Immuno-histochemical detection of MRPs in human lung cells in culture

The transport of molecules across membranes is an essential function of all living organisms and a large number of specific transporters have evolved to carry out this function. The largest transporter gene family is the ATP-binding cassette (ABC) transporter superfamily. The multidrug resistance-associated protein (MRP) family is comprised of nine related ABC transporters. The intra-cellular distribution of the different MRP isoforms in relation to their physiological and non physiological function is still a point of discussion. For this purpose we used normal human lung cells (bronchial epithelial cells, NHBEC, and peripheral lung cells, PLC) as well as tumor cell cultures as test tools to investigate the intracelluar localization of these proteins under classical culture conditions and under air-liquid interface by means of indirect fluorescence microscopy. Characterization of the cultured cells as lung epithelial cells was performed by means of immuno-histochemical analysis. MRP1 and MRP3 were localised to the cellular membrane in all tested lung cell types. In contrast to that MRP2, MRP4 and MRP5 could be described as intracellular proteins in NHBEC and PLC. All MRP1-MRP5 isoforms could be characterized in A549 tumor cell line as membrane proteins. In order to imitate the physiological in vivo circumstances in the lung, we have established a dry/wet method (air-liquid interface) for cell cultivation so that cultured cells have the option to polarize between air and basal membrane and this might influence the distribution pattern of MRP1 and MRP2 in NHBEC. Using confocal laser scanning techniques we could show that in cells kept under dry/wet conditions MRP1 was found to be localised to baso-lateral cell regions while MRP2 was localised to all cell regions. Under classical culture conditions MRP1 was not localized to particular membrane regions and MRP2 was found to be an intracellular protein.