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Journal of Nutritional Biochemistry 29 (2016) 1 11

REVIEWS: CURRENT TOPICS

Nutritional therapy for nonalcoholic fatty liver disease

Paola Dongiovanni a , Claudia Lanti b , Patrizia Riso b,, Luca Valenti a, c

a
Internal Medicine and Metabolic Diseases, Fondazione IRCCS Ca Granda Ospedale Maggiore Policlinico, 20122 Milano, Italy
b
Department of Food, Environmental and Nutritional Sciences (DeFENS), Division of Human Nutrition, Universit degli Studi di Milano, 20133 Milano, Italy
c
Department of Pathophysiology and Transplantation (DEPT), Universit degli Studi di Milano, 20122 Milano, Italy

Received 21 July 2015; received in revised form 26 August 2015; accepted 26 August 2015

Abstract

Following the epidemics of obesity, nonalcoholic fatty liver disease (NAFLD) has become the leading cause of liver disease in western countries. NAFLD is the
hepatic manifestation of metabolic syndrome and may progress to cirrhosis and hepatocellular carcinoma. To date, there are no approved drugs for the treatment
of NAFLD, and the main clinical recommendation is lifestyle modification, including increase of physical activity and the adoption of a healthy eating behavior. In
this regard, studies aimed to elucidate the effect of dietary interventions and the mechanisms of action of specific food bioactives are urgently needed.
The present review tries to summarize the most recent data evidencing the effects of nutrients and dietary bioactive compounds intake (i.e., long-chain PUFA,
Vitamin E, Vitamin D, minerals and polyphenols) on the modulation of molecular mechanisms leading to fat accumulation, oxidative stress, inflammation and
liver fibrosis in NAFLD patients.
2015 Elsevier Inc. All rights reserved.

Keywords: Nonalcoholic fatty liver disease (NAFLD); Food bioactives; Molecular mechanisms; In vitro studies; Animal models; Clinical trials

1. Introduction mechanism by which selected macro-/micronutrients and food

Nonalcoholic fatty liver disease (NAFLD), also known as hepatic
steatosis, is dened by liver fat deposition in the absence of excessive
alcohol intake [1]. Following the epidemics of obesity, NAFLD has
become the leading cause of liver disease (prevalence, 2034%) [2,3],
and it is epidemiologically associated with the metabolic syndrome
and insulin resistance (IR) [46]. NAFLD is an umbrella term used to
described a histological spectrum ranging from simple steatosis,
dened by a concentration of hepatic triglycerides (TGs) exceeding 5%
of liver weight, to nonalcoholic steatohepatitis (NASH) characterized
by hepatocellular damage, lobular necroinammation and brogen-
esis [7,8]. NASH may evolve to cirrhosis and then to end-stage liver
failure or hepatocellular carcinoma [9,10]. Genetic variants play a
major role in disease predisposition [11] by interacting with
nutritional and other environmental factors, typically hypercaloric
diet and lack of physical activity. To date, there are no approved drugs
for the treatment of NAFLD, and the main clinical recommendation as
an initial step is lifestyle modication.
Systematic reviews on the role of specic nutrients and phyto-
chemicals on NAFLD and related outcomes have recently been
published [12,13]. In this review, we will specically focus on the
Corresponding author. Division of Human Nutrition, Department of
Food, Environmental and Nutritional Sciences (DeFENS), Universit degli
Studi di Milano, 20133 Milano, Italy. Tel.: + 39-02-50316726; fax: + 39-02-
50316721.
E-mail address: Patrizia.riso@unimi.it (P. Riso).
bioactives exert a benecial effect on the hepatic outcomes of NAFLD.
2. Pathophysiology of NAFLD
Fatty liver results from an unbalance between TG accumulation
and removal and represents the safest way to store free fatty acids
(FFAs) in the liver [6,14]. Excess hepatocellular TG derives from
several sources including dietary fatty acids, increased peripheral
lipolysis due to adipose tissue IR and elevated hepatic de novo
lipogenesis due to hyperinsulinemia. Indeed, the major determinant of
NAFLD is systemic IR [4,15]. Reduction of lipid secretion through very
low-density lipoproteins (VLDL) and a decreased fatty acids oxidation
are also involved in hepatic fat accumulation [5].
The development of NASH has been explained by the occurrence of
multiple so-called second-hits leading to the activation of inam-
mation in the context of hepatic steatosis [16,17]. The initial hit
leading to the development of fatty liver renders hepatocytes
susceptible to other multiple hepatotoxic insults including (a)
peroxidation; (b) oxidative stress secondary to free radicals produced
during - and omega-oxidation of FFAs; (c) inammation triggered by
endotoxin engaging Toll-like receptor-4 in Kupffer cells (KCs) and
hepatocytes due to increased intestinal permeability; (d) qualitative
and quantitative changes in gut microbiota [18,19]; (e) hepatic stellate
cells (HSCs) activation; and (f) mitochondrial dysfunction. All these
conditions lead in the end to inammation, cellular damage and
activation of brogenesis [20].

http://dx.doi.org/10.1016/j.jnutbio.2015.08.024
0955-2863/ 2015 Elsevier Inc. All rights reserved.
2 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
3. NAFLD management
The usual management of NAFLD includes lifestyle counseling to
achieve a gradual weight reduction and an increase in physical
activity. Patients are encouraged to lose 8% of their body weight. An
intensive lifestyle intervention focused on diet, exercise and behavior
modication with a goal of 710% weight reduction that leads to
signicant improvement in liver histology in patients with NASH [21].
Indeed, weight loss improves steatosis [22], reduces hepatic inam-
mation and hepatocellular injury [21,23] and improves cardiovascular
risk prole. However, weight loss through energy restriction is
difcult to achieve and sustain [24]. Physical activity and exercise
also effectively decrease steatosis. Cross-sectional and prospective
studies have shown that physical activity decreases intrahepatic lipids
[25,26]. Both aerobic and resistant exercises have been shown to
improve liver function, independently of weight loss [2729].
In addition to total energy intake, the composition of the diet also
affects the metabolic and endocrine functions and overall energy
balance [30]. Most recommendations encourage the consumption of
diets rich in fruits and vegetables for prevention of chronic disease,
and NAFLD is not an exception. Such diets would provide signicant
amount of bioactive components with known benecial effects due in
part to their antiinammatory properties [31].
General recommendations include a reduction in the intakes of
total fat, saturated fatty acids, trans fatty acids and fructose. Indeed,
high fructose intake has been associated with increased risk of NAFLD
and liver damage [3234]. Dietary fructose (consumed in the form of
soft drinks) has been implicated in the pathogenesis of NAFLD [35].
Mice with ad libitum access to fructose solution showed signicantly
higher levels of hepatic lipid accumulation, lipid peroxidation and
endotoxin levels in the portal blood compared to controls and mice fed
with glucose solution [36].
Conversely, an increase in the intakes of polyunsaturated fatty
acids (PUFAs) and monounsaturated fatty acids is advised. Moreover,
there is recommendation to include long-chain n-3 fatty acids to
reduce the risk of NAFLD.
A few trials were conducted to evaluate the impact of specic dietary
patterns on liver damage in patients with NAFLD. In this regard,
Mediterranean diet led to similar weight loss but induced a more
marked reduction of liver enzymes and of IR compared to a low-fat high
carbohydrate diet [30]. Indeed, the diet of patients with NASH is usually
enriched in saturated fat and cholesterol, whereas it is poor in
polyunsaturated fat, bers and antioxidant vitamins C and E [31]. In
addition to an imbalance in fat intake, higher odds of inammation were
associated with higher carbohydrate intake in NASH patients [37].
Apart from lifestyle modication, statins (lipid-lowering drugs),
glitazones (insulin sensitizers), antioxidants and metformin have
been used as therapies for NAFLD [38,39]. Glitazones improve steatosis
at the expense of an increase of weight, and the long-term safety of
their utilization is still not clear. Randomized clinical trials with
antioxidants (Vitamin E and N-acetylcysteine) have given conicting
results, suggesting that their effect may be different depending on age,
dosage and lifestyle modications [39]. A few studies have tested
metformin in nondiabetic patients but with inconsistent results
[40,41].
All these ndings emphasize the difculties to achieve success in
NAFLD clinical setting and attract attention to the importance of
alternative approaches for the prevention of liver damage progression
in NAFLD.
4. Promising food bioactives
In this review, we will focus our attention on the most promising
bioactive compounds studied in the last years for their possible
benecial effects on the prevention and treatment of NAFLD. We have
selected the compounds that have been most investigated in in vitro
and in vivo studies, especially if there is accompanying evidence of
efcacy clinical trials. Evaluated bioactives and their putative
mechanisms of action are listed in Table 1. In the following paragraphs,
we will review the most important evidence supporting their activity
and discuss the evidence supporting the mechanisms of their
benecial effects.
4.1. Omega-3 PUFAs
Long-chain omega-3 (n-3) fatty acids have been proposed as
potential treatment for NAFLD. These fatty acids are present in large
quantities in sh oil, axseed and some nuts. They can be synthesized
in vivo by the human body from -linolenic acid and mainly occur as
eicosapentaenoic acid (EPA) and decosahexaenoic acid (DHA), which
are both antiinammatory. Omega-3 PUFA supplementation amelio-
rates hepatic steatosis in animal models and in human studies [42,43].
Clinical trials, investigating the therapeutic effect of omega-3 in
patients with NAFLD suggested benecial effects on hepatic fat
accumulation, liver function tests, fasting blood glucose and serum
TGs [4446]. A randomized, double-blind, placebo-controlled trial of
DHA, EPA or DHA + EPA supplementation has shown that EPA
enrichment in the peripheral blood is linearly associated with
decreased liver fat percentage in patients with NAFLD [47]. Two
controlled clinical trials performed in NAFLD children also demon-
strated that omega-3 supplementation for 624 months reduced
hepatic steatosis, IR, circulating TG and ALT levels [48,49]. Moreover, a
recent meta-analysis conrmed the benecial effect of omega-3 on
steatosis [50]. Conversely, evidence about necroinammation and
brosis progression in NASH after omega-3 supplementation is still
lacking [51]. However, clinical studies are ongoing, and there is a
strong mechanistic rationale for supporting such an effect. Indeed,
DHA specically binds with high afnity to the G protein-coupled
receptor 120 (GPR120) that mediates potent insulin-sensitizing
effects in vivo by repressing macrophage-induced tissue inammation
[52]. In pediatric NAFLD, DHA treatment reduced liver damage, the
number of inammatory macrophages and increased GPR120 expres-
sion in hepatocytes. Modulation of GPR120 plays a key role in the
regulation of the cell-to-cell cross-talk that drives inammatory
response, hepatic progenitor cell activation and hepatocyte survival
[53,54].
In HepG2 hepatoma cells, the expression of fatty acid synthase
(FAS) and sterol regulatory element binding protein 1c involved in de
novo lipogenesis was suppressed by DHA or EPA supplementation
[55]. Moreover, PUFA supplementation modulated the antioxidant
defense increasing SOD, GST and GPX activity [56].
In high-fat diet (HFD) fed mice, dietary intake of EPA reduced
steatosis by reducing hepatic cholesterol, TG and FFAs [57]. Moreover,
EPA intake seems to abrogate HFD-induced modulation in genes
involved in hepatocellular lipid metabolism. These include up-
regulation of Srebp-1c, which induces the lipogenic program, FAS
and acyl-coenzymeA-carboxylase-1 and the decrease of expression of
carnitinepalmitoyltransferase (CPT1), which transports FFAs to the
mitochondria and promotes -oxidation [58]. Several studies have
demonstrated that EPA decreases steatosis and brosis progression by
reducing TG synthesis and the expression of brogenic genes, and
indeed, it represents an established treatment for hypertriglyc-
eridemia [59,60]. EPA supplementation is associated with decreased
hepatic ROS production and activation of AMP-activated protein
kinase (AMPK) and Peroxisome-proliferator activated receptor-
(PPAR), which stimulates lipid catabolism. In mice fed with HFD
and steatogenic choline-decient diets, DHA supplementation re-
duced hepatic steatosis, inammation, brosis and lipid peroxidation
[61,62]. Increased activity of superoxide dismutase (SOD) and down-
regulation of Srebp-1c seem to account for the inhibitory effect of DHA.
 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111 3

Table 1
Food bioactives for the prevention of nonalcoholic fatty liver: promising compounds and mechanisms
Nutrient/Bioactive Experimental model Mechanism

Omega-3 In vitro Lipogenesis [55]

PUFAs HepG2 cells Antioxidant defence system [56]
In vivo Inammation [52,61,62]
HFD-fed mice, ricesh medaka (Oryzias latipes) Hepatic cholesterol, TG, FFAs [57]
Wistar rat Lipogenesis [58,64]
Lipolysis [58]
Steatosis and brosis [5962]
Lipid peroxidation [61]
Patients Steatosis, FFAs, fasting glucose [44,45]
Adults and children with NAFLD IR, hepatic steatosis, ALT, [49]
Inammatory macrophage [53]
Vitamin E In vitro Lipid peroxidation [78]
Human broblast, rat hepatic stellate cells, human Collagen up-regulation [79]
and mouse hepatocellular carcinoma, Hepa 1-6, Lipogenesis [80]
HepG2 cells -oxidation [80]
In vivo CD36 receptor [81]
Wistar rats, obese (ob/ob) mouse model of NASH, Oxidative stress [82]
guinea pigs Lipid peroxidation [84,85]
Fibrogenesis[84], inammation [82,83], lipid uptake [81]
Patients Steatosis, inammation, brosis [72]
Adults with NASH and NAFLD
Children with NAFLD
Vitamin D In vitro Detoxifying enzyme [100]
HepG2 cells Inammation [103]
Hepatic stellate cells FXR and LXR [100]
Fibrosis [102]
In vivo Lipogenesis [103]
Balb/C mice Lipolysis [103]
D-depleted rat
Patients hs-CRP, serum MDA[97]
Adults with NAFLD
Polyphenol In vitro TG synthesis [112]
HepG2 cells Oxidative stress [113]
Primary rat hepatocyte Lipolysis [114]
Lipogenesis [115]
In vivo De novo lipogenesis [116]
db/db mice fed with MCD Glucose metabolism [118]
Mice fed with HFD Inammation [118]
Liver lipid accumulation [118]
Patients Oxidative stress, inammation [108]
NAFLD patients Body weight, FFA, steatosis [109]
ACNs In vitro Lipogenesis [124]
HepG2 cells Lipolysis [125]
Primary rat hepatocyte Oxidative stress [126]
ROS production [126,128]
In vivo Lipogenesis [127]
Obese mice Steatosis [128]
ApoE3 Leiden mice Inammation, oxidative stress, brosis [129]
Patients Liver enzymes [122]
Adults with NAFLD Oxidative stress, apoptosis [123]
Silybin/silybinin In vitro Lipid accumulation, resistin[139]
HepG2 Inammation, brogenesis [140]
HSC
In vivo IR, ALT [141]
db/db mice fed with MCD Inammation, oxidative stress [142]
Obese fed with MCD-diet mice Visceral fat, gluconeogenesis [143]
Lipolysis [143]
Patients Liver enzymes and histology [137,138]
Adults with NAFLD
Resveratrol In vitro Lipogenesis [148]
HepG2 cells TG synthesis [147]
Primary rat hepatocyte
In vivo Liver weight, TG accumulation [149]
Mice fed HFD-diet Lipogenesis [153]
Zucker rats Lipolysis [154]
Patients Liver enzymes, IR, inammation [144,145]
Adults with NAFLD
Minerals In vitro Selenium
Human hepatoblastoma (C3A) Fibrosis, inammation [162]
HepG2 cells Antioxidant enzymes[162]
Iron
Trasferrin receptor [169]
In vivo Copper deciency
Rabbit Lipolysis, antioxidant system [159]

(continued on next page)

4 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
Table 1 (continued)
Nutrient/Bioactive Experimental model
Sprague-Dawley rats
Abbreviations: necroinammatory score (NAS), sterol regulatory binding protein (Srebp), acetyl-CoA carboxylase (ACC), carnitinepalmitoyltransferase (CPT1), radical oxygen species
(ROS), AMP-activated proten kinase (AMPK), peroxisome proliferator-activated receptor (PPAR), cytochrome P450 3A4 (CYP3A4), transforming growth factor (TGF), tumor necrosis
factor (TNF), cluster of differentiation 36 (CD36), Farnesoid X receptor (FXR), liver X receptor (LXR), C-reactive protein (CRP), malondialdehyde (MDA), glutathione peroxidase (GPX),
thioredoxin reductase (TrxR1).
DHA was also found to ameliorate hepatic steatosis through the down-
regulation of other nuclear receptors involved in TG synthesis such as
Peroxisome proliferator activated receptor and Retionid X receptor
(RXR). Several unsaturated fatty acids, including DHA, have the
capacity to specically bind and activate the RXR, since PUFAs have
been shown to t into the ligand-binding pocket of the RXR crystal
[63]. Finally, both DHA and EPA are able to restore adiponectin levels
which contribute to improve hepatic insulin sensitivity [64].
In summary, omega-3 fatty acids may be a possible therapy for
NAFLD. They have several potential mechanisms of action, the most
relevant is the regulation of hepatic gene expression, thereby switching
intracellular metabolism from lipogenesis and storage to fatty acid
oxidation and catabolism, and activation of antiinammatory pathways.
4.2. Vitamin E
Vitamin E is the generic name for eight lipophilic isoforms: four
tocopherols (, , , -Toc) and four tocotrienols (, , , -T3). Toc is
present in a variety of foods as vegetable oil and nuts whereas T3-
containing foods are limited as palm oil and cereal grains [65]. Both
Toc and T3 have a chroman ring structure with an isoprene side chain.
Toc have a saturated isoprene side chain; conversely, T3 have three
unsaturated bonds in the side chain [66]. Although all forms of Vitamin
E have an antioxidant effect on lipid peroxidation, the most important
for human health are and -Toc due to their dietary abundance.
Despite all the isoforms reach the liver, only -Toc binds -tocopherol
transfer protein that is located in the cytosol of hepatocytes.
Afterwards, it is incorporated into nascent VLDLs and released into
the blood circulation [67]. Since oxidative stress is a major feature of
NASH [6870], Vitamin E has been investigated in this condition.
Clinical trials with Vitamin E supplementation in NASH patients
yielded promising results. In the PIVENS (Pioglitazone, Vitamin E or
Placebo for Non-alcoholic Steatohepatitis) trial, high-dose Vitamin E
(800 IU/day) reduced hepatic necroinammation and facilitated
resolution of NASH, as compared to placebo, but there was not an
improvement in brosis score [71]. A combination of atorvastatin,
vitamin C and high-dose Vitamin E for 4 years reduced hepatic
steatosis (by 71%) in subjects with NAFLD [72]. In another study, 12
patients with NASH and 10 with NAFLD received 300-mg/day -toc
for 1 year. Steatosis, inammation and brosis were improved after
-toc treatment in NASH patient [73].
Few data are available on the effect of antioxidants/Vit E in
paediatric NAFLD. Diet and physical exercise in biopsy-proven NAFLD
children led to a signicant improvement of liver function and glucose
metabolism beyond any antioxidant therapy [74]. In the Treatment of
NAFLD in Children trial, Vitamin E improved hepatocellular ballooning
and resolved NASH more frequently compared to placebo [75].
However, it should be noted that a meta-analysis of randomized
controlled trials showed that high-dosage Vitamin E is associated with
an increase in total mortality, and Vitamin E supplementation has
been associated with increased risk of haemorrhagic stroke and
Mechanism
Lipogenesis [158]
Selenium
TG levels [163]
Cholesterol levels [161]
Antioxidant defenses [163]
Iron
IR and dyslipidemia [168]
prostate cancer [76]. Vitamin E may have prooxidant effect at high
doses disrupting the natural balance of antioxidant systems and
increasing vulnerability to oxidative damage. High-dose Vitamin E
supplements (N or =400 IU/d) have been reported to increase all-
cause mortality suggesting that the dose-dependent effect of Vitamin
E should be carefully considered [77].
The mechanism of action is not limited to its antioxidant
properties, determining reduced mitochondrial damage, but also to
the indirect inuence on other pathways as highlighted in in vitro and
in vivo studies.
Cultured human broblast and rat HSCs treated with Vitamin E
showed a reduction of lipid peroxidation and the inhibition of collagen
gene transcription [78,79]. In addition, T3 supplementation inhibited
TG accumulation in human and mouse hepatoma cells through the
down- and up-regulation of genes involved in lipogenesis and
-oxidation, respectively. In particular, T3 reduced Srebp2 and
Apoliprotein B100 enhancing VLDL efux [80]. The uptake of fatty
acids in the liver is crucial for the establishment and development of
NAFLD. In a recent study, the expression of the fatty acid carrier CD36
was reduced by both Toc and atorvastatin [72,81].
In rats fed with HFD, Vitamin E reduced oxidative stress, protein
nitrosylation and tissue TNF-alpha levels [82]. A 5-week dietary supple-
mentation with either - or -Toc in genetically obese Lepob/obmice
decreased LPS-triggered lipid peroxidation, inammation and hepatic
damage [83]. Moreover, Vitamin E inhibited hepatic TGF-1 gene
expression and protected against liver brosis in rats [84]. The
amelioration of steatohepatitis and the reduction in lipid peroxidation
were also observed in mice and rats fed with a methionine choline-
decient diet [85]. Conversely, in Wistar rats fed with methionine
choline-decient diet supplemented with Vitamin E, despite the
reduction of lipid peroxidation, inltration of inammatory cells, lipid
deposition and brosis were not prevented [67].
In conclusion, Vitamin E supplementation could be considered a
therapeutic tool in NAFLD management. However, the optimal
benetrisk ratio has to be determined for the specic individual.
4.3. Vitamin D
Vitamin D refers to a group of fat-soluble secosteroid hormones
which are involved in the regulation of mineral and skeletal
homeostasis. Vitamin D derives from both dietary sources, such as
oily sh (Vitamin D2) and from dermal synthesis (Vitamin D3). In the
skin, ultraviolet radiation converts 7-dehydrocholesterol to pre-
Vitamin D3 and then to Vitamin D3. In the liver, Vitamin D is
metabolized to 25-hydroxyvitamin D [86] or calcidiol. Calcidiol is then
transported to the kidney where it is further hydroxylated to the active
form (1,25-dihydroxyvitamin D) or calcitriol [87]. The production of
calcitriol is regulated by hormonal cues (such as parathyroid hormone
and FGF-19) and serum calcium and phosphate levels [88].
Besides calcium and bone homeostasis, Vitamin D also regulates
cell proliferation and differentiation and has immunomodulatory,
 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
antiinammatory and antibrotic properties. Evidence showed that
Vitamin D deciency contributes to the development of IR and NAFLD
[86]. This is relevant since up to 55% of adolescents in the US were
reported to be Vitamin D decient [89]. Obese children are more likely
to be sedentary with reduced sunlight exposure, and often they
consume caloric foods low in vitamin content [90,91]. Moreover,
Vitamin D levels are related to the histological severity of steatosis,
necroinammation and brosis [87,9295].
However, short-term Vitamin D supplementation did not consis-
tently improve NAFLD histological features or dyslipidemia in affected
subjects [96], while longer-term supplementation was associated with
reduced inammation and lipid peroxidation [97].
The biological activity of calcitriol is mediated by binding and
transactivation of the nuclear Vitamin D receptor (VDR) [98,99]. In
hepatoma cells, VDR directly induces detoxifying enzymes and
regulate bile acid homeostasis [100]. Vitamin D could avoid excessive
inammatory response in hepatic macrophages since these immune
cells express both VDR and 1 hydroxylase [101].
HSCs are the main producers of extracellular matrix components
playing a pivotal role in liver brosis. VDR is expressed in HSCs and
orchestrates myobroblast trans-differentiation and the brogenic
program. VDR ligands inhibit HSC activation by TGF and abrogate
brotic gene expression, whereas VDR knockout mice spontaneously
develop hepatic brosis. Mechanistic studies revealed that activation
of VDR signalling antagonizes a wide range of TGF/SMAD-dependent
transcriptional responses on probrotic genes in HSCs, suggesting that
the dynamic VDR/SMAD circuit could represent a possible target for
antibrotic therapy [102]. In vivo studies tried to investigate further
applications of Vitamin D as therapy in NAFLD. In rodents, Vitamin D
signaling was involved in maintaining hepatic lipid homeostasis, and
Vitamin D depletion promoted NASH development [103,104]. On the
other hand, Vitamin D supplementation attenuated HFD-induced
hepatic steatosis in a dose-dependent manner along with improve-
ment in serum lipid prole, by decreasing lipogenesis and promoting
FFAs oxidation [105].
Although the potential use of Vitamin D in NAFLD has become more
intriguing with preclinical model of steatosis, further studies are
necessary to better clarify the biological activity of Vitamin D and the
clinical impact of supplementation.
4.4. Polyphenols
Fruits, vegetable and beverages including fruit juices, wine, tea,
coffee and chocolate are important sources of bioactive compounds
such as polyphenols.
Polyphenols are a group of phytochemicals mostly investigated for
their potential role in the prevention and treatment of oxidative stress
and inammation. Polyphenols are secondary metabolites of plants;
they are characterized by the presence of at least one aromatic ring in
their structure, linked to different chemical group as phenolic,
hydroxyl or carbon groups. They can be classied based on their
source, biological function and chemical structure.
Polyphenols are generally subdivided in avonoids and nonavo-
noids depending on their chemical structure. Flavonoids are the most
abundant in the diet and include avonols (e.g. quercetin and
kaempferol, avones (e.g., luteolin, apigenin), avan-3-ols (e.g.,
catechins), avanones (e.g., hesperetin, naringenin), isoavones
(e.g., genistein), anthocyanidins (i.e., cyanidin, malvidin, pelargonidin,
delphinidin, peonidin, petunidin) and proanthocyanidins (i.e., con-
densed tannins). Nonavonoids are represented by stilbenes, phenolic
acids and hydroxycinnamates [106,107].
Several in vitro and in vivo studies investigated polyphenols
properties related to NAFLD despite having few clinical evidences of
the benecial effect in NAFLD treatment. In a randomized, placebo-
controlled, double-blind trial, participants received 250 ml of bayberry
5
juice twice daily for 4 weeks. The consumption of polyphenols-rich
bayberry juice reduced the levels of oxidative (i.e., protein carbonyl
groups), inammatory (i.e., TNF and interleukin-8) and apoptotic
(i.e., tissue polypeptide-specic antigen and cytokeratin-18 fragment
M30) biomarkers in young individuals with NAFLD [108]. Another
study demonstrated that a Hibiscus sabdariffa L. extract rich in
polyphenols (1.43% of avonoids, 2.5% anthocyanins (ACNs) and
1.7% phenolic acid), administered in capsules, was able to decrease
body weight, serum FFAs and improve liver steatosis in overweight
subject [109].
It must be considered that avonoids are a very heterogenic group
of compounds with numerous benecial effects; indeed, they could
target different pathways possibly involved in the pathogenesis of
liver diseases [110]. First of all, avonoids could control de novo
lipogenesis, inhibiting lipogenic (ACC, SREBP-1, FAS, LXR) and
increasing lypolitic enzymes (AMPK, PPAR, CPT-1). Secondly,
avonoids are very effective scavengers since they can protect or
enhance the endogenous antioxidant defense. Lastly, avonoids have
antiinammatory properties inhibiting NFB pathway [110]. In HepG2
hepatoma cells, polyphenols reduce the activity of hydroxymethilglu-
taryl-CoA lyase [111] and the activity of acyl-coenzyme A diacylglyc-
erol acyltransferase, which catalizes the nal step in TG synthesis
[112]. In particular, within polyphenols family, quercitin inhibits TAG
accumulation and promote cell proliferation in HepG2, while SOD,
GPX and CAT activities are up-regulated [113]. Primary rat hepato-
cytes treated with phenolic fraction show a down-regulation of ACC
and hydroxymethilglutaryl-CoA reductase (HMGCR), regulating cho-
lesterol synthesis and activities [114]. Caffeic acid addition in HepG2
cells induces hepatic lipolysis and reduces hepatic lipogenesis up-
regulating AMPK and PPAR and decreasing ACC, SREBP-1 and FAS
protein expression [115].
Concerning in vivo studies, the investigation of polyphenols
properties have been performed using different poliphenols-rich
extracts in heterogeneous animal models. In genetically obese db/db
mice, a polyphenol extract reduced the activity of lipogenic enzymes
involved in de novo fatty acid biosynthesis [116]. Moreover, in mice
fed with HFD enriched in polyphenols, the up-regulation of fatty acid
and TG synthesis-related genes (FAS, SCD1) was reversed [117]. In
diet-induced obese C57BL/mice, a polyphenol-rich Rutgers Scarlet
Lettuce improves glucose metabolism, liver lipid accumulation and
reduced TNF- [118]. In mice with fatty liver, induced by orally
supplementation of high-fat milk, a polyphenol-rich Chrysanthemum
morifolium extract decreased lipid accumulation and hepatic PPAR
gene expression [119]. In the light of this promising evidence,
supplementation of polyphenols may represent a useful approach
for the management of patients with NAFLD, but additional research is
required to conrm this initial data.
4.4.1. Anthocyanins
ACNs, a subclass of avonoids, have been largely investigated for
their potential protective effect in the prevention and treatment of
different diseases. ACNs are the principal components of the red, blue
and purple pigments of the majority of ower petals, fruits and
vegetables such as blueberries, blackberries, raspberries, strawberries,
blackcurrants, elderberries, grapes, cranberries, red cabbage, red
radishes and spinach. ACNs in plants mainly exist in glycosidic
forms; a total of more than 500 ACNs are known depending on the
hydroxylation, methoxylation patterns on the B ring and glycosylation
with different sugar units [120]. The colour of ACNs is pH dependent,
that is, red in acidic and blue in basic conditions, and they are
chemically stable in acidic solutions [107]. To evaluate their benecial
effects on human health, it must be considered that ACNs are rapidly
metabolized and their presence in the circulation is limited to a few
hours. Despite their low absorption and rapid metabolism, regular
intake of ACNs may ameliorate hyperglycaemia, modulate endothelial
6 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
function and decrease inammation [121]. Moreover, their role has
been investigated in the prevention of oxidative stress by scavenging
reactive oxygen species and free radicals [122] and their role in the
modulation of lipid metabolism and fat deposition [108,123] in
different tissues, including the liver. As we recently reviewed [121],
ACNs can reduce hepatic lipid accumulation, but their impact on
NAFLD has yet to be understood. Until now, only few clinical studies on
humans are available, and they diverge for ACNs source, doses and
clinical features of patients. Suda et al. showed an effect of ACN intake
(200-mg acylated ACNs from purple sweet potato) in the reduction of
liver enzymes (e.g., gamma-glutamyltransferases) in subjects with
borderline levels of one or more hepatic markers [122]. In a recent
study, NAFLD patients received either puried ACNs (320 mg/d)
derived from bilberry and black currant or placebo for 12 weeks.
Individuals receiving ACNs showed a decrease in plasma alanine
aminotransferase, cytokeratin-18 fragment and myeloperoxidase and
an overall improvement of IR [123].
Several in vitro studies, performed mainly in HepG2 cells
supplemented with oleic acid and/or palmitic acid, highlighted three
different mechanisms of action by which ACNs could prevent the
progression of liver dysfunction/damage: inhibition of lipogenesis
(i.e., reducing SREBP1c), promotion of lipolysis (i.e., inducing PPAR
activity with activation of AMPK pathway) and reduction of oxidative
stress (i.e., induction of antioxidant enzymes). Mulberry ACNs (0.1,
0.3, 0.5 mg/ml) supplementation in HepG2 reduced lipogenesis
(SREBP-1, FAS, ACC and A-FABP), cholesterol biosynthesis (SREBP-2
and HMGCR) and TG biosynthesis and enhanced fatty acid -oxidation
(PPAR and CPT-1) [124]. Cyanidin 3-O-glucoside reduced cellular
lipid concentration in HepG2 cells by rewiring the expression of genes
involved in lipid metabolic pathway as PPAR [125], whereas in
primary mice hepatocytes it decreased intracellular ROS production
acting as free radical scavenger and enhanced PI3K/Akt activation
[126].
In vivo studies were performed in different experimental models of
NAFLD and metabolic syndrome and evaluated different outcomes as
lipid metabolism, oxidative stress and liver damage. Obese mice
supplemented with 200 mg/kg per day of ACN fraction extracted from
purple sweet potato showed a reduced hepatic fat accumulation
associated with a decreased hepatic lipogenesis [127]. Moro juice with
an ACN content of 85 mg/l administered ad libitum to mice prevented
fatty liver suppressing LXR- expression and activity [128]. Moreover,
1 g of puried ACN from bilberry and blackcurrant administered to
mice ameliorated hepatic steatosis, inammation, oxidative stress as
well as brosis [129]. ACN-rich bilberry (Vaccinium myrtillus L.) extract
was tested in E3Leiden mice fed with high-fat/high-cholesterol diet.
The ACN extract reduced NASH development, attenuating both
steatosis and inammation, and reduced hepatic brosis. These effects
were associated with a decreased hepatic free cholesterol accumula-
tion and cholesterol crystal formation. On the basis of these data, ACN-
rich food could be useful for the prevention of liver diseases as NAFLD,
even if additional studies are needed to deeply characterize the
molecular mechanism of the different extracts.
4.4.2. Silibinin
Silymarin and its major constituent, Silibinin, are avonoid
compounds extracted from the medicinal plant Silybum marianum
(milk thistle). Extracts have traditionally been used for the treatments
of liver disease [130134].
Only few controlled randomized studies has been conducted in
patients with NAFLD. These studies suggested that silymarin could
reduce steatosis severity, liver ballooning and brosis, acting also as
efcient insulin sensitizer and lowering aminotransferase levels in
both short- and long-term trials [135]. In a study where patients
received Vitamin E, L-gluthatione, L-cysteine, L-methionine and Sily-
bum Marianum, it was observed that silymarin was effective in
reducing the biochemical and ultrasonographic changes induced by
NAFLD. These data were in agreement with those obtained by other
authors [136]. Similarly, in a multicenter, double-blind clinical trial,
patients with steatosis received silybinin combined with phosphati-
dylcholine and Vitamin E for 12 months. Combined treatment was
associated with an improvement in liver enzymes, IR and liver
histology without increase in body weight [137]. In another recent
study, the intake of 210-mg/day silymarin orally for 8 weeks
decreased hepatic enzymes in patients with NASH [138].
Few in vitro studies investigated silibinin properties. In hepatoma
cells, silibinin prevented lipid accumulation and resistin induction by
fatty acids targeting the insulin signaling pathway [139]. In HSCs
isolated from human liver, silybin inhibited dose dependently cell
proliferation, cell motility and de novo synthesis of extracellular
matrix components. Silibinin was also conrmed to act as a potent
antioxidant and antiinammatory agent [140].
As concern in vivo studies, in obese db/db mice fed with MCD,
silibinin decreased IR, serum ALT and NAFLD histological activity. This
was associated with reduced oxidative stress and inammation, due to
lower isoprostanes, 8-OHG and TNF- expression and restored
mitochondrial reduced glutathion levels [141]. Again, in obese mice
fed with MCD and in rats fed with HFD, silibinin improved hepatic
oxidative and nitrosative stress mediated by iNOS and inammation
modulating the expression and the activity of lipid metabolic enzymes
[142]. This resulted in improvement in liver damage. In HFD rats,
silibilin ameliorated IR mainly by reducing visceral fat, up-regulating
lipolysis and inhibiting gluconeogenesis [143].
These ndings are a rst step in the comprehension of the plausible
mechanisms of action of silibilinin, but further work is necessary to
better characterize the possible use of these polyphenolic compounds.
4.4.3. Resveratrol
Resveratrol (trans-3,4,5-trihydroxystilbene) is a stilbene occur-
ring naturally in several plants and provided in the diet by various food
stuffs such as grapes, berries, red wine and nuts. Evidences have
shown its health benets, such as improvement of insulin sensitivity
and glucose tolerance, reduction of serum lipids and suppression of
inammation and oxidative stress. Moreover, resveratrol is able to
modify lipid metabolism and more specically to induce a reduction in
liver TG content [144]. Few studies were performed in humans with
conicting results. Recently, patients with NAFLD who received
1500-mg resveratrol capsule twice daily for 12 weeks had an
improvement of liver enzymes, IR and inammation [145]. Similarly,
patients receiving 50-mg resveratrol twice daily for 3 months
associated with lifestyle modication had a higher attenuation of
inammatory markers and hepatocellular apoptosis compared to
placebo treatment [144]. Conversely, 8 weeks administration of 3000-
mg resveratrol led to increased liver enzymes in NAFLD patients [146].
Few in vitro studies investigated the potential hepatoprotective
effect of resveratrol. In primary rat hepatocytes, resveratrol supple-
mentation (25 mol/l) reduced ACC activity [147]. In a study
performed in HepG2 cells exposed to high concentration of glucose,
resveratrol supplementation reduced triacylglycerol accumulation by
increasing AMPK activity and down-regulating SREBP-1c and ACC
activity [148].
In vivo studies revealed that resveratrol could reduce liver weight
and TG content. In mice treated with 22.4-mg resveratrol/kg,
histological examination revealed a reduced accumulation of large
lipid droplets [149]. Recent studies in mice fed with HFD showed that
resveratrol protected the liver from fat accumulation by activating
Sirt1 [150,151]. Sirt1, a NAD-dependent deacetylase, stimulates the
activity of FOXO1 [152], which may in turn indirectly inhibit SREBP1
expression [153,154], suggesting that the AMPK/Sirt1 axis could
down-regulate genes of the lipogenic pathways (FAS, ACC) and up-
regulate genes of the lypolitic pathway (PPAR, CPT-1). As concern in
 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111 7

Silibinin
Omega-3 Minerals
Minerals Omega-3
Polyphenols Vit E
Anthocyanins
Vit D

Fibrosis
-ox
TGs ROS
HSCs

Omega-3
Minerals TGs
Polyphenols IL-6
Anthocyanins DNL TNF
Resveratrol Srebp, Fas, Acc IL-1
Vit D and E
KC

Silibinin
Omega-3
FFAs Polyphenols
LDL Anthocyanins
M
Resveratrol
Vit D and E

IL-6
TNF
IL-1

Fig. 1. Molecular mechanisms explaining the hepatoprotective effect of food bioactives. Development of NAFLD/NASH is induced by different risk factors, such as western-type diet, physical
inactivity and genetic predisposition. In the presence of obesity and IR, there is an increased ux of FFAs to the liver. These FFAs are stored as TG in lipid droplets leading to hepatic fat
accumulation or undergo -oxidation increasing oxidative stress and the inammatory pathway.The damaged hepatocyte leads to a further increase of inammatory signalling (IL-1, TNFa,
IL-6) and the recruitment of circulating and residual macrophages (KCs). All of these mechanisms can directly induce the activation of HSCs, the major cell type involved in extracellular
matrix deposition and liver brosis. The bioactive compounds may exert benecial effects on NAFLD development and progression by inhibiting lipogenesis, -oxidation of FFAs,
inammation and HSCs activation. In the cartoon, we have listed the food bioactives indicating the putative mechanisms by which they may improve liver damage in NAFLD.

the antioxidant activity, in Zucker rats supplemented with two
different doses of resveratrol (15 and 45 mg/kg body weight), it was
able to reduce oxidative damage in liver measured as hepatic
thiobarbituric acid and oxidized glutathione (GSSG) levels [153].
In conclusion, recent evidence demonstrated that resveratrol is
effective in decreasing de novo lipogenesis in the liver and it could be
used in the prevention of liver diseases.
5. Minerals
It is generally recognised a progressive decay in the homeostasis of
trace minerals in patients with NAFLD; this may reect an increased
oxidative stress and inammation condition. In particular, minerals
such as copper, selenium and iron have been investigated for their
possible contribution to the development and treatment of liver
diseases as NAFLD.
5.1. Copper
Copper has a role in antioxidant defence, lipid peroxidation and
mitochondrial function. Copper deciency has been linked to different
metabolic disorders as hypercholesterolemia, increased blood pres-
sure and glucose intolerance both in rodents [155,156] and in humans
[157]. Since inadequate copper availability may increase lipid
accumulation in the liver, its potential contribution to the develop-
ment of NAFLD has been investigated. Hepatic copper concentrations
were reduced in NAFLD and inversely correlated with steatosis, NASH
and IR [157]. Development of hepatic steatosis and IR in response to
dietary copper restriction in rats suggests that copper availability has a
causal role in the development of NAFLD [157].
Interestingly, fructose feeding exacerbates complications of copper
deciency. In rats, fructose consumption impaired copper status and
precipitated copper deciency possibly inhibiting its absorption
through the intestinal epithelium. Moreover, copper deciency and
fructose appear to act together to accelerate hepatic fat accumulation
and liver damage [156,158]. According with histological ndings,
copper deciency markedly suppressed CPT-1 and up-regulated FAS
expression [158]. In addition, copper deciency also contributed to an
impaired antioxidant defence system considering that the activity of
Cu/Zn SOD depends on adequate copper availability [159].
5.2. Selenium
Selenium deciency could be considered a dietary condition
correlated with oxidative stress in patients with liver diseases [160].
Selenium levels have been associated with cardiovascular disease, and
8 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
its supplementation leads to decrease in total cholesterol and TG levels
[161]. Unfortunately, selenium status on NAFLD has not been yet
investigated.
Human hepatoblastoma cells supplemented with selenite showed
a reduced TGF1-induced collagen and IL-8 production and maxi-
mized the expression of antioxidant enzymes in response to FFAs
overload [162]. In experimental models selenium supplementation
decreased TG levels, protected low-density lipoprotein (LDL) from
oxidation by restoring the antioxidant properties of the LDL-
associated enzyme Paraoxonase 1 [163].
Considering that in vitro and in vivo selenium supplementation has
shown a potential effect in the reduction of oxidative stress, it is
important to take into account the possible clinical implication in
subjects with liver disease who have selenium deciency.
5.3. Iron
Iron is essential to the life of all mammalian organisms. It has a key
role in oxygen transport and in enzymes involved in mitochondrial
respiration, DNA biosynthesis and the citric acid cycle via the
capability to change its redox state. However, this characteristic also
renders excess iron detrimental, mostly via the formation of reactive
oxygen species, which may lead to severe organ damage. Iron
perturbations are frequently observed in patients with obesity, IR
and NAFLD. The term dysmetabolic iron overload syndrome (DIOS) is
now most frequently used to describe the typical association of hepatic
steatosis with mild-to-moderate iron deposition in liver biopsies and
increased serum ferritin in patients with NAFLD [164].
Hyperferritinemia is usually associated with NASH and the severity
of liver damage, whereas iron depletion, achieved by phlebotomy, in
patients with mild iron overload could have benecial effects more
than lifestyle modications alone in normalizing IR and liver enzymes
[165,166]. Moreover, iron depletion up-regulated glucose uptake and
increased insulin receptor expression and signaling in hepatocytes in
vitro and in vivo[167], whereas dietary iron supplementation induced
IR and dyslipidemia [168]. To investigate the mechanisms underlying
DIOS, we recently examined the effect of FFAs on hepatic iron
metabolism. The main nding was that exposure of hepatocytes to
FFAs, leading to steatosis, was associated with a subversion of iron
metabolism characterized by increased expression of transferrin
receptor and facilitation of iron storage [169]. Additional studies are
warranted to evaluate the potential of iron reductive therapy on
clinical outcomes in NAFLD patients.
6. Conclusions
NAFLD, a disease caused by an unhealthy eating behaviour and
lifestyle has become the leading cause of liver diseases in the western
countries. To date, there are no approved drugs for this indication, and
the main clinical recommendation as an initial step is lifestyle
modication including both improvement of dietary pattern and
increased physical activity. The identication of the molecular
mechanisms leading to fat accumulation, oxidative balance impair-
ment and liver brosis is expected to improve both diagnostic and
therapeutic approaches (Fig. 1). Food bioactive compounds, which
modulate the activation of genes involved in lipogenesis, brogenesis,
lipid peroxidation and inammation, represent a new attractive
therapeutic approach for this condition.
Acknowledgements
The authors would like to thank Dr Cristian Del Bo and the
Metabolic Liver Diseases Research Lab for careful reading of the
manuscript and suggestions. Luca Valenti and Patrizia Riso designed
the study and critically reviewed the paper literature and approved
the nal manuscript. Paola Dongiovanni wrote the paper draft and
contributed to critical discussion of results. Claudia Lanti performed
the literature search and contributed to the writing of the paper draft.
The authors declare no conict of interest.
References
[1] Angulo P. Nonalcoholic fatty liver disease. N Engl J Med 2002;346:122131.
[2] Browning JD, Szczepaniak LS, Dobbins R, Nuremberg P, Horton JD, Cohen JC, et al.
Prevalence of hepatic steatosis in an urban population in the United States:
impact of ethnicity. Hepatology 2004;40:138795.
[3] Blachier M, Leleu H, Peck-Radosavljevic M, Valla DC, Roudot-Thoraval F. The
burden of liver disease in Europe: a review of available epidemiological data.
J Hepatol 2013;58:593608.
[4] Marchesini G, Brizi M, Morselli-Labate AM, Bianchi G, Bugianesi E, McCullough
AJ, et al. Association of nonalcoholic fatty liver disease with insulin resistance.
Am J Med 1999;107:4505.
[5] Fabbrini E, Mohammed BS, Magkos F, Korenblat KM, Patterson BW, Klein S.
Alterations in adipose tissue and hepatic lipid kinetics in obese men and women
with nonalcoholic fatty liver disease. Gastroenterology 2008;134:42431.
[6] Cohen JC, Horton JD, Hobbs HH. Human fatty liver disease: old questions and
new insights. Science 2011;332:151923.
[7] Kleiner DE, Brunt EM, Van Natta M, Behling C, Contos MJ, Cummings OW, et al.
Design and validation of a histological scoring system for nonalcoholic fatty liver
disease. Hepatology 2005;41:131321.
[8] Day CP. From fat to inflammation. Gastroenterology 2006;130:20710.
[9] Bugianesi E, Leone N, Vanni E, Marchesini G, Brunello F, Carucci P, et al.
Expanding the natural history of nonalcoholic steatohepatitis: from cryptogenic
cirrhosis to hepatocellular carcinoma. Gastroenterology 2002;123:13440.
[10] Liu YL, Patman GL, Leathart JB, Piguet AC, Burt AD, Dufour JF, et al. Carriage of the
PNPLA3 rs738409 C N G polymorphism confers an increased risk of non-alcoholic
fatty liver disease associated hepatocellular carcinoma. J Hepatol 2014;61:
7581.
[11] Dongiovanni P, Anstee QM, Valenti L. Genetic predisposition in NAFLD and
NASH: impact on severity of liver disease and response to treatment. Curr Pharm
Des 2013;19:521938.
[12] Eslamparast T, Eghtesad S, Poustchi H, Hekmatdoost A. Recent advances in
dietary supplementation, in treating non-alcoholic fatty liver disease. World J
Hepatol 2015;7:20412.
[13] Aguirre L, Portillo MP, Hijona E, Bujanda L. Effects of resveratrol and other
polyphenols in hepatic steatosis. World J Gastroenterol 2014;20:736680.
[14] Yamaguchi K, Yang L, McCall S, Huang J, Yu XX, Pandey SK, et al. Inhibiting
triglyceride synthesis improves hepatic steatosis but exacerbates liver damage
and fibrosis in obese mice with nonalcoholic steatohepatitis. Hepatology 2007;
45:136674.
[15] Marchesini G, Brizi M, Bianchi G, Tomassetti S, Bugianesi E, Lenzi M, et al.
Nonalcoholic fatty liver disease: a feature of the metabolic syndrome. Diabetes
2001;50:184450.
[16] Day CP. Steatohepatitis: a tale of two "hits"? Gastroenterology 1998;114:8425.
[17] Tilg H, Moschen AR. Evolution of inflammation in nonalcoholic fatty liver
disease: the multiple parallel hits hypothesis. Hepatology 18361846;52:2010.
[18] Valenti L, Ludovica Fracanzani A, Fargion S. The immunopathogenesis of
alcoholic and nonalcoholic steatohepatitis: two triggers for one disease?
Semin Immunopathol 2009;31(3):35969.
[19] Miele L, Valenza V, La Torre G, Montalto M, Cammarota G, Ricci R, et al. Increased
intestinal permeability and tight junction alterations in nonalcoholic fatty liver
disease. Hepatology 2009;49(6):187787.
[20] Anstee QM, Daly AK, Day CP. Genetics of alcoholic and nonalcoholic fatty liver
disease. Semin Liver Dis 2011;31(2):12846.
[21] Promrat K, Kleiner DE, Niemeier HM, Jackvony E, Kearns M, Wands JR, et al.
Randomized controlled trial testing the effects of weight loss on nonalcoholic
steatohepatitis. Hepatology 2010;51:1219.
[22] Sullivan S, Kirk EP, Mittendorfer B, Patterson BW, Klein S. Randomized trial of
exercise effect on intrahepatic triglyceride content and lipid kinetics in
nonalcoholic fatty liver disease. Hepatology 2012;55:173845.
[23] Harrison SA, Fecht W, Brunt EM, Neuschwander-Tetri BA. Orlistat for overweight
subjects with nonalcoholic steatohepatitis: a randomized, prospective trial.
Hepatology 2009;49:806.
[24] Centis E, Moscatiello S, Bugianesi E, Bellentani S, Fracanzani AL, Calugi S, et al.
Stage of change and motivation to healthier lifestyle in non-alcoholic fatty liver
disease. J Hepatol 2014;58:7717.
[25] St George A, Bauman A, Johnston A, Farrell G, Chey T, George J. Independent
effects of physical activity in patients with nonalcoholic fatty liver disease.
Hepatology 2009;50:6876.
[26] Perseghin G, Lattuada G, De Cobelli F, Ragogna F, Ntali G, Esposito A, et al.
Habitual physical activity is associated with intrahepatic fat content in humans.
Diabetes Care 2007;30:6838.
[27] Johnson NA, Sachinwalla T, Walton DW, Smith K, Armstrong A, Thompson MW,
et al. Aerobic exercise training reduces hepatic and visceral lipids in obese
individuals without weight loss. Hepatology 2009;50:110512.
 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
[28] Sreenivasa Baba C, Alexander G, Kalyani B, Pandey R, Rastogi S, Pandey A, et al.
Effect of exercise and dietary modification on serum aminotransferase levels in
patients with nonalcoholic steatohepatitis. J Gastroenterol Hepatol 2006;21:
1918.
[29] Hallsworth K, Fattakhova G, Hollingsworth KG, Thoma C, Moore S, Taylor R, et al.
Resistance exercise reduces liver fat and its mediators in non-alcoholic fatty liver
disease independent of weight loss. Gut 2011;60:127883.
[30] Ryan MC, Desmond P, Wilson A. Reply to: "might some of the beneficial effects of
the Mediterranean diet on non-alcoholic fatty liver disease be mediated by
reduced iron stores?". J Hepatol 2013;59:640.
[31] Musso G, Gambino R, De Michieli F, Cassader M, Rizzetto M, Durazzo M, et al.
Dietary habits and their relations to insulin resistance and postprandial lipemia
in nonalcoholic steatohepatitis. Hepatology 2003;37:90916.
[32] Ferder L, Ferder MD, Inserra F. The role of high-fructose corn syrup in metabolic
syndrome and hypertension. Curr Hypertens Rep 2010;12:10512.
[33] Chiu S, Sievenpiper JL, de Souza RJ, Cozma AI, Mirrahimi A, Carleton AJ, et al.
Effect of fructose on markers of non-alcoholic fatty liver disease (NAFLD): a
systematic review and meta-analysis of controlled feeding trials. Eur J Clin Nutr
2014;68:41623.
[34] Vos MB, Lavine JE. Dietary fructose in nonalcoholic fatty liver disease.
Hepatology 2013;57:252531.
[35] Ma J, Fox CS, Jacques PF, Speliotes EK, Hoffmann U, Smith CE, et al. Sugar-
sweetened beverage, diet soda, and fatty liver disease in the Framingham Heart
Study cohorts. J Hepatol 2015.
[36] Bergheim I, Weber S, Vos M, Kramer S, Volynets V, Kaserouni S, et al. Antibiotics
protect against fructose-induced hepatic lipid accumulation in mice: role of
endotoxin. J Hepatol 2008;48:98392.
[37] Toshimitsu K, Matsuura B, Ohkubo I, Niiya T, Furukawa S, Hiasa Y, et al. Dietary
habits and nutrient intake in non-alcoholic steatohepatitis. Nutrition 2007;23:
4652.
[38] Lomonaco R, Sunny NE, Bril F, Cusi K. Nonalcoholic fatty liver disease: current
issues and novel treatment approaches. Drugs 2013;73:114.
[39] Musso G, Gambino R, Cassader M, Pagano G. A meta-analysis of randomized
trials for the treatment of nonalcoholic fatty liver disease. Hepatology 2010;52:
79104.
[40] Duseja A, Das A, Dhiman RK, Chawla YK, Thumburu KT, Bhadada S, et al.
Metformin is effective in achieving biochemical response in patients with
nonalcoholic fatty liver disease (NAFLD) not responding to lifestyle interven-
tions. Ann Hepatol 2007;6:2226.
[41] Rakoski MO, Singal AG, Rogers MA, Conjeevaram H. Meta-analysis: insulin
sensitizers for the treatment of non-alcoholic steatohepatitis. Aliment Pharma-
col Ther 2010;32:121121.
[42] Zhu FS, Liu S, Chen XM, Huang ZG, Zhang DW. Effects of n-3 polyunsaturated
fatty acids from seal oils on nonalcoholic fatty liver disease associated with
hyperlipidemia. World J Gastroenterol 2008;14:6395400.
[43] Masterton GS, Plevris JN, Hayes PC. Review article: omega-3 fatty acids - a
promising novel therapy for non-alcoholic fatty liver disease. Aliment Pharmacol
Ther 2010;31:67992.
[44] Capanni M, Calella F, Biagini MR, Genise S, Raimondi L, Bedogni G, et al.
Prolonged n-3 polyunsaturated fatty acid supplementation ameliorates hepatic
steatosis in patients with non-alcoholic fatty liver disease: a pilot study. Aliment
Pharmacol Ther 2006;23:114351.
[45] Spadaro L, Magliocco O, Spampinato D, Piro S, Oliveri C, Alagona C, et al. Effects of
n-3 polyunsaturated fatty acids in subjects with nonalcoholic fatty liver disease.
Dig Liver Dis 2008;40:1949.
[46] Tanaka N, Sano K, Horiuchi A, Tanaka E, Kiyosawa K, Aoyama T. Highly purified
eicosapentaenoic acid treatment improves nonalcoholic steatohepatitis. J Clin
Gastroenterol 2008;42:4138.
[47] Scorletti E, Bhatia L, McCormick KG, Clough GF, Nash K, Calder PC, et al.
Design and rationale of the WELCOME trial: A randomised, placebo
controlled study to test the efficacy of purified long chainomega-3 fatty
acid treatment in non-alcoholic fatty liver disease [corrected]. Contemp Clin
Trials 2014;37:30111.
[48] Nobili V, Bedogni G, Alisi A, Pietrobattista A, Rise P, Galli C, et al. Docosahexaenoic
acid supplementation decreases liver fat content in children with non-alcoholic
fatty liver disease: double-blind randomised controlled clinical trial. Arch Dis
Child 2011;96:3503.
[49] Nobili V, Alisi A, Della Corte C, Rise P, Galli C, Agostoni C, et al. Docosahexaenoic
acid for the treatment of fatty liver: randomised controlled trial in children. Nutr
Metab Cardiovasc Dis 2013;23:106670.
[50] Parker HM, Johnson NA, Burdon CA, Cohn JS, O'Connor HT, George J. Omega-3
supplementation and non-alcoholic fatty liver disease: a systematic review and
meta-analysis. J Hepatol 2012;56:94451.
[51] Argo CK, Patrie JT, Lackner C, Henry TD, de Lange EE, Weltman AL, et al. Effects of
n-3 fish oil on metabolic and histological parameters in NASH: a double-blind,
randomized, placebo-controlled trial. J Hepatol 2015;62:1907.
[52] Oh DY, Talukdar S, Bae EJ, Imamura T, Morinaga H, Fan W, et al. GPR120 is an
omega-3 fatty acid receptor mediating potent anti-inflammatory and insulin-
sensitizing effects. Cell 2010;142:68798.
[53] Nobili V, Carpino G, Alisi A, De Vito R, Franchitto A, Alpini G, et al. Role of
docosahexaenoic acid treatment in improving liver histology in pediatric
nonalcoholic fatty liver disease. PLoS One;9:e88005.
[54] Valenti L, Nobili V. Deciphering the role of omega3 fatty acids in nonalcoholic
steatohepatitis. J Pediatr Gastroenterol Nutr 2014;59:4234.
9
[55] Nagao K, Nakamitsu K, Ishida H, Yoshinaga K, Nagai T, Mizobe H, et al.
Comparison of the lipid-lowering effects of four different n-3 highly unsaturated
fatty acids in HepG2 cells. J Oleo Sci 2014;63:97985.
[56] Di Nunzio M, Valli V, Bordoni A. Pro- and anti-oxidant effects of polyunsaturated
fatty acid supplementation in HepG2 cells. Prostaglandins Leukot Essent Fatty
Acids 2011;85:1217.
[57] Shiba S, Tsunoda N, Wakutsu M, Muraki E, Sonoda M, Tam PS, et al. Regulation of
lipid metabolism by palmitoleate and eicosapentaenoic acid (EPA) in mice fed a
high-fat diet. Biosci Biotechnol Biochem 2011;75:24013.
[58] Matsumoto T, Terai S, Oishi T, Kuwashiro S, Fujisawa K, Yamamoto N, et al.
Medaka as a model for human nonalcoholic steatohepatitis. Dis Model Mech
2010;3:43140.
[59] Kajikawa S, Imada K, Takeuchi T, Shimizu Y, Kawashima A, Harada T, et al.
Eicosapentaenoic acid attenuates progression of hepatic fibrosis with inhibition
of reactive oxygen species production in rats fed methionine- and choline-
deficient diet. Dig Dis Sci 2011;56:106574.
[60] Kajikawa S, Harada T, Kawashima A, Imada K, Mizuguchi K. Highly purified
eicosapentaenoic acid ethyl ester prevents development of steatosis and hepatic
fibrosis in rats. Dig Dis Sci 2010;55:63141.
[61] Poudyal H, Panchal SK, Ward LC, Brown L. Effects of ALA, EPA and DHA in high-
carbohydrate, high-fat diet-induced metabolic syndrome in rats. J Nutr Biochem
2013;24:104152.
[62] Tang X, Li ZJ, Xu J, Xue Y, Li JZ, Wang JF, et al. Short term effects of different
omega-3 fatty acid formulation on lipid metabolism in mice fed high or low fat
diet. Lipids Health Dis 2012;11:70.
[63] Lengqvist J, Mata De Urquiza A, Bergman AC, Willson TM, Sjovall J, Perlmann T,
et al. Polyunsaturated fatty acids including docosahexaenoic and arachidonic
acid bind to the retinoid X receptor alpha ligand-binding domain. Mol Cell
Proteomics 2004;3:692703.
[64] Vemuri M, Kelley DS, Mackey BE, Rasooly R, Bartolini G. Docosahexaenoic Acid
(DHA) But Not Eicosapentaenoic Acid (EPA) Prevents Trans-10, Cis-12
Conjugated Linoleic Acid (CLA)-Induced Insulin Resistance in Mice. Metab
Syndr Relat Disord 2007;5:31522.
[65] Birringer M, Pfluger P, Kluth D, Landes N, Brigelius-Flohe R. Identities and
differences in the metabolism of tocotrienols and tocopherols in HepG2 cells. J
Nutr 2002;132:31138.
[66] Takitani K, Miyazaki H, Yoden A, Tamai H. Children's toxicology from bench to
bedLiver Injury (2): mechanism of antioxidant therapy for nonalcoholic fatty
liver disease. J Toxicol Sci 2009;34(Suppl. 2):SP2238.
[67] Miyazaki H, Takitani K, Koh M, Yoden A, Tamai H. The alpha-tocopherol status
and expression of alpha-tocopherol-related proteins in methionine-choline
deficient rats treated with vitamin E. J Clin Biochem Nutr 2014;54:1907.
[68] Sanyal AJ, Campbell-Sargent C, Mirshahi F, Rizzo WB, Contos MJ, Sterling RK,
et al. Nonalcoholic steatohepatitis: association of insulin resistance and
mitochondrial abnormalities. Gastroenterology 2001;120:118392.
[69] Seki S, Kitada T, Yamada T, Sakaguchi H, Nakatani K, Wakasa K. In situ detection
of lipid peroxidation and oxidative DNA damage in non-alcoholic fatty liver
diseases. J Hepatol 2002;37:5662.
[70] MacDonald GA, Bridle KR, Ward PJ, Walker NI, Houglum K, George DK, et al. Lipid
peroxidation in hepatic steatosis in humans is associated with hepatic fibrosis
and occurs predominately in acinar zone 3. J Gastroenterol Hepatol 2001;16:
599606.
[71] Sanyal AJ, Chalasani N, Kowdley KV, McCullough A, Diehl AM, Bass NM, et al.
Pioglitazone, vitamin E, or placebo for nonalcoholic steatohepatitis. N Engl J Med
2010;362:167585.
[72] Foster T, Budoff MJ, Saab S, Ahmadi N, Gordon C, Guerci AD. Atorvastatin and
antioxidants for the treatment of nonalcoholic fatty liver disease: the St Francis
Heart Study randomized clinical trial. Am J Gastroenterol 2011;106:717.
[73] Thuma PE, Mabeza GF, Biemba G, Bhat GJ, McLaren CE, Moyo VM, et al. Effect of
iron chelation therapy on mortality in Zambian children with cerebral malaria.
Trans R Soc Trop Med Hyg 1998;92:2148.
[74] Nobili V, Manco M, Devito R, Ciampalini P, Piemonte F, Marcellini M. Effect
of vitamin E on aminotransferase levels and insulin resistance in children
with non-alcoholic fatty liver disease. Aliment Pharmacol Ther 2006;24:
155361.
[75] Lavine JE, Schwimmer JB, Van Natta ML, Molleston JP, Murray KF, Rosenthal P,
et al. Effect of vitamin E or metformin for treatment of nonalcoholic fatty liver
disease in children and adolescents: the TONIC randomized controlled trial.
JAMA 2011;305:165968.
[76] Albanes D, Till C, Klein EA, Goodman PJ, Mondul AM, Weinstein SJ, et al. Plasma
tocopherols and risk of prostate cancer in the Selenium and Vitamin E Cancer
Prevention Trial (SELECT). Cancer Prev Res (Phila) 2014;7:88695.
[77] Miller III ER, Pastor-Barriuso R, Dalal D, Riemersma RA, Appel LJ, Guallar E. Meta-
analysis: high-dosage vitamin E supplementation may increase all-cause
mortality. Ann Intern Med 2005;142:3746.
[78] Houglum K, Brenner DA, Chojkier M. d-alpha-tocopherol inhibits collagen alpha
1(I) gene expression in cultured human fibroblasts. Modulation of constitutive
collagen gene expression by lipid peroxidation. J Clin Invest 1991;87:22305.
[79] Nieto N, Friedman SL, Greenwel P, Cederbaum AI. CYP2E1-mediated oxidative
stress induces collagen type I expression in rat hepatic stellate cells. Hepatology
1999;30:98796.
[80] Zaiden N, Yap WN, Ong S, Xu CH, Teo VH, Chang CP, et al. Gamma delta
tocotrienols reduce hepatic triglyceride synthesis and VLDL secretion. J
Atheroscler Thromb 2010;17:1019932.
10 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
[81] Podszun MC, Grebenstein N, Spruss A, Schlueter T, Kremoser C, Bergheim I, et al.
Dietary alpha-tocopherol and atorvastatin reduce high-fat-induced lipid
accumulation and down-regulate CD36 protein in the liver of guinea pigs. J
Nutr Biochem 2014;25:5739.
[82] Raso GM, Esposito E, Iacono A, Pacilio M, Cuzzocrea S, Canani RB, et al.
Comparative therapeutic effects of metformin and vitamin E in a model of non-
alcoholic steatohepatitis in the young rat. Eur J Pharmacol 2009;604:12531.
[83] Chung MY, Yeung SF, Park HJ, Volek JS, Bruno RS. Dietary alpha- and gamma-
tocopherol supplementation attenuates lipopolysaccharide-induced oxidative
stress and inflammatory-related responses in an obese mouse model of
nonalcoholic steatohepatitis. J Nutr Biochem 2010;21:12006.
[84] Tzanetakou IP, Doulamis IP, Korou LM, Agrogiannis G, Vlachos IS, Pantopoulou A,
et al. Water Soluble Vitamin E Administration in Wistar Rats with Non-alcoholic
Fatty Liver Disease. Open Cardiovasc Med J 2012;6:8897.
[85] Phung N, Pera N, Farrell G, Leclercq I, Hou JY, George J. Pro-oxidant-mediated
hepatic fibrosis and effects of antioxidant intervention in murine dietary
steatohepatitis. Int J Mol Med 2009;24:17180.
[86] Alvarez JA, Ashraf A. Role of vitamin d in insulin secretion and insulin sensitivity
for glucose homeostasis. Int J Endocrinol 2010:351385.
[87] Zuniga S, Firrincieli D, Housset C, Chignard N. Vitamin D and the vitamin D
receptor in liver pathophysiology. Clin Res Hepatol Gastroenterol 2011;35:
295302.
[88] Smith HJ, Meremikwu M. Iron chelating agents for treating malaria. Cochrane
Database Syst Rev 2000:CD001474.
[89] Smotkin-Tangorra M, Purushothaman R, Gupta A, Nejati G, Anhalt H, Ten S.
Prevalence of vitamin D insufficiency in obese children and adolescents. J Pediatr
Endocrinol Metab 2007;20:81723.
[90] Bradlee ML, Singer MR, Qureshi MM, Moore LL. Food group intake and central
obesity among children and adolescents in the Third National Health and
Nutrition Examination Survey (NHANES III). Public Health Nutr 2010;13:
797805.
[91] Kwok RM, Torres DM, Harrison SA. Vitamin D and nonalcoholic fatty liver disease
(NAFLD): is it more than just an association? Hepatology 2013;58:116674.
[92] Kasapoglu B, Turkay C, Yalcin KS, Carlioglu A, Sozen M, Koktener A. Low vitamin
D levels are associated with increased risk for fatty liver disease among non-
obese adults. Clin Med 2013;13:5769.
[93] Targher G, Bertolini L, Scala L, Cigolini M, Zenari L, Falezza G, et al. Associations
between serum 25-hydroxyvitamin D3 concentrations and liver histology in
patients with non-alcoholic fatty liver disease. Nutr Metab Cardiovasc Dis 2007;
17:51724.
[94] Kitson MT, Roberts SK. D-livering the message: the importance of vitamin D
status in chronic liver disease. J Hepatol 2012;57:897909.
[95] Barchetta I, Carotti S, Labbadia G, Gentilucci UV, Muda AO, Angelico F, et al. Liver
vitamin D receptor, CYP2R1, and CYP27A1 expression: relationship with liver
histology and vitamin D3 levels in patients with nonalcoholic steatohepatitis or
hepatitis C virus. Hepatology 2012;56:21807.
[96] Foroughi M, Maghsoudi Z, Ghiasvand R, Iraj B, Askari G. Effect of Vitamin D
Supplementation on C-reactive Protein in Patients with Nonalcoholic Fatty Liver.
Int J Prev Med 2014;5:96975.
[97] Sharifi N, Amani R, Hajiani E, Cheraghian B. Does vitamin D improve liver
enzymes, oxidative stress, and inflammatory biomarkers in adults with non-
alcoholic fatty liver disease? A randomized clinical trial. Endocrine 2014;47:
7080.
[98] Nishida S, Ozeki J, Makishima M. Modulation of bile acid metabolism by 1alpha-
hydroxyvitamin D3 administration in mice. Drug Metab Dispos 2009;37:
203744.
[99] Theodoropoulos C, Demers C, Petit JL, Gascon-Barre M. High sensitivity of rat
hepatic vitamin D3-25 hydroxylase CYP27A to 1,25-dihydroxyvitamin D3
administration. Am J Physiol Endocrinol Metab 2003;284:E13847.
[100] Sakuma T, Miyamoto T, Jiang W, Kakizawa T, Nishio SI, Suzuki S, et al. Inhibition
of peroxisome proliferator-activated receptor alpha signaling by vitamin D
receptor. Biochem Biophys Res Commun 2003;312:5139.
[101] Chen Z, Gao C, Hua Y, Keep RF, Muraszko K, Xi G. Role of iron in brain injury after
intraventricular hemorrhage. Stroke 2011;42:46570.
[102] Ding N, Yu RT, Subramaniam N, Sherman MH, Wilson C, Rao R, et al. A vitamin D
receptor/SMAD genomic circuit gates hepatic fibrotic response. Cell 2013;153:
60113.
[103] Kong M, Zhu L, Bai L, Zhang X, Chen Y, Liu S, et al. Vitamin D deficiency
promotes nonalcoholic steatohepatitis through impaired enterohepatic
circulation in animal model. Am J Physiol Gastrointest Liver Physiol 2014;
307:G88393.
[104] Roth CL, Elfers CT, Figlewicz DP, Melhorn SJ, Morton GJ, Hoofnagle A, et al.
Vitamin D deficiency in obese rats exacerbates nonalcoholic fatty liver disease
and increases hepatic resistin and Toll-like receptor activation. Hepatology
2012;55:110311.
[105] Yin Y, Yu Z, Xia M, Luo X, Lu X, Ling W. Vitamin D attenuates high fat diet-
induced hepatic steatosis in rats by modulating lipid metabolism. Eur J Clin
Invest 2012;42:118996.
[106] Del Rio D, Rodriguez-Mateos A, Spencer JP, Tognolini M, Borges G, Crozier A.
Dietary (poly)phenolics in human health: structures, bioavailability, and
evidence of protective effects against chronic diseases. Antioxid Redox Signal
2013;18:181892.
[107] Tsao R. Chemistry and biochemistry of dietary polyphenols. Nutrients 2010;2:
123146.
[108] Guo H, Zhong R, Liu Y, Jiang X, Tang X, Li Z, et al. Effects of bayberry juice on
inflammatory and apoptotic markers in young adults with features of non-
alcoholic fatty liver disease. Nutrition 2014;30:198203.
[109] Chang HC, Peng CH, Yeh DM, Kao ES, Wang CJ. Hibiscus sabdariffa extract inhibits
obesity and fat accumulation, and improves liver steatosis in humans. Food
Funct 2014;5:7349.
[110] Van De Wier B, Koek GH, Bast A, Haenen GR. The Potential of Flavonoids in the
Treatment of Non-alcoholic Fatty Liver Disease. Crit Rev Food Sci Nutr 2015;0.
[111] Nakagawa S, Kojima Y, Sekino K, Yamato S. Effect of polyphenols on 3-hydroxy-
3-methylglutaryl-coenzyme A lyase activity in human hepatoma HepG2 cell
extracts. Biol Pharm Bull 2013;36:19026.
[112] Liu Y, Wang D, Zhang D, Lv Y, Wei Y, Wu W, et al. Inhibitory effect of blueberry
polyphenolic compounds on oleic acid-induced hepatic steatosis in vitro. J Agric
Food Chem 2011;59:1225463.
[113] Vidyashankar S, Sandeep Varma R, Patki PS. Quercetin ameliorate insulin
resistance and up-regulates cellular antioxidants during oleic acid induced
hepatic steatosis in HepG2 cells. Toxicol In Vitro 2013;27:94553.
[114] Priore P, Caruso D, Siculella L, Gnoni GV. Rapid down-regulation of hepatic lipid
metabolism by phenolic fraction from extra virgin olive oil. Eur J Nutr 2015.
[115] Liao CC, Ou TT, Huang HP, Wang CJ. The inhibition of oleic acid induced hepatic
lipogenesis and the promotion of lipolysis by caffeic acid via up-regulation of
AMP-activated kinase. J Sci Food Agric 2014;94:115462.
[116] Tsuruta Y, Nagao K, Kai S, Tsuge K, Yoshimura T, Koganemaru K, et al.
Polyphenolic extract of lotus root (edible rhizome of Nelumbo nucifera)
alleviates hepatic steatosis in obese diabetic db/db mice. Lipids Health Dis
2011;10:202.
[117] Park HJ, Jung UJ, Lee MK, Cho SJ, Jung HK, Hong JH, et al. Modulation of lipid
metabolism by polyphenol-rich grape skin extract improves liver steatosis and
adiposity in high fat fed mice. Mol Nutr Food Res 2013;57:3604.
[118] Cheng DM, Pogrebnyak N, Kuhn P, Poulev A, Waterman C, Rojas-Silva P, et al.
Polyphenol-rich Rutgers Scarlet Lettuce improves glucose metabolism and liver
lipid accumulation in diet-induced obese C57BL/6 mice. Nutrition 2014;30:
S528.
[119] Cui Y, Wang X, Xue J, Liu J, Xie M. Chrysanthemum morifolium extract attenuates
high-fat milk-induced fatty liver through peroxisome proliferator-activated
receptor alpha-mediated mechanism in mice. Nutr Res 2014;34:26875.
[120] Fang J. Bioavailability of anthocyanins. Drug Metab Rev 2014;46:50820.
[121] Valenti L, Riso P, Mazzocchi A, Porrini M, Fargion S, Agostoni C. Dietary
anthocyanins as nutritional therapy for nonalcoholic fatty liver disease. Oxid
Med Cell Longev 2013:145421.
[122] Suda I, Ishikawa F, Hatakeyama M, Miyawaki M, Kudo T, Hirano K, et al. Intake of
purple sweet potato beverage affects on serum hepatic biomarker levels of
healthy adult men with borderline hepatitis. Eur J Clin Nutr 2008;62:607.
[123] Zhang PW, Chen FX, Li D, Ling WH, Guo HH. A CONSORT-Compliant,
Randomized, Double-Blind, Placebo-Controlled Pilot Trial of Purified Anthocy-
anin in Patients With Nonalcoholic Fatty Liver Disease. Medicine (Baltimore)
2015;94:e758.
[124] Chang JJ, Hsu MJ, Huang HP, Chung DJ, Chang YC, Wang CJ. Mulberry
anthocyanins inhibit oleic acid induced lipid accumulation by reduction of
lipogenesis and promotion of hepatic lipid clearance. J Agric Food Chem 2013;
61:606976.
[125] Jia Y, Kim JY, Jun HJ, Kim SJ, Lee JH, Hoang MH, et al. Cyanidin is an agonistic
ligand for peroxisome proliferator-activated receptor-alpha reducing hepatic
lipid. Biochim Biophys Acta 2013;1831:698708.
[126] Jiang X, Tang X, Zhang P, Liu G, Guo H. Cyanidin-3-O-beta-glucoside protects
primary mouse hepatocytes against high glucose-induced apoptosis by
modulating mitochondrial dysfunction and the PI3K/Akt pathway. Biochem
Pharmacol 2014;90:13544.
[127] Hwang YP, Choi JH, Han EH, Kim HG, Wee JH, Jung KO, et al. Purple sweet potato
anthocyanins attenuate hepatic lipid accumulation through activating adenosine
monophosphate-activated protein kinase in human HepG2 cells and obese mice.
Nutr Res 2011;31:896906.
[128] Salamone F, Li Volti G, Titta L, Puzzo L, Barbagallo I, La Delia F, et al. Moro orange
juice prevents fatty liver in mice. World J Gastroenterol 2012;18:38628.
[129] Tang X, Shen T, Jiang X, Xia M, Sun X, Guo H, et al. Purified anthocyanins from
bilberry and black currant attenuate hepatic mitochondrial dysfunction and
steatohepatitis in mice with methionine and choline deficiency. J Agric Food
Chem 2015;63:5525561.
[130] Cheung CW, Gibbons N, Johnson DW, Nicol DL. Silibinina promising new
treatment for cancer. Anticancer Agents Med Chem 2010;10:18695.
[131] Huseini HF, Larijani B, Heshmat R, Fakhrzadeh H, Radjabipour B, Toliat T, et al.
The efficacy of Silybum marianum (L.) Gaertn. (silymarin) in the treatment of
type II diabetes: a randomized, double-blind, placebo-controlled, clinical trial.
Phytother Res 2006;20:10369.
[132] Al-Anati L, Essid E, Reinehr R, Petzinger E. Silibinin protects OTA-mediated TNF-
alpha release from perfused rat livers and isolated rat Kupffer cells. Mol Nutr
Food Res 2009;53:4606.
[133] Jayaraj R, Deb U, Bhaskar AS, Prasad GB, Rao PV. Hepatoprotective efficacy of
certain flavonoids against microcystin induced toxicity in mice. Environ Toxicol
2007;22:4729.
[134] Esser-Nobis K, Romero-Brey I, Ganten TM, Gouttenoire J, Harak C, Klein R, et al.
Analysis of hepatitis C virus resistance to silibinin in vitro and in vivo points to a
novel mechanism involving nonstructural protein 4B. Hepatology 2013;57:
95363.
 P. Dongiovanni et al. / Journal of Nutritional Biochemistry 29 (2016) 111
[135] Milosevic N, Milanovic M, Abenavoli L, Milic N. Phytotherapy and NAFLDfrom
goals and challenges to clinical practice. Rev Recent Clin Trials 2014;9:195203.
[136] Flora K, Hahn M, Rosen H, Benner K. Milk thistle (Silybum marianum) for the
therapy of liver disease. Am J Gastroenterol 1998;93:13943.
[137] Loguercio C, Andreone P, Brisc C, Brisc MC, Bugianesi E, Chiaramonte M, et al.
Silybin combined with phosphatidylcholine and vitamin E in patients with
nonalcoholic fatty liver disease: a randomized controlled trial. Free Radic Biol
Med 2012;52:165865.
[138] Solhi H, Ghahremani R, Kazemifar AM, Hoseini Yazdi Z. Silymarin in treatment of
non-alcoholic steatohepatitis: A randomized clinical trial. Caspian J Intern Med
2014;5:912.
[139] Zhang Y, Hai J, Cao M, Pei S, Wang J, Zhang Q. Silibinin ameliorates steatosis and
insulin resistance during non-alcoholic fatty liver disease development partly
through targeting IRS-1/PI3K/Akt pathway. Int Immunopharmacol 2013;17:71420.
[140] Trappoliere M, Caligiuri A, Schmid M, Bertolani C, Failli P, Vizzutti F, et al. Silybin,
a component of sylimarin, exerts anti-inflammatory and anti-fibrogenic effects
on human hepatic stellate cells. J Hepatol 2009;50:110211.
[141] Salamone F, Galvano F, Marino Gammazza A, Paternostro C, Tibullo D, Bucchieri
F, et al. Silibinin improves hepatic and myocardial injury in mice with
nonalcoholic steatohepatitis. Dig Liver Dis 2012;44:33442.
[142] Salamone F, Galvano F, Cappello F, Mangiameli A, Barbagallo I, Li Volti G. Silibinin
modulates lipid homeostasis and inhibits nuclear factor kappa B activation in
experimental nonalcoholic steatohepatitis. Transl Res 2012;159:47786.
[143] Yao J, Zhi M, Gao X, Hu P, Li C, Yang X. Effect and the probable mechanisms of
silibinin in regulating insulin resistance in the liver of rats with non-alcoholic
fatty liver. Braz J Med Biol Res 2013;46:2707.
[144] Faghihzadeh F, Adibi P, Rafiei R, Hekmatdoost A. Resveratrol supplementation
improves inflammatory biomarkers in patients with nonalcoholic fatty liver
disease. Nutr Res 2014;34:83743.
[145] Chen S, Zhao X, Ran L, Wan J, Wang X, Qin Y, et al. Resveratrol improves insulin
resistance, glucose and lipid metabolism in patients with non-alcoholic fatty
liver disease: a randomized controlled trial. Dig Liver Dis 2015;47:22632.
[146] Chachay VS, Macdonald GA, Martin JH, Whitehead JP, O'Moore-Sullivan TM, Lee
P, et al. Resveratrol does not benefit patients with nonalcoholic fatty liver
disease. Clin Gastroenterol Hepatol 2014;12 2092.e12103.e6.
[147] Gnoni GV, Paglialonga G. Resveratrol inhibits fatty acid and triacylglycerol
synthesis in rat hepatocytes. Eur J Clin Invest 2009;39:2118.
[148] Shang J, Chen LL, Xiao FX, Sun H, Ding HC, Xiao H. Resveratrol improves non-
alcoholic fatty liver disease by activating AMP-activated protein kinase. Acta
Pharmacol Sin 2008;29:698706.
[149] Baur JA, Pearson KJ, Price NL, Jamieson HA, Lerin C, Kalra A, et al. Resveratrol
improves health and survival of mice on a high-calorie diet. Nature 2006;444:
33742.
[150] Deng XQ, Chen LL, Li NX. The expression of SIRT1 in nonalcoholic fatty liver
disease induced by high-fat diet in rats. Liver Int 2007;27:70815.
[151] Pfluger PT, Herranz D, Velasco-Miguel S, Serrano M, Tschop MH. Sirt1 protects
against high-fat diet-induced metabolic damage. Proc Natl Acad Sci U S A 2008;
105:97938.
[152] Frescas D, Valenti L, Accili D. Nuclear trapping of the forkhead transcription
factor FoxO1 via Sirt-dependent deacetylation promotes expression of glucoge-
netic genes. J Biol Chem 2005;280:2058995.
[153] Zang M, Xu S, Maitland-Toolan KA, Zuccollo A, Hou X, Jiang B, et al. Polyphenols
stimulate AMP-activated protein kinase, lower lipids, and inhibit accelerated
11
atherosclerosis in diabetic LDL receptor-deficient mice. Diabetes 2006;55:
218091.
[154] Kamei Y, Miura S, Suganami T, Akaike F, Kanai S, Sugita S, et al. Regulation of
SREBP1c gene expression in skeletal muscle: role of retinoid X receptor/liver X
receptor and forkhead-O1 transcription factor. Endocrinology 2008;149:
2293305.
[155] Aigner E, Theurl I, Haufe H, Seifert M, Hohla F, Scharinger L, et al. Copper
availability contributes to iron perturbations in human nonalcoholic fatty liver
disease. Gastroenterology 2008;135:6808.
[156] Fields M, Holbrook J, Scholfield D, Smith Jr JC, Reiser S. Effect of fructose or starch
on copper-67 absorption and excretion by the rat. J Nutr 1986;116:62532.
[157] Aigner E, Strasser M, Haufe H, Sonnweber T, Hohla F, Stadlmayr A, et al. A role for
low hepatic copper concentrations in nonalcoholic fatty liver disease. Am J
Gastroenterol 2010;105:197885.
[158] Song M, Schuschke DA, Zhou Z, Chen T, Pierce Jr WM, Wang R, et al. High fructose
feeding induces copper deficiency in Sprague-Dawley rats: a novel mechanism
for obesity related fatty liver. J Hepatol 2012;56:43340.
[159] Prohaska JR, Geissler J, Brokate B, Broderius M. Copper, zinc-superoxide
dismutase protein but not mRNA is lower in copper-deficient mice and mice
lacking the copper chaperone for superoxide dismutase. Exp Biol Med
(Maywood) 2003;228:95966.
[160] Machado MV, Ravasco P, Jesus L, Marques-Vidal P, Oliveira CR, Proenca T, et al.
Blood oxidative stress markers in non-alcoholic steatohepatitis and how it
correlates with diet. Scand J Gastroenterol 2008;43:95102.
[161] Kang BP, Bansal MP, Mehta U. Hyperlipidemia and type I 5'-monodeiodinase
activity: regulation by selenium supplementation in rabbits. Biol Trace Elem Res
2000;77:2319.
[162] Clarke C, Baghdadi H, Howie AF, Mason JI, Walker SW, Beckett GJ. Selenium
supplementation attenuates procollagen-1 and interleukin-8 production in fat-
loaded human C3A hepatoblastoma cells treated with TGFbeta1. Biochim
Biophys Acta 2010;1800:6118.
[163] Kaur HD, Bansal MP. Studies on HDL associated enzymes under experimental
hypercholesterolemia: possible modulation on selenium supplementation.
Lipids Health Dis 2009;8:55.
[164] Dongiovanni P, Fracanzani AL, Fargion S, Valenti L. Iron in fatty liver and in the
metabolic syndrome: a promising therapeutic target. J Hepatol 2011;55:92032.
[165] Valenti L, Fracanzani AL, Dongiovanni P, Bugianesi E, Marchesini G, Manzini P,
et al. Iron depletion by phlebotomy improves insulin resistance in patients with
nonalcoholic fatty liver disease and hyperferritinemia: evidence from a case-
control study. Am J Gastroenterol 2007;102:12518.
[166] Valenti L, Fracanzani AL, Bugianesi E, Dongiovanni P, Galmozzi E, Vanni E,
et al. HFE genotype, parenchymal iron accumulation, and liver fibrosis in
patients with nonalcoholic fatty liver disease. Gastroenterology 2010;138:
90512.
[167] Dongiovanni P, Valenti L, Ludovica Fracanzani A, Gatti S, Cairo G, Fargion S. Iron
depletion by deferoxamine up-regulates glucose uptake and insulin signaling in
hepatoma cells and in rat liver. Am J Pathol 2008;172:73847.
[168] Dongiovanni P, Ruscica M, Rametta R, Recalcati S, Steffani L, Gatti S, et al. Dietary
iron overload induces visceral adipose tissue insulin resistance. Am J Pathol
2013;182:225463.
[169] Dongiovanni P, Lanti C, Gatti S, Rametta R, Recalcati S, Maggioni M, et al. High fat
diet subverts hepatocellular iron uptake determining dysmetabolic iron
overload. PLoS One 2015;10:e0116855.