Year of Publication

2013

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Medicine

Department

Physiology

First Advisor

Dr. Kathryn E. Saatman

Abstract

Traumatic brain injury (TBI) is a devastating health problem based on its high incidence, economic burden, and lack of effective pharmacological treatment. Individuals who suffer an injury often experience lifelong disability. TBI results in abrupt, initial cell damage leading to delayed neuronal death. The calcium-activated proteases, calpains, are known to contribute to this secondary neurodegenerative cascade. Prolonged activation of calpains results in proteolysis of numerous cellular substrates including cytoskeletal components, membrane receptors, and cytosolic proteins, contributing to cell demise despite coincident expression of calpastatin, the specific inhibitor of calpains.

A comprehensive analysis using two separate calpastatin transgenic mouse lines was performed to test the hypothesis that calpastatin overexpression will reduce posttraumatic calpain activity affording neuroprotection and behavioral efficacy. Increased calpastatin expression was achieved using transgenic mice that overexpress the human calpastatin (hCAST) construct under control of a neuron-specific calcium-calmodulin dependent kinase II alpha or a ubiquitous prion protein promoter. Both transgenic lines exhibited enhanced calpastatin expression within the brain, extending into peripheral tissues under the prion protein promoter. hCAST overexpression significantly reduced protease activity confirmed by reductions in acute calpain-mediated substrate proteolysis in the cortex and hippocampus following controlled cortical impact brain injury. Aspects of posttraumatic motor and cognitive behavioral deficits were also lessened in hCAST transgenic mice compared to their wildtype littermates. However, volumetric analyses of neocortical contusion revealed no histological neuroprotection at either acute or long-term time points in either transgenic line. Partial hippocampal neuroprotection observed at a moderate injury severity in neuron-specific calpastatin overexpressing transgenic mice was lost after severe TBI. Greater levels of calpastatin under the prion protein promoter line failed to protect against hippocampal cell loss after severe brain injury.

This study underscores the effectiveness of calpastatin overexpression in reducing calpain-mediated proteolysis and behavioral impairment after TBI, supporting the therapeutic potential for calpain inhibition. However, the reduction in proteolysis without accompanied neocortical neuroprotection suggests the involvement of other factors that are critical for neuronal survival after contusion brain injury. Augmenting calpastatin levels may be an effective method for calpain inhibition and may have efficacy in reducing behavioral morbidity after TBI and neurodegenerative disorders.

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Neurosciences Commons

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