Longitudinal DNA Methylation Analysis of Human Exertional Heat Illness Reveals Divergent Epigenetic Trajectories of Recovery and Pathology

Background: Exertional heat stress induces a spectrum of illnesses, from mild heat exhaustion (EHE) to life-threatening heat stroke (EHS). While an epigenetic basis for heat acclimation "memory" is proposed, the dynamic molecular changes defining the severity and progression of human exertional heat illness remain poorly understood. Study objective: The primary objective of this study was to characterize and compare the longitudinal dynamics of whole-blood DNA methylation across the clinical spectrum of exertional heat illness. Hypothesis. We hypothesized that the severity of exertional heat illness would be encoded by distinct and persistent epigenetic signatures in peripheral blood. Methods: We conducted a longitudinal analysis of whole-blood DNA methylation in a human cohort diagnosed with EHE (n=32), Exertional Heat Injury (EHI, n=16), or EHS (n=39). Methylation profiles were assessed at initial diagnosis (T0) and across four subsequent time-bins representing acute, sub-acute, and later recovery phases ( >8 days). Data: We employed differential methylation analysis, time-course clustering, and pathway enrichment to define the epigenetic landscape of each condition. Summary of Results: At diagnosis, EHS and EHI displayed thousands of differentially methylated probes (DMPs) compared to EHE, with largely distinct signatures (67% and 15% unique DMPs, respectively), indicating severity-specific initial responses. Longitudinal analysis revealed that while EHE profiles normalized rapidly, EHS and EHI exhibited dramatic and persistent methylation changes over time. Temporal clustering identified divergent regulatory programs. EHI was characterized by transient upregulation of pathways related to neuronal protection (BDNF), stress regulation (FKBP5), and adaptive immunity, followed by sustained downregulation of synaptic and T-cell signaling pathways, suggesting a shift toward a resource-conserving recovery state. In contrast, EHS showed a delayed and sustained upregulation of maladaptive pathways, including inflammasome activation (CARD8, GSDMD), proteotoxic stress (HSP90AB1), and oxidative strain (TXNRD1), reflecting a progression toward systemic immune and cellular collapse. Conclusions: Exertional heat stress engraves divergent, severity-dependent epigenetic trajectories in peripheral blood. EHI is marked by adaptive methylation programs geared toward repair and recovery, while EHS triggers a persistent, pathological signature of systemic inflammation. These findings provide a molecular framework for understanding heat illness progression, offer a rich source for potential biomarkers to predict clinical outcomes, and highlight novel therapeutic targets to mitigate the long-term consequences of severe heat injury. Funding sources- This study was funded by US Army Medical Research and Development Command (USAMRDC).Disclaimer Material has been reviewed by the Walter Reed Army Institute of Research. There is no objection to its presentation and/or publication. The opinions or assertions contained herein are the private views of the author, and are not to be construed as official, or as reflecting true views of the Department of the Army or the Department of Defense. The investigators have adhered to the policies for protection of human subjects as prescribed in AR 70–25/DoD Instruction (DoDI) 3216.02. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.