Using Drosophila Melanogaster as an Integrated Model to Study Traumatic Brain Injury

Overview

Background: Traumatic brain injury (TBI) is a leading international cause of morbidity and mortality. The neurologic outcome of TBI is significantly influenced by the cellular and molecular responses (secondary injury) to the initial impact, which are poorly understood. The goals of this project were to: 1) To assess the level of apoptosis by immunolabeling cleaved Caspase-3 in brains of Drosophila subjected to TBI, 2) To utilize qRT-PCR to quantify the level of expression of 2 anti-microbial peptides (AMP) genes (Attacin C and Diptericin B) after TBI, thereby assessing the response of the innate immune system, and 3) To detect the loss of intestinal barrier function in Drosophila following TBI, by the presence of a non-absorbable blue food dye outside of the digestive tract after feeding. Methods: A “high-impact trauma” (HIT) device was built, which used a spring-based mechanism to propel flies against the wall of a vial, causing mechanical damage to the brain. An antibody to cleaved-Caspase-3 was used as a general apoptotic marker in whole Drosophila brains. The immune response was assessed by quantifying Anti-Microbial Peptides (AMP) gene expression, using qRT-PCR. Loss of intestinal barrier function was detected by the presence of a non-absorbable blue food dye (FD&C blue dye no. 1) outside of the digestive tract after feeding. Results: Increased apoptosis was detected in Drosophila brains subjected to TBI, by measuring cleaved caspase. TBI flies showed an enhanced innate immune response as measured by increased gene expression of the anti-microbial peptide, Diptericin B. More flies with TBI had the “Smurf” phenotype, compared with controls, demonstrating that impaired gut permeability is a non-neuronal effect of severe TBI. Conclusion: TBI causes distinct biochemical and physiological alterations. An improved understanding of these secondary sub-cellular mechanisms of TBI is a vital prerequisite for developing effective interventions.