Transport rather than diffusion-dependent route for nitric oxide gas activity in alveolar epithelium.
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.
Brahmajothi
Mulugu V
MV
Department of Pediatrics, Duke University Medical Center, Durham, NC 27710, USA.
Mason
S Nicholas
SN
Whorton
A Richard
AR
McMahon
Timothy J
TJ
Auten
Richard L
RL
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
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