Figure 4.
Recruited and resident MPs exert distinct functions in nerve regeneration. (a) TPLSM images show the qualitative defect in myelin degeneration in CCR2-deficient compared with CCR2 proficient mice. Images are zoomed in representative region from Fig. 3 b. Scale bar = 15 µm. (b) Comparative quantification (mean ± 95% CI) of the myelin surface between WT and Ccr2−/− mice based on TPLSM images of nerve cryosections (n = 3–4 mice per time point processed independently; P values of two-way ANOVA with Bonferroni multiple comparison tests between WT and Ccr2−/− mice are indicated). (c) Fluorescent images of NF200 staining of sciatic nerves in WT mice show the axon degeneration and regeneration after CCI. Scale bars = 100 µm for the upper image and 50 µm for lower images. (d) Quantification (mean ± 95% CI) of the NF200 staining between WT and Ccr2−/− mice based on fluorescent images of nerve cryosections (n = 3–5 mice per time point, processed independently; two-way ANOVA with Bonferroni multiple comparison tests unveiled no significant difference between WT and Ccr2−/− mice). (e) TPLSM image (left panel) shows example of foamy/phagocytic ECFP+ cell morphologies compared with EGFP+ cells (scale bar = 20 µm) and (right panel) shows the quantification of the proportion of phagocytic cells among ECFP+ cells (middle panel) and EGFP+ cells (right panel) following CCI (n = 3–4 mice per time point, processed independently; P values of one-way ANOVA with Bonferroni multiple comparison tests are indicated). (f) Schema of anti-CSF1R treatment to prevent resident MPs accumulation 30 d after CCI. (g) Representative TPLSM images (scale bar = 20 µm) and quantification of cell numbers show the complete abrogation of EGFP+ MP accumulation in anti-CSF1R treated mice 30 d after CCI (n = 3 mice representative of two independent experiments; P values of two-way ANOVA with Bonferroni’s multiple comparison test are indicated). (h) Quantification of myelin roundness and surface show qualitatively reduced myelin regeneration after anti-CSF1R treatment (n = 3 mice, representative of two independent experiments; P values of unpaired t tests are indicated). (i) Left: Fluorescent images of NF200 staining of sciatic nerve cryosection in WT mice treated or not with anti-CSF1R 30 d after CCI. Scale bars = 500 µm for left images and 100 µm for right images. (j) Quantification (mean ± 95% CI) of NF200 staining based on fluorescent images of nerve cryosections (n = 3 mice, representative of two independent experiments; P values of unpaired t test are indicated). See also Fig. S3, Video 1, and Video 2. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Recruited and resident MPs exert distinct functions in nerve regeneration. (a) TPLSM images show the qualitative defect in myelin degeneration in CCR2-deficient compared with CCR2 proficient mice. Images are zoomed in representative region from Fig. 3 b. Scale bar = 15 µm. (b) Comparative quantification (mean ± 95% CI) of the myelin surface between WT and Ccr2−/− mice based on TPLSM images of nerve cryosections (n = 3–4 mice per time point processed independently; P values of two-way ANOVA with Bonferroni multiple comparison tests between WT and Ccr2−/− mice are indicated). (c) Fluorescent images of NF200 staining of sciatic nerves in WT mice show the axon degeneration and regeneration after CCI. Scale bars = 100 µm for the upper image and 50 µm for lower images. (d) Quantification (mean ± 95% CI) of the NF200 staining between WT and Ccr2−/− mice based on fluorescent images of nerve cryosections (n = 3–5 mice per time point, processed independently; two-way ANOVA with Bonferroni multiple comparison tests unveiled no significant difference between WT and Ccr2−/− mice). (e) TPLSM image (left panel) shows example of foamy/phagocytic ECFP+ cell morphologies compared with EGFP+ cells (scale bar = 20 µm) and (right panel) shows the quantification of the proportion of phagocytic cells among ECFP+ cells (middle panel) and EGFP+ cells (right panel) following CCI (n = 3–4 mice per time point, processed independently; P values of one-way ANOVA with Bonferroni multiple comparison tests are indicated). (f) Schema of anti-CSF1R treatment to prevent resident MPs accumulation 30 d after CCI. (g) Representative TPLSM images (scale bar = 20 µm) and quantification of cell numbers show the complete abrogation of EGFP+ MP accumulation in anti-CSF1R treated mice 30 d after CCI (n = 3 mice representative of two independent experiments; P values of two-way ANOVA with Bonferroni’s multiple comparison test are indicated). (h) Quantification of myelin roundness and surface show qualitatively reduced myelin regeneration after anti-CSF1R treatment (n = 3 mice, representative of two independent experiments; P values of unpaired t tests are indicated). (i) Left: Fluorescent images of NF200 staining of sciatic nerve cryosection in WT mice treated or not with anti-CSF1R 30 d after CCI. Scale bars = 500 µm for left images and 100 µm for right images. (j) Quantification (mean ± 95% CI) of NF200 staining based on fluorescent images of nerve cryosections (n = 3 mice, representative of two independent experiments; P values of unpaired t test are indicated). See also Fig. S3, Video 1, and Video 2. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

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