Microvascular barrier integrity is dependent on bioavailable nitric oxide (NO) produced locally by endothelial NO synthase (eNOS). Under conditions of limited substrate or cofactor availability or by enzymatic modification, eNOS may become uncoupled, producing superoxide in lieu of NO. This study was designed to investigate how eNOS-dependent superoxide production contributes to endothelial barrier dysfunction in inflammatory lung injury and its regulation. C57BL/6J mice were challenged with intratracheal LPS. Bronchoalveolar lavage fluid was analyzed for protein accumulation, and lung tissue homogenate was assayed for endothelial NOS content and function. Human lung microvascular endothelial cell (HLMVEC) monolayers were exposed to LPS in vitro, and barrier integrity and superoxide production were measured. Biopterin species were quantified, and coimmunoprecipitation (Co-IP) assays were performed to identify protein interactions with eNOS that putatively drive uncoupling. Mice exposed to LPS demonstrated eNOS-dependent increased alveolar permeability without evidence for altered canonical NO signaling. LPS-induced superoxide production and permeability in HLMVEC were inhibited by the NOS inhibitor nitro-l-arginine methyl ester, eNOS-targeted siRNA, the eNOS cofactor tetrahydrobiopterin, and superoxide dismutase. Co-IP indicated that LPS stimulated the association of eNOS with NADPH oxidase 2 (Nox2), which correlated with augmented eNOS S-glutathionylation both in vitro and in vivo. In vitro, Nox2-specific inhibition prevented LPS-induced eNOS modification and increases in both superoxide production and permeability. These data indicate that eNOS uncoupling contributes to superoxide production and barrier dysfunction in the lung microvasculature after exposure to LPS. Furthermore, the results implicate Nox2-mediated eNOS-S-glutathionylation as a mechanism underlying LPS-induced eNOS uncoupling in the lung microvasculature.
Pharmacology, Toxicology and Pharmaceutical Science
2
5%
Business, Management and Accounting
1
2%
Other
2
5%
Unknown
11
27%
Attention Score in Context
Attention Score in Context
This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 14 January 2015.
All research outputs
#24,089,247
of 34,361,833 outputs
Outputs from American Journal of Physiology: Lung Cellular & Molecular Physiology
#3,252
of 3,943 outputs
Outputs of similar age
#200,320
of 305,208 outputs
Outputs of similar age from American Journal of Physiology: Lung Cellular & Molecular Physiology
#30
of 49 outputs
Altmetric has tracked 34,361,833 research outputs across all sources so far. This one is in the 28th percentile – i.e., 28% of other outputs scored the same or lower than it.
So far Altmetric has tracked 3,943 research outputs from this source. They typically receive a little more attention than average, with a mean Attention Score of 6.6. This one is in the 15th percentile – i.e., 15% of its peers scored the same or lower than it.
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We're also able to compare this research output to 49 others from the same source and published within six weeks on either side of this one. This one is in the 32nd percentile – i.e., 32% of its contemporaries scored the same or lower than it.
