Figure 5: Detection of apoptosis by TUNEL, cavernous caspase-3 immunohistochemical staining and western blot. (a) The TUNEL method; (b) bar graph of apoptotic index; (c) cavernous caspase-3 immunohistochemical staining; (d) bar graph of caspase-3 positive area; (e) western blot of the Akt/Bad/Bax/caspase-3 pathways; (f) bar graph of relative p-AKT/AKT expression; (g) bar graph of relative p-Bad/Bad expression; (h) bar graph of relative Bax/Bcl-2 expression; (i) bar graph of relative caspase-3 activity; scale bars = 50 μm; n = 6 in each group. Representative micrographs of TUNEL show apoptotic cells as black-brown dots (×400). Representative micrographs of cavernous caspase-3 immunohistochemical staining show apoptotic cells as brown areas (×200). Bar graph depicts caspase-3 activity in penile tissues measured by a Caspase-3 Activity Assay Kit, which was calculated according to a standard curve and normalized by the protein concentration. Data of western blot analysis and caspase-3 activity assay are shown as the fold changes over the control group. Bar graphs represent mean ± standard deviation. *P < 0.05 and **P < 0.01, the indicated group compared with the Sham group; #P < 0.05 and ##P < 0.01, the indicated group compared with the BCNI group. BCNI: bilateral cavernous nerve injury; TUNEL: terminal dexynucleotidyl transferase(TdT)-mediated dUTP nick end labeling; Bcl-2: B-cell lymphoma 2; Bax: Bcl-2-associated X protein; Bad: Bcl-2-associated death promoter; p-Bad: phosphor-Bad; AKT: the protein kinase B; p-AKT: phosphor-AKT.