Publications (79) View all
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Article: Interaction of the cardiovascular risk marker asymmetric dimethylarginine (ADMA) with the human cationic amino acid transporter 1 (CAT1).
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ABSTRACT: Elevated plasma concentrations of endogenously formed asymmetric (ADMA) and symmetric dimethyl-l-arginine (SDMA) are associated with adverse clinical outcomes. Our aim was to investigate the cellular uptake properties of ADMA by the human cationic amino acid transporter 1 (CAT1; SLC7A1). Human embryonic kidney cells (HEK293) stably overexpressing CAT1 (HEK-CAT1) and vector-transfected control cells (HEK-VC) were established to determine cellular uptake of labeled [(3)H]ADMA and [(3)H]l-arginine. Uptake of ADMA and l-arginine were significantly (p<0.001) higher in HEK-CAT1 than in HEK-VC at all investigated concentrations. Apparent V(max) values of cellular ADMA and l-arginine uptake by CAT1 were 26.9 ± 0.8 and 11.0 ± 0.2 nmol mg protein(-1) min(-1), respectively. K(m) values were 183 ± 21 μmoll(-1) (ADMA) and 519 ± 36 μmoll(-1) (l-arginine). Uptake of ADMA was inhibited by l-arginine and SDMA with IC(50) values (95% CI) of 227 (69-742) μmoll(-1) and 273 (191-390) μmoll(-1), respectively. ADMA and SDMA inhibited CAT1-mediated uptake of l-arginine with IC(50) values of 758 (460-1251) μmoll(-1) and 789 (481-1295) μmoll(-1), respectively. Efflux of ADMA was significantly increased in HEK-CAT1 cells as compared to HEK-VC (p<0.05). CAT1 mediates the cellular uptake of ADMA. In its physiological concentration range ADMA is unlikely to impair CAT1-mediated transport of l-arginine. Conversely, high (but still physiological) concentrations of l-arginine can inhibit CAT1-mediated cellular uptake of ADMA.Journal of Molecular and Cellular Cardiology 06/2012; 53(3):392-400. · 5.17 Impact Factor -
Article: Deletion of the mammalian INDY homolog mimics aspects of dietary restriction and protects against adiposity and insulin resistance in mice.
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ABSTRACT: Reduced expression of the Indy (I'm Not Dead, Yet) gene in D. melanogaster and its homolog in C. elegans prolongs life span and in D. melanogaster augments mitochondrial biogenesis in a manner akin to caloric restriction. However, the cellular mechanism by which Indy does this is unknown. Here, we report on the knockout mouse model of the mammalian Indy (mIndy) homolog, SLC13A5. Deletion of mIndy in mice (mINDY(-/-) mice) reduces hepatocellular ATP/ADP ratio, activates hepatic AMPK, induces PGC-1α, inhibits ACC-2, and reduces SREBP-1c levels. This signaling network promotes hepatic mitochondrial biogenesis, lipid oxidation, and energy expenditure and attenuates hepatic de novo lipogenesis. Together, these traits protect mINDY(-/-) mice from the adiposity and insulin resistance that evolve with high-fat feeding and aging. Our studies demonstrate a profound effect of mIndy on mammalian energy metabolism and suggest that mINDY might be a therapeutic target for the treatment of obesity and type 2 diabetes.Cell metabolism 08/2011; 14(2):184-95. · 17.35 Impact Factor -
Article: Molecular mechanism of renal tubular secretion of the antimalarial drug chloroquine.
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ABSTRACT: The antimalarial drug chloroquine is eliminated to a significant extent by renal tubular secretion. The molecular mechanism of renal chloroquine secretion remains unknown. We hypothesized that organic cation transporter 2 (OCT2) and multidrug and toxin extrusion protein 1 (MATE1), localized in the basolateral and luminal membranes of proximal tubule cells, respectively, are involved in chloroquine transport. The interaction of chloroquine with both transporters was investigated using single-transfected human embryonic kidney 293 (HEK293)-MATE1 cells in uptake experiments and single-transfected Madin-Darby canine kidney II (MDCK)-OCT2 and MDCK-MATE1 cells as well as double-transfected MDCK-OCT2-MATE1 cells grown as polarized monolayers on transwell filters. In HEK293-MATE1 cells, chloroquine competitively inhibited MATE1-mediated metformin uptake (K(i) = 2.8 μM). Cellular accumulation of chloroquine was significantly lower (P < 0.001) and transcellular chloroquine transport was significantly increased (P < 0.001) in MDCK-MATE1 and MDCK-OCT2-MATE1 cells compared to vector control cells after basal addition of chloroquine (0.1 to 10 μM). In contrast, no difference in cellular accumulation or transcellular transport of chloroquine was observed between MDCK-OCT2 and vector control cells. In line with an oppositely directed proton gradient acting as a driving force for MATE1, basal-to-apical transport of chloroquine by MDCK-OCT2-MATE1 cells increased with decreasing apical pH from 7.8 to 6.0. Transcellular transport of chloroquine by MDCK-OCT2-MATE1 cells was inhibited by cimetidine, trimethoprim, and amitriptyline. Our data demonstrate that chloroquine is a substrate and potent competitive inhibitor of MATE1, whereas OCT2 seems to play no role in chloroquine uptake. Concomitantly administered MATE1 inhibitors are likely to modify the renal secretion of chloroquine.Antimicrobial Agents and Chemotherapy 07/2011; 55(7):3091-8. · 4.84 Impact Factor -
Article: Expression and localization of the uptake transporters OATP2B1, OATP3A1 and OATP5A1 in non-malignant and malignant breast tissue.
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ABSTRACT: Organic anion transporting polypeptides (OATPs, gene family SLCO/SLC21) mediate the uptake of multiple endogenous substances such as estrogens and estrogen metabolites and of several widely prescribed drugs (e.g. statins, antibiotics and anticancer agents) into cells. Since several anticancer agents have been identified as substrates for OATPs, these transporters may also have an impact on cancer treatment. Expression of OATPs has been identified in colon, pancreatic and gastric carcinomas but to date little is known about the expression and localization of OATP family members in non-malignant breast tissue and breast cancer. We therefore analyzed the mRNA expression of all human OATP family members and further evaluated the mRNA amount of the highly-expressed OATPs OATP2B1, OATP3A1 and OATP5A1 in 10 paired samples of normal breast tissue and breast cancer. Furthermore, the tissue-specific localization of these OATPs was investigated. The results demonstrated that the mRNA expression of OATPs in normal and malignant breast tissue shows a high interindividual variability and that no significant differences in the mRNA amount between normal and malignant tissue could be detected. Furthermore, we localized OATP3A1 and OATP5A1 to the plasma membrane of epithelial cells of the lactiferous ducts in normal breast tissue. In breast cancer, both OATPs are highly expressed in the plasma membrane and in the cytoplasm. Since estrogen and estrogen metabolites as well as anticancer agents are substrates for OATPs these results indicate the possibility of OATP-mediated uptake of hormones during breast cancer development and an impact of certain OATPs on chemotherapeutic cancer treatment.Cancer biology & therapy 03/2011; 11(6):584-91. · 2.64 Impact Factor -
Article: Influence of non-steroidal anti-inflammatory drugs on organic anion transporting polypeptide (OATP) 1B1- and OATP1B3-mediated drug transport.
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ABSTRACT: The transporter-mediated uptake of drugs from blood into hepatocytes is a prerequisite for intrahepatic drug action or intracellular drug metabolism before excretion. Therefore, uptake transporters, e.g., members of the organic anion transporting polypeptide (OATP) family are important determinants of drug pharmacokinetics. Highly and almost exclusively expressed in hepatocytes are the OATP family members OATP1B1 (SLCO1B1) and OATP1B3 (SLCO1B3). Drug substrates of OATP1B1 and OATP1B3 include antibiotics and HMG-CoA reductase inhibitors (statins). It has been demonstrated that administration of two or more drugs that are substrates for these hepatic uptake transporters may lead to transporter-mediated drug-drug interactions, resulting in altered transport kinetics for drug substrates. In this study we investigated whether non-steroidal anti-inflammatory drugs (NSAIDs) and paracetamol interact with OATP1B1 and OATP1B3 using the standard substrate BSP and the drug substrate pravastatin. Using human embryonic kidney cells stably expressing OATP1B1 or OATP1B3, we demonstrated that bromosulfophthalein uptake was inhibited by diclofenac, ibuprofen. and lumiracoxib. Of interest, pravastatin uptake was stimulated by these NSAIDs, and for ibuprofen we determined activation constants (EC₅₀ values) of 64.0 and 93.1 μM for OATP1B1- and OATP1B3-mediated uptake, respectively. Furthermore, we investigated whether NSAIDs were also substrates for OATP1B1 and OATP1B3 and demonstrated that only diclofenac was significantly transported by OATP1B3, whereas all other NSAIDs investigated were not substrates for these uptake transporters. These results demonstrated that drugs may interact with transport proteins by allosteric mechanisms without being substrates and, therefore, not only uptake inhibition but also allosteric-induced modulation of transport function may be an important mechanism in transporter-mediated drug-drug interactions.Drug metabolism and disposition: the biological fate of chemicals 03/2011; 39(6):1047-53. · 3.74 Impact Factor
