Gut 2003;52:300-303
© 2003 by BMJ Publishing Group &
British Society of Gastroenterology
Antidepressant induced cholestasis: hepatocellular redistribution of
multidrug resistant protein (MRP2)
P
Milkiewicz1, A P Chilton2, S G
Hubscher3 and E Elias2
1 Liver and Hepatobiliary Unit, Queen Elizabeth
Hospital, Birmingham, UK, and Department of Gastroenterology, Pomeranian Medical
School, Szczecin, Poland
2 Liver and Hepatobiliary Unit, Queen
Elizabeth Hospital, Birmingham, UK
3 Department of Pathology,
University of Birmingham, UK
Correspondence to:
Dr E Elias, Liver Unit, Queen
Elizabeth Hospital, 3rd floor, Nuffield House, Edgbaston, Birmingham B15 2TH,
UK;
elwyn.elias{at}uhb.nhs.uk
Accepted for publication
23 September 2002
 |
ABSTRACT |
Background:
We report two cases of antidepressant induced cholestasis.
Case reports: We describe the first reported case of acute
cholestasis due to citalopram (selective serotonin reuptake
inhibitor) occurring in a patient who also experienced obstetric
cholestasis in association with each of three pregnancies; in a
second patient cholestasis developed due to dothiepin (tricyclic
antidepressant), and six years later due to paroxetine. In both cases
liver biopsies showed features of a "pure" cholestasis with total
resolution within 1–6 months after withdrawal of the causative
drug. Immunostaining for the canalicular transporter, multidrug
resistant protein 2 (MRP2), responsible for biliary secretion of
several organic anions including bilirubin glucuronides, showed
sustained expression in both biopsies as well as relocalisation with
appearance of strong staining of the basolateral membrane of the
hepatocyte. This finding has also not been reported previously.
Conclusions: We postulate that intracellular redistribution
of MRP2 may reflect an adaptive compensatory mechanism which
helps in the elimination of the drug or its cholestatic metabolites
from the hepatocyte back to the sinusoidal space and subsequent
excretion in urine. Changes seen in these two patients differ
from findings previously reported in rats where downregulation
of mrp2 occurs in response to experimentally induced cholestasis.
We speculate that the rat is more advanced than humans in its
ability to downregulate canalicular transporter expression as
protection against progressive intrahepatic cholestasis.
Keywords: antidepressants; cholestasis; multidrug resistant protein;
MRP2
Abbreviations: MRP2, multidrug resistant protein 2; SSRI,
selective serotonin reuptake inhibitor; LFTs, liver function tests; ALP,
alkaline phosphatase; AST, aspartate transaminase; INR, international normalised
ratio; SBA, serum bile acids; TBS, Tris buffered saline; OC, obstetric
cholestasis
Antidepressive drugs may occasionally cause impairment of liver
function. Both tricyclic antidepressant and monoamine oxidase
inhibitors have been reported to induce prolonged1
or even fatal jaundice.2,3
Selective serotonin reuptake inhibitors (SSRI), such as paroxetine,
have been shown to induce severe acute or chronic hepatitis.4,5
The mechanisms by which antidepressive drugs induce cholestatic
changes are not known. Recent cloning of proteins involved in the
secretion of bile and its constituents has helped to provide a better
understanding of the processes involved in the pathogenesis of some
cholestatic disorders.6
However, data on expression of these transporters in drug induced
cholestasis do not exist.6
In this paper, we describe two cases of antidepressant induced
cholestasis. The first patient developed acute cholestasis during
treatment with citalopram, an SSRI, which is the first report
of such a cholestatic reaction. The second patient developed
features of cholestasis after dothiepin (tricyclic antidepressant)
and six years later he presented with similar symptoms associated
with treatment with paroxetine. Neither the occurrence of two
episodes of cholestasis following two different antidepressants
in one patient nor acute cholestasis caused by paroxetine have
been reported previously. We also looked (for the first time in
drug induced cholestasis) at the hepatocellular distribution of
multidrug resistant protein 2 (MRP2), one of the key canalicular
proteins responsible for the transport of several organic anions,
including bilirubin glucuronides, from the hepatocyte to bile.
|
PATIENTS |
Patient No 1
A
30 year old female presented initially with symptoms of obstetric
cholestasis (OC) which developed in two successive pregnancies
and responded well to treatment with ursodeoxycholic acid. In
October 1999 she was found to have clinical symptoms of depression
and was given fluoxetine. As there had been no significant
improvement in her depression over a 12 month period she was changed
to citalopram. The initial dose of 10 mg daily was increased to
20 mg after four weeks. She presented one month later with symptoms
of jaundice and pruritus. Citalopram was her only treatment at
the onset of cholestasis. Her liver function tests (LFT) at
presentation were as follows: bilirubin 75 µmol/l (normal <22
µmol/l); alkaline phosphatase (ALP) 637 U/l (normal 120–360 U/l);
aspartate transaminase (AST) 33 U/l (normal 5–35 U/l); albumin 43 g/l
(normal 35–50 g/l); and international normalised ratio (INR) 0.9
(normal 0.8–1.2). Autoantibody screening (ANA, AMA, SMA, ds DNA,
ssDNA) was negative as was viral screening for hepatitis A, B, and C.
Abdominal ultrasound was normal. A liver biopsy showed severe
canalicular cholestasis, mainly centrilobular in distribution. There
was no evidence of inflammation, bile duct loss, or any
underlying chronic liver damage. Her symptoms resolved within a
couple of months after citalopram was withdrawn. In June 2000 her
LFTs showed complete normalisation and were as follows:
bilirubin 6 µmol/l; ALP 277 U/l; AST 13 U/l; serum bile acids
(SBA) 15; albumin 46 g/l; and INR 1.0 (fig 1A
).
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Figure 1 Values for alkaline
phosphatase (ALP; normal 120–360 U/l) and bilirubin (BIL; normal
<22 µmol/l) in both patients during their episodes of
cholestasis. Pred., prednisolone.
| |
Patient No 2
A
63 year old male with depression presented in May 1991 with a three
week history of malaise, pruritus, and subsequent jaundice. Prior to
the onset of cholestasis he had been treated with dothiepin at a dose
of 75 mg daily for three months. Dothiepin was his only treatment at
that time. His LFTs at presentation were as follows: bilirubin 161
µmol/l; ALP 600 U/l; AST 40 U/l; albumin 42 g/l; and INR 1.0. SBA
were not done. Autoantibody screening (ANA, AMA, SMA, ds DNA, ssDNA)
was negative as was viral screening for hepatitis A and B. Abdominal
ultrasound was normal. Liver biopsy showed severe pure cholestasis
without inflammation, bile duct loss, or features of chronic liver
damage. Dothiepin had been stopped at presentation and his
symptoms persisted for a couple of weeks. His pruritus was treated
with cholestyramine and rifampicin with no effect. In June 1991
he was placed on a short course of oral prednisolone and his
symptoms resolved within the next four weeks. His biochemistry in
September 1991 showed complete normalisation with a bilirubin level
of 9 µmol/l, ALP 300 U/l, AST 25U/l, and albumin 46 g/l (fig
1B
).
In October 1997 he received fluoxetine and remained on it for 12
months. It did not affect his depression and therefore his local
general practitioner switched him to paroxetine at a dose of 20 mg
daily. Two months later he developed symptoms of cholestasis and
presented with jaundice and pruritus. His LFTs showed bilirubin 260
µmol/l, ALP 544 U/l, AST 36 U/l, SBA 456 (normal <15), albumin 43
g/l, and INR 1.0. As in the first episode of cholestasis,
autoantibody and viral screening were negative and abdominal
ultrasound was normal. Paroxetine was stopped and his symptoms
resolved within a couple of months. His LFTs in May 1999 were
as follows: bilirubin 7 µmol/l; ALP 255 U/l; AST 35 U/l; SBA
15; albumin 43; and INR 1.0. He was recently seen in our clinic in
May 2001 (fig 1C). His
LFTs remain entirely normal and with respect to his liver, he is
asymptomatic.
|
IMMUNOHISTOCHEMISTRY |
Methods
Formalin
fixed paraffin embedded liver sections were subjected to antigen
retrieval by microwave treatment for 30 minutes. Before the primary
antibody was applied, sections were pretreated with 3% hydrogen
peroxidase in methanol to block endogenous peroxidase activity and
with 10% normal horse serum. Tissues were then exposed to monoclonal
anti-MRP2 antibody (Alexis, UK) diluted 1:20 in Tris buffered saline
(TBS) for one hour at room temperature. After triple washing in TBS,
biotinylated antimouse secondary antibody (1:150) was applied for one
hour, followed by detection using the
avidin-biotin-immunoperoxidase method (ABC Kit, Vector, UK) for 30
minutes. To obtain a brown reaction product, the slides were exposed
to 3,3 diaminobenzidine (Vector UK Ltd) and finally a counterstain
with Mayer’s haematoxylin (Vector UK Ltd) was applied.
Results
Histologically normal liver tissue
obtained from donor liver used for transplantation was used as a
normal control. This showed diffuse canalicular immunostaining for
MRP2 with no obvious zonal variation in staining intensity (fig 2A).
Immunostaining was also present in the epithelium of bile ducts and
ductules. This was mainly luminal with focal basolateral staining
being evident.
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Figure 2 Immunohistochemical
staining reaction for multidrug resistant protein 2 (MRP2). (A)
Normal liver; (B, C) biopsy from patient No 2; (D) biopsy from
patient No 2. In normal liver, MRP2 expression was present in
canaliculi and bile duct epithelium. In the liver biopsy from
patient No 2 (B, C) there was stronger staining of MRP2 in dilated
canaliculi in acinar zones 2–3. Focal basolateral membranous
immunoreactivity was present in perivenular hepatocytes (C).
Basolateral membranous immunoreactivity was also present in
periportal hepatocytes (D, arrow).
| |
Immunostaining
for MRP2 in both patients was very similar. Diffuse canalicular
immunoreactivity was present, with stronger staining seen in dilated
canaliculi in the perivenular regions (fig 2B).
In addition, membranous staining of hepatocytes was present in
periportal and perivenular regions (fig 2C, D), a
feature not seen in normal liver. Biliary epithelial staining was
present although in patient No 2 staining appeared to be weaker
than in normal liver.
|
DISCUSSION |
Both
patients described here manifested clinical, biochemical, and
histological features of cholestasis associated with antidepressive
drugs. The second episode of cholestasis in patient No 2 and
symptoms of cholestasis in patient No 1 occurred after exposure
to two different SSRI, paroxetine and citalopram. Interestingly,
before the onset of cholestasis both patients were treated for
12 months with another SSRI, fluoxetine, without any side effects
but also without significant improvement of their depression.
Symptoms of cholestasis in both cases started suddenly approximately
two months after the introduction or incremental dose increase
of either paroxetine or citalopram. Features of cholestasis
have not yet been described in association with the above mentioned
drugs. To the best of our knowledge there is only one case report
of SSRI induced cholestasis published in the literature. In
their letter, Cosme et al described a 49 year old man in whom
symptoms of cholestasis occurred five months after introduction
of fluoxetine.7
Unfortunately, that patient had also been simultaneously treated with
several other drugs, including sulpiride, an agent which has been
described as causing cholestasis8,9
The first episode of cholestasis in patient No 2 was associated with
the tricyclic antidepressant dothiepin. His liver biopsy showed
features of a pure cholestasis, without impairment of liver
architecture. This is in contrast with previously described
cases of tricyclic antidepressant induced liver damage where
cholestasis was associated with progressive inflammatory
infiltration, fibrosis, and bile duct loss.1
The striking zonal distribution of canalicular plugging supports
the theory that in these cholestatic drug reactions biliary
bile acids are insufficient for solubilisation of biliary solutes
which form intracanalicular precipitates in lobular zones 3 and
2 but sufficient to prevent canalicular precipitates accumulating in
zone1.10
The severe degree of bile plugging seen in both our patients is not
unusual in several human cholestatic conditions, including those
caused by sepsis, drugs, and haemolysis, but not classically seen in
animal models of cholestasis. This suggests species differences in
the control and regulation of biliary transporter proteins. In the
rat, downregulation of mrp2, the transporter responsible for
canalicular secretion of bilirubin, has been shown to be an early
response to a variety of cholestatic insults in models using
endotoxin, oestradiol, and bile duct ligated rats.11
This downregulation would protect the rodent liver against the kind
of biliary plugging seen in our patients. Regulation of mrp2 and ntcp
expression in the rat occurs via RXR/RAR and HNF-1, downregulation
occurring in response to bile acid ligands as well as in response to
cytokines, and is associated with nuclear factor 
B stimulated
upregulation of Mdr1b. Our immunohistological observations provide no
evidence in support of MRP2 downregulation in our severely
cholestatic patients. The high serum bile acid level of >400 is of
an order we see in patients with complete deficiency of expression of
BSEP and suggests that biliary plugging has resulted from
maintained levels of bilirubin and other organic anion secretion
despite poor biliary bile acid output. In turn, it suggests that
human MRP2 expression is not as susceptible to downregulation as
it is in the rat, which may therefore be perceived as more
advanced in its hepatoprotective repertoire of gene expression
which confers protection against xenobiotics.
Our immunostaining study performed on both biopsies showed significant
redistribution of one of the key canalicular transporters, multidrug
resistant protein 2 (MRP2). MRP2 is a recently cloned protein
responsible for the canalicular transport of several organic
anions, including bilirubin. We have tested a wide range of
antibodies against hepatocellular transporters but it was only
MRP2 antibody which worked in paraffin embedded sections. Despite
the fact that in both biopsies analysed cholestasis was induced
by two different agents, MRP2 stainings showed striking similarities,
especially in their strong basolateral membrane reaction, not
seen in normal liver. Redistribution of MRP2 into the basolateral
membrane has been observed in vitro in HepG2 cells.12
Under physiological conditions, transport of MRP2 from the Golgi
apparatus to the canaliculus occurs via a sinusoidal membrane, as
demonstrated by Boyer and Soroka.13
One may assume that membranous staining observed by us may be
explained by cholestasis induced disturbance of the polarity of
hepatocytes. We have recently shown however that in another
cholestatic condition, primary biliary cirrhosis, MRP2 maintained its
characteristic and specifically canalicular distribution without any
detectable staining of the basolateral membrane of the hepatocyte14
which would be indicative of lost hepatocellular polarity. It seems
therefore more likely that basolateral membrane expression of MRP2
reflects an adaptive compensatory mechanism which helps in
elimination of the drug or its cholestatic metabolites from the
hepatocyte back to the sinusoidal space, and then the subsequent
excretion of the drug (metabolite) in urine. Our finding that MDR3, a
phospholipid flippase, is uncharacteristically redistributed to the
basolateral membrane of the hepatocyte in the periphery of cirrhotic
nodules in advanced primary biliary cirrhosis, in association with
upregulation of MRP3 which is able to excrete bile acids into the
blood, was interpreted by us as an adaptive response of the
cholestatic hepatocyte facilitating extrusion of bile acids and
"biliary" phospholipid into plasma.14
We can postulate that the parallel uncharacteristic basolateral
expression of MRP2 in our patients is a response of the hepatocyte to
an extreme degree of canalicular obstruction. Whether the
redistribution occurs prior to insertion of the transporter into the
canalicular membrane or subsequently by lateral diffusion from the
canalicular domain via tight junctions rendered incompetent as a
consequence of the drug toxicity per se, or secondary to plugging,
remains to be determined.
It is likely that antidepressant induced cholestasis may occur in
predisposed individuals. The observation that patient No 1 developed
OC in addition to citalopram jaundice and patient No 2 developed
cholestatic jaundice in response to two drugs of different classes
strongly supports the hypothesis that they represent genetically
predisposed individuals who are susceptible to cholestatic jaundice
which can be precipitated by a variety of stimuli. The predisposed
liver thus responds with a limited repertoire to diverse insults. We
were not able to identify predisposing factors in patient No 2.
Patient No 1 however presented a prior history of OC. In fact, she
became pregnant soon after recovery from her citalopram induced
cholestasis and again developed features of OC. OC can be associated
with malfunction of other canalicular transporters, or a mutation
causing disturbed trafficking of MDR3 to the canalicular membrane, as
recently shown by us and others.15,16
Whether a molecular defect(s) leading to OC predisposes to SSRI
induced cholestasis remains to be elucidated.
|
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