1. Elicited and steady-state transport of skin self-antigens in PYNOD-KO mice.
Elicited and steady-state transport of skin self-antigens in PYNOD-KO mice. (A) Mice received an s.c. injection of 20 μl of Fluoresbrite and control PBS in the left and right hind footpads, respectively. Eight hours later, popliteal LN cells were isolated, stained with allophycocyanin-conjugated anti-CD11c mAb, and examined by flow cytometry. The left panels indicate Fluoresbrite incorporation and CD11c expression profiles. Numbers in rectangular areas indicate the percentage of CD11+ cells. The right panels indicate the Fluoresbrite fluorescence intensity of cells in the rectangular areas at left. Numbers indicate the percentage of Fluoresbrite+ cells in CD11c+ cells. Representative flow cytometry profiles from three independent experiments using nine total WT and nine total PYNOD-KO mice are shown. The upper right graph shows the average of two independent experiments using six total WT and six total PYNOD-KO mice. (B) Mice were intranasally administrated with 1% Fluoresbrite containing 1 μg of LPS. Eighteen hours later, single cells isolated from lungs and mediastinal LNs were stained with biotin-conjugated anti-mouse CD11c Ab and allophycocyanin-conjugated streptavidin. Fluoresbrite+CD11c+ cells were detected by flow cytometry. The two-dimensional profiles indicate gating strategy for CD11c+ cells, and the numbers in the gated region indicate the percentage of CD11c+ cells in total cells. The histograms indicate Fluoresbrite fluorescence profiles of CD11c+ cells, and the numbers indicate the percentage of Fluoresbrite+ cells in CD11c+ cells. The upper right graph shows the cumulative data from four independent experiments using 10 total WT and 6 total PYNOD-KO mice. (C) PYNOD-KO mice were crossed twice with KitL-Tg mice to generate epidermis-pigmented PYNOD-KO/KitL-Tg mice. Upper panels show skin regional inguinal LNs and mesenteric LNs of 7-wk-old PYNOD+/−/KitL-Tg mice and PYNOD-KO/KitL-Tg mice. Lower graphs show the amount of melanin in skin regional LNs (sum of submandibular, axillary, brachial, inguinal, popliteal LNs taken from one side of the body, left) and the amount of melanin in ear skin (15.7 mm2, right). Representative findings of LNs from five independent experiments are shown. The amount of melanin represents pooled data from five independent experiments. Mesenteric LNs (upper panels) and the amount of melanin in non-Tg mice [Tg(−), lower graphs] are shown as controls. Numbers in each column indicate the number of mice sampled. (A–C) No significant differences were observed between WT and PYNOD-KO mice. ***p > 0.005, ****p > n.s., not significant.
Shinsuke Nakajima et al. ImmunoHorizons 2018;2:
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