Expression of xenobiotic transporters in the human renal proximal tubule cell line RPTEC/TERT1.

The kidney is a major target for drug-induced injury, primarily due the fact that it transports a wide variety of chemical entities into and out of the tubular lumen. Here, we investigated the expression of the main xenobiotic transporters in the human renal proximal tubule cell line RPTEC/TERT1 at an mRNA and/or protein level. RPTEC/TERT1 cells expressed OCT2, OCT3, OCTN2, MATE1, MATE2, OAT1, OAT3 and OAT4. The functionality of the OCTs was demonstrated by directional transport of the fluorescent dye 4-Di-1-ASP. In addition, P-glycoprotein activity in RPTEC/TERT1 cells was verified by fluorescent dye retention in presence of various P-glycoprotein inhibitors. In comparison to proliferating cells, contact inhibited RPTEC/TERT1 cells expressed increased mRNA levels of several ABC transporter family members and were less sensitive to cyclosporine A. We conclude that differentiated RPTEC/TERT1 cells are well suited for utilisation in xenobiotic transport and pharmacokinetic studies.

[1]  J. Slattery,et al.  ABCC2-Mediated Biliary Transport of 4-Glutathionylcyclophosphamide and Its Contribution to Elimination of 4-Hydroxycyclophosphamide in Rat , 2004, Journal of Pharmacology and Experimental Therapeutics.

[2]  Paul Jennings,et al.  Lactate is an ideal non-invasive marker for evaluating temporal alterations in cell stress and toxicity in repeat dose testing regimes. , 2011, Toxicology in vitro : an international journal published in association with BIBRA.

[3]  T. Cihlar,et al.  The antiviral nucleotide analogs cidofovir and adefovir are novel substrates for human and rat renal organic anion transporter 1. , 1999, Molecular pharmacology.

[4]  D. Perri,et al.  The kidney--the body's playground for drugs: an overview of renal drug handling with selected clinical correlates. , 2003, The Canadian journal of clinical pharmacology = Journal canadien de pharmacologie clinique.

[5]  F. Roch-Ramel,et al.  Renal handling of drugs and xenobiotics , 2003 .

[6]  J. Wijnholds,et al.  Characterization of the transport of nucleoside analog drugs by the human multidrug resistance proteins MRP4 and MRP5. , 2003, Molecular pharmacology.

[7]  D. Keppler,et al.  Expression of organic cation transporters OCT1 (SLC22A1) and OCT3 (SLC22A3) is affected by genetic factors and cholestasis in human liver , 2009, Hepatology.

[8]  A. Limonciel,et al.  Evidence for a role of claudin 2 as a proximal tubular stress responsive paracellular water channel. , 2014, Toxicology and applied pharmacology.

[9]  P. Pávek,et al.  Interactions with selected drug renal transporters and transporter-mediated cytotoxicity in antiviral agents from the group of acyclic nucleoside phosphonates. , 2013, Toxicology.

[10]  N. Longo,et al.  Glycosylation of the OCTN2 carnitine transporter: study of natural mutations identified in patients with primary carnitine deficiency. , 2011, Biochimica et biophysica acta.

[11]  T. Cihlar,et al.  Cytotoxicity of antiviral nucleotides adefovir and cidofovir is induced by the expression of human renal organic anion transporter 1. , 2000, Journal of the American Society of Nephrology : JASN.

[12]  F. Russel,et al.  The breast cancer resistance protein transporter ABCG2 is expressed in the human kidney proximal tubule apical membrane. , 2008, Kidney international.

[13]  A. Yonezawa,et al.  Organic cation transporter OCT/SLC22A and H(+)/organic cation antiporter MATE/SLC47A are key molecules for nephrotoxicity of platinum agents. , 2011, Biochemical pharmacology.

[14]  H. Koepsell,et al.  Regulation of human organic cation transporter hOCT2 by PKA, PI3K, and calmodulin-dependent kinases. , 2003, American journal of physiology. Renal physiology.

[15]  P. Galle,et al.  Downregulation of organic cation transporters OCT1 (SLC22A1) and OCT3 (SLC22A3) in human hepatocellular carcinoma and their prognostic significance , 2012, BMC Cancer.

[16]  Paul Jennings,et al.  Development of an in vitro renal epithelial disease state model for xenobiotic toxicity testing. , 2015, Toxicology in vitro : an international journal published in association with BIBRA.

[17]  H. Koepsell,et al.  Polyspecific Organic Cation Transporters: Structure, Function, Physiological Roles, and Biopharmaceutical Implications , 2007, Pharmaceutical Research.

[18]  D. Keppler,et al.  Localization, substrate specificity, and drug resistance conferred by conjugate export pumps of the MRP family. , 2000, Advances in enzyme regulation.

[19]  F. Baas,et al.  Multidrug-resistance protein 5 is a multispecific organic anion transporter able to transport nucleotide analogs. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[20]  S. Moestrup,et al.  Evidence that epithelial glycoprotein 330/megalin mediates uptake of polybasic drugs. , 1995, The Journal of clinical investigation.

[21]  Fanfan Zhou,et al.  Role of Glycosylation in the Organic Anion Transporter OAT1* , 2004, Journal of Biological Chemistry.

[22]  O. Griesbeck,et al.  The organic cation transporter 3 (OCT3) as molecular target of psychotropic drugs: transport characteristics and acute regulation of cloned murine OCT3 , 2013, Pflügers Archiv - European Journal of Physiology.

[23]  Shufeng Zhou,et al.  Human Multidrug Resistance Associated Protein 4 Confers Resistance to Camptothecins , 2005, Pharmaceutical Research.

[24]  Stephen H Wright,et al.  Functional influence of N-glycosylation in OCT2-mediated tetraethylammonium transport. , 2006, American journal of physiology. Renal physiology.

[25]  Daxi Sun,et al.  Substrate overlap between Mrp4 and Abcg2/Bcrp affects purine analogue drug cytotoxicity and tissue distribution. , 2007, Cancer research.

[26]  Wooin Lee,et al.  Transporters and renal drug elimination. , 2004, Annual review of pharmacology and toxicology.

[27]  Bas J Blaauboer,et al.  Application of integrated transcriptomic, proteomic and metabolomic profiling for the delineation of mechanisms of drug induced cell stress. , 2013, Journal of proteomics.

[28]  J. Cherrington,et al.  Adefovir and tenofovir susceptibilities of HIV-1 after 24 to 48 weeks of adefovir dipivoxil therapy: genotypic and phenotypic analyses of study GS-96-408. , 2001 .

[29]  Fanfan Zhou,et al.  The Role of N-Linked Glycosylation in Protein Folding, Membrane Targeting, and Substrate Binding of Human Organic Anion Transporter hOAT4 , 2005, Molecular Pharmacology.

[30]  H. Daniel,et al.  The proton oligopeptide cotransporter family SLC15 in physiology and pharmacology , 2004, Pflügers Archiv.

[31]  Yuichi Sugiyama,et al.  Functional Involvement of Multidrug Resistance-Associated Protein 4 (MRP4/ABCC4) in the Renal Elimination of the Antiviral Drugs Adefovir and Tenofovir , 2007, Molecular Pharmacology.

[32]  T. Ludwig,et al.  Cisplatin nephrotoxicity is critically mediated via the human organic cation transporter 2. , 2005, The American journal of pathology.

[33]  S. Masuda,et al.  Protective effect of concomitant administration of imatinib on cisplatin-induced nephrotoxicity focusing on renal organic cation transporter OCT2. , 2009, Biochemical pharmacology.

[34]  Thomas Ludwig,et al.  Nephrotoxicity of platinum complexes is related to basolateral organic cation transport. , 2004, Kidney international.

[35]  M C Willingham,et al.  Cellular localization of the multidrug-resistance gene product P-glycoprotein in normal human tissues. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[36]  B. Ryffel,et al.  Identification of the multidrug resistance-related P-glycoprotein as a cyclosporine binding protein. , 1989, Molecular pharmacology.

[37]  Shufeng Zhou,et al.  Upregulation of Rat Renal Cortical Organic Anion Transporter (OAT1 and OAT3) Expression in Response to Ischemia/Reperfusion Injury , 2008, American Journal of Nephrology.

[38]  T. Komatsu,et al.  Characterization of the human MATE2 proton-coupled polyspecific organic cation exporter. , 2011, The international journal of biochemistry & cell biology.

[39]  H. Koepsell,et al.  Regulation of the human organic cation transporter hOCT1 , 2004, Journal of cellular physiology.

[40]  I. Tamai,et al.  Competitive interaction of cyclosporins with the Vinca alkaloid-binding site of P-glycoprotein in multidrug-resistant cells. , 1990, The Journal of biological chemistry.

[41]  K. Inui,et al.  Interaction and transport characteristics of mycophenolic acid and its glucuronide via human organic anion transporters hOAT1 and hOAT3. , 2007, Biochemical pharmacology.

[42]  F. Russel,et al.  The MRP4/ABCC4 gene encodes a novel apical organic anion transporter in human kidney proximal tubules: putative efflux pump for urinary cAMP and cGMP. , 2002, Journal of the American Society of Nephrology : JASN.

[43]  A. Yonezawa,et al.  Disruption of multidrug and toxin extrusion MATE1 potentiates cisplatin-induced nephrotoxicity. , 2010, Biochemical pharmacology.

[44]  O. Ogawa,et al.  Identification and functional characterization of a new human kidney-specific H+/organic cation antiporter, kidney-specific multidrug and toxin extrusion 2. , 2006, Journal of the American Society of Nephrology : JASN.

[45]  Paul Jennings,et al.  Cyclosporine A induces senescence in renal tubular epithelial cells. , 2007, American journal of physiology. Renal physiology.

[46]  H. Koepsell,et al.  Characterization of regulatory mechanisms and states of human organic cation transporter 2. , 2006, American journal of physiology. Cell physiology.

[47]  T. Cihlar,et al.  Nonsteroidal anti-inflammatory drugs efficiently reduce the transport and cytotoxicity of adefovir mediated by the human renal organic anion transporter 1. , 2000, The Journal of pharmacology and experimental therapeutics.

[48]  Paul Jennings,et al.  Delineation of the Key Aspects in the Regulation of Epithelial Monolayer Formation , 2013, Molecular and Cellular Biology.

[49]  M. Hediger,et al.  Molecular cloning of PEPT 2, a new member of the H+/peptide cotransporter family, from human kidney. , 1995, Biochimica et biophysica acta.

[50]  Paul Jennings,et al.  hTERT alone immortalizes epithelial cells of renal proximal tubules without changing their functional characteristics. , 2008, American journal of physiology. Renal physiology.

[51]  Qiang Zhang,et al.  The Role of Dileucine in the Expression and Function of Human Organic Anion Transporter 1 (hOAT1). , 2011, International journal of biochemistry and molecular biology.

[52]  J. Schuetz,et al.  MRP4: A previously unidentified factor in resistance to nucleoside-based antiviral drugs , 1999, Nature Medicine.

[53]  Hiroshi Omote,et al.  A human transporter protein that mediates the final excretion step for toxic organic cations. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[54]  D. Miller,et al.  P-glycoprotein-mediated secretion of a fluorescent cyclosporin analogue by teleost renal proximal tubules. , 1995, The American journal of physiology.

[55]  V. Ganapathy,et al.  Structure, function, and regional distribution of the organic cation transporter OCT3 in the kidney. , 2000, American journal of physiology. Renal physiology.

[56]  G. Burckhardt,et al.  New clues for nephrotoxicity induced by ifosfamide: preferential renal uptake via the human organic cation transporter 2. , 2011, Molecular pharmaceutics.

[57]  A. Enomoto,et al.  Role of human organic anion transporter 4 in the transport of ochratoxin A. , 2002, Biochimica et biophysica acta.

[58]  P. Jennings,et al.  Formation of ochratoxin A and aflatoxins following the use of propionic acid as a grain preservative for storing damp barley , 2004, Mycotoxin Research.

[59]  P. McNamara,et al.  Localization and Functional Characterization of the Rat Oatp4c1 Transporter in an In Vitro Cell System and Rat Tissues , 2012, PloS one.

[60]  F. Russel,et al.  Mechanisms of renal anionic drug transport. , 2008, European journal of pharmacology.

[61]  Paul Jennings,et al.  Oxidative stress induced by potassium bromate exposure results in altered tight junction protein expression in renal proximal tubule cells , 2012, Archives of Toxicology.

[62]  E. Lock,et al.  Transcriptomic alterations induced by Ochratoxin A in rat and human renal proximal tubular in vitro models and comparison to a rat in vivo model , 2011, Archives of Toxicology.

[63]  Annette Kopp-Schneider,et al.  Application of RPTEC/TERT1 cells for investigation of repeat dose nephrotoxicity: A transcriptomic study. , 2015, Toxicology in vitro : an international journal published in association with BIBRA.

[64]  Stephan Aiche,et al.  Mechanism of cisplatin proximal tubule toxicity revealed by integrating transcriptomics, proteomics, metabolomics and biokinetics. , 2015, Toxicology in vitro : an international journal published in association with BIBRA.

[65]  F. Luft,et al.  Megalin Deficiency Offers Protection from Renal Aminoglycoside Accumulation* , 2002, The Journal of Biological Chemistry.

[66]  H. Pavenstädt,et al.  Transport Mechanisms and Their Pathology-Induced Regulation Govern Tyrosine Kinase Inhibitor Delivery in Rheumatoid Arthritis , 2012, PloS one.

[67]  S. Wright,et al.  Molecular and cellular physiology of renal organic cation and anion transport. , 2004, Physiological reviews.

[68]  A. Limonciel,et al.  A Review of the Evidence that Ochratoxin A Is an Nrf2 Inhibitor: Implications for Nephrotoxicity and Renal Carcinogenicity , 2014, Toxins.

[69]  Hideyuki Saito,et al.  Gene expression levels and immunolocalization of organic ion transporters in the human kidney. , 2002, Journal of the American Society of Nephrology : JASN.

[70]  O. Takikawa,et al.  OCTN2VT, a splice variant of OCTN2, does not transport carnitine because of the retention in the endoplasmic reticulum caused by insertion of 24 amino acids in the first extracellular loop of OCTN2. , 2007, Biochimica et biophysica acta.