Fig. 5: Disease modelling of the HNF4A/PMM2 genetic condition with gastric MRAs.
a, Brightfield images of PMM2 fundus, body and antrum organoids compared with healthy control (HC) body organoids. Scale bars, 100 μm. b, Full-grown organoid diameters were measured in three HCs and a minimum of two PMM2 organoid lines. Relative fold changes are shown compared with the average for PMM2 antrum. The data points represent single organoids and the means are shown as a bar. PMM2H represents organoids from hyperplastic polyp. Statistical significance was determined by t-test analysis for the fundus and body and one-way ANOVA for the antrum. c, Ki-67+ percentages in whole-mount-stained HC organoids (n = 3) versus PMM2 organoids (n = 4). The data points represent the single-patient average and the error bars represent means ± s.d. Statistical significance was determined by one-way ANOVA. No significant differences were found. d, Ki-67+ percentages in whole-mount-stained MRA regions (n = 5 for HCs (three patients) and n = 3 for PMM2 (two patients)). The data represent means ± s.d. Statistical significance was determined by one-way ANOVA. F, fundus; B, body; and A, antrum. e, Whole-mount immunostaining of the antral region of a HC MRA and a PMM2-MRA for Ki-67 (magenta), PDX1 (yellow) and MUC5AC (white). The asterisk identifies the luminal side of the MRA. Scale bars, 50 μm. f, Imaging tiling of a haematoxylin-and-eosin-stained section of a PMM2-MRA. The red arrows indicate cell aggregates. Scale bar, 500 μm. g, Toluidine blue staining (scale bar, 200 μm) and matching TEM images of PMM2 tissue samples and TEM images of PMM2-MRAs (scale bars, 10 μm), with magnifications of mucin granules (scale bars, 2 μm; n = 1). h, Whole-mount staining of HC and PMM2 organoids for nuclei (cyan) and MUC5AC (yellow). Scale bar, 20 μm. i, MUC5AC RT-qPCR data (n = 6 for HC organoids (three patients), n = 4 for PMM2 organoids (two patients) n = 5 for HC MRAs (three patients) and n = 4 for PMM2 MRAs (two patients)). j, ATP4B RT-qPCR data (n = 6 for HC organoids (three patients), n = 4 for PMM2 organoids (two patients), n = 3 for HC MRAs (three patients) and n = 4 for PMM2-MRAs (two patients)). In i and j, the data represent means ± s.d. and statistical significance was determined by one-way ANOVA. Blue, green and red data points represent samples from the fundus, body and antrum, respectively. k, WES analysis of the ATPase β protein ratio (glycosylated/precursor) in fundus, body and antrum PMM2 organoids, MRAs and SRAs. Single data points are shown, where circles represent organoids (n = 4; two patients), rhomboids represent SRAs (n = 6; two patients) and triangles represent MRAs (n = 6; two patients). Statistical significance was determined by Kruskal–Wallis test. No significant differences were found. l, Acridine orange (AO) 650 nm/550 nm PMM2-MRA ratio quantification (fundus, body and antrum), relative to the antrum control. The treatments included histamine (100 μM), omeprazole (100 μM) and histamine + omeprazole (100 μM). The circles indicate the average of single patients and the data represent means ± s.d. (n = 3; two patients). m, Acridine orange 650 nm/550 nm ratio timelapse quantification of single cells (fundus, body and antrum) in PMM2-MRAs at four timepoints (every 20 min for 1 h). The treatment groups were as in l. The data points indicate an average of 12 cells (two patients). Statistical significance was determined by one-way ANOVA for the t3 timepoint.
