AbstractThe molecular signalling pathways that regulate inflammation and the response to hypoxia share significant crosstalk and appear to play major roles in high‐altitude acclimatization and adaptation. Several studies demonstrate increases in circulating candidate inflammatory markers during acute high‐altitude exposure, but significant gaps remain in our understanding of how inflammation and immune function change at high altitude and whether these responses contribute to high‐altitude pathologies, such as acute mountain sickness. To address this, we took an unbiased transcriptomic approach, including RNA sequencing and direct digital mRNA detection with NanoString, to identify changes in the inflammatory profile of peripheral blood throughout 3 days of high‐altitude acclimatization in healthy sea‐level residents (n = 15; five women). Several inflammation‐related genes were upregulated on the first day of high‐altitude exposure, including a large increase in HMGB1 (high mobility group box 1), a damage‐associated molecular pattern (DAMP) molecule that amplifies immune responses during tissue injury. Differentially expressed genes on the first and third days of acclimatization were enriched for several inflammatory pathways, including nuclear factor‐κB and Toll‐like receptor (TLR) signalling. Indeed, both TLR4 and LY96, which encodes the lipopolysaccharide binding protein (MD‐2), were upregulated at high altitude. Finally, FASLG and SMAD7 were associated with acute mountain sickness scores and peripheral oxygen saturation levels on the first day at high altitude, suggesting a potential role of immune regulation in response to high‐altitude hypoxia. These results indicate that acute high‐altitude exposure upregulates inflammatory signalling pathways and might sensitize the TLR4 signalling pathway to subsequent inflammatory stimuli. imageKey points Inflammation plays a crucial role in the physiological response to hypoxia. High‐altitude hypoxia exposure causes alterations in the inflammatory profile that might play an adaptive or maladaptive role in acclimatization. In this study, we characterized changes in the inflammatory profile following acute high‐altitude exposure. We report upregulation of novel inflammation‐related genes in the first 3 days of high‐altitude exposure, which might play a role in immune system sensitization. These results provide insight into how hypoxia‐induced inflammation might contribute to high‐altitude pathologies and exacerbate inflammatory responses in critical illnesses associated with hypoxaemia.