Figure 1.

Long-term intestinal culture. (a) Time-course analysis of short-term air-liquid interface culture of neonatal small intestine. Stereomicroscopy shows the progressive growth of intestinal cultures, forming cyst-like structures in the collagen gel. Arrowheads indicate growth of individual spheres. Scale bars, 1 mm. (b) Long-term stereomicroscopy of air-liquid interface cultures of neonatal small intestine in collagen gel. The same field is visualized. Scale bars, 5 mm. (c) Histological analysis of small intestinal cultures (day 10). The wall of the intestinal spheres consists of a polarized epithelial monolayer and outer lining myofibroblasts (arrowheads). Tall-columnar absorptive enterocytes are visible on H&E staining. Alcian blue (Al-B) staining shows goblet cells with secreted mucus in the lumen. Chromogranin A immunohistochemistry reveals the presence of enteroendocrine cells. Lysozyme immunohistochemistry shows Paneth cells. PCNA staining reveals active proliferation of jejunal cultures at day 217. Scale bars, 10 μm. (d) Stereomicroscopy of long-term colon culture. Scale bars, 1 mm. The same field of view is depicted in both photographs. (e) Histology of long-term colonic cultures. Differentiated colonic epithelial cells are visible on H&E staining (top; day 92, middle; day 357). PCNA staining (bottom) shows continued active proliferation of cultured colonic mucosa at day 357. (f) Electron micrographs of jejunal culture (day 14). Goblet cells (G) contain mucus granules (M) in their apical cytoplasm. The epithelium is lined by myofibroblasts (MF). The apical surface of absorptive cells (A) shows microvilli (MV) ending on the terminal web (TW). Note the well organized tight junction (arrowhead) and desmosome (arrow) at the basolateral domain anchoring adjacent epithelial cells. Enteroendocrine cells (EE) contain secretory granules (SG) in their basal cytoplasm.