Fig. S5.

(A) Development of CD34⁺ CSCs is not affected by the intestinal microbiota. Immunofluorescence analysis of gp38 (green) and CD34 (red) expression by colon lamina propria cells of 3-wk-old B6 mouse bred in SPF (Left) or GF (Right) conditions. Arrows indicate double positive CD34⁺ CSCs. Representative images of three independent experiments are shown. At Right, percentage of CD34⁺ CSCs, as measured by FACS, in the colon lamina propria of SPF, GF or Myd88^−/−TRIF^−/− mice. n = 4 from two independent experiments, values are mean ± SD. (Scale bars: 50 µm.) (B) Gremlin1 expression is detected in the SML and in proximity to crypts. Immunofluorescence analysis of Gremlin1 (green) and CD34 (red) expression by colon stromal cells of mice at the indicated age (E, days after conception; P, days postnatal). Insets show expression of gremlin1 in the SML at 2 wk (P14) and in the pericryptal region at 3 wk after birth (P21). Red signal in intestinal lamina propria of E16.5 embryos and 1-wk-old mice (P7) indicate CD34⁺ blood vessels. Images are representative of two independent experiments. C, crypt; L, lumen; ns, not significant; SML, submucosal layer. DAPI stains nuclei. (Scale bar: 50 μm.)