Fig. 2: Spatial patterns of PD-L1+ cells in mouse and human SCCs demonstrate that infiltrating leukocytes are the primary source of PD-L1 within tumors.

a Percentage of PD-L1⁺ A223 and LY2 cells after stimulation with 2 ng/ml recombinant TGFβ (orange triangles, n = 4) or PBS vehicle (black circles, n = 4) as measured by flow cytometry. b tdTomato expression (purple bars: positive, blue bars: negative) in PD-L1⁺ live cells isolated from mTmG mice bearing A223 tumors and treated with TGFβ trap control isotype control 3×/week for 1 week, as quantified by flow cytometry (n = 5 per treatment group). c Analysis of CD11b and CD11c expression of tdTomato⁺ cells from c as quantified by flow cytometry (CD11b⁺ CD11c⁻: blue bars, CD11b⁺ CD11c⁺: purple bars, CD11b⁻: orange bars, n = 5 per treatment group). d Representative tumor from multispectral imaging analysis of 23 human HNSCC tumors stained for pan-cytokeratin (cyan), CD68 (magenta), CD11c (yellow), PD-L1 (red), and DAPI (blue). Arrowheads represent example phenotypes using the same color code. e Table of total number of cells of each indicated phenotype across all samples in (d) (horizontal bars), and counts of cells that overlap one or more phenotypes (vertical bars). The central matrix displays the combinations graphically with connected dots indicating overlapping phenotypes. Unpaired two-tailed t tests were performed for (a–c), and all error bars represent the SEM. Raw unstacked values in (b) and (c) are represented by the indicated symbols.