The activity of the aryl hydrocarbon receptor in T cells tunes the gut microenvironment to sustain autoimmunity and neuroinflammation
Fig 2
Ahr knockout in CD4+ cells does not influence T cell differentiation.
(A) Naïve CD4+ T cells were isolated from cohoused Cd4creAhrfl/fl and littermate control animals and treated to promote differentiation. Flow cytometry gating strategy to measure percent of cells expressing the transcription factors specific to each cell type is shown. (B) Representative histogram and quantification of Tbet+ cells after in vitro differentiation of naïve CD4+ cells to TH1 cell type, (n = 6 biological replicates/group; N = 2 experiments; unpaired t test [p = 0.8715]) (C) GATA3 staining of TH2 differentiated cells, (n = 6 biological replicates/group; N = 2 experiments; unpaired t test [p = 0.9356]), (D) RORγt staining of TH17 cells (n = 8 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.3989]) (E), and FoxP3 staining for Tregs (n = 8 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.4078]). (F) ELISA analysis of culture supernatant after 24 h of anti-CD3 stimulation in TH1cells (n = 9 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.3924]) (G) TH2 cells (n = 8 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.5917]) (H) TH17 cells (n = 7–8 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.4013]) (I) or Treg cells. (n = 8 biological replicates/group; N = 3 experiments; unpaired t test [p = 0.2278]) (J) Gating strategy for (K) flow cytometry of single-cell suspensions of Peyer’s patches from cohoused Cd4creAhrfl/fl and Ahrfl/fl mice. (Multiple t tests with Welch correction [p = 0.070748, 0.081520, 0.325104, 0.080798]) (L) Analysis of cell compositions from mesenteric lymph nodes (multiple t tests with Welch correction [p = 0.127038, 0.179526, 0.444670, 0.118472]) (M) and the inguinal lymph nodes from cohoused Cd4creAhrfl/fl and Ahrfl/fl mice (multiple t tests with Welch correction [p = 0.381964, 0.224164, 0.664161, 0.465348]). (N–S) Luminex assay on whole spinal cord homogenate of separately housed Cd4creAhrfl/fl and Ahrfl/fl mice at the peak of disease (day 16 after immunization; n = 8 mice/group; N = 2 experiments; unpaired t test), (N) [p = 0.3087], (O) [p = 0.4580], (P) [p = 0.8313], (Q) [p = 0.4941], (R) [p = 0.0215], (S) [p = 0.9333]. Error bars represent standard error from the mean. Raw data can be found in Supporting information (S1 Data).