Alcohol 34 (2004) 9–19

Alcoholic fatty liver: its pathogenesis and mechanism of

progression to inflammation and fibrosis
Charles S. Lieber*
Bronx Veterans Affairs Medical Center, Bronx, NY 10468, USA
Mount Sinai School of Medicine, New York, NY 10029, USA
Received 30 March 2004; received in revised form 16 July 2004; accepted 20 July 2004

Abstract
Liver disease in the alcoholic is due not only to malnutrition but also to ethanol’s hepatotoxicity linked to its metabolism by means of
the alcohol dehydrogenase and cytochrome P450 2E1 (CYP2E1) pathways and the resulting production of toxic acetaldehyde. In addition,
alcohol dehydrogenase–mediated ethanol metabolism generates the reduced form of nicotinamide adenine dinucleotide (NADH), which
promotes steatosis by stimulating the synthesis of fatty acids and opposing their oxidation. Steatosis is also promoted by excess dietary
lipids and can be attenuated by their replacement with medium-chain triglycerides. Through reduction of pyruvate, elevated NADH also
increases lactate, which stimulates collagen synthesis in myofibroblasts. Furthermore, CYP2E1 activity is inducible by its substrates, not
only ethanol but also fatty acids. Their excess and metabolism by means of this pathway generate release of free radicals, which cause oxidative
stress, with peroxidation of lipids and membrane damage, including altered enzyme activities. Products of lipid peroxidation such as
4-hydroxynonenal stimulate collagen generation and fibrosis, which are further increased through diminished feedback inhibition of collagen
synthesis because acetaldehyde forms adducts with the carboxyl-terminal propeptide of procollagen in hepatic stellate cells. Acetaldehyde
is also toxic to the mitochondria, and it aggravates their oxidative stress by binding to reduced glutathione and promoting its
leakage. Oxidative stress and associated cellular injury promote inflammation, which is aggravated by increased production of the
proinflammatory cytokine tumor necrosis factor-alpha in the Kupffer cells. These are activated by induction of their CYP2E1 as well as
by endotoxin. The endotoxin-stimulated tumor necrosis factor-alpha release is decreased by dilinoleoylphosphatidylcholine, the active
phosphatidylcholine (PC) species of polyenylphosphatidylcholine (PPC). Moreover, defense mechanisms provided by peroxisome proliferator-
activated receptor alpha and omega fatty acid oxidation are readily overwhelmed, particularly in female rats and also in women who
have low hepatic induction of fatty acid–binding protein (L-FABPc). Accordingly, the intracellular concentration of free fatty acids may
become high enough to injure membranes, thereby contributing to necrosis, inflammation, and progression to fibrosis and cirrhosis. Even-
tually, hepatic S-adenosylmethionine and PCs become depleted in the alcoholic, with impairment of their multiple cellular functions, which
can be restored by PC replenishment. Thus, prevention and therapy opposing the development of steatosis and its progression to more
severe injury can be achieved by a multifactorial approach: control of alcohol consumption, avoidance of obesity and of excess dietary
long-chain fatty acids, or their replacement with medium-chain fatty acids, and replenishment of S-adenosylmethionine and PCs by using
PPC. Progress in the understanding of the pathogenesis of alcoholic fatty liver and its progression to inflammation and fibrosis has resulted
in prospects for their better prevention and treatment. 쑖 2005 Elsevier Inc. All rights reserved.
Keywords: Alcoholic steatosis; Alcoholic steatohepatitis; Cytochrome P450 2E1; Obesity; Dilinoleoylphosphatidylcholine; S-adenosylmethionine; Oxidative
stress; TNF-α; Acetaldehyde

1. Introduction dominant metabolism in the liver associated with oxidation–

reduction (redox) changes and oxidative stress. Redox
Progress in the understanding of the pathogenesis of alco-
changes are mediated by alcohol dehydrogenase (ADH), and
holic liver disease was achieved when it was discovered
oxidative stress is generated mainly by the activity of the
that alcohol affects the liver through not only nutritional
microsomal ethanol oxidizing system (MEOS) and its key
disturbances but also its direct toxicity because of its pre-
enzyme cytochrome P450 2E1 (CYP2E1), which releases
free radicals.
* Corresponding author. Bronx Veterans Affairs Medical Center, 130 The clinical course and ultimate outcome of alcoholic
West Kingsbridge Road (151-2), Bronx, NY 10468, USA. Tel.: ⫹1-718-
741-4244; fax: ⫹1-718-733-6257.
liver disease are dismal. In a prospective survey of 280
E-mail address: liebercs@aol.com (C.S. Lieber). patients with alcoholic liver injury, Chedid et al. (1991)
Editor: T.R. Jerrells found that, within 48 months of follow-up, 30% of those
0741-8329/05/$ – see front matter 쑖 2005 Elsevier Inc. All rights reserved.
doi: 10.1016/j.alcohol.2004.07.008
10 C.S. Lieber / Alcohol 34 (2004) 9–19

with a fatty liver, more than half of those with cirrhosis, and
two-thirds of those with cirrhosis plus alcoholic hepatitis
died. This outcome is more severe than that of many
cancers, yet it is attracting much less concern, among both
the public and the medical profession. This may be due, at
least in part, to the prevailing and pervasive perception
that not much can be done about this major public health
issue. Pathogenic concepts of alcoholic liver disease, how-
ever, are evolving, and elucidation of the biochemical effects
of ethanol allows for a more optimistic outlook in terms of
diagnosis and treatment.
2. Pathogenesis of alcoholic and non-alcoholic fatty liver
2.1. Alcohol and nutrition
Unlike other drugs, ethanol is a substantial source of
energy, with 7.1 kcal (29.7 kJ) per gram, a value that exceeds
the energy content of carbohydrates or proteins. On average,
ethanol accounts for half an alcoholic’s caloric intake. It
therefore displaces normal nutrients, causing malnutrition
(Fig. 1), including deficiencies of folate, thiamine, and other
vitamins. Secondary malnutrition also occurs through malab-
sorption due to gastrointestinal complications, such as pan-
creatic insufficiency and impaired hepatic metabolism of
nutrients (Fig. 1). In addition, alcohol promotes the degrada-
tion of nutrients, as exemplified by its effects on vitamin A
(Lieber, 1992).
Because, experimentally, nutritional deficiencies cause
liver damage, it was postulated that this is also the mecha-
nism whereby alcoholic liver disease develops. As a result,
the prevailing therapy was nutritional with enriched diets.
This, however, did not prevent the development of the first
stage of alcoholic liver disease, namely alcoholic fatty liver.
Therefore, the question was raised whether alcohol may
have some direct toxic effect on the liver independent of
malnutrition (Fig. 1). This hypothesis was initially rejected
because, as stated by Best et al. (1949, p. 1005), “there is
no more evidence of a specific toxic effect of pure ethyl
alcohol upon liver cells than there is for one due to sugar”.
This was based on experiments in rats given adequate diets
with alcohol in drinking water. They did not develop any
liver damage unless the diet was made deficient. However,
rats do not drink enough alcohol when given in drinking
water for significant blood concentrations to develop. Their
aversion was overcome by incorporating the added ethanol
into totally liquid, nutritionally adequate diets containing
35% of calories as alcohol. Under those conditions, an obvi-
ous fatty liver developed compared with the findings in
control animals pair-fed with an isocaloric diet in which
alcohol was replaced by carbohydrates (Lieber et al., 1963).
Similar results were obtained in human volunteers (Lieber
et al., 1963, 1965). Results of these studies clearly estab-
lished that, even in the presence of an adequate diet, alcohol
can produce a fatty liver and at blood concentrations that
do not necessarily produce intoxication. Increases in die-
tary long-chain triglycerides enhanced the alcohol-induced
steatosis, and this effect was attenuated by medium-chain
triglycerides (Lieber et al., 1967). Decreasing of dietary fat
reduced steatosis; however, below a concentration of 10%
of calories as dietary fat, there was a reversal of this effect,
probably owing to the high carbohydrate content of the
corresponding diet (Lieber & DeCarli, 1970). These effects
of alcohol were shown to be due to its metabolism in the liver.

Fig. 1. Interaction of the direct toxicity of ethanol with malnutrition resulting from primary or secondary deficiencies. Secondary malnutrition may be
caused by either (1) maldigestion and malabsorption or (2) impaired utilization due to decreased activation or increased degradation of nutrients. Both
direct toxicity of ethanol and malnutrition (whether primary or secondary) may affect function and structure of the liver and gastrointestinal tract.
 C.S. Lieber / Alcohol 34 (2004) 9–19 11

2.2. Toxic effects of hepatic alcohol oxidation
Oxidation of ethanol through the ADH pathway produces
acetaldehyde, which is converted to acetate. Both reac-
tions reduce nicotinamide adenine dinucleotide (NAD) to
its reduced form (NADH) (Fig. 2). Excess NADH causes a
number of metabolic disorders (Lieber, 1992), including
inhibition of the Krebs cycle and of its fatty acid oxidation.
The inhibition of fatty acid oxidation favors steatosis and
hyperlipidemia. The effects of ethanol were reproduced in
vitro by an alternative NADH-generating system (sorbitol–
fructose) and were blocked by an H⫹ acceptor (methylene
blue) (Lieber et al., 1959; Lieber & Schmid, 1961). The
preventive effect of methylene blue against ethanol-induced
fat accumulation was confirmed subsequently (Galli et al.,
1999). This rat model of alcoholic liver disease consistently
reproduced a fatty liver, but it did not progress to inflam-
mation, fibrosis, and cirrhosis. This is probably due to the
fact that, even with the liquid diet, the alcohol intake is
limited to 35% of total calories, whereas alcoholics in whom
cirrhosis develops usually consume at least 50% of their
calories as alcohol. Such a high intake of alcohol was subse-
quently achieved in non-human primates, namely baboons
(Lieber & DeCarli, 1974a). In that animal model, it was also
shown that, in addition to the metabolic abnormalities caused
by the ADH activity, a new pathway of ethanol metabolism,
namely MEOS, plays a key role in the progression of the
disease.
2.3. Microsomal ethanol oxidizing system
As reviewed elsewhere (Lieber, 1997, 1999), the discov-
ery of the MEOS was prompted by the observation that, after
chronic ethanol consumption, there is an adaptive increase of
ethanol metabolism, which could not be explained on the
basis of ADH, therefore raising the possibility of the exis-
tence of an additional pathway. The first indication of a
possible interaction of ethanol with the endoplasmic reticu-
lum of the hepatocyte (also called microsomal fraction when
obtained by ultracentrifugation) was provided by the obser-
vation that, in rats and human beings, ethanol feeding results
in a proliferation of the smooth endoplasmic reticulum (SER)
(Iseri et al., 1964, 1966; Lane & Lieber, 1966), which was
confirmed by an increase in the amount of enzyme activities
of the hepatic microsomal membranes (Ishii et al., 1973a),
including a rise in microsomal glucose-6-phosphatase activ-
ity after chronic ethanol administration (Ishii et al., 1973b).
This increase in SER resembled that seen after the adminis-
tration of a wide variety of hepatotoxins (Meldolesi, 1967),
barbiturates, and other therapeutic agents (Conney, 1967) and
food additives (Lane & Lieber, 1967). Because many of the
substances that induce a proliferation of the SER are metab-
olized, at least in part, by the cytochrome P450 enzyme
system that is located in the SER, the possibility that ethanol
may also be metabolized by a similar process was raised.
Indeed, such a system, named MEOS, was demonstrated in
liver microsomes in vitro and found to be inducible by

Fig. 2. Ethanol oxidation by (A) alcohol dehydrogenase (ADH) and nicotinamide adenine dinucleotide (NAD⫹); (B) the hepatic microsomal ethanol
oxidizing system [(MEOS), which involves cytochrome P450 2E1 and the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH)];
(C) a combination of NADPH oxidase and catalase; and (D) xanthine oxidase and catalase.
12 C.S. Lieber / Alcohol 34 (2004) 9–19
chronic ethanol feeding in vivo (Lieber & DeCarli, 1968,
1970). The proposal that this new pathway plays a significant
role in ethanol metabolism initiated a decade of lively debate:
Some invoked that ADH was contaminating the liver micro-
somes (Isselbacher & Carter, 1970), whereas others asserted
that this microsomal ethanol oxidation was due to a hydrogen
peroxide–dependent reaction promoted by contaminating
catalase (Ziegler, 1972). Indeed, it had been postulated that
the combination of hydrogen peroxide generation from the
reduced form of nicotinamide adenine dinucleotide phos-
phate (NADPH) oxidase and of catalase could account for
microsomal ethanol oxidation (Roach et al., 1969; Tephly
et al., 1969) (Fig. 1), especially because a hydrogen perox-
ide–generating system (glucose–glucose oxidase) can be
substituted for NADPH. Actually, the latter is not unex-
pected, because not only do microsomes contain catalase,
but commercial glucose oxidase (used to generate NADPH)
is contaminated with catalase. It also had been reported that
microsomes obtained from acatalasemic mice fail to oxidize
ethanol (Vatsis & Schulman, 1973), but this claim was
subsequently retracted (Vatsis & Schulman, 1974). Hepatic
microsomes of acatalasemic mice subjected to heat inactiva-
tion displayed decreased catalase activity, but NADPH-
dependent MEOS remained active and unaffected (Lieber &
DeCarli, 1974b; Teschke et al., 1975). Eventually, MEOS
was solubilized and separated from ADH and catalase activi-
ties by diethylaminoethyl cellulose column chromatography
(Teschke et al., 1972, 1975).
It was established that the key enzyme of the MEOS
is the ethanol-inducible CYP2E1, which was found to be
increased fourfold to 10-fold in liver biopsy samples obtained
from recently drinking subjects (Tsutsumi et al., 1989),
with a corresponding rise in mRNA (Takahashi et al.,
1993). This induction contributes to the metabolic tolerance
to ethanol that develops in the alcoholic (in addition to its
CNS tolerance).
Finally, successful reconstitution of the system was ac-
complished with either partially purified or highly purified
microsomal P-450 obtained from alcohol-treated (Ohnishi &
Lieber, 1977) or phenobarbital-treated (Miwa et al., 1978)
rats. In a new nomenclature system, it was proposed that
the ethanol-inducible form be designated as CYP2El (Nelson
et al., 1993). On the basis of findings of these and various
other studies, and regardless of the original claim to the
contrary (Thurman et al., 1972), it was finally agreed by
the principal contenders involved that catalase cannot ac-
count for the microsomal ethanol oxidation (Teschke et al.,
1977; Thurman & Brentzel, 1977), and that the MEOS is
distinct from ADH and catalase and dependent on cyto-
chromes P450, as reviewed elsewhere (Lieber, 1997, 1999).
Despite the discovery of CYP2E1 and its prevailing role
in microsomal ethanol oxidation, the term MEOS was main-
tained because cytochromes P450 other than CYP2E1 (such
as CYP1A2 and CYP3A4) (Salmela et al., 1998) may also
contribute to ethanol metabolism in the microsomes. Thus,
the term MEOS characterizes total microsomal ethanol
oxidation, not only that catalyzed by CYP2E1.
The rat CYP2E1 gene was isolated, characterized, and
localized to chromosome 7 (Umeno et al., 1988b), and the
human gene was isolated, characterized, and localized to
chromosome 10 (Umeno et al., 1988a). In rabbits, two genes
may be involved (Khani et al., 1988).
Enhanced concentrations of both hepatic CYP2E1 pro-
tein and mRNA were found in actively drinking patients
(Takahashi et al., 1993). As revealed from results of the
studies cited above and reviewed elsewhere (Lieber, 1997,
1999), the most common cause for CYP2E1 induction is
early alcoholic liver injury, such as alcoholic steatosis and
alcoholic steatohepatitis. Cytochrome P450 2E1, however,
is also induced in nonalcoholic steatohepatitis, which can be
understood on the basis of the physiologic role of CYP2E1.
Indeed, there seems to be a dual role of CYP2E1 (Fig. 3),
namely one of detoxification and one of nutritional support.
That CYP2E1 contributes to the defense mechanisms of the
body against the penetration of toxic xenobiotics is suggested
by its location and inducibility at port of entries into the body,
and by its broad substrate specificity.
In terms of its nutritional role, CYP2E1 is inducible by
fasting in the rat (Koop & Tierney, 1990). The increase
may be due, at least in part, to ketones. Indeed, in rats
(Casazza et al., 1984), rabbits (Koop & Casazza, 1985), and
human beings (Reichard et al., 1986), acetone seems to
be actively used, being metabolized by a microsomal acetone
monooxygenase identified as CYP2E1 (Casazza & Veech,
1985; Johansson et al., 1986). Acetone is both an inducer
and a substrate of CYP2E1 (Koop, 1992; Yang et al., 1990).
Cytochrome P450 2E1 was also found to be induced in
obese rats (Raucy et al., 1991). The role of CYP2E1 in fatty
acid metabolism supports the concept of a nutritional role
for CYP2E1. Indeed, in addition to its ethanol-oxidizing
Fig. 3. Physiologic and toxic roles of cytochrome P450 2E1 (CYP2E1),
the main cytochrome P450 of the microsomal ethanol oxidizing system
(MEOS). Many endogenous and xenobiotic compounds, including ethanol,
ketones, and fatty acids, are substrates for CYP2E1 and induce its activity
through various mechanisms (see text), resulting in an array of beneficial as
well as harmful effects. NADPH ⫽ Reduced form of nicotinamide adenine
dinucleotide phosphate. Reprinted from C. S. Lieber, Microsomal ethanol-
oxidizing system (MEOS): the first 30 years (1968–1998)—a review,
Alcoholism: Clinical and Experimental Research 23(6), 991–1007, 1999,
with permission of Lippincott, Williams & Wilkins.
 C.S. Lieber / Alcohol 34 (2004) 9–19
activity, CYP2E1 catalyzes fatty acid ω-1 and ω-2 hydroxyl-
ations (Adas et al., 1998; Amet et al., 1994; Laethem
et al., 1993). Ethanol feeding also results in an increased
activity of CYP4A1 (Ma et al., 1993). The CYP4A subfam-
ily catalyzes ω-hydroxylation at the terminal carbon of fatty
acids. Further oxidation of the ω and ω-1 hydroxyacids
by alcohol and ADH results in the production of dicarboxylic
and oxocarboxylic acids. The oxidation of ω-hydroxyacids
is principally catalyzed by class 3 ADH, which has a low
affinity for ethanol, whereas the ω-l hydroxyacids are likely
to be preferentially oxidized by class 1 ADH, which has a
high affinity for ethanol (Boleda et al., 1993).
It is of interest that dicarboxylic acids (products of the
CYP4A-mediated pathway) play a regulatory role in the
hepatic disposition of nonesterified fatty acids by activating
the peroxisome proliferator-activated receptor alpha, which
increases the transcription of key fatty acid–disposal path-
ways, such as the microsomal CYP4A1, the peroxisomal
acyl-coenzyme A oxidase, and the liver fatty acid–binding
protein in the cytosol (L-FABPc) (Kaikaus et al., 1993).
This, in turn, results in enhanced microsomal omega-oxida-
tion and peroxisomal beta-oxidation of fatty acids and in
a greater stimulatory effect of L-FABPc on microsomal acyl-
glycerol synthesis (Burnett et al., 1979).
The resulting changes exacerbate the sex difference in
the alcohol-induced lipid abnormalities and in the vulnera-
bility to alcoholic liver disease. It was known that sex dif-
ferences in ethanol distribution (Jones & Jones, 1976),
bioavailability (Frezza et al., 1990a), and hepatic metabo-
lism (Mishra et al., 1989), with increased production of acet-
aldehyde (Eriksson et al., 1996), may contribute to women’s
vulnerability to alcohol consumption. In addition, sex differ-
ences in alcohol-induced derangements of hepatic lipid
metabolism, affecting the disposition of potentially toxic
fatty acids, have also been reported in animal experiments
(Shevchuk et al., 1991). Indeed, chronic alcohol administra-
tion increased omega-oxidation more effectively in male than
in female rats (Ma et al., 1993), explaining the potentially
deleterious accumulation of nonesterified fatty acids in the
liver of the alcohol-fed female rats (Shevchuk et al., 1991).
This was also associated with a much smaller ethanol-induced
increase in cytosolic L-FABPc and microsomal esterification
in female than in male rats, whereas the inhibition of mito-
chondrial beta-oxidation was similar in both sexes. Related
findings have been observed in human beings (Ma et al.,
1999) and contribute to the greater vulnerability of women
to the development of alcoholic liver injury.
In any event, the induction of CYP2E1 has been shown
to play a key role in the pathogenesis of alcoholic liver
injury, including alcoholic steatohepatitis, because of the
oxidative stress it generates (Lieber, 1997). In addition,
CYP2E1 is invariably elevated in the liver of patients with
nonalcoholic steatohepatitis (Weltman et al., 1998) because
fatty acids (which increase in obesity) and ketones (which
increase in diabetes) are also substrates for CYP2E1 (vide
13
supra and Fig. 3); their excess up-regulates CYP2E1. Al-
though the pathogenesis of non-alcoholic fatty liver disease
and nonalcoholic steatohepatitis has not yet been fully eluci-
dated, a popular mechanism is the “two hit” theory (James &
Day, 1998), with the first hit being the accumulation, by
several causes (e.g., obesity), of fatty acids in the liver.
The second hit is the peroxidation of these fatty acids because
of the oxidative stress produced by different factors such as
CYP2E1 induction (Angulo & Lindor, 2001).
The damage caused by oxidative stress in both alcoholic
steatohepatitis and nonalcoholic steatohepatitis includes mi-
tochondrial injury, which, in turn, exacerbates the oxidative
stress (Lieber, 1997). That mitochondrial damage is a key
component of alcoholic liver injury is well established
(Lieber, 1992). However, nonalcoholic steatohepatitis is also
associated with mitochondrial structural defects, whereas
non-alcoholic fatty liver disease is not. This mitchondrial
dysfunction contributes to the oxidative stress in nonalco-
holic steatohepatitis (Sanyal et al., 2001).
Like many other useful adaptive systems, when the adap-
tation becomes excessive, adverse consequences prevail.
Cytochrome P450 2E1 leaks oxygen radicals as part of its
operation (Fig. 3), and when they exceed the cellular defense
systems they result in oxidative stress with its pathologic
consequences. This is true when excess alcohol has to be
metabolized, as in alcoholic steatohepatitis, or when
CYP2E1 is confronted by an excess of ketones and fatty
acids associated with diabetes, obesity, or both, resulting in
nonalcoholic steatohepatitis.
3. Progression of fatty liver to inflammation and fibrosis
The oxidative stress caused by CYP2E1 induction and
mitochondrial injury results in lipid peroxidation and mem-
brane damage. In addition, the acetaldehyde produced by
the oxidation of ethanol has toxic effects, inhibiting the
repair of alkylated nucleoproteins (Espina et al., 1988),
decreasing the activity of key enzymes, and markedly reduc-
ing oxygen use in mitochondria damaged by long-term etha-
nol consumption (Baraona et al., 1988; Lieber et al., 1989).
Impaired oxygen use was also confirmed in baboons in vivo
(Lieber et al., 1989). The impaired oxidation capacity of the
mitochondria may, in turn, interfere with the oxidation of
acetaldehyde (Hasumura et al., 1976), leading to a vicious
circle of progressive acetaldehyde accumulation and greater
mitochondrial injury. Moreover, acetaldehyde promotes
cell death by depleting the concentration of reduced glutathi-
one (GSH), inducing lipid peroxidation, and increasing
the toxic effect of free radicals. By binding to the tubulin
of microtubules, acetaldehyde blocks the secretion of
proteins. The increases in protein, lipid, water (Wondergem
& Davis, 1994), and electrolytes cause hepatocytes to
enlarge or “balloon,” a hallmark of alcoholic liver disease.
Acetaldehyde–protein adducts promote collagen production
and may also act as neoantigens, which stimulate an immune
response (Hoerner et al., 1986; Niemela et al., 1987).
14 C.S. Lieber / Alcohol 34 (2004) 9–19
Lipid peroxidation products such as 4-hydroxynonenal
stimulate fibrosis, which is also increased through decreased
feedback inhibition of collagen synthesis because acetalde-
hyde forms adducts with the carboxyl-terminal propeptide of
procollagen (Ma et al., 1997).
Oxidative stress promotes inflammation, which is aggra-
vated by an increase of the proinflammatory cytokine tumor
necrosis factor-alpha (TNF-α) in the Kupffer cells. Kupf-
fer cells are a major source of cytokines. They also harbor
CYP2E1, and its increase after chronic alcohol consumption
(Koivisto et al., 1996) may act as a major stimulator. Indeed,
in both acute and chronic liver diseases, Kupffer cells
become activated to produce cytokines and reactive oxygen
radicals (Batey et al., 1998). Of potential therapeutic interest
is the observation that dilinoleoylphosphatidylcholine
(DLPC) decreases stellate cell activation (Poniachik et al.,
1999) and that it selectively modulates the lipopolysaccha-
ride-induced activation of Kupffer cells by decreasing the
production of the cytotoxic TNF-α, while potentiating the
release of the protective interleukin-1 beta (IL-1β) (Oneta
et al., 1999). This dual action of DLPC on cytokines may
provide a potent mechanism against liver injury. In addition
to the reduction in TNF-α, the enhanced release of IL-1β
could oppose the hepatotoxicity of TNF-α, either directly
or indirectly through an IL-1β–related increase in the toler-
ance to TNF-α–mediated toxicity.
It has also been shown that ethanol induces transforming
growth factor-alpha (TGF-α) production in hepatocytes,
leading to stimulation of collagen synthesis by hepatic stel-
late cells (Kato et al., 2003). These results support the notion
that TGF-α derived from ethanol-exposed hepatocytes may
contribute to the development of hepatic fibrosis in alcoholic
liver disease. This effect was specifically inhibited by anti–
TGF-α antibodies. It is noteworthy that, experimentally,
DLPC also decreased transforming growth factor-beta 1
(TGF-β1)–induced collagen mRNA by inhibiting p38 mito-
gen-activated protein kinase in hepatic stellate cells (Cao
et al., 2002). These are activated through induction of their
CYP2E1, through acetaldehyde, and by endotoxin. That
TNF-α is decreased by DLPC, the active phosphatidylcho-
line (PC) species of polyenylphosphatidylcholine (PPC), was
also shown by Cao et al. (2002).
4. Antifibrotic therapy through correction of altered nu-
trient activation or of the deficiency in selective nutrients
The nutritional approach to liver disease has been im-
proved by recognizing the importance of not only providing
a sufficient intake of nutrients but also correcting the impair-
ment in the activation of key nutrients by alcohol.
4.1. Methionine and S-adenosylmethionine
4.1.1. Pathogenesis of the deficiency and its consequences
Correction of methionine deficiency has been proposed
for the treatment of liver diseases, especially the alcoholic
variety, but excess methionine was shown to have some
adverse effects, as reviewed elsewhere (Lieber, 2000).
Whereas in some patients with alcoholic liver disease circu-
lating methionine concentrations are within normal limits, in
others elevated concentrations were observed. It was reported
that the blood clearance of methionine after an oral load
was slowed, supporting the notion of impaired metabolism.
Indeed, for most of its functions, methionine must be acti-
vated to S-adenosylmethionine (SAMe) and, in cirrhotic
livers, Duce et al. (1988) reported a decrease in the activity of
the enzyme involved, namely SAMe synthetase, also called
methionine adenosyltransferase (Lieber, 2001) (Fig. 1). Fur-
thermore, long-term ethanol consumption in non-human pri-
mates was associated with a significant depletion of hepatic
SAMe (Lieber et al., 1990), which may have a number of
adverse effects because SAMe is the principal methylating
agent in various transmethylation reactions, which are im-
portant for nucleic acid and protein synthesis, membrane
fluidity, and the transport of metabolites and transmission of
signals across membranes. By impairing methyltransferase
activity, SAMe depletion exacerbates the membrane injury
induced by alcohol. Furthermore, SAMe plays a key role in
the synthesis of polyamines and provides a source of cysteine
for GSH production (Fig. 4).
4.1.2. Therapeutic applications of S-adenosylmethionine,
including clinical trials
In baboons, correction of the ethanol-induced hepatic
SAMe depletion with oral SAMe administration resulted in
a corresponding attenuation of ethanol-induced liver injury,
as shown by a less striking GSH depletion, lesser increases
Fig. 4. Link between enhanced lipid peroxidation and accelerated acetalde-
hyde production as well as increased free radical generation by the induced
microsomes, with sites of possible therapeutic interventions. Metabolic
blocks caused by liver disease (a, b) or folate (c), B12 (c), or B6 (d) deficiencies
are illustrated, with corresponding depletions in S-adenosylmethionine,
phosphatidylcholine, and reduced glutathione (GSH). New therapeutic ap-
proaches include down-regulation of microsomal enzyme induction,
especially of cytochrome P450 2E1 (CYP2E1); decrease of free radicals
with antioxidants; and replenishment of S-adenosylmethionine and of
phosphatidylcholine. ADH ⫽ Alcohol dehydrogenase. Reprinted from
Journal of Hepatology, 32, C. S. Lieber, Alcoholic liver disease: new
insights in pathogenesis lead to new treatments, 113–128, Copyright (2000),
with permission from Elsevier.
 C.S. Lieber / Alcohol 34 (2004) 9–19
in plasma aspartate aminotransferase and glutamic dehydro-
genase activities, and fewer megamitochondria (Lieber et al.,
1990). Treatment with SAMe is also beneficial in human
beings, as shown in a prospective, multicenter, double-blind,
placebo-controlled trial (Frezza et al., 1990b) performed in
220 inpatients with chronic liver diseases (chronic active
hepatitis and cirrhosis, including primary biliary cirrhosis)
in whom serum markers of cholestasis and subjective symp-
toms significantly improved after SAMe treatment. Oral ad-
ministration of SAMe at 1,200 mg/day for 6 months also
resulted in a significant increase of hepatic GSH concentra-
tions in patients with alcoholic as well as non-alcoholic
liver disease (Vendemiale et al., 1989). Noteworthy is a 2-
year randomized, placebo-controlled, double-blind, multi-
center clinical trial of SAMe in patients with alcoholic
liver cirrhosis (Child class A and B) in whom SAMe improved
survival (29% vs. 12%, P ⫽ .025) or delayed liver trans-
plantation (Mato et al., 1999).
4.2. Polyenylphosphatidylcholine and other antioxidants
4.2.1. Pathogenesis of the deficiency and its consequences
Characteristic features of alcoholic liver injury include
striking membrane alterations with associated phospholipid
depletion (Lieber, 1992) as well as scarring (or fibrosis).
Fibrosis is characterized by the accumulation of excess colla-
gen after chronic ethanol consumption. This is due, at least
in part, to acetaldehyde, which stimulates collagen produc-
tion in hepatic stellate cells (Moshage et al., 1990) (vide
supra). Furthermore, fibrosis is related to associated meta-
bolic effects of ethanol. Indeed, the hepatic oxidation of
ethanol to acetaldehyde in the ADH pathway and the further
oxidation of acetaldehyde to acetate reduce NAD to NADH,
thereby producing striking redox changes that have an
impact on collagen metabolism. Indeed, the rise in NADH
shifts pyruvate to lactate, which stimulates collagen synthe-
sis in cultured myofibroblasts (Savolainen et al., 1984). This
finding seemed to indicate that ethanol-induced redox
changes may play a role, which was confirmed by the obser-
vation that methylene blue, a scavenger of reducing equiva-
lents, totally inhibits the stimulatory effect of acetaldehyde
on alpha1(I) procollagen and fibronectin gene expression in
stellate cells (Casini et al., 1991).
Some progress was made concerning possible therapeutic
effects of agents that oppose the alcohol-induced oxidative
stress. Indeed, contrary to ADH, the CYP2E1 is strikingly
inducible by chronic ethanol consumption, which results in
an increased production of not only acetaldehyde but also
free radicals (Lieber, 1997). Replenishment of GSH can be
achieved by administration of precursors of cysteine (one of
the amino acids of this tripeptide), such as acetylcysteine
or SAMe (Lieber, 1999; Lieber et al., 1990). Experimentally,
CYP2E1 has also been down-regulated by PPC (Aleynik
et al., 1999) and by its main component DLPC (Aleynik
& Lieber, 2001), a potentially beneficial therapeutic approach
(vide supra). The resulting oxidative stress is exacerbated
15
by acetaldehyde-mediated impairments of the antioxidant
systems, including depletion of GSH (Fig. 4). This results,
as reviewed elsewhere (Lieber, 1999), in peroxidation of
lipids, including PCs. They form the backbone of the mem-
branes, which are strikingly altered, as illustrated by the
blebbing of the plasma membranes in alcohol-fed rats
(Yamada et al., 1985), as well as the distortion of the mito-
chondrial membranes in baboons and human beings consum-
ing alcohol, even when their diets are nutritionally adequate
(Lieber, 1992). Cytochrome P450 2E1 generates several
species of active oxygen (Fig. 3), and GSH offers one of
the mechanisms for the scavenging of these toxic free radi-
cals (Fig. 4). Eventually, however, GSH is depleted, and the
oxidative stress promotes injury by inactivation of enzymes
and peroxidation of lipids. In patients with cirrhosis, hepatic
depletion of alpha-tocopherol (Leo et al., 1993), a major
antioxidant, potentiates this effect.
4.2.2. Therapeutic approaches with PPC, including clini-
cal trials
Phosphatidylcholines were replenished in the membranes
by administration of PPC, a mixture of polyunsaturated
PCs rich in DLPC, selected for its high bioavailability.
Also, PPC totally prevented the associated septal fibrosis
and cirrhosis in baboons fed adequate diets (Lieber et al.,
1994b), with an associated reduction in the number of acti-
vated stellate cells. These had been found to be strikingly
increased in alcohol-fed baboons, contributing to the pro-
gression of fibrosis (Mak et al., 1984). A similar transforma-
tion of stellate cells was observed in patients with alcoholic
liver disease (Mak & Lieber, 1988), and an attenuation of
the activation of stellate cells to myofibroblast-like cells was
also observed in vitro (Poniachik et al., 1999). In addition,
the activity of membrane-bound enzymes was normalized:
Cytochrome oxidase, a key enzyme of the mitochondrial
electron-transport chain, which requires a normal phospho-
lipid milieu for optimal activity and is severely depressed
by chronic ethanol consumption, was restored to normal
by addition of PC in vitro (Arai et al., 1984) or by PPC
supplementation in vivo (Navder et al., 1997), with improve-
ment of hepatic mitochondrial respiration.
One concern was that PPC and DLPC, because of their
polyunsaturated nature, may aggravate the oxidative stress,
but the opposite was found, both in vitro and in vivo. In
alcohol-fed baboons, PPC not only prevented septal fibrosis
and cirrhosis (Lieber et al., 1994b), but it also resulted in a
protection against oxidative stress, as determined by normal-
ization of 4-hydroxynonenal, F2-isoprostanes, and GSH
concentrations (Lieber et al., 1997).
Even in an experimental model devoid of oxidative stress,
namely fibrosis and cirrhosis induced by the injection of
heterologous albumin in rats, PPC was nevertheless protec-
tive (Ma et al., 1996). This may reflect, at least in part, that
both PPC (Li et al., 1992) and DLPC (Lieber et al., 1994b)
stimulate collagenase production in stellate cells, resulting
16 C.S. Lieber / Alcohol 34 (2004) 9–19
in a decrease of collagen accumulation. The activity of phos-
phatidylethanolamine methyltransferase, a key enzyme for
the regeneration of hepatic PC and which is depressed in
alcoholic liver disease (Fig. 4), was also restored with PPC
treatment (Lieber et al., 1994a).
All these favorable effects of PPC provided a basis for
some of the reported therapeutic effects of unsaturated phos-
pholipids in the treatment of liver diseases, including in
patients with alcoholic hepatitis (Bird et al., 1991). Further-
more, PC interfered with collagen formation from stellate
cells (Lieber et al., 1994b), which have a predominant role in
alcohol-induced fibrogenesis. Subsequently, PPC improved
liver test results in patients with hepatitis C (Niederau et al.,
1998) and ameliorated alcoholic liver disease in a pilot study
of patients with underlying fibrosis (Lieber et al., 2000).
This was followed by a randomized, prospective, double-
blind, placebo-controlled clinical trial conducted in 20
Veterans Affairs Medical Centers with 789 patients (97%
male; mean age, 48.8 years) averaging 16 drinks per day
(1 drink ⫽ 14 g of alcohol) for 19 years (Lieber et al., 2003b,
2003c). Evaluation of a baseline liver biopsy sample con-
firmed the presence of perivenular or septal fibrosis or
incomplete cirrhosis. Alcohol intake was reduced in both
groups to approximately 2 1/2 drinks per day as a result
of the “Brief Intervention” approach (Lieber et al., 2003b,
2003c). Accordingly, there was no further progression of
the fibrosis and, therefore, no way to test whether PPC could
oppose such a progression, except in a subgroup who were
still consuming 6 or more drinks a day and showed benefi-
cial effects against fibrosis. Ascites, an important secondary
clinical measure of liver disease, was also observed less
frequently during monitoring of PPC-treated patients. Fur-
thermore, improvement in concentrations of transaminases
and bilirubin favoring PPC was seen at some time points in
the subgroups of drinkers who were seropositive for hepatitis
C virus.
4.2.3. Silymarin and other antioxidants
Silymarin was found to be effective against various liver
injuries in rodents. In patients with alcoholic liver disease,
results of some randomized controlled trials with silymarin
showed beneficial effects such as improved survival (Ferenci
et al., 1989). However, findings of other studies did not
verify such an effect (Parés et al., 1998). To clarify this issue,
silymarin was studied under controlled conditions in non-
human primates, and it was found to oppose the alcohol-
induced oxidative stress and to retard the development of
alcohol-induced hepatic fibrosis (Lieber et al., 2003a). The
negative outcome observed in some clinical trials possibly
reflects poor compliance. Thus, in view of the innocuity of
silymarin, it might be justified to verify its effect in additional
clinical studies. Theoretically, other antioxidants such as
alpha-tocopherol might be useful. Indeed, in patients with
cirrhosis, diminished hepatic vitamin E concentrations have
been observed (Leo et al., 1993). However, therapeutic
results were disappointing (de la Maza et al., 1995).
5. Conclusions
Much progress has been made in the understanding of the
pathogenesis of alcoholic liver disease, resulting in improved
prevention and therapy, with promising prospects for even
more effective treatments. The most successful approaches
that one can expect to evolve are those that deal with the
fundamental cellular disturbances resulting from excessive
alcohol consumption. Two pathologic concepts are emerging
as particularly useful therapeutically. Whereas it continues
to be important to replenish nutritional deficiencies, when
present, it is crucial to recognize that, because of the alcohol-
induced disease process, some of the nutritional require-
ments change. This is exemplified by methionine: It normally
is one of the essential amino acids for human beings, but
it needs to be activated to SAMe, a process impaired by
the disease. Thus, SAMe, rather than methionine, is the
compound that must be supplemented in the presence of
significant liver disease. Indeed, SAMe was found to attenu-
ate mitochondrial lesions in baboons, replenish GSH, and
significantly reduce mortality in patients with Child A or B
cirrhosis. Similarly, PPC corrects the ethanol-induced he-
patic phospholipid depletion, restores fully or partially
phosphatidylethanolamine methyltransferase activity, and
opposes oxidative stress. It also deactivates hepatic stellate
cells, whereas its DLPC increases collagenase activity, result-
ing in prevention of ethanol-induced septal fibrosis and
cirrhosis in the baboon. Furthermore, CYP2E1, when ex-
cessively induced, becomes harmful and should be down-
regulated. PPC is one of the substances with anti-CYP2E1
properties that is now emerging (Aleynik & Lieber, 2001).
A quarter of all patients with alcoholic liver disease are
seropositive for hepatitis C virus, with an even higher inci-
dence in some urban areas. In addition to antiviral medica-
tions, agents that oppose oxidative stress and fibrosis are
being tested for hepatitis C treatment because these two
processes contribute much to the pathologic changes and
mortality associated with the virus.
Thus, effective prevention and therapy against steatosis
and its progression to more severe injury can be achieved
by a multifactorial approach: control of alcohol consump-
tion, avoidance of obesity and excess dietary long-chain fatty
acids, or their replacement with medium-chain triglycerides,
and replenishment of SAMe and the PCs.
Acknowledgments
Original studies were supported, in part, by NIH grants
AA11115 and AT001583; the Department of Veterans Af-
fairs; and the Kingsbridge and C.D. Smithers Research Foun-
dations. I am grateful to Ms. Y. Rodriguez for the skillful
typing of this document.
References
Adas, F., Berthou, F., Picart, D., Lozac’h, P., Beaugé, F., & Amet, Y. (1998).
Involvement of cytochrome P450 2E1 in the (ω–1)-hydroxylation
 C.S. Lieber / Alcohol 34 (2004) 9–19
of oleic acid in human and rat liver microsomes. J Lipid Res 39,
1210–1219.
Aleynik, M. K., Leo, M. A., Aleynik, S. I., & Lieber, C. S. (1999). Polyenyl-
phosphatidylcholine opposes the increase of cytochrome P-4502E1 by
ethanol and corrects its iron-induced decrease. Alcohol Clin Exp Res
23, 96–100.
Aleynik, M. K., & Lieber, C. S. (2001). Dilinoleoylphosphatidylcholine
decreases ethanol-induced cytochrome P4502E1. Biochem Biophys Res
Commun 288, 1047–1051.
Amet, Y., Berthou, F., Goasduff, T., Salaun, J. P., Le Breton, L., & Menez,
J. F. (1994). Evidence that cytochrome P450 2E1 is involved in the
(ω-1)-hydroxylation of lauric acid in rat liver microsomes. Biochem
Biophys Res Commun 203, 1168–1174.
Angulo, P., & Lindor, K. D. (2001). Insulin resistance and mitochondrial
abnormalities in NASH: a cool look into a burning issue. Gastroenter-
ology 120, 1281–1285 (Editorial).
Arai, M., Gordon, E. R., & Lieber, C. S. (1984). Decreased cytochrome
oxidase activity in hepatic mitochondria after chronic ethanol consump-
tion and the possible role of decreased cytochrome aa3 content and
changes in phospholipids. Biochim Biophys Acta 797, 320–327.
Baraona, E., Matsumura, T., Hernandez, R., Kubota, S., Soong, J., Kawano,
S., Kato, S., Inatomi, N., Sato, S., & Lieber, C.S. (1988). Chronic
alcohol intake impairs liver oxygenation by decreasing O2 utilization.
In K. Kuriyama, A. Takada, & H. Ishii (Eds.), Biomedical and Social
Aspects of Alcohol and Alcoholism: Proceedings of the Fourth Congress
of the International Society for Biomedical Research on Alcoholism
(pp. 387–390). Amsterdam, The Netherlands: Elsevier Science Publish-
ers BV.
Batey, R., Cao, Q., Madsen, G., Pang, G., Russell, A., & Clancy, R.
(1998). Decreased tumor necrosis factor-α and interleukin-1α production
from intrahepatic mononuclear cells in chronic ethanol consumption and
upregulation by endotoxin. Alcohol Clin Exp Res 22, 150–156.
Best, C. H., Hartroft, W. S., Lucas, C. C., & Ridout, J. H. (1949). Liver
damage produced by feeding alcohol or sugar and its prevention by
choline. Br Med J 2, 1001–1006.
Bird, G. L. A., Panos, M. Z., Polson, R., Johnson, R., Portman, B., Neu-
berger, J., & Williams, R. (1991). Activity of polyunsaturated phosphati-
dylcholine in HbsAg negative chronic active hepatitis and acute
alcoholic hepatitis. Z Gastroenterol 29, 21–24.
Boleda, M. D., Saubi, N., Farrés, J., & Parés, X. (1993). Physiological
substrates for rat alcohol dehydrogenase classes: aldehydes of lipid
peroxidation, ω-hydroxyfatty acids, and retinoids. Arch Biochem Bio-
phys 307, 85–90.
Burnett, D. A., Lysenko, N., Manning, J. A., & Ockner, R. K. (1979).
Utilization of long chain fatty acids by rat liver: studies of the role of
fatty acid binding protein. Gastroenterology 77, 241–249.
Cao, Q., Mak, K. M., & Lieber, C. S. (2002). DLPC decreases TGF-
β1-induced collagen mRNA by inhibiting p38 MAPK in hepatic stellate
cells. Am J Physiol Gastrointest Liver Physiol 283, G1051–G1061.
Casazza, J. P., Felver, M. E., & Veech, R. L. (1984). The metabolism of
acetone in rat. J Biol Chem 259, 231–236.
Casazza, J. P., & Veech, R. L. (1985). The production of 1,2-propanediol
in ethanol treated rats. Biochem Biophys Res Commun 129, 426–430.
Casini, A., Cunningham, M., Rojkind, M., & Lieber, C. S. (1991). Acetalde-
hyde increases procollagen type I and fibronectin gene transcription
in cultured rat fat-storing cells through a protein synthesis-dependent
mechanism. Hepatology 13, 758–765.
Chedid, A., Mendenhall, C. L., Gartside, P., French, S. W., Chen, T., Rabin,
L., and the VA Cooperative Study Group. (1991). Prognostic factors
in alcoholic liver disease. Am J Gastroenterol 86, 210–216.
Conney, A. H. (1967). Pharmacological implications of microsomal enzyme
induction. Pharmacol Rev 19, 317–366.
de la Maza, M. P., Petermann, M., Bunout, D., & Hirsch, S. (1995). Effects
of long-term vitamin E supplementation in alcoholic cirrhotics. J Am
Coll Nutr 14, 192–196.
Duce, A. M., Ortiz, P., Cabrero, C., & Mato, J. M. (1988). S-adenosyl-L-
methionine synthetase and phospholipid methyltransferase are inhibited
in human cirrhosis. Hepatology 8, 65–68.
17
Eriksson, C. J. P., Fukunaga, T., Sarkola, T., Lindholm, H., & Ahola,
L. (1996). Estrogen-related acetaldehyde elevation in women during
alcohol intoxication. Alcohol Clin Exp Res 20, 1192–1195.
Espina, N., Lima, V., Lieber, C. S., & Garro, A. J. (1988). In vitro and in
vivo inhibitory effect of ethanol and acetaldehyde on O6-methylguanine
transferase. Carcinogenesis 9, 761–766.
Ferenci, P., Dragosics, B., Dittrich, H., Frank, H., Benda, L., Lochs, H.,
Meryn, S., Base, W., & Schneider, B. (1989). Randomized controlled
trial of silymarin treatment in patients with cirrhosis of the liver. J
Hepatol 9, 105–113.
Frezza, M., di Padova, C., Pozzato, G., Terpin, M., Baraona, E., & Lieber,
C. S. (1990a). High blood alcohol levels in women. The role of decreased
gastric alcohol dehydrogenase activity and first-pass metabolism. N
Engl J Med 322, 95–99.
Frezza, M., Surrenti, C., Manzillo, G., Fiaccadori, F., Bortolini, M., & Di
Padova, C. (1990b). Oral S-adenosylmethionine in the symptomatic
treatment of intrahepatic cholestasis. A double-blind, placebo-controlled
study. Gastroenterology 99, 211–215.
Galli, A., Price, D., & Crabb, D. (1999). High-level expression of rat class
I alcohol dehydrogenase is sufficient for ethanol-induced fat accumula-
tion in transduced HeLa cells. Hepatology 29, 1164–1170.
Hasumura, Y., Teschke, R., & Lieber, C. S. (1976). Characteristics of
acetaldehyde oxidation in rat liver mitochondria. J Biol Chem 251,
4908–4913.
Hoerner, M., Behrens, U. J., Worner, T., & Lieber, C. S. (1986). Humoral
immune response to acetaldehyde adducts in alcoholic patients. Res
Commun Chem Pathol Pharmacol 54, 3–12.
Iseri, O. A., Gottlieb, L. S., & Lieber, C. S. (1964). The ultrastructure of
ethanol induced fatty liver. Fed Proc 23, 579A (Abstract).
Iseri, O. A., Lieber, C. S., & Gottlieb, L. S. (1966). The ultrastructure of
fatty liver induced by prolonged ethanol ingestion. Am J Pathol 48,
535–555.
Ishii, H., Joly, J.-G., & Lieber, C. S. (1973a). Effect of ethanol on the
amount and enzyme activities of hepatic rough and smooth microsomal
membranes. Biochim Biophys Acta 291, 411–420.
Ishii, H., Joly, J.-G., & Lieber, C. S. (1973b). Increase of microsomal
glucose-6-phosphatase activity after chronic ethanol administration.
Metabolism 22, 799–806.
Isselbacher, K. J., & Carter, E. A. (1970). Ethanol oxidation by liver
microsomes: evidence against a separate and distinct enzyme system.
Biochem Biophys Res Commun 39, 530–537.
James, O. F. W., & Day, C. P. (1998). Non-alcoholic steatohepatitis (NASH):
a disease of emerging identity and importance. J Hepatol 29, 495–501.
Johansson, I., Eliasson, E., Norsten, C., & Ingelman-Sundberg, M. (1986).
Hydroxylation of acetone by ethanol- and acetone-inducible cytochrome
P-450 in liver microsomes and reconstituted membranes. FEBS Lett
196, 59–64.
Jones, B. M., & Jones, M. K. (1976). Male and female intoxication levels
for three alcohol doses or do women really get higher than men?
Alcohol Technical Report 5, 544–583.
Kaikaus, R. M., Chan, W. K., Ortiz de Montellano, P. R., & Bass, N. M.
(1993). Mechanisms of regulation of liver fatty acid–binding protein.
Mol Cell Biochem 123, 93–100.
Kato, J., Sato, Y., Inui, N., Nakano, Y., Takimoto, R., Takada, K., Kobune,
M., Kuroiwa, G., Miyake, S., Kohgo, Y., & Niitsu, Y. (2003). Ethanol
induces transforming growth factor-α expression in hepatocytes, leading
to stimulation of collagen synthesis by hepatic stellate cells. Alcohol
Clin Exp Res 27(8 Suppl), 58S–63S.
Khani, S. C., Porter, T. D., & Coon, M. J. (1988). Isolation and partial
characterization of the gene for cytochrome P-450 3a (P-450ALC) and
a second closely related gene. Biochem Biophys Res Commun 150,
10–17.
Koivisto, T., Mishin, V. M., Mak, K. M., Cohen, P. A., & Lieber, C. S.
(1996). Induction of cytochrome P-4502E1 by ethanol in rat Kupffer
cells. Alcohol Clin Exp Res 20, 207–212.
Koop, D. R. (1992). Oxidative and reductive metabolism by cytochrome
P450 2E1. FASEB J 6, 724–730.
18 C.S. Lieber / Alcohol 34 (2004) 9–19
Koop, D. R., & Casazza, J. P. (1985). Identification of ethanol-inducible
P-450 isozyme 3a as the acetone and acetol monooxygenase of rabbit
microsomes. J Biol Chem 260, 13607–13612.
Koop, D. R., & Tierney, D. J. (1990). Multiple mechanisms in the regulation
of ethanol-inducible cytochrome P450IIE1. Bioessays 12, 429–435.
Laethem, R. M., Balazy, M., Falck, J. R., Laethem, C. L., & Koop, D. R.
(1993). Formation of 19(S)-, 19(R)-, and 18(R)-hydroxyeicosatetraenoic
acids by alcohol-inducible cytochrome P450 2E1. J Biol Chem 268,
12912–12918.
Lane, B. P., & Lieber, C. S. (1966). Ultrastructural alterations in human
hepatocytes following ingestion of ethanol with adequate diets. Am J
Pathol 49, 593–603.
Lane, B. P., & Lieber, C. S. (1967). Effects of butylated hydroxytoluene
on the ultrastructure of rat hepatocytes. Lab Invest 16, 342–348.
Leo, M. A., Rosman, A. S., & Lieber, C. S. (1993). Differential depletion
of carotenoids and tocopherol in liver disease. Hepatology 17, 977–986.
Li, J., Kim, C. I., Leo, M. A., Mak, K. M., Rojkind, M., & Lieber, C. S.
(1992). Polyunsaturated lecithin prevents acetaldehyde-mediated he-
patic collagen accumulation by stimulating collagenase activity in cul-
tured lipocytes. Hepatology 15, 373–381.
Lieber, C. S. (1992). Medical and Nutritional Complications of Alcoholism:
Mechanisms and Management. New York: Plenum Press.
Lieber, C. S. (1997). Cytochrome P-4502E1: its physiological and patholog-
ical role. Physiol Rev 77, 517–544.
Lieber, C. S. (1999). Microsomal ethanol-oxidizing system (MEOS): the
first 30 years (1968–1998)—a review. Alcohol Clin Exp Res 23,
991–1007.
Lieber, C. S. (2000). Alcoholic liver disease: new insights in pathogenesis
lead to new treatments. J Hepatol 32(suppl 1), 113–128.
Lieber, C. S. (2001). Alcoholic liver injury: pathogenesis and therapy in
2001. Pathol Biol (Paris) 49, 738–752.
Lieber, C. S., Baraona, E., Hernández-Muñoz, R., Kubota, S., Sato, N.,
Kawano, S., Matsumura, T., & Inatomi, N. (1989). Impaired oxygen uti-
lization: a new mechanism for the hepatotoxicity of ethanol in sub-
human primates. J Clin Invest 83, 1682–1690.
Lieber, C. S., Casini, A., DeCarli, L. M., Kim, C. I., Lowe, N., Sasaki,
R., & Leo, M. A. (1990). S-adenosyl-L-methionine attenuates alcohol-
induced liver injury in the baboon. Hepatology 11, 165–172.
Lieber, C. S., & DeCarli, L. M. (1968). Ethanol oxidation by hepatic
microsomes: adaptive increase after ethanol feeding. Science 162,
917–918.
Lieber, C. S., & DeCarli, L. M. (1970). Quantitative relationship between
amount of dietary fat and severity of the alcoholic fatty liver. Am J
Clin Nutr 23, 474–478.
Lieber, C. S., & DeCarli, L. M. (1974a). An experimental model of alcohol
feeding and liver injury in the baboon. J Med Primatol 3, 153–163.
Lieber, C. S., & DeCarli, L. M. (1974b). Oxidation of ethanol by hepatic
microsomes of acatalasemic mice. Biochem Biophys Res Commun 60,
1187–1192.
Lieber, C. S., DeCarli, L. M., & Schmid, R. (1959). Effects of ethanol on
fatty acid metabolism in liver slices. Biochem Biophys Res Commun
1, 302–306.
Lieber, C. S., Jones, D. P., & DeCarli, L. M. (1965). Effects of prolonged
ethanol intake: production of fatty liver despite adequate diets. J Clin
Invest 44, 1009–1021.
Lieber, C. S., Jones, D. P., Mendelson, J., & DeCarli, L. M. (1963). Fatty
liver, hyperlipemia and hyperuricemia produced by prolonged alcohol
consumption, despite adequate dietary intake. Trans Assoc Am Physi-
cians 76, 289–300.
Lieber, C. S., Lefèvre, A., Spritz, N., Feinman, L., & DeCarli, L. M.
(1967). Difference in hepatic metabolism of long- and medium-chain
fatty acids: the role of fatty acid chain length in the production of the
alcoholic fatty liver. J Clin Invest 46, 1451–1460.
Lieber, C. S., Leo, M. A., Aleynik, S. I., Aleynik, M. K., & DeCarli,
L. M. (1997). Polyenylphosphatidylcholine decreases alcohol-induced
oxidative stress in the baboon. Alcohol Clin Exp Res 21, 375–379.
Lieber, C. S., Leo, M. A., Aleynik, S. I., Lowe, N., Aleynik, M. K., &
Paronetto, F. (2000). Increasing circulating level of dilinoleoylphospha-
tidylcholine is associated with protection against alcohol induced oxi-
dative stress and liver fibrosis in man. Hepatology 32, 386A (Abstract).
Lieber, C. S., Leo, M. A., Cao, Q., Ren, C., & DeCarli, L. M. (2003a).
Silymarin retards the progression of alcohol-induced hepatic fibrosis
in baboons. J Clin Gastroenterol 37, 336–339.
Lieber, C. S., Robins, S. J., & Leo, M. A. (1994a). Hepatic phosphatidyletha-
nolamine methytransferase activity is decreased by ethanol and in-
creased by phosphatidylcholine. Alcohol Clin Exp Res 18, 592–595.
Lieber, C. S., Robins, S. J., Li, J., DeCarli, L. M., Mak, K. M., Fasulo, J. M., &
Leo, M. A. (1994b). Phosphatidylcholine protects against fibrosis and
cirrhosis in the baboon. Gastroenterology 106, 152–159.
Lieber, C. S., & Schmid, R. (1961). The effect of ethanol on fatty acid
metabolism: stimulation of hepatic fatty acid synthesis in vitro. J Clin
Invest 40, 394–399.
Lieber, C. S., Weiss, D. G., Groszmann, R., Paronetto, F., & Schenker, S.,
for the Veterans Affairs Cooperative Study 391 Group. (2003b). I.
Veterans Affairs Cooperative Study of polyenylphosphatidylcholine
in alcoholic liver disease: effects on drinking behavior by nurse/physician
teams. Alcohol Clin Exp Res 27, 1757–1764.
Lieber, C. S., Weiss, D. G., Groszmann, R., Paronetto, F., & Schenker, S.,
for the Veterans Affairs Cooperative Study 391 Group. (2003c). II.
Veterans Affairs Cooperative Study of polyenylphosphatidylcholine in
alcoholic liver disease. Alcohol Clin Exp Res 27, 1765–1772.
Ma, X., Baraona, E., Goozner, B. G., & Lieber, C. S. (1999). Gender
differences in medium-chain dicarboxylic aciduria in alcoholic men and
women. Am J Med 106, 70–75.
Ma, X., Baraona, E., & Lieber, C. S. (1993). Alcohol consumption enhances
fatty acid ω-oxidation, with a greater increase in male than in female
rats. Hepatology 18, 1247–1253.
Ma, X., Svegliati-Baroni, G., Poniachik, J., Baraona, E., & Lieber, C. S.
(1997). Collagen synthesis by liver stellate cells is released from its
normal feedback regulation by acetaldehyde-induced modification of
the carboxyl-terminal propeptide of procollagen. Alcohol Clin Exp Res
21, 1204–1211.
Ma, X., Zhao, J., & Lieber, C. S. (1996). Polyenylphosphatidylcholine
attenuates non-alcoholic hepatic fibrosis and accelerates its regression.
J Hepatol 24, 604–613.
Mak, K. M., Leo, M. A., & Lieber, C. S. (1984). Alcoholic liver injury in
baboons: transformation of lipocytes to transitional cells. Gastroenterol-
ogy 87, 188–200.
Mak, K. M., & Lieber, C. S. (1988). Lipocytes and transitional cells in
alcoholic liver disease: a morphometric study. Hepatology 8, 1027–
1033.
Mato, J. M., Cámara, J., Fernández de Paz, J., Caballerı́a, L., Coll, S.,
Caballero, A., Garcı́a-Buey, L., Beltrán, J., Benita, V., Caballerı́a, J., Solà,
R., Moreno-Otero, R., Barrao, F., Martı́n-Duce, A., Correa, J. A., Parés,
A., Barrao, E., Garcı́a-Magaz, I., Puerta, J. L., Moreno, J., Boissard,
G., Ortiz, P., & Rodés, J. (1999). S-Adenosylmethionine in alcoholic
liver cirrhosis: a randomized, placebo-controlled, double-blind, multi-
center clinical trial. J Hepatology 30, 1081–1089.
Meldolesi, J. (1967). On the significance of the hypertrophy of the smooth
endoplasmic reticulum in liver cells after administration of drugs.
Biochem Pharmacol 16, 125–131.
Mishra, L., Sharma, S., Potter, J. J., & Mezey, E. (1989). More rapid
elimination of alcohol in women as compared to their male siblings.
Alcohol Clin Exp Res 13, 752–754.
Miwa, G. T., Levin, W., Thomas, P. E., & Lu, A. Y. H. (1978). The direct
oxidation of ethanol by a catalase- and alcohol dehydrogenase–free
reconstituted system containing cytochrome P-4501. Arch Biochem
Biophys 187, 464–475.
Moshage, H., Casini, A., & Lieber, C. S. (1990). Acetaldehyde selectively
stimulates collagen production in cultured rat liver fat-storing cells but
not in hepatocytes. Hepatology 12(3 Pt 1), 511–518.
Navder, K. P., Baraona, E., & Lieber, C. S. (1997). Polyenylphosphatidyl-
choline attenuates alcohol-induced fatty liver and hyperlipemia in rats.
J Nutr 127, 1800–1806.
 C.S. Lieber / Alcohol 34 (2004) 9–19
Nelson, D. R., Kamataki, T., Waxman, D. J., Guengerich, F. P., Estabrook,
R. W., Feyereisen, R., Gonzalez, F. J., Coon, M. J., Gunsalus, I. C.,
Gotoh, O., Okuda, K., & Nebert, D. W. (1993). The P450 superfamily:
update on new sequences, gene mapping, accession numbers, early trivial
names of enzymes, and nomenclature. DNA Cell Biol 12, 1–51.
Niederau, C., Strohmeyer, G., Heintges, T., Peter, K., & Gopfert, E., for
Leich Study Group. (1998). Polyunsaturated phosphatidyl-choline and
interferon alpha for treatment of chronic hepatitis B and C: a multi-
center, randomized, double-blind, placebo-controlled trial. Hepatogas-
troenterology 45, 797–804.
Niemela, O., Klajner, F., Orrego, H., Vidins, E., Blendis, L., & Israel, Y.
(1987). Antibodies against acetaldehyde-modified protein epitopes in
human alcoholics. Hepatology 7, 1210–1214.
Ohnishi, K., & Lieber, C. S. (1977). Reconstitution of the microsomal
ethanol-oxidizing system: qualitative and quantitative changes of cyto-
chrome P-450 after chronic ethanol consumption. J Biol Chem 252,
7124–7131.
Oneta, C. M., Mak, K. M., & Lieber, C. S. (1999). Dilinoleoylphosphatidyl-
choline selectively modulates lipopolysaccharide-induced Kupffer cell
activation. J Lab Clin Med 134, 466–470.
Parés, A., Planas, R., Torres, M., Caballerı́a, J., Viver, J. M., Acero, D., Panés,
J., Rigau, J., Santos, J., & Rodés, J. (1998). Effects of silymarin in
alcoholic patients with cirrhosis of the liver: results of a controlled,
double-blind, randomized and multicenter trial. J Hepatol 28, 615–621.
Poniachik, J., Baraona, E., Zhao, J., & Lieber, C. S. (1999). Dilinoleoylphos-
phatidylcholine decreases hepatic stellate cell activation. J Lab Clin
Med 133, 342–348.
Raucy, J. L., Lasker, J. M., Kraner, J. C., Salazar, D. E., Lieber, C. S., &
Corcoran, G. B. (1991). Induction of cytochrome P450IIE1 in the obese
overfed rat. Mol Pharmacol 39, 275–280.
Reichard, G. A. Jr., Skutches, C. L., Hoeldtke, R. D., & Owen, O. E.
(1986). Acetone metabolism in humans during diabetic ketoacidosis.
Diabetes 35, 668–674.
Roach, M. K., Resse, W. N. Jr., & Creaven, P. J. (1969). Ethanol oxidation
in the microsomal fraction of rat liver. Biochem Biophys Res Commun
36, 596–602.
Salmela, K. S., Kessova, I. G., Tsyrlov, I. B., & Lieber, C. S. (1998).
Respective roles of human cytochrome P-4502E1, 1A2, and 3A4 in
the hepatic microsomal ethanol oxidizing system. Alcohol Clin Exp Res
22, 2125–2132.
Sanyal, A. J., Campbell-Sargent, C., Mirshahi, F., Rizzo, W. B., Contos,
M. J., Sterling, R. K., Luketic, V. A., Shiffman, M. L., & Clore, J. N.
(2001). Nonalcoholic steatohepatitis: association of insulin resistance
and mitochondrial abnormalities. Gastroenterology 120, 1183–1192.
Savolainen, E. R., Leo, M. A., Timpl, R., & Lieber, C. S. (1984). Acetalde-
hyde and lactate stimulate collagen synthesis of cultured baboon liver
myofibroblasts. Gastroenterology 87, 777–787.
Shevchuk, O., Baraona, E., Ma, X. L., Pignon, J. P., & Lieber, C. S.
(1991). Gender differences in the response of hepatic fatty acids and
cytosolic fatty acid–binding capacity to alcohol consumption in rats.
Proc Soc Exp Biol Med 198, 584–590.
Takahashi, T., Lasker, J. M., Rosman, A. S., & Lieber, C. S. (1993). Induction
of cytochrome P-4502E1 in the human liver by ethanol is caused by
a corresponding increase in encoding messenger RNA. Hepatology 17,
236–245.
19
Tephly, T. R., Tinelli, F., & Watkins, W. D. (1969). Alcohol metabolism:
role of microsomal oxidation in vivo. Science 166, 627–628.
Teschke, R., Hasumura, Y., Joly, J.-G., & Lieber, C. S. (1972). Microsomal
ethanol-oxidizing system (MEOS): purification and properties of a rat
liver system free of catalase and alcohol dehydrogenase. Biochem Bio-
phys Res Commun 49, 1187–1193.
Teschke, R., Hasumura, Y., & Lieber, C. S. (1975). Hepatic microsomal
alcohol-oxidizing system in normal and acatalasemic mice: its dissocia-
tion from the peroxidatic activity of catalase-H2O2. Mol Pharmacol 11,
841–849.
Teschke, R., Matsuzaki, S., Ohnishi, K., DeCarli, L. M., & Lieber, C. S.
(1977). Microsomal ethanol oxidizing system (MEOS): current status
of its characterization and its role. Alcohol Clin Exp Res 1, 7–15.
Thurman, R. G., & Brentzel, H. J. (1977). The role of alcohol dehydrogenase
in microsomal ethanol oxidation and the adaptive increase in ethanol
metabolism due to chronic treatment with ethanol. Alcohol Clin Exp
Res 1, 33–38.
Thurman, R. G., Ley, H. G., & Scholz, R. (1972). Hepatic microsomal
ethanol oxidation. Hydrogen peroxide formation and the role of catalase.
Eur J Biochem 25, 420–430.
Tsutsumi, M., Lasker, J. M., Shimizu, M., Rosman, A. S., & Lieber, C. S.
(1989). The intralobular distribution of ethanol-inducible P450IIE1 in
rat and human liver. Hepatology 10, 437–446.
Umeno, M., McBride, O. W., Yang, C. S., Gelboin, H. V., & Gonzalez, F. J.
(1988a). Human ethanol-inducible P450IIE1: complete gene sequence,
promoter characterization, chromosome mapping, and cDNA-directed
expression. Biochemistry 27, 9006–9013.
Umeno, M., Song, B. J., Kozak, C., Gelboin, H. V., & Gonzalez, F. J.
(1988b). The rat P450IIE1 gene: complete intron and exon sequence,
chromosome mapping, and correlation of developmental expression
with specific 5′ cytosine demethylation. J Biol Chem 263, 4956–4962.
Vatsis, K. P., & Schulman, M. P. (1973). Absence of ethanol metabolism
in “acatalatic” hepatic microsomes that oxidize drugs. Biochem Biophys
Res Commun 52, 588–594.
Vatsis, K. P., & Schulman, M. P. (1974). An unidentified constituent of
ethanol oxidation in hepatic microsomes. Fed Proc 33, 554A (Abstract).
Vendemiale, G., Altomare, E., Trizio, T., Le Grazie, C., Di Padova, C.,
Salerno, M. T., Carrieri, V., & Albano, O. (1989). Effects of oral
S-adenosyl-L-methionine on hepatic glutathione in patients with liver
disease. Scand J Gastroenterol 24, 407–415.
Weltman, M. D., Farrell, G. C., Hall, P., lngelman-Sundberg, M., & Liddle,
C. (1998). Hepatic cytochrome P450 2E1 is increased in patients with
nonalcoholic steatohepatitis. Hepatology 27, 128–133.
Wondergem, R., & Davis, J. (1994). Ethanol increases hepatocyte water
volume. Alcohol Clin Exp Res 18, 1230–1236.
Yamada, S., Mak, K. M., & Lieber, C. S. (1985). Chronic ethanol consump-
tion alters rat liver plasma membranes and potentiates release of alkaline
phosphatase. Gastroenterology 88, 1799–1806.
Yang, C. S., Yoo, J. S., Ishizaki, H., & Hong, J. Y. (1990). Cytochrome
P450IIE1: roles in nitrosamine metabolism and mechanisms of regula-
tion. Drug Metab Rev 22, 147–159.
Ziegler, D. M. (1972). Discussion. In R. W. Estabrook, R. J. Gillette, &
K. C. Lieberman (Eds.), Microsomes and Drug Oxidations (p. 458).
Baltimore: Williams & Wilkins.