Figure 3

Local E2 injection activates Ptch1 and Gli1 expression in injured nerves. Ptch1-LacZ mice and Gli1-LacZ mice received local injections of E2 or saline (placebo) and PLGA 1 week before sciatic nerve-crush injury; subsequent assessments were performed in mice killed 3 days after injury, when both Ptch1 and Gli1 mRNA expression were significantly upregulated in wild-type mice (Figures 2c and d). (a, b) The activation of Ptch1-LacZ protein expression was evaluated by staining (a) whole-mount tissues and (b) cross sections of the injured nerves from Ptch1-LacZ mice with X-gal (blue). (c, d) The activation of Gli1-LacZ expression was evaluated in X-gal–stained sections of (c) epineural and (d) endoneural tissue from the injured nerves of Gli1-LacZ mice. (e) Gli1 mRNA expression was evaluated in Gli1-LacZ mice treated with placebo, with E2 injections, or with injections of E2 and the E2-receptor blocker ICI; measurements were performed via qRT-PCR, normalized to endogenous 18S rRNA levels, and presented relative to the values obtained in placebo-treated mice. (f) X-gal–stained sections from uninjured limbs and from the injured limbs of placebo-treated and E2-treated mice were co-stained for expression of the endothelial-specific marker CD31 (brown) to identify endothelial cells that expressed Gli1 (blue). ^*P<0.01 vs uninjured; ^†P<0.01 vs placebo or E2+ICI.