A104-07 CD8 T Cell Heterogeneity in Critical Illness Is Marked by the Emergence of a CXCR4-Hi IL7R-Lo Effector Population Associated With Respiratory Failure

Abstract Rationale Acute respiratory failure in critical illness arises amid profound immune dysregulation, yet whether specific immune cell states accompany impaired gas exchange remains poorly defined in humans. CD8 T cells play a critical role in host defense and tissue remodeling, but their heterogeneity has not been extensively studied in human critical illness. To address these gaps, we performed single-cell transcriptomic profiling to resolve CD8 T cell heterogeneity and assess its relationship to respiratory dysfunction. Methods We performed single-cell RNA sequencing on peripheral blood mononuclear cells from critically ill patients (n = 38) in the medical and surgical ICUs at Vanderbilt and healthy controls (n = 9). CD8 T cells were annotated, and Leiden clustering was applied to resolve subpopulations. We interrogated these subsets using proportionality assessment, differential gene expression analysis, gene set enrichment analysis (GSEA), and in silico metabolic flux modeling. Finally, features of CD8 T cell subsets were related to relevant clinical metrics. Results Unsupervised analysis resolved seven CD8 T cell subsets, including expected naive, central memory, effector/memory, EMRA (terminal effectors re-expressing CD45RA), and two innate-like populations. A discrete effector subset characterized by high CXCR4 and low IL7R expression emerged in critical illness (8.06% vs. 0.11% in healthy controls; p = 0.0004). By global gene expression this CXCR4hi IL7Rlo population was most closely related to EMRA cells. These CXCR4hi IL7Rlo cells exhibited reduced cytotoxic programs (GZMA, GZMH, NKG7) and inflammatory chemokine receptors (CX3CR1) alongside increased expression of exhaustion markers (HAVCR2, LAG3, TOX) and inhibitory genes (DUSP1, DUSP2) compared to EMRA. GSEA demonstrated marked de-enrichment of oxidative phosphorylation (NES −1.96; q 0.0001), tricarboxylic acid cycle activity (NES −1.47; q = 0.047), and glycolysis (NES −1.65; q = 0.002). Differential expression highlighted increased DDIT4 and reduced PGAM1, suggesting coordinated regulation of mTOR signaling and glycolytic flux. In silico metabolic flux modeling revealed broadly diminished predicted reaction activity across virtually all major metabolic pathways. Finally, ICU patients who exhibited high frequencies of CXCR4hi IL7Rlo effectors had more impaired alveolar gas exchange compared to those with low frequencies (PaO2/FiO2 124 vs. 219, respectively; p = 0.08). Conclusions In critical illness, we observed the emergence of a CXCR4hi IL7Rlo effector CD8 T cell population marked by impaired cytotoxicity and hypometabolic programming. This population associated with respiratory dysfunction, suggesting potential utility as a prognostic biomarker and/or therapeutic target. Overall, these data demonstrate the value of high-dimensional immune profiling for resolving disease-emergent cellular profiles relevant to critical illness. This abstract is funded by: NIGMS (R33GM144915), NHLBI (T32HL094296), NCATS (KL2TR002245), Vanderbilt Center for Immunobiology