Journal: PLoS ONE

Article Title: Early Systemic Granulocyte-Colony Stimulating Factor Treatment Attenuates Neuropathic Pain after Peripheral Nerve Injury

doi: 10.1371/journal.pone.0043680

Figure Lengend Snippet: Immune cell subpopulations of tissue fluid were quantified by flow cytometry. (A) At 48 h, increased expression of markers, such as, PMN cell marker (stained using anti-PMN-FITC), and β-endorphin marker (stained using anti-β-endorphin-PE), on leukocytes is observed in G-CSF–treated rats with CCI than in vehicle-treated rats with CCI (n = 6 per group). CD45-PE-Cy5 and ED2-PE antibodies were used to react with all hematopoietic cells and macrophages, respectively. Low staining with the respective control antibodies, confirmed a specific antibody staining of opioid-containing PMN cells. In contrast, low staining with ED2-PE and PMN-FITC antibodies (0.3–3.4%), confirmed a specific staining of PMN cells by PMN-FITC antibody. (B) At 48 h, the number of β-endorphin–containing PMN cells in G-CSF–treated rats with CCI is significantly higher than that in vehicle-treated rats with CCI. Data are shown as the mean±SEM, n = 6 per group; t test, * p <0.05.

Article Snippet: For intracellular staining, the cells were prepared and incubated with rabbit anti-rat β-endorphin-PE polyclonal antibody that can recognize opioid peptides (β-endorphin; 1∶500; Bioss).

Techniques: Flow Cytometry, Expressing, Marker, Staining

Journal: PLoS ONE

Article Title: Early Systemic Granulocyte-Colony Stimulating Factor Treatment Attenuates Neuropathic Pain after Peripheral Nerve Injury

doi: 10.1371/journal.pone.0043680

Figure Lengend Snippet: (A–C) Confocal laser scanning microscopy of PMN cells and β-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 β-endorphin peptides were observed in the vehicle-treated sham operation rats. However, at 12 h, the count of β-endorphin-containing PMN cells was significantly higher in the G-CSF-treated CCI rats than in the vehicle-treated CCI rats. Scale bar: 100 µm. (D) Confocal high-power field laser scanning pictures showing a PMN cell with a typical segmented nucleus, which stained positive for β-endorphin peptides. Green: PMN cells; red: β-endorphin (END)-containing cells; blue (DAPI): cell nuclei. Scale bar: 10 µm. (E) The number of β-endorphin-containing PMN cells per section as calculated at different time points. The count of β-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.

Article Snippet: For intracellular staining, the cells were prepared and incubated with rabbit anti-rat β-endorphin-PE polyclonal antibody that can recognize opioid peptides (β-endorphin; 1∶500; Bioss).

Techniques: Confocal Laser Scanning Microscopy, Staining