Sex-Specific Transcriptome Responses in Circulating Immune Cells During Exercise-Heat Stress and Acclimation
Authors: Bowie Jacob , Oh Soohyun , Szymanski Michael , Struder Jeb , Filep Erica , Morrisey-Basler Margaret , Thornton Staci 
Published: May 2024

Objective: To examine the sex-based differences in heat stress and acclimation we used transcriptomic analysis to test the hypotheses that 1) gene expression varies by biological sex before and after acute heat exposure and 2) circulating immune cell transcriptome responses differ between naïve and acclimated state. Methods: Participants (3F/3M, VO2max >40/45 ml×kg×min−1) completed two heat tolerance tests (HTT), termed HTT1 (120min walking at 1.39 m/s at 2% grade) and HTT2 (~70% vVO2max until rectal temperature (Trec) reached 39.5°C) before and after a 5-day heat acclimation (HA1-5, daily 60min run/jog at Trec 38.5-39.5°C), in a hot environment (40°C, 40%RH). Blood was collected in PAXgene blood RNA tubes PRE and POST HTT1, HTT2, HA1, and HA5 with a recovery (REC) timepoint ~30min after HTT2. Paired end sequencing of ribodepleted total RNA from 84 samples (14 timepoints) was performed on a NovaSeq 6000 (Illumina, Inc., San Diego, CA). All samples passed quality checks with Fastp read trimming retaining 98.2% of reads with a GC content of 49.1%. HISAT2 indexed and mapped an average of 75.9M reads to GrCh38.105. Expression was quantified with htseq-counts, excluding genes with <3 mapped fragments per sample resulting in 26,671 genes. Principal component analysis indicated 36% of variation could be attributed to sex. Differential expression was assessed using DESeq2's generalized linear model, applying a Benjamini-Hochberg correction with an FDR of 0.1 and an adjusted p-value threshold of 0.1. DE genes were analyzed for functional annotation and pathways. Results: In total, we detected 8611 unique DE genes composed of protein coding (6241), lncRNA (1498), and 636 pseudogenes. Males and Females exhibited significant differences in gene expression at baseline, in naïve state (PREHA) with >1555 genes showing sex differences (FDR<0.1). Reactome pathways differing between M and F included cytokine signaling, response, and regulation, cell senescence, cell growth and death, cell cytoskeleton and extracellular structure and innate immunity processes (p<0.05). Interestingly, heat acclimation increased the responsiveness of the transcriptome while reducing the overall magnitude of the response. PREHA HTT2 had 86 DE genes at POST rising to 3170 at REC (vs. PRE, FDR<0.1). While POSTHA HTT2 had 602 DE genes at POST rising to 1236 at REC (vs. PRE, FDR<0.1). GO terms protein serine kinase activity, GTPase binding, and GDP binding were present at PREHA, but absent POSTHA. Conclusion: Acute exercise-heat stress induces marked changes in circulating immune cell transcriptional landscape, while acclimation reduces this response to the same challenge in an adapted state. Sex differences exist and are the subject of ongoing analyses. Funding: DoD BA200299. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.