Fig. 4

Cancer cell identification and lineage tracing with mitochondrial mutations. A Heatmap showing the mitochondrial mutations in cells of patient #1 that cultured in chemical-defined medium. Red represents mutant site, blue represents wild type, and gray represents site with sequence reads lower than 9. L-Org, long-term cultured organoids; S-Org, short-term cultured organoids. Chem, chemical-defined medium. Cond, conditioned medium. B Heatmap showing the mitochondrial mutations in tumor organoid cells of patient #1 that cultured in chemical-defined medium. Cells of three single organoid spheres and a mixture of several organoids were showed. Bar plot showing the corresponding expression levels of intestinal cell type markers. OLFM4, pluripotent cell marker; MKI67, cell proliferation marker; LYZ, Paneth cell marker; MUC2, goblet cell marker; CA2, enterocyte marker. C Heatmap showing the mitochondrial mutations in long-term-cultured normal organoid cells of patient #1 that cultured in chemical-defined medium. Bar plot showing the corresponding expression levels of intestinal cell type markers. D Heatmap showing the mitochondrial mutations in cells of patient #6 that cultured in both chemical-defined medium and conditioned medium. Red represents mutant site, blue represents wild type, and gray represents site with reads lower than 9. Chem, chemical-defined medium; Cond, conditioned medium. E Scatterplot showing the expression pattern of the epithelial cell marker EPCAM and classical mesenchymal cell marker VIM in each individual epithelial cell. The ratio of cells that co-expressed EPCAM and VIM is shown at the bottom right. F Box plot showing the expression levels of EPCAM, SPARC, and VIM in VIM-positive epithelial cells. The color of bar represents different cell types and the red background highlight cells in tumor. The black lines of box plot represent median values, the box limits indicate upper and lower quartiles, and the whiskers correspond to 1.5× the interquartile range. G Immunofluorescent staining of VIM and EPCAM on tumor- and normal-derived organoids. Three cells are shown at higher magnification at the lower right. Cell #a expressed only EPCAM, while Cell #b co-expressed VIM and EPCAM. H The schematic diagram summarizes the mitochondrial mutations and cell type changes of tumor cells in short-term and long-term cultures in two cultural media