Fig. 4.
Effect of OATP1B3 and OATP1B1 expression on the intracellular AR-67 amounts in HeLa-pIRES/OATP1B3 and RKO-pIRES/OATP1B1 cells. (A) HeLa-pIRES/OATP1B3 and (B) RKO-pIRES/OATP1B1 cells were incubated with 0.5, 1.0, 1.5, 2.0, and 3.0 µM of AR67 carboxylate for 1 minute to study the dose-dependent uptake of the carboxylate form. The estimated Km values for OATP1B3 and OATP1B1 were 0.32 (0.0–0.8316) μM and 2.58 (0.0–6.253) μM, respectively. They were obtained by using the Michaelis-Menten equation to fit the transporter-mediated uptake of AR-67, as described in the Materials and Methods section, and are reported as the mean (95% confidence interval). (C) HeLa-pIRES/OATP1B3 and (D) RKO-pIRES/OATP1B1 cells were incubated with 1 μM of AR-67 carboxylate for 5 and 10 minutes, respectively (open bars). The inhibitory effect of BSP (50 μM) was studied by pretreating for 10 minutes before exposure to AR-67 carboxylate as described previously for all cell lines (solid bars). Intracellular AR-67 was quantified using HPLC, as determined in the Materials and Methods section. Data are represented as the mean (n = 3) ± S.D. Statistical analysis was performed using unpaired t test, statistical significance for **P < 0.01; ***P < 0.001.