(A) Schematic representation of BCCs cultured with MSCs to obtain hybrid-enriched population. (B) Representative flow cytometry live imaging stream pictures showing a Ki67^lo hybrid cell. GFP-labeled MDA-MB-231 (GFP-231) BCCs were cultured with DsRed-labeled MSCs (DsRed-MSCs) for 72 hours and stained with Ki67 antibody. Ki67^lo cells were sorted and fixed, and nuclei were stained with DAPI. An overlay of all fluorescence channels shows a multinucleated GFP⁺/DsRed⁺ hybrid cell. Cell phase image (bright-field [BF]) is included to display cell morphology. (C) Cell-cycle analysis by flow cytometry in the coculture of GFP-231 BCCs with DsRed-MSCs after 72 hours showing the emergence of a polyploid population in GFP⁺/DsRed⁺ (hybrid) compared with GFP⁺ cells. Bar graph shows the quantification of the flow cytometry analyses. (D) ELISA analyses of IL-6, CCL2, OPN, THBS1, and uPAR in supernatants of GFP-231 BCCs and MSCs diluted 1:1 (GFP-231) or in coculture (GFP-231 + DsRed-MSC). (E) Senescence-associated β-Gal (SA–β-Gal) activity analyzed by flow cytometry using DsRed-labeled MDA-MB-231 cells (DsRed-231) and MSC (unlabeled) in single cultures and cocultures for 1 week. Bar graph shows the percent of SA–β-Gal/DsRed⁺ cells. (F) Lung metastases from mice injected intracardially with GFP-231 or GFP⁺/FAP⁺ hybrid cells sorted by FACS, stained with H&E and SA–β-Gal by IHC (n = 3 per group). Bar graph shows the quantification of the SA–β-Gal in 5 different fields per condition (n = 4 per group) using ImageJ. Scale bars: 20 µm. For all panels, data are shown as individual values with mean ± SEM. In C, D, and F, 2-tailed unpaired Student’s t test was employed; in E, 1-way ANOVA with Tukey’s multiple comparison test was employed. *P < 0.05; **P < 0.005; ***P < 0.0005; ****P < 0.0001.