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Fig. 6. ISEMFs express Ptch1 and are responsive to Shh stimulation. (A,B) ISEMF cell lines from rat jejunum (MIC101), ileum (MIC216), colon (MIC316), and human colon (CCD-18Co) respond to 24- or 72-hour treatment with Shh-conditioned medium (CM) from Cos7 cells transfected with pcDNA3.1-Shh (Shh CM) or pcDNA3.1 (control CM). Q-RT-PCR analysis reveals upregulation of Gli target genes Ptch1 (A) and Gli1 (B). Error bars represent s.e.m. of triplicate analysis. (C) X-gal stain marks nuclear LacZ expression from Ptch1 locus in cells beneath intervillus epithelium in P0 Ptch1+/nLacZ small intestine. (D-F) Confocal microscopy of immunofluorescence for {alpha}-SMA (red) and desmin (green) identifies ISEMFs beneath epithelium. False-color overlay of X-gal stain (blue) reveals that ISEMF cells surrounding precrypts (arrows) express Ptch1. X-gal-positive nuclei are also found in scattered surface SMCs of the muscularis extrema (C,D, arrowhead) and in rare desmin-positive cells in the lamina propria (F, arrowhead). Scale bar: 20 µm.