title: Involvement of LAT1 and LAT2 in the high- and low-affinity transport of L-leucine in human retinal pigment epithelial cells (ARPE-19 cells).
abstract: System L, which is encoded by LAT1 and LAT2, is an amino acid transport system that transports neutral amino acids, including several essential amino acids in an Na+-independent manner. Due to its broad substrate selectivity, system L has been proposed to mediate the transport of amino-acid-related drugs across the blood-tissue barriers. We characterized L-leucine transport and its corresponding transporter in a human retinal pigment epithelial cell line (ARPE-19 cells) as an in vitro model of the outer blood-retinal barrier. [3H]L-leucine uptake by ARPE-19 cells took place in an Na+-, Cl(-)-independent and saturable manner with K(m) values of 8.71 and 220 microM. This process was more potently cis-inhibited by substrates of LAT1 than those of LAT2. [3H]L-leucine efflux from ARPE-19 cells was trans-stimulated by substrates of LAT1 and LAT2 through the obligatory exchange mechanism of system L. Although RT-PCR analysis demonstrated that LAT1 and LAT2 mRNA are expressed in ARPE-19 cells, the LAT1 mRNA concentration is 42-fold higher than that of LAT2. Moreover, immunoblot analysis demonstrated that LAT1 is expressed in ARPE-19 cells. In conclusion, although the transport function of LAT1 is greater than that of LAT2, LAT1 and LAT2 are involved in L-leucine transport in ARPE-19 cells.
authors.affiliation: Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Yamamoto
authors.forename: Shin-Ichi
authors.initials: S
authors.lastname: Akanuma
authors.forename: Masanori
authors.initials: M
authors.lastname: Tachikawa
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Hosoya
mesh_terms: D027182:Amino Acid Transport System y+ / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D000596:Amino Acids / Q000378:metabolism / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D001813:Blood-Retinal Barrier / Q000378:metabolism*; D018929:Cell Culture Techniques; D002460:Cell Line; D002713:Chlorine / Q000494:pharmacology; D003470:Culture Media; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000378:metabolism*; D027301:Fusion Regulatory Protein 1, Light Chains / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D015151:Immunoblotting; D027283:Large Neutral Amino Acid-Transporter 1 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D007930:Leucine / Q000378:metabolism* / Q000493:pharmacokinetics; D008954:Models, Biological; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012964:Sodium / Q000494:pharmacology; D013379:Substrate Specificity
chemical_list: D027182:Amino Acid Transport System y+; D000596:Amino Acids; D003470:Culture Media; D027301:Fusion Regulatory Protein 1, Light Chains; D027283:Large Neutral Amino Acid-Transporter 1; C420646:SLC7A8 protein, human; D002713:Chlorine; D012964:Sodium; D007930:Leucine