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Abstract
Reduction in Posttraumatic Calpain-Mediated Cleavage of Spectrin and Sodium Channel Protein with Calpastatin Overexpression
Journal of Neurotrauma, v 28(6), P244
20 Jun 2011
Abstract
Calpains, calcium-activated neutral proteases, are activated early after traumatic brain injury (TBI), with subsequent proteolysis of substrates including cytoskeletal components and membrane receptors. The prolonged activity of calpains after trauma suggests that the action or levels of its endogenous inhibitor, calpastatin, may be insufficient to fully inhibit the proteolytic activity of calpains. Therefore, we hypothesize that calpastatin overexpression via the prion promoter (PrP-hCAST) will reduce calpain activity and calpain-mediated substrate proteolysis after TBI. Previously, we showed that cortical protease activity via fluorogenic assay was lower in homogenates obtained from transgenic mice (n = 6) compared to wildtype (WT) mice (n = 5; p < 0.001). Here we demonstrate that hCAST overexpression results in reduced posttraumatic cytoskeletal and membrane channel cleavage. Contusion brain injury was created with a controlled cortical impact (CCI) device, transiently indenting the exposed cortex to a 1.0mm depth. Immunoblots for alpha-spectrin revealed that PrP-hCAST mice (n = 6) exhibited significantly less cortical and hippocampal calpain-mediated proteolysis than WT (n = 5-7) at 6 hours (p < 0.0.01) and 24 hours (p < 0.005) post-injury. Brain injury resulted in a reduction in cortical full-length voltage-gated sodium channel (NaCh) protein, coupled with the appearance of fragments detected by pan, Nav1.2 I-II loop and C-terminus-specific antibodies. PrP-hCAST mice (n = 4) exhibited a significant reduction in NaCh breakdown products detected by pan, Nav1.2 I-II loop, and Nav1.2 C-terminal antibodies compared to WT (n = 4) at 6 h. At 24 h post-CCI, an 85 kDa Nav1.2 I-II loop fragment was also attenuated in PrP-hCAST mice (n = 4) compared to WT (n = 5); however, other sodium channel fragments were equivalent. Considering the structural and functional importance of spectrin and NaCh dynamics in neurons, preservation of these proteins after injury may contribute to a reduction in neuronal damage or dysfunction. Future studies will continue to investigate sparing of calpain substrates with hCAST overexpression, supporting the therapeutic potential for augmenting endogenous calpastatin levels.
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Details
- Title
- Reduction in Posttraumatic Calpain-Mediated Cleavage of Spectrin and Sodium Channel Protein with Calpastatin Overexpression
- Creators
- Kathleen M. Schoch - University of KentuckyCatherine R von Reyn - University of PennsylvaniaGlenn C. Telling - University of KentuckyDavid F. Meaney - University of PennsylvaniaKathryn E Saatman - University of Kentucky
- Publication Details
- Journal of Neurotrauma, v 28(6), P244
- Conference
- 29th Annual National Neurotrauma Symposium, 29th (Hollywood Beach, Florida, United States, 10 Jul 2011–13 Jul 2011)
- Publisher
- Mary Ann Liebert
- Number of pages
- 1
- Resource Type
- Abstract
- Academic Unit
- School of Biomedical Engineering, Science, and Health Systems
- Other Identifier
- 991019289315704721