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AJP - Gastrointestinal and Liver Physiology, Vol 272, Issue 6 1499-G1508, Copyright © 1997 by American Physiological Society
ARTICLES |
S. C. Linn, P. J. Morelli, I. Edry, S. E. Cottongim, C. Szabo and A. L. Salzman
Division of Critical Care Medicine, Children's Hospital Medical Center, Cincinnati, Ohio 45229, USA.
The inducible form of nitric oxide synthase (iNOS) is expressed during inflammation of the intestine and may contribute to tissue injury. We have examined iNOS transcriptional regulation in DLD-1 cells, a human intestinal epithelial line that produces large amounts of nitric oxide and iNOS mRNA in response to a combination of the proinflammatory cytokines interleukin-1 beta (IL-1 beta) and interferon-gamma (IFN-gamma). Levels of iNOS mRNA are extremely low in unstimulated DLD-1 cells but increase dramatically after cytokine treatment. Nuclear run-on analyses demonstrated that transcriptional activation, which accounts for a portion of this increase, is dependent on both IL-1 beta and IFN-gamma and requires de novo protein synthesis. Transfection of DLD-1 cells with reporter constructs containing deletions of the iNOS promoter showed that sequences located between 8.7 and 10.7 kb upstream of the transcription initiation site are necessary for cytokine responsiveness. This region contains potential binding sites for several cytokine-induced transcription factors and was shown to function in either orientation when placed upstream of a basal iNOS promoter segment terminating at-1.1 kb. The extremely distal location of the cytokine-responsive region contrasts with the reported positions of elements involved in the regulation of iNOS transcription in other cell types. Our data also suggest that posttranscriptional events could play a significant role in regulating iNOS gene expression in human intestinal epithelia.
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