TY - JOUR
T1 - Role of oxidants in NF-κB activation and TNF-α gene transcription induced by hypoxia and endotoxin
AU - Chandel, Navdeep S.
AU - Trzyna, Wendy C.
AU - McClintock, David S.
AU - Schumacker, Paul T.
PY - 2000/7/15
Y1 - 2000/7/15
N2 - The transcription factor NF-κB stimulates the transcription of proinflammatory cytokines including TNF-α. LPS (endotoxin) and hypoxia both induce NF-κB activation and TNF-α gene transcription. Furthermore, hypoxia augments LPS induction of TNF-α mRNA. Previous reports have indicated that antioxidants abolish NF-κB activation in response to LPS or hypoxia, which suggests that reactive oxygen species (ROS) are involved in NF-κB activation. This study tested whether mitochondrial ROS are required for both NF-κB activation and the increase in TNF-α mRNA levels during hypoxia and LPS. Our results indicate that hypoxia (1.5% O2) stimulates NF-κB and TNF- α 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-κB, TNF-α gene transcription, and increases in ROS levels. Rotenone, an inhibitor of mitochondrial complex I, abolished the increase in ROS signal, the activation of NF-κB, and TNF-α gene transcription during hypoxia. LPS stimulated NF-κB and TNF-α 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-κB and TNF-α gene transcription, indicating that LPS activates NF-κB and TNF-α gene transcription through a ROS-independent mechanism. These results indicate that mitochondrial ROS are required for the hypoxic activation of NF-κB and TNF-α gene transcription, but not for the LPS activation of NF-κB.
AB - The transcription factor NF-κB stimulates the transcription of proinflammatory cytokines including TNF-α. LPS (endotoxin) and hypoxia both induce NF-κB activation and TNF-α gene transcription. Furthermore, hypoxia augments LPS induction of TNF-α mRNA. Previous reports have indicated that antioxidants abolish NF-κB activation in response to LPS or hypoxia, which suggests that reactive oxygen species (ROS) are involved in NF-κB activation. This study tested whether mitochondrial ROS are required for both NF-κB activation and the increase in TNF-α mRNA levels during hypoxia and LPS. Our results indicate that hypoxia (1.5% O2) stimulates NF-κB and TNF- α 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-κB, TNF-α gene transcription, and increases in ROS levels. Rotenone, an inhibitor of mitochondrial complex I, abolished the increase in ROS signal, the activation of NF-κB, and TNF-α gene transcription during hypoxia. LPS stimulated NF-κB and TNF-α 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-κB and TNF-α gene transcription, indicating that LPS activates NF-κB and TNF-α gene transcription through a ROS-independent mechanism. These results indicate that mitochondrial ROS are required for the hypoxic activation of NF-κB and TNF-α gene transcription, but not for the LPS activation of NF-κB.
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U2 - 10.4049/jimmunol.165.2.1013
DO - 10.4049/jimmunol.165.2.1013
M3 - Article
C2 - 10878378
AN - SCOPUS:0034661990
VL - 165
SP - 1013
EP - 1021
JO - Journal of Immunology
JF - Journal of Immunology
SN - 0022-1767
IS - 2
ER -