672416.fig.002a
672416.fig.002b
Figure 2: Effect of piceatannol on the invasion of AH109A hepatoma cells (A), reactive oxygen species- (ROS-) potentiated invasive activity and intracellular peroxide levels (B). (A) Piceatannol was dissolved in ethanol. The piceatannol solution was added to the culture medium at a final ethanol concentration of 0.2%. Piceatannol was suspended in a 0.3% carboxymethyl cellulose sodium salt (CMC) aqueous solution at concentrations of 0.25, 0.5, and 1 mg/mL. Piceatannol suspension or 0.3% CMC alone (vehicle control) (1 mL/100 g body weight) was intubated to male Donryu rats which had been fasted overnight, and blood was collected 2 h after oral administration. The invasive activity was determined by AH109A-M cell coculture method in vitro (A(a)) and ex vivo (A(b)). Each value represents the mean ± SEM of ten areas. Values not sharing a common letter are significantly different at 𝑃 < 0 . 0 5 . (B) AH109A cells were cultured for 4 h in the absence or presence of 10 μM piceatannol with or without 25 μM hydrogen peroxide (H2O2). AH109A cells were then washed once with 10% CS/MEM and seeded on the M-cell monolayer in 10% CS/MEM without piceatannol and ROS. After cultured for 24 h, invaded cells and colonies underneath M-cells were counted with a phase-contrast microscope. Each value represents the mean ± SEM of ten areas. Values not sharing a common letter are significantly different at 𝑃 < 0 . 0 5 (B(a)). To evaluate the effect of piceatannol on intracellular peroxide levels, AH109A cells were treated with 100 μM H2O2 as ROS source. After ROS treatment for 1 hour, DCFH-DA was added, incubated for 20 minutes, and analyzed with a flow cytometer. Basal and ROS-potentiated intracellular peroxide levels are indicated as solid and dotted lines, respectively. Data are from representative experiment repeated three times with similar results (B(b)).