Fig 2: Conventional dendritic cells accumulate in the dural meninges during chronic brain infection and sample CSF-derived protein.Mice were infected with 10 cysts of the ME49 strain of T. gondii intraperitoneally (i.p.) and analyzed at 6 weeks post-infection. (a) Confocal microscopy was used to image whole-mount dural meninges dissected from the skullcaps of naïve or chronically infected CD11cYFP reporter mice. Images are representative of three independent experiments. Scale bar, 2000 µm. (b) Representative images of CD11c+ cells (yellow) in the region surrounding the dural sinuses. Scale bar, 150 µm. (c) Quantification of area coverage by CD11c+ cells in the region surrounding the dural sinuses. Data compiled from three experiments (n = 10-11 mice per group). (d-e) Representative images (d) and quantification (e) of CD11c+ cells (yellow) present within LYVE1+ meningeal lymphatic vessels (white). Data compiled from three experiments (n = 10-11 mice per group). Scale bar, 50 µm. (f) Quantification by spectral flow cytometry of total cDC1 number (CD45+Lin-CD11chiMHC IIhiCD64-CD26+XCR1+SIRPa-) in the dural meninges of naïve, acutely infected, or chronically infected C57BL/6 mice. Data compiled from two experiments (n = 7 mice per group). (g) Quantification by spectral flow cytometry of total cDC2 number (CD45+Lin-CD11chiMHC IIhiCD64-CD26+XCR1-SIRPa+) in the dural meninges of naive, acutely infected, or chronically infected C57BL/6 mice. Data compiled from two experiments (n = 7 mice per group). (h) Quantification of T. gondii gDNA in brain and dural meninges by real-time PCR. Data compiled from two experiments (n = 3-8 mice per group). (i–k) Quantification of the geometric mean fluorescence intensity (MFI) of CD80 (i), CD86 (j), and MHC class II (k) expressed by cDC1s and cDC2s in the dural meninges. Data compiled from two experiments (n = 7 mice per group). (l) Schematic diagram illustrating intra-cisterna magna (i.c.m.) injection of DQ-OVA into the CSF. (m) Representative images of MHC class II-expressing antigen-presenting cells (magenta) co-labeling with fluorescent DQ-OVA cleavage products (green) in the vicinity of (closed arrows) or present within (open arrows) meningeal lymphatic vessels (LYVE1+, white). Three independent experiments were performed. Scale bar, 50 µm. Data are represented as mean values ± s.e.m. For c and e, statistical significance was measured using randomized block ANOVA (two-way), with p<0.001 (***). For (f and g) statistical significance was measured using a one-way ANOVA with post-hoc Tukey multiple comparison testing, with p<0.05 (*), p<0.01 (**), and p<0.001 (***). For (h–k) statistical significance was measured using a two-way ANOVA, with Tukey’s multiple comparison test to assess differences across timepoints [p<0.01 (##) and p<0.001 (###)] and Sidak’s multiple comparison test to assess differences between tissues (h) or cell type (i–k) [ns = not significant, p<0.05 (*), p<0.01 (**), and p<0.001 (***)].