Fig. 6: ADAMTS18 promotes villus tip vessel and epithelial cell integrity.

a Villus tip endothelial cell fenestration extends to the villus core side in Adamts18^−/− mice. Electron micrographs of villus tip vessels from wild-type and Adamts18^−/− mice. Zoomed areas from the epithelial side (black box) or villus core side (red box) of villus tip vessels are shown below. Quantification for the percentage of fenestra observed on each side of the vessels (p = 0.0017, n = 4 mice). b, c Increased villus tip vessel leakage in Adamts18^−/− mice. b 100 nm fluorescent beads were injected i.p. and visualized in the intestine by wholemount imaging. In wild-type animals beads (red) accumulated sparsely throughout the villus but were concentrated around venules (green) in villi and the submucosal layer. c Analysis of bead distribution in the villus tip of wild-type and Adamts18^−/− mice. Quantification of the bead area/villus tip vessel area (p = 0.001, n = 4–5 mice). d Amino acid concentration is increased in blood of Adamts18^−/− mice. Blood was sampled from the (left) portal vein of fasted mice or (right) hearts of mice gavaged with high protein solution. (Left, Tyr, p = 0.0051; Met, p = 0.0011; Ser, p = 0.0063; n = 5–9, right, Tyr, p = 0.0188; Gln, p = 0.0005; Glu, p = 0.0046; Asp, 0.0018; n = 3–5 mice; 2 technical replicates/sample displayed). e Villus tip “holes” in Adamts18^−/− mice. LAMA5 staining (red) allows visualization of holes in the basement membrane. Quantification of percentage of villi with basement membrane holes in adult Adamts18^−/− mice or littermate wild-type controls (p = 0.0112, n = 5–6 mice). f Holes run completely through the basement membrane (red, LAMA5) at the villus tips and are covered by epithelial cells (green, EpCAM). Cross-sectional view of a villus tip hole in an Adamts18^−/− mouse; white, VEGFR2; blue, DAPI. Quantification of hole location along the villus/crypt axis, 100 denotes the villus tip. g Bifurcated villi are more abundant in adult Adamts18^−/− mice than littermate wild-type controls. Whole-mount immunostaining for LAMA5 (red) and VEGFR2 (green). Quantification of percentage of villi with split villi in adult Adamts18^−/− mice or littermate wild-type controls (p = 0.0147, n = 5–6 mice). h Scheme for proposed mechanism of VTT-derived ADAMTS18 limiting fibronectin and VEGFA to maintain the polarized endothelial cell phenotype at the intestinal villus tip (created with biorender.com). Scale bars: 50 μm: b (villus) e, g; 20 μm: b (submucosal), c, f; 2 μm: a. All values shown as mean ± SD. Source data are provided as Source Data and Supplementary Data files.