Fig. 5

The proportion of GZMA^high NK cells was increased and the proportion of exhausted T cells was decreased in the combination therapy. (A) UMAP of CD4⁺ T cells, CD8⁺ T cells, and NK cells were showed on the left, and relative proportion of these three clusters in four different treatments were showed on the right. (B) UMAP of GZMA^high NK, HLA-DRB1^high NK, XCL1^high NK, and NKT cells were showed on the left, and relative proportion of these four clusters in four different treatments were showed on the right. (C) Violin plot showing expression of GZMA, CD3D, HLA-DRB1, XCL1, NCAM1, and FCGR3A in four NK clusters. (D) Violin plot showing the highest granzyme and perforin signature score in GZMA^high NK cells compared with that in other subclusters. ^****P < 0.0001. (E) Metascape analysis showing the top representative biological pathways of three NK subclusters based on DEGs. (F) Immunofluorescence staining indicates the co-expression of GZMA, CMC1, and DAPI (nuclei) on GZMA^high NK cells in PD-1+SMI group. (G) Granzyme A levels in the co-culture of NK cells and LUSC NCI-H226 cells following different treatment. ^*P < 0.05, n = 2. (H) Chord diagram showing the ligand-receptor pair TNFSF10-TNFRSF11B involved in interactions between GZMA^high NK cells and tumor subclusters (FSTL1⁺ tumor cells, XIST⁺ tumor cells, S100A4⁺ tumor cells, NNMT⁺ tumor cells, FOSB⁺ tumor cells, and NEAT1⁺ tumor cells), and between XCL1^high NK cells and the same tumor subclusters, also the ligand-receptor FASL-FAS pair was inferred between three NK subclusters and FSTL1⁺ tumor cells, and between three NK subclusters and NEAT1⁺ tumor cells. (I) UMAP of thirteen T cell subclusters were showed on the left. Relative proportion of these T subclusters in different treatments were showed on the right. (J) Relative proportion of CXCL13⁺ CD8⁺ T_EX, ENTPD1⁺ CD8⁺ T_EX, and PDCD1⁺ CD4⁺ T_EX cells in different treatments