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

Abstract

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 [(3)H]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.

Pharmacological characterization of a high-affinity p-tyramine transporter in rat brain synaptosomes

“…The somewhat more selective inhibitor quinidine (pan OCT inhibition, no PMAT inhibition)4849 gave similar effects to decynium-22, increasing the apparent p -tyramine release under both basal (Fig. 5a; F = 106.6 (3, 58), P  < 0.0001, n = 4) and depolarizing (Fig.…”

“…Quinidine, an inhibitor of OCT1-3, but not PMAT4849, had similar effects to decynium-22 (Fig. 5a,b), suggesting that PMAT may not be involved in transporting p -tyramine.…”

Expression of organic cation transporter 1 (OCT1): unique patterns of indirect regulation by nuclear receptors and hepatospecific gene regulation

“…mammary gland, eye, adipose tissue, and immune cells (Gilchrist & Alcorn, 2010;Ito et al, 2014;Lee et al, 2013;Minuesa et al, 2008;More et al, 2010;MorenoNavarrete et al, 2011;Nishimura & Naito, 2005;Zhang et al, 2008). OCT1 protein was also localized in ileum at the brush-border luminal as well as at lateral membranes of the enterocytes (Han et al, 2013;Muller et al, 2005), at the luminal membrane of bronchial epithelial cells (Ingoglia et al, 2015;Lips et al, 2005) and in endothelial cells of brain microvessels (Lin et al, 2010). In human eye, OCT1 was detected in cornea, iris-ciliary body, and blood-retina barrier (Zhang et al, 2008) ( Table 1).…”

“…In contrast to rodents, human OCT1 might be involved in the reabsorption of ultra-filtrated cations in human kidney (Jonker & Schinkel, 2004). In human lung, OCT1 is proposed to mediate absorption of some drugs from the bronchi (Ingoglia et al, 2015). In human brain, OCT1 may facilitate passage of endogenous substrates and drugs across the blood-brain barrier (Lin et al, 2010).…”

P-Glycoprotein Transport of Neurotoxic Pesticides

“…Little is known about the uptake transporters of these pesticides. Organic cation transporters are thought to mediate MPP 1 uptake in humans and rodents (Ahmadimoghaddam et al, 2012;Belzer et al, 2013;Ingoglia et al, 2015;Wu et al, 2015), and a organic anion transporting polypeptide may be involved in diazinon uptake in zebrafish (Popovic et al, 2014). However, it is likely that other uptake transporters of pesticides will be identified in the future.…”