Dear Dr. Boos,

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Academic Editor

PLOS ONE

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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2. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #1: This is an original paper that examines the relationship between certain cardiovascular risk factors including visceral fat, heart rate variability, physical functioning in patients with combat related traumatic injury or without injury or amputation and subendocardial viability ratio. They find reduced SEVR in injured amputees and other fascinating associations. Some of this seems like common sense. For example, an amputee is not going to be able to necessarily exercise in the same fashion as a non injured amputee and therefore coronary reserve is likely to be better in the non injured individual. My major criticism is that we are not provided information about other important cardiovascular risk factors here: Cholesterol, LDL, HDL, triglyceride values; information on diabetes, Hemoglobin A1C, hypertension, medicines. For the HRV measures how long was the pulse or ECG measured? How were arrhythmias handled? Data on depression/anxiety and emotional stress are not provided, but it is well known that psychological stress can contribute to cardiovascular events. What medicines were the participants on .... were any on vasodilators including things like calcium channel blockers, nitrates, PDE5 inhibitors... Viagra like drugs, since these can all cause vasodilation including of the coronary arteries. There should be some discussions related to these issues or data provided if available on medicines.

Reviewer #2: The authors present a valuable analysis using the ADVANCE cohort to investigate the mediating pathways linking combat-related traumatic injury (CRTI) to long-term reductions in myocardial blood flow reserve (MBFR), as measured by the Subendocardial Viability Ratio (SEVR). The study addresses a clinically important question regarding the long-term cardiovascular health of service personnel. The use of a robust, prospective cohort and the application of mediation analysis are strong methodological points. The manuscript is generally well-structured. However, several critical details concerning the mediation model's reporting, methodology, and interpretation need to be fully clarified and addressed before moving forward.

Major Comments:

1- The core methodology of this manuscript is mediation analysis. It is crucial to report the analysis with explicit detail. Given that the cohort was frequency-matched (by age, rank, and deployment), please detail how these matching factors were included in the mediation models (as covariates or via a more specialized approach).

2- The results must clearly state the full statistical picture of the mediation. What was the calculated Indirect Effect (IE) through the identified mediator(s)? Explicitly state whether the finding represents full mediation (CDE non-significant) or partial mediation (CDE significant). This is a required step for interpreting mediation models.

3- The legend for Figure 2 mentions an exposure-mediator interaction. If this interaction term was statistically significant, the standard interpretation of a simple indirect effect is no longer valid. The analysis must then focus on Conditional Indirect Effects (i.e., how the mediation effect changes at different levels of the moderator, which here appears to be the mediator). Please elaborate on this analysis.

4- Did the authors run separate single-mediator models for each factor, or did they perform a parallel or serial multiple mediation analysis? If separate models were run, please justify this choice and explain how the issue of Type I error (false positives) from running multiple tests was addressed.

5- Which version of SEVR was used? Please explain if LV pressure was not used in the limitations.

6- SEVR is a dynamic measure and highly sensitive, which can be influenced by the mediators themselves (e.g., fitness via 6MWD, autonomic control via RMMSD). Please confirm that the SEVR measurements were standardized (e.g., performed under the same rest/fasting conditions) and specify if blood pressure and heart rate at the time of SEVR measurement were included as time-varying covariates in the outcome model. If not, this should be discussed as a limitation.

7- The authors mentioned sex (male) in the methods once. Did the study only include male servicemen? Did you also include females? If yes, what was any sex-difference observed in mediator analysis?

Minor Comments:

1- Given that the mediators were assessed approximately eight years post-injury/deployment, and the outcome (SEVR) was measured later, there is a risk of reverse causality or confounding due to post-injury lifestyle changes. Please include a dedicated limitations section addressing the temporal relationship and the challenge of establishing true causality in this context.

2- Please confirm and state whether the analysis includes all 1018 servicemen mentioned, or if there was subsequent drop-out or missing data for the SEVR measurement. If the sample size is smaller for the final analysis, please state the N used in the Methods/Results.

Reviewer #3: Please see the full review in the attached PDF.

Summary:

This manuscript entitled “Mediating Factors in the Relationship between Combat-related Traumatic Injury and Myocardial Blood Flow Reserve: the ADVANCE Cohort Study” investigates whether specific cardiovascular risk factors mediate the relationship between combat-related traumatic injury (CRTI) and myocardial blood flow reserve, represented by the subendocardial viability ratio (SEVR), using longitudinal data from the ADVANCE cohort. The study leverages a valuable dataset of UK servicemen (n=1,018, 504 with CRTI, of whom 140 were amputees, and 514 uninjured men, frequency-matched, by age, rank and deployment) with a follow-up duration for more than a decade after deployment. From the standpoint of public health significance, innovation, and scientific contribution, this paper represents a substantive and meaningful addition to the emerging literature on the long-term cardiovascular consequences of military trauma. The manuscript is suitable for publication pending modifications and clarifications described below:

Significance and Impact

The primary strength of this work lies in addressing an understudied question with substantial clinical implications: why combat-injured veterans exhibit elevated cardiovascular risk even years after injury. While prior papers from the same ADVANCE cohort established associations between CRTI, adverse cardiometabolic profiles, and reduced SEVR, this study advances the field by investigating the underlying mechanisms. Understanding modifiable mediators such as visceral fat mass, heart-rate variability, and physical function is crucial for designing targeted interventions and informing long-term rehabilitation strategies for injured service members. Given the large population of veterans with blast injuries, limb loss, and related trauma, these insights could have high translational relevance for both defense health systems and civilian trauma rehabilitation.

In terms of the approach, this study’s main novelty lies in its rigorous application of counterfactual mediation analysis. Prior literature linking traumatic injury to cardiovascular risk has largely been retrospective, cross-sectional, or limited to mortality outcomes. By contrast, to strengthen causal inference, this manuscript uses longitudinal measurements: baseline (~8 years post-injury) for mediators and follow-up (~3 years later) for SEVR. The approach allows the authors to quantify the extent to which specific physiological domains (autonomic balance, adiposity, inflammation, physical activity, and physical function) explain the CRTI–SEVR association. The subgroup analysis focusing on traumatic amputation is particularly innovative and important, offering mechanistic hypotheses tailored to a clinical subgroup known to be at heightened cardiovascular risk.

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what does this mean? ). If published, this will include your full peer review and any attached files.

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Reviewer #1: No

Reviewer #2: Yes: Rashid Alavi

Reviewer #3: No

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Attachments

Attachment

Submitted filename: Review.pdf

Mediating Factors in the Relationship between Combat-related Traumatic Injury and Myocardial Blood Flow Reserve: the ADVANCE Cohort Study

PONE-D-25-50727R1

Dear Dr. Boos,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Niema M. Pahlevan, PhD, FAHA

Academic Editor

PLOS One

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #2: All comments have been addressed

Reviewer #3: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions??>

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #2: Yes

Reviewer #3: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #2: Yes

Reviewer #3: (No Response)

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5. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: All my comments have been addressed or discussed as limitations. No further comments. The manuscript can be published now.

Reviewer #3: (No Response)

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what does this mean? ). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy

Reviewer #2: No

Reviewer #3: No

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