Brief hypoxia differentially regulates LPS-induced IL-1beta and TNF-alpha gene transcription in RAW 264.7 cells.

Abstract

Episodes of tissue hypoxia and reoxygenation frequently occur during gram-negative bacteremia that progresses to septic shock. However, few studies have evaluated modulation by hypoxia and reoxygenation of the proinflammatory cytokine gene expression that is normally induced by gram-negative bacteremia or endotoxemia. In buffer-perfused organs, hypoxia downregulates Escherichia coli-induced expression of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta in the liver but upregulates these cytokines in the lungs. To identify molecular mechanisms underlying these events, we investigated the effects of brief (1.5-h) hypoxia on TNF-alpha and IL-1beta expression in cultured RAW 264.7 cells during their continuous exposure to lipopolysaccharide (LPS) endotoxin derived from E. coli (serotype 055:B5) for up to 24 h. IL-1beta and TNF-alpha concentrations in cell lysates and culture supernatants were measured by ELISA, and steady-state mRNA was measured by Northern analysis. LPS-induced IL-1beta synthesis was downregulated by hypoxia at both the protein and mRNA levels despite no change in cellular redox status as measured by levels of GSH. In contrast, LPS-induced TNF-alpha production was unaffected by hypoxia as assessed by cell lysate mRNA and lysate and supernatant protein levels. Nuclear runoff analysis showed that downregulation of IL-1beta gene expression by hypoxia occurred transcriptionally. Allopurinol or catalase treatment did not alter modulation of LPS-induced IL-1beta expression by hypoxia, suggesting that this suppression was not caused by reactive oxygen species. Cycloheximide pretreatment suggested that hypoxia-induced downregulation of IL-1beta expression did not require de novo protein synthesis.

• Authors: M M Ndengele, C J Bellone, A J Lechner, G M Matuschak
• Journal: American journal of physiology. Lung cellular and molecular physiology (Am J Physiol Lung Cell Mol Physiol) Vol. 278 Issue 6 Pg. L1289-96 (Jun 2000) ISSN: 1040-0605 [Print] United States
• PMID: 10835336 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Carbocyanines
• Interleukin-1
• Lipopolysaccharides
• RNA, Messenger
• Tumor Necrosis Factor-alpha
• Catalase

Topics

• Animals
• Carbocyanines (pharmacology)
• Catalase (pharmacology)
• Cell Line
• Homeostasis
• Hypoxia (genetics)
• Interleukin-1 (antagonists & inhibitors, genetics)
• Lipopolysaccharides (pharmacology)
• Mice
• RNA, Messenger (antagonists & inhibitors, metabolism)
• Transcription, Genetic
• Tumor Necrosis Factor-alpha (genetics, metabolism)