Figure S4.

Sciatic denervation induces an autoimmune response to peripheral nerve antigens. (a) Cellularity of popLNs 1 wk after unilateral sciatic denervation with or without broader ethanol disinfection; n = 3 mice; data are representative of two independent experiments; two-way ANOVA; Šídák’s post-test. (b) An image of popLNs 1 wk after unilateral sciatic denervation with daily topical hypochlorite treatment. (c) Cellularity of popLNs in WT or Myd88-KO mice a week after unilateral sciatic denervation. n = 5 or 6 mice; data are representative of two independent experiments; two-way ANOVA; Šídák’s post-test. (d) Quantification of CD25⁺ Foxp3⁺ regulatory T cells in the naive, sham-operated, or denervated popLNs 7 d after surgery; n = 6 from two independent experiments; one-way ANOVA; Tukey’s post-test. (e) Human epithelial cell (HEp2) slides incubated with sera generated from different time points after unilateral sciatic denervation. Scale bar, 200 µm. (f) Immunoreactivity of serum (green) harvested from mice 0 or 20 d after unilateral sciatic denervation to the intact sciatic nerve. Sections were costained with antibodies directed against CD31 (red). Scale bar, 100 μm. Images are representative of two independent experiments. (g) Immunoreactivity of serum (green) harvested from mice 20 d after unilateral sciatic denervation to the intact sciatic nerve. Sections were costained with antibodies directed against MRP14 (red) and neurofilament (NF; blue). Scale bar, 30 μm. Images are representative of two independent experiments. (h and j) Staining of serum harvested from mice 20 d after unilateral sciatic denervation on the sciatic nerve together with antibodies directed against NF and MPZ. Scale bars, 20 µm. (i and k) Immunoreactivity of serum harvested from mice after unilateral sciatic denervation to the sciatic nerve (i) and popLNs (k). Sections were costained with antibodies directed against CD31 (vasculature), MRP-14 (Schwann cell), NF (nerve), or JAM-C (noncompact myelin). Scale bars, 20 µm (i); and 200 (intact) and 300 (denervated; j) µm. Images are representative of two independent experiments. (l) Quantification of immunoreactivity of sera harvested from mice and normalized to total IgG levels after unilateral sciatic denervation to intact and denervated popLN from before (D0) or 3 wk (D20) after unilateral sciatic denervation on popLNs; n = 4 or 5 mice; data are representative of two independent experiments; two-way ANOVA; Šídák’s post-test. Myd88^−/− mice were 8–12 wk old and bred in the animal facility. Their age- and sex-matched controls and all other mice in this figure were 8–12-wk-old WT males purchased from Charles River or Janvier Labs. All data are presented as mean ± SEM; *, P < 0.05; **, P < 0.01; ****, P < 0.0001. Ab, antibody; MFI, mean fluorescence intensity; Sx, sciatic denervation; Treg, regulatory T cell; W, week.