Figure 3

Combined treatment with FTS and MCP induce apoptosis and G1 cell cycle arrest. (a and b) For cell cycle analysis, ARO cells (5×10⁴) were plated and treated for 72 h either with FTS, MCP, or FTS+MCP. (a) Representative histograms of FACS analysis of treated cells by PI staining. (b) Quantification of the FACS analysis: amount of cells in G1, S and G2M and sub-G1 cell cycle phases of control (black bar), FTS-treated (gray bar), MCP-treated (upward diagonal bar) and FTS+MCP-treated ARO cells (white bar) are shown. Amount of cells in phases G1 and sub-G1 were significantly higher in FTS+MCP-treated cells compared with untreated cells. Amount of cells in phases S was significantly lower in FTS-treated cells, and FTS+MCP-treated cells compared with untreated cells (c and d) To examine whether combined treatment of FTS and MCP induces apoptosis, ARO cells (2.5×10⁵) were treated either with FTS, MCP, or FTS+MCP for 48 h. (c) Representative dot plot of FACS analysis of the treated cells by Annexin-V and PI staining. Numbers in the quadrants represent the percentage of ARO cells within each quadrant. (d) Quantification of the FACS analysis: Left side bars: Apoptotic ARO cells (high Annexin V, low PI) in untreated cells (control-black bar), FTS-treated (grey bar), MCP-treated (upward diagonal bar) and FTS+MCP treated ARO cells (white bar). Right side bars: Necrotic ARO cells (high Annexin V, high PI) in untreated cells (control-black bar), FTS-treated (grey bar), MCP-treated (upward diagonal bar) and FTS+MCP treated ARO cells (white bar). Apoptosis was significantly increased in cells treated with FTS+MCP compared with control. Results are presented as means±S.E.M.; one-way ANOVA analysis revealed significant differences between Annexin-V and PI staining; post hoc analysis was performed by the HSD test; *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001. (a and b: n=5; c and d: n=3); FTS, S-farnesylthiosalicylic acid; MCP, modified citrus pectin; PI, phosphatidylinositide.