Acute and Chronic Molecular Signatures and Associated Symptoms of Blast Exposure in Military Breachers

Abstract

Injuries from exposure to explosions rose dramatically during the Iraq and Afghanistan wars, which motivated investigations of blast-related neurotrauma and operational breaching. In this study, military "breachers" were exposed to controlled, low-level blast during a 10-day explosive breaching course. Using an omics approach, we assessed epigenetic, transcriptional, and inflammatory profile changes in blood from operational breaching trainees, with varying levels of lifetime blast exposure, along with daily selfreported symptoms (with tinnitus, headaches, and sleep disturbances as the most frequently reported). Although acute exposure to blast did not confer epigenetic changes, specifically in DNA methylation, differentially methylated regions (DMRs) with coordinated gene expression changes associated with chronic lifetime cumulative blast exposures were identified. The accumulative effect of blast showed increased methylation of PAX8 antisense transcript with coordinated repression of gene expression, which has been associated with sleep disturbance. DNA methylation analyses conducted in conjunction with reported symptoms of tinnitus in the low vs. high blast incidents groups identified DMRS in KCNE1 and CYP2E1 genes. KCNE1 and CYP2E1 showed the expected inverse correlation between DNA methylation and gene expression, which have been previously implicated in noise related hearing loss.Although no significant transcriptional changes were observed in samples obtained at the onset of the training course relative to chronic cumulative blast, we identified a large number of transcriptional perturbations acutely pre-versus post-blast exposure. Acutely, 67 robustly differentially expressed genes (fold change ≥ 1.5), including UFC1 and YOD1, ubiquitin-related proteins were identified. Inflammatory analyses of cytokines and chemokines revealed dysregulation of MCP-1, GCSF, HGF, MCSF, and RANTES acutely following blast exposure. These data show the importance of an omics approach, revealing that transcriptional and inflammatory biomarkers capture acute lowlevel blast overpressure exposure, whereas DNA methylation marks encapsulate chronic long-term symptoms.

DNA Methylation Patterns of Chronic Explosive Breaching in U.S. Military Warfighters

“…We previously reported that DNA methylation changes do not track with acute exposure to blast (21), and our present findings further support this in that DNA methylation appears to not be altered acutely pre-vs. post-blast exposure. As noted, for this experiment we used biospecimens from one site, FLWB, and examined DNA methylation changes in day-2 of training, pre vs. post blast exposure (approximately within 2 h post blast exposure, see Figure 1A for training protocol).…”

“…In the present study of military breachers, we set out to replicate previous findings, investigating effects of acute and chronic blast exposure on DNA methylation and reported symptoms at baseline across two military training sites-representing the largest DNA methylation study of breachers to date. In previous work, we found no significant DNA methylation changes associated with acute blast exposure (3-days post blast) (21). In the present study, we investigated whether blast induced DNA methylation changes could be detected at a more proximal time point, within 2-h post-blast exposure.…”

“…We have previously shown that lifetime cumulative blast exposure is capable of altering DNA methylation in our investigations of military breachers with high relative to low lifetime blast exposures at baseline among military service members participating in a 10-days explosive breaching training course (21). Here, we also investigate at baseline, alterations in DNA methylation in 59 military trainees from two independent cohorts over a 3-days period during the explosive breaching course at Fort Leonard Wood (FLWA and FLWB sites, Figure 1A).…”

“…Based on concerns for potential injury by the cadre of breachers and instructors, the Department of Defense (DOD) has been conducting studies on the bio-effects from repeated exposure, and findings from our own group (21) and others (22,23) have begun to show that blast exposure during training is capable of inducing changes in DNA methylation and gene expression in military breachers (22) that track with the physiological symptoms of blast injury (24). In the present study of military breachers, we set out to replicate previous findings, investigating effects of acute and chronic blast exposure on DNA methylation and reported symptoms at baseline across two military training sites-representing the largest DNA methylation study of breachers to date.…”

Factors Contributing to Increased Blast Overpressure inside Modern Ballistic Helmets

“…The blast wave propagation inside combat helmets is a complex problem, with the pressure focusing under the helmet dependent on several factors: 1) the shape of the helmet; 2) orientation relative to the source of the blast wave, and 3) the suspension system (i.e., number of pads, their distribution under the helmet and material properties). The mounting evidence suggests an association between occupational low-level blast exposure and acute adverse neurological effects [26], and related serum biomarkers changes [27,28]. The role of the helmet in the development of these effects is unclear at this time.…”