Cav-1 has a differential location in fibroblasts lacking AhR. (A) T-FGM AhR+/+ fibroblasts transfected with a siRNA against AhR (si-AhR) or with an scrambled sequence, and T-FGM AhR?/? fibroblasts transfected with a pcDNA-AhR or with an empty expression vector (E.V.) were analyzed for Cav-1 expression by immunofluorescence using an Alexa 488-conjugated secondary antibody. (B) Quantification of the cellular distribution of Cav-1 in T-FGM cells. Cav-1 was considered membranal (dashed bars) when present within a 2 μm distance from the cell border or cytosolic (gray bars) when expressed from 2 μm up to the cell nucleus. Measurements were taken by triplicate in two cultures of each genotype. (C) The ability of the pcDNA AhR expression construct to rescue receptor expression in AhR?/? fibroblasts was determined by immunoblotting. (D) Primary dermal fibroblasts obtained from the skin of AhR+/+ and AhR?/? newborn mice were also analyzed as above. Regions of interest (ROI) were selected (red line in upper panels) and the levels of fluorescence analyzed by using the "ROI profiles" tool of the Fluoview software. The intensity per pixel was plotted for Cav-1 (green). Cav-1 was detected using a Fluoview F1000 confocal microscope. The nuclear signal is represented in dark blue (DAPI staining). The experiments were done at least three times in independent T-FGM and dermal fibroblasts cultures. Data are shown as mean ± s.d.