Fig. 4.

The involvement of c-Jun N-terminal kinase (JNK) in irinotecan and etoposide-induced cell death. (A) Cells were transiently transfected with control or MAP kinase phosphatase-1 (MKP-1) specific siRNA (siMKP-1) for 48 hr, after which they were incubated in the presence or absence of irinotecan or etoposide (10 mg/L) for 15 min. Western blot analysis was used to determine the protein expression of phospho-JNK, total JNK, caspase-3, and cleaved caspase-3 in total cell lysates. (B) Cells were transiently transfected with control or siMKP-1 for 48 hr, after which they were incubated in the presence or absence of SP600125 (JNK inhibitor, 10 µmol/L) for 1 hr, and then treated in the presence or absence of irinotecan (10 mg/L) for an additional 24 hr. The cell viability was then evaluated by a WST-1 assay. The results are presented as the mean±SEM (n=4). Tukey's post-hoc test was applied to significant group effects in ANOVA, ^**p<0.01, ^***p<0.001. (C) Cell viability was measured by flow cytometry after tetramethylrhodamine ethyl ester (TMRE; 100 nmol/L) staining. The labeled numbers indicate live cells. Images are representative of three independent experiments. p-JNK, phosphorylated JNK; t-JNK, total JNK; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.