Immune checkpoint TIM-3 defines hyperactivated NK cells and predicts fatal outcome in severe fever with thrombocytopenia syndrome

Background Severe fever with thrombocytopenia syndrome (SFTS) is a viral hemorrhagic fever with high mortality, primarily driven by immune dysregulation. This study aimed to characterize the expression profile of TIM-3 expression on peripheral NK cells in SFTS, explore the role of TIM-3 ⁺ NK cells in disease progression, and evaluate soluble TIM-3 (sTIM-3) and Galectin-9 (sGalectin-9) as prognostic biomarkers. Methods Public single-cell RNA sequencing (scRNA-seq) datasets were analyzed using weighted gene co-expression network analysis (WGCNA). Peripheral blood samples from 21 SFTS patients and 14 healthy donors collected at the First Affiliated Hospital, Zhejiang University School of Medicine, were analyzed by flow cytometry, functional assays, and serological assessments. NK cell cytotoxicity was assessed by granzyme B, perforin, IFN-γ, and TNF-α production. Serum sTIM-3 and sGalectin-9 were measured by Cytometric Bead Array. Results TIM-3 ⁺ NK cells were significantly increased in SFTS patients, especially in fatal cases. WGCNA identified HAVCR2 (encoding TIM-3) as a mortality-associated hub gene. TIM-3 ⁺ NK cells exhibited enhanced granzyme B and perforin expression, while TIM-3 blockade significantly reduced IFN-γ and TNF-α production. Elevated sTIM-3 and sGalectin-9 levels were correlated with fatal outcomes. Conclusions Activated TIM-3 ⁺ NK cells were associated with fatal outcomes in SFTS. TIM-3 ⁺ NK cell proportion, sTIM-3, and sGalectin-9 may serve as novel prognostic biomarkers and therapeutic targets.