Extended Data Fig. 2: MSCs lose their osteogenic capacity upon aging.

(a) Comparison of the proliferation rate of young and aged MSCs, after 1,3,5, and 7 days in culture, using the CyQuANT NF Cell Proliferation Assay kit. Results of n = 3 biologically independent experiments are shown and data are presented as mean ± SEM. (b) Representative images and quantification of Colony-Forming Unit assay for young and aged MSCs, after culturing them for 10-14 days. Results of n = 6 biologically independent experiments are shown and data are presented as mean ± SEM. (c-d) Representative images (c) and quantification (d) after Oil Red-O staining of young and aged cells, 9 days after induction of adipogenesis. Results of n = 3 biologically independent experiments are shown and data are presented as mean ± SEM. Scale bars, 500um. (e-f) Representative images (e) and quantification (f) of Alizarin Red S staining of young and aged MSCs, 12 days after osteogenesis induction. Images were acquired using bright-field microscopy. Scale bars, 500um. Results of n = 3 biologically independent experiments are shown and data are presented as mean ± SEM. (g) GO-term analysis for down-regulated genes upon ageing, identified by RNA-seq. n = 3 biologically independent replicates. (h-j) Measurement of bone density and bone volume of the cortical and trabecular bone compartments from the femurs of young and old animals; Results of n = 9 biologically independent experiments of young mice and n = 8 biological replicates for old mice are shown and data are presented as mean ± SEM. Statistical significance in (a), (b), (d), (f), (i) and (j) was determined using two-sided unpaired t-Test.