Septic pulmonary microvascular endothelial cell injury: role of alveolar macrophage NADPH oxidase.
A significant role for alveolar macrophages (AM) in the pathophysiology of sepsis-induced acute lung injury (ALI) has been shown; however, the mechanisms behind AM-related lung injury remain relatively uncertain. We examined the role of AM nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in pulmonary endothelial cell septic injury. NADPH oxidase is one of the major sources of cellular reactive oxygen species and has been implicated in endothelial injury in ALI. Pulmonary microvascular endothelial cells (PMVEC) monolayers were grown on Transwell inserts and incubated with wild-type and NADPH oxidase-deficient AM in the presence or absence of cytomix (equimolar TNF-alpha, IL-1beta, and IFN-gamma). Injury to the monolayers was assessed by trans-PMVEC Evans blue (EB)-labeled albumin flux. We found AM under cytomix stimulation caused significant EB-albumin flux across the PMVEC monolayers, and this effect was attenuated by the genetic deletion of AM NADPH oxidase. The pharmacological inhibition of AM NADPH oxidase with apocynin and PR-39 also significantly reduced AM-dependent PMVEC injury. In the AM-PMVEC cocultures, we also assessed PMVEC injury through measurement of protein oxidation and lipid peroxidation. AM were shown to cause a significant increase in these markers of PMVEC injury, which was also attenuated by the inhibition of NADPH oxidase or through the use of NADPH oxidase-deficient AM. PMVEC NADPH oxidase was shown not to significantly contribute to PMVEC injury in our studies. From our findings we have concluded that AM NADPH oxidase is crucial for the septic increase in pulmonary vascular permeability.
Farley
K S
KS
Division of Respirology, Centre for Critical Illness Research, Lawson Health Research Institute, University Of Western Ontario, London, Ontario, Canada.
Wang
L
L
Mehta
S
S
D055371:Acute Lung Injury / Q000201:enzymology* / Q000473:pathology; D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000473:pathology; D008168:Lung / Q000098:blood supply / Q000201:enzymology / Q000473:pathology; D016676:Macrophages, Alveolar / Q000201:enzymology* / Q000473:pathology; D008297:Male; D008562:Membrane Glycoproteins / Q000172:deficiency / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008833:Microcirculation; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress; D018805:Sepsis / Q000201:enzymology* / Q000473:pathology; D012709:Serum Albumin / Q000378:metabolism
D008562:Membrane Glycoproteins; D012709:Serum Albumin; C488842:Cybb protein, mouse; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1