Recent advances in clinical practice
Artificial and bioartificial liver
University of Technology of
Compiègne, UMR CNRS 6600 –
Biomechanics and
Bioengineering, Compiègne,
France
Correspondence to:
Dr C Legallais, UTC, UMR CNRS
6600 Biomechanics and
Bioengineering, BP 20529,
60205 Compiègne Cedex,
France; cecile.legallais@utc.fr
1690
devices: present and future
B Carpentier, A Gautier, C Legallais
ABSTRACT
Liver failure is associated with high morbidity and
mortality without transplantation. There are two types of
device for temporary support: artificial and bioartificial
livers. Artificial livers essentially use non-living compo-
nents to remove the toxins accumulated during liver
failure. Bioartificial livers have bioreactors containing
hepatocytes to provide both biotransformation and
synthetic liver functions. We review here the operating
principles, chemical effects, clinical effects and compli-
cations of both types, with specific attention paid to
bioartificial systems. Several artificial support systems
have FDA marketing authorisation or are CE labelled, but
the improvement they provide in terms of patient clinical
outcome has not yet been fully demonstrated. At present,
different bioartifical systems are being investigated
clinically on the basis of their promises and capacity to
provide and replace most liver functions. However,
important issues such as cost, cell availability, main-
tenance of cell viability and functionality throughout
treatment, and regulatory issues, as well as difficult
challenges, including implementing cell-housing devices at
the patient’s bedside on an emergency basis, have
delayed their appearance in intensive care units and on
the market. Bioreactors are, nevertheless, when com-
bined with artificial components, a pragmatic approach for
future treatment of liver failure.
Liver failure results from the liver’s inability to
perform its normal functions. It is a severe clinical
syndrome in which the liver’s metabolic functions
– detoxification, biotransformation, excretion and
synthesis – are severely impaired, leading to the
accumulation of lethal toxins in the patient and
the onset of life-threatening complications and
manifestations. The main detoxification and bio-
transformation functions of the liver are detailed in
the Appendix.
Liver failure is associated with a rise in numerous
endogenous substances such as bilirubin, ammo-
nia, glutamine, lactate, aromatic amino acids, free
fatty acids, phenol, mercaptans, benzodiazepines
and proinflammatory cytokines. These toxins are
known to play an important role in the pathogen-
esis of liver failure.1–5 The loss of liver function
appears to result from an overload of hepatotoxic
substances that progressively saturate available
detoxification pathways, leading to the accumula-
tion of other toxins and the production of
cytokines. Moreover, the accumulation of toxins
may further impair the patient’s liver as a result of
hepatocellular apoptosis and necrosis.
The most frequent complications of liver failure
are hepatic encephalopathy (HE), coagulopathy,
jaundice, cholestasis, pruritus, ascites, immune
disorders, sepsis and kidney failure.1 6
In acute liver failure (ALF), liver function is
normal 2–8 weeks before the onset of the disease.
In acute-on-chronic liver failure (A-on-C LF), liver
function decreases abruptly in a patient already
suffering from chronic liver insufficiency.1 6 The
most frequent causes of ALF are intoxications,
especially acetaminophen intoxication, viral infec-
tions and hepatic ischaemia. Sometimes the cause
remains unknown.7 8 A-on-C LF usually occurs in
patients with cirrhosis, following infectious dis-
orders, toxin exposure or gastrointestinal bleeding.
Both are associated with high morbidity and
mortality without transplantation.
Extracorporeal liver support devices have there-
fore been developed in the last few decades in order
to either bridge patients to liver transplantation or
allow the native liver to recover from injury. They
may also be valuable when primary non-function
occurs after liver transplantation or when a large
hepatic resection leaves too little liver in reserve.3 9
There are two types of liver assist devices:
artificial and bioartificial livers. Artificial liver
(AL) devices use non-living components to cleanse
the blood or plasma of its toxins. Removal is based
on physical/chemical gradients and adsorption.
Bioartificial liver (BAL) devices contain a cell-
housing bioreactor, the role of which is to replace
the primary and most important liver functions
(oxidative detoxification, biotransformation, excre-
tion and synthesis).
In this paper, the first part focuses on the current
therapies available in intensive care units (ICUs)
for treating or supporting patients with liver
failure. The operating principles of ALs, the effects
of ALs on chemical and clinical parameters and
their market status will be noted and discussed.
The second part will focus on future treatments
for liver failure patients. Special attention will be
given to BAL devices that are currently included in
clinical trials. Their operating principles, chemical
and clinical effects and side effects and complica-
tions will be reviewed.
LIVER SUPPORT FOR PATIENTS WITH LIVER
FAILURE: STATE OF THE ART
Social and economic data on liver support therapies
The World Health Organization estimates that
10% of the world’s population has chronic liver
disease, including 25 million Americans. Fulminant
hepatic failure generally ends in death within 96 h
without transplantation. This pathology affects
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around five people per million in Western coun-
tries; ie, approximately 2500 people per year.
Liver transplantation is the only efficient treat-
ment for patients with acute or fulminant organ
failure. The shortage of liver donors has resulted in a
high death rate in potential graft patients on a
waiting list for a suitable donor (fig 1). In the last two
decades, the expanding gap between the number of
patients on the waiting list for transplantation and
the number of transplants available has highlighted
the need for a temporary liver support system that
could be used on an emergency basis.
Moreover, not all patients are candidates for
transplants and substitute solutions must be found
to face this issue. Liver transplantation is a risky
treatment that requires life-long immunosuppres-
sion treatment. Xenotransplantation is not yet well
controlled and is subject to ethical questions.10
Partial transplantation of the liver within people
from the same family (living donors) or from
cadavers has nevertheless led to substantial progress.
Approaches, clinical results and market status of
the main artificial liver devices
Operating principles
Most ALs aim to replace detoxification functions
and use membrane separation associated with
columns or suspensions of sorbents – including
charcoal, anion-exchange or cation-exchange resins
– that selectively remove toxins and/or regenerate
dialysate or plasma filtrate. The treatment units
(TUs) that may be included in liver assist devices
are shown in fig 2. ALs do not contain bioreactors.
TUs are implemented either in parallel or in series
within the extracorporeal circuit (figs 3 and 4).
The Liver Dialysis device
Liver Dialysis (HemoCleanse, Lafayette, Indiana,
USA) – originally called Biologic-DT – was devel-
oped by Ash et al.11 12 The patient’s blood is moved
from a central vein via a single lumen multi-holed
catheter using a push–pull procedure, resulting in a
flow rate of 200–250 ml/min.13 Simultaneously, a
2-litre suspension of powdered activated charcoal
and cation exchange resin (TU 7) is pumped from a
sorbent bag through the dialysate side of a flat-
plate membrane dialyser (TU 4: 5 kDa molecular
weight cut-off (MWCO) cellulosic membranes)
and returned to the sorbent bag.
Liver Dialysis is currently being redesigned. A
method for immobilising the powdered charcoal in
a block is being evaluated for increased toxin
binding capacity and better regeneration of dialy-
sate.14 15 The next system should be able to support
any type of hollow fibre (TUs 1 to 4).
The Molecular Adsorbent Recirculating System
The Molecular Adsorbent Recirculating System
(MARS; Gambro Lundia, Lund, Sweden) was initi-
ally developed by Stange and Mitzner.1 16 The
patient’s blood is drawn from the femoral vein to a
high-flux albumin-coated polysulfone haemodialyser
(TU 6: surface area of 2.1 m2).9 17 Toxins (both
Gut 2009;58:1690–1702. doi:10.1136/gut.2008.175380
Recent advances in clinical practice
albumin-bound and soluble) are transferred to the
counter-current human albumin-enriched dialysate
(TU 5).18 The exogenous albumin dialysate is then
regenerated in a closed loop by dialysis (TU 4 using a
low-flux polysulfone membrane) against a conven-
tional bicarbonate-buffered dialysate and adsorption
through charcoal and anion-exchange resin columns
(TU 7).9 Blood and albumin dialysate flow rates are
usually set between 150 and 250 ml/min, and
bicarbonate dialysate between 300 and 500 ml/min.
The Prometheus device
Prometheus was designed by Fresenius Medical
Care (Bad Homburg, Germany).19 20 The patient’s
blood is drawn at a flow rate of 200 ml/min to an
albumin-permeable polysulfone filter (TU 2: Albu-
Flow, MWCO 250 kDa), generating an albumin-
containing filtrate. The endogenous albumin fil-
trate subsequently flows through a neutral resin
and an anion exchanger in series (TU 7). The blood
leaving the Albu-Flow is consequently dialysed in a
conventional high-flux polysulfone dialyser (TU 4),
against a 500 ml/min bicarbonate dialysate.
Clinical results and complications associated with the
treatment
Liver Dialysis, MARS and Prometheus treatments
are usually performed 6–8 h daily for several days
or until the patient’s blood pressure and liver
function have improved and the encephalopathy
has cleared.1 9 21 However, the optimal timing for
the initiation, frequency and duration of the
treatments, plus the biochemical and clinical
parameters on which to base the decision, remain
partially unanswered issues. The most important
randomised controlled trials to have been con-
ducted on the safety and efficacy of Liver Dialysis,
MARS and Prometheus are summarised in table 1.
The Liver Dialysis device
The largest prospective randomised trial with Liver
Dialysis was carried out on 56 patients.11 22
Compared with controls and standard medical
therapy (SMT), Liver Dialysis was responsible for
improved neurological status and blood pressure in
patients with either A-on-C LF or ALF and a higher
occurrence of positive outcome in patients with A-
on-C LF and stage III–IV encephalopathy (suffi-
cient recovery of liver function to allow hospital
discharge or adequate clinical improvement to
allow transplantation). In patients with liver failure
complicated with kidney failure (type I hepatorenal
syndrome), no continuous veno-venous haemofil-
tration was required as Liver Dialysis was able to
remove up to 3 litres of fluid throughout each
treatment without causing hypotension.
Liver Dialysis was associated with certain risks,
including an increased degree of bleeding in
patients with active bleeding and disseminated
intravascular coagulation – due to activation and
aggregation of the patients’ platelets – and clotting
of the blood circuit during treatment.
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 Recent advances in clinical practice
Figure 1 1994 to 2005 statistics from the European Liver Transplant Registry: time
course of cadaveric and living organ transplants, persons on the waiting list and
subsequent death while on the waiting list.
The Molecular Adsorbent Recirculating System
Significant reductions in serum bilirubin, bile acids,
ammonia, urea, lactate and creatinine levels have
repeatedly been documented with MARS ther-
apy.1 3 4 6 9 17 31–35 Total bilirubin and conjugated
bilirubin are reduced, whereas no changes in
unconjugated bilirubin levels are observed.34 MARS
does not interfere with valuable molecules such as
albumin,17 36 coagulation factors and electrolytes.6 33
MARS does not alter blood cell counts and arterial
blood gases and does not generate haemodynamic
instability.6 Improvement in mean arterial pressure
(MAP), systemic vascular resistance, cardiac output
and cerebral blood flow (due to reduced cerebral
oedema) have repeatedly been demon-
strated.17 31 32 37 38 Improvement in HE grade during
MARS treatment has frequently been observed
regardless of the aetiology of the liver fail-
ure.6 17 21 31 38–43 Improvement in HE was not neces-
sarily associated with significant changes in cytokine
and ammonia levels.24
Clinical outcome depends greatly on the aetiol-
ogy of the liver failure. Survival rates at the time of
discharge vary between 60% and 70% in patients
with either ALF or A-on-C LF of various aetiolo-
gies.32 43 In one study in which MARS therapy was
used to treat 56 patients with ALF of various
aetiologies, the survival rate was 88% at 6 months
and 84% at 1 year. The highest incidence of liver
recovery was observed in patients with ALF due to
intoxication.3 Some authors have reported the
successful support of patients with ALF before
liver transplantation.44 The impact of MARS on
the outcome of patients with A-on-C LF is
controversial. Heeman et al conducted a prospec-
tive randomised controlled study in 23 patients
with cirrhosis and with A-on-C LF.27 Renal
dysfunction, HE and 30-day survival improved
with MARS compared to SMT.
A prospective, randomised, controlled, multi-
centre trial – the largest published so far – has
1692
recently been completed to assess the efficacy,
safety and tolerability of MARS treatment in
patients with advanced cirrhosis associated with
severe HE.23 Besides safety and tolerability, higher,
earlier and more frequent improvements in HE
grade could be demonstrated when compared to
SMT.
Further randomised studies are necessary before
any strong conclusions can be drawn. Even though
many case-report studies have shown encouraging
patient outcome, the effects of MARS on clinical
outcome is not yet proven for ALF and A-on-C LF
patients. However, trends for improved survival
rates have shown up in several randomised studies.
Randomised, controlled trials in patients with
ALF (the FULMAR trial; preliminary data pre-
sented at the American Association for the Study
of Liver Diseases meeting in November 2008) and
A-on-C LF (the MARS RELIEF trial) are ongoing.
Adverse events or complications have sometimes
been noted, such as mild thrombocytopenia,9 17 45
and disseminated intravascular coagulation with
significant bleeding.42 MARS is associated with the
same risks as any other extracorporeal filtration
procedure requiring catheterisation.32 MARS seems
to be relatively safe but should be used with
caution in patients with a coagulopathy.
The Prometheus device
A significant decrease in both albumin-bound and
water-soluble toxins has repeatedly been demon-
strated throughout treatment with
Prometheus.2 19 46 47 The Prometheus treatment
improves serum levels of conjugated bilirubin, bile
acids, ammonia, creatinine, urea and blood
pH.2 19 46–48 Krisper et al have compared the effects
of MARS and Prometheus on toxin removal. They
observed a higher efficiency and delivered dose
with Prometheus compared to MARS. However,
this was not responsible for significant differences
in plasma levels.2 Substances such as albumin,
fibrinogen, coagulation factors, other plasma pro-
teins, electrolytes and cytokines (interleukin 6 and
tumour necrosis factor a) remained unchanged
during the treatment with Prometheus.20 46 48
Even though no randomised, controlled studies
have been reported yet, treatment with
Prometheus seems to have beneficial effects on
patient outcome.19 46 Further investigations must
be completed, especially in the course of a multi-
centre randomised study named HELIOS (ongoing
study).
Decreased blood pressure, clotting of the sec-
ondary circuit and a slight increase in leucocyte
counts were the only side effects or complications
reported for the Prometheus therapy.19 48
Market status of the major artificial liver devices
In 1997, Liver Dialysis was the first AL to receive
approval from the US Food and Drug
Administration (FDA) for use in the treatment of
encephalopathy secondary to liver failure, under
the name of Biologic-DT. Liver Dialysis is not
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 Recent advances in clinical practice

Figure 2 Treatment units (TUs): the principal unitary treatment blocks that may be included within an extracorporeal liver
support system. Each extracorporeal liver support system is based on a combination of three or more blocks that are added
either in series or in parallel within the circuit. The aim and methods on which each block is based are indicated. TU 1:
plasmapheresis – based either on centrifugation or microporous membrane filtration. TU 2: plasma fractionation – filtration
based on a very high permeability membrane (MWCO .70–100 kDa). TU 3: haemofiltration – filtration based on a medium
permeability membrane (15 kDa , MWCO ,70 kDa). TU 4: haemodialysis – diffusion through a low-to-medium
permeability membrane. A buffered dialysate present in the extracapillary space is used for the removal of water soluble
toxins. TU 5: albumin dialysis – same as TU 4, with albumin solution on the dialysate side. TU 6: aided transfer – same as TU
5, with an albumin-coated membrane to help the albumin-bound toxins cross the membrane. TU 7: adsorption on various
substrates to remove protein-bound toxins, such as bilirubin or bile acids. TU 8: bioreactor hosting cells for biotransformation
and synthesis functions. TU 9: oxygen supply. MWCO, molecular weight cut-off.

currently marketed in the USA as it is being
redesigned. MARS and Prometheus have received
the CE label to allow their sale in Europe. Although
they are used in many countries, they are not yet
available in the USA.
FUTURE TREATMENT OF LIVER FAILURE:
BIOARTIFICIAL LIVER SYSTEMS
The aim of BALs is to provide both liver detoxifica-
tion and synthetic functions, using a combination
of physical and chemical procedures, and bioreac-
tors hosting cells (fig 2). The advantages and
disadvantages of the various most commonly used
cell sources are summarised in table 2.
Four configurations of BAL are currently under
investigation: hollow fibre cartridges or chambers
(ELAD, HepatAssist, MELS), monolayer cultures,
perfused matrices (BLSS, AMC-BAL) within dedi-
cated devices and microencapsulation-based sys-
tems.49–51 Only the devices currently under clinical
investigation are described in detail in this paper.
The treatment units that make up a BAL are
implemented either in parallel or in series within
the extracorporeal circuit (figs 3 and 4).

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 Recent advances in clinical practice

Figure 3 Schematic
principles of extracorporeal
systems for the treatment
of liver failure patients.
Liver support systems are
designed in such a way
that liver failure toxins can
be removed from the
patient – and
biotransformation and
synthesis can occur when
bioartificial livers (BALs)
are considered – by
combining essential and
optional treatment units
together. Blood is drawn
from a central vein and
directed towards the
treatment units (TUs) (see
fig 2) before being returned
to the patient. Different
strategies have been
chosen for each device.
Treatment units are either
connected in series or in
parallel to the others,
depending on the device.

Bioreactor specifications for bioartificial liver
applications
Several requirements must be satisfied in order to
ensure full efficiency of the bioreactor. Functional
liver cells (table 3) must be isolated from the
external environment and immobilised on or
within a favourable substrate (fig 5).
The substrate should allow the preservation of
cell morphology and metabolism throughout the
treatment. Oxygen and nutrients should be acces-
sible to the cells in appropriate concentrations. A
porous material (not necessarily the substrate)
should act as a barrier between the patient’s blood
or plasma and the hepatocytes located within the

Figure 4 Summary of the

treatment units integrated
within each artificial or
bioartificial liver support
device. Refer to fig 2 to
identify each treatment unit
block.

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 Recent advances in clinical practice

Table 1 Randomised controlled trials on the safety and efficacy of artificial livers and bioartificial livers
Number of
Liver assist device patients Inclusion criteria Treatment Clinical results Authors

Liver Dialysis 56 A-on-C LF Liver Dialysis vs SMT Improved neurological status and MAP Ash et al11 22
ALF Increased degree of bleeding in patients
with active bleeding and DIVC

MARS 70 HE grade 3 or 4 MARS (6 h per day, 5 days) + Higher, earlier and more frequent Hassanein et al23
SMT or SMT alone improvement of HE compared to SMT
18 A-on-C LF due to alcohol abuse MARS (four sessions of 8 h, Improvement of HE Sen et al24
7 days) + SMT or SMT alone Unchanged MAP and renal function
No significant change in plasma
cytokines and ammonia levels
27 Hypoxic liver failure after MARS (three consecutive Improved survival El Banayosy et al25
cardiogenic shock sessions at least) vs SMT
Bilirubin levels greater than Further study with larger patient cohort
8 mg/ml needed
18 A-on-C LF MARS + SMT Decreased serum bilirubin levels Laleman et al26
Alcoholic cirrhosis and or Prometheus + SMT or Improvement in MAP and SVRI
superimposed alcoholic SMT alone – three Reduction of the hyperdynamic
hepatitis consecutive days (6 h circulation
sessions)
24 Decompensated cirrhosis MARS (2 weeks, up to 10 Improved 30-day survival Heemann et al27
Bilirubin greater than 20 mg/dl sessions) + SMT vs SMT Decrease in plasma bile acids and
not responding to prior SMT alone bilirubin
Improvement in renal dysfunction and HE

Prometheus 24 Decompensated cirrhosis Single 6 h treatment with No differences in systemic Dethloff et al28
Prometheus (study group), haemodynamics
MARS or haemodialysis No improvement in MAP
No adverse effects
Decrease in platelet count
18 A-on-C LF MARS + SMT or Prometheus Decreased serum bilirubin levels (more Laleman et al26
Alcoholic cirrhosis and + SMT or effective than MARS)
superimposed alcoholic SMT alone – three No improvement in MAP, SVRI and
hepatitis consecutive days (6 h hyperdynamic circulation
sessions)
HepatAssist 171 Fulminant or subfulminant HepatAssist (6 h daily) + SMT Improved survival at 30 days (71% vs Demetriou et al29
hepatic failure and primary non- vs SMT alone 62%). Good safety
function following liver
transplantation

ELAD 24 Acute liver failure with ELAD (median period of 72 h) Good biocompatibility Ellis et al30
potentially recoverable lesion or vs SMT DIVC or hypersensitivity reaction (2
fulfilment of criteria for cases)
transplantation Survival comparable to SMT
Better indices of prognosis required

BLSS None conducted yet

AMC-BAL None conducted yet
MELS None conducted yet
A-on-C LF, acute-on-chronic liver failure; ALF, acute liver failure; AMC-BAL, Academic Medical Center – Bioartificial Liver; BLSS, Bioartificial Liver Support System; DIVC, disseminated
intravascular coagulation; ELAD, Extracorporeal Liver Assist Device; HE, hepatic encephalopathy; MAP, mean arterial pressure; MARS, Molecular Adsorbent Recirculating System; MELS,
Molecular Extracorporeal Liver Support; SMT, standard medical therapy; SVRI, systemic vascular resistance index.

bioreactor in order to isolate the cells from the
immune factors (immunoglobulins, MW 150 kDa)
and leucocytes, and to avoid immune rejection
(table 4). Smaller particles such as toxins and
metabolites or synthesised proteins (albumin and
coagulation factors for instance) should be free to
cross the barrier.
The perfusion system is more or less com-
plex (table 5). An initial detoxification step
using charcoal and/or ion exchange resins could be
performed prior to the passage of the blood or
plasma through the bioreactor in order to avoid
the death of the pool of supporting cells.
An oxygenator can also be included upstream in the
circuit to improve oxygen delivery (fig 4).
Operating principles, and the chemical and clinical
effects of bioartificial livers
Here, we focus only on the devices that are
currently under clinical investigation. Their market
status and stage of advancement are detailed later
in the paper. HepatAssist and ELAD are the only
BAL systems on which randomised and controlled
trials have been performed (table 1).

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 Recent advances in clinical practice

Table 2 Advantages and disadvantages of potential cell sources for bioartificial livers
Cell source Advantages Disadvantages

Autologous cells harvested i. No immune reactions i. Limited availability

from the patient ii. Low risk of infection with ii. Heterogeneity in quality and behaviour
pathogens

Allogenic cells collected form i. High availability (above all, in i. Risk of disease transmission
donors emergencies) ii. Risk of immune response
ii. Pooling possible from different
donors

Allogenic cell lines (after i. High availability i. Loss of functions

genetic mutation making indefinite ii. Production of a large number of ii. Potential tumorigenicity
proliferation cells
possible)
iii. Infinite growth capability

Xenogenic cells isolated from i. High availability i. Risk of animal pathogen transmission
different species ii. Production of a large number of ii. Risk of immunogenic rejection
cells iii. Regulatory issues

The HepatAssist device HepatAssist was relatively well tolerated.

The HepatAssist device developed by Demetriou et
al initially used 56109 to 76109 cryopreserved
porcine hepatocytes constrained within the extra-
capillary compartment of a hollow fibre bioreactor
(TU 1 and 8).52 The cells are attached to micro-
carriers. The patient’s blood is separated into
plasma (TU 1) collected in a special bag used as a
buffer reservoir. The plasma passes through a
column of activated charcoal (TU 7) and an
oxygenator (TU 9) before it is circulated within
the bioreactor (fig 6a).
In a phase I clinical trial, 39 patients with ALF
were treated with HepatAssist. Thirty-two
patients were successfully bridged to liver trans-
plantation and six recovered without requiring a
graft. Survival rate at 1 month was 90%.53 Based on
these results, a multicentre phase II/III prospective
randomised controlled trial was initiated in which
HepatAssist therapy was compared to standard
medial therapy in patients with ALF or primary
non-function.29 One hundred and seventy-one
patients with ALF (fulminant or sub-fulminant)
or primary non-function after liver transplantation
were enrolled in this two-armed study. Eighty-six
patients received SMT, whereas 85 patients were
treated with the HepatAssist system. The survival
rate at 30 days was 71% and 62% in the entire
population (p = 0.26) and 73% and 59% in the ALF
subgroup (p = 0.12) with HepatAssist treatment
and SMT, respectively. It was thus possible to
demonstrate the safety of the system, but not
improved survival.
In another controlled study,54 10 out of 13
selected patients with ALF were treated with
HepatAssist as a bridge to transplantation. Two
patients improved spontaneously and one was
transplanted before the HepatAssist treatment
could be initiated. Ten patients received one to
three HepatAssist treatment sessions. A significant
improvement in neurological status was observed
using the Glasgow Coma Scale (6.5 (SD 3.7) and
9.6 (SD 4.4) before and after treatment, respec-
tively). Meanwhile, a significant decrease in
bilirubin and transaminase levels was observed.
HepatAssist-related complications were transient
episodes of haemodynamic instability and bleeding
in some patients. Two patients died after being
transplanted, whereas eight survived with a mean
follow-up of 18–32 months.
Due to the use of a microporous membrane,
mass transfers between the plasma and the
immobilised hepatocytes were high. No immuno-
logical response was reported despite the pore size
which should not prevent immunoglobulin trans-
fer. In a retrospective study carried out in 28
patients previously treated with the HepatAssist
system,55 absence of porcine endogenous retrovirus
(PERV) infection was observed. All patients were
negative for PERV up to 5 years after treatment
with HepatAssist as demonstrated by polymerase
chain reaction analysis of peripheral blood mono-
nuclear cells.
The Extracorporeal Liver Assist Device
The Extracorporeal Liver Assist Device (ELAD)
developed by Sussman et al56 uses 200 g of C3A
human hepatoblastoma cells located in the extra-
capillary compartment of hollow fibre cartridges
(TUs 3 and 8). Blood is first separated into plasma.
Prior to entering the cell-containing bioreactor,
plasma is directed towards activated charcoal (TU
7) and an oxygenator (TU 9) (fig 6a).
The first clinical applications have recently been
studied to demonstrate the safety of the system. In
one case study,57 a patient with fulminant liver
failure of unknown aetiology was treated with the
ELAD system. Continuous treatment for 6 days
was associated with improved neurological status
and clinical parameters. Treatment was discontin-
ued when evidence of native liver function
recovery occurred. However, the patient died from
septic shock a few days later.
The short-term safety of the ELAD system was
assessed in another clinical pilot study in which
eleven patients were enrolled.56 No safety problems
such as haemodynamic instability, complement
activation or deterioration of vital organ function

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Table 3 Biological components in bioartificial liver devices
System Cell type Cell source Cell amount
HepatAssist Porcine Cryopreserved 56109 to 76109
ELAD Human, tumour cell line C3A 200–400 g
BLSS Porcine Freshly isolated 70–120 g
MELS Porcine/human Freshly isolated Up to 600 g
AMC-BAL Porcine Freshly isolated 106109
AMC-BAL, Academic Medical Center – Bioartificial Liver; BLSS, Bioartificial Liver Support System; ELAD,
Extracorporeal Liver Assist Device; MELS, Modular Extracorporeal Liver Support.
were associated with ELAD treatment. Metabolic
support was observed in 10 patients suffering from
late-stage liver failure.
A pilot, controlled, two-armed study was per-
formed to evaluate ELAD treatment in ALF
patients with a significant chance of survival (17
patients) or fulfilling criteria for transplantation
(seven patients).30 The two groups were formed
based on predicted outcome. In each group,
patients were either treated with ELAD plus
SMT or with SMT alone. No acceleration in
platelet consumption and haemodynamic instabil-
ity were associated with ELAD treatment.
However, two patients were withdrawn from the
study because of exacerbated pre-existing dissemi-
nated intravascular coagulation or hypersensitivity
reaction. Encephalopathy grade worsened in 58%
of the control patients versus 25% of the patients
treated with ELAD, showing some benefit with
ELAD support. Survival rates for the control and
ELAD-treated patients were similar in ALF patients
with potentially recoverable lesions (75% vs 78%,
respectively). In patients who satisfied the trans-
plantation criteria, survival was higher in the ELAD
group compared to the controls (33% vs 25%,
respectively). This study did not show any
improvement in either the chemical parameters
or the clinical outcome.
Based on these results, the device was modified
and used in a clinical trial to assess safety.58 The
70 kDa cut-off membrane was replaced by a
120 kDa cut-off membrane (TU 2). Four hollow
fibre cartridges with 100 g of C3A hepatoblastoma
cells were used for each treatment. Compared to
the initial settings, the flow rate through each
cartridge was set at 500 instead of 150–200 ml/
min. Five patients with fulminant liver failure
received continuous ELAD therapy with the
modified version of the system as a bridge to and
throughout liver transplantation. They were all
successfully bridged and transplanted. Survival rate
at 30 days was 80%. No adverse events or
complications were reported. Patients’ haemody-
namic conditions did not worsen in the course of
the treatment. The cells in the bioreactors were
metabolically active throughout the treatment, as
evidenced by oxygen consumption. Larger rando-
mised multicentre trials should now therefore be
performed.
The Bioartificial Liver Support System
The Bioartificial Liver Support System (BLSS)59 60
developed by Patzer et al59 is a haemofiltration
Gut 2009;58:1690–1702. doi:10.1136/gut.2008.175380
Recent advances in clinical practice
device (TUs 3 and 8) that contains approximately
100 g of primary porcine hepatocytes within the
extracapillary compartment of a hollow fibre
bioreactor (membrane cut-off 100 kDa). Whole
blood is perfused through the fibres after being
heated through a heat exchanger and oxygenated
using an oxygenator (TU 9). Treatment with BLSS
lasts for approximately 12 h (fig 6a).
The first clinical use of the BLSS was performed
in a patient with fulminant liver failure.61 Four
patients with ALF were treated with the BLSS
device in a phase I clinical trial.60 Inclusion criteria
were ALF of any aetiology associated with ence-
phalopathy deteriorating beyond grade 2. A single
support session consisted of a 12-h extracorporeal
BLSS treatment. A second treatment session could
be given if necessary. Ammonia and total bilirubin
levels decreased by 33% and 6%, respectively,
compared with pre-treatment values. Kidney and
neurological functions did not improve signifi-
cantly during or after BLSS therapy. Transient
hypotension in one patient was the only adverse
event observed. No PERV infection could be
detected up to 1 year post-therapy.
The Academic Medical Center – Bioartificial Liver
The AMC-BAL developed by Chamuleau’s team62
consists of a plasmapheresis system (TU 1) and a
separate polycarbonate housing that contains a
three-dimensional non-woven hydrophilic poly-
ester matrix (TU 8). Ten billion (106109) primary
porcine hepatocytes are seeded within the matrix
which is wound around a polycarbonate core.
Hollow fibres are regularly distributed amid the
matrix layers for oxygen supply and CO2 removal
(TU 9). Plasma is perfused through the chamber,
between the matrix layers.
Because of legislation issues on xenotransplanta-
tion in European countries, the system is being
redesigned with a human-derived hepatocyte cell
line (fig 6b).63 However, the system has been tested
Figure 5 Principle of bioartificial livers. Nutrients and
oxygen cross the membrane from the plasma to the
hepatocyte-housing compartment to maintain cell
activities. Plasma toxins are transferred to the
hepatocytes for elimination or further transformation into
metabolites. Metabolites and synthesised substances are
returned to the plasma stream.
1697
 Recent advances in clinical practice
Table 4 Membrane types in bioartificial liver devices
Retained cells/
System Membrane type Membrane cut-off substances
HepatAssist Polysulfone 3000 kDa Blood cells
ELAD Cellulose acetate 70 kDa Above albumin
BLSS Cellulose acetate 100 kDa Above albumin/large
molecules
MELS Polyethersulfone 400 kDa Very large molecules
AMC-BAL Polysulfone Hepatocytes in direct contact – Market status of major bioartificial liver devices
with plasma
AMC-BAL, Academic Medical Center – Bioartificial Liver; BLSS, Bioartificial Liver Support System; ELAD,
Extracorporeal Liver Assist Device; MELS, Modular Extracorporeal Liver Support.
for safety and efficacy. In one case study, a patient
was successfully bridged to liver transplantation
after 35 h of treatment with the AMC-BAL.64
Biochemical and clinical parameters improved
during therapy. No severe adverse events or porcine
endogenous retrovirus activity in the patient’s
blood or blood cells were reported.
In a phase I clinical trial, the AMC-BAL was used
as a bridge to liver transplantation in patients with
ALF.65 Seven patients with grade III or IV
encephalopathy and meeting the criteria for
transplantation were treated with AMC-BAL
treatment sessions of 8–35 h. Three patients
received two additional treatment sessions with
two different BALs. Six patients were successfully
bridged to transplantation. One patient recovered
liver function after two treatment sessions and did
not need transplantation. No severe adverse events
were reported.
The Modular Extracorporeal Liver Support device
The Modular Extracorporeal Liver Support (MELS)
device was developed by Sauer et al.66 It is made of
several units: a bioreactor (TU 8), a detoxification
module performing single pass albumin dialysis
(SPAD) (TU 5) and a haemodiafiltration module
(TUs 3 and 4). The bioreactor itself contains three
bundles of hollow fibre membrane interwoven into
a three-dimensional capillary network. Two bun-
dles of polysulfone membrane with a cut-off of
400 kDa are used to perfuse the cells with the
patient’s plasma (TU 2) (fig 6a). The third set of
fibres is used for in situ oxygen supply (TU 9). The
extracapillary space is loaded with up to 500–600 g
of human hepatocytes. The cells attach to the
fibres and form aggregates. Prior to therapy, a
stand-by phase of 21 days allows the cells to adapt
to the new environment, tissue reformation and
quality controls.
In one case study, a patient with primary non-
function after transplantation was treated with
the MELS system as a bridge towards a new
transplantation.67 The bioreactor was loaded with
470 g from a discarded liver (viability 60%; 5–10%
of non-parenchymal cells) and integrated into a
circuit including continuous single pass albumin
dialysis and continuous veno-venous haemodiafil-
tration. MELS therapy was performed for 79 h.
Total plasma bilirubin levels were significantly
reduced (21.1 mg/dl at the initiation of therapy vs
1698
10.1 mg/dl after treatment). Ammonia decreased
from 100 mmol/l to 22.7 mmol/l. The patient’s
neurological status improved significantly from
coma stage IV to coma stage I. Coagulation factors
were improved. Kidney function was also recov-
ered after treatment. No adverse events were
reported.
HepatAssist is now presented as HepaMate
(HepaLife Technologies, Boston, Massachusetts,
USA). It contains 146109 cryopreserved hepato-
cytes. A new monitor (HepaDrive) is proposed by
the company. It is now in a new phase III clinical
trial in the USA.
ELAD is manufactured by Vital Therapies,
which was founded in 2003 and is based in San
Diego, California, USA, with a subsidiary in
Beijing, China. ELAD manufacturing is carried
out in San Diego in a facility that is compliant
with good manufacturing practice (GMP). Patient
enrolment has begun for a randomised, controlled,
multi-centre, phase II clinical trial that will
investigate ELAD as a treatment for patients with
ALF, with three protocols. The study is planned to
be expanded to 15 sites in the USA and Europe
within 2009.
Excorp Medical (Minneapolis, Minnesota, USA),
manufactures BLSS. This corporation is headquar-
tered and registered in the state of Minnesota,
USA. It has a Wholly Foreign Owned Entity in
JiangSu Province, China and a Wholly Foreign
Owned Subsidiary in Hong Kong, China. In
association with the University of Pittsburgh
Medical Center, patients with ALF or A-on-C LF
– regardless of underlying cause or aetiology – are
to be enrolled in the phase I/II clinical trial. The
company anticipates that the phase III efficacy
study will be conducted in six centres in the USA
and will involve up to 150 patients. Clinical trials
are also planned in China.
Hep-Art Medical Devices is a small spin-off
company of the Academic Medical Center in
Amsterdam which develops the AMC-BAL. To
comply with EU standards, several functional first-
grade human liver cell lines are currently being
developed.
How can bioartificial livers be implemented in
clinical practice?
Cell-housing devices are obliged to meet several
requirements before being used in everyday med-
ical practice. First, they must not be harmful to the
patient and must be relatively safe. Any side effects
must be minor compared to the clinical benefits.
The regulatory issues and international rules are
stated and explained in the next section.
The use of living components within extracor-
poreal assist devices results in major challenges:
preservation of sustained cell metabolism and
function, safety and applicability at the bedside
on an emergency basis are the main issues that
need to be addressed before a bioartificial liver can
Gut 2009;58:1690–1702. doi:10.1136/gut.2008.175380
 Recent advances in clinical practice

Table 5 Perfusion characteristics in bioartificial liver devices

Blood Plasma Bioreactor
filtration rate filtration rate flow rate Oxygenation
System Perfusion (ml/min) (ml/min) (ml/min) location Anticoagulation

HepatAssist Plasma 90–100 50 400 Pre-bioreactor Citrate

ELAD Blood/plasma 150–200 – 15–200 Pre-bioreactor Heparin
ultrafiltrate
BLSS Blood 100–200 – 100–250 Pre-bioreactor Heparin
MELS Plasma 150–300 31 100–200 Inside bioreactor Heparin
AMC-BAL Plasma 100 40–50 150 Inside bioreactor Heparin
AMC-BAL, Academic Medical Center – Bioartificial Liver; BLSS, Bioartificial Liver Support System; ELAD, Extracorporeal Liver Assist Device; MELS,
Modular Extracorporeal Liver Support.

reach the market. The first point brings us back to
the cell source.
Safety regarding cell preparation could be
achieved provided all operations are conducted in
accordance with good manufacturing practice
(GMP). If primary cells are to be used, special
attention must be paid with regard to the harvest-
ing and isolation procedures. Nowadays, cell banks
are able to meet such requirements.
The bioartificial device should be readily avail-
able to clinicians. This is one of the most difficult
and costly issues that has to be addressed. There
are several options: cell-containing bioreactors can
be supplied with culture medium for several days
before use or frozen and thawed right before use.
When needed, the bioreactor is removed from its
feeding circuit or thawed and included within the
extracorporeal device to support patients with liver
failure. The first solution is extremely expensive
since the bioreactors have to be fed with culture
medium and looked after. There are risks, and
careful attention would have to be paid to
maintaining the aseptic conditions throughout
the whole process. Cryopreservation may seem
the ideal solution; however, many cells may die
during freezing or/and defrosting. Cell metabolism
and detoxification capacities must remain high
enough to treat the patient.
Regulatory issues
The World Health Organization (WHO) recently
stated that a harmonised international regulatory
approach would be ideal for the successful transla-
tion of tissue engineering research to the clinic and
market place. In the USA, BALs, considered as
Tissue-Engineered Medical Products (TEMP), are
examined by the Office of Combination Products
(OCP), or by the Office of Orphans Products
(OOP). The Public Health Service (PHS) requests
proof of the safety, purity and potency of
biological products before introduction into inter-
state commerce. However, pre-market clinical
studies are allowed and include three phases: phase
I, feasibility; phase II, proper and safe dosing, and
potential efficacy; and phase III, pivotal study with
well controlled clinical trial design.68
In the EU, BALs would be treated as Advanced
Therapies Medicinal Products (ATMP). On 30
December 2008, new European legislation laid
down the rules on how ATMP are to be authorised,
supervised and monitored to ensure that they are
safe and effective.69 The legislation establishes

Figure 6 Exchange principles between the hepatocytes and plasma or blood in the (a) HepatAssist, MELS, ELAD,
BLSS and (b) AMC-BAL systems. In (a), plasma or blood is perfused through the lumen of hollow fibres whose structure
acts as a selective barrier for the hepatocytes placed in the outer compartment. In the MELS device, another hollow fibre
network is specifically added for cell oxygenation. In (b), plasma is in direct contact with the hepatocytes. The hollow
fibre membrane is dedicated to oxygen supply. AMC-BAL, Academic Medical Center – Bioartificial Liver; BLSS,
Bioartificial Liver Support System; ELAD, Extracorporeal Liver Assist Device; MELS, Modular Extracorporeal Liver
Support.

Gut 2009;58:1690–1702. doi:10.1136/gut.2008.175380 1699

 Recent advances in clinical practice
Table 6 Advantages and disadvantages of artificial and bioartificial liver support
Type of device Advantages
Artificial systems i. Relatively easy to use
ii. Limited cost for system conception and ii. Limited efficacy
patient treatment
Bioartificial systems i. All hepatic functions ensured
ii. Expected clinical results more
promising
within the European Medicines Agency a new
committee dedicated to advanced therapies. The
Committee for Advanced Therapies (CAT) plays a
central role in the scientific assessment of advanced
therapy products.
CONCLUSION
Artificial liver and bioartificial liver devices each
represents a potentially useful option for the
treatment of patients with liver failure (table 6).
Both ALs and BALs have been intensely studied
in the last few decades. ALs have proven to be a
useful option in many cases for improving bio-
chemical parameters and clinical symptoms but
the benefits in terms of patient outcome have not
yet been fully demonstrated. Detoxification alone,
as provided by an AL device, may improve the
condition of patients with liver failure if applied
early. As demonstrated with the limited success of
plasma exchange, detoxification alone may not be
sufficient in most cases. BAL devices have not yet
reached the market and are still subject to
cumbersome studies.
Some patients could be efficiently treated with
BAL systems, proving the potential for the concept.
However, their use is still very confidential. To us,
their transfer from the laboratory bench to the
patient’s bedside is hindered by three types of
obstacle: (1) scientific challenges, including the
absence of a readily available functional cell source,
the loss of cell viability and functionality before
and throughout treatment, problematic large-scale
cell culture in a dynamic setting, and the current
limitations of cryopreservation procedures (to
provide ICUs with ready-to-use devices on an
emergency basis); (2) regulatory matters; and (3)
cost issues due to inappropriate storage procedures,
complexity of use and the number of patients to
include in clinical studies before authorisation.
These important challenges delay the appear-
ance of BAL systems in ICUs and on the market.
Bioreactors combined with artificial components
nevertheless represent a pragmatic approach for
future treatment of liver failure patients and
should be investigated further. On the one hand,
special attention needs to be paid to identifying/
isolating a readily available and functional source
of cells and to improving hepatocyte entrapment.
Bioreactor configurations that are not hollow-fibre-
based should be considered to improve both large-
scale cell culture prior to therapy and mass
transfers during treatment. On the other hand,
1700
Disadvantages
i. Only detoxification
i. Cell source still under discussion
ii. Complexity of implementing living components
iii. High cost for design, operation and patients treatment
iv. Heavy logistics
regulatory issues need to be reconsidered. A better
chance to develop should be given to these new
and promising products. This could be achieved by
encouraging multi-disciplinary academic teams, as
well as small and/or leading companies, to
accompany such developments.
Acknowledgements: Certain parts of several of the figures were
produced using Servier Medical Art (www.servier.com).
Competing interests: None.
Provenance and peer review: Commissioned; externally peer
reviewed.
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2001/83/EC and Regulation (EC) No 726/2004
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 Recent advances in clinical practice
APPENDIX: BIOTRANSFORMATION FUNCTIONS OF
THE LIVER
The liver is a highly complex metabolic organ that participates in
nutrient absorption, storage and delivery according to the body’s
demands. It also plays a key role in the biotransformation,
detoxification and elimination of endogenous waste, medicines and
exogenous toxins (fig A1). Furthermore, it contributes to blood flow
control and antibacterial immune defence.
Ammonia metabolism – the main toxic waste resulting from protein
catabolism – mainly takes place in the liver. Ammonia is captured
and transformed into urea by periportal hepatocytes before being
eliminated by the kidney. Perihepatic (or centrolobular) hepatocytes,
localised close to the hepatic venules, are able to transform ammonia
into glutamine.
Bile acids – synthesised by the liver from cholesterol – are liberated
into the intestine in the bile. The liver is also able to capture bile acids
circulating within the blood after their intestinal reabsorption. A small
amount (5%) of the secreted bile acids is eliminated in the stool. This
is the main way of eliminating cholesterol from the body.
Bilirubin is the degradation product of haem, a prosthetic group
included within haemoglobin and haemoproteins such as myoglobin
and cytochromes. Haem is transformed into biliverdin and then into
bilirubin by haem oxygenase, the activity of which is predominantly in
Figure A1 The main
biotransformation and
detoxification functions of
the liver, which, in
coordination with the
kidneys and intestine, plays
a central role in the
elimination of endogenous
and exogenous toxins.
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the spleen, Kupffer cells and, to a lesser degree, hepatocytes.
Bilirubin molecules are insoluble and thus albumin-bound in the plasma.
In the liver, bilirubin is captured, conjugated with glucuronic acids to
make it soluble and eliminated in the bile. Within the intestine, bilirubin
molecules are deconjugated and hydrogenated into urobilinogen by
colon bacteria. A small amount is reabsorbed by the intestine to be
eliminated once again in the bile and, to a lesser degree, in urine. Most
of it is eliminated in the stool. The oxidation of urobilinogen to urobilin is
responsible for the coloration of stools and urine.
Xenobiotics such as exotoxins and drugs must also be eliminated.
Most hydrosoluble xenobiotics are eliminated by the kidneys. In
contrast, liposoluble toxins must be solubilised before being
eliminated. These modifications are mainly made by the liver and,
to a lesser degree, by the lungs, kidneys and intestine through two
types of reaction: phase I or oxidation reactions (super-family of P450
cytochromes or CYP) and phase II or conjugation reactions (such as
UDP-glucuronyl-transferases, for example). The former increase the
molecule’s solubility through the addition of polar groups, whereas
the latter enable the linkage of hydrosoluble endogenous molecules –
such as glucuronic acids, glutathione, sulfates and amino acids – to
oxidised molecules (simultaneously reducing their pharmacological
activity and increasing their hydrosolubility). Those molecules can
subsequently be eliminated in the bile or by the kidneys.
Gut 2009;58:1690–1702. doi:10.1136/gut.2008.175380