Fig. 5. Repression of p65 activation by IL-4/STAT6. HeLa cells transiently transfected with p65-dsRed and either EGFP or EGFP-STAT6 were pretreated with 10 ng ml^-1 IL-4 or medium for 40 minutes. Subsequent stimulation with 10 ng ml^-1 TNF was followed by confocal microscopy for 80 minutes. (A) Fluorescence images of cells expressing p65-dsRed (left; red fluorescence) and EGFP-STAT6 (right; green fluorescence) treated with IL-4 and TNF, showing marked inhibition of p65-dsRed translocation compared with cells expressing EGFP treated with TNF (Fig. 2B). Positions of nuclei are highlighted in grey in the first EGFP-STAT6 image. Times after TNF treatment are shown in minutes. (B) Quantification of p65-dsRed translocation inhibition by activation and expression of EGFP-STAT6. p65-dsRed localization was analysed and quantified as described in Fig. 3. Each treatment was performed in triplicate with a minimum of 16 cells per experiment. Data plotted represent mean nuclear:cytoplasmic ratio per cell±s.d._(n–1) for each experiment.

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