FIGURE 3.

Inhibition of sAPPα enhances the cytotoxic effect of gemcitabine on pancreatic cancer cells. A, 10,000 CD18 cells were plated in 24-well culture dishes and treated with 5 μm batimastat, 100 ng/ml gemcitabine, or a combination of both for 24 h. Cytotoxicity assay using MTT reagent was performed and samples analyzed at A₅₄₀. B, 10,000 MiaPaCa2 cells were plated in 24-well culture dishes and treated with the indicated drugs, and the extent of cytotoxicity was determined using MTT reagent at A₅₄₀. C, CD18 cells were plated in 24-well plates and treated with the indicated drugs for 24 h and cytotoxicity was measured using the MTT reagent at A₅₄₀. Each experiment was performed more than three times and (*) indicates significant difference with p value less than 0.05 from an unpaired two-tailed Student's t test. D, 10,000 CD18 cells were plated in 24-well plates and treated with 5 μm batimastat, 5 μm GI254023X, 100 ng/ml gemcitabine, or combinations for 24 h. MTT reagent was used to determine cytotoxicity at A₅₄₀. E, 10,000 MiaPaCa2 cells were plated in 24-well plates and treated with 5 μm batimastat, 5 μm GI254023X, 100 ng/ml gemcitabine or combinations for 24 h. MTT reagent was used to determine cytotoxicity at A₅₄₀. F, CD18 and MiaPaCa2 cells were treated with 5 μm batimastat, 100 ng/ml gemcitabine or a combination of the drugs for 24 h, and cell lysates collected for analysis of PARP cleavage. The same blot was reprobed for actin, which served as a loading control. G, cleavage of PARP was quantified from three independent experiments and normalized to actin using the ImageJ software as represented by the bar graph. Each of the cytotoxicity and quantification experiments were performed more than three times, and (*)(#)(●)(■) indicate significant difference with p value less than 0.05 from an unpaired two-tailed Student's t test.