1. Effect of D‐Asp treatment on myelination and remyelination in cerebellar organotypic slices
Effect of D‐Asp treatment on myelination and remyelination in cerebellar organotypic slices ASchematic diagram showing D‐Asp exposure protocol in organotypic slices.BMaximum intensity projection of z‐stack confocal images displaying MBP (red) and NF200 (green) immunoreactivities in seven DIV cerebellar organotypic slices cultured in the absence (b–d) or in the presence of 100 μM D‐Asp (f–h). Panels (d and h) display colocalized points (white). Panels (a and e) show representative low magnification images of seven DIV cerebellar slices cultured cultures in the absence (a) or in the presence of 100 μM D‐Asp (e). Scale bars in (a and e): 200 μm; in (b–d) and (f–h): 50 μm.CScatter plot histogram analysis of the myelination index in seven DIV cerebellar slices cultured in the absence or in the presence of 100 μM D‐Asp.DWestern blotting analysis of MBP protein levels from homogenates of organotypic cerebellar slices cultured in the absence or in the presence of 100 μM D‐Asp. Data were normalized on the basis of β‐actin and expressed as percentage of controls.ESchematic diagram showing D‐Asp exposure protocol in organotypic slices after LPC exposure.FMaximum intensity projection of z‐stack confocal images displaying MBP (red) and NF200 (green) immunoreactivities in control cerebellar organotypic slices (b–d), and in cerebellar slices at 6 days after LPC exposure (dpl) in the absence (f–h) or in the presence 100 μM D‐Asp (j–l). Panels (d, h, and l) display colocalized points (white). Panels (a, e, and i) show representative low magnification images of seven DIV cerebellar slices cultured cultures in the absence (a) or in the presence of 100 μM D‐Asp (e). Scale bars in (a, e, i): 200 μm; in (b–d, f–h, and j–l): 50 μm.GWestern blotting and quantitative analysis of MBP protein levels from homogenates of organotypic cerebellar slices at 6 dpl cultured in the absence or in the presence of 100 μM D‐Asp.HScatter plot histogram analysis of the remyelination index in cerebellar explants at 6 dpl after LPC exposure cultured in the absence or in the presence of 100 μM D‐Asp. Data were normalized to vehicle control.IMaximum intensity projection of z‐stack confocal images displaying MBP (red) and NF200 (green) immunoreactivities in control cerebellar organotypic slices (b–d), and in cerebellar slices at 10 dpl in the absence (f–h) or in the presence of 100 μM D‐Asp (j–l). Panels (d, h, and l) display colocalized points (white). Panels (a, e, and i) show representative low magnification images of seven DIV cerebellar slices cultured cultures in the absence (a) or in the presence of 100 μM D‐Asp (e). Scale bars in (a, e, i): 200 μm; in (b–d, f–h, and j–l): 50 μm.JScatter plot histogram analysis of the remyelination index in cerebellar explants at 10 dpl cultured in the absence or in the presence of 100 μM D‐Asp. Data were normalized to vehicle control.Data information: The values represent the means ± SEM. Level of significance was determined by using: in (C) two‐tailed Student's t‐test, *P < 0.05 versus control (n = 4 animals, 3–4 slices per group); (D) two‐tailed Student's t‐test, *P < 0.05 versus controls (n = 3); (G) one‐way ANOVA P = 0.003 followed by Bonferroni post hoc test, *P < 0.05 versus control, ˄P < 0.05 versus LPC (n = 3); (H) one‐way ANOVA P < 0.001 followed by Bonferroni post hoc test, *P < 0.05 versus controls, ˄P < 0.05 versus LPC (n = 4 animals, 3–4 slices per group); (J) one‐way ANOVA P < 0.001 followed by Bonferroni post hoc test, *P < 0.05 versus controls, ˄P < 0.05 versus LPC (n = 4 animals, 3–4 slices per group). See the exact P‐values from comparisons tests in Appendix Table S2.Source data are available online for this figure.
Valeria de Rosa et al. EMBO Mol Med. 2018;emmm
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