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9. Are the lungs of deployers seeking treatment for respiratory complaints histopathologically distinct from:

Question 9
1. PECOT QUESTION

PECOT: Are the lungs of military personnel deployed to the SWATO and who are seeking treatment for respiratory complaints histopathologically distinct from: (a) healthy individuals, (b) non-exposed military, and (c) patients with other known respiratory diseases?

Why is this question important?

ILDs are difficult to diagnose and are often misdiagnosed.[cite] Additionally, depending on individual differences (e.g., genetics, immunological history, other exposures, etc.) these diseases may manifest differently in different people. Thus, it is important to determine whether there are common underlying histopathological features (as opposed to diagnoses) in the lungs of military personnel exposed to hazards in the theater of war that distinguish them from both healthy people and non- deployed military. If these histopathological features are also known to be related to different ILDs, then this increases our confidence in a causal relation between exposure to hazards in the theater of operations and the diseases of interest.

2. Conclusion

We did not identify any research comparing histopathological findings of US military deployed to the Southwest Asia Theater of Operations (SWATO) and non-deployed US military. However, we did identify research comparing US military deployed to SWATO and (a) healthy individuals and (b) patients with other respiratory diseases. Additionally, we identified one study that compared exposed versus unexposed Iraqi soldiers. Taken together, the available research indicates that:

  • The lungs of deployed soldiers seeking treatment are histologically distinct from healthy individuals and characterized by chronic inflammation and small airway thickening (fibrosis),
  • Comparisons between exposed and non-exposed non-US soldiers indicate large differences in terms of the presence of fibrogenic markers, and
  • While the histopathological pattern of deployed military shares some features with other respiratory diseases, deployment-related distal lung disease (DDLD)1,2 appears to be distinct from other respiratory diseases. In particular, DDLD appears to have a different profile from smoking-related bronchiolitis.

Evidence Level: Considering evidence for the primary hypotheses, combined with uncertainty and evidence for the alternative hypotheses, the evidence level for this conclusion is Moderate while the evidence level for the alternative explanations is Low.

3. KNOWN BIASES

There are known biases in the research on military exposures. We summarize the direction and magnitude of these biases in the table below.

In summary, there are competing biases that may affect the reported effects. While lack of adjustment for smoking may overestimate the effect, exposure misclassification bias is likely to underestimate the effect. Based on the best estimates, these two would approximately balance out. Thus, for comparisons to non-military, the presence of known biases poses little concern. For deployed to non-deployed military comparisons, the additional bias of comparator selection may lead to a net underestimate of the effect.

4. SUMMARY

The six included studies compared deployed military (seeking treatment for unexplained dyspnea) to various other groups:

  • Healthy non-military controls: Zell-Baran 2021, Zell-Baran 2022, Gutor 2021, Gutor 2022, Rose 2022, Aghanouri 2004
  • Healthy military controls: Aghanouri 2004
  • Other disease states in non-military: Rose 2022, Gutor 2022, Aghanouri 2004

We were interested in markers of inflammation as well as markers of parenchyma wall changes or scarring.

4.1 DEPLOYERS VERSUS NON-MILITARY HEALTHY CONTROLS

It should come as no surprise that formerly deployed military personnel seeking treatment would have distinct histopathological findings than healthy controls. However, the importance of this comparison is to determine in precisely which ways the lungs of treatment seeking deployers and healthy individuals differ.

In comparing deployed to healthy individuals, data from three studies (Aghanouri 2004, Gutor 2021, Zell-Baran 2021, Zell-Baran 2022) provided results for a number of different inflammation and airway thickening measures. Because multiple measures were taken from the same study, a formal meta-analysis was not appropriate. However, in order to compare the histopathological findings of deployed to healthy individuals, we created a forest plot of the standardized mean differences between groups. The results are presented below (statistically significant differences indicated with an asterisk).

Figure 1 . Comparison of Histological Differences in Deployed Military Versus Healthy Controls

As can be seen, histopathological results showed higher levels of inflammation and airway thickening across measures and across areas of the lung. In work by Gutor and colleagues (using both cluster analysis [2021] and principle components analysis [2022], the key features that differentiate deployed military from healthy individuals were:

  • Evidence of inflammation (e.g., presence of higher levels of CD8 [possibly indicating chronic inflammatory lung diseases] and CD4 cells [possibly indicating an active immune response]),
  • Wall thickness in distal small airways (indicating inflammation and possible remodeling),
  • Reduced density of capillaries in the interalveolar septa (the walls that separate alveoli and vital for facilitating gas exchange),
  • Pathology of the pleura (the membrane that covers the surface of the lungs and lines the inner chest wall).

Rose and colleagues (2022) report a similar pattern of increased evidence of inflammation and fibrosis. Comparing n=65 military patients who were deployed between 2001 and 2007 to healthy controls, they find:

  • Substantially higher rates of lymphocytic inflammation in the bronchioles (90.8% v 10%), the interstitial airspace (41.5% v 0%), and pleura (89.2% v 10%).
  • Substantially higher rates of peribronchiolar metaplasia (fibrosis near the terminal bronchioles an alveolar ducts: 67.7% versus 0%), smooth muscle hypertrophy of the bronchioles (69.2% v 0%), and moderate/severe emphysema (46.2% versus 0%).

Similarly, Agranouri 2004, reported a statistically significant increase in the expression of transforming growth factor TGF-β1 in bronchoalveolar lavage (BAL) of deployers compared to both healthy controls (MD p<0.003). The presence of TGF-β1 is notable since research has shown overexpression of TGF-β1 in macrophages, mesenchymal, and mesoendothelial cells in pulmonary fibrosis. This indicates that, at least following exposure to sulfur mustard gas, fibrogenic factors are activated during inflammation which can then progress to bronchiolitis and other chronic lung diseases.

Taken together there is strong evidence of a pattern of histopathological features in the lungs of deployed military personnel seeking treatment compared to healthy individuals. These features center on evidence of chronic inflammation and fibrotic activity.

4.2 EXPOSED MILITARY VERSUS HEALTHY MILITARY

Only one study compared to deployed military confirmed to be exposed to airborne hazards (sulfur mustard) and military who were not exposed.8 Similar to what was reported when comparing exposed military to healthy controls, the TGF-β1 in exposed military personnel was substantially higher than non-exposed military (mean difference = 30.33, p=0.001). This translates to a standardized effect of d=1.37, a large effect.

Aghanouri et al 2004 do not indicate whether the unexposed military personnel were deployed, only that they were confirmed to have not been exposed. Additionally, the focus was specifically on mustard sulfur exposure, unlike the US context where there is no specific airborne hazard specified. While indirect, this study provides the closest comparison between deployed versus non-deployed military in the US context.

4.3 DEPLOYED MILITARY VERSUS OTHER DISEASE STATES

While it is clear that the histopathological profile of deployed soldiers seeking care is different than healthy individuals, are there any differences between this profile and other disease states (like COPD or smoking-related respiratory bronchiolitis)? In other words, are deployed soldiers seeking treatment displaying a garden variety respiratory disorder, or is there something unique about their disease?

Gutor and colleagues (2022) compare deployed soldiers to patients with sporadic constrictive bronchiolitis and to patients with COPD (stages I-II and III-IV). They find that while deployers are similar to patients with CoPD in terms some characteristics, they were different from patients with COPD in others.

Table 1 . Comparison of Histopathological Features of Deployers v COPD Patients

When the genes that differentiate deployed soldiers from both normal controls and patients with COPD are mapped onto biochemical pathways, they point to a profile characterized by persistent inflammation (chronic T-cell activation) leading to increased smooth muscle thickness and collagen content compared to both healthy individuals and those with COPD.

Rose and colleagues (2022) compare similar histopathological measures of deployed soldiers seeking treatment to civilian patients with other diseases ( Figure 2 ). Note that while deployers are similar to individuals with hypersensitivity pneumonitis and obliterative bronchiolitis ( Figure 2 (c) green boxes) in terms of bronchiolar and pleural inflammation, deployers have a much higher rate of smooth muscle hypertrophy (airway thickening) than either of these respiratory diseases ( Figure 2 (a)). Notably, the patterns in deployers are also different than individuals with smoking-related bronchiolitis, both in terms of higher inflammation in both the bronchioles and pleura as well as much lower rates of respiratory bronchiolitis ( Figure 2 (c)).

Figure 2 . Comparison of Histopathological Features of Deployers v Other Respiratory Disease Patients

Finally, in Aghanouri and colleagues (2004), while exposed military personnel had significantly higher rates of TGF-β1 (indicating fibrosis) than either unexposed military or healthy civilians, there was no difference in TGF-β1 levels when compared to civilians diagnosed with idiopathic pulmonary fibrosis.

Summary: Taken together, the available research indicates that (a) the lungs of deployed soldiers seeking treatment are histologically distinct from healthy individuals and characterized by chronic inflammation and small airway thickening (fibrosis), (b) comparisons between exposed and non-exposed non-US soldiers indicate large differences in terms of the presence of fibrogenic markers, and (c) while this pattern shares some features with other respiratory diseases, deployment-related distal lung disease (DDLD) 1,2 appears to be distinct from other respiratory diseases. In particular, DDLD appears to have a different profile from smoking-related bronchiolitis.

5 UNCERTAINTY

Deployers versus Non-Deployers: Unfortunately, no research directly comparing the histopathological profiles of deployers versus non deployers was identified. Thus, we can only reason that if the lungs of non-deployed military share more in common with healthy civilians, then many of the patterns we see above should carry over to the comparison between deployers and non-deployers. If, however, there are military exposures that are common to both deployers and non-deployers but distinct from civilians, then the differences presented above may be somewhat attenuated. Indeed, there is evidence 14 that even non-deployed military are at risk for many different airborne hazards, though these are at a lower rate than deployed military personnel ( Figure 3 ).

Figure 3 . Self-Reported Airborne Hazard Exposure: Deployed v Non-Deployed

As noted above, the Aghanouri et al 2004 comparison provides the closest analogue to the US comparison between deployers and non deployers. Because the Aghanouri 2004 focuses on a specific exposure and compares to military confirmed to be unexposed, we would expect the measured differences to be more precise than in the US context where the difference in type and severity of exposures among deployers cannot be confirmed (thus biasing toward the null) and where non- deployed military also report increased occupational risk of many airborne exposures (which would also bias toward the null). Thus, it is reasonable to speculate that the histomorphological differences between deployed and non-deployed in the US context would more approximate the large difference seen in the Iraqi context.

Mechanistic Pathways for Different Interstitial Diseases: Though there is mechanistic evidence to indicate a causal relation between airborne hazards present in the SWATO and the combined inflammatory and fibrotic pathologies, it is unclear how differences in exposures between deployed and non-deployed would result in disproportions in risk of some specific diseases (e.g., organizing pneumonia).

6 STRENGTH OF EVIDENCE

Following recent guidance regarding the baseline evidence level for study designs of this type used in complex environmental exposure
research 15,16 , studies start at a moderate confidence level. Then, following the GRADE approach of rating up or rating down based on
characteristics of the body of research, we reach a final strength of evidence rating for each of the three related PECOT questions.

7 ALTERNATIVE EXPLANATIONS

The most plausible alternative explanations for differences in the histomorphology between deployed versus non-deployed military are:

  • Differences in prevalence of smoking (there is evidence that deployers are more likely to smoke than non-deployers)
  • Differences in occupational or environmental exposures following separation from military service.

Regarding smoking, there is only one study 10 where differences between deployers and the comparison arm of major concern. For the others, smoking is either statistically adjusted for 12,13 , only non-smokers are examined 8 , or confounding was reduced via design (i.e., constructing all smoker comparison groups 9 or comparison groups with different proportions of smokers 11 ). Given that there was no reduction in effect in the studies that adjusted for smoking, the identified differences in effect are very unlikely to be due to smoking. Additionally, in the study that compared deployers to a group of smokers identified as having smoking-related bronchiolitis, the histopathological patterns for inflammation and thickening were different. This seems to indicate that, at the histological level, lung damage due to smoking has a different feature profile than damage due to airborne exposures in the SWATO.

Regarding differences in occupational and environmental exposures, there is no available evidence to suggest that, following service, formerly deployed versus non-deployed veterans have differential exposure to pathogens shown to be associated with ILD.

8 OVERVIEW TABLE

* CB: constrictive bronchiolitis; cHP: chronic hypersensitivity pneumonitis; RB: respiratory bronchiolitis: OB: post-transplant and autoimmune obliterative bronchiolitis
‡ In the Aghanouri 2004, the authors excluded smokers from the case group (exposed military). For the control groups, no explicit  exclusion criteria were listed, though the authors state “All exclusion criteria were observed for case and control groups and for final tests”, leading to the inference that there were no smokers included for any of the comparison arms.
§ In the Aghanouri 2004, it is not entirely clear that the cases (exposed to sulfur mustard gas) were only military. Authors state that both military and civilian personnel were exposed during attacks on Iran during the Iraq-Iran war (1984 –1988). However, the case sample is entirely male while the civilian and IPF arms contain both males and females. This leads us to believe that the exposed group was largely, if not entirely, military personnel.

9 RISK OF BIAS

The risk of bias assessment for the included articles is presented below for each type of study design (see Figure 4 and Figure 5 ).

9.1 COHORT DESIGN

Figure 4 . Cohort Design Risk of Bias

9.2 CASE-CONTROL DESIGN

Figure 5 . Case-Control Design Risk of Bias

While there were few concerns with the cohort studies, the primary concern with one of the case- control studies (Gutor et al 2021) was with confounding due to smoking. The case arm contained 33% smokers while controls had no smokers. A subanalysis comparing non smokers in both arms could have been done but was not. This problem was avoided in Gutor 2022 where a sample of smokers was collected as one of the comparison arms, allowing for comparisons to both smoker and non-smoker case arms.

10 METHODOLOGICAL NOTE

Meta-analytic procedures were conducted using the meta package in R.

11 REFERENCES

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2. Krefft SD, Wolff J, Zell-Baran L, et al. Respiratory Diseases in Post-9/11 Military Personnel Following Southwest Asia Deployment. J Occup Environ Med. May 2020;62(5):337-343. doi:10.1097/JOM.0000000000001817

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