SiglecF ^hi TANs directly promote tumorigenesis. (A) Pearson’s correlation of tumorigenesis-related gene signatures with Siglecf-Hi–like TAN signature in the TCGA-LIHC dataset. (B–E) Sort-purified SiglecF^– and SiglecF^hi TANs were co-cultured with RIL175 cells. Schematic diagram of the sorting and co-culture strategy (B). Limiting dilution assay: representative images of spheroids formed and stem cell frequency. Scale bar represents 50 μm (C). Clonogenic assay: representative images of colonies formed and colony counts. Scale bar represents 3.5 mm (D). Scratch wound assay: representative images of the wound at indicated time points and percentage of the wound area. Scale bar represents 200 μm (E). (F–H) RIL175 cells alongside SiglecF⁻ or SiglecF^hi TANs were subcutaneously injected into C57BL/6J mice. Schematic diagram of the adoptive transfer strategy (F). Representative images and volumes of tumors at the endpoint. Scale bar represents 1 cm (G). Representative FACs plots and percentage of Ki-67⁺ tumor cells (H). Each symbol represents TANs isolated from two to three mice (D, E, G, and H). Data are mean ± SEM (D, E, G, and H) and represent two to three independent experiments (C, E, G, and H) or are pooled from two independent experiments (D). *P < 0.05, **P < 0.01, ***P < 0.001 by Pearson’s correlation test (A), Pearson’s chi-square test with a 95% confidence interval (C), one-way ANOVA (D and E), and unpaired Student’s t test (G and H).