Summarize this. Exposure to war results in a plethora of biological impacts, described below and summarised in Figure 1. The physiological effects of war trauma seen in veterans with PTSD include co-morbid chronic pain (which may reduce alongside PTSD treatment) (52, 53). These findings were applicable to both male and female veterans (53), although the lack of data pertaining to female veterans often makes sex-specific effects difficult to explore. Other pathologies incurred from war include a higher incidence of peptic ulcers in veterans and increases in dyspepsia and heartburn (2). Australian combat prisoners of war (POWs) in World War II (WWII) Japanese camps exhibited an increased rate of gastrointestinal ulcers (1). Children exposed to war exhibit an increased likelihood of irritable bowel syndrome (IBS), a condition also more common in veteran populations (4). An increased risk of obesity was observed in male veterans but, when adjusted for, were no more likely to exhibit hypertension or diabetes (7).

One group found no change in POW medical mortality (54) but, later, found a clear increase at 5–14 years after WWII, which reduced after this point. This indicates an individual's age at the time of war ending may influence mortality (55). Costa et al. also highlighted an age-related effect; after 30, the age at which a POW became imprisoned resulted in a range of differing effects during ageing. Survivors with the worst conditions showed reduced mortality versus non-POWs (or POWs when conditions were not so poor) 35 years post-imprisonment (56). An established increased risk of mortality through ischemic heart disease was found to be even more prevalent after 75 years of age (57, 58). POWs aged younger than 30 years showed an increased risk of cardio- and cerebrovascular disease as well as morbidity from the former (56). As an increased risk of cardiovascular disease is also seen in those prenatally exposed to famine, the malnutrition experienced by POWs may therefore convolute specific impacts of war (e.g., combat exposure) (59, 60). During WWII, the genocide of six million European Jews and other ethnic and societal minorities occurred. Studies on individuals exposed to the Shoah provide information on those experiencing the effects of war through torture, and genocide, but also severe famine. Despite these impacts, small-scale Holocaust studies did not reveal an increased risk of mortality at two timepoints [20–41 (61) and 40–50 (62) years after WWII].

As with other civilian populations, veterans (as well as those exposed to the Holocaust 63), despite often showing increased resilience, are at risk of mental health conditions such as depression and anxiety, which negatively impact on quality of life (64–69). A reduced expression of brain-derived neurotrophic factor (BDNF) is exhibited in depression (70) and has been found to be dysregulated in PTSD/traumatic brain injury (TBI) in veterans (71). In addition, one study revealed the increased methylation of the BDNF promoter region is associated with a PTSD diagnosis in Vietnam War veterans (72). Furthermore, older veterans with PTSD exhibit a twofold increase in the risk for dementia (73, 74). Veterans showed increased levels of depression 40 years on compared to non-POWs; however, in this study, there were no differences in alcohol issues or anxiety (54).

As mentioned above regarding civilian groups with PTSD, lowered cortisol has also been shown in combat veterans and Holocaust survivors with PTSD (75–77). The hormone and neurotransmitter noradrenaline, the mediator of the HPA-stress system, was present at the baseline in higher levels in the cerebrospinal fluid (CSF) from male combat veterans with PTSD, an effect not replicated in blood plasma (78). Blood plasma again was reported by Yahyavi et al. to show normal levels of adrenaline and noradrenaline in the plasma of veterans with PTSD (and their offspring) but decreased cortisol in those with PTSD (79). A study using CSF samples from 52 veterans with trauma revealed no significant difference in noradrenaline levels between those with or without PTSD/trauma when taking the medication prazosin (which treats high blood pressure and heart failure). However, in those not taking the drug, a significantly positive correlation between behavioural symptoms (e.g., depression and insomnia) and higher noradrenaline levels was revealed (80). The authors suggest trauma exposure may influence responsiveness to noradrenaline, and these data reveal other medication may influence hormone levels.

Above, we have discussed the physiological ramifications of war and conflict exposure, such as gastrointestinal upset (included in Figure 1). We will further expand upon this in the next section while specifically discussing epigenetic alterations (summarised in Table 1), predominantly pertaining to gene methylation. Monoamine oxidase A (MAO-A) is an enzyme critically involved in catalysing adrenaline, noradrenaline, dopamine and serotonin (102). Hypermethylation of the MAO-A gene is linked to PTSD (35) and was suggested as a tool to assess the severity of the disorder in male patients with PTSD due to war exposure; a sex-linked effect was seen, although the female sample size was comparatively small (see Table 1) (98). Gene hypermethylation often results in reduced expression, in this case potentially impeding catabolic activity on noradrenaline resulting in increased circulating levels upon stress exposure. Such dysregulation of stress hormones is a hallmark of PTSD.

In combat veteran studies, those with PTSD exhibited NR3C1 hypermethylation when compared to those without PTSD, an effect not seen in those exposed to childhood trauma (31, 35, 100). The impact of the Tutsi genocide in Rwanda was studied in 25 pregnant women and found higher CpG methylation in the nuclear receptor subfamily 3 group C member 2 (NR3C2; a mineralocorticoid receptor, also binds glucocorticoids) gene (103). The increased methylation of spindle and kinetochore-associated protein 2 (SKA2) plays a role in GR chaperoning, providing negative feedback to the HPA by removing the receptor from the cytoplasm and to the nucleus (104). Blood methylation of SKA2 and reduced expression, along with aberrant glucocorticoid signalling, was identified as a potential indicator for suicide risk (105, 106). In veterans, a positive correlation of SKA2 methylation in the blood was seen with severity of PTSD symptoms (96).

Sleep difficulties are often reported by veterans (8), including those with PTSD (107), making these epigenetic changes an area of interest. Indeed, an increase in methylation on the circadian clock gene, LHX1, has been seen in veterans with PTSD (81). Impacts on sleep-related genes were found in military personnel with chronic blast exposure, including differential methylation of NTSR1 and SPON1 (5), and differentially methylated regions (DMRs) for CCDC68 and COMT (also linked to sleep difficulties experienced by the individuals). An earlier study by the same group found, concurrent with impacts on sleep, chronic exposure to blasts resulted in the increased methylation of the paired box gene 8 (PAX8) antisense transcript, related to repressed gene expression (6). However, another study found decreased PAX8 methylation in veterans with PTSD (88). PAX8 is associated with thyroid function (108) as well as sleep duration (109). Sleep after exposure to emotional information was shown to consolidate memory up to 4 years later, so initial sleep disruption may be beneficial to reduce PTSD severity (110). However, this is a short-term benefit, given that persistently poor sleep impedes mental health (111). To alleviate PTSD, attempts to restore effective sleep should be undertaken; gene methylation may offer a useful marker of success.

Chronic blast exposure also revealed the differential methylation of auditory function genes (KCNN3, SOD3, MUC3, GALR1 and WDR45B) which were associated with tinnitus, and the DMRs within FMOD and TNXB were linked to headache and pain (5). Symptoms of tinnitus in low versus high blast exposure groups were also linked to DMRs in auditory genes KCNE1 and CYP2E1, showing an inverse correlation with methylation and expression (6). Finally, pre- versus post-blast exposure showed at least a 1.5-fold difference in expression in 67 genes, such as UFC1 and YOD1 (ubiquitin related proteins; others have been linked to TBI). The dysregulation of cytokine and chemokines was also exhibited in MCP-1, GCSF, HGF, MCSF, and RANTES after acute exposure to blasts (6). Blast exposure can result in neurological impacts, such as brain swelling and neurotrauma; these epigenetic markers are evidence of both the psychological and physical impacts of war (5, 6).

PTSD has been associated with increased immunological inflammation (112). A number of immunological changes were characterised in a study on combat veterans with PTSD (99). First, there was a significant increase in the numbers of (1) T-helper 1 (Th1; CD4+), (2) Th17, (3) cytotoxic T (CD8+), and (4) B cells. It was found that an increase in Th1 percentage was correlated with severity of PTSD score, as well as a reduction in the number of T-regulatory cells. These data suggest a heightened inflammatory response with reduced regulation. Reflecting this, upregulated levels of the proinflammatory cytokines interferon gamma (IFN-γ) and IL-17 were seen in the plasma. Zhou et al. then revealed that these changes were associated with the downregulation of the miRNA, miR-125a (as well as miR-181c), and went on to show the inhibitory effect of miR-125a on IFN-γ release in vitro (99). This signalling molecule has been linked to the upregulation of another proinflammatory cytokine, NF-κB, in human cancer cell lines (113). In rats, NF-κB is necessary for memory reconsolidation (e.g., to allow for alterations of negative associations) (114, 115). This suggests that the cytokine cascade in PTSD is dysregulated. Regular amounts of miR-125a would control the inflammatory response; by reducing IFN-γ and increasing NF-κB, a reduction of trauma responses may occur.

Further exploring the immunological impacts, the largest epigenetics study to date was performed on mixed veteran and civilian populations with and without PTSD (see Table 1) (97). The aryl-hydrocarbon receptor repressor (AHRR), a gene that regulates transcription, revealed four CpG sites in which DNA methylation was lower in individuals with PTSD versus the controls. This was found to be associated with the reduced kynurenine levels also seen in individuals with PTSD (97). The authors suggest that the reduced levels of kynurenine (a metabolite that promotes T-regulatory cell differentiation) may explain why increased inflammation is seen in individuals with PTSD. This form of gene methylation appears in both military and civilian populations with PTSD, which suggests that the HSE is not evident in this instance.

Differential expression of 25 genes associated with HPA, immune and/or cerebral function was seen in survivors of the 9/11 attack with PTSD (101). The altered expression of FKBP5 was revealed as an acute state marker for PTSD as well as major histocompatibility complex (MHC) Class II (vs. lifetime PTSD). In addition, the reduced expression of Stat five B inhibitor (STAT5B; a GR inhibitor 116) and nuclear factor I/A (key for astrocytic function 117) was seen in PTSD. These genes reveal that epigenetic impacts from PTSD impact on both the HPA axis and the immune system.

Rutten et al. compared the longitudinal changes of genome-wide blood DNA methylation profiles of two different subgroups of soldiers and identified the genes associated with stress in a military context (included in Table 1) (88). In the first cohort, consisting of Dutch military personnel, they discovered 17 differentially methylated positions (DMPs) and 12 DMRs in individuals after they had been deployed to a combat zone. The DMPs and DMRs were then used to explore replication in a different cohort of US marines. In this group, a decrease in DNA methylation at the genes ZFP57, RNF39, and HIST1H2APS2 was associated with an increase in PTSD symptoms over time. They found that the association between trauma exposure and PTSD symptoms was mediated by DNA methylation; those who did not exhibit PTSD symptoms showed increased methylation, but those who did develop PTSD symptoms exhibited decreased DNA methylation at the DMP and DMRs. Altered methylation on DUSP22 and PAX8 genes, earlier identified, revealed decreased methylation in both, with no change in expression to the former but increased expression in the latter (6, 88).

PTSD is often defined solely by symptom severity, despite the heterogenous nature of the symptoms themselves. To challenge this perception, Yang and colleagues defined the epigenetic biotypes of veterans/active-duty personnel and propensity for PTSD (118). They revealed two main subsets with opposing symptoms: G1 (faster recovery from PTSD, lower methylation vs. controls) and G2 (increased risk of PTSD, higher methylation vs. controls), enriched for individuals also exhibiting major depressive disorder (MDD). That study highlights the complex aetiology of PTSD and variation within military subsets. However, the authors did not discuss immunological components or pro-inflammatory processes, which also contribute to PTSD (99, 112, 119). The identification of if, and how, G1 and G2 express different immunological profiles would inform on whether immune function also differs with opposing symptoms.

Therefore, there is a large body of research exploring a plethora of epigenetic changes upon the individual exposed to war (listed with details in Table 1), which we also attempt to summarise here. As noted earlier, the FKBP5 and NR3C1 genes are epigenetically modified; in addition, research on war and conflict exposure has revealed epigenetic impacts to players in auditory, sleep and immune systems.