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Inward Translocation of the Phospholipid Analogue Miltefosine across Caco-2 Cell Membranes Exhibits Characteristics of a Carrier-mediated Process

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Lipids

Abstract

Miltefosine (hexadecylphosphocholine, HePC) is the first effective oral agent for the treatment of visceral leishmaniasis. The characteristics of HePC incorporation into the human intestinal epithelial cell line Caco-2 were investigated in order to understand its oral absorption mechanism. The results provide evidence for the involvement of a carrier-mediated mechanism, since the association of HePC at the apical pole of Caco-2 cells was (1) saturable as a function of time with a rapid initial incorporation over 5 min followed by a more gradual increase; (2) saturable as a function of concentration over the range studied (2–200 μM) with a saturable component which followed Michaelis–Menten kinetics (apparent K m 15.7 μmol/L, V max 39.2 nmol/mg protein/h) and a nonspecific diffusion component; (3) partially inhibited by low temperature and ATP depletion, indicating the temperature and energy-dependence of the uptake process. Moreover, we demonstrated, by an albumin back-extraction method, that HePC is internalized via translocation from the outer to the inner leaflet of the plasma membrane and that HePC may preferentially diffuse through intact raft microdomains. In conclusion, our results suggest that incorporation of HePC at the apical membrane of Caco-2 cells may occur through a passive diffusion followed by a translocation in the inner membrane leaflet through an active carrier-mediated mechanism.

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Acknowledgments

The authors are very grateful to Christophe Dugave for [3H]HePC synthesis and would like to thank Hélène Chacun for valuable help in radioactivity studies.

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Authors and Affiliations

  1. Laboratoire de Physico-chimie, Pharmacotechnie et Biopharmacie, UMR CNRS 8612 Faculté de Pharmacie, Univ. Paris-Sud 11, IFR 141, Tour D5, 2éme étage, 5 rue J.B. Clément, Châtenay-Malabry, Cedex, 92296, France

    Cécile Ménez & Gillian Barratt

  2. Laboratoire de Pharmacie Clinique, UPRES 2706 Faculté de Pharmacie, Univ. Paris-Sud 11, IFR 141, Châtenay-Malabry, 92296, France

    Marion Buyse & Robert Farinotti

Authors
  1. Cécile Ménez
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  2. Marion Buyse
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  4. Gillian Barratt
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Correspondence to Gillian Barratt.

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Ménez, C., Buyse, M., Farinotti, R. et al. Inward Translocation of the Phospholipid Analogue Miltefosine across Caco-2 Cell Membranes Exhibits Characteristics of a Carrier-mediated Process. Lipids 42, 229–240 (2007). https://doi.org/10.1007/s11745-007-3026-8

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  • DOI: https://doi.org/10.1007/s11745-007-3026-8

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