FIGURE 3.

Characterization of small intestinal and liver stem cell populations in progeroid Ercc1^{Δ /−} mice. (a) Enhanced green fluorescent protein (EGFP) immunostaining in crypts of the small intestine from 15‐week‐old mutant and wt mice. Nuclei counterstained with DAPI. Bars 10 μm. (b) Quantitation of intestinal crypt cells expressing high levels of GFP (GFP^hi) by flow cytometry of 15‐week‐old wt (n = 4) and Ercc1^{Δ /−} (n = 3) mice. (c) Quantitation of the total EGFP‐expressing (EGFP⁺) cell population in small intestinal crypts of wt (n = 4) and mutant (n = 3) mice by flow cytometry. (d) Ratio of crypt cells expressing low (EGFP^low, i.e., downstream progenitor population) versus high levels of EGFP (GFP^hi, i.e., stem cells) in the intestine of the indicated mice. Each column represents collectively the values for individual crypts for 3 mice per genotype. (e) Quantitation of the GFP^hi intestinal population that expresses Ki67 proliferation marker from immunofluorescently stained tissue of 15‐week‐old mice of the indicated genotypes (n = 3 mice per genotype). Note the increase of proliferative stem cells in progeroid Ercc1 mutant mice. (f) Immunofluorescence images of paraffin‐embedded intestinal tissue from 15‐week‐old Lgr5^EGFP Ercc1^{Δ /−} and wt mice, stained for EGFP and Ki67. Bars 10 μm. (g) Immunofluorescent staining for Sox9 expression (green) in bile duct cells of liver sections from 15‐week‐old Lgr5^EGFP Ercc1^{Δ /−} and wt mice, along with Ki67 (red). Bar 10 μm. (h) Percentage of Sox9+ bile cell population per total cells in liver ducts of 15‐week‐old Ercc1^{Δ /−} and wt mice, quantified from 4μm liver sections stained as in (g) (n = 3 mice). (i) Average number of cells per bile duct in livers of 15‐week‐old Ercc1^{Δ /−} and wt mice, quantified from 4 μm liver sections. Data represent means with SD from quantitation of cells in round bile ducts from 3 mice per genotype and at least 10 bile ducts per mouse. (j) EGFP‐expressing cells in the liver of Lgr5^EGFPErcc1^{Δ /−} (n = 3) and control (n = 2) mice. (k) EGFP + Lgr5‐marked stem cells in liver of 15‐week‐old Lgr5^EGFPErcc1^{Δ /−} and control mice stained for GFP. Bars 10 μm. Data: mean ± SEM unless otherwise specified. *p < 0.05, ****p < 0.0001