Fluorescence-Based Transport Assays Revisited in a Human Renal Proximal Tubule Cell Line

Apical transport is key in renal function, and the activity of efflux transporters and receptor-mediated endocytosis is pivotal in this process. The conditionally immortalized proximal tubule epithelial cell line (ciPTEC) endogenously expresses these systems. Here, we used ciPTEC to investigate the activity of three major efflux transporters, viz., breast cancer resistance protein (BCRP), multidrug resistance protein 4 (MRP4), and P-glycoprotein (P-gp), as well as protein uptake through receptor-mediated endocytosis, using a fluorescence-based setup for transport assays. To this end, cells were exposed to Hoechst33342, chloromethylfluorescein-diacetate (CMFDA), and calcein-AM in the presence or absence of model inhibitors for BCRP (KO143), P-gp (PSC833), or MRPs (MK571). Overexpression cell lines MDCKII-BCRP and MDCKII-P-gp were used as positive controls, and membrane vesicles overexpressing one transporter were used to determine substrate and inhibitor specificities. Receptor-mediated endocytosis was inv...