Figure 1

Induced P-gp-mediated drug resistance of endothelial cells. P-gp cell surface expression was analyzed with flow cytometry in HMEC (Panel a) or HUVEC cells (Panel b). Endothelial cells expressed P-gp after the induction by Dox treatment. Parental (thick black line), HMECd1 and HUVECd3 (thin black line), and HMECd2 (−−) cells were incubated with 10 μg/ml 4E3. Incubation with control IgG2a gave similar histograms for the three cell lines (filled grey histogram). Histograms are representative of four separate experiments. Panel c: The western blot of P-gp levels in HMECd1, HMECd2 and their parental cells. The data for the ratio were obtained with three repeated blots. *: p < 0.05 in comparison with the controls. Panel d: The western blot of ABCG2 levels in these cells. The data for the ratio were obtained with three repeated blots. *: p < 0.05 versus the controls. Panel e: qPCR (primer Hs01067802_m1) results of P-gp mRNA levels in treated or nontreated HMEC-1, HMECd1, and HMECd2. Cyclosporine A (C), Verapamil (V), Fumitremorgin C (F), and Diethylstibesterol (D) were used to treat the cells. The results were obtained from three independent experiments. *: p < 0.05 versus the nontreated cells. Panel f: qPCR (Hs01053790_m1) results of ABCG2 mRNA levels in treated or nontreated HMEC-1, HMECd1, and HMECd2. Cyclosporine A (C), Verapamil (V), Fumitremorgin C (F), and Diethylstibesterol (D) were used to treat the cells. The results were obtained from three independent experiments. *: p < 0.05 versus the nontreated cells. Panel g: Correlation between P-gp surface expression and its efflux function. During the establishment of resistant HMEC cell lines, the P-gp surface expression and the Rho efflux were regularly analyzed by flow cytometry, as shown in Figure 2a-d (R² = 0.9301).