Hyperoxia-induced NF-kappaB activation occurs via a maturationally sensitive atypical pathway.
NF-kappaB activation is exaggerated in neonatal organisms after oxidant and inflammatory insults, but the reason for this and the downstream effects are unclear. We hypothesized that specific phosphorylation patterns of IkappaBalpha could account for differences in NF-kappaB activation in hyperoxia-exposed fetal and adult lung fibroblasts. After exposure to hyperoxia (>95% O(2)), nuclear NF-kappaB binding increased in fetal, but not adult, lung fibroblasts. Unique to fetal cells, phosphorylation of IkappaBalpha on tyrosine 42, rather than serine 32/36 as seen in TNF-alpha-exposed cells, preceded NF-kappaB nuclear translocation. In fetal cells stably transfected with an NF-kappaB-driven luciferase reporter, hyperoxia significantly suppressed reporter activity, in contrast to increased reporter activity after TNF-alpha incubation. Targeted gene profiling analysis showed that hyperoxia resulted in decreased expression of multiple genes, including proapoptotic factors. Transfection with a dominant-negative IkappaBalpha (Y42F), which cannot be phosphorylated on tyrosine 42, resulted in upregulation of multiple proapoptotic genes. In support of this finding, caspase-3 activity and DNA laddering were specifically increased in fetal lung fibroblasts expressing Y42F after exposure to hyperoxia. These data demonstrate a unique pathway of NF-kappaB activation in fetal lung fibroblasts after exposure to hyperoxia, whereby these cells are protected against apoptosis. Activation of this pathway in fetal cells may prevent the normal pattern of fibroblast apoptosis necessary for normal lung development, resulting in aberrant lung morphology in vivo.
Wright
Clyde J
CJ
Division of Neonatology, Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
Zhuang
Tiangang
T
La
Ping
P
Yang
Guang
G
Dennery
Phyllis A
PA
D019943:Amino Acid Substitution; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D001483:Base Sequence; D002454:Cell Differentiation; D002460:Cell Line; D004247:DNA / Q000235:genetics; D005333:Fetus / Q000166:cytology / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D018496:Hyperoxia / Q000378:metabolism*; D020672:I-kappa B Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D008168:Lung / Q000166:cytology* / Q000378:metabolism*; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry
D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606206:Nfkbia protein, rat; D011994:Recombinant Proteins; D000072000:NF-KappaB Inhibitor alpha; D014443:Tyrosine; D004247:DNA; D008156:Luciferases