Characteristics of L-carnitine transport in cultured human hepatoma HLF cells.
The recently cloned organic cation transporter, OCTN2, isolated as a homologue of OCTN1, has been shown to be of physiological importance in the renal tubular reabsorption of filtered L-carnitine as a high-affinity Na+ carnitine transporter in man. Although the mutation of the OCTN2 gene has been proved to be directly related to primary carnitine deficiency, there is little information about the L-carnitine transport system in the liver. In this study, the characteristics of L-carnitine transport into hepatocytes were studied by use of cultured human hepatoma HLF cells, which expressed OCTN2 mRNA to a greater extent than OCTN1 mRNA. The uptake of L-carnitine into HLF cells was saturable and the Eadie-Hofstee plot showed two distinct components. The apparent Michaelis constant and the maximum transport rate were 6.59+/-1.85 microM (mean+/-s.d.) and 78.5+/-21.4 pmol/5 min/10(6) cells, respectively, for high-affinity uptake, and 590+/-134 microM and 1507+/-142 pmol/5 min/10(6) cells, respectively, for low-affinity uptake. The high affinity L-carnitine transporter was significantly inhibited by metabolic inhibitors (sodium azide, dinitrophenol, iodoacetic acid) and at low temperature (4 degrees C). Uptake of [3H]L-carnitine also required the presence of Na+ ions in the external medium. The uptake activity was highest at pH 7.4, and was significantly lower at acidic or basic pH. L-Carnitine analogues (D-carnitine, L-acetylcarnitine and gamma-butyrobetaine) strongly inhibited uptake of [3H] L-carnitine, whereas beta-alanine, glycine, choline, acetylcholine and an organic anion and cation had little or no inhibitory effect. In conclusion, L-carnitine is absorbed by hepatocytes from man by an active carrier-mediated transport system which is Na+-, energy- and pH-dependent and has properties very similar to those of the carnitine transporter OCTN2.
Yokogawa
K
K
Department of Pharmacology and Pharmaceutics, Graduate School of Natural Science and Technology, Kanazawa University, Japan.
Miya
K
K
Tamai
I
I
Higashi
Y
Y
Nomura
M
M
Miyamoto
K
K
Tsuji
A
A
D001693:Biological Transport, Active / Q000502:physiology; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D002331:Carnitine / Q000031:analogs & derivatives / Q000493:pharmacokinetics*; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008660:Metabolism / Q000187:drug effects*; D012333:RNA, Messenger / Q000145:classification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012964:Sodium / Q000494:pharmacology*; D013696:Temperature; D013997:Time Factors; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D012964:Sodium; D002331:Carnitine