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. 2018 Apr 16;53(1):113–123. doi: 10.3892/ijo.2018.4373

Figure 3.

Figure 3

OOS induces cell death as well as cell cycle retardation in vitro. (A) GLC-8 cells were treated for 24 h with OOS (1:25), harvested and stained with Annexin V and PI to determine apoptotic cell death. (B) Representation of the viable and non-viable populations of the experiment shown in (A). (C) Quantification of the levels of apoptosis-related proteins after OOS treatment. GLC-8 cells were treated with OOS as in (A) and the levels of the indicated proteins were analyzed by conventional WB. (D) Cell cycle profile of the OOS-treated cells. GLC-8 cells were treated as in (A), harvested, fixed and the DNA content of the living cells was determined by PI staining. (E) OOS treatment caused a decrease in the percentage of cells in the S phase. To measure that population, GLC-8 cells were treated for 24 h with OOS at 1:25 and then incubated in the presence of BrDU for another 3 h. BrDU incorporation was measured using a colorimetric assay as described and normalized to that of the untreated controls. (F) Action of OOS on p27 and retinoblastoma protein (RB) protein levels. GLC-8 cells were treated for 24 h with OOS (1:25 and 1:50 dilution) and cell extracts were prepared to analyze p27 and RB by WB. GAPDH was used as a loading control. OOS, Ocoxin® oral solution; WB, western blotting; PI, propidium iodide.