The Role of NRF2 Transcription Factor in Metabolic Syndrome

Abstract

Metabolic syndrome includes several diseases that are associated with metabolic abnormalities such as obesity, dyslipidemia, hypertension, type 2 diabetes, obesity, cardiac diseases, and insulin resistance. In order to maintain cellular homeostasis, it is necessary to regulate the signaling pathways involved in controlling oxidative stress. Nuclear factor erythroid-2 factor 2 (NRF2) is a transcription factor largely expressed in several tissues and cells and participates in the oxidative stress regulation signaling pathways. NRF2 also mediates transcriptional regulation of a variety of target genes to signalize and regulate acute and chronic stress pathways in metabolic syndrome. Deregulation of NRF2 could contribute to a worst prognosis/profile of individuals with metabolic syndrome. Therefore, NRF2 and its activators might play a role in its treatment, highlighted as targets for modulation by pharmacological agents.

1. Introduction

Metabolic syndrome (MetS) is associated with an increased risk of developing cardiovascular diseases (CVDs) and type 2 diabetes (T2D) and is characterized by several metabolic changes, among which the following stand out: obesity, insulin resistance, hypertension, high levels of triglycerides and fasting glucose, and decreased levels of high-density cholesterol (HDL-c) [1, 2]. The MetS global prevalence can vary from 12.5 to 31.4% being higher in the Eastern Mediterranean Region and Americas [3]. MetS was also associated with a higher risk of incidence of cardiovascular mortality in prospective analyses, increasing for each additional characteristic of MetS observed in the patient [4]. Cardiovascular diseases account for most noncommunicable diseases (NCD) deaths, about 17.9 million people annually, followed by cancers (9.3 million), chronic respiratory diseases (4.1 million), and diabetes (2.0 million including kidney disease deaths caused by diabetes) [5].

Inflammation and oxidative stress are precursors to several complications associated with the findings of MetS, mainly due to increased adiposity, insulin resistance, hypertension, and hyperlipidemia. Oxidative stress is an imbalance between oxidant and antioxidant molecules, leading to changes in signaling pathways, control of redox processes, and cellular damage [6]. The imbalance between prooxidant and antioxidant factors in the body can lead to an increased production of reactive oxygen species (ROS) and nitrogen species (RNS) [7]. ROS play important signaling roles under physiological conditions, once cells deliberately generate ROS through the respiratory chain and other metabolic processes, activating molecular signaling and immune defense mechanisms [8]. Also, ROS regulate several cellular functions such as cell growth, contraction, dilation, migration of vascular cells, and muscle tone [9]. Due to their importance in various physiological functions, certain ROS levels have beneficial effects such as in the process of glucose metabolism, adipogenesis, and adipocyte differentiation [10]. However, increased ROS leads to mitochondrial dysfunction, lipid peroxidation, protein, and DNA damage, compromising antioxidation pathways in metabolic syndrome [11]. Nuclear factor erythroid 2-related factor 2 (NRF2) is known as a master regulator of transcriptional activation of components of antioxidant systems. NRF2 transduces chemical signals to regulate a battery of cytoprotective genes [12]. Target genes include superoxide dismutase (SOD), catalase, glutathione S-transferase (GST), nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase, quinone oxidoreductase 1 (NQO1), γ-glutamate cysteine ligase (GCL), and heme oxygenase-1 (HO-1) [10].

Changes in metabolism and gene expression of redox homeostasis activate NRF2 and other stress response pathways. Furthermore, NRF2 has been identified as a regulator of drug or xenobiotic detoxification, target genes involved in proteasomal and autophagic function, iron, lipid, and carbohydrate metabolism, and DNA repair [13]. In homeostatic conditions, NRF2 binds to its main inhibitor, Kelch-like ECH-associated protein 1 (Keap1), with two specific coupling sites form, a cyclic E3-ubiquitin ligase and Cullin-3/Rbx1 complex. So, NRF2 goes into the ubiquitin proteasome to be degraded. In addition to being an NRF2 inhibitor, Keap 1 acts as a cysteine ​​thiol-rich redox sensor [12]. In the process of oxidative stress, oxidative factors can act and alter the chemical conformation of Keap1 cysteine, thus inhibiting ubiquitination and leading to excessive activation of NRF2 [14]. NRF2 can be phosphorylated by several enzymes, such as protein kinase C (PKC), which phosphorylates it at the Ser40 position, disrupting its interaction with Keap1. ROS can also increase NRF2 translocation to the nucleus, through promotion of the oxidation of cysteine residues in Keap1, allowing the release of NRF2, which will initiate the antioxidant transcription process [15].

Several studies correlate NRF2 with various regulatory functions of genes involved in cell proliferation and differentiation, inflammatory response, and lipid metabolism [12]. It has already confirmed the association between the etiology of MetS, oxidative stress, and chronic inflammation, which are under important control of NRF2 [11]. These data show the importance of investigating the role of NRF2 in this context, which involves changes in metabolic pathways in different organs.

2. NRF2 and metabolic syndrome

The involvement of NRF2 in the development of MetS is based on the interaction of NRF2 with other transcription factors or with the regulation of the expression of molecules that play a role in metabolism. Therefore, the impacts of Nrf2 on MetS require a systemic influence, rather than being controlled by a specific organ. Alterations in NRF2 have been shown to impair adipogenesis through peroxisome proliferator-activated receptor (PPAR), a master regulator of adipocyte differentiation and adipogenesis involved in MetS-related diseases, such as obesity and dyslipidemia [16]. It is known that oxidative stress plays a significant role in inducing insulin resistance, especially in association with the intake of high-fat diets (HFD) and obesity. On the other hand, in pancreatic β cells, excessive oxidative stress suppresses the transcription of insulin-related genes. Once NRF2 regulates redox homeostasis, many studies have been focused on deletion of the NRF2 gene, as well as its overexpression and its pharmacological activators [10].

Liu et al. [17] investigated about the role of NRF2 in nonalcoholic fatty liver disease, a hepatic manifestation of MetS, and suggested that deletion of NRF2 in mice may lead to hepatic insulin resistance due to activation of nuclear factor kappa B (NF-κB), a molecule involved in the phosphorylation of insulin receptor substrates. Consequently, NRF2 activation mitigated the progression of the disease and proved to be a therapeutic strategy. A significant decrease in mitochondrial fatty acid oxidation was observed in the absence of NRF2 and accelerated when NRF2 was constitutively active. These findings have implications for chronic disease conditions, including MetS [18].

2.1 Obesity and dyslipidemia

Commonly recognized for its energy storage function, adipose tissue is also responsible for endocrine functions. Adipocytes can release active biological compounds (adipokines), such as leptin, which act in the control of body weight and play inflammatory functions. This tissue also has different types of cells, such as immune and endothelial cells, which produce different types of hormones, with different functions such as glucose control and lipid metabolism, thus acting as a regulator of metabolic homeostasis [19].

Adipose tissue serves as a pivotal factor in the generation of systemic oxidative stress. The oxidation of fatty acids, within mitochondria and peroxisomes, stands as a significant contributor to oxidative stress, catalyzing the production of ROS. The overconsumption of oxygen also triggers the generation of free radicals within the mitochondrial respiratory chain, intrinsically linked with oxidative phosphorylation [20]. So, it is clear that deregulation in the adipose metabolism can lead to ROS impairment and break down the homeostasis.

Obesity is correlated with low-grade chronic and systemic inflammatory processes. Different studies associate these inflammation mechanisms with severe damage to various organs and tissues and are directly linked to several metabolic diseases [21]. Obese individuals or those with marked weight gain present a phenotypic change in white adipose tissue (WAT) with the appearance of inflamed and dysfunctional adipocytes that secrete pro-inflammatory cytokines [21, 22, 23]. These individuals may also have dyslipidemia, which is characterized by a lipid disorder changes in plasma lipoproteins. These alterations in the lipid profile include reduced levels of high-density lipoprotein (HDL) cholesterol, elevated small dense low-density lipoprotein (LDL), and hypertriglyceridemia [24].

Nonalcoholic fatty liver disease (NAFLD), associated with obesity and T2D, is characterized by liver damage caused by a dysfunctional diet, associated with the excess triglycerides in hepatocytes. So, in obese individuals, factors, such as NAFLD, induce oxidative stress in hepatocytes, thus increasing cellular damage. In addition, hepatic oxidative stress was attenuated with an antioxidant diet in rats with NAFLD, through modulation of the KEAP1/NRF2 pathway [14, 25]. Ma et al. [26] noted that wine devoid of sulfur dioxide effectively alleviated dyslipidemia induced by a HFD, influencing transcription factors, such as NRF2. Then, NRF2 activation enhances lipid metabolism and fatty acid β-oxidation by upregulating crucial genes, such as carnitine palmitoyltransferase I and PARP alpha. Ginsenoside Rg1 (antioxidant molecule extracted from ginseng) significantly improved chronic hepatic inflammatory injuries and fibrosis induced by lipopolysaccharides. This process involves the dissociation of NRF2 from Keap1, subsequently activating NRF2 pathway, which further inhibits NLRP3, NLRP1, and AIM2 inflammasomes. It is important to highlight that the activation of inflammasomes can be mediated by an increase in ROS, contributing to the inflammatory process also present in CVDs [27].

2.2 Hypertension

Reactive oxygen species and changes in radical production can break down redox-sensitive signaling and play an important role in the development of cardiovascular diseases and hypertension, through alterations in the renin-angiotensin system (RAS) and the increased RAS activation and Ang II production lead to an imbalance of redox pathways and to the activation of ROS-producing sources [28].

The harmful effects of the oxidative stress manifest themselves in tissues, inducing endothelial injuries and promoting the enlargement and thickening of the heart walls, especially in the left ventricle. This phenomenon contributes to the development of hypertension and other cardiovascular diseases (CVD) [29]. Increased peripheral resistance, a hallmark of arterial hypertension, arises mainly from structural and functional changes in small resistance arteries, manifested through vascular remodeling and endothelial dysfunction. These changes can result in narrowing the lumen due to a reduced outer diameter (eutrophic remodeling) or an invasion of the lumen by a thicker layer (hypertrophic remodeling) [30].

The interconnection between endothelial function and vascular remodeling is primarily attributed to the decreased availability of nitric oxide (NO), resulting in the promotion of predominantly hypertrophic vascular remodeling [31]. NO is an important vasodilator produced in the endothelium by the catalytic conversion of L-arginine into L-citrulline, through the enzyme endothelial nitric oxide synthase (eNOS). A series of cofactors are involved in this reaction, and their disruption leads to a monomeric form of the enzyme, which is uncoupled, and, instead of NO, superoxide is formed. In both hypertension and atherosclerosis, this uncoupled form of eNOS has been found. Increasingly, evidence indicates that NRF2 activation can trigger vasoprotective effects, reducing the bioavailability of ROS and increasing NO levels by eNOS [15]. In a study using rat pheochromocytoma cells, it was observed that NO can activate NRF2 by S-nitrosylation of Keap1 and, alternatively, by PKC-catalyzed phosphorylation of NRF2 [32]. Therefore, downregulation of NRF2 in hypertension may contribute to decreased antioxidant capacity, resulting in increased oxidative stress and consequent vascular dysfunction.

2.3 Diabetes

Oxidative stress is also causal in the development of β cell dysfunction, diabetes and insulin resistance [33, 34]. Diabetic patients have been shown increased cellular levels of ROS and ROS-induced DNA damage through a variety of mechanisms. An increased glycemic load in diabetes overwhelms the Krebs cycle, resulting in the inhibition of electron transfer within the mitochondrial membrane. Then, the accumulation of free radicals and ROS could lead to diabetic alterations such as the depletion of natural antioxidant molecules and damage to vascular cells, as well as alterations in gene and protein expression, blood flow, and endothelial cell permeability [33, 35, 36].

The excess of glucose is related to ROS dependent and alterations in NRF2 concentration are involved in the diabetes physiopathology and its complications. In both mouse placental tissue and human placenta cells, both increase and decrease of NRF2 were linked to gestational diabetes and implications for the health of the mother and offspring [37]. Proteins involved in oxidative stress, NRF2-linked phase II enzyme detoxification, and glutathione synthesis showed alterations in gestational diabetes, leading to increased oxidative stress, mitochondrial superoxide generation, protein carbonylation, and DNA damage [38]. In addition, alterations to NRF2 function have been implicated in the onset and progression of type 1 diabetes, through inhibition of the Keap1/NRF2 signaling pathway [39].

Regarding T2DM, the NRF2 has been shown to play a role in mediating diabetic organ-related complications. An important organ-related complication is the pancreatic β cell dysfunction and death induced by ROS. In mice, the genetic NRF2 induction reduced β cell damage, whereas the genetic NRF2 depletion markedly enhanced β cell damage [40, 41]. In addition to the pancreas, NRF2 has a role in diabetes in adipose tissue, skeletal muscle, heart, liver, and kidneys, with the complication of diabetic nephropathy [42].

The treatment of patients with T2DM is challenging since there is no cure available, but glycemia can be controlled by pharmacological therapy [43]. The NRF2 activation results in the activation of other antioxidant enzymes and cytoprotective genes, making it an attractive therapeutic target for treatment of diabetic complications [44]. Antioxidant therapies are comparatively more effective for treating diabetes from which NRF2 and its related factors are one of the potential activators. Moreover, antioxidant-based treatments serve as potential therapeutics for protection against the enhanced oxidative stress, as well as inflammation in diabetes mellitus [45, 46, 47, 48, 49]. The diabetic cardiomyopathy induced by T2DM has two main pathogenesis: the oxidative stress status in cardiomyocytes and lipid toxicity [50]. In the late diabetic cardiomyopathy stage, the Nrf2 is significantly downregulated, and then therapies focused on restoring the expression and activity of NRF2 could prevent diabetes-associated injuries in cardiomyocytes [50, 51, 52].

2.4 Coronary artery disease

Coronary artery disease (CAD) presents several common complications such as arrhythmias, acute coronary syndrome, congestive heart failure, mitral regurgitation, pericarditis, aneurysm formation, and mural thrombi. Recent research has challenged the protective effects of HDL due to difficulties in raising serum levels and is now focusing on triglyceride-rich lipoproteins (TGRL), in addition to LDL, as causes of atherosclerosis. Traditional risk factors for coronary artery disease have been well combated over the years, such as smoking, LDL levels, and high blood pressure, for example, due to population guidelines and low-cost therapies. However, TGRL are examples of emerging targets as they have greater correlations with the inflammatory state than LDL itself. TGRL cause inflammation, considering, for example, the content of one of its components, apolipoprotein C-III [53, 54].

The pathophysiology of CAD involves both the inflammatory response of the vascular endothelium and oxidative stress. Activation of the NRF2 pathway is associated with a reduction in myocardial infarct size and a delay in the progression to heart failure. The use of NRF2 activation also demonstrated protective effects against ischemia/reperfusion injury. Oxidative stress and inflammation are closely interconnected processes with oxidative stress-induced inflammation predominantly driven by NF-κB activation. It plays a crucial role in regulating the inflammatory response by controlling the gene expression of inflammatory cytokines, which are the main contributors to cardiovascular diseases in humans [54, 55].

Several mechanisms lead to the complex connection between the NRF2 and NF-kB pathways, which occur by inhibiting each other. IκB-α is an inhibitor of the NF-κB transcription factor as it prevents, for example, the binding of NF-kB to DNA, making its action impossible. NRF2 prevents proteasomal degradation of IκB-α and inhibits nuclear translocation of NF-κB. This occurs due to the increase in HO-1 levels stimulated by NRF2 and subsequent increase in the expression of phase II enzymes that block IκB degradation. NRF2 acts to decrease the expression of pro-inflammatory cytokines such as vascular cell adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), and some interleukins, all involved in atherosclerosis. Understanding this intricate interaction is essential to unravel the inflammatory mechanisms involved in the pathogenesis of CAD and develop more specific therapeutic interventions [56, 57, 58].

3. NRF2 and therapy

Most pharmacological activators of NRF2 are electrophilic substances capable of modifying cysteine residues present in Keap1, ultimately resulting in changes in its conformation through oxidation or alkylation processes [15]. Some of these new therapies and medications have been considered to control MetS through regulation of Keap1 protein and NRF2 (Figure 1).

Figure 1.

Effect of oxidative stress and antioxidant drugs used in the treatment of metabolic syndrome.

Cannabidiol (CBD) is a nontoxic compound derived from cannabis, known for its anti-inflammatory, anti-apoptotic, and antioxidant properties. Interest in the use of CBD has grown, including studies focusing on cardiovascular diseases. In mice, it was observed increased translocation of NRF2 from the cytoplasm to the nucleus via Keap1 inhibition, recognized as a beneficial effect of CBD, preventing acute myocardial infarction after exhaustive exercise [59].

In an environment of oxidative stress NRF2 is released from Keap1, allowing the nuclear translocation of NRF2 and the increase of expression of antioxidant proteins. The action of drugs on MetS components (obesity, diabetes, hypertension, and coronary artery disease) may lead to the decrease of Keap1, thus increasing the activation of NRF2 and HO-1, NRF2 nuclear translocation, and activation of antioxidant protein expression. The drugs also act to reduce inflammatory cytokines and NF-Kb, which in normal oxidative conditions act to inhibit NRF2, impairing translocation.

Resveratrol, administered to hypertensive rats, improved NRF2 activity by combating oxidative stress in proximal tubular epithelial cells, improving inflammation, and attenuating the development of hypertension [60]. Lopes et al. [30] observed a vasoprotective function of NRF2 in hypertension when evaluating hypertensive rats under conditions of vascular oxidative stress related to vascular dysfunction. Bardoxolone and L-sulforaphane activate NRF2 when administered to animals, reducing the formation of ROS that are induced by angiotensin II, and reestablishing the endothelial function in the cells. The impact of oxidative stress, induced by ANG II infusion, was also investigated by Wang et al. [61], in mice, analyzing the effects of treatment with tert-butylhydroquinone (tBHQ), that triggered the activation of NRF2, preventing increased oxidative stress, microvascular ROS, endothelial dysfunction, enhanced microvascular remodeling, contractility, and hypertension. In a study by Gao et al. [62], Qing brick tea (QBT), a traditional and popular Chinese drink, was administered to mice with obesity induced by monosodium glutamate, proving effective in protecting against metabolic syndrome. QBT treatment attenuated oxidative stress and IR and markedly stimulated the NRF2/HO-1 and insulin signaling pathways. A study based on the transcriptome network analysis after using methylglyoxal, conducted on the visceral adipose tissue of hereditary hypertriglyceridemic rats, a nonobese model of metabolic syndrome, revealed that the downregulation of NRF2 expression resulted in the over-representation of genes associated with insulin signaling, lipid metabolism, and angiogenesis [63].

Some drugs already used to control glycemia act regulating the activity of NRF2. Among these, Sitagliptin is a drug indicated for use combined with metformin or sulfonylurea, for treating T2DM. Recent studies indicate that Sitagliptin can also have a regulatory effect on autophagy and protect the cardiovascular system. In the cell, it acts to reduce NF-kB, which, in turn, regulates inflammatory cytokines and helps to inhibit the overexpression of NRF2 [64, 65]. Metformin is the first-line pharmacotherapy for T2DM. Studies carried out with mice to understand the extent of the drug’s use have shown that its long-term use not only helps in the treatment of T2DM but also acts as a cardioprotector against heart diseases associated with MetS. These studies indicate that Metformin increases the expression of Nrf2, inducing an improvement in the nuclear translocation of Nrf2 [65, 66]. It reduces excessive ROS generation, limiting cell apoptosis, and increasing signaling through the ROS-mediated PI3K/Akt and NRF2/HO-1 pathways and the levels of antioxidants [66].

Diabetic nephropathy is an important diabetes complication characterized by inflammatory response and renal dysfunction [48]. Sulforaphane (SFN) is an organosulfur compound that exhibits anticancer and antidiabetic properties and has an indirect antioxidant effect inducing expression of several enzymes via the Keap1/NRF2 pathway [67, 68]. Then, SFN has particular interest in the role via induction of NRF2 of preventing diabetic nephropathy and renal damage [69]. The upregulation of NRF2 of diabetic nephropathy mice model, by treatment with fenofibrate, led to regulation of other antioxidant molecules and delayed the progression of diabetic nephropathy [46]. Baicalin (BAI), a flavonoid component extracted from the traditional Chinese medicine S. baicalensis, showed to inhibit oxidative stress in hyperglycemic environments through regulation of NRF2 pathways. It also decreases blood lipids and inflammation markers and suppresses the excessive production of ROS in cardiovascular diseases [50, 70, 71].

4. Conclusion

Considering the role of NRF2 in activating genes involved in combating oxidative stress and inflammation, it is clear that its inhibition, which often occurs due to increased Keap1 activity, can significantly increase the chances of developing metabolic syndrome. This association is justified by the participation of these genes, activated by this important transcription factor, in several metabolic pathways related especially to endothelial function, insulin signaling, and lipid catabolism. So, it impacts the set of clinical findings that constitute metabolic syndrome. Therefore, several drugs that act on NRF2 activation are promising for improving or preventing chronic metabolic diseases that are not only common but also increasing, assumed as one of the diseases of the century.

Conflict of interest

The authors declare no conflict of interest.