Fig. 7
From: Partial loss of Sorting Nexin 27 resembles age- and Down syndrome-associated T cell dysfunctions
SNX27 partial deletion impairs MTOC polarization in CD4+ T cells but not in CTL. A Isolated naïve CD4+ T cells from SNX27fl/fl and CD4-Cre-SNX27fl/+ mice were set to interact with CMAC-labeled (blue), anti-CD3-coated (2.5 µg /ml) P815 cells for 15 min. Cells were then fixed with 2% PFA and co-stained for F-actin (phalloidin-A488) and MTOC (anti-pericentrin) followed by anti-rabbit IgG-Cy5. Confocal optical sections were acquired at 0.2 µm intervals. Representative confocal sections are depicted. Scale bar = 5 µm. Quantification of (B) MTOC polarity index and (C) percentage of cell conjugates with a polarized MTOC is shown. The polarity index is calculated as the ratio between the distance from the MTOC to the IS and the distance from the IS to the distal pole of the cell. When the relative distance was < 0.25 µm, MTOC was considered to be polarized. Ratio values are displayed as dot plots, with each dot representing an individual cell. Data shown as mean ± SEM; ****p < 0.0001; unpaired t test; n = 71 SNX27fl/fl cells, 95 CD4-Cre-SNX27fl/+ cells in (B), and n = 5 mice in (C). D Ex vivo-differentiated CTL from SNX27fl/fl and CD4-Cre-SNX27fl/+ mice were incubated with CMAC-labeled (blue), anti-CD3 coated (2.5 µg /ml) P815 target cells during 15 min and stained as described in (A). Quantification of (E) MTOC polarity index and (F) percentage of cell conjugates with a polarized MTOC is shown. Data shown as mean ± SEM; unpaired t-test; n = 107 SNX27fl/fl cells, 104 CD4-Cre-SNX27fl/+ cells in (E), and n = 5 mice in (F). SNX Sorting nexin, CTL Cytotoxic T lymphocytes, SEM Standard error of the mean