BACKGROUND/AIMS: The human iNOS promoter contains a consensus sequence for binding the hypoxia inducible factor. The aim of this study was to see whether iNOS gene expression is triggered by oxygen tension in rat hepatocytes exposed in vivo to high (periportal) and low (perivenous) oxygen tension. METHODS: Hepatocytes transfected or not with a plasmid containing rat iNOS promoter linked to chloramphenicol acetyltransferase were cultured at 21% and 5% oxygen tension. In normal hepatocytes, iNOS protein, mRNA and activity were detected. In transfected cells, chloramphenicol acetyltransferase activity was measured. RESULTS: In cells cultured in a hypoxic environment, both iNOS protein and mRNA increased, whereas the nitrite level in the medium decreased. However, electron paramagnetic resonance analysis and in vitro iNOS activity indicated that iNOS was active. Transfection experiments showed that the expression of chloramphenicol acetyltransferase driven by iNOS promoter was increased in cells maintained at low oxygen tension. CONCLUSIONS: Our experiments show that in rat hepatocytes: 1) iNOS is induced by low oxygen tension; 2) the modification occurs at the transcriptional level; 3) the enzyme at 5% oxygen is able to catalyze the synthesis of NO, although no nitrites are accumulated in the medium. These findings could have physiopathological relevance, e.g. in determining the resistance of perivenous hepatocytes to ischemia injury.

Oxygen regulation of rat hepatocyte iNOS gene expression.

CRAVANZOLA, Carlo;GRILLO, Maria Angelica;COLOMBATTO, Sebastiano
2000

Abstract

BACKGROUND/AIMS: The human iNOS promoter contains a consensus sequence for binding the hypoxia inducible factor. The aim of this study was to see whether iNOS gene expression is triggered by oxygen tension in rat hepatocytes exposed in vivo to high (periportal) and low (perivenous) oxygen tension. METHODS: Hepatocytes transfected or not with a plasmid containing rat iNOS promoter linked to chloramphenicol acetyltransferase were cultured at 21% and 5% oxygen tension. In normal hepatocytes, iNOS protein, mRNA and activity were detected. In transfected cells, chloramphenicol acetyltransferase activity was measured. RESULTS: In cells cultured in a hypoxic environment, both iNOS protein and mRNA increased, whereas the nitrite level in the medium decreased. However, electron paramagnetic resonance analysis and in vitro iNOS activity indicated that iNOS was active. Transfection experiments showed that the expression of chloramphenicol acetyltransferase driven by iNOS promoter was increased in cells maintained at low oxygen tension. CONCLUSIONS: Our experiments show that in rat hepatocytes: 1) iNOS is induced by low oxygen tension; 2) the modification occurs at the transcriptional level; 3) the enzyme at 5% oxygen is able to catalyze the synthesis of NO, although no nitrites are accumulated in the medium. These findings could have physiopathological relevance, e.g. in determining the resistance of perivenous hepatocytes to ischemia injury.
32(4)
567
573
VARGIU C; BELLIARDO S; CRAVANZOLA C; GRILLO MA; COLOMBATTO S
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2318/39050
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