Fig. 7: CD154-dependent endogenous CD19⁺TIM-1⁺ Breg are enriched in blood and tumours of patients with cutaneous squamous cell carcinoma.

a Summarised data of % CD73^-CD25⁺CD71⁺ B cells in CD19⁺ B-cell subsets within peripheral blood of healthy human donors (n = 10). Each dot is an individual response. Representative FACS plots are presented in Supplementary Fig. 9a, b. b Summarised data of % TIM-1⁺ B cells in CD19⁺ B-cell subsets within peripheral blood of healthy human donors (n = 10). Each dot is an individual response. Representative FACS plots are presented in Supplementary Fig. 9c. c Summarised data of % CD154⁺ B cells in CD19⁺ B cell subsets within peripheral blood of healthy human donors (n = 10). Each dot is an individual response. Representative FACS plots are presented in Supplementary Fig. 9d. d Representative FACS plots and summarised data of % CD4⁺TNFa⁺ expression when FACS-sorted CD73⁻CD25⁺CD71⁺ enriched, CD73⁻CD25⁻CD71⁻, TIM-1^hi and TIM-1^lo B cells were cultured with autologous CD4⁺ T cells and plate-bound anti-CD3 mAb [0.5 µg/ml] ± blocking anti-CD154 mAb [10 µg/ml] for 3 days. Gating is on FMO controls. Data from experiments performed with cells from different healthy donors (n = 5) are presented. Each dot is an individual response. e Summarised data of % CD73⁻CD25⁺CD71⁺, CD24^hiCD38^hi and CD24^hiCD27⁺ B cell subsets in whole CD19⁺ B cells in peripheral blood of healthy human donors (n = 10) and age-matched patients with squamous cell carcinoma of the skin (SCC, n = 8). Each dot is an individual response. f Summarised data of % TIM-1⁺ and CD154⁺ B cells within CD73⁻CD25⁺CD71⁺ B cell subsets in peripheral blood of healthy human donors (n = 10) and age-matched patients with SCC (n = 8). Each dot is an individual response. g Hematoxylin and eosin (H&E) stained histological section demonstrating a moderately differentiated cutaneous SCC arising on the dorsum of the right hand of an 81-year-old male (hashed line delineates tumour boundary; higher magnification image demonstrating representative histological features in top right-hand corner). h Double immunohistochemistry staining for pSTAT3 (red) and CD20 (brown) demonstrates the peritumoral accumulation of CD20⁺ B-cell clusters (arrows) within the tumour and in proximity to its invasive margin (hashed line). i Higher magnification of the annotated B-cell cluster in (h) (hashed box), demonstrating double positive cells (arrows). j On adjacent section, a single cell double stained for PAX5 (red) and TIM-1 (brown) is identified (arrow; higher magnification image of cell in top right-hand corner). Histological sections in (g–j) are of an SCC tumour from one of five patients. Histological SCC sections of remaining four patients are presented in Supplementary Fig. 10a–d. Error bars in (a–c, e, f) represent Median + interquartile range. Error bars in (d) represent Mean ± SD. One-way ANOVA with Dunn’s multiple comparisons test (a–d) and 2-tailed Mann–Whitney test (e, f) were used. Source data are provided as a Source Data file.