Fig. 3
Comparison of rbNPs with other pdNPs, DOXIL® or rb-sup. (A) The number of colon26 cells 24 h after the addition of grape, ginger, and lemon NPs, and rbNPs at varying concentrations. Colon26 cells are incubated with 0.1−10 × 109 NPs/mL, and the cell number is measured at 24 h using CCK8 assay. Results are expressed as the mean ± SD of four samples. #p < 0.01 vs. NT group. (B) The number of colon26 cells 24 h after the addition of rbNPs or DOXIL®. Colon26 cells are incubated with 0.1−10 × 109 NPs/mL of rbNP or DOXIL®, and the cell number is measured as described earlier. Results are expressed as the mean ± SD of three samples. #p < 0.01 vs. DOXIL®. (C) The number of colon26 cells 24 h after addition of rbNPs and rb-sup. Colon26 cells are incubated with 1,000 µg/mL of rbNP or rb-sup, and the cell number is measured at 24 h using CCK8 assay. Results are expressed as the mean ± SD of three samples. *p < 0.05. #p < 0.01. (D) Western blot analysis of β-catenin, cyclin D1, and β-actin in colon26 cells. Colon26 cells are treated with 1,000 µg/mL rbNP or rb-sup for 24 h, and the cellular proteins are extracted for the analysis The bands of each protein are visualized using Invitrogen iBright Imaging Systems. (E) DNA fragmentation of colon26 cells after addition of rbNPs, rb-sup, or actinomycin D. Colon26 cells are incubated with 1 µM actinomycin D or 1,000 µg/mL rbNP or rb-sup. The DNA of the cells is extracted and subjected to 3% agarose gel electrophoresis, followed by visualization using Invitrogen iBright Imaging Systems. (F) Confocal images of colon26 cells stained with DAPI. Colon26 cells are incubated with 1,000 µg/mL rb-sup or rbNP for 24 h at 37 °C, fixed with paraformaldehyde, and the nuclei of the cells are stained with DAPI. Scale bars indicate 50 μm (low magnification) and 10 μm (high magnification). White arrows indicate chromatin condensation