Nonalcoholic steatohepatitis can be ameliorated by reactivating the CNC-bZIP transcription factor Nrf2


Fat in the liver typically develops when an individual consumes more fat and sugar than the body can handle. It is usually observed in people who are overweight or obese. If fat represents more than 5% of the liver mass, then it is a fatty liver. Whilst this condition probably does not cause any immediate harm, the extra fat may render the liver vulnerable to injury and inflammation that results in non-alcoholic steatohepatitis (NASH). The long-term outlook of people with NASH is not well understood, though emerging epidemiological evidence from Scottish National Health Service records indicates that NASH is a risk factor for cancer. At present, there is no proven effective medication for NASH. Since it is now a common health condition, NASH has raised considerable interest amongst scientists seeking to understand the pathophysiology of the disease.

Mechanistically, NASH has been linked to endoplasmic reticulum stress, mitochondrial dysfunction, oxidative stress, lipotoxicity and iron overload, all of which can be countered by activation of the CNC-bZIP transcription factor NF-E2 p45-related factor 2 (Nrf2). Many studies have revealed Nrf2 to be a master regulator of gene expression that enables cells to adapt to the presence of a wide spectrum of electrophiles and oxidative stressors by upregulating proteins with antioxidant, detoxification, anti-inflammatory and mitochondrial metabolic functions. In a paper published in the peer-reviewed journal Free Radical Biology and Medicine, University of Dundee scientists Dr Boushra Bathish, Dr Holly Robertson, Professor John Dillon, Professor Albena Dinkova-Kostova, and Professor John Hayes have provided an expert opinion review of the various mechanisms that contribute to NASH, the molecular regulation of Nrf2 transcriptional activity and the biochemical actions of the genes controlled by Nrf2. Moreover, they have evaluated in vivo evidence that activation of Nrf2 can arrest, and possibly reverse, the development of NASH. It is noteworthy that Professor Hayes and colleagues are internationally recognized for their research into the molecular mechanisms that regulate Nrf2 and the potential of pharmacological manipulation of Nrf2 activity to treat NASH.

According to the authors, Nrf2-knockout mice are especially vulnerable to NASH, whilst genetic or pharmacological activation of Nrf2 boosts resistance to NASH. Importantly, they also discuss evidence that the development and progression of NASH is accompanied by downregulation of Nrf2, with the likelihood that Nrf2 downregulation represents an integral component of the pathogenesis of the disease. Recognition that Nrf2 downregulation accompanies NASH, suggests that restoring Nrf2 activity to its normal levels might provide a viable therapeutic strategy. Puzzlingly, during the early stages of the disease, diets that cause NASH appear to activate Nrf2. Such upregulation of Nrf2 activity during the initial onset of disease occurs because the oxidative stress and lipid peroxidation components, generated in the fatty liver, inactivate the Nrf2 repressor Keap1, which under non-stressed conditions continuously targets Nrf2 for proteasomal degradation by mediating its ubiquitylation. In the longer term however, persistent consumption of NASH-inducing diets results in a significant reduction in Nrf2 activity, under conditions when Keap1 is unlikely to repress Nrf2, suggesting that Nrf2 is subject to additional mechanisms of suppression that are poorly understood. The authors have detailed alternative mechanisms of Nrf2 suppression, such as ubiquitylation of Nrf2 directed by β-TrCP that is initiated by activation of MAPKs, or ubiquitylation of Nrf2 upon activation of Hrd1/SYVN1 by endoplasmic reticulum stress, and have proposed that these additional ways of ubiquitylating Nrf2 probably contribute to the progression of NASH.

An appealing and perhaps successful method for treating NASH is the activation of Nrf2, possibly in conjunction with other medications. In view of the broad cytoprotective effects of Nrf2 and its role in maintaining redox homeostasis, it is apparent that numerous pathways that contribute to the aetiology of NASH will probably become blunted when Nrf2 is activated. It is well recognized that soft electrophiles that block the ubiquitin ligase substrate adaptor function of Keap1 can induce the expression of Nrf2-target genes, making such compounds potential therapeutic agents for activating Nrf2 and treating NASH. It may also be necessary to use medications that oppose β-TrCP or inhibit MAPKs, and so restore Nrf2 activity to typical cytoprotective levels when MAPKs are activated and downregulate Nrf2. Other ways of treating NASH might include restoring normal homeostatic Nrf2 activity by inhibiting its ubiquitylation by Hrd1/SYVN1 or by preventing competition for the recruitment of coactivators to Nrf2 by ATF3 or by preventing the competition by Bach1 for the binding to Nrf2 recognition sites in gene promoters. Interestingly, some of the proteins that repress Nrf2 are activated by TGF-β signaling, which suggests that the progression of NASH is dictated by crosstalk between Nrf2 and TGF-β signaling.

In a nutshell, Professor Hayes and colleagues have summarized the molecular mechanisms that regulate Nrf2 activity and the pathways it controls, and have discussed the potential therapeutic applications of targeting Nrf2 in NASH. It is hoped in the future further studies to understand and explore the therapeutic impact of Nrf2 will improve the prognosis for NASH patients.

Nonalcoholic steatohepatitis can be ameliorated by reactivating the CNC-bZIP transcription factor Nrf2 - Medicine Innovates
Panel A shows that the expression of antioxidant and cytoprotective genes that are regulated by Nrf2 through an antioxidant response element (ARE) can be downregulated by three broad mechanisms: i) proteasomal degradation of Nrf2 directed by Keap1, β-TrCP or Hrd1 (also called synoviolin 1), presented on a grey background; ii) inhibition of recruitment of RNA pol II and CBP/p300 to Nrf2-small Maf (sMaf) heterodimers bound to ARE sequences, presented on a pale-blue background; iii) increased competition of binding to ARE sequences by the negatively-acting transcription factors Bach1 and Fra-1, or competition for heterodimerisation with sMaf proteins by Bach1, presented on a pink background. Compounds and pharmacological agents that can modulate the different regulators of Nrf2 are depicted in red boxes. Treatment with Ezetimibe (see, top left-hand side), that increases p62/SQSTM1-directed degradation of Keap1 by autophagy, or thiol-reactive agents such as TBE-31 that block the Cul3 substrate-adaptor function of Keap1, or the Keap1 protein-Nrf2 protein interaction inhibitor NK-252, all inhibit the turnover of Nrf2 by Keap1-Cul3 and have all been shown to prevent diet-stimulated NASH in rodents. The DSGIS-containing phosphodegron in Nrf2 that is created by GSK-3 and is subsequently recognised by β-TrCP, likely requires MAP kinases to prime Nrf2 for phosphorylation by GSK-3 (see, top of cartoon), and so inhibition of these protein kinases will stabilise Nrf2 protein and are thus likely to inhibit NASH. The Hrd1 inhibitor LS-102 (see, right-hand side) has been reported to prevent CCl4-initiated liver fibrosis, and it may possibly also prevent NASH. Treatment with Resolvin D1 (see, right-hand side) has been reported to inhibit NASH and so likely prevents NF-kB p65 from blocking the recruitment of RNA pol II and CBP/p300 to Nrf2 located on gene promoters. Treatment with the Bach1 inhibitor hemin can increase the expression of certain Nrf2-target genes, and it may also protect against NASH.
Nonalcoholic steatohepatitis can be ameliorated by reactivating the CNC-bZIP transcription factor Nrf2 - Medicine Innovates
Panel B shows the progressive downregulation of Nrf2 during progression of NASH, from an initial upregulation of Nrf2 as an adaptive response to oxidative stress at the onset of disease, to a marked suppression of Nrf2 during severe disease. It is proposed that activation of JNK and MAP kinases may be responsible for Nrf2 downregulation during mid-stage chronic disease, and that Bach1, ATF3, Hrd1 and NF-B p65 are responsible for Nrf2 downregulation during late-stage chronic disease, which might also include liver fibrosis. According to this scenario (bottom half), early-stage NASH could be treated by agents that inhibit Keap1, mid-stage NASH could be treated by agents that inhibit repression of Nrf2 by Keap1 or β-TrCP (e.g., inhibitors of JNK and MAPK), and late-stage NASH could be treated by agents that inhibit repression of Nrf2 by Keap1, β-TrCP or TGF-β (or proteins that TGF-β signalling activates such as NF-kB, ATF3 and Hrd1). Despite the hierarchical depiction in the cartoon of mechanisms that downregulate Nrf2, it is possible that an agent that is specific for a single mechanism may fully restore Nrf2 to normal homeostatic levels. The figure was created with

About the author

Boushra Bathish is a postdoctoral researcher who is studying redox signalling in liver fibrosis and the potential of pharmacological activation of the cytoprotective transcription factor Nrf2 as a therapeutic approach to liver fibrosis. She completed her doctoral research at the University of Otago in New Zealand, where she studied the enzymology of peroxidasin in the extracellular matrix. Her research interests are redox signalling and extracellular matrix biology and pathology.


About the author

John Hayes holds the chair of Molecular Carcinogenesis in the Medical School at the University of Dundee, where he was until recently associate dean of Postgraduate Student Studies and deputy director of the Medical Research Institute. His research has focused for many years on the mechanisms of adaptation to oxidative stress, with a particular focus on the mechanisms by which the antioxidant transcription factor NRF2 is regulated. Prof Hayes initially trained in the School of Biological Sciences at the University of Edinburgh, and gained a PhD from the Medical School of the same university in 1980. In 1981, he was appointed a Lecturer within the University Department of Clinical Chemistry at Edinburgh. Here, he undertook research into the enzymology and protein chemistry of the glutathione S-transferase (GST) enzyme superfamily in rodents and human, becoming particularly interested in the contribution of inducible class Alpha GST to chemoprevention against the liver carcinogen aflatoxin B1. In October 1992, Dr Hayes moved as a Reader to the University of Dundee where he researched the molecular biology underpinning induction of detoxification enzymes that resulted in his laboratory identifying a previously unrecognized family of aflatoxin B1-metabolising aldo-keto reductases (AKR). In January 1997, the University of Dundee promoted him to a personal chair. Since then, Prof Hayes has directed his research towards the mechanisms by which GST, AKR and glutathione- and thioredoxin-based antioxidant genes are regulated by the transcription factor NRF2 (i.e., NF-E2 p45-related factor 2); NRF2 is a master regulator of intracellular redox homeostasis. The Hayes lab is currently engaged in research into how activation of NRF2 provides protection against established and ongoing non-alcoholic steatohepatitis and liver fibrosis, as well as exploring why NRF2 is downregulated during degenerative disease. During his academic career, Prof Hayes has been regarded a leading researcher in the GST, AKR, NRF2 and oxidative stress fields. He has participated in the organization of many international scientific conferences and has given numerous invited lectures. Prof Hayes was elected a Fellow of the Royal Society of Edinburgh (equivalent to the National Academy of Scotland) in May 2008, and a Fellow of the Society of Biology in September 2008. Further information about publications and citations can be found at the sites listed below.

 UoD Discovery page
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Bathish B, Robertson H, Dillon JF, Dinkova-Kostova AT, Hayes JD. Nonalcoholic steatohepatitis and mechanisms by which it is ameliorated by activation of the CNC-bZIP transcription factor Nrf2. Free Radical Biology and Medicine. 2022 Jun 18.

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