Figure 5

Confocal analysis of ISBn treated growing Caco-2 cells stained for iNOS, hsp27, and vimentin. Panel A: Subcellular co-localization of iNOS (red) and vimentin filaments (green) in Caco-2 cells without ISBn (ctr) and with Pol, Res, and Pol-Res combination treatment. The cells were incubated with anti-vimentin, anti-HSP27 and anti-iNOS antibodies. Images were obtained by confocal microscopy. Panel B: Subcellular co-localization of HSP-27 (red) and vimentin filaments (green) in Caco-2 cells without ISBn (ctr) and with Pol, Res, and Pol-Res combination treatment The cells were incubated with anti-vimentin, anti-HSP27 and anti-iNOS antibodies. Images were obtained by confocal microscopy. Panel C: The expression levels of the proteins are reported as intensities quantified with Image J software. Panel D: Caspase-3 activity on growing and differentiated Caco-2 cells without ISBn (ctr) and with Pol, Res, and Pol-Res combination treatment .Caspase-3 activity was determined by incubating whole-cell extracts with 40 μM caspase-3 substrate and measuring production of hydrolysed 7-amido-4-methyl-coumarin (AMC) groups using a multi-label plate reader. The results are representative of four separate experiments; summary data shown are means ± SEM.