Introduction: The properties of airway surface liquid (ASL) are controlled by the coordinated activity of ion channels and transporters that mediate transepithelial absorption and secretion of ions. ASL homeostasis is dramatically altered in cystic fibrosis (CF) due to CFTR dysfunction that produces ASL dehydration and acidification, resulting in mucus-stasis, reduced airway clearance and favoring bacterial infections and inflammation. Monocarboxylates can be transported by an electrogenic Na⁺-coupled system that corresponds to SMCTs (SLC5 family) and an electroneutral H⁺-coupled system that correspond to MCTs, (SLC16 family). Previous work demonstrated the presence of MCT2 on the apical surface and MCT4 on the basolateral surface in human bronchial epithelium. But there is no evidence of SMCTs activity in this tissue. In here, we describe for first time that SMCT1/SCL5A8 is functionally expressed in mouse tracheal epithelium. We hypothesized that the activation of monocarboxylate transport might down regulate airway clearance as it is known that increased Na⁺ absorption reduce ASL, while increased H⁺ secretion acidify ASL. Then we tested if mucociliary clearance is affected by MCT and SMCT activity in mouse airways.

Methods: Short-circuit current in mouse tracheas was measured in Ussing chambers, expression of transporters by qRT-PCR, particle track speed (PTS) and mucus transport velocity (MTV) by videomicroscopy. Localization of MCT2 and MCT4 was evaluated by immunofluorescence. Animals (C57BL6/J) were housed at CECs-Animal facility under controlled temperature and humidity with free access to water and food. All protocols were approved by the IACUC (#CECs-2022-04), in accordance with relevant guidelines and regulations.

Results: 10mM apical L-lactate and D-lactate induced a negative current (I_SC ~ -35 µA·cm^-2; n=4-5 for each group) that was dependent on apical Na⁺. qRT-PCR assay determined that SLC5A8/SMCT1 was highly expressed in airway epithelium compared to SLC5A12/SMCT2. In addition, L-lactate, which is transported by SMCT and MCT induced an increase in PTS (2.1±0.1 control vs 3.28±0.4 µm·s^-1 n=3 each group; p<0.05 rank sum test) and inhibited by 1 µM of the MCT1/2 inhibitor AR-C155858 (1.9±0.1 µm·s^-1). On the other hand, D-lactate that is exclusively transported by SMCT1 did not affected PTS (2.03±0.1 µm·s^-1; p>0.05 compared to controls; n=3 each group), indicating that SMCT1-dependent Na⁺ absorption didn’t modify airway clearance. Furthermore, analysis of MTV determined that L-lactate but not D-lactate increased the velocity of mucus (1.05±0.1 control vs 1.29±0.1 µm·s^-1 for L-lactate and 1.07±0.02 µm·s^-1 for D-lactate; n=3 each group; p<0.05 rank sum test) and the increase of L-lactate was prevented by 1 µM of AR-C155858 (0.98±0.1 µm·s^-1). Surprisingly, immunofluorescence determined that MCT2 and MCT4 are localized in basolateral membrane. Thus, our results indicate that the effect on PTS and MTV induced by L-lactate possibly is mediated by MCT1. Preliminary data indicated that in presence of MCT1 inhibitor (AZD3965) this effect on MTV was diminished.

Discussion: Monocarboxylate uptake by MCTs removes H⁺ from ASL, alkalizing the airway surface and improving airway clearance. The use of MCTs transportable substrates might help alleviate mucostasis in muco-obstructive diseases and will be tested in animal models of these diseases.