Epigenetic silencing of Lgr5 induces senescence of intestinal epithelial organoids during the process of aging

Senescence-related changes in intestinal epithelial organoids derived from aged mice. a Representative images of a single stem cell expanding into organoids derived from intestinal epithelia of young (4 weeks) and aged (54 weeks) mice. Scale bars: 250？μm. b Success rate of organoid establishment from young mice (4–27 weeks, n？=？10) and aged mice (54–130 weeks, n？=？15). We defined the establishment of organoids as successful if it was possible to culture them over 5 passages. *p？<？0.05. c TUNEL assay for detection of apoptosis. Apoptotic cell death was observed in intestinal epithelial organoids after trypsin treatment without the Rho-kinase inhibitor Y-27632. Scale bars: 100？μm. d Cell numbers in intestinal epithelial organoids derived from young (4 weeks) and aged (54 weeks) mice during the culture course. Results of 5 experiments were plotted as mean？±？SD. ***p？<？0.001. e Relative expression levels of Cdkn1a (p21) and Cdkn2a (p16) and cell cycle analysis of intestinal epithelial organoids established from young (4 weeks) and aged (54 weeks) mice. Results of 5 experiments were plotted as mean？±？SD. *p？<？0.05, **p？<？0.01. f The numbers of intestinal crypts in young (10 weeks) and aged (120 weeks) mice. HE staining of intestinal crypts in young and aged mice are shown. Scale bars: 100？μm. The number of intestinal crypts (/mm) was significantly decreased in aged mice in comparison to young mice. Results of 5 experiments were plotted as mean？±？SD. *p？<？0.0