Fig 2: The effect of PVT1 on GBC growth in vivo.A The efficiency of sh-PVT1 in GBC-SD cells examined by qRT-PCR. B The tumor tissues from nude mice of GBC-SD/sh-NC and GBC-SD/sh-PVT1 groups. C The tumor volumes in two groups were evaluated every week until 4 weeks. D The tumor weight from two groups. E The relative expression of PVT1 in tumor tissues from two groups detected by qRT-PCR. F The Ki67 expression in tumors from two groups determined by immunohistochemical staining. *p < 0.05, **p < 0.01, ***p < 0.001.

Fig 3: Evaluation of the mouse external auditory canal (EAC) into which EGF-expression vector was transfected. (A) Otoendoscopic views of empty or EGF-expression vector-transfected ears. The area within the dotted lines indicates the region of protuberant tumor formation in the EAC in which the EGF-expression vector was transfected. (B) Morphological and immunohistochemical analysis of empty or EGF-expression vector-transfected ears. Hematoxylin and eosin (H&E) staining revealed a thickened epithelium of the EAC into which EGF-expression vector was transfected, while a normal appearance was observed in the EAC into which empty vector was transfected. Immunohistochemical analysis revealed EGF-positive (+)/EGFR+/PDPK1+ cells were detected in the lower layers of the thickened epithelium of the EGF group. Ki67+ cells were also detected in the EGF+/EGFR+/PDPK1+ layers in the section of the EGF group. Negative control was obtained by normal mouse (Ms) IgG and normal rabbit (Rb) IgG instead of first antibodies. (C) Labeling indexes (LIs) of EGF, EGFR, PDPK1, and Ki67 in the epithelium of the EAC in which empty, or EGF-expression vector was transfected (n = 4, each). Asterisks: tympanic membrane. Dashed lines: basement membrane. Arrows: positive cells. Scale bar: 20 µm. Error bars: 95% confidence interval. ***p <.001, ****p <.0001

Fig 4: Loss of p19Arf enhances fibroblast proliferation and tumor cell proliferation during leucine deprivation (LD). (A) Quantification of trypan blue negative WT or p19Arf-/- lung fibroblasts on the days indicated after normal media (left) or LD media (right). Fold change is relative to day 0 cell counts. (B) Representative images of EdU uptake (16-h pulse) by WT or p19Arf-/- lung fibroblasts after LD for the indicated days. Scale bars: 50 µm. (C) Representative images of EdU uptake by WT or p19Arf-/- lung fibroblasts during LD plus mitomycin C (Mito C). Fold change is relative to day 0. Scale bars: 50 µm. (D) Quantification of Annexin V+ WT or p19Arf-/-fibroblasts in the presence of complete or LD media. N=3. (E) Representative images of dissociated cells from tumoroids stained for Ki67 Quantification is shown on the right. Scale bars: 50 µm. N=5; *, P<0.05; ***, P<0.001; ns, not significant.

Fig 5: iHUCOs retain the colitic phenotype upon omental transplantation.a Schematic representation of the omental transplantation study. iHN and iHUC organoids were encapsulated in a TS-HA hydrogel (see “Methods” section), and each bead was transplanted into the omentum of NSG mice for 90 days. b Representative H&E-stained beads collected at 90 days, confirming the formation of colon-like crypts in iHN and iHUC organoids while retaining the phenotypes: simple columnar epithelia in normal crypts and disorganized epithelium in UC. c Summarized percentages for types of epithelial structure in normal and UC epithelium. The median value is indicated in the center of the box plot. The whiskers above and below the plot represent 1-standard deviation (SD) above and below the mean of the data, respectively. d Representative IHC for Ki67 in normal and UC organoid-derived crypts confirming the greater rate of proliferation in UC. e Percentage distribution of cells expressing Ki67 in normal and UC epithelium. f, g Representative AB-PAS staining in normal and UC organoid-derived crypts, highlighting a lack of acid mucin and significantly lower number of goblet cells in UC. h Representative IHC for chromogranin A in iHNO and iHUCO colon-like epithelium, indicating a significantly higher number of EECs in UC. i Violin plot of cell percentages positive for chromogranin A. j, l Representative IHC for ß-catenin and E-cadherin showing greater protein expression in normal vs. UC colon-like epithelium. k, m Violin plots of cells positive for ß-catenin and E-cadherin according to cellular compartment: membrane extended to cytoplasm and nucleus (Cyt + Nuc) or plasma membrane only (Mem). n, o Representative IHC for RhoA revealing higher expression in UC vs. normal organoid-derived colon, with quantification of the percentage of cells positive for RhoA. Scale bar, 40 µm. n = 3 each. For all comparisons, the unpaired, non-parametric two-sided Mann–Whitney U test was used. Source data are provided as a Source Data file.