Fig. 6: Induction of InfFib and Wnt5a expression via hypoxia.
From: Hypoxia-induced Wnt5a-secreting fibroblasts promote colon cancer progression
a (Left) Force-directed graph embedding for mouse colon cancer fibroblast single-cell RNA-seq (scRNA-seq) data related to Fig. 4a. Cell type labels were from Fig. 4d. (Right) PAGA plot on force-directed graph embedding. b Force-directed graph embedding depicting diffusion pseudotime. c Force-directed graph embedding colored by CytoTRACE scores. A lower score indicates more differentiated states. d Changes of marker gene expression, CytoTRACE score, and hypoxia activity along PAGA path during pseudotime progression are shown. Marker genes for InfFib, Il1rl1 and Mmp10; BMPsFib, Syt13 and Lama1. DPT, diffusion pseudotime. e, f Pathway activities in each fibroblast subtype from mouse (e) or human (f) scRNA-seq data related to Fig. 4d or 3g, respectively, were inferred and are represented in heatmaps. g, h FFPE (g) or fresh-frozen (h) tissue sections from AOM/DSS treated mice after injection of pimonidazole were stained with indicated antibodies and DAPI. Scale bars, g 100 μm, h 20 μm. i Scatter plots showing the correlation between signature scores of hypoxia (x-axis) and (left) InfFib or (right) BMPsFib proportion estimated in Fig. 3l (y-axis) were obtained from TCGA-COAD patient dataset. A regression line was fitted using linear regression, and the shaded area represents the 95% confidence interval. The Spearman R- and P-values are shown on the top. j WEHI-YH2 cells were treated with CoCl2 for 24 h and subjected to bulk RNA-seq. Signature scores for each subtype are represented as strip plots. (n = 3; Unpaired t-test, Two-tailed). k WEHI-YH2 cells were exposed to CoCl2 or 1% O2 for indicated duration. The mRNA levels of the indicated genes are represented in bar plots. (n = 3; One-way ANOVA, Holm-Sidak’s multiple comparisons test). l WEHI-YH2 cells were exposed to 20% or 1% O2 for 24 h, and Wnt5a was precipitated from conditioned media. The precipitates (CM) and total cell lysates (Lysate) were probed with different antibodies: the former was probed for anti-Wnt5a antibody, and the latter was probed for anti-Hsp90 antibody as a loading control. m Tissue sections from control or Wnt5a cKO-derived tumors (n = 3 different tumors) were stained with anti-hypoxyprobe antibody and DAPI. The results are shown as the percentage of hypoxyprobe-positive area compared to the total area proximity to the lumen (within 75 μm from the edge) and presented as a bar plot. Scale bars, 100 μm. (Unpaired t-test, Two-tailed). ***, P < 0.001; **, P < 0.01; *, P < 0.05; ns, P > = 0.05; Source data are provided as a Source Data file.
