Fig 1: Activation markers CD69 and CD44 are increased under metabolic stress on hepatic CD4 T cells, but not CD8 T cells, while this activation is inhibited in PD1+ CD4 T cells. (A) CD25 expression on blood and liver CD8 T cells under SC or WD for 24 weeks. (n = 5). (B) CD69 expression on blood and on hepatic CD8 T cells after 24 weeks treatment with SC or WD. (n = 5), (***p < 0.001). (C) CD69 expression on blood and hepatic CD4 T cells after 24 weeks SC and WD feeding. (n = 5, *p < 0.05). (D) CD44 expression on blood and intrahepatic CD4 T cells under SC or WD treatment for 24 weeks. (n = 5, **p < 0.01 and ***p < 0.001). (E) CD25 expression on blood and hepatic CD4 T cells under SC or WD. (n = 5), (*p < 0.05 and ***p < 0.001). (F) CD44 and CD69 expression on PD1+ or PD1- CD4 T cells after SC or WD feeding. (n = 5), (*p < 0.05; **p < 0.01; and ***p < 0.001).
Fig 2: Tagln2-/- DCs do not optimally support T cell activation in vivo. a Schematic diagram of the experimental setup for this figure (left). WT or Tagln2-/- DCs were pulsed with pOVA (323–339), stained with CMFDA-green, and injected into the footpad of OTII recipient mice. A representative photograph of excised popliteal LNs (right). Scale bar, 1 µm. b Flow cytometric plots show activation of migrating DCs in vivo at 24 h post-injection. DCs were isolated from the popliteal LNs of recipient mice and stained with anti-CD40 or CD80. The percent of CD11c+ cells in the draining LNs is presented as a bar graph. c DC-mediated T cell activation in vivo. At 24 h post-injection of DCs, OTII CD4+ T cells were isolated from the popliteal LNs of recipient mice and stained with anti-CD69 or CD25. All data represent the mean of three experiments ± SEM. NS, not significant. *P < 0.01
Fig 3: Dynamic changes of lung NK cells in the development of asthma. (A) NK cells were identified as a CD3- DX5+ cell population in a lymphocyte morphology gate on the basis of FSC and side scatter characteristics (violet box, left panel). (B) NK cells as a percentage of total lymphocytes were enumerated. (C) Absolute NK cell numbers obtained from lungs of BALB/c mice. (D) Representative flow cytometry plots showing the expression of CD69 on CD3- DX5+ NK cells from the lungs obtained at day 18. (E) Representative flow cytometry plots showing the expression of IFN-?, IL-5 and IL-13 on CD3-DX5+ NK cells from the lungs obtained at day 18 (n=6). ***P<0.001, vs. control group. NK, natural killer; FSC, forward scatter; IFN, interferon; IL, interleukin; NS, not significant.
Fig 4: CpG immunization induces APC maturation in the DLN. BALB/cByJ mice were immunized intranasally. (A) Representative FACS plots from the DLN at day 1 post immunization. Innate immune populations were enumerated at day 1 (B) and day 3 (E) by flow cytometry. CD8+ DCs and CD103+ DCs were enumerated at day 1 (C) and day 3 (F). Gated innate immune populations were assessed for their surface expression of the costimulatory molecule CD86 (D,G). CD69 upregulation on the surface of CD4 and CD8 T cells in the draining lymph nodes 3 days after immunization (H). Graphs represent two separate experiments (n = 3–5 mice per group) with data given in mean percentage ± SD. *p < 0.05 **p < 0.01, ***p < 0.001.
Fig 5: CpG immunization induces APC maturation in the lung. BALB/cByJ mice were immunized intranasally. (A) Representative FACS plots from the lung at day 1 show (i) interstitial macrophages (F4/80+), (ii) alveolar macrophages (F4/80+ CD11c+), and (iii) conventional DCs (CD11c+). DCs, interstitial macrophages, and alveolar macrophage populations were enumerated at day 1 (B) and day 3 (F) in the lungs. Innate immune cell populations were assessed for their surface expression of the costimulatory molecule CD86 by measuring the median fluorescence intensity (MFI) (C,G). The influx of CD103+ dendritic cells (D,H) and TLR9+ (E,I) cells was also enumerated after immunization. CD69 upregulation on CD4 and CD8 T cells in the lungs 3 days after immunization (J). Graphs represent two separate experiments (n = 3–5 mice per group) with data given in mean percentage ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.
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