Ishizuya-Oka A (2007), Regeneration of the amphibian intestinal epithe...

XB-ART-35433

Dev Growth Differ 2007 Feb 01;492:99-107. doi: 10.1111/j.1440-169X.2007.00913.x.

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Regeneration of the amphibian intestinal epithelium under the control of stem cell niche.

Ishizuya-Oka A .

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The epithelium of the mammalian digestive tract originates from stem cells and undergoes rapid cell-renewal throughout adulthood. It has been proposed that the microenvironment around the stem cells, called 'niche', plays an important role in epithelial cell-renewal through cell-cell and cell-extracellular matrix interactions. The amphibian intestine, which establishes epithelial cell-renewal during metamorphosis, serves as a unique and good model for studying molecular mechanisms of the stem cell niche. By using the organ culture of the Xenopus laevis intestine, we have previously shown that larval-to-adult epithelial remodeling can be organ-autonomously induced by thyroid hormone (TH) and needs interactions with the surrounding connective tissue. Thus, in this animal model, the functional analysis of TH response genes is useful for clarifying the epithelial-connective tissue interactions essential for intestinal remodeling at the molecular level. Recent progress in culture and transgenic technology now enables us to investigate functions of such TH response genes in the X. laevis intestine and sheds light on molecular aspects of the stem cell niche that are common to the mammalian intestine.

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Species referenced: Xenopus laevis
Genes referenced: bmp4 chrd fabp2 mmp11 shh

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	Fig. 1. Comparison between the amphibian intestine from larval to adult form and the mammalian intestine. During amphibian metamorphosis, the simple larval intestine is remodeled into the more complex one, similar to the mammalian intestine. The brush border of the amphibian larval intestine is longer than that in the adult intestine and in the mammalian intestine. The amphibian adult epithelium undergoes cell-renewal along the trough-crest axis of intestinal folds, similar to that along the crypt-villus axis in the mammalian intestine. All of the mammalian intestinal epithelial cells originate from stem cells in the crypt, where several kinds of cells act as niche players.
	Fig. 2. Adult epithelial primordia surrounded by subepithelial fibroblasts during Xenopus laevis metamorphosis. (A) Primordia of the adult intestinal epithelium (AE) consist of undifferentiated cells, whose cytoplasm is poor in cell organelles. They appear as islets between the degenerating larval epithelium (LE) and the connective tissue (CT). (B) Higher magnification of the site indicated by the box in (A). Subepithelial fibroblast (Fb) makes cell contact (arrowhead) with the adult epithelial cell through the basal lamina (bl), which becomes thick and amorphous. (C) Fibroblasts just beneath the adult epithelial primordia possess well-developed rough endoplasmic reticulum (rer). Bars, 1 μm.
	Fig. 3. Schematic drawing showing thyroid hormone (TH) response genes that are expressed either in the epithelium or the connective tissue during Xenopus laevis intestinal remodeling. Some of the genes, including Shh and ST3, are directly upregulated by TH, whereas the others are indirectly upregulated.
	Fig. 4. Strategy for the functional analysis of thyroid hormone (TH) response genes. If genes encode the secretory protein, their effects can be examined by adding the encoded protein and its antagonist to the medium in the organ culture system (A), where tubular intestines isolated from tadpoles are slit open lengthwise and cultured at 26°C. If genes encode the transcription factor, a gene transfer system using electroporation (B) is available. In the intestine transfected with green fluorescent protein (GFP)-expressing plasmid and then cultured, its expression attains its maximum around day 3, when the GFP protein is detectable uniformly in the epithelium. Intestines isolated from transgenic frogs (C) are also useful for the organ culture system. CT, connective tissue; E, epithelium; M, muscle.
	Fig. 5. BMP-4 mRNA expression in the Xenopus laevis intestine during metamorphic climax (A) and the stage 57 tadpole intestines cultured for 5 days in the presence of thyroid hormone (TH) (B–D). (A) When adult epithelial primordia (AE) grow in size, signals in the connective tissue (CT) become stronger with a gradient toward the primordia. (B) Intact intestine. BMP-4 expression is upregulated in the connective tissue just beneath the epithelium. (C, D) Epithelium-free intestines in the absence (C) and presence of Shh (500 ng/mL) (D). Shh induces BMP-4 expression only in the connective tissue. M, muscle. Scale bars, 20 μm.
	Fig. 6. Effects of exogenous BMP-4 and Chordin proteins on the X. laevis intestine at stage 57 and cultured for 5 days (A–C) and 7 days (D–F) in the presence of thyroid hormone (TH). Sections were stained with methyl green-pyronin Y (A–C) and immunostained with anti-IFABP antibodies (D–F). (A, D) Control intestines cultured without BMP-4 and Chordin. Primordia (arrowheads) of the adult epithelium (AE) become detectable between the larval epithelium (LE) and the connective tissue (CT) (A) and then differentiate into the absorptive epithelium expressing IFABP (D). (B, E) In the presence of BMP-4 (100 ng/mL). The single layer of the adult epithelium is precociously differentiated on day 5 (B) and its immunoreactivity for IFABP is much higher (E) than that in the control intestine (D). (C, F) In the presence of Chordin (1 μg/mL). Adult progenitor cells positive for pyronin Y (arrows) are fewer (C) than those in the control intestine on day 5 (A) and IFABP expression remains undetectable until day 7 (F). Scale bars, 20 μm.