Fig 5. The down-regulation of MRP2 expression by shRNA renders Caco-2 cells more sensitive to etoposide.

(A) Western blot analysis was used to assess the down-regulation of MRP2 protein expression in the presence of two shRNAs directed against the protein. Down-regulation was achieved by lentiviral infection of Caco-2 cells as previously described [31]. shRNA directed against MRP2 (sh305 and sh307) abolished protein expression by 90–100% comparatively to cells expressing a non-targeting shRNA (shNT). (B) Caco-2 cells stably expressing shNT or shMRP2 (#305) were incubated with the cytotoxic drug etoposide for 84 h. Sensitivity to the anti-cancer drug was determined by the MTT cell viability assay. A dose-response curve was fitted to the data to determine the toxicity (IC₅₀ value) of the drugs. The non-linear survival curves are presented as the mean ± SEM of four experiments performed in triplicate. Statistical significance was calculated using multiple t-test comparisons, where * p < 0.05 as compared with shNT. Inhibition of human shMRP2 expression reduced the resistance of Caco-2 cells to etoposide compared to control cells. IC₅₀ and RR values are presented in Table 1.