Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line
Background/Aims: Stabilization of the hypoxia-inducible factor (HIF-1α) is proposed to provide a protective host-response to C. difficile intoxication. Here, we aimed to elucidate whether nitric oxide and/or reactive oxygen species produced during C. difficile toxin exposure could influence HIF-1α s...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Cell Physiol Biochem Press GmbH & Co KG
2013-08-01
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| Series: | Cellular Physiology and Biochemistry |
| Subjects: | |
| Online Access: | http://www.karger.com/Article/FullText/354448 |
| _version_ | 1811219681465335808 |
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| author | Joshua Y. Lee Simon A. Hirota Louise E. Glover Glen D. Armstrong Paul L. Beck Justin A. MacDonald |
| author_facet | Joshua Y. Lee Simon A. Hirota Louise E. Glover Glen D. Armstrong Paul L. Beck Justin A. MacDonald |
| author_sort | Joshua Y. Lee |
| collection | DOAJ |
| description | Background/Aims: Stabilization of the hypoxia-inducible factor (HIF-1α) is proposed to provide a protective host-response to C. difficile intoxication. Here, we aimed to elucidate whether nitric oxide and/or reactive oxygen species produced during C. difficile toxin exposure could influence HIF-1α stability and initiate protection against epithelial cell damage. Methods/Results: HIF-1α and inducible nitric oxide synthase (iNOS) proteins were up-regulated whereas factor-inhibiting HIF-1 (FIH-1) protein was down-regulated in Caco-2 epithelial cell monolayers with in vitro toxin exposure. We demonstrate using the biotin-switch assay that the stabilization of HIF-1α protein occurred via iNOS-dependent nitrosylation. Inhibition of iNOS activity by selective inhibitor (1400W) attenuated HIF-1α stabilization and exacerbated toxin-dependent disruptions in Caco-2 monolayer morphology and tight junctional integrity in vitro. Treatment of Caco-2 cell monolayers with N-actylcysteine (NAC), a scavenger of reactive oxygen species (ROS), attenuated toxin-dependent increases in iNOS and HIF-1α protein levels but had no effect on FIH-1 responses. In addition, mice that were exposed to C. difficile toxin in vivo also demonstrated a significant increase in HIF-1α protein and nitrosylation levels. Conclusion: Taken together, these data suggest that important synergistic actions exist between nitric oxide and ROS to stabilize HIF-1α and its innate, protective actions in the context of C. difficile toxin-mediated epithelial injury. |
| first_indexed | 2024-04-12T07:30:02Z |
| format | Article |
| id | doaj.art-24681ee2f2944602bad318ab2c2da620 |
| institution | Directory Open Access Journal |
| issn | 1015-8987 1421-9778 |
| language | English |
| last_indexed | 2024-04-12T07:30:02Z |
| publishDate | 2013-08-01 |
| publisher | Cell Physiol Biochem Press GmbH & Co KG |
| record_format | Article |
| series | Cellular Physiology and Biochemistry |
| spelling | doaj.art-24681ee2f2944602bad318ab2c2da6202022-12-22T03:42:05ZengCell Physiol Biochem Press GmbH & Co KGCellular Physiology and Biochemistry1015-89871421-97782013-08-0132241743010.1159/000354448354448Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell LineJoshua Y. LeeSimon A. HirotaLouise E. GloverGlen D. ArmstrongPaul L. BeckJustin A. MacDonaldBackground/Aims: Stabilization of the hypoxia-inducible factor (HIF-1α) is proposed to provide a protective host-response to C. difficile intoxication. Here, we aimed to elucidate whether nitric oxide and/or reactive oxygen species produced during C. difficile toxin exposure could influence HIF-1α stability and initiate protection against epithelial cell damage. Methods/Results: HIF-1α and inducible nitric oxide synthase (iNOS) proteins were up-regulated whereas factor-inhibiting HIF-1 (FIH-1) protein was down-regulated in Caco-2 epithelial cell monolayers with in vitro toxin exposure. We demonstrate using the biotin-switch assay that the stabilization of HIF-1α protein occurred via iNOS-dependent nitrosylation. Inhibition of iNOS activity by selective inhibitor (1400W) attenuated HIF-1α stabilization and exacerbated toxin-dependent disruptions in Caco-2 monolayer morphology and tight junctional integrity in vitro. Treatment of Caco-2 cell monolayers with N-actylcysteine (NAC), a scavenger of reactive oxygen species (ROS), attenuated toxin-dependent increases in iNOS and HIF-1α protein levels but had no effect on FIH-1 responses. In addition, mice that were exposed to C. difficile toxin in vivo also demonstrated a significant increase in HIF-1α protein and nitrosylation levels. Conclusion: Taken together, these data suggest that important synergistic actions exist between nitric oxide and ROS to stabilize HIF-1α and its innate, protective actions in the context of C. difficile toxin-mediated epithelial injury.http://www.karger.com/Article/FullText/354448CDIEpithelial permeabilityIntestinal inflammationHypoxia-inducible factorNitrosylationiNOS |
| spellingShingle | Joshua Y. Lee Simon A. Hirota Louise E. Glover Glen D. Armstrong Paul L. Beck Justin A. MacDonald Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line Cellular Physiology and Biochemistry CDI Epithelial permeability Intestinal inflammation Hypoxia-inducible factor Nitrosylation iNOS |
| title | Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line |
| title_full | Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line |
| title_fullStr | Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line |
| title_full_unstemmed | Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line |
| title_short | Effects of Nitric Oxide and Reactive Oxygen Species on HIF-1α Stabilization Following Clostridium Difficile Toxin Exposure of the Caco-2 Epithelial Cell Line |
| title_sort | effects of nitric oxide and reactive oxygen species on hif 1α stabilization following clostridium difficile toxin exposure of the caco 2 epithelial cell line |
| topic | CDI Epithelial permeability Intestinal inflammation Hypoxia-inducible factor Nitrosylation iNOS |
| url | http://www.karger.com/Article/FullText/354448 |
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