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
- 725 Downloads
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.
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.
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.
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 WORDSabsorption enhancer drug uptake epithelial cell tight junctions
Medium-chain fatty acids
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 18.104.22.168).
- 21.Zenger V, Laurate C. US Food and Drug Administration. High laurate canola oil. BNF No.25. 1995. http://www.fda.gov/Food/FoodScienceResearch/Biotechnology/Submissions/ucm161141.htm. Accessed 28 February 2014.
- 37.Sirotkin H, Morrow B, Saint-Jore B, Puech A, Das Gupta R, Patanjali SR, et al. Identification, characterization, and precise mapping of a human gene encoding a novel membrane-spanning protein from the 22q11 region deleted in velo-cardio-facial syndrome. Genomics. 1997;42(2):245–51.PubMedCrossRefGoogle Scholar