Tumor necrosis factor-alpha increases airway smooth muscle oxidants production through a NADPH oxidase-like system to enhance myosin light chain phosphorylation and contractility.
Tumor necrosis factor plays a critical role in airway smooth muscle hyperresponsiveness observed in asthma. However, the mechanisms underlying this phenomenon are poorly understood. We investigated if tumor necrosis factor-stimulated airway smooth muscle produced reactive oxygen species, leading to muscular hyperresponsiveness. Tumor necrosis factor increased intracellular and extracellular oxidants production in guinea pig airway smooth muscle cells and tissue homogenates. This production was abolished by inhibitors of NADPH oxidase (diphenylene iodinium or apocynin) and was enhanced by NADPH, whereas inhibitors of mitochondrial respiratory chain, nitric-oxide synthase, cyclooxygenase, and xanthine oxidase had no effect. NADPH oxidase subunits p22(phox) and p47(phox) were detected in smooth muscle cells and tissue homogenates by Western blot, immunohistochemistry, and spectral analysis. Furthermore, oxidants production was significantly reduced by transient transfection of smooth muscle cells with p22(phox) antisense oligonucleotides. Intracellular antioxidants and diphenylene iodinium abolished tumor necrosis factor-induced muscular hyperresponsiveness and increased in phosphorylation of the myosin light chain. Finally, NADPH oxidase subunits p22(phox) and p47(phox) were also detected in human airway smooth muscle. Collectively, these results demonstrate that tumor necrosis factor-stimulated airway smooth muscle produces oxidants through a NADPH oxidase-like system, which plays a pivotal role in muscle hyperresponsiveness and myosin light chain phosphorylation.
Thabut
Gabriel
G
INSERM U408, Faculté de Médecine Xavier Bichat 75870 Paris Cedex 18, France.
El-Benna
Jamel
J
Samb
Abdoulaye
A
Corda
Stephano
S
Megret
Jerôme
J
Leseche
Guy
G
Vicaut
Eric
E
Aubier
Michel
M
Boczkowski
Jorge
J
D000818:Animals; D000838:Anions; D015153:Blotting, Western; D002478:Cells, Cultured; D003573:Cytochrome b Group / Q000378:metabolism; D006168:Guinea Pigs; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D026901:Membrane Transport Proteins; D009130:Muscle, Smooth / Q000166:cytology / Q000378:metabolism*; D018994:Myosin Light Chains / Q000737:chemistry / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D009252:NADPH Dehydrogenase / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017382:Reactive Oxygen Species; D014132:Trachea / Q000378:metabolism; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
D000838:Anions; D003573:Cytochrome b Group; D026901:Membrane Transport Proteins; D018994:Myosin Light Chains; D016376:Oligonucleotides, Antisense; D010750:Phosphoproteins; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D009249:NADP; C036888:cytochrome b558; D019255:NADPH Oxidases; C086986:CYBA protein, human; C060505:neutrophil cytosolic factor 1; D009252:NADPH Dehydrogenase; D010100:Oxygen