The molecular signaling 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, however significant gaps remain in our understanding of how inflammation and immune function change at high altitude and if these responses contribute to high-altitude pathologies, such as Acute Mountain Sickness. To address this, we used 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 three days of high-altitude acclimatization in healthy sea-level residents (N = 15; 5 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 danger associated molecular pattern (DAMP) molecule which amplifies immune responses during tissue injury. Differentially expressed genes on the first and third days of acclimatization was enriched for several inflammatory pathways including NF-kB and toll-like receptor (TLR) signaling. 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 AMS scores and pulse 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 signaling pathways and may sensitize the TLR4 signaling pathway to subsequent inflammatory stimuli.