Epigenetic Memory and Phenotype Change Observed in Mouse Skeletal Muscle 30 Days after Exertional Heat Stroke
Authors: Murray Kevin , Sheikh Laila , Laitano Orlando , Iwaniec John , Garcia Christian , Robinson Gerard , Hammamieh Rasha , Campbell Ross , Yang Ruoting , Clanton Thomas 
Published: June 2021

Exposure to exercise and extreme environments, such as exertional heat stroke (EHS), can induce a long‐term epigenetic memory of stress that is stored as altered DNA‐methylation and/or histone posttranslational modifications. These changes can ultimately affect gene and protein expression as well as cellular function, which may lead to greater disease susceptibility. Previously, we have observed that a single bout of EHS can drastically alter the DNA methylome in monocytes lasting up to 30 days; however, whether these epigenetic changes are reflected in skeletal muscle has not been characterized.PURPOSETo determine whether a single exposure to EHS, in a preclinical mouse model, induces a prolonged epigenetic memory in skeletal muscle identified by stable alterations in DNA methylation.METHODSMice were either subjected to a standardized EHS protocol using a forced running wheel (environmental temp: 37.5°C) or a matched exercise control trial (22.5°C). The EHS mice achieved peak core temps of ~42.2°C, accompanied by transient loss of consciousness. Mice were sacrificed after 30 days of recovery. The gastrocnemius was excised, DNA was extracted and tested using bisulfite DNA sequencing (N=8/group). Additionally, intact solei muscles were excised (N=12/group) and tested for Ca2+ dysregulation in response to caffeine.RESULTSAlterations in DNA methylation were found to be 3.5x more frequent in gastrocnemius muscle compared to monocytes; some of these changes resided within gene regulatory regions, such as the 3′UTR and 5′UTR regions. Changes to DNA methylation were observed to take place at many genes of interest, including RyR1&2, Cacna1(s&c), Atp2a2, Pde4d, Casq2, Stim1, Orai1, Saraf, Nos1, Nox4, Tnnc1, and various calmodulin genes. Caffeine responsiveness was tested in ex vivo solei muscle from 0.25 up to 4.0mM. Most EHS mice did not exhibit contracture throughout the protocol compared to control (EHS: 10/12; control: 2/12; p=0.003). A follow up group (N=4) expanding the caffeine test beyond 4.0mM was performed (8.0mM, 16, & 32). We observed that all EHS mice elicited contracture at 8.0mM, while control had an average of 3.75mM.CONCLUSIONSThe data demonstrate that EHS induces robust alterations in the epigenome of skeletal muscle indicative of a memory up to 30 days. Ex vivo caffeine solei muscle tests determine that skeletal muscle becomes less sensitive to caffeine after exposure to EHS. DNA epigenetic alterations extend to regions implicated in gene regulation. Whether the observed epigenetic alterations function as a maladaptive or highly calculated adaptive response to EHS is still unknown.Support or Funding InformationBK and Betty Stevens Endowment and DOD Grant W81XWH‐15‐2‐0038 The opinions or assertions contained herein are the private views of the author(s) and are not to be construed as official or reflecting the views of the Army or the DoD.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.