Images: G-CSF treatment increases the number of migrated b-endorphin-containing PMN cells in injured nerves. (A-C) Confocal laser scanning microscopy of PMN cells and b-endorphin peptides in the right sciatic nerve of vehicle-treated sham rats (A1-3), vehicle-treated CCI rats (B1-3), and G-CSF treated CCI rats (C1-3) 12 h after CCI, respectively. No apparent PMN cells or b-endorphin peptides were observed in the vehicle-treated sham operation rats. However, at 12 h, the count of b-endorphin-containing PMN cells was significantly higher in the G-CSF-treated CCI rats than in the vehicle-treated CCI rats. Scale bar: 100 mum. (D) Confocal high-power field laser scanning pictures showing a PMN cell with a typical segmented nucleus, which stained positive for b-endorphin peptides. Green: PMN cells; red: b-endorphin (END)-containing cells; blue (DAPI): cell nuclei. Scale bar: 10 mum. (E) The number of b-endorphin-containing PMN cells per section as calculated at different time points. The count of b-endorphin-containing PMN cells was higher in the CCI rats treated with G-CSF (black bars) compared to those treated with vehicle (grey bars) between 12-48 h after nerve injury. Data are shown as the means±SEM, n = 5 per group; one-way ANOVA, *p<0.05, **p<0.01: CCI + G-CSF group compared to CCI + vehicle group; #p<0.05, ##p<0.01: CCI + G-CSF or CCI + vehicle group compared to vehicle-treated sham operation control. doi:10.1371/journal.pone.0043680.g004