Figure 4. Effects of PI3 Kinase Pathway Inhibitors on IL13-mediated Potentiation of Oxidative Stress-Induced Cell Death.

(A) MN9D cells grown in 96 well dishes were untreated or treated with 1, 5 or 10 ng/ml IL-13 alone, in the presence of 80 μM H₂O₂, 2.5 μM tBOOH or 10 μM LY294002 or as combination as indicated. Cell survival was measured after 24 hr with the MTT assay. Results are presented as the fold change in cell death as compared to H₂O₂ or tBOOH alone. The experiments were done in quadruplicate and repeated 5 times. (B) MN9D cells grown in 96 well dishes were untreated or treated with 1, 5 or 10 ng/ml IL-13 alone or in the presence of 80 μM H₂O₂, 2.5 μM tBOOH, 100 nM rapamycin or a combination as indicated. Cell survival was measured after 24 hr with the MTT assay. Results are presented as the fold change in cell death as compared to H₂O₂ or tBOOH alone. The experiments were done in quadruplicate and repeated 4 times. * p < 0.05; ** p < 0.01; *** p < 0.001 relative to no LY294002 or no rapamycin. (C) and (D) MN9D cells were untreated or treated with 10 ng/ml IL-13 alone or in the presence of 100 nM rapamycin, 80 μM H₂O₂, 2.5 μM tBOOH or a combination as indicated for 10 min, 2 hr and 4 hr. Cell lysates were prepared and equal amounts of protein were analyzed by SDS-PAGE and immunoblotting with antibodies to phospho and total Stat6. Representative Western blots are shown. Similar results were obtained in 3 independent experiments. (E) Human SH-SY5Y dopaminergic cells grown in 96 well dishes were untreated or treated with 1, 2.5 or 5 ng/ml IL-13 alone or in the presence of 40 μM H₂O₂, 2.5 μM tBOOH, 100 nM rapamycin or a combination as indicated. Cell survival was measured after 24 hr with the MTT assay. Results are presented as the fold change in cell death as compared to H₂O₂ or tBOOH alone. The experiments were done in quadruplicate and repeated 4 times. ** p < 0.01; *** p < 0.001 relative to no rapamycin.