Fig. 3

Effect of PC1 treatment on macrophage activation and cytokine levels in the sciatic nerve. a, b mRNA levels of CD68 and the percentage of CD68 positive area in the sciatic nerve of CTR, BTZ 14 days (day 14 corresponding to c.d. 2,4 mg/kg), BTZ 28 days (day 28, corresponding to c.d. 4,8 mg/kg), and PC1 (BTZ + PC1) mice. Quantitative analysis of CD68 signal was computed as integrated optical density for arbitrary areas (6 sections per animal, 5 animals per group). c Representative immunofluorescence images of CD68 in the sciatic nerve sections of the same experimental groups. One-way ANOVA was used for statistical evaluation, followed by Bonferroni’s test for multiple comparisons. **p < 0.01, ***p < 0.001 vs CTR; °p < 0.05 vs BTZ day 28. d Immunofluorescence double-staining shows the colocalization (yellow, arrowhead) of PK2 (green) with CD68 (activated macrophages, red) in the sciatic nerve of CTR, BTZ day 14, BTZ day 28, and BTZ + PC1 mice (PC1). Cell nuclei were counterstained with DAPI (blue fluorescence). e–h The mRNA levels of TLR4 and of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 respectively. i The mRNA of the anti-inflammatory cytokine IL-10. All the measurements were performed 14 days after the first BTZ administration (c.d. 2.4 mg/kg), before starting PC1 treatment (CTR and BTZ groups), and at the end of the BTZ protocol (c.d. 4.8 mg/kg) (CTR, BTZ, BTZ + PC1 groups). mRNA levels, determined by real-time qPCR, were expressed in relation to GAPDH and presented as fold-increases over the levels of CTR animals (at the same time point). (a, e–i) Data represent mean ± SD of 6 mice/group. At day 14, statistical analysis was performed by means of t test while at day 28 by using one-way ANOVA followed by Bonferroni’s post-test. *p < 0.05, **p < 0.01, ***p < 0.001 vs vehicle/CTR (at the same time point); °p < 0.05, °°°p < 0.001 vs BTZ day 28