Functional characterization of the organic cation transporters (OCTs) in human airway pulmonary epithelial cells

• Published in: Biochimica et biophysica acta Vol. 1848; no. 7; pp. 1563 - 1572
• Main Authors: Ingoglia, Filippo, Visigalli, Rossana, Rotoli, Bianca Maria, Barilli, Amelia, Riccardi, Benedetta, Puccini, Paola, Dall'Asta, Valeria
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.07.2015
• Subjects: 1-Methyl-4-phenylpyridinium - metabolism; 1-Methyl-4-phenylpyridinium - pharmacokinetics; Biological Transport - drug effects; Biological Transport - genetics; Cationic drug transporters; Cell Line; Cell Line, Tumor; Corticosterone - pharmacology; Dinoprostone - pharmacology; Epithelial Cells - metabolism; Humans; Hydrogen-Ion Concentration; Kinetics; Lung - cytology; OCT1; OCT2; OCT3; Organic Cation Transport Proteins - genetics; Organic Cation Transport Proteins - metabolism; Organic Cation Transporter 1 - genetics; Organic Cation Transporter 1 - metabolism; Organic Cation Transporter 2; Organic cation transporters; Pulmonary epithelium; Quinidine - pharmacology; RNA Interference; Time Factors; PBS; OCT; PGE2; Organic cation transporters; MPP; OCT1; OCT2; OCT3; Cationic drug transporters; TEER; Pulmonary epithelium; EBSS; ALI
• ISSN: 0005-2736; 0006-3002; 1879-2642
• DOI: 10.1016/j.bbamem.2015.04.001

Organic cation transporters (OCT1–3) mediate the transport of organic cations including inhaled drugs across the cell membrane, although their role in lung epithelium hasn't been well understood yet. We address here the expression and functional activity of OCT1–3 in human airway epithelial cells A549, Calu-3 and NCl-H441. Kinetic and inhibition analyses, employing [3H]1-methyl-4-phenylpyridinium (MPP+) as substrate, and the compounds quinidine, prostaglandine E2 (PGE2) and corticosterone as preferential inhibitors of OCT1, OCT2, and OCT3, respectively, have been performed. A549 cells present a robust MPP+ uptake mediated by one high-affinity component (Km~50μM) which is identifiable with OCT3. Corticosterone, indeed, completely inhibits MPP+ transport, while quinidine and PGE2 are inactive and SLC22A3/OCT3 silencing with siRNA markedly lowers MPP+ uptake. Conversely, Calu-3 exhibits both a high (Km<20μM) and a low affinity (Km>0.6mM) transport components, referable to OCT3 and OCT1, respectively, as demonstrated by the inhibition analysis performed at proper substrate concentrations and confirmed by the use of specific siRNA. These transporters are active also when cells are grown under air–liquid interface (ALI) conditions. Only a very modest saturable MPP+ uptake is measurable in NCl-H441 cells and the inhibitory effect of quinidine points to OCT1 as the subtype functionally involved in this model. Finally, the characterization of MPP+ transport in human bronchial BEAS-2B cells suggests that OCT1 and OCT3 are operative. These findings could help to identify in vitro models to be employed for studies concerning the specific involvement of each transporter in drug transportation. Proposed contribution of OCT1‐3 for MPP+ uptake in airway epithelial cells [Display omitted] •MPP+ transport in A549 cells is operated by OCT3.•MPP+ transport in Calu-3 cells is operated by OCT1 and OCT3.•NCl-H441 cells display a very modest saturable uptake of MPP+.•BEAS-2B cells display a very modest saturable uptake of MPP+, with kinetic features similar to Calu-3 cells.