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Pharmaceutical Research

, Volume 31, Issue 9, pp 2539–2548 | Cite as

Laurate Permeates the Paracellular Pathway for Small Molecules in the Intestinal Epithelial Cell Model HT-29/B6 via Opening the Tight Junctions by Reversible Relocation of Claudin-5

  • Isabel Dittmann
  • Maren Amasheh
  • Susanne M. Krug
  • Alexander G. Markov
  • Michael Fromm
  • Salah Amasheh
Research Paper

ABSTRACT

Purpose

To mechanistically analyze effects of the medium-chain fatty acid laurate on transepithelial permeability in confluent monolayers of the intestinal epithelial cell line HT-29/B6, in context with an application as an absorption enhancer improving transepithelial drug permeation.

Methods

Transepithelial resistance and apparent permeability for paracellular flux markers was measured using Ussing-type chambers. Two-path impedance spectroscopy was employed to differentiate between transcellular and paracellular resistance, and confocal imaging and Western blotting was performed.

Results

Laurate resulted in a substantial and reversible decrease in transepithelial resistance by 50% which was attributed to a decrease in paracellular resistance. Simultaneously, an increase in permeability for fluorescein (330 Da) was detected, while permeabilities for 4 kDa FITC-dextran and sulpho-NHS-SS-biotin (607 Da) remained unaltered. Confocal laser-scanning microscopy revealed a marked reduction of claudin-5, while other tight junction proteins including tricellulin, a protein preventing the paracellular passage of macromolecules, were not affected.

Conclusions

Laurate induces an increase in paracellular permeability for molecules up to a molecular mass of 330 Da by retrieval of claudin-5 from tight junctions without affecting tricellular contacts and the paracellular passage of macromolecules. We hereby provide, for the first time, a mechanistical explanation of laurate-induced permeability enhancement on molecular level.

KEY WORDS

absorption enhancer drug uptake epithelial cell tight junctions 

ABBREVIATIONS

FITC

Fluorescein isothiocyanate

MCFA

Medium-chain fatty acids

NHS

n-hydroxysuccinimide

TER

Transepithelial resistance

TJ

Tight junctions

Notes

ACKNOWLEDGMENTS AND DISCLOSURES

We thank Detlef Sorgenfrei, In-Fah M. Lee, and Anja Fromm for their expert technical assistance. This work was supported by grants of the Deutsche Forschungsgemeinschaft (Grants DFG FOR 721/2, DFG SFB 852), and the St. Petersburg State University (Grant SPbGU 1.37.118.201).

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Isabel Dittmann
    • 1
  • Maren Amasheh
    • 2
  • Susanne M. Krug
    • 1
  • Alexander G. Markov
    • 3
  • Michael Fromm
    • 1
  • Salah Amasheh
    • 4
  1. 1.Institute of Clinical PhysiologyCharitéBerlinGermany
  2. 2.Department of Gastroenterology Infectiology and Rheumatology, Division of Nutritional MedicineCharitéBerlinGermany
  3. 3.Department of General PhysiologySt. Petersburg State UniversitySt. PetersburgRussian Federation
  4. 4.Institute of Veterinary Physiology, Department of Veterinary MedicineFreie Universität BerlinBerlinGermany

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