Deoxycholic acid induces reactive oxygen species accumulation and promotes colorectal cancer cell apoptosis through the CaMKII-Ca2+ pathway

Figure 1 Deoxycholic acid promotes apoptosis in SW480 and DLD-1 cells. A: Flow cytometry results and statistical plots for SW480 and DLD-1 cells; B: Western blot results and statistical plots of apoptosis-related proteins in SW480 and DLD-1 cells, and a histogram of the Cleaved-PARP/PARP ratio analysis. Control represents DMSO treatment, and the experimental group was treated with 100 μM deoxycholic acid for 24 hours. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. PE: Phycoerythrin; DCA: Deoxycholic acid.

Figure 2 Deoxycholic acid promotes intracellular and mitochondrial reactive oxygen species accumulation in SW480 and DLD-1 cells. A: Flow cytometry results and statistics of intracellular reactive oxygen species (ROS) in SW480 and DLD-1 cells; B: Confocal microscopy results of intra-mitochondrial ROS in SW480 and DLD-1 cells. ^aP < 0.01. FITC: Fluorescein isothiocyanate; DCA: Deoxycholic acid.

Figure 3 Deoxycholic acid promotes the increase of intracellular and mitochondrial calcium ion content in SW480 and DLD-1 cells. A and B: Flow cytometry results and statistics of intracellular calcium content in SW480 and DLD-1 cells, respectively; C and D: Flow cytometry results and statistics of intracellular calcium content in mitochondria of SW480 and DLD-1 cells, respectively; E: Confocal microscopy results of intracellular calcium content in mitochondria of SW480 and DLD-1 cells; F: Western blot results and statistics of p-CaMKII protein in SW480 and DLD-1 cells. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. PE: Phycoerythrin; FITC: Fluorescein isothiocyanate; DCA: Deoxycholic acid.

Figure 4 Effect of intracellular calcium chelator BAPTA-AM on intracellular and mitochondrial Ca²⁺ and reactive oxygen species. A: Flow cytometry results of intracellular calcium content in SW480 cells; B: Flow cytometry results of intracellular calcium content in mitochondria of SW480 cells; C and D: Flow cytometry results and statistics of the calcium ion content in the mitochondria of SW480 cells; E and F: Flow cytometry results and statistical graphs of reactive oxygen species (ROS) content in mitochondria of SW480 cells. The effect of deoxycholic acid on the mitochondrial Ca²⁺ and ROS in SW480 cells after chelating intracellular calcium ions at 0, 1, and 2.5 μM in three replicates of BAPTA-AM treatment, respectively. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. PE: Phycoerythrin; FITC: Fluorescein isothiocyanate; DCA: Deoxycholic acid.

Figure 5 Effect of deoxycholic acid on apoptosis of SW480 cells after chelation of intracellular calcium ions. The graphs show the flow cytometry results and statistical plots of SW480 cell apoptosis. ^aP < 0.05, ^bP < 0.01. PE: Phycoerythrin; DCA: Deoxycholic acid.