Fig. 8: Local intramuscular CCL2 neutralisation or IgG treatment ameliorates NMJ pathology and reduces immune cell infiltration in the GC muscle of hTDP-43^Tg/Tg mice.

A Schematic of experimental design. B–D Representative images of NMJ denervation in the gastrocnemius (GC) muscle of untreated (B), IgG-treated (C) and CCL2-neutralising antibody-treated (D) late symptomatic postnatal day 19 (P19) hTDP-43^Tg/Tg mice. Red arrows indicate fully innervated NMJs, as revealed by the presynaptic NEFH/SV2a staining. Scale bar represents 40 µm. E–G Representative images of the CD45⁺ leukocyte infiltration in the innervation zone of the GC muscle in untreated (E), IgG-treated (F) and CCL2-neutralising antibody-treated (G) late symptomatic hTDP-43^Tg/Tg mice. Scale bar represents 40 µm. H Qualitative analysis of NMJ innervation, representing the ratio of fully innervated NMJs in the GC muscle. Analysis was performed with n = 7 (untreated) and n = 4 (IgG, CCL2) mice/group and one-way ANOVA with Fisher’s LSD post-hoc, no treatment vs. IgG: p < 0.0001, no treatment vs. CCL2: p < 0.0001. On average 30-40 NMJs were analysed per muscle. I–K Quantitative morphometric analysis of the NMJs revealed improvement in axonal diameter (I), nerve terminal area (J) and overlap (K) variables. The exact p values and statistical analyses are detailed in Table 2. L, M Quantitative analysis of CD45⁺ (L) and CCR2⁺ (M) cell count in the late symptomatic GC muscle of hTDP-43^Tg/Tg mice. Analysis was performed on n = 8 (untreated) and n = 4 (IgG, CCL2) mice/group. Data analysis: (L) CD45 analysis: Kruskal-Wallis test and uncorrected Dunn’s post-hoc [Shapiro-Wilk normality test, ALS untreated group: p = 0.0091, ALS IgG group: p = 0.7802, ALS CCL2 group p = 0.3820], no treatment vs. IgG: p = 0.0114, no treatment vs. CCL2: p = 0.0053; (M) CCR2 analysis: one-way ANOVA with Fisher’s LSD post-hoc, no treatment vs. IgG: p < 0.0001, no treatment vs. CCL2: p = 0.0002. * = p < 0.05; ** = p < 0.01; *** = p < 0.001. Data is presented as mean ± sem.