Grants and Contracts Details
Description
Traumatic brain injury (TBI) afflicts approximately 1.4 million individuals in the U.S. each year.
Following TBI, a variety of cellular mediators contribute to neuronal death and dysfunction including the
cysteine proteases, calpains. Prolonged activation of calpains occurs within neurons due to a rapid and
sustained rise of intracellular free calcium. Although an endogenous inhibitor of calpains, calpastatin, is coexpressed,
the sustained activation of these calcium-dependent proteases suggests endogenous calpastatin
levels may be insufficient. The overall hypothesis of this proposal is that overexpression of calpastatin will
reduce the proteolytic activity of calpains and associated neuronal death after trauma, thereby attenuating
motor and cognitive deficits. Calpastatin overexpression will be induced in two ways-transgenic
overexpression of human calpastatin (hCAST) (Aim 1) and calpastatin expression via lentiviral vector delivery
into brain regions vulnerable to TBI (Aim 2).
Aim 1 will use a novel transgenic mouse line with human calpastatin under control of the ubiquitous
prion promoter. This mouse line exhibits a 9-fold greater expression of calpastatin in the cortex and
hippocampus compared to wildtype mice. hCAST transgenic and wildtype littermates will be subjected to
severe controlled cortical impact (CCI) injury or sham treatment. To confirm that calpastatin overexpression
decreases calpain activity, cortical and hippocampal homogenates will be evaluated for calpain-mediated
cytoskeletal and membrane protein breakdown via immunoblot. Both motor and cognitive functions will be
assessed after injury, after which mice will be euthanized for analysis of hippocampal neurodegeneration and
cortical tissue damage to assess the neuroprotective actions of hCAST overexpression. The expectation is that
hCAST transgenic mice will have reduced posttraumatic calpain proteolytic activity, offering a neuroprotective
advantage. Aim 2 establishes an alternative approach to hCAST overexpression through lentiviral vector
delivery. After injection of controllentivirus or calpastatin lentivirus into the cortex or hippocampus, mice will be
subjected to 1.0mm CCI brain injury or sham treatment and assessed as in Aim 1. Targeting of calpastatin to
vulnerable neuronal regions prior to injury should spare affected neurons and reduce behavioral deficits.
Project Description
Status | Finished |
---|---|
Effective start/end date | 1/5/11 → 1/4/12 |
Funding
- National Institute of Neurological Disorders & Stroke: $31,408.00
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