The localization of the C-terminal SHH protein (precursor form only, Fig. 1) was similar to N-terminal SHH, with two notable differences. Precursor SHH protein was abundant in the perineurium of the nerves, but was not present in the Schwann cells or the veins, as was the active form of SHH protein. SHH/PECAM1 and PTCH1/PECAM1 dual staining shows that SHH and PTCH1 do not colocalize with PECAM1, indicating that SHH and PTCH1 are not present in endothelial cells.
Shh and Ptch1 RNA Expression Increased Significantly by RT-PCR Analysis Following CN Injury
Relative changes in RNA expression of Shh and Ptchl were quantified by RT-PCR in control and CN-injured penes (Fig. 3). Shh and Ptchl RNA expression significantly increased 1.25-fold and 2-fold, respectively, 21 days after CN injury (P-values = 0.01 and 0.002, respectively). Shh RNA expression remained unchanged when assayed at 7 (control = 0.25 6 0.05, CN-injured = 0.39 6 0.1, P-value = 0.13) and 14 days post-CN injury (control = 0.46 6 0.1, CN-injured = 0.32 6 0.08, P-value = 0.12). Bmp4 expression was also measured 21 days after CN injury. Bmp4 expression was unchanged following CN injury (control = 0.62 6 0.08, CN-injured = 0.71 6 0.09, P-value = 0.25).
FIG. 3. Shh and Ptch1 RNA expression in control and CN-injured rats. Relative abundance of Shh and Ptch1 RNA expression measured by semiquantitative RT-PCR analysis in control and CN-injured Sprague Dawley rat penes 21 days after CN injury. Shh and Ptch1 expression were significantly increased 21 days after CN injury (P-value — 0.01 and 0.002, respectively). Asterisks identify significant changes in expression. C, Control;CN, cavernous nerve injury.