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Suppression of the Nrf2-dependent antioxidant response by glucocorticoids and 11?-HSD1-mediated glucocorticoid activation in hepatic cells.

ABSTRACT:

Background

Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a key transcription factor regulating a plethora of detoxifying enzymes and antioxidant genes involved in drug metabolism and defence against oxidative stress. The glucocorticoid receptor (GR) is a ligand-induced transcription factor involved in the regulation of energy supply for metabolic needs to cope with various stressors. GR activity is controlled by glucocorticoids, which are synthesized in the adrenal glands and regenerated mainly in the liver from inactive cortisone by 11?-hydroxysteroid dehydrogenase-1 (11?-HSD1).

Methods and principal findings

Using transfected HEK-293 cells and hepatic H4IIE cells we show that glucocorticoids, activated by 11?-HSD1 and acting through GR, suppress the Nrf2-dependent antioxidant response. The expression of the marker genes NQO1, HMOX1 and GST2A was suppressed upon treatment of 11?-HSD1 expressing cells with cortisone, an effect that was reversed by 11?-HSD1 inhibitors. Furthermore, our results demonstrate that elevated glucocorticoids lowered the ability of cells to detoxify H(2)O(2). Moreover, a comparison of gene expression in male and female rats revealed an opposite sexual dimorphism with an inverse relationship between 11?-HSD1 and Nrf2 target gene expression.

Conclusions

The results demonstrate a suppression of the cellular antioxidant defence capacity by glucocorticoids and suggest that elevated 11?-HSD1 activity may lead to impaired Nrf2-dependent antioxidant response. The gender-specific differences in hepatic expression levels of 11?-HSD1 and Nrf2 target genes and the impact of pharmacological inhibition of 11?-HSD1 on improving cellular capacity to cope with oxidative stress warrants further studies in vivo.

SUBMITTER: Kratschmar DV

PROVIDER: S-EPMC3350474 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Publications

Suppression of the Nrf2-dependent antioxidant response by glucocorticoids and 11β-HSD1-mediated glucocorticoid activation in hepatic cells.

Kratschmar Denise V DV Calabrese Diego D Walsh Jo J Lister Adam A Birk Julia J Appenzeller-Herzog Christian C Moulin Pierre P Goldring Chris E CE Odermatt Alex A

PloS one 20120511 5

<h4>Background</h4>Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a key transcription factor regulating a plethora of detoxifying enzymes and antioxidant genes involved in drug metabolism and defence against oxidative stress. The glucocorticoid receptor (GR) is a ligand-induced transcription factor involved in the regulation of energy supply for metabolic needs to cope with various stressors. GR activity is controlled by glucocorticoids, which are synthesized in the adrenal glands and reg ...[more]

PMID: 22606287

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Project description:BackgroundIntracellular metabolism of glucocorticoid hormones plays an important role in the pathogenesis of metabolic syndrome and regulates, among many physiological processes, collagen metabolism in skin. At the peripheral level the concentration of active glucocorticoids is mainly regulated by the 11?-hydroxysteroid dehydrogenase 1 (11?-HSD1) enzyme, involved in the conversion of cortisone into the biologically active hormone cortisol. Cortisol interacts with the glucocorticoid receptor and regulates the expression of different classes of genes within the nucleus. Due to its implication in glucocorticoid metabolism, the inhibition of 11?-HSD1 activity has become a dominant strategy for the treatment of metabolic syndrome. Moreover, inhibitors of this target enzyme can be used for development of formulations to counteract skin ageing. Here we present the construction of two yeast cell based assays that can be used for the screening of novel 11?-HSD1 inhibitors.ResultsThe yeast Saccharomyces cerevisiae is used as a host organism for the expression of human 11?-HSD1 as well as a genetically encoded assay system that allows intracellular screening of molecules with 11?-HSD1 inhibitory activity. As proof of concept the correlation between 11?-HSD1 inhibition and fluorescent output signals was successfully tested with increasing concentrations of carbenoxolone and tanshinone IIA, two known 11?-HSD1 inhibitors. The first assay detects a decrease in fluorescence upon 11?-HSD1 inhibition, whereas the second assay relies on stabilization of yEGFP upon inhibition of 11?-HSD1, resulting in a positive read-out and thus minimizing the rate of false positives sometimes associated with read-outs based on loss of signals. Specific inhibition of the ABC transporter Pdr5p improves the sensitivity of the assay strains to cortisone concentrations by up to 60 times.ConclusionsOur yeast assay strains provide a cost-efficient and easy to handle alternative to other currently available assays for the screening of 11?-HSD1 inhibitors. These assays are designed for an initial fast screening of large numbers of compounds and enable the selection of cell permeable molecules with target inhibitory activity, before proceeding to more advanced selection processes. Moreover, they can be employed in yeast synthetic biology platforms to reconstitute heterologous biosynthetic pathways of drug-relevant scaffolds for simultaneous synthesis and screening of 11?-HSD1 inhibitors at intracellular level.