Acute hypoxia simultaneously induces the expression of gp91phox and endothelial nitric oxide synthase in the porcine pulmonary artery.
BACKGROUND
The effect of hypoxia on the formation of superoxide (O2-), the expression of gp91phox and endothelial NO synthase (eNOS) were studied in pig intact pulmonary artery (PA) segments and PA vascular smooth muscle cells (PAVSMCs) and PA endothelial cells (PAECs).


METHODS
Segments and cells were incubated under hypoxic conditions for 2 hours (with or without enzyme inhibitors) and the formation of O2- measured spectrophotometrically. Protein expression was assessed using Western blotting and immunocytochemistry.


RESULTS
Hypoxia promoted the formation of O2- in PA segments, PAVSMCs and PAECs, an effect inhibited by diphenylene iodonium and apocynin (NAD[P]H oxidase inhibitors). Hypoxia induced O2- formation was enhanced by inhibition of eNOS and augmented by endotoxin and cytokines and re-oxygenation. Hypoxia also promoted the expression of gp91phox and eNOS. In intact PA segments hypoxia induced the expression of nitrotyrosine and eNOS in the endothelium.


CONCLUSIONS
The simultaneous upregulation of NAD[P]H oxidase and eNOS in response to hypoxia in the PA results in the simultaneous formation of O2-, NO, and ONOO-. This may represent either a protective mechanism designed to counter the pro-oxidant effect of hypoxia or a novel pathological mechanism underlying the progression of acute respiratory distress syndrome (ARDS).
Muzaffar
S
S
Bristol Heart Institute, Bristol Royal Infirmary, Bristol BS2 8HW, UK.
Shukla
N
N
Angelini
G D
GD
Jeremy
J Y
JY
D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000378:metabolism; D000860:Hypoxia / Q000378:metabolism*; D007150:Immunohistochemistry; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III; D009573:Nitrites / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011651:Pulmonary Artery / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D013552:Swine
D016207:Cytokines; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D009573:Nitrites; D013481:Superoxides; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D010100:Oxygen