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B Activation and TNF-
Gene Transcription Induced by Hypoxia and Endotoxin1
Section of Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, IL 60637
The transcription factor NF-
B stimulates the transcription of
proinflammatory cytokines including TNF-
. LPS (endotoxin) and
hypoxia both induce NF-B activation and TNF- gene transcription.
Furthermore, hypoxia augments LPS induction of TNF- mRNA. Previous
reports have indicated that antioxidants abolish NF-B activation in
response to LPS or hypoxia, which suggests that reactive oxygen species
(ROS) are involved in NF-B activation. This study tested whether
mitochondrial ROS are required for both NF-B activation and the
increase in TNF- mRNA levels during hypoxia and LPS. Our results
indicate that hypoxia (1.5% O2) stimulates NF-B and
TNF- gene transcription and increases ROS generation as measured by
the oxidant sensitive dye 2',7'-dichlorofluorescein diacetate in murine
macrophage J774.1 cells. The antioxidants
N-acetylcysteine and pyrrolidinedithiocarbamic acid
abolished the hypoxic activation of NF-B, TNF- gene
transcription, and increases in ROS levels. Rotenone, an inhibitor of
mitochondrial complex I, abolished the increase in ROS signal, the
activation of NF-B, and TNF- gene transcription during hypoxia.
LPS stimulated NF-B and TNF- gene transcription but not ROS
generation in J774.1 cells. Rotenone, pyrrolidinedithiocarbamic acid,
and N-acetylcysteine had no effect on the LPS
stimulation of NF-B and TNF- gene transcription, indicating that
LPS activates NF-B and TNF- gene transcription through a
ROS-independent mechanism. These results indicate that mitochondrial
ROS are required for the hypoxic activation of NF-B and TNF- gene
transcription, but not for the LPS activation of
NF-B.
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