Figure 2. 3D co-culturing with senescent ESCs negatively affects surrounding cells. (A) Experimental scheme of co-culturing of young mCherry-labeled ESCs with unlabeled young or senescent ones in 3D condition. (B) Representative photographs of spheroids formed from a mixture of unlabeled and mCherry-labeled ESCs. Scale bars of all images are 500 μm. (C–E) Growth curves, cell size and autofluorescence of mCherry-labeled ESCs, respectively. Cells were cultured in spheroids during 4 d, trypsinized and cultured for additional 5 d up to analysis by FACS. Forward scatter (FS) reflects the average cell size. Values are M ± S.D. (N=3 for (C) and (D), N=2 for (E)). ** – p<0.01, *** – p<0.005 by Student’s t-test.