graphs show the mean ± s. e. m. an unpaired two - sided student ’ s t - test ( k ( right and middle ), n ) or two - sided one - way anova with bonferroni post hoc multiple comparison test ( d, e, k ( left ), l, m ) was used to determine statistical significance. one control value was set to 1. source data because the pathways enriched in the mitotracker - high fibroblasts are often activated in cafs, we analyzed the dataset for skin caf marker genes. many of them were indeed overexpressed in the mitotracker - high and, to a lesser extent, in the mitotracker - low population. among them were inhba ( encoding the protumorigenic cytokine activin a ), il6 ( encoding interleukin-6 ), acta2 ( encoding α smooth muscle actin ) and col1a1 ( encoding coli, alpha 1 subunit ) ( fig. 3d ). this was verified by reverse transcription ( rt ) – qpcr in independent coculture experiments ( extended data fig. 2f ). pdgfra and the caf markers pdgfrb, s100a4 ( encoding fsp1 ), fap and cd74 ( ref. ) were also upregulated in the mitotracker - high population ( fig. 3e ). to determine whether the mitotracker - high cells ( mito - cafs ), correspond to inflammatory cafs ( icafs ), myof ibroblastic cafs ( mycafs ) or antigen - presenting cafs ( apcafs ), we compared their expression profile to those of published caf datasets. mito - cafs overexpressed genes characteristic of both icafs and mycafs ( fig. 3f – h ). a proteomic analysis using the same fibroblast populations revealed clear clustering of the groups ( extended data fig. 3a ), increased abundance of caf markers and isg - encoded proteins and activation of proinflammatory pathways in the mitotracker - high population ( fig. 3i, extended data fig. 3b – d and supplementary table 1 ). many of the observed gene expression changes were reflected by protein abundance changes ( fig. 3j ). mitotracker - high fibroblasts also exhibited functional characteristics of protumorigenic cafs, including increased proliferation and higher concentrations of intracellular adenosine triphosphate ( atp ) and mitochondrial reactive oxygen species ( ros ) ( fig. 3k ). increased proliferation and caf marker expression in sorted fibroblasts were also observed when hpfs in cocultures received mitochondria from a431 cells expressing su9 – rfp ( extended data fig. 3e – g ), further confirming the reliability of the mitotracker approach in our setting. lastly, the cm from mitotracker - high hpfs promoted cancer cell proliferation and transwell migration more efficiently than cm from mitotracker - low hpfs. this was observed for a431 cells ( fig. 3l, m ), primary skin squamous cell carcinoma ( scc ) cells and scc13 ( ref. ) and ha – ras - transformed hacat cells ( hacat - ras ) ( extended data fig. 3h – j ). furthermore, the decellularized extracellular matrix ( decm ) produced by mitotracker - high fibroblasts induced the formation of larger a431 colonies ( fig. 3n ). to specifically test the role of cancer cell mitochondria in fibroblast reprogramming, we isolated and purified mitochondria from cancer cells and transplanted them directly into hpfs using mitoception. while mitochondria directly move from the cytoplasm of the donor cells to the cytoplasm of recipient cells during tnt - mediated transfer, mitoception induces the rapid uptake of purified mitochondria through the plasma membrane, most likely through an endocyt ic pathway. the uptake using mitoception was confirmed by detection of mitotracker green fluorescence and by an increase in mtdna content in the ‘ mitocepted ’ fibroblasts ( fig. 4a, b ), which was in a similar range to that described for mitocepted endothelial cells ( 13 % ). the mitotracker staining likely overestimates the uptake because cancer cell mitochondria fuse with the mitochondria of recipient hpfs, as seen after transplantation of mitotracker green - labeled or su9 – rfp - expressing mitochondria from a431 cells into hpfs prestained with mitotracker red or expressing tom20 – gfp, respectively ( fig. 4c ). we found a substantial colocalization of the mitocepted and the endogenous mitochondria using confocal microscopy. as expected, it was more pronounced with mitotracker green because the dye labels the entire mitochondria. although an additional effect of dye leakage cannot be excluded, the findings obtained with mitotracker - labeled and, in particular, with genetically labeled mitochondria demonstrate that the mitocepted mitochondria were released into the cytoplasm. fig. 4caf reprogramming through transplantation of cancer cell mitochondria. a, representative fluorescence images of hpfs mitocepted with mitotracker green - stained a431 mitochondria ( mitocepted hpfs ) or mock treatment, counterstained with hoechst ( blue ) ( n = 3 cultures per group ). b, qpcr for the mtdna encoding trna - leu ( uur ) relative to the nucdna encoding b2m using dna from mitocepted ( mc ) or mock - treated ( ctrl ) hpfs. relative mtdna content ( based on ct values ) is indicated ( n = 3 cultures per group ). c, representative confocal image in the xyz plane showing hpfs prestained with mitotracker red and mitocepted with mitotracker green - labeled a431 mitochondria ( left ) and tom20 – gfp - expressing hpfs mitocepted with mitochondria from a431 su9 – rfp cells ( right ). yellow staining indicates mitochondrial fusion. d, percentage of ki67 mitocepted or control hpfs among all cells ( n = 3 cultures per group ). e, rt – qpcr for inhba, il6, acta2 and col1a1 relative to rpl27 using rna from mitocepted or mock - treated hpfs ( n = 3 cultures per group ). f, fluidfm experimental setup, adapted from a previous study. g, image of fluidfm - mediated injection of mitochondria into hpfs. h, percentage of ki67 fibroblasts in hpfs injected with a431 - derived mitochondria using fluidfm or mock treatment ( n = 3 cultures per group ). i, rt – qpcr for inhba using rna from hpfs subjected to mitoception with mitochondria from hacat, hacat - ras or a431 cell lines ( n = 3 cultures per cell line ). j, rt – qpcr for inhba using rna from ( 1 ) mock - treated hpfs or hpfs subjected to mitoception with mitochondria ( 2 ) from keratinocytes of a healthy individual, ( 3 ) from normal keratinocytes of a person with scc or ( 4 ) malignant cancer cells of the same person with scc. right, representative fn1 – coli immunoflu orescence stainings with quantification of staining intensity in the decm produced by hpfs after mitoception with mitochondria from the different primary donor cells ( n = 3 mitoceptions per cell type ). k, percentage of ki67 hpfs subjected to mitoception with different amounts of mitochondria isolated from a431 cells. numbers on the x axis show the ratio of donor a431 cells ( used for mitochondrial isolation ) and recipient hpfs ( n = 3 cultures per group ). graphs show the mean ± s. e. m. an unpaired two - sided student ’ s t - test ( b, d, e, h ) or two - sided one - way anova with bonferroni post hoc multiple comparison test ( i – k ) was used to determine statistical significance. scale bars, 100 μm ( a, j ), 10 μm ( c ) and 25 μm ( g ). source data a, representative fluorescence images of hpfs mitocepted with mitotracker green - stained a431 mitochondria ( mitocepted hpfs ) or mock treatment, counterstained with hoechst ( blue ) ( n = 3 cultures per group ). b, qpcr for the mtdna encoding trna - leu ( uur ) relative to the nucdna encoding b2m using dna from mitocepted ( mc ) or mock - treated ( ctrl ) hpfs. relative mtdna content ( based on ct values ) is indicated ( n = 3 cultures per group ). c, representative confocal image in the xyz plane showing hpfs prestained with mitotracker red and mitocepted with mitotracker green - labeled a431 mitochondria ( left ) and tom20 – gfp - expressing hpfs mitocepted with mitochondria from a431 su9 – rfp cells ( right ). yellow staining indicates mitochondrial fusion. d, percentage of ki67 mitocepted or control hpfs among all cells ( n = 3 cultures per group ).