Oxidative stress and redox signaling mechanisms of alcoholic liver disease: Updated experimental and clinical evidence


  • Hong ZHU,

    1. Department of Pharmacology, Edward Via College of Osteopathic Medicine, Virginia Tech Corporate Research Center
    2. Department of Biomedical Sciences and Pathobiology, Virginia Polytechnic Institute and State University
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  • Zhenquan JIA,

    1. Department of Biology, University of North Carolina at Greensboro, Greensboro, North Carolina, USA
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  • Hara MISRA,

    1. Department of Pharmacology, Edward Via College of Osteopathic Medicine, Virginia Tech Corporate Research Center
    2. Department of Biomedical Sciences and Pathobiology, Virginia Polytechnic Institute and State University
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  • Y Robert LI

    Corresponding author
    1. Department of Pharmacology, Edward Via College of Osteopathic Medicine, Virginia Tech Corporate Research Center
    2. Department of Biomedical Sciences and Pathobiology, Virginia Polytechnic Institute and State University
    3. Virginia Tech-Wake Forest University School of Biomedical Engineering and Sciences, Blacksburg, Virginia
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Y Robert LI, Department of Pharmacology, Edward Via College of Osteopathic Medicine, Virginia Tech Corporate Research Center, 1861 Pratt Drive, Blacksburg, VA 24060, USA. Email: yli@vcom.vt.edu


Alcoholic liver disease (ALD) is a major cause of morbidity and mortality in the United States and Europe. The spectrum of ALD ranges from fatty liver to alcoholic hepatitis and cirrhosis, which may eventually lead to hepatocellular carcinoma. In developed countries as well as developing nations, ALD is a major cause of end-stage liver disease that requires liver transplantation. The most effective therapy for ALD is alcohol abstinence; however, for individuals with severe ALD and those in whom alcohol abstinence is not achievable, targeted therapies are absolutely necessary. In this context, advances of our understanding of the pathophysiology of ALD over the past two decades have contributed to the development of therapeutic modalities (e.g., pentoxifylline and corticosteroids) for the disease although the efficacy of the available treatments remains limited. This article is intended to succinctly review the recent experimental and clinical findings of the involvement of oxidative stress and redox signaling in the pathophysiology of ALD and the development of mechanistically based antioxidant modalities targeting oxidative stress and redox signaling mechanisms. The biochemical and cellular sources of reactive oxygen and nitrogen species (ROS/RNS) and dysregulated redox signaling pathways associated with alcohol consumption are particularly discussed to provide insight into the molecular basis of hepatic cell dysfunction and destruction as well as tissue remodeling underlying ALD.


In chemistry, an alcohol is any organic compound in which a hydroxyl functional group (-OH) is bound to a carbon atom, usually connected to other carbon or hydrogen atoms. Although alcohol is a general term for any alcoholic compound, in this article the term refers specifically to ethyl alcohol or ethanol (CH3CH2OH). Alcohol has been a part of human culture since the beginning of recorded history. Excessive use of alcohol contributes substantially to the global burden of disease (4% of total mortality) and is thus one of the largest avoidable risk factors.1 In the United States, alcoholism is also a major public health problem, and it is estimated that over 100 million Americans are alcoholic. Alcohol abuse accounts for 100 000 to 200 000 deaths annually in the USA, of which over 20 000 are attributable directly to end-stage hepatic cirrhosis and many more are the result of automobile accidents.2 Because of this, it is recommended that alcohol consumption be moderate: ≤2 drinks daily in men and ≤1 drink in women. In the USA a ‘standard’ drink is any drink that contains about 0.6 fluid ounces or 14 g of ‘pure’ ethanol. It is of note that ethanol contains 7 calories per gram.

Although excessive alcohol consumption is associated with a variety of disorders, alcoholic liver disease (ALD) has the greatest health impact. As depicted in Figure 1, ALD encompasses a spectrum of hepatic disorders including (i) fatty liver (also known as steatosis, due to abnormal accumulation of lipids in hepatic tissue); (ii) alcoholic hepatitis (due to inflammation and necrosis) and (iii) cirrhosis (due to excessive fibrosis).3 These are not necessarily distinct stages of evolution of the disease, but rather multiple stages that may be present simultaneously in a given individual. Over 90% of heavy drinkers develop fatty liver and, of those, 10–35% develop alcoholic hepatitis. About 8–20% of chronic alcoholics develop liver cirrhosis, which may eventually lead to hepatocellular carcinoma.4

Figure 1.

Spectrum of alcoholic liver disease (ALD). It is of note that although cirrhosis is usually the primary pathophysiological process that leads to the development of hepatocellular carcinoma, ALD may progress into liver cancer without necessarily going through cirrhosis.


Multiple pathways are involved in the genesis and progression of alcoholic fatty liver, alcoholic hepatitis and cirrhosis.

Alcoholic fatty liver results from the excessive accumulation of triglyceride (TG) in hepatocytes. Alcohol is metabolized in hepatocytes through oxidation to acetaldehyde and subsequently from acetaldehyde to acetate catalyzed by various enzymes or enzymatic systems, including the alcohol dehydrogenase pathway, cytochrome P450 2E1 (CYP2E1) system and catalase (Fig. 2).5 The oxidative metabolism of alcohol generates an excess of the reduced form of nicotinamide adenine dinucleotide (NADH), resulting in an increased ratio of NADH to NAD+ in hepatocytes. This altered NADH/NAD+ ratio in hepatocytes leads to the inhibition of fatty acid oxidation and the promotion of lipogenesis. In addition, alcohol promotes lipid metabolism through the inhibition of peroxisome proliferator-activated receptor (PPAR)-α and adenosine monophosphate kinase, and via the stimulation of sterol regulatory element-binding protein 1 (SREBP1), a membrane-bound transcription factor.6,7 In combination, these effects result in a fat-storing metabolic remodeling of the liver, which is manifested as an excessive accumulation of lipids in hepatocytes. Although fatty liver resolves with abstinence, it predisposes the individuals who continue to drink excessively to more serious forms of ALD, that is, alcoholic hepatitis and cirrhosis.

Figure 2.

Sequential metabolism of ethanol to acetaldehyde and acetate. As illustrated, the metabolism of ethanol occurs in the liver (also in many other types of tissues) via three enzymatic systems, alcohol dehydrogenase (ADH), cytochrome P450 2E1 (CYP2E1), and catalase, which are primarily located in cytosol, endoplasmic reticulum membrane (microsomes), and peroxisomes, respectively. Aldehyde dehydrogenase (ALDH) is located in mitochondria and cytosol. NAD+, oxidized form of nicotinamide adenine dinucleotide; NADH, nicotinamide adenine dinucleotide; NADPH, nicotinamide adenine dinucleotide phosphate (reduced).

Alcoholic hepatitis is characterized by inflammation and necrosis.8 It is believed that alcohol compromises the intestinal barrier, which leads to an increase in its permeability and the subsequent presence of gut bacteria-derived lipopolysaccharide (LPS) in the portal blood. LPS stimulates the Kupffer cells and possibly other types of cells in the liver, resulting in an excessive release of inflammatory cytokines and reactive oxygen and nitrogen species (ROS/RNS). The dysregulated inflammatory responses are thought to be contributed primarily to the hepatocyte injury and tissue necrosis. In addition, the accumulation of lipids also promotes inflammation. Progression of these detrimental processes may eventually cause an excessive accumulation of collagen in the extracellular matrix (ECM), resulting in liver fibrosis and cirrhosis. The crucial role of inflammation in ALD is supported by the demonstrated therapeutic efficacy of anti-inflammatory drugs (e.g., glucocorticosteroids) in animal models and certain patient subgroups.8,9 In addition to dysregulated inflammation, recent work has demonstrated that oxidative stress also plays an important role in the development of alcoholic hepatitis and cirrhosis as well as in fatty liver. Since the general aspects of oxidative stress mechanism of ALD have been reviewed previously,6,10 we aimed to focus on the discussion of the new findings from both animal experiments and human studies with an emphasis on novel oxidative stress and redox signaling pathways and the mechanistically based modalities targeting these molecular pathways for the intervention of ALD.


Studies using animal models have contributed greatly to our understanding of how ALD develops and how the severity of liver injury is influenced by factors other than alcohol, such as nutrition, oxygen deprivation (as occurs with sleep apnea and smoking), and gene regulation. In this regard, the intragastric feeding model resulting in liver lesions that mimic human ALD has been widely used.11,12 In addition, the chronic feeding of a liquid diet containing ethanol is also a commonly used model of ALD.13 Animal experiments have provided important information on the pathophysiology of ALD, including the causative involvement of oxidative stress, and have also contributed to the development of new therapeutic approaches.

Evidence for a causative role of oxidative stress in experimental ALD

There are four lines of experimental evidence which support the causative involvement of oxidative stress in the development of ALD in animal models.

Increased formation of ROS/RNS

The ability of acute and chronic alcohol treatment to increase the production of ROS/RNS as well as other free radical species (e.g., 1-hydroxyethyl radical) has been demonstrated in a variety of systems, including cell cultures and experimental animals, as well as in humans.6,10 Detailed cellular sources and mechanisms of ROS/RNS formation will be described later in this article.

Depletion of antioxidants and increased biomarkers of oxidative damage

Although acute exposure to alcohol may cause the transient induction of certain antioxidant genes, excessive chronic exposure usually results in decreased levels of antioxidant defenses both in hepatic tissue and in blood. Of note, an early change after alcohol treatment is the depletion of mitochondrial glutathione in hepatocytes, which appears to play an important part in alcohol-induced liver injury.14 In line with the augmented formation of ROS/RNS and decreased levels of antioxidant defenses, increased levels of oxidative stress biomarkers are observed in experimental animals exposed to alcohol. These biomarkers include lipid peroxidation products (e.g., malondialdehyde, 4-hydroxy-2-nonenal, F2-isoprostanes), oxidative DNA-base modifications (e.g., 8-hydroxy-2’-deoxyguanosine) and the protein adducts derived from reactive aldehydes, as well as 1-hydroxyethyl radical. Notably, the levels of the oxidative stress biomarkers are correlated with the severity of alcohol-induced liver injury. Thus, oxidative stress biomarkers are frequently used to assess liver injury as well as protection against it by potential therapeutic agents, including antioxidant compounds.6,10

Protection by exogenous antioxidant compounds

The ability of alcohol to cause oxidative stress makes it rational to use antioxidant compounds to protect against liver injury. Indeed, various compounds with antioxidant properties have been shown to be protective against alcohol-induced liver injury in animal models. These include antioxidant vitamins, glutathione-augmenting agents (e.g., glutathione esters, N-acetylcysteine, S-adenosyl-L-methionine), antioxidant mimetics, probucol and dietary polyphenols.15–19 In addition, the chemical induction of endogenous antioxidants also ameliorated alcohol-induced liver injury.20,21 These findings are consistent with the concept that oxidative stress plays a causal role in experimental ALD.

Protection by genetic overexpression of endogenous antioxidant genes and sensitization by antioxidant gene knockout

In line with protection by exogenous antioxidant compounds, the overexpression of endogenous antioxidant proteins (e.g., manganese superoxide dismutase [MnSOD], copper-zinc [Cu-Zn] superoxide dismutase [SOD] and metallothionein) via transgenic approaches or gene delivery blunted alcohol-induced liver injury in experimental animals.22–24 Of note, protective effects were also observed with the systemic administration of antioxidant proteins, such as thioredoxin.25 Conversely, gene knockout of endogenous antioxidant proteins (e.g., Cu-Zn SOD, catalase, glutathione peroxidase-1 and metallothionein) sensitized the experimental animals to alcohol-induced liver injury.26–29 More recently, it has been reported that the ablation of either sulfiredoxin or peroxiredoxin-1 in mice markedly aggravated alcohol-induced oxidative hepatic injury.30 Notably, peroxiredoxin-1 was found to be closely associated with CYP2E1 on the cytosolic side of the endoplasmic reticulum membrane of the hepatocytes. The close proximity of peroxiredoxin-1 and CYP2E1 was suggested to be responsible for the peroxiredoxin-1-mediated removal of ROS generated by CYP2E1.30

The above observations using antioxidant gene overexpression and knockout models provide the most compelling evidence for a causal relationship between oxidative stress and the development of ALD. The crucial beneficial function of endogenous antioxidant defenses in ALD is further strengthened by the finding that the targeted disruption of nuclear factor-erythroid 2 p45-related factor 2 (Nrf2), a central regulator of antioxidant genes, markedly aggravated alcohol-induced liver injury in mice, as indicated by increased liver necrosis, inflammation, oxidative stress and mortality.31 Consequently, activation of Nrf2 signaling by pharmacological agents or phytochemicals has been proposed as an important strategy for protecting against liver injury induced by alcohol as well as other hepatotoxicants32 (see below for further discussion of Nrf2 signaling in the molecular pathophysiology of ALD).

Sources of ROS/RNS


As oxidative damage takes place predominantly in hepatocytes following alcohol administration, hepatocytes may thus be a major source of ROS/RNS as well as of other free radical species. Indeed, several intracellular pathways have been shown to contribute to the increased production of reactive species in hepatocytes. These include mitochondria, CYP2E1, NAD(P)H oxidase (NOX), and inducible nitric oxide synthase (iNOS).

  • • Mitochondria. As discussed earlier, the metabolism of alcohol results in increased levels of NADH, which provide electrons for the mitochondrial electron transport chain, leading to an increased one-electron reduction of oxygen to superoxide. In addition, the formation of acetaldehyde was shown to cause mitochondrial damage, which may also lead to the increased formation of superoxide from the electron transport chain.33,34

Recently, the redox-active protein p66Shc was found to be associated with mitochondria and to generate ROS via electron transfer from cytochrome c.35 This redox-active protein potentiated ethanol-induced oxidative stress and mitochondrial damage in hepatocytes.36 Ethanol treatment of mice also caused induction of p66Shc in hepatic tissue, while mice deficient in p66Shc showed increased resistance to ethanol-induced liver oxidative stress, steatosis and necrosis as compared with the wild-type counterparts.36 Interestingly, the deletion of p66Shc led to a marked augmentation of ethanol-mediated induction of MnSOD in hepatic tissue and cultured hepatocytes, suggesting that p66Shc may also act as a transcriptional suppressor of liver MnSOD.36 Hence, the mitochondria-associated, ethanol-inducible, redox-active protein p66Shc may function as a novel source of ROS generation during ethanol consumption, contributing to the oxidative liver injury underlying ALD.

  • • CYP2E1. CYP enzymes, especially CYP2E1 are known to produce ROS. The ability of CYP2E1 to produce ROS is augmented during alcohol administration as this enzyme is highly inducible by alcohol. Both cell experiments and animal studies in vivo have demonstrated that CYP2E1 as an important source of ROS that participates in alcohol-induced liver injury. The knockout of CYP2E1 blunted alcohol-induced oxidative stress and liver injury in mice, whereas transgenic overexpression of the enzyme aggravated alcohol-induced hepatic damage, pointing to a causative involvement of CYP2E1 in the pathophysiology of ALD.37
  • • NOX and iNOS. Although NOX and iNOS of inflammatory cells are primarily responsible for the augmented formation of ROS/RNS during inflammatory responses, alcohol treatment also resulted in the activation of both NOX and iNOS in hepatocytes, leading to the increased production of both superoxide and nitric oxide.38,39 The reaction of these two radical species generates the potent oxidant, peroxynitrite, which has been suggested to cause mitochondrial damage underlying experimental ALD.40,41

Inflammatory cells

As stated above, the activation of NOX and iNOS in inflammatory cells results in large amounts of ROS/RNS formation. Both hepatic Kupffer cells and infiltrated inflammatory cells have been found to be activated to produce ROS/RNS following alcohol administration.42 The activation of Kupffer cells by LPS derived from gut bacteria is an early event leading to ALD. In this regard, the depletion or inactivation of Kupffer cells ameliorated alcohol-induced inflammation, oxidative stress and liver injury in experimental animals.42 Alcohol-induced oxidative stress and hepatic injury were also blunted in mice deficient in either p47phox, an essential component of the NOX enzyme complex, or iNOS, indicating a causative role for NOX and iNOS in the pathophysiology of ALD.43–45

Hepatic stellate cells (HSCs)

In addition to hepatocytes and inflammatory cells, HSCs may also contribute to the formation of ROS during alcohol exposure. Treatment with alcohol or its metabolite acetaldehyde was shown to elicit increased ROS formation from NOX in HSCs. ROS thus formed as well as the ROS derived from other cellular sources, such as CYP2E1, may play an important part in the transformation of HSCs into myofibroblasts as well as the increased production of collagen, a process critically involved in liver fibrosis.6,46 Mechanistically, it was demonstrated that the induction of collagen expression by acetaldehyde in HSCs was dependent on PPAR-γ phosphorylation induced by a hydrogen peroxide-mediated activation of the profibrogenic c-Abl signaling pathway.47 This finding also highlighted the intimate association between acetaldehyde and ROS in the molecular pathophysiology of ALD.

Molecular mechanisms of oxidative stress-mediated injury

The key molecular mechanisms underlying oxidative stress-mediated liver injury in ALD can be summarized in the following three schemes: (i) mitochondrial damage; (ii) the perpetuation of inflammation via dysregulated redox signaling and (iii) the transformation of HSCs and ECM remodeling associated with dysregulated redox signaling.6,48 In addition, recent studies have suggested there is a crucial role for dysregulated Nrf2 signaling in molecular alterations leading to experimental ALD.

Mitochondrial damage

Alcohol administration causes the increased formation of ROS in mitochondria.33,34 Such locally generated ROS, as well as that derived from other cellular sources, cause mitochondrial dysfunction, energy depletion and, ultimately, cell death.49 The increased production of ROS/RNS also results in the damage of other cellular consumption, including membrane lipids, enzymes and nucleic acids, contributing to cell injury. Notably, alcohol consumption increases hepatic iron accumulation, which may further facilitate the production of the highly reactive hydroxyl radical, causing damage to mitochondria and other cellular constituents.50 The key role of mitochondrial oxidative stress injury in ALD is further supported by the finding that either the overexpression of the mitochondrial antioxidant enzyme MnSOD or the administration of exogenous antioxidant compounds targeted to mitochondria, such as mito-coenzyme Q, led to the attenuation of oxidative liver injury underlying ALD.23,51

Perpetuation of inflammation via dysregulated redox signaling

As noted earlier, the activation of inflammatory cells results in the formation of large quantities of ROS/RNS. ROS/RNS thus formed also cause dysregulated redox homeostasis, leading to the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and the subsequent overexpression of proinflammatory cytokines and adhesion molecules.52,53 NF-κB-regulated chemokine monocyte chemotactic protein (MCP)-1 was recently shown to be essential for continuous inflammatory responses in liver tissue and the development of hepatic steatosis in alcohol-treated mice.54 Hence, the perpetuation of inflammation and inflammatory injury may be an important molecular mechanism underlying the development and progression of ALD. Since the metabolism of alcohol also leads to the increased ROS formation independent of the activation of inflammatory cells (for example, the CYP2E1 and mitochondria-derived ROS), ROS may also contribute to the initiation of the inflammatory responses by activating NF-κB and other redox-sensitive signaling molecules. Oxidative stress may thus contribute to the genesis of ALD. This notion is supported by observations that the administration of antioxidant compounds or the overexpression of endogenous antioxidant proteins is frequently able to prevent the development of ALD in experimental animals.

Transformation of HSCs and ECM remodeling associated with dysregulated redox signaling

ROS/RNS are able to induce the transformation of HSCs into myofibroblasts, leading to the excessive production and deposition of collagen in the ECM. Reactive aldehydes derived from lipid peroxidation, such as 4-hydroxy-2-nonenal, have also been shown to stimulate collagen production from stellate cells.55 In addition, ROS/RNS are capable of inducing the proliferation and activation of other cells, such as periportal fibroblasts and fibrocytes. Furthermore, ROS/RNS may cause the activation of matrix metalloproteinases, resulting in the remodeling of the ECM.56 Progression of these molecular and cellular events eventually leads to excessive hepatic fibrosis and cirrhosis.57

Hepatic Nrf2 signaling

A study in the early 1990s has demonstrated a crucial role for a cis-acting element named antioxidant response element (ARE) in the regulation of antioxidant gene expression.58 In 1997, Itoh et al. identified Nrf2 as a critical transcription factor that interacts with the ARE, leading to the increased expression of various antioxidant and cytoprotective genes.59 Subsequent studies from multiple research groups have established that Nrf2 plays a central role in regulating both constitutive and inducible expression of a wide variety of antioxidant and anti-inflammatory genes in mammalian tissues/cells.60–62 As noted earlier, the targeted disruption of Nrf2 in mice led to the aggravation of ethanol-induced liver injury as evidenced by augmented hepatic oxidative stress, inflammatory responses, steatosis and necrosis. The ethanol-mediated hepatic mitochondrial damage was also exacerbated in Nrf2-knockout mice. These exacerbated deleterious responses led to accelerated liver failure and death caused by ethanol in the Nrf2-knockout mice.31 Notably, short-term ethanol treatment resulted in the induction of several antioxidant and anti-inflammatory enzymes, including heme oxygenase-1, NAD(P)H, quinine oxidoreductase-1 and glutathione S-transferase (GST) in liver tissue in an Nrf2-dependent manner.31 Hence, the induction of antioxidant and anti-inflammatory enzymes via the Nrf2 signaling might act as an early compensatory or adaptive mechanism to suppress ethanol-induced oxidative injury. The inflammation in liver tissue and the ablation of this protective compensatory mechanism in Nrf2-knockout mice led to a dramatic acceleration of ethanol-induced liver injury, hepatic failure and death in mice.31 In this context, a recent study reported that ethanol-induced upregulation of heme oxygenase-1 expression was mediated by both Nrf2 signaling and hypoxia-inducible factor (HIF)-1α and the upregulation of heme oxygenase-1 attenuated inflammatory cytokine expression in ALD.63 Nrf2 was also found to be induced by the overexpression of CYP2E1 in liver tissue and cultured hepatocytes, and the augmented Nrf2-dependent antioxidants attenuated CYP2E1-mediated oxidative hepatic injury.64

Taken together, the above observations demonstrate a crucial role for Nrf2 signaling (and possibly other signaling pathways) in suppressing the oxidative and inflammatory pathophysiology of ALD (Fig. 3). Hence, the disruption of the normal functionality of Nrf2 signaling may be a potential mechanism leading to increased susceptibility to ALD. Likewise, the augmentation of hepatic Nrf2 signaling by pharmacological approaches or gene therapy, as described earlier, may represent an effective strategy for the intervention of the oxidative stress pathophysiology of ALD.

Figure 3.

Involvement of nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) signaling in the pathophysiology of alcoholic liver disease (ALD). Activation of Nrf2 signaling and Nrf2-dependent antioxidants and anti-inflammatory enzymes may act as a compensatory protective mechanism in ALD. Consequently, compromised Nrf2 functionality may aggravate the pathophysiology of ALD. CYP2E1, cytochrome P450 2E1; LPS, lipopolysaccharide; ROS, reactive oxygen species.

Summary of the role of oxidative stress in experimental ALD

Alcohol causes the augmented formation of ROS/RNS from multiple sources, including hepatocytes, inflammatory cells and other cell types in liver tissue. The augmented production of ROS/RNS and the consequent depletion of antioxidants together lead to the marked oxidative stress that contributes to both the genesis and progression of ALD in animal models (Fig. 4). The causative involvement of oxidative stress in ALD is further supported by the demonstrated efficacy of both exogenous and endogenous antioxidants in protecting against alcohol-induced liver injury. These findings have prompted the investigation of the causative role of oxidative stress in human ALD and the antioxidant-based modalities for intervention in the disease.

Figure 4.

Schematic illustration of the overall involvement of oxidative/electrophilic stress and dysregulated inflammation in the pathophysiology of alcoholic fatty liver disease. CYP2E1, cytochrome P450 2E1; LPS, lipopolysaccharide; ROS, reactive oxygen species.


Evidence for the role of oxidative stress

Increased ROS/RNS formation and biomarkers of oxidative stress

The excessive consumption of alcohol led to increases in biomarkers of oxidative stress in plasma and urine, including the lipid peroxidation products F2-isoprostanes and 4-hydroxy-2-nonenal.65,66 Oxidative stress biomarkers were closely associated with the severity of liver injury. Notably, acute alcohol consumption in healthy individuals also resulted in increases in oxidative stress biomarkers, suggesting that oxidative stress may precede the development of ALD.67 In addition, levels of antioxidants, such as glutathione, vitamin C and E decreased in ALD patients, compared with normal individuals. As discussed above, the metabolism of alcohol forms 1-hydroxyethyl radical in experimental systems. The protein adduct of the 1-hydroxyethyl radical was also detected in patients with alcoholic cirrhosis.68

Antioxidant gene polymorphisms

Studies have demonstrated that polymorphisms of GST, especially the null-genotypes of GSTM1 and GSTT1, were associated with an increased risk of developing ALD as well as with hepatocellular carcinoma.69,70 These findings are consistent with the notion GST enzymes play an important role in the detoxification of reactive aldehydes, including those participate in alcohol-induced liver injury. In addition to GST, a Val-Ala polymorphism in MnSOD was associated with an elevated risk of developing cirrhosis in French alcoholics and increased rates of hepatocellular carcinoma development and death in cirrhotic patients.71 Notably, the polymorphisms in antioxidant genes may interact with those in other xenobiotic biotransforming enzymes, resulting in an augmented risk of developing ALD. For example, the combination of the GSTM1-null genotype and a polymorphism of CYP2E1 (causing increased CYP2E1 activity) resulted in an increased risk of developing alcoholic liver cirrhosis.72 The combination of GG-myeloperoxidase (MPO) genotype (leading to high MPO expression) and at least one Ala-MnSOD allele markedly elevated the risk of hepatocellular carcinoma occurrence and death in patients with alcoholic cirrhosis.73

Antioxidant intervention

Although the experimental evidence supporting the protective role of antioxidants in alcohol-induced liver injury is extensive, the effectiveness of antioxidant therapy remains unclear in ALD patients. Randomized controlled clinical trials on the use of antioxidant vitamin E or an antioxidant cocktail (containing antioxidant vitamins, N-acetylcysteine and coenzyme Q, etc.) have failed to show their efficacy in improving the disease conditions in ALD patients.74–76 The causes of this apparent discrepancy between experimental studies and clinical trials are not clear. Potential reasons may include the doses of antioxidants used in the trials, the duration and time-window of the intervention and the patients' oxidative stress and antioxidants status, as well as the lack of a profound understanding of the pharmacokinetic and pharmacodynamic activities of the antioxidant compounds in patients with ALD. In this context, S-adenosyl-L-methionine is a well-characterized antioxidant compound that functions as a methylating agent, a precursor of glutathione and a modulator of proinflammatory cytokine expression. A previous randomized controlled trial demonstrated that patients with ALD treated with S-adenosyl-L-methionine showed improved mortality and a decreased need for liver transplantation.77 Although S-adenosyl-L-methionine has promising effects, the exact contribution of the antioxidant activity of this compound to the therapeutic activity in ALD remains to be determined. Moreover, the clinical efficacy of S-adenosyl-L-methionine as a therapy for ALD needs to be further established in large-scale well-designed trials.

Summary and future perspectives on studies of ALD in humans

Compared with the substantial experimental evidence supporting a causative role of oxidative stress in ALD and the benefits of antioxidant-based modalities in disease intervention in animal models, evidence from studies in humans is less compelling. Nevertheless, there is accumulating evidence indicating the possible causative involvement of oxidative stress in the pathophysiology of human ALD. Future studies should focus on developing novel antioxidant compounds with well-characterized pharmacokinetic and pharmacodynamic profiles, and testing them in selected populations with overt oxidative stress and antioxidant deficiency. Such a subpopulation targeting strategy has been shown to be effective in the intervention of other oxidative stress-associated diseases, such as cardiovascular disorders and diabetes.78–80 In addition to therapeutic intervention, future efforts should also be devoted to the development of antioxidant-based strategies for primary prevention of ALD in susceptible or high-risk individuals.


The study was approved by Grant from National Institutes of Health (No. DK81905) to Hong ZHU, and Grant from Association for International Cancer Research and National Institutes of Health (No. HL93557) to Zhenquan JIA.