Fig. 3
Effects of AsIII and Tetra, alone or in combination, on the cell cycle profiling and the expression level of cell cycle related-proteins in MDA-MB-231 cells. a–d After treatment with various concentrations of AsIII (5, 10 and 15 µM), and Tetra (3.5, 4 and 4.5 µg/ml), alone or in combination (5 µM AsIII + 3.5 µg/ml Tetra, 10 µM AsIII + 4 µg/ml Tetra), for 48 h, cell cycle profiling was performed by FACSCanto flow cytometer as described under “Materials and methods”. Analyzed data and profiles for each G0/G1 and G2/M phase using Diva software and ModFit LT™ ver. 3.0. are shown in the gray area. Cells at S phase are shown as shaded area. A representative FACS histogram from three separate experiments is shown. Significant difference between control and treatment with AsIII and Tetra, alone or in combination, are shown (*p < 0.05, ‡p < 0.01, §p < 0.001, †p < 0.0001 vs. control, #p < 0.05, $p < 0.01 vs. AsIII alone; &p < 0.05, ¶p < 0.01 vs. Tetra alone). e Representative image of the expression profile of each protein is shown from three independent experiments. The densitometry of protein bands was analyzed using a program, NIH ImageJ 1.52a. The values under each image represent the ratios between each key molecule and β-actin protein expression levels, which were further compared with those of control group (untreated cells). As, AsIII; Tetra, tetrandrine. Since enough cells cannot be collected in the group treated with 15 µM AsIII in combination with 4.5 µg/ml Tetra due to its strong cytotoxicity, cell cycle and western blot analysis were not conducted