Fig 1: Combined deletion of Men1, Atrx, and Pten in pancreatic islet cells triggers the development of pancreatic neuroendocrine tumors in mutant mice. (A) Schematic illustrating conditional mutant alleles utilized in the generation of PanNET mouse model. To assess the effects of combined Men1LoxP/LoxP, Atrx LoxP/LoxP, and Pten LoxP/LoxP (MAP) ablation on neoplastic transformation of the pancreatic neuroendocrine cells. Tamoxifen-induced recombination of mutant alleles is mediated using pancreatic neuroendocrine cells-specific Pdx1-CreER strain. (B) Experimental design to investigate tumorigenesis in the MAP model following tamoxifen-induced mutant gene recombination (three intraperitoneal injections of 1 mg tamoxifen per mouse every other day) at 8 months and endpoint. (C) Representative macroscopic pathology, HE-stained sections and immunohistochemical (IHC) staining with indicated antibodies of pancreas tissue from normal (wildtype) and MAP mouse model at 8 months and endpoint. IHC stainings for insulin and Ki67 (a marker of proliferating cells) are shown. Scale bars, 1 cm (macroscopic), 200 µm (histology), 50 µm (Ki67 IHC), representative of n = 10 mice for each experimental group. (D) Quantification of neuroendocrine area per total pancreas area in wildtype and MAP mutant mice at indicated time points reveals significant expansion of neuroendocrine compartment. P-values were calculated by one-way ANOVA with Tukey’s multiple comparisons test. (E) Analysis of Ki67 index in normal pancreatic islets and MAP mouse model tumors at 8 months and endpoint, revealing a significant increase in tumor cell proliferation. P values were calculated by one-way ANOVA with Tukey’s multiple comparisons test. (F) Survival analysis of MAP mutant mice (n = 10, median survival of 363 days; see Supplementary Fig. 3A for detailed analysis of the clinical spectrum of disease observed in MAP mutant model). (G) Immunoblotting analysis with indicated antibodies of representative protein lysates of tissue biopsies from wildtype pancreas and MAP model tumors. Vinculin is shown as a loading control. (H) Fasting blood glucose levels in MAP mutant and wildtype (WT) control mice (males and females) at 8 months of age. P values were calculated by Student’s t test.