Primary porcine proximal tubular cells as a model for transepithelial drug transport in human kidney.
BACKGROUND
Kidney proximal tubular cells play a major role in the transport of endogenous and exogenous compounds. A multitude of different transporters are expressed starting with multidrug ABC transporters (e.g. abcb1, abcc1-6), slc22a6-8 (organic anion transporters) and slc22a1-3 (organic cation transporters). For transport studies of renal drug transport, cell lines like MDCK and LLC-PK1 are often used to overexpress and study one or two transporters, such as abcb1 or abcc1-6. However, the use is limited since under physiological conditions xenobiotics are transported through different transporters at the same time. Therefore, a primary in vitro model expressing functionally different transporters simultaneously, as it is the case in vivo, would be of great benefit.


METHODS
Primary proximal tubular cells were isolated from porcine kidney. Cells were cultured under selective culturing conditions leading to specific growth of primary proximal tubular cells. Expression of important proximal transporters was checked at mRNA level with RT-PCR, at protein level with immunocytochemistry and functionally by transport and uptake assays.


RESULTS
A model of primary proximal tubular cells was established expressing the most important transporters: abcb1, abcc1, abcc2, slc22a8, slco1a2, slc15a1, slc5a2 and slc4a4. In freshly isolated cells, slc22a1 and slc22a6 were expressed, but were down-regulated in culture. Abcb1, abcc1, abcc2 and slc4a4 were detected at protein level with immunostaining. Functional activity was confirmed for abcb1, abcc1/2, slc22a8, slc15a1/2 and slc5a1/2. The tightness of the monolayers of this model was better than in previously established in vitro models.


CONCLUSIONS
This primary cell culture model might be an interesting tool to investigate proximal tubular transport and to predict toxicity and drug interactions since it expresses functionally several transporters simultaneously.
Schlatter
Philipp
P
Department of Clinical Pharmacology and Toxicology, University Hospital, Basel, Switzerland.
Gutmann
Heike
H
Drewe
Juergen
J
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004553:Electric Conductivity; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism*; D018613:Microscopy, Confocal; D008954:Models, Biological; D000089762:Multidrug Resistance-Associated Protein 2; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine
C000715087:ABCC2 protein, human; D002352:Carrier Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D012333:RNA, Messenger