Fig. 3

Fibroblast subsets in HPSCC tumor and adjacent normal tissues. a UMAP plot of fibroblast cells colored by cell and sample type. b Heatmap showing signature DEGs among four fibroblast subsets. c Violin plots showing marker gene expression in fibroblast subsets. d Bubble heatmap showing marker gene expression in Elastic Fbs. Dot size indicates fraction of expressing cells, colored according to expression normalized based on z-scores. e Representative images showing mIHC staining of panCK, FAP, ⍶-SMA, and PLA2G2A in HPSCC samples, in individual and merged channels. Scale bar represents 50 μm. f Absolute infiltration proportion of mCAFs comparing normal (n = 43) and tumor tissues (n = 43) in the TCGA-HNSC cohort. g Kaplan–Meier curve of the OS in the TCGA-HNSC cohort stratified by the optimal cut-off for FAP expression and mCAF infiltration. h Representative images of the IHC staining of HPSCC tumor samples with a high and low FAP expression. i Comparing OS (Kaplan–Meier curves) among patients with HPSCC who either lowly or highly express FAP. j FaDu or SNU1076 cells were incubated for 48 h with normal and CM of NFs and CAFs; E-cadherin, N-cadherin, vimentin, Twist, and GADPH expression was evaluated using immunoblotting. Cropped blots are used here and the full-length gel images are available in Additional file 3 (Fig. S7). k FaDu or SNU1076 cell invasion in CM relative to complete growth medium measured after 48 h. Photographs are representative of randomly chosen fields. l Representative images showing wound healing of FaDu or SNU1076 cells in CM of NFs and CAFs relative to complete growth medium, at 0, 12, 24, 48, and 72 h after wound infliction. Significance in (g) and (i) was assessed with two-sided log-rank tests. In (j–l), data are shown as mean ± SEM, with n = 3 paracancerous tissues and n = 4 tumor tissue columns. Differences were determined using unpaired t-tests (*P < 0.05; **P < 0.01; ***P < 0.001)