Localization of SHH and PTCH1 in the Adult Penis by IHC
Antibodies that recognize the active and precursor forms of SHH protein and PTCH1 were used to determine their localization in adult Sprague Dawley penis tissue. N-terminal SHH protein (active and precursor forms) was localized within the corpora cavernosal sinusoidal tissue, a layer adjacent to the tunica, Schwann cells of the nerves, veins (Fig. 1), and the urethra. Colocalization of SHH and ACTA1 by dual IHC identifies SHH in smooth muscle cells of the corpora cavernosal sinusoidal tissue.
SHH is present to a lesser extent in cells residing between the sinusoids of the corpora. These cells which stain for SHH protein colocalize with P4HB, a fibroblast marker (Fig. 2). The localization of PTCH1 protein was very similar to SHH (Figs. 1 and 2), with staining present within the corpora cavernosal sinusoidal tissue, a layer adjacent to the tunica, Schwann cells of the nerves, veins and the urethra. Colocalization of PTCH1 and ACTA1 by dual IHC identify PTCH1 in smooth muscle cells of the corpora cavernosal sinusoidal tissue.
FIG. 1. IHC analysis of SHH (N-19 and C-18) and PTCH1 in adult Sprague Dawley rat penes. a) The precursor and active forms of SHH protein recognized by SHH (N-19) are abundant within corpus cavernosal sinusoidal tissue and a layer under the tunica (left), nerves of the dorsal nerve bundle, and veins (middle). n, Nerves;v, vein. b) The localization of PTCH1 is similar to the active form of SHH. c) The localization of the precursor SHH protein (C-18) is similar to that of the active form. However, it is not present in the veins and is restricted to the perineurium of the nerves. Elastin fibers in the veins autofluoresce and do not stain for SHH protein. Original magnification X60.
FIG. 2. Dual IHC for SHH/ACTA1, SHH/ PECAM1, SHH/P4HB, PTCH1/ACTA1, and PTCH1/PECAM1 in adult Sprague Dawley rat penes. SHH and PTCH1 proteins coloc-alize with ACTA1 in the corpus cavernosal sinusoidal tissue. In addition, SHH protein is present in cells located between the sinusoids. These cells stain for P4HB, a fibroblast marker. SHH and PTCH1 proteins are shown in green, and ACTA1, PECAM1, and P4HB proteins are presented in red. Yellow represents dual staining. Neither SHH nor PTCH1 colocalize with PECAM1, indicating that SHH and PTCH1 are not present in endothelial cells. Original magnification X400 (ACTA1 and PECAM1) and X1000 (P4HB).