Age and Diet Affect Genetically Separable Secondary Injuries that Cause Acute Mortality Following Traumatic Brain Injury in Drosophila

Abstract

Outcomes of traumatic brain injury (TBI) vary because of differences in primary and secondary injuries. Primary injuries occur at the time of a traumatic event, whereas secondary injuries occur later as a result of cellular and molecular events activated in the brain and other tissues by primary injuries. We used a Drosophila melanogaster TBI model to investigate secondary injuries that cause acute mortality. By analyzing mortality percentage within 24 hr of primary injuries, we previously found that age at the time of primary injuries and diet afterward affect the severity of secondary injuries. Here, we show that secondary injuries peaked in activity 1–8 hr after primary injuries. Additionally, we demonstrate that age and diet activated distinct secondary injuries in a genotype-specific manner, and that concurrent activation of age- and diet-regulated secondary injuries synergistically increased mortality. To identify genes involved in secondary injuries that cause mortality, we compared genome-wide mRNA expression profiles of uninjured and injured flies under age and diet conditions that had different mortalities. During the peak period of secondary injuries, innate immune response genes were the predominant class of genes that changed expression. Furthermore, age and diet affected the magnitude of the change in expression of some innate immune response genes, suggesting roles for these genes in inhibiting secondary injuries that cause mortality. Our results indicate that the complexity of TBI outcomes is due in part to distinct, genetically controlled, age- and diet-regulated mechanisms that promote secondary injuries and that involve a subset of innate immune response genes.

Expansion of a fly TBI model to four levels of injury severity reveals synergistic effects of repetitive injury for moderate injury conditions

“…Animals subjected to TBI experience both primary injury from the TBI event itself and secondary injuries related to cellular and molecular events instigated by the primary injury. The secondary injury period in flies reportedly peaks between 1 h and 8 h post-injury and persists for at least 24 h (Katzenberger et al 2016). We hypothesized that the relative contributions of primary and secondary injuries on mortality would differ when comparing a single severe injury (90° × 1HIT) to repetitive injury at less severe angles.…”

“…The exact secondary mechanisms underlying the fast synergistic effects we observed are thus far unknown. Moreover, we do not know if synergistic effects at mild to moderate TBI conditions in our model drive other TBI-related consequences observed in flies such as changes in lifespan and inflammation (Katzenberger et al 2013, 2015, 2016; Barekat et al 2016; Anderson et al 2018). However, our fly model offers an unparalleled platform for rapidly, and systematically, testing candidate factors or pathways for their involvement in TBI outcomes across injury severities, number, and inter-injury interval.…”

“…It was previously reported that dividing the MI 24 by the number of HITs resulted in no differences when comparing across HIT number (Katzenberger et al 2013). This result was used as evidence that the main factor influencing MI 24 across multiple HITs was the likelihood of suffering a critical injury for each HIT, and that secondary injury mechanisms were negligible for injuries spaced closely together as the secondary injury window does not peak until 1-8 h after injury (Katzenberger et al 2013, 2016). By contrast, we found differences when comparing median MI 24 /HIT values even at these close (15 second) inter-injury intervals (Figs.…”

“…Why might 2HITs at the 80° deflection angle, but not 3HITs at 70°, cause a different pattern of death than 1HIT at 90°? One possibility is the presence of separate secondary injury mechanisms, one operating on a short time-scale (seconds) and the other the previously defined mechanism operating on a longer time-scale (hours) (Katzenberger et al 2016). By this model, the administration of 2HITs at 80° causes significant secondary injuries on the longer time-scale and greater death after 8+ h compared to 1HIT at 90° (Fig.…”

“…Importantly, use of each method results in classic post-TBI symptoms including impaired locomotion, shortened lifespan, neurodegeneration, intestinal barrier disruption, and activation of immune and autophagy processes (Katzenberger et al 2013; Barekat et al 2016; Anderson et al 2018). While the Bead Ruptor method offers potential advantages in the ease of scaling primary injuries and inter-experiment standardization, the HIT method is simple, cost-effective, and better characterized to date (Katzenberger et al 2013, 2015, 2016; Barekat et al 2016; Anderson et al 2018).…”