Data-based mathematical modeling of vectorial transport across double-transfected polarized cells.
Vectorial transport of endogenous small molecules, toxins, and drugs across polarized epithelial cells contributes to their half-life in the organism and to detoxification. To study vectorial transport in a quantitative manner, an in vitro model was used that includes polarized MDCKII cells stably expressing the recombinant human uptake transporter OATP1B3 in their basolateral membrane and the recombinant ATP-driven efflux pump ABCC2 in their apical membrane. These double-transfected cells enabled mathematical modeling of the vectorial transport of the anionic prototype substance bromosulfophthalein (BSP) that has frequently been used to examine hepatobiliary transport. Time-dependent analyses of (3)H-labeled BSP in the basolateral, intracellular, and apical compartments of cells cultured on filter membranes and efflux experiments in cells preloaded with BSP were performed. A mathematical model was fitted to the experimental data. Data-based modeling was optimized by including endogenous transport processes in addition to the recombinant transport proteins. The predominant contributions to the overall vectorial transport of BSP were mediated by OATP1B3 (44%) and ABCC2 (28%). Model comparison predicted a previously unrecognized endogenous basolateral efflux process as a negative contribution to total vectorial transport, amounting to 19%, which is in line with the detection of the basolateral efflux pump Abcc4 in MDCKII cells. Rate-determining steps in the vectorial transport were identified by calculating control coefficients. Data-based mathematical modeling of vectorial transport of BSP as a model substance resulted in a quantitative description of this process and its components. The same systems biology approach may be applied to other cellular systems and to different substances.
Bartholomé
Kilian
K
Institute for Physics, University of Freiburg, Freiburg, Germany.
Rius
Maria
M
Letschert
Katrin
K
Keller
Daniela
D
Timmer
Jens
J
Keppler
Dietrich
D
D000818:Animals; D001646:Bile / Q000378:metabolism; D001693:Biological Transport, Active / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology*; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004285:Dogs; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D015233:Models, Statistical; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism; D029362:Organic Anion Transporters, Sodium-Independent / Q000235:genetics / Q000378:metabolism; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3; D014162:Transfection
D002352:Carrier Proteins; D018076:DNA, Complementary; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D029362:Organic Anion Transporters, Sodium-Independent; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3; C000715087:ABCC2 protein, human; C421209:SLCO1B3 protein, human