Sterling J , Fu L , Matsuura K .

Intramural NIH HHS

	Figure 2. Morphological changes in the Xenopus tropicalis intestine during natural metamorphosis.The intestine of Xenopus tropicalis tadpoles at stage 54 to 66 was isolated and stained with MGPY and photographed with a light microscope. A–C and G–I: a cross-section of the intestine at the indicated stage. D–F and J–L: the enlarged photo of the boxed area in A–C and G–I, respectively. Note that during metamorphosis, the MGPY staining became weaker in larval epithelium as the cells undergo degeneration. At the climax of metamorphosis, the newly formed, proliferating adult epithelial islets were strongly stained by MGPY (arrows). The connective tissue (CT) and muscles (Mu) increased dramatically during metamorphosis. Ep: epithelium; Lu: lumen; Ty: typhlosole; Scale bars: 50 µm.
	Figure 3. T3 induces intestinal remodeling in premetamorphic X. tropicalis tadpoles.Stages 54 X. tropicalis tadpoles were treated with 10 nM T3 at 25°C. The intestine was isolated from the tadpoles at day 0, 1, 3, and 5, respectively. The intestine was fixed, sectioned, and stained with MGPY as in Fig. 2. A–D: a cross-section of the intestine after 0, 1, 3, 5 days of T3 treatment, respectively. E–H: the enlarged image of the boxed area for the corresponding tissues in A–D, respectively. Note that after 3 days of T3 treatment, the MGPY staining became weaker in larval epithelium as the cells undergo apoptosis. At the same time, the newly formed, proliferating adult epithelial islets were strongly stained by MGPY (arrows). The connective tissue (CT) and muscles (Mu) increased dramatically during metamorphosis. Ep: epithelium; Lu: lumen; Ty: typhlosole. Scale bars: 50 µm.
	Figure 4. The typhlosole is present in the entire small intestine of premetamorphic X. tropicalis tadpoles (A) but only in the anterior small intestine of premetamorphic X. laevis tadpoles (B).A schematic diagram of the intestine from the anterior to the posterior is shown on the left (dashed lines indicate the boundaries of typhlosole with the intestines). On the right of each panel shows representative MGPY-stained cross-sections of the intestine from indicated regions of the intestine of premetamorphic tadpoles at stage 54. In the middle is a schematic drawing of the cross-section showing the presence or absence of the typhlosole. Note the presence of the typhlosole in the posterior half of the small intestine in X. tropicalis but not X. laevis.
	Figure 5. Reductions in the length of typhlosole and the small intestine of premetamorphic X. laevis and X. tropicalis tadpoles upon T3 treatment.Stage 54 X. laevis or tropicalis tadpoles were treated with 10 nM T3 at 18°C (Xenopus laevis tadpoles) or 25°C (Xenopus tropicalis tadpoles) for the indicated days and the intestine was isolated. The length of the typhlosole and the small intestine were measured. Note that the lengths of the intestine was reduced upon T3 treatment and that after 4–5 days of treatment, the typhlosole was no longer identifiable due to metamorphic changes in the intestine, resembling that at the climax of metamorphosis. The faster changes for X. tropicalis tadpoles were in part due to the higher temperature at which the animals needed to be reared.
	Figure 6. Intestinal epithelial apoptosis occurs during natural and T3-induced metamorphosis in X. tropicalis.The intestine from tadpoles at premetamorphic stage 54 (ST54), the climax of metamorphosis (ST61), or ST54 but treated with 5 nM T3 for 3 days (ST54+ T3), were analyzed by TUNEL assay (A–C) or TUNEL without the enzyme TdT as a the negative control (D–F). The biotin-dUTP labeled apoptotic cells were visualized with Texas-Red labeled Streptavidin and the nuclei were stained with DAPI. The boxed areas were shown at a higher magnification (a-f) with the apoptotic cells indicated with yellow arrows in either merged (DAPI+Texas Red) and single channel of red fluorescence (Texas-Red) images. Scale bar: 50 µm.
	Figure 7. ST3 is upregulated while IFABP is downregulated during X. tropicalis intestinal metamorphosis.Total RNA were extracted from intestines of tadpoles at the indicated stages and analyzed by qRT-PCR. The expression levels for ST3 or IFABP were normalized against the control gene rpl8 and represented as means with with standard deviations in arbitrary units.

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