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Pharm Res (2015) 32:665679 DOI 10.1007/s11095-014-1494-0
RESEARCH PAPER
The Characterization of the Human Nasal Epithelial Cell Line RPMI 2650 Under Different Culture Conditions and Their Optimization for an Appropriate in vitro Nasal Model
Mateja Erdani Kreft & Urka Dragin Jerman & Eva Lasi &
Tea Laninik Riner & Neli Hevir-Kene & Luka Peternel &
Katja Kristan
Received: 11 March 2014 /Accepted: 15 August 2014 /Published online: 22 August 2014 # Springer Science+Business Media New York 2014
ABSTRACTPurpose The further characterization of the cell line RPMI 2650 and the evaluation of different culture conditions for an in vitro model for nasal mucosa.
Methods Cells were cultured in media MEM or A-MEM at air-liquid (A-L) or liquid-liquid (L-L) interfaces for 1 or 3 weeks. Different cryopreservation methods and cell culture techniques were evaluated with immunolabelling of junctional proteins, ultra-structural analysis using electron microscopy, transepithelial electrical resistance (TEER) measurements, permeation studies with dextran and jacalin, and gene expression profiling of 84 drug transporters.
Results Cell proliferation and differentiation depended on the used medium. The established epithelia expressed occludin, claudin-1, and E-cadherin under all conditions. Cells grown at the A-L interface formed more layers and exhibited a higher TEER and lower dextran and jacalin permeability than at the L-L interface, where cells morphologically exhibited a more differentiated phenotype. The expression of ABC and SLC transporters depended on culture duration and interface.
Conclusions The RPMI 2650 cells form a polarized epithelium resembling nasal mucosa. However, different culture conditions have a significant effect on cell ultrastructure, barrier integrity, and
gene expression, and should be considered when using this cell line as an in vitro model for drug permeability studies and screening of nasal drug candidates.
KEY WORDS drug transporters . microscopy . nasal epithelial cell model . permeability . TEER
ABBREVIATIONSA Surface areaABC ATP-binding cassetteA-L Air-liquid interfacec0 Initial donor concentrationdQ/dt Flux of dextran-FITC or jacalin-fluorescein across the RPMI 2650 cell barrierL-L Liquid-liquid interfacePapp Apparent permeability coefficient
SEM Scanning electron microscopy SLC Solute carrierTEER Transepithelial electrical resistance TEM Transmission electron microscopy TJ Tight junction
INTRODUCTION
The nasal administration of aerosol therapeutics is becoming an appealing alternative in drug delivery. This non-invasive method offers higher bioavailability with its avoidance of first-pass metabolism, intestinal efflux transporters, and degradation in the digestive system, as well...