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Abstract:
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Traumatic Brain Injury (TBI) annually affects 1.5 million individuals in the United States and associated costs soar to over $56 billion. Despite its widespread burden, relatively little is known about how to reduce the damage that occurs after TBI, and most attempts to develop successful therapies have failed. A large percentage of TBI patients are over the age of 75, but it remains to be discovered how a person’s age impacts the level and nature of the biological responses to the injury: edema, inflammation, necrosis, and apoptosis. The purpose of this project was to aid in the development of a murine model of age-related TBI damage and repair. Methods: Brains from mice with unilateral cortical injuries were sectioned on a cryostat. Sections were processed using immunohistochemical procedures with antibodies to astrocyte marker glial fibrillary acidic protein (GFAP), inflammatory mediator phosphorylated c-Jun N-terminal kinase (pJNK), and the apoptosis marker activated Caspase-3. Tissues were also processed with Fluoro-Jade B, a marker for degenerating neurons. The methods were fine-tuned to provide a high signal/background ratio. Results: A significantly improved signal/background ratio was obtained following repeated optimizing trials, and procedural details were formulated to allow reproducible, highly specific results. High-contrast Fluoro-Jade B signals specific to neurons and highly localized anti-GFAP and anti-activated Caspase-3 signals were obtained in injured tissues. Conclusions: Fluoro-Jade B is useful for identifying dying neurons following TBI, while GFAP and Caspase-3 antibodies are useful markers for studying signals related to tissue damage. Further study will characterize the locations and quantify the extent of these signals and seek to determine their relationship to injury severity and age. These data should prove useful in advancing our understanding of the mechanisms of age-related TBI damage and repair. |
