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Glutamine attenuates nitric oxide synthase expression and mitochondria membrane potential decrease in interleukin-1β-activated rat hepatocytes

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Abstract

Background

Mitochondrial dysfunction induced by nitric oxide (NO) overproduction is involved in the pathogenesis of organ failure during many severe diseases. Recently, several experiments have reported that glutamine (Gln) modifies inducible nitric oxide synthase (iNOS) gene expression which is thought to be responsible for its beneficial effects in critical illnesses.

Aim of the study

This study was designed to evaluate the effects of Gln on NO production, iNOS expression and mitochondria membrane potential (Δψm) variation in interleukin (IL)-1β-activated rat hepatocytes. The mechanical effects of nuclear factor kappaB (NF-κB) were also investigated.

Methods

Primary cultured rat hepatocytes were isolated by an in situ collagenase perfusion method. The cultured hepatocytes were treated with IL-1β (1 nmol/l) alone or with Gln in concentrations ranging from 2 to 10 mmol/l for 24 h. The concentrations of alanine aminotransferase (ALT) and nitrite in the culture medium were detected by biochemical methods. The levels of iNOS protein and mRNA in the hepatocytes were analyzed by western blot and real-time RT-PCR. The Δψm of the hepatocytes was assessed with flow cytometry after incubating with fluorescent probe JC-1. Th binding activity of NF-κB in hepatocytes was investigated using electrophoretic mobility shift assay.

Results

IL-1β significantly enhanced iNOS protein and mRNA level in the cultured rat hepatocytes which consisted with the increased NO production and ALT releasing in the culture medium. These effects were inhibited by the treatment of glutamine in a dose-dependent manner. The Δψm of hepatocytes was markedly decreased after IL-1β stimulation which was significantly attenuated by Gln at 5 and 10 mmol/l. NF-κB activity was increased by IL-1β stimulation and this effect was augmented by Gln at 5 and 10 mol/l.

Conclusions

These observations indicate that Gln has a capability to protect mitochondria function in hepatocytes under inflammatory stress. Although the molecular mechanism remains to be clarified, our findings suggest this effect may be related to down-regulation of iNOS gene expression through an NF-κB independent pathway.

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The authors have no commercial associations or sources of support that might pose a conflict of interest.

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Correspondence to Wen-hao Tang.

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Lu, J., Wang, Xy. & Tang, Wh. Glutamine attenuates nitric oxide synthase expression and mitochondria membrane potential decrease in interleukin-1β-activated rat hepatocytes. Eur J Nutr 48, 333–339 (2009). https://doi.org/10.1007/s00394-009-0018-x

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  • DOI: https://doi.org/10.1007/s00394-009-0018-x

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