used by the bacteria as a nutrient source, and confirms that low glucose concentration in the ASL is required for the antimicrobial activity of airway epithelia in vivo. Discussion In this study, we describe a novel mechanism that allows human airway epithelia to generate a transepithelial glucose concentration gradient. Moreover, this gradient results in an ASL that contains a low glucose concentration. Our studies suggest that this low concentration of glucose plays an important role in limiting the growth of bacteria and maintaining the sterility on the surface of human airway epithelia and leads us to speculate that the concentration of other carbon sources in ASL may also be low. We propose a novel mechanism in human airway epithelia that generates a transepithelial glucose concentration gradient by expressing 2 distinct facilitated diffusion transporters in polarized localizations. This is in contrast to mechanisms in epithelia of the human ileum or alveolar epithelia in the distal lung, where Na+- glucose cotransport plays a central role in transepithelial glucose transport. GLUT-1, which we observed in the basolateral membrane, is the most ubiquitously expressed facilitated diffusion glucose transporter in humans; it has a Km between 3 and 7 mM and has a proposed function of cellular insulin-independent basal glucose uptake [18,37]. GLUT-10, which we observed in the apical membrane of human airway epithelia, has previously been examined because of a potential link to Diabetes Mellitus Type 2, but studies failed to prove an association [38–40]. Interestingly, GLUT-10 has a very low Km of approximately 0.3 mM, the lowest in the group of facilitated diffusion glucose transporters [18,22]. This Km is very close to the glucose concentration in both human ASL [10] and rat lung cells [41] (0.5 mM). Glucose is exclusively supplied to the airways from circulating blood, reaching the basolateral side of epithelial cells, where uptake of glucose can occur. Glucose can reach the ASL through both the paracellular and transcellular pathways at a specific rate, but the epithelial cells have the capacity to absorb it through the apical membrane at an equal or higher rate (Figure 3D). What is the driving force that allows epithelial absorption of glucose from both the apical and basolateral compartments? Our data supports interesting conclusions. Intracellular glucose, under normal conditions, is constantly phosphorylated by hexokinase in Airway Glucose and Innate Immunity an ATP-dependent reaction, creating flux into a chemical ‘‘fourth compartment’’. This would maintain an intracellular non- phosphorylated glucose concentration that is lower than that of blood, and a glucose concentration gradient that can be the driving force for basolateral uptake. Any glucose reaching the ASL would therefore be absorbed at a rate that would keep it at concentration equilibrium with intracellular non-phosphorylated glucose (Figure 6, A). Data supporting this idea come from studies performed in isolated lung cells grown in suspension, in which the intracellular concentration of glucose was found to be ,0.5 mM when the extracellular glucose was 5 mM or higher [41]. Disruption of this homeostatic mechanism could occur at different levels. An increase in blood glucose concentration, such as in diabetes mellitus, could result in an increased rate of glucose flux into ASL with correspondingly increased glucose concentra- tion (Figure 6, B). This could explain the increased rate of ventilator-associated MRSA pneumonia [11] in ICU patients with hyperglycemia and the poor outcomes in hyperglycemic patients hospitalized for community acquired pneumonia [3]. This mechanism could also be disrupted by fluctuations of tight junction permeability (Figure 6, C), such as those induced by hypersensitivity reactions, exposure to toxic particles, or other inflammatory stimuli, including viral infections such as influenza or the common cold, which have been shown to increase the risk of bacterial pneumonia and correlate with detection of nasal glucose [1,10,42–45]. Also, even though environmental exposures to glucose are uncommon (confectioners lung), the presence of sugars in formulations that are nebulized into the airway could affect the homeostatic mechanism we have described. Finally, disruption of apical transport of glucose –as shown in our in vitro data– could impair generation of the glucose concentration gradient (Figure 6, D). It has been shown that mutations in the gene encoding GLUT-10 are responsible for Arterial Tortuosity Syndrome [46,47], a rare autosomal recessive disorder, with a phenotype characterized by arterial elongation, tortuosity and aneurysms, and a high mortality rate (40% before the age of 5 years). The cause of early deaths has not been studied extensively, but a lethal case of spontaneous bilateral Staphylococcus aureus bronchopneumonia in a 4 year-old child has been reported [48]. The question of whether this infection was a consequence of impaired glucose transport in the airways remains to be answered. Figure 6. Alterations in transepithelial glucose transport. Diagram of transepithelial glucose transport under normal conditions (A), hyperglycemia (B), increased tight junction permeability (C) or impaired apical glucose transporter function (D). Gray solid arrows represent glucose uptake capacity across the apical (top) and basolateral (bottom) membranes. Dotted arrows represent transcellular bidirectional fluxes of glucose. Arrow width and font size are modified in each figure according to the primary alteration in glucose transport rate and resulting basolateral and apical concentration, respectively. GLUT-10 is present in the apical membrane and GLUT-1 is present in the basolateral membrane. Intracellular glucose (G) is phosphorylated by hexokinase (HK) for subsequent glycolysis. G6P = glucose-6-phosphate. doi:10.1371/journal.pone.0016166.g006 PLoS ONE | www.plosone.org 6 January 2011 | Volume 6 | Issue 1 | e16166