We have previously shown that MDR1 P-glycoprotein (P-gp) is expressed on the microvillous membrane (mvm) of the human placenta and that it plays an important role in the capacity of cultured cytotrophoblast cells to efflux xenobiotics (Atkinson et al. 2003a). Recently we demonstrated raised expression of P-gp in placental villous tissue in explant culture transduced with MDR1 using a retroviral vector (Atkinson et al. 2003b). However after transduction P-gp was distributed diffusely in the explant and not discretely localised to the mvm. Here we test the hypothesis that diffuse P-gp distribution after transduction alters the capacity of the tissue to efflux xenobiotics.
Placentas were collected by procedures approved by the local research ethics committee. Small villous fragments dissected from normal term placentas within 1hour of delivery were cultured for 7 days in CMRL 1066 medium. For retroviral transduction the explants were co-cultured throughout with the retroviral packaging cell line GPAM MDR1. On day 7 the explants were used to measure the time course of 3H-vinblastine accumulation in the presence and absence of cyclosporin A (cycA, an inhibitor of multi-drug resistance proteins).
Time course data for control explants confirms an increased accumulation of 3H-vinblastine in the presence of cycA consistent with inhibition of P-gp and reduced efflux (Fig. 1). Transduced explants show higher accumulation of 3H-vinblastine in the presence of cycA compared to controls as demonstrated by a significantly higher cycA-sensitive accumulation i.e. lower efflux (Fig. 2).
These experiments confirm P-gp activity in cultured placental villous explants and suggest a decreased capacity of the tissue to efflux xenobiotics following retroviral transduction, despite an increased overall expression of P-gp. The increased accumulation is likely related to the broader distribution of P-gp following transduction. This study therefore demonstrates the importance of the normal mvm localization of P-gp in xenobiotic efflux from the trophoblast and illustrates the consequences of any disruption to the trafficking of this protein.
This work was funded by MRC