Fig. 3

High insulin level accelerates palmitic acid-induced apoptosis in endometrial cancer cells. A, B Apoptosis of Ishikawa and AN3CA cells detecting by Annexin V-FITC/PI labelling flow cytometry after saturated fatty acid and/or insulin treatment. The proportion of apoptotic cells was detected after treatment with 0.5 mM palmitic acid (PA) or 0.5 mM stearic acid (SA) for 12 h in Ishikawa cells and treatment for 18 h in AN3CA cells. The proportion of apoptotic cells were also compared after treatment with 0.5 mM PA alone or 0.5 mM PA plus 100 nM insulin, 0.5 mM SA alone or 0.5 mM SA plus 100 nM insulin. Saturated fatty acid (PA and SA) significantly induced apoptosis in EC cells, and the pro-apoptotic effect could be enhanced by excessive insulin. C, D Analysis of Annexin-V-positive cell ratio *P < 0.05. E–G Immunoblot of caspase-3, cleaved caspase-3, Bcl-2, Bax and GAPDH after saturated fatty acid and/or insulin treatment. Ishikawa cells are treated with 0.5 mM PA or 0.5 mM SA alone, and PA or SA plus 100 mM insulin for 12 h. Histogram plot shows cleaved caspase-3/caspase-3 and Bax/Bcl-2 intensity relative to vehicle *P < 0.05. H Apoptotic effect of insulin on PA or SA-treated (0.5 mM) Ishikawa cells, determined by TUNEL assay (red)