Figure 2.

L. johnsonii-derived nanovesicles promote AHR nuclear translocation in βlox5 cells. Cells were incubated with NV10 (A–F) or 100 nM TCDD (G–L) and visualized by confocal microscopy. Cells were fixed and incubated with rabbit anti-AHR antibody tagged with PE-Texas red (red pixels) and the nucleus was stained with DAPI (blue pixels) as described in the methods section. (A, G) Time 0 (2D image) and (B, H) 3D z-stack image showing both stains or (C, I) AHR only channel. (D, J) 2D image, (E, K) 3D z-stack image showing both stains, (F, L) AHR only channel after 45 minutes of incubation. Yellow arrows indicate the localization of AHR in the cytoplasm and around the nuclei at time 0 and inside the nuclei after 45 minutes of incubation. (M–P) Heatmap showing the colocalization of both stains (DAPI and Texas red) at time 0 (M, O) and 45 minutes (N, P), when cells were treated with NVs (M, N) or TCDD (O, P). The upper right quadrant (number 2) shows the pixels that are stained with both stains simultaneously. The figures shown are representative of at least three biological triplicates. TCDD denotes 2,3,7,8–tetrachlorodibenzo-p-dioxin; NV10 denotes nanovesicles from L. johnsonii at 10¹⁰ particles/ml.