title: Transport rather than diffusion-dependent route for nitric oxide gas activity in alveolar epithelium.
abstract: The pathway by which inhaled NO gas enters pulmonary alveolar epithelial cells has not been directly tested. Although the expected mechanism is diffusion, another route is the formation of S-nitroso-L-cysteine, which then enters the cell through the L-type amino acid transporter (LAT). To determine if NO gas also enters alveolar epithelium this way, we exposed alveolar epithelial-rat type I, type II, L2, R3/1, and human A549-cells to NO gas at the air liquid interface in the presence of L- and D-cysteine+/-LAT competitors. NO gas exposure concentration dependently increased intracellular NO and S-nitrosothiol levels in the presence of L- but not D-cysteine, which was inhibited by LAT competitors, and was inversely proportional to diffusion distance. The effect of L-cysteine on NO uptake was also concentration dependent. Without preincubation with L-cysteine, NO uptake was significantly reduced. We found similar effects using ethyl nitrite gas in place of NO. Exposure to either gas induced activation of soluble guanylyl cylase in a parallel manner, consistent with LAT dependence. We conclude that NO gas uptake by alveolar epithelium achieves NO-based signaling predominantly by forming extracellular S-nitroso-L-cysteine that is taken up through LAT, rather than by diffusion. Augmenting extracellular S-nitroso-L-cysteine formation may augment pharmacological actions of inhaled NO gas.
authors.affiliation: Department of Pediatrics, Duke University Medical Center, Durham, NC 27710, USA.
authors.forename: Mulugu V
authors.initials: MV
authors.lastname: Brahmajothi
authors.forename: S Nicholas
authors.initials: SN
authors.lastname: Mason
authors.forename: A Richard
authors.initials: AR
authors.lastname: Whorton
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: McMahon
authors.forename: Richard L
authors.initials: RL
authors.lastname: Auten
mesh_terms: D027241:Amino Acid Transport System y+L / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive; D001692:Biological Transport / Q000187:drug effects*; D003545:Cysteine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D004058:Diffusion; D004848:Epithelium / Q000378:metabolism* / Q000473:pathology; D006162:Guanylate Cyclase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007930:Leucine / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology; D011650:Pulmonary Alveoli / Q000473:pathology; D051381:Rats; D026403:S-Nitrosothiols / Q000378:metabolism; D013237:Stereoisomerism
chemical_list: D027241:Amino Acid Transport System y+L; D009363:Neoplasm Proteins; D026403:S-Nitrosothiols; C115131:SLC43A1 protein, human; D009569:Nitric Oxide; C021315:S-nitrosocysteine; D006162:Guanylate Cyclase; D007930:Leucine; D003545:Cysteine