Growing Self-Organizing Mini-Guts from a Single Intestinal Stem Cell: Mechanism and Applications

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Science  07 Jun 2013:
Vol. 340, Issue 6137, pp. 1190-1194
DOI: 10.1126/science.1234852

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  1. Fig. 1 Histological location and biological interaction of intestinal stem cells and their niche.

    (A) Scheme of intestinal epithelial structure and stem cells. Spatial gradients of Wnt, BMP, and EGF signals are formed along the crypt axis. (B) Cartoon of the stem cell niche. Lgr5+ intestinal CBC cells intimately adhere to Paneth cells and receive signals for stem cell maintenance. (C) Three signals (EGF, Notch, and Wnt) are essential for intestinal epithelial stemness, whereas BMP negatively regulates stemness. For full Wnt activation in the intestinal epithelium, R-spondin–Lgr4/5 signal is required. Currently, the source of R-spondin is unknown.

  2. Fig. 2 Mini-gut culture system.

    (A) Lgr5+ CBC cells genetically labeled by EGFP are sorted and embedded in Matrigel. The culture medium consists of EGF, Noggin, and R-spondin. FACS, fluorescence-activated cell sorting. (B) Time course of organoid growth. A single stem cell forms a symmetric cyst structure. The symmetry is broken by bud formation. The budding structure resembles a crypt. Lgr5+ CBC cells are depicted in yellow, and Paneth cells are shown in blue. (C) Scheme showing the engraftment of intestinal oganoids. Organoids adhere to a de-epithelialized wound bed. Organoids form a flat epithelial layer, followed by crypt reconstruction. Wnt-5A+ mesenchymal cells support crypt structure formation in the damaged area. Donor organoids are depicted in red.

  3. Fig. 3 Mechanism of the self-organizing architecture of a mini-gut.

    (A) An organoid derived from Axin2-LacZ knock-in mice. Axin2-LacZ (blue) expression recapitulates Wnt activation. (B) Wnt activation induces local proliferation and EphB expression. Local cell expansion and Eph-Ephrin repulsive force generate bud formation. Wnt-producing Paneth cells are depicted in blue; Wnt-activated cells are shown in pink.

  4. Fig. 4 Basic and clinical applications of an epithelial mini-gut.

    An epithelial mini-gut is efficiently established from a single (3 to 5 mm2) endoscopic biopsy sample. EDTA chelation releases ~3000 crypts from a biopsy sample. An epithelial mini-gut grows logarithmically and expands 1000-fold within a month. Three applications of epithelial mini-guts are as follows: (i) As an experimental tool. Genetic manipulation, gene expression analysis, live imaging, and other standard biological analyses can be employed for normal and patient-derived epithelial mini-guts. (ii) As a diagnostic tool. Patient-derived epithelial mini-guts recapitulate in vivo intestinal epithelial functions and genetic signatures. Efficient expansion of pure epithelial cells provides a high-quality source for deep sequencing or functional assays. (iii) As a therapeutic tool. Epithelial mini-gut transplantation may become a feasible regenerative therapy.