Fig. 3

Dexamethasone increases retinal stem cell sphere size, number, and self-renewal but inhibits growth and insulin expression of pancreatic multipotent progenitor spheres. a–c Quantification of sphere diameter after a 7-day free-floating clonal sphere assay with actin-GFP mouse-derived secondary RSPCs in 0.1% DMSO vehicle or 1 μM Dex. a The number of cell colonies less than 80 μm in diameter was not significantly different between 0.1% DMSO vehicle control and 1 μM Dex treatment. N = 8 per condition. b Cell colonies greater than 80 μm in diameter increased in number by 1.63-fold with Dex treatment. (t test t (₁₄) = − 5, p < .001; N = 8 per condition). c RSC sphere colonies 80 μm or above in diameter demonstrated an overall increase in size with Dex treatment (t test t (₁₄) = 2.56, p < .05; N = 8 per condition). *Significantly different from 0.1% DMSO control. d, e Quantification of the number of tertiary RSC sphere colonies grown in drug-free media after prior exposure of secondary cells to the indicated compounds at 1 μM or 10 μM during a 7-day free-floating clonal sphere assay. Exposure to Dex and Pred increased the number of RSC spheres after passaging by 2.2-fold and 2-fold, respectively, at 1 μM (d; one-way ANOVA F (_{2, 15}) = 4.02, p = 0.04; Fisher LSD post hoc test, *p < 0.05; N = 6 per group), and increased RSC sphere number by 5.2-fold and 4-fold, respectively, at 10 μM (e; one-way ANOVA F (_{2, 6}) = 9.6, p = 0.014; Fisher LSD post hoc test, *p < 0.05; N = 3 per group). *Significantly different from 0.1% DMSO control. f–h Quantification of adult PMP spheres after a 7-day free-floating clonal sphere assay. f The total number of spheres ≥ 30 μm was significantly reduced by all concentrations of Dex tested (one-way ANOVA F (_{4, 10}) = 4.76, p = 0.02; post hoc test, Holm-Sidak post hoc test, *p < 0.05; N = 3 experiments). g The number of spheres 30–49 μm in diameter was not influenced by Dex treatment at any concentration tested. N = 3 experiments. h The number of spheres ≥ 50 μm was significantly reduced by all concentrations of Dex tested (one-way ANOVA F (_{4, 10}) = 16.12, p < 0.001; Holm-Sidak post hoc test, *p < 0.05; N = 3 experiments). *Significantly different from indicated conditions. i–l The intensity of MIP-GFP expression in PMP spheres. i Quantification of the intensity of MIP-GFP expression in PMP spheres (one-way ANOVA F (_{2, 6}) = 18.35, p = 0.0028; Holm-Sidak post hoc test, *p < 0.05; N = 3 replicates per condition). j–l Representative confocal projection images of MIP-GFP expression in PMP spheres. Data are mean ± SEM