Drug Delivery Characteristics of the Progenitor Bronchial Epithelial Cell Line VA10
- 630 Downloads
To determine the integrity and permeability properties of the immortalized human VA10 bronchial epithelial cell line for its suitability as an in vitro drug permeation model.
Cells were grown under liquid-covered culture (LCC) or air-liquid interface (ALI) culture, characterized using electron microscopy and immunostaining. Integrity was measured using transepithelial electrical resistance (TER) and permeability of fluorescein sodium (Flu-Na). General permeability was established with dextrans and model drugs and P-glycoprotein (P-gp) function determined with bidirectional flux of rhodamine-123.
ALI culture resulted in 2–3 cell layers with differentiation towards ciliated cells but LCC showed undifferentiated morphology. VA10 cells formed TJ, with higher TER in LCC than ALI (∼2500 vs. ∼1200 Ω*cm2) and Flu-Na permeability ∼1–2 × 10−7 cm/s. ALI cultured cells expressed P-gp and distinguished between compounds depending on lipophilicity and size, consistent with previous data from Calu-3 and 16HBE14o-cell lines.
ALI cultured cell layers capture the in vivo-like phenotype of bronchial epithelium and form functional cell barrier capable of discriminating between compounds depending on physiochemical properties. The VA10 cell line is an important alternative to previously published cell lines and a relevant model to study airway drug delivery in vitro.
KEY WORDSair-liquid interface culture airway permeability differentiation drug delivery human bronchial epithelial cells
surface area (cm2)
basolateral to apical
bovine serum albumin
ciliary beating frequency
cystic fibrosis transmembrane conductance regulator
fluorescein isothiocyanate labeled dextran
Hanks balanced salt solution
human papilloma virus-16
normal human bronchial epithelial
apparent permeability (cm/s)
phosphate buffered saline
retinoblastoma tumor suppressor protein
scanning electron microscopy
transepithelial electrical resistance (Ω*cm2)
ACKNOWLEDGMENTS AND DISCLOSURES
Financial support from the Eimskip Fund of University of Iceland, the University of Iceland Research Fund, the Landspitali University Hospital Science Fund and the Bergthóru and Thorsteins Scheving Thorsteinssonar Fund is gratefully acknowledged. We thank Professor Magnus Karl Magnusson for critical discussion and good advice, Sigrún Kristjánsdóttir at the Pathology Department of Landspitali University Hospital for her contribution to the paraffin prepared samples and Bergthóra S. Snorradóttir at the University of Iceland for help with the HPLC.
Video of beating cilia of differentiated VA10 cells cultured at ALI for 14 days was taken with Leica DMI3000 inverted microscopy, 40x objective and DIC filter, focus directed at apical surface of the cell layer. Before imaging, the surface of the cells was immersed in PBS. The ciliary beating can clearly be seen on individual cells and patches of ciliated cells that cover 10-15% of the surface area. (WMV 1794 kb)
- 2.Guidance for industry: Waiver of in vivo bioavailability and bioequivalence studies for immediate-release solid oral dosage forms based on a biopharmaceutics classification system. August 2000, CDER/FDA.Google Scholar
- 3.Note for guidance on the investigation of bioavailability and bioequivalence. December 2000, EMEA.Google Scholar
- 6.Shen BQ, Finkbeiner WE, Wine JJ, Mrsny RJ, Widdicombe JH. Calu-3: a human airway epithelial cell line that shows cAMP-dependent Cl- secretion. Am J Physiol Lung Cell Mol Physiol. 1994;266(5):L493–501.Google Scholar
- 19.Ehrhardt C, Kneuer C, Fiegel J, Hanes J, Schaefer U, Kim K-J, et al. Influence of apical fluid volume on the development of functional intercellular junctions in the human epithelial cell line 16HBE14o- implications for the use of this cell line as an in vitro model for bronchial drug absorption studies. Cell Tissue Res. 2002;308(3):391–400.PubMedCrossRefGoogle Scholar
- 30.Otton A. Cell culture forensics of Calu-3: a human lung epithelial cell line. Ethn Dis. 2009;19(2):S78–9.Google Scholar
- 36.Yamaya M, Finkbeiner WE, Chun SY, Widdicombe JH. Differentiated structure and function of cultures from human tracheal epithelium. Am J Physiol Lung Cell Mol Physiol. 1992;262(6):L713–24.Google Scholar
- 41.Kim K-J. Bioelectrical characterization of cultured epithelial cell (mono)layers and excised tissues. In: Lehr CM, editor. Cell culture models of biological barriers. CRC Press; 2002. p. 41–51.Google Scholar
- 46.Conradi RA, Burton PS, Borchardt RT. Physico-chemical and biological factors that influence a drug’s cellular permeability by passive diffusion. In: Pliška V, Testa B, van de Waterbeemd H, editors. Lipophilicity in drug action and toxicology. Weinheim: Wiley-VCH Verlag GmbH; 2008. p. 233–52.Google Scholar