Extended Data Fig. 3: Molecular and functional characterization of GINS organoids derived from multiple donors.
From: Stomach-derived human insulin-secreting organoids restore glucose homeostasis
a, Schematic diagram and representative images of cells at key stages in GINS organoid formation. Ngn3ER-hGSCs: human gastric stem cells that incorporated a Ngn3 and estrogen receptor (ER) fusion gene (Ngn3ER); 4-OH-TAM: 4-OH Tamoxifen; Lenti-CMV-PM, lentiviral integration of a polycistronic Pdx1-Mafa co-expression cassette. b, Representative immunofluorescent staining of corpus GINS organoids derived from three different donors, and co-localization of INS and CPPT in GINS cells. a, b, Repeated independently 3 times with similar results. c, To assess CPPT+ mono-hormonal cells, a cocktail of GCG, SST and GHRL antibodies were stained together with CPPT in day-21 GINS organoids. Right panel shows immunofluorescent staining of CPPT (red) and a combination of GCG, GHRL and SST (green). left panel shows quantification of mono-hormonal CPPT+ cells (n = 10 organoids from donor #6). d, Relative expression of endocrine hormone genes including INS, GCG, SST, and GHRL in day-18 GINS organoids derived from four different donors in comparison with human islets. n = 4 (donor#6) or 3 (donor#7, #9, #10, or islet donors) separate batches of samples for each donor. e, Relative expression of key β-cell markers in GINS organoids derived from different donors in comparison with human islets. n = 4 (donor#6) or 3 (donor#7, #9, #10, or islet donors) separate batches of samples for each donor. f, Glucose-stimulated insulin secretion of GINS organoids at different time points (n = 3 independent samples from donor #6 for each batch of differentiation) or donor #9 (n = 3 independent samples, day-18). g, Insulin secretion of day-18 GINS organoids from donor #6 incubated with the indicated concentrations of glucose with or without 10 nM glibenclamide (Glib) or 0.5 mM diazoxide (Dzx) as indicated (n = 4 independent samples). c-g, Data presented as mean ± s.d.; one-way ANOVA (d, e, g) or repeated-measures two-way ANOVA with Holm–Sidak’s multiple comparisons test for different time points (f) or one-tailed paired t-test for donor#9 (f).
