Fig. 3: Nociceptor-deficient mice present higher levels of neutrophil infiltration in the skin after HSV-1 infection.

Control DTA and Nav_1.8-DTA mice were infected with HSV-OVA-TK⁻ by flank scarification. Immune responses in the skin were analysed 6 days pi by flow cytometry. a tSNE analysis of CD45⁺ cells with the CD11c, MHC-II, CD206, CD11b, Ly6G, CD24, c-kit, CD4 and γδ TCR markers. The colours in the expression-level heatmaps (right panel) represent the median intensity values for each marker. b Flow cytometry gating strategy used to identify immune cell subsets: the CD45⁺ cell population was selected after the exclusion of doublets and dead cells. DCs were gated as CD11c^high MHC-II⁺ cells and split into subsets on the basis of their expression of CD24 and CD11b. cDC1 was defined as CD24⁺ CD11b⁻ DC, cDC2 as CD11b⁺ CD24⁻ DC and Langerhans cells (LC) as CD24⁺ CD11b⁺ DC. Neutrophils were gated as CD11b⁺ Ly6G⁺ cells. Eosinophils were then gated as CD11b⁺ CD24⁺ cells. Monocytes and macrophages were defined as CD11b⁺ CD24⁻ cells and subdivided into macrophages (CD11b⁺ CD24⁻ CD206⁺ CD64⁺) and monocytes (CD11b⁺ CD24⁻ CD206⁻ Ly6C^high/low). Finally, within the non-DC subset, lymphoid cells were gated as CD11b⁻ γδ TCR⁺ cells and subdivided into CD4⁺ T cells (CD4⁺ γδ TCR⁻), γδ T cells (CD4⁻ γδ TCR⁺) and other lymphoid cells (CD4⁻ γδ TCR⁻). c Distribution of the different immune cell subsets in the tSNE analysis: neutrophils (CD11b⁺ Ly6G⁺, in red), dendritic cells (CD11c^high MHC-II⁺, in green), monocytes (CD11b⁺ Ly6G⁻ CD24⁻ CD206⁻, in pink), macrophages (CD11b⁺ Ly6G⁻ CD24⁻ CD206⁺, in blue), eosinophils (CD11b⁺ Ly6G⁻ CD24+, in orange), mast cells (CD11b⁺ c-kit⁺, in black), CD4⁺ T cells (CD11b⁻ TCRγδ⁻ CD4⁺, in cyan), γδ T cells (CD11b⁻ TCRγδ⁺, in dark yellow) and other lymphoid cells (CD11b⁻ TCRγδ⁻ CD4⁻, in grey). d Relative clustering of skin immune cells from the indicated mouse strains (control DTA or Nav_1.8-DTA) used for the tSNE analysis. e Percentages of neutrophils among CD45⁺ skin cells from DTA and Nav_1.8-DTA mice 6 days pi. f Absolute numbers of immune cells subsets in the skin of DTA and Nav_1.8-DTA mice 6 days pi. The data presented are from three independent experiments (n = 8–11 mice per group) and are shown as mean ± SEM; P values were obtained by using a Mann–Whitney test (two-tailed).