Characteristics of L-Carnitine Transport in Cultured Human Hepatoma HLF Cells

Abstract

Abstract 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 re-absorption 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 (mean ± s.d.) and 78.5 ± 21.4 pmol/5 min/106 cells, respectively, for high-affinity uptake, and 590 ± 134 μM and 1507 ± 142 pmol/5 min/106 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°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 γ-butyrobetaine) strongly inhibited uptake of [3H] L-carnitine, whereas β-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.