Fig. 1.

Growth and differentiation of isolated pancreatic progenitor cells in pancreatic spheres. (A) Immunohistologic detection of Sox9⁺ cells (green), the major population of epithelia in E11.5 fetal mouse pancreas. Sox9^neg glucagon⁺ or Sox9^neg Insulin⁺ endocrine cells (white) are observed. (Scale bar, 40 μm.) (B) FACS of enzymatically dissociated E11.5 pancreas from Sox9-eGFP; Ngn3-tdT dual reporter mice. Progenitor cells are the Sox9-eGFP⁺ Ngn3-tdT^neg. (C) A schematic summarizing conditions for culturing purified pancreatic progenitors. (Scale bar, 50 μm.) (D) Immunohistology of pancreatic spheres. (Scale bar, 40 μm.) (E) Microscopic images of MIP-GFP expression in pancreatic spheres exposed to ambient (21%) or physiological (5%) oxygen. (Scale bar, 50 μm.) (F) Immunohistology of pancreatic spheres cultured in physiological oxygen. Arrows indicate MafA⁺ nuclei. (Scale bar, 25 μm.) (G) C-peptide content measured by ELISA, normalized to DNA from purified MIP-GFP⁺ cells (n = 3). (H) Insulin secretion after glucose or potassium challenge measured by ELISA (n = 6). (I) Transmission electron micrograph revealing cells with β-cell dense core vesicles (Left) and α cell vesicles (Right). Magnified images are shown in insets. Figure is a composite to create the full image. (Scale bar, 1 μm.) (J) Quantitative RT-PCR analysis of Ngn3 gene deletion by Cre recombinase-based inactivation of conditional Ngn3 alleles (n = 3). Error bars, SEM.