of Brain-Derived αII-Spectrin in Cerebral Spinal Fluid after Middle Cerebral Artery Occlusion in Rats

Abstract

Summary:Preclinical studies have identified numerous neuroprotective drugs that attenuate brain damage and improve functional outcome after cerebral ischemia. Despite this success in animal models, neuroprotective therapies in the clinical setting have been unsuccessful. Identification of biochemical markers common to preclinical and clinical cerebral ischemia will provide a more sensitive and objective measure of injury severity and outcome to facilitate clinical management and treatment. However, there are currently no effective biomarkers available for assessment of stroke. Nonerythroid ␣II-spectrin is a cytoskeletal protein that is cleaved by calpain and caspase-3 proteases to signature ␣II-spectrin breakdown products (␣II-SBDPs) after cerebral ischemia in rodents. This investigation examined accumulation of calpain-and caspase-3-cleaved ␣II-SBDPs in cerebrospinal fluid (CSF) of rodents subjected to 2 hours of transient focal cerebral ischemia produced by middle cerebral artery occlusion (MCAO) followed by reperfusion. After MCAO injury, full-length ␣II-spectrin protein was decreased in brain tissue and increased in CSF from 24 to 72 hours after injury. Whereas ␣II-SBDPs were undetectable in sham-injured control animals, calpain but not caspase-3 specific ␣II-SBDPs were significantly increased in CSF after injury. However, caspase-3 ␣II-SBDPS were observed in CSF of some injured animals. These results indicate that ␣II-SBDPs detected in CSF after injury, particularly those mediated by calpain, may be useful diagnostic indicators of cerebral infarction that can provide important information about specific neurochemical events that have occurred in the brain after acute stroke.

αII-Spectrin Breakdown Products (SBDPs): Diagnosis and Outcome in Severe Traumatic Brain Injury Patients

“…Detection of these proteins in blood or CSF offers little insight into neurochemical alterations that mediate brain damage after TBI. In contrast, SBDP levels are related to specific cell death processes within cells (necrosis versus apoptosis, and activation of calpain versus caspase-3; Pike et al, 2004;Ringger et al, 2004;Wang et al, 1998). Using ELISA assays, this prospective observational study of 40 patients provides the first quantitative extension of previous findings employing Western blot analyses collected in humans following a severe brain injury Farkas et al, 2005;Pineda et al, 2007;).…”

“…Recently investigators have described aII-spectrin breakdown products (SBDPs) as potential biomarkers for brain injury in rats and humans (Pike et al, 2004(Pike et al, , 2001Pineda et al, 2007;Wang et al, 2005). aII-spectrin is primarily found in neurons, and is abundant in axons and presynaptic terminals (Riederer et al, 1986), and the protein is processed to breakdown products (SBDP) of molecular weights 150 kDa (SBDP150) and 145 kDa (SBDP145) by calpain, and is also cleaved to a 120-kDa product (SBDP120) by caspase-3.…”

Biomarker evidence for mild central nervous system injury after surgically-induced circulation arrest

“…Indeed, the CSF level of 14-3-3β correlates with the magnitude of acute neurodegeneration as monitored by quantitative histopathology, and is an even better predictor of brain damage than the duration of the ischemia. Other forms of acute CNS damage such as traumatic brain injury also lead to marked increases in CSF levels of the two 14-3-3 isoforms, pNFH and calpain-derived fragments of α-spectrin and tau Pike et al, 2004;. Calpain is a family of calcium-activated cysteine proteases that has been causally linked to the acute necrotic neurodegeneration that is triggered by intraneuronal calcium overload and occurs in response to ischemia, trauma, and excitotoxicity (Siman et al, 1989;Roberts-Lewis and Siman, 1993;Saatman et al, 1996;Zhang et al, 2003).…”

Exploring the Role of Biomarkers for the Diagnosis and Management of Traumatic Brain Injury Patients

“…A signature of caspase-3 and calpain-2 activation is cleavage of several common proteins such as cytoskeletal II-spectrin. 106 In a rat model, mean levels of both ipsilateral cortex (IC) and cerebral spinal fluid (CSF) spectrin breakdown products (SBDP) at 2, 6, and 24 h after two levels of controlled cortical impact (1.0 mm and 1.6 mm of cortical deformation) were significantly elevated by injury using immunoblotting. 107 Using the same proteomic Western blot technique, levels of spectrin breakdown products (SBDP's) have been reported in CSF from adults with severe TBI and they have shown a significant relationship with severity of injury and clinical outcome.…”