Fig. 3. Modulation of IB1/JIP-1 regulates apoptosis in INS-1 cells. (A) Reduced IB1/JIP-1 content in INS-1 cells is associated with DNA laddering. DNA laddering in INS-1 cells producing different levels of IB1/JIP-1 shows a large increase in nucleosomal band staining in cells exposed to cytokines. Cells producing lower amounts of IB1 show an increase in the intensity of nucleosomal bands in the absence and in the presence of cytokines (IL-1ß, TNF- and IFN-). (B) Quantitative assessment shows a 400% increase in the intensity of nucleosomal bands in the presence of cytokines, whereas no change was observed in cells overproducing IB1. By contrast, cells decreasing their IB1 content show an increase in the DNA laddering pattern in the presence (500%) and the absence (400%) of cytokines. (C) Apoptosis rate in INS-1 cells is dependent on the IB1/JIP-1 content. Cells infected or not with the different adenovirus constructs were treated with cytokines or not for 2 days. Cells were finally stained with Hoechst 33342 and propidium iodide, and counted. The ratio between apoptotic and normal cells is indicated. Columns represent mean ± s.e.m. of three independent experiments performed in triplicate. **, °° or P<0.01; ***P<0.001. (D) To confirm the protective effect of IB1 in INS-1 cells, western blots of cleaved caspase were carried out. Cleaved caspase was increased in cells infected with control adenovirus (Ad-GFP) in the presence of cytokines. By contrast, overproduction of IB1 decreased the presence of caspase 3 cleavage.