(A–D) Immunohistochemical staining of Ptc1-LacZ adult hearts with Abs against cardiac actin (B, green) and β-galactosidase (C, red) demonstrating PTC1 expression in cardiomyocytes (white arrows) and large coronary vessels (white arrowheads). A shows the corresponding differential interference contrast (DIC) microscopy image. (E–H) Immunohistochemical staining of Ptc1-LacZ adult hearts with Abs against PECAM (E, blue), SMA (F, green), and β-galactosidase (G, red). PTC1 expression was detected in both cardiomyocytes and perivascular smooth muscle cells. White arrowheads denote colocalization of PTC1 and SMA expression. (I–L) Immunohistochemical staining with Abs against PECAM (I, blue), vimentin (J, green), and SHH (K, red) revealed that SHH is expressed in adventitial and interstitial fibroblasts. White arrowheads denote colocalization of SHH and vimentin expression. The asterisk indicates area of vimentin and PECAM coexpression. (M–P) Immunostaining with PECAM (M, blue), SHH (N, green), and β-galactosidase (O, red) demonstrates a close association among SHH-expressing interstitial fibroblasts, PTC1-expressing cardiomyocytes, and blood vessels (white arrowhead). In E–G, I–K, and M–O, florescent signals are overlaid on top of DIC images. D, H, L, and P show merged images of A–C, E–G, I–K, and M–O, respectively. Original magnification, ×400 (A–D, I–L, and M–P); ×200 (E–H).