750 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastroDepartment of Liver and Gastrointestinal Diseases, Biodonostia Research Institute, Donostia… [600382]

750 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastroDepartment of Liver
and Gastrointestinal
Diseases, Biodonostia
Research Institute,
Donostia University
Hospital, University of
the Basque Country
(UPV-EHU), CIBERehd,
IKERBASQUE, Paseo
del Doctor Beguiristain,
20014 San Sebastián,
Spain ( M.J.P., L.B.,
J.M.B.). Division of
Gastroenterology and
Hepatology, Mayo Clinic
College of Medicine,
200 First Street SW,
Rochester, MN 55905,
USA ( T.V.M., N.F.L.).
Department
of Physiology and
Pharmacology,
Experimental
Hepatology and Drug
Targeting (HEVEFARM),
Biomedical Research
Institute of Salamanca
(IBSAL), Campus
Miguel de Unamuno,
University of
Salamanca, 37007
Salamanca, Spain
(J.J.M.). Department
of Gastroenterology,
“Università Politecnica
delle Marche”, Piazza
Roma 22, 60121,
Ancona, Italy ( M.M.).
Correspondence to:
J.M.B
jesus.bana les@
biodonostia.orgPolycystic liver diseases: advanced insights
into the molecular mechanisms
Maria J. Perugorria, Tatyana V. Masyuk, Jose J. Marin, Marco Marzioni, Luis Bujanda,
Nicholas F. LaRusso and Jesus M. Banales
Abstract | Polycystic liver diseases are genetic disorders characterized by progressive bile duct dilatation
and/or cyst development. The large volume of hepatic cysts causes different symptoms and complications
such as abdominal distension, local pressure with back pain, hypertension, gastro-oesophageal reflux and
dyspnea as well as bleeding, infection and rupture of the cysts. Current therapeutic strategies are based on
surgical procedures and pharmacological management, which partially prevent or ameliorate the disease.
However, as these treatments only show short-term and/or modest beneficial effects, liver transplantation
is the only definitive therapy. Therefore, interest in understanding the molecular mechanisms involved in
disease pathogenesis is increasing so that new targets for therapy can be identified. In this Review, the
genetic mechanisms underlying polycystic liver diseases and the most relevant molecular pathways of hepatic
cystogenesis are discussed. Moreover, the main clinical and preclinical studies are highlighted and future
directions in basic as well as clinical research are indicated.
Perugorria, M. J. et al. Nat. Rev. Gastroenterol. Hepatol. 11, 750–761 (2014); published online 30 September 2014;
doi:10.1038/nrgas tro.2014.155
Introduction
Polycystic liver diseases (PCLDs) are genetic disorders
characterized by bile duct dilatation and/or cyst develop –
ment derived from the bile duct epithelial cells, cholan –
giocytes. PCLDs are inherited in a dominant or recessive
form and can develop alone or in association with poly –
cystic kidney diseases (PKDs).1 The most common symp –
toms and complications of PCLDs are hypertension, back
pain, bloating and abdominal discomfort, dyspnea, gastro-
oesophageal reflux, bleeding, infection and cyst rupture.
Patients progressively worsen and surgical procedures such
as aspiration, sclerotherapy, fenestration and/or segmental
hepatic resection are commonly used in the management
of patients with PCLDs, but have short-term beneficial
effects. High rates of recurrence and complications make
liver transplantation the only curative treatment. As the
possibilities to prevent and cure PCLDs are limited, this
Review appraises the most up-to-date research into
the  underlying molecular mechanisms of PCLDs and the
i dentification of potential therapeutic targets.
Genetic mechanisms of PCLDs
The formation of multiple cysts scattered throughout
the liver alone (autosomal dominant polycystic liver
disease [ADPLD]), or in association with similar kidney
lesions (autosomal dominant polycystic kidney disease
[ADPKD]) and autosomal recessive polycystic kidney
disease (ARPKD; Caroli disease and congenial hepatic
fibrosis [CHF] in infants), is the result of germ line and/or somatic mutations (listed in Table 1, along with disease fre –
quency). Three genes, PRKCSH,2,3 SEC634 and LRP55 are
associated with the aetiology of ADPLD, whereas PKD16
and PKD27 have been identified as causative genes for
ADPKD. Mutations in PKHD1 are responsible for ARPKD
as well as Caroli disease and CHF.8,9 The genetic diagnosis
of ADPKD have revealed that mutations in PKD1 are more
frequent (~85%) than PKD2 (~15%) and are also associated
with a more severe phenotype.10 On the other hand, 20% of
patients with ADPLD show mutations in PRKCSH , SEC63
or LRP5, with PRKCSH being the most frequent (occurring
in ~15% of all ADPLD patients);11,12 however, the type of
mutation (if any) present in the remaining 80% of patients
with ADPLD remains unknown.
PKD1 encodes the mechanoreceptor polycystin -1 and
PKD2 encodes the nonselective calcium channel poly –
cystin -2, which are coupled in the ciliary membrane to
form a functional complex. Activation of polycystin -1
facilitates calcium uptake through polycystin -2,13 a large
pool of which is also present in the endoplasmic reticu –
lum.14 PRKCSH and SEC63 encode two proteins resident
in the endoplasmic reticulum. PRKCSH (also known as
hepatocystin) is the β subunit of glucosidase 2, a hetero –
dimer complex with N-linked glycan-processing activ –
ity that is involved in the maturation and/or folding of
glyco proteins.2,3 SEC63 is a component of the protein
translocation machinery required for transport of glyco –
proteins into and out of the endoplasmic reticulum
(Table 1).4 LRP5 encodes a transmembrane protein that
acts as a co-receptor with Frizzled protein members to
transduce Wnt signalling.5 PKHD1 encodes fibrocystin Competing interests
The authors declare no competing interests.
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NATURE REVIEWS | GASTROENTEROLOGY & HEPATOLOGY VOLUME 11 | DECEMBER 2014 | 751(also known as polyductin), a plasma membrane protein
localized in the primary cilium that has a role in tubulo –
genesis and maintaining the architecture of the epithelial
duct lumen (Table 1).15
To date, the Online Mendelian Inheritance in Man
(OMIM) database includes 46 mutations that affect genes
involved in the development of PCLDs (Table 2), which
are functionally and clinically important. Moreover, four
new mutations in the LRP5 gene have been described, but
are not yet included in the OMIM database. Additional
genetic variations that result in changes to protein
functions have been identified but their pathogenic
c onsequences have yet to be elucidated.
Although ADPLD and ADPKD are inherited in a domi –
nant fashion, how heterozygous mutations lead to disease
is unclear. Previous reports have indicated that mutations
in a single allele do not have severe consequences for
cholangiocyte function and that PCLD will only develop
after loss of heterozygosity.16 Indeed, PRKCSH loss of Key points
șProteins encoded by the genes that cause polycystic liver diseases are
predominantly localized in the primary cilium, plasma membrane and/or the
endoplasmic reticulum of cholangiocytes
șCurrent treatments are based on surgical procedures and/or pharmacological
management; however, their beneficial effects are modest, leaving liver
transplantation as the only definitive remedy
șElucidating the molecular mechanisms involved in the pathogenesis of these
disorders is crucial in order to identify new potential targets for therapy
șHepatic cystogenesis is characterized by ductal plate malformation,
abnormalities of the cholangiocyte primary cilium, centrosome
amplification, hyperproliferation, hypersecretion, matrix-metalloprotease
hyperactivity, angiogenesis, epigenetic alterations and atypical levels of key
intracellular mediators
șPreclinical studies have revealed new potential therapeutic targets that need
to be validated in future clinical trialsheterozygosity has been found in a high proportion of
the cysts in patients carrying a germ line mutation in this
gene,17 whereas only a small proportion of SEC63 -mutated
cysts acquire SEC63 loss of heterozygosity.16 Second-hit
mechanisms, such as loss of heterozygosity, have also been
reported in cysts of patients with ADPKD who have PKD1
germ line mutations18 and in people with PKD2 germline
mutations.19 Additionally, transheterozygous mutations in
other genes associated with PCLDs have been suggested.20
The importance of somatic second-hit mutations in the
pathogenesis of PCLDs has been reviewed elsewhere.21
Hepatic cystogenesis
Accumulating evidence suggests that pathophysio logical
alterations in ductal plate remodelling, the primary cilium
and in many intracellular signalling pathways and cel –
lular functions (proliferation, angiogenesis, secretion,
cell–matrix interaction) account for hepatic cystogenesis.
Given the multifactorial nature of these defects, their
r elative involvement in cyst growth is discussed below.
Embryology and ductal plate malformation
Hepatic cystogenesis in PCLDs is associated with ductal
plate malformation, which is a defect of ductal plate
remodelling.1,22–26 Development of the human liver starts
on the eighteenth day of gestation when the hepatic
diverticulum is formed.27 At the sixth week of gestation,
hepatoblasts immediately adjacent to the portal tract mes –
enchyme flatten and establish the ductal plate, a layer of
biliary-type cuboidal cells expressing the biliary markers
CK19 and SOX9, as well as high levels of cadherin -1.23,28
Ductal plate malformation is defined as embryological
arrest of ductal plate development29 and falls into three
categories: the inability of biliary precursor cells to differ –
entiate; defects in maturation of primitive bile ducts; and
abnormal bile duct enlargement.24,30
Table 1 | Genes, proteins and animal models of polycystic liver diseases
Mutated genes Protein Localization Function Animal models
ADPLD (~1:100,000)
PRKCSH Glucosidase 2 subunit  β,
protein kinase C substrate
80K-H or hepatocystinER N-linked glycan-processing
enzyme in the endoplasmic
reticulumPrkcshflox/flox:pCXCreER
mice and zebrafish
SEC63 Translocation protein
SEC63 homologueER Translocation of proteins in
the endoplasmic reticulumSec63flox/flox:pCXCreER
mice and zebrafish
LRP5 Low density lipoprotein
receptor-related protein 5Plasma membrane Canonical Wnt signalling Lrp5KO mouse
ADPKD (~1:500–1:1,000)
PKD1 Polycystin-1 Primary cilium, plasma
membrane and cell
junctionsMechanoreceptor involved
in calcium signallingPkd1flox/:pCxCreERTM
(Pkd1cKO ) and zebrafish
PKD2 Polycystin-2 Primary cilium and
endoplasmic reticulumNonselective calcium
channelPkd2flox/:pCxCreERTM
(Pkd2cKO ) and Pkd2WS25/–
ARPKD, CHF or CD (~1:20,000)
PKHD1 Fibrocystin or polyductin Primary cilium Tubulogenesis and/or
maintenance of bile
duct architecturePCK rat, and Pkhd1del2/del2
mouse
Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; ADPLD, autosomal dominant polycystic liver disease; ARPKD, autosomal recessive
polycystic kidney disease; CD, Caroli disease; CHF, congenital hepatic fibrosis; ER, endoplasmic reticulum.
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752 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastroARPKD and CHF are both characterized by the pres –
ence of ductal plate remnants along the portal tract
margins. In patients with ARPKD, liver histology depicts
incompletely developed ductal plates,31 whereas in CHF
bile ducts are embedded in fibrous stroma and form
cystic lesions (called von Meyenburg complexes).29 Both
ADPKD and ADPLD are associated with ductal plate
malformation; however, is not clear to which category
of malformatio n they belong.24Hepatoblast commitment to biliary lineage and ductal
plate remodelling is controlled by a network of the follow –
ing: signalling molecules including Notch,32–34 transforming
growth factor β,35,36 Wnt37–39 and fibroblast growth factor;40–42
transcription factors;43–50 and microRNAs51 (Box 1). Many
components of ductal plate remodelling have also been
implicated in the regulation of ciliary function and are
aberrantly expressed in cystic cholangiocytes, thus linking
ductal plate malformation and hepatic cystogenesis (Box 1).Table 2 | Mutations in genes causing polycystic liver diseases resulting in change of function with clinical significance
Gene Locus Nucleotide change Amino acid change OMIM number Phenotype MIM number
PRKCSH 19p13.2 IVS16, A -G, –2
IVS4, G -C, +1
2-bp del, IVS16GT , +1
c.1240C>T
c.1269C>G
1-bp ins, 216A–
–
–
p.Gln414Ter
p.Tyr423Ter
–177060.0001
177060.0002
177060.0003
177060.0004
177060.0005
177060.0006174050
SEC63 6q21 c.173G>A
1-bp ins, 442A
IVS8ds, G -A, +1
c.1702_1704delp.Trp58Ter
–
–
p.Glu568del608648.0001
608648.0002
608648.0003
608648.0004174050
LRP5 11q13.2 c.1360G>A
c.3562C>T
c.4587G>C
c.4651G>Ap.Val454Met
p.Arg1188Trp
p.Arg1529Ser
p.Asp1551AsnNot assigned
Not assigned
Not assigned
Not assignedNot assigned
PKD1 16p13.3 IVSds, G -C, +1
c.12124C>T
15-bp del
c.12682C>T
c.11512C>T
c.12261T>A
c.11457C>A
12036G -A
28-bp del, nt6434
IVS14, G -A, –1
c.971G>T
c.2534T>C
c.5764C>T
2-bp del, 5224AG
c.12420G>A
Gly2579del, 8 -bp del–
p.Gln4042Ter
–
p.Arg4228Ter
p.Gln3838Ter
p.Cys4087Ter
p.Tyr3819Ter
–
–
–
p.Arg324Leu
p.Leu845Ser
p.Gln1922Ter
–
p.Trp4140Ter
–601313.0001
601313.0002
601313.0003
601313.0004
601313.0005
601313.0006
601313.0007
601313.0008
601313.0009
601313.00010
601313.00011
601313.00012
601313.00013
601313.00014
601313.00015
601313.00016173900
PKD2 4q22.1 c.1139G>A
c.2224C>T
c.1213C>T
1-bp ins, 693C
c.1390C>T
1-bp ins, 2160A
1-bp ins, 197 -203C
c.1532A>T
2-bp del/1 -bp ins, nt1934
Ex3dup
c.305_306insGAGp.Trp380Ter
p.Arg742Ter
p.Gln405Ter
–
p.Arg464Ter
–
–
p.Asp511Val
–
–
p.Glu102_Val103insArg173910.0001
173910.0002
173910.0003
173910.0004
173910.0005
173910.0006
173910.0007
173910.0008
173910.0009
173910.00010
173910.00011613095
PKHD1 6p12.2 c.107C>T
c.4991C>T
c.9053C>T
c.5221G>A
c.8011C>T
c.10658T>C
c.1486C>T
c.10412T>G
IVS46ds, A -G, +653p.Thr36Met
p.Ser1664Phe
p.Ser3018Phe
p.Val1741Met
p.Arg2671Ter
p.Ile3553Thr
p.Arg496Ter
p.Val3471Gly
–606702.0001
606702.0002
606702.0003
606702.0004
606702.0005
606702.0006
606702.0007
606702.0008
606702.0009263200
Data were obtained in June 2014 from the Online Mendelian Inheritance in Man (OMIM) of the National Center for Biotechnology Information (NCBI). Four new
mutations in LRP5 have been described but the OMIM number and the phenotype (MIM number) have not yet been assigned. The mutations listed consist of
single nucleotide polymorphisms resulting in an amino acid change, exon duplications, small deletions and insertions, and splice site mutations. Abbreviations:
bp, base pair; del, small deletion; ds, donor splice site; ER, endoplasmic reticulum; Ex3dup, exon 3 duplication; ins, insertion; IVS, intervening sequence.
Permission obtained from the Online Mendelian Inheritance in Man, OMIM® . McKusick-Nathans Institute of Genetic Medicine, John Hopkins University
(Baltimore, MD), September 2014. World Wide Web URL: http://omim.org/ .
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NATURE REVIEWS | GASTROENTEROLOGY & HEPATOLOGY VOLUME 11 | DECEMBER 2014 | 753Cholangiocyte abnormalities
Primary cilium and centrosomes
Cholangiocytes, the epithelial cells that line the bile ducts,
are key liver cells involved in the regulation of the flow
and composition of bile. Cholangiocytes contain a single
primary cilium that extends from the apical membrane
into the bile duct lumen, which is formed by an axoneme
and a centriole-derived basal body.52 The primary cilium,
an antenna-like bulge, functions as a sensory organelle
detecting changes in bile flow, composition and osmolar –
ity and has an important role in cholangiocyte physiology
and pathophysiology (Figure 1a).52,53 Hepatic cystogenesis
is thought to be associated with disturbances in ciliary sen –
sation that result from structural and functional changes
owing to aberrant expression of PCLD-related and c iliary-
associated proteins. Indeed, shortened, u nusually long or
entirely absent cilia are present in the cystic cholangio –
cytes of animal models with PCLD47,54–56 and patients
with ADPKD (Figure 1b).57 Abnormalities in the primary
cilium are accompanied by atypical centrosome position –
ing, supernumerary centrosomes and multipolar spin –
dles.48,49,58 Notably, cholangiocytes with hyperamplified
centrosomes represent a small but notably abnormal
portion of cells lining liver cysts. In this regard, a link
between centrosome amplification and renal cystogenesis
was reported, emphasizing the importance of centrosome
abnormality in hepatorenal cystogenesis.59
The absence of PCLD-related proteins (fibro –
cystin,54,55,60 polycystin -1 and polycystin -253,61,62) and
cAMP-associated G-protein-coupled receptor TGR5,63
or the overexpression of the calcium channel TRPV4,64,65 causes functional abnormalities in cholangiocyte cilia and
enhances proliferation and fluid secretion, which con –
tributes to progressive cyst growth.1,25,66–68 Interestingly,
a strong piece of evidence that might implicate cilia in
the pathogenesis of cyst growth comes from outstand –
ing research carried out in animal models of PCLDs.
Polycystin -1, polycystin -2, fibrocystin, hepatocystin and
SEC63 seem to interact together in a complex network;
hepatocystin and SEC63 are necessary for the adequate
expression of polycystin -1 and polycystin -2 functional
complex, and importantly, polycystin -1 was defined as
the rate-limiting component that determines cyst forma –
tion.69 Thus, polycystin -1 expression levels seem to be
involved in determining the severity of ADPKD, ARPKD
and ADPLD phenotypes, providing a direct link between
cyst growth and the primary cilium.70 In addition, the use
of proteasome inhibitors has been suggested as a therapy,
which might inhibit cystogenesis in patients with ADPLD
by increasing the steady-state levels of polycystin -1 and
also by inducing apoptosis due to increased toxic levels
of unfolded proteins in cyst-lining cells.70 On the other
hand, studies carried out in experimental animal models
of ADPKD suggest that, in the absence of polycystin -1
or polycystin -2, signalling mechanism present in the
remaining cilium are required to promote cyst growth,
whereas total absence of the cilium results in inhibition
of cystogenesis.71 Evidence supporting the concept that
ciliary dysfunctions underlie hepatic cystogenesis is
s ummarized in Box 2.
Proliferation and angiogenesis
Normal cholangiocytes are quiescent with almost no
expression of proliferating cell nuclear antigen (PCNA)
detectable by immunohistochemistry,72 whereas cystic
cholangiocytes show intense PCNA staining.72–75 How –
ever, proliferation of cystic cholangiocytes in patients with
ADPLD is insignificant, with few Ki67 positive cells,76
which suggests that other events such as secretion, cell–
matrix interactions and ductal plate malformations could
have a major role in ADPLD pathogenesis. The hyper –
proliferative phenotype of cystic cholangiocytes is regulated
by growth factors and hormones present in the cystic fluid
and/or secreted by cystic cholangiocytes (Figure 1c).72,77–79
The growth factors epidermal growth factor (EGF),
vascular endothelial growth factor (VEGF) and insulin-
like growth factor 1 (IGF1) all participate in autocrine
and/or paracrine loops that stimulate cellular prolifera –
tion. Cholangiocytes from the PCK rat, an animal model
of ARPKD, show more pronounced hyperproliferative
features in response to Egf than normal rat cholangio –
cytes.80 Egf-stimulated hyperproliferation was linked to
an overexpression of mitogen-activated protein kinase
kinase 5 (Map2k5 also known as Mek5) and subsequent
phosphorylation of Erk5, which was abolished by molec –
ular or pharmacological targeting of either Mek5 or the
Egf receptor with gefitinib.80 However, the role of EGF in
hepatic cystogenesis is uncertain because a different study
demonstrated that chronic administration of EGF receptor
inhibitors (EKI -785 or EKB -569) had no effect on hepatic
cystogenesis in PCK rats.81Box 1 | Regulators of ductal plate remodelling
Signaling pathways
șNotch receptors and Notch-processing enzymes are expressed in primary cilia
and regulate their length.32–34
șTGF- β signalling also regulates biliary commitment of hepatoblasts.23 TGF- β and its
receptor are overexpressed in cystic cholangiocytes and renal epithelial cells.35,36
șWnt signalling is implicated in both ciliary sensation and cystogenesis.37–39
șFGF signalling regulates cilia length and hepatoblast differentiation towards the
biliary lineage. Patients with ADPKD have raised FGF23 levels.40–42
Transcription factors23,24
șHNF1 β, 4 and 6
șHHEX
șC/EBP α
șSmad2 and Smad3, Onecut 2
șFOXM1b
șSALL4
șTBX3
Examples include deletion of Hnf6 and Hhex in mice results in accumulation
of ductal plate remnants, cyst development and cilia absence in cystic
cholangiocytes.23,43,44 In addition, in the pancreas, Hnf6 stimulates the expression
of Pkhd1 and Cys1 genes, mutations of which cause hepatic cystogenesis and
affect ciliary length.45 In a further example, mice deficient in Cys1,46,47Hnf‑1 β48,49
or C/EBP α50 exhibit ductal plate malformation.
miRNAs
miRNA -30 family. miR -30a depletion in zebrafish affects bile duct morphogenesis.51
Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; C/EBP α, CCAAT/
enhancer binding protein α; FGF, fibroblast growth factor; FOXM1b, forkhead box factor M1b;
HHEX, hematopoietically expressed homeobox; HNF, hepatocyte nuclear factor; miRNA,
microRNA; SALL4, spalt-like transcription factor 4; TBX3, T-box transcription factor 3; TGF,
transforming growth factor.
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754 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastroVEGF and angiopoietin 1 are pleiotropic growth factors
that have key roles in hepatic cytogenesis promoting
cyst growth and their vascular supply. The expression of
VEGF and angiopoietin 1 and their respective receptors
(that is, VEGFR1, VEGFR2 and TIE2) are upregulated
in cholangio cytes of patients with ADPKD.78 In addi –
tion, VEGF is found in liver cyst fluids of patients with
ADPKD.82 VEGF stimulates proliferation of cystic chol –
angiocytes in patients with ADPKD, in the Pkd2WS25/–
and Pkd2flox/:pCxCreER (Pkd2cKO) mouse models of
ADPKD74,78,82 and promotes liver cyst growth in Pkd2cKO
mice but not in Pkd1flox/:pCxCreER (Pkd1cKO) mice.74 The
effects of VEGF secretion and VEGFR2 signalling are
dependent on protein kinase A (PKA) and/or extracellu –
lar signal regulated kinase 1/2 (ERK1/2).74 In addition, the
effect of VEGF is synergized by angiopoietin 1.78 Biliary
cysts are surrounded by vascular networks that are critical
for hepatic cystogenesis. Different intracellular signalling
pathways and growth factors present in the cystic fluid,
such as VEGF and IL-8, promote proliferation of endothe –
lial cells (Figure 1c),83 which suggests that autocrine and
paracrine mechanisms could be involved in neovasculari –
zation.78 The potential therapeutic value of targeting VEGF
in patients with PCLDs is supported by the fact that the
VEGFR2 inhibitor, SU5416, blunts liver cyst growth in
animal models.74,82
Another potential therapeutic target is IGF1, a promitotic
factor present in the cystic fluid of patients with ADPKD.72
IGF1, its main receptor IGF1R and their downstream
effectors phosphorylated AKT and phosphorylated mam –
malian target of rapamycin (mTOR) are all overexpressed
in the cystic epithelium.72 Moreover, the proliferation of
cholangio cytes stimulated by cystic fluid, 17β-estradiol or
IGF1 was inhibited by an IGF1R antagonist.72
As well as being downstream of IGF1 signalling, AKT
is an activator of mTOR, through which it regulates the
expression of hypoxia inducible factor 1 α (encoded by
HIF1α ), a major transcriptional activator of VEGF . As
mTOR is involved in both VEGF and IGF1 signalling
pathways it was thought that mTOR could be a useful therapeutic target for treating patients with PCLD. The
expression of phosphorylated mTOR is increased in
the liver cystic epithelium of Pkd2cKO mice;73 inhibiting
mTOR in vivo with sirolimus decreased IGF1-stimulated
HIF1α accumulation, VEGF secretion in cystic cholangio –
cytes and cyst growth.73 Accordingly, sirolimus and SU5416
(VEGFR2 inhibitor) inhibited the proliferation of cholan –
giocytes stimulated by IGF1 in Pkd2cKO mice.73 However,
the inhib ition of mTOR by sirolimus did not attenuate
hepatic and renal cystogenesis in PCK rats,84 highlight –
ing the necessity to clarify in clinical trials the potential
therapeutic role of mTOR inhibitors in the different forms
of PCLDs. Interestingly, a reduction in liver volume was
observed in a clinical trial using sirolimus as an immuno –
suppressant after renal transplantation in 16 patients with
ADPKD (Table 3).85 However, in two independent clini –
cal trials, chronic everolimus (an mTOR inhibitor derived
from sirolimus) treatment in patients with ADPKD did
not slow the progression of renal impairment and kidney
growth.86,87 Unfortunately, effects of everolimus on liver
volume were not examined in these trials. In a 2013
clinical trial, the efficacy of combining everolimus and
octreotide (a somatostatin analogue) was compared with
octreotide monotherapy. Everolimus did not enhance the
beneficial effect of octreotide in reducing liver volume in
patients with PCLDs (Table 3).88 Overall, the role of mTOR
inhibitors in the treatment of patients with PCLDs has not
f ulfilled expectations and is not clinically recommended.
The potential therapeutic value of sorafenib, a tyrosine
kinase inhibitor that might block the action of all EGF,
VEGF and IGF1 receptors, has been tested for its ability
to attenuate the proliferation of cystic cholangiocytes.
However, evidence indicates that, paradoxically, sorafenib
transactivates Raf1 thereby promoting cystogenesis in
Pkd2cKO mice.89
Peroxisome proliferator-activated receptor γ (PPAR -γ)
is also involved in hepatic cystogenesis by inhibiting genes
involved in proliferation, inflammation and fibrosis. The
PPAR -γ agonist, pioglitazone, inhibited kidney and liver
disease in PCK rats,90 but its use in clinical practice is Figure 1 | Cellular alterations and molecular mechanisms involved in hepatic cystogenesis. a | The normal structure of both
a cholangiocyte and a primary cilium. b | The primary cilium and gene expression levels of key intracellular mediators can
be altered in polycystic liver diseases. c | Different growth factors and cytokines stimulate the proliferation of cystic
cholangiocytes and endothelial cells in an autocrine and/or paracrine fashion. Moreover, d | oestrogens and e | changes in
intracellular calcium and cAMP levels might also induce the proliferation of cystic cholangiocytes. Hepatic cystogenesis is
associated with f | alterations in fluid secretion and g | extracellular matrix remodelling. h | Global downregulation of
microRNAs occurs in cystic cholangiocytes, which facilitates the proliferation of cystic cholangiocytes. The relative
involvement of each pathway in different forms of hepatic cystogenesis has been highlighted in the main body text. The
central image is of human liver tissue with cysts. Abbreviations: 17 βES, 17 β oestradiol; AC, adenylate cyclase; AE2, anion
exchanger 2; ANG-1, angiopoetin-1; AQP1, aquaporin 1; cAMP , cyclic adenosine monophosphate; Cdc25A, cell division cycle
25A; Cdks, cyclin dependent kinase; CFTR, cystic fibrosis transmembrane conductance regulator; EPAC, rap guanine
nucleotide exchange factor 3; ER, oestrogen receptor; ER α, oestrogen receptor α; ER β, oestrogen receptor β; ERK1/2,
extracellular signal regulated kinase 1/2; Gs, Gs protein; HIF1 α, hypoxia inducible factor 1 α; IGF1, insulin-like growth factor
1; MEK, mitogen-activated protein kinase kinase 1; miR-15a, microRNA 15a; MMP , matrix metalloproteinase; mTOR,
mammalian target of rapamycin; pAKT, phosphorylated v-akt murine thymoma viral oncogene homolog 1; PC-1, polycystin-1;
PC-2, polycystin-2; PI3K, phosphatidylinositol 4,5-bisphosphate 3-kinase; PKA, protein kinase A; pmTOR, phosphorylated
mTOR; pVEGFR2, phosphorylated vascular endothelial growth factor receptor 2; SEC63, SEC63 homolog; SR, serotonin
receptor; TGR5, g-protein coupled bile acid receptor-1; TIE-2, TEK tyrosine kinase; TRPV4, transient receptor potential
cation subfamily V member 4; VEGF, vascular endothelial growth factor. Permission obtained from BMJ Publishing Group
Ltd. © Urribarri, A. D. et al. Gut http://dx.doi.org/10.1136 /gutjnl-2013-305281 . ◀
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NATURE REVIEWS | GASTROENTEROLOGY & HEPATOLOGY VOLUME 11 | DECEMBER 2014 | 755HIF1 α VEGF
ProliferationCyclinspAKTPI3K
ERK1/2MEKIGF-1ANG-1
TIE-2VEGF
pmTORVEGF
VEGF
ProliferationpVEGFR2VEGFR2VEGF
IL-8RIL-8Endothelial
cellLiver cyst
/f_luid
Aberrantly expressed proteins
TRPV4, mTOR, IGF-1, VEGF,
Cdc25A, Cdks, TGR5 etc.Malformed,
shortened
unusually long
or entirely
absent cilla
Atypical centrosome
positioning, supernumerary
centrosomes, multipolar
spindles and extra cillaa
Normal cholangiocyteTGR5
Ca2+PC-2PC-1
NucleusFibrocystin
TRPV4H2O Cl–
AE2
Cl–HCO2–AQP1CFTRApical
membrane
HepatocystinSEC63
Endoplasmic
reticulumPC-2Ca2+
ProliferationpAKTPI3KIGF-1
pmTORERαERβ17βES
Ca2+
ProliferationCyclinsCa2+
ERK1/2MEKPKA/EPACcAMP
H2OCl–
AE2
Cl–HCO2–
AE2Enhanced
/f_luid
secretion
cAMP
SecretinPKA
SR ACGSER
Extracellular
matrix
degradation
MMPsProliferationCyclinsIL-8
17βESIL-6IL-6IL-8
MMPsCdc25AmiR-15a
Basolateral expressionIL-6Rhc
d e
fb
g
discouraged because of the increased risk of bladder cancer
associated with this drug.90,91 However, telmisartan—an
angiotensin II type 1 receptor antagonist that has off-target
PPAR -γ agonist properties but fewer associated risks—
reduced liver cystogenesis in PCK rats,92 which suggests that
PPAR -γ might be a potential therapeutic target in PCLDs.
Although PCLDs affect both men and women, the
latter usually exhibit a more severe phenotype. A number of clinical observations suggest that oestrogens are key
regulators of hepatic cystogenesis.93–95 Cholangiocytes
that line bile ducts of healthy individuals and unaffected
bile ducts of patients with ADPKD do not express the
oestro gen receptors α or β.72 By contrast, cystic cholangio –
cytes in patients with ADPKD are positive for oestrogen
receptors α and β. Oestrogens exert promitotic effects on
cystic cholangiocytes either directly or by inducing IGF1
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756 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastrosecretion (Figure 1d);96 these effects can be partially inhib –
ited by oestrogen receptor antagonists.72 All these data
support the rationale for studying the potential therapeutic
value of antioestrogen therapies (such as t amoxifen) for
the t reatment of female patients with PCLDs.
cAMP and calcium levels
Intracellular signalling abnormalities associated with
increased cAMP levels and decreased [Ca2+]i levels
underlie the hyperproliferative phenotype of cystic chol –
angiocytes and represent potential therapeutic targets
(Figure 1e). Raised cAMP levels in PCK rat cholangio –
cytes97 stimulates proliferation via two intracellular effec –
tors involved in the Mek/Erk pathway—rap guanine
nucleotide exchange factor 3 (Rapgef3, also known as
Epac) and Pka.98 Octreotide treatment decreased these
raised cAMP levels in the cholangiocytes and serum of
PCK rats, reducing liver weight, cyst volume, hepatic
fibrosis and mitotic indices.97 A study has shown that
pasireotide, a more potent somatostatin analogue than
octreotide with broader receptor specificity and a longer
half-life, is more effective than octreotide in reduc –
ing hepatorenal cystogenesis in PCLD rodent models.99
Several clinical trials have evaluated the effects of octreo –
tide100–102 and lanreotide103–105 in patients with PCLDs
(Table 3). Both drugs moderately decreased liver volume (by ~5%) in patients with ADPLD and ADPKD, and
improved quality of life.61,101,106 In addition, a clinical trial
(NCT01670110107) to evaluate the therapeutic potential of
pasireotide in patients with ADPKD and ADPLD is now
underway at the Mayo Clinic. For these reasons, current
pharmacological strategies are based on the chronic
administration of somatostatin analogues, which are
clinically recommended but might result in gastrointesti –
nal adverse effects such as diarrhoea, abdominal cramps,
flatulence, bloating, gas and injection site granulomas.101,105
Cholangiocytes from both PCK rats and Pkd2cKO mice
are also characterized by diminished levels of [Ca2+]i.98,108
Restoration of intracellular calcium by a calcium iono –
phore inhibited both the basal and cAMP/Pka-stimulated
proliferation of PCK rat cholangiocytes via the Pi3k/
Akt pathway, whereas cAMP/Epac-stimulated prolifera –
tion was not affected.98 Thus, cAMP might regulate the
proliferation of cystic cholangiocytes through calcium-
dependent (Pka) and calcium-independent (Epac) mech –
anisms.98 Restoration of intracellular calcium could be
a useful therapeutic approach in treating patients with
PCLD. Pharmacological activation of the calcium-entry
channel Trpv4 (which is overexpressed in PCLDs) inhib –
ited the proliferation of PCK rat cholangiocytes in vitro ,
but did not affect hepatic cystogenesis in vivo .65 The
in vivo study was limited by the low sublethal dose of the
Trvp4 activator used (GSK1016790A), as it induces acute
circulatory collapse when administered at higher doses.
Therefore, we believe that new pharmacological strategies
are needed to evaluate in vivo the potential therapeutic role
of intracellular calcium restoration in PCLDs.
Secretion
Cholangioctes have a key role in the fluidization and
alkalinization of the primary bile generated by hepato –
cytes, which are controlled by nucleotides, bile acids and
hormones, such as secretin.109–110 Secretin interacts with
its receptor (localized to the basolateral membrane of
cholangiocytes), which leads to increased cAMP levels
and further activation of PKA. PKA then mediates the
exocitosis of intracellular vesicles containing the chloride
channel, cystic fibrosis transmembrane conductance regu –
lator (CFTR), the chloride/bicarbonate exchanger, anion
exchanger 2 (AE2) and the water channel, aquaporin 1
(AQ1), which results in bicarbonate-rich choleresis.109–111
Increased fluid secretion is one of the contributing
mechanisms of bile duct dilatation and cyst expansion in
PCLDs (Figure 1f). PCK rat cholangiocytes cultured in a
3D collagen matrix showed enhanced expansion under
basal conditions in response to secretin and hypo tonicity.
Observed alterations were associated with abnormal
expression and location of Cftr, anion exchanger 2 and
aquaporin 1.68 These carriers are preferentially localized
to the apical membrane of normal rat cholangiocytes, but
they were mainly found overexpressed and mislozalized
at the basolateral membrane of PCK rat cholangiocytes.
The basolateral presence of Cftr or anion exchanger 2
inhibitors blocked secretin-stimulated hypersecretion in
cysts of PCK rats but not in normal cystic structures,68
which suggests that targeting these aberrant mechanisms Box 2 | Ciliary dysfunction underlies hepatic cystogenesis
șDisappearance of fibrocystin (PKHD1) from cholangiocyte cilia leads to ciliary
malformations and accelerated cyst expansion.54,55 ,60
șSomatic inactivation of Pkd2 in mice results in hepato-cystic phenotype.56
șPolycystin -1 and polycystin -2 function as ciliary sensors of cell injury activating
the cholangiocyte proliferation as a reparative mechanism.62
șCiliary structural malformations (shortened, unusually long or entirely absent
cilia) are present in cystic cholangiocytes of PCLD animal models47,54 –56 and
patients with ADPKD.57
șCiliary defects are linked to basal body abnormalities (that is atypical
centrosome positioning, supernumerary centrosomes, multipolar spindles
and extra cilia).58
șHnf1 β deficient mice lack cholangiocyte cilia due to mispositioning of the
basal body resulting in ductal plate malformation and biliary dysgenesis that
resembles the lesions observed in patients with ARPKD.30,49 ,50
șAbnormal ciliary structure is associated with enhanced fluid secretion and
ion transport.62,66 –68
șAbsence of polycystin -1 in cholangiocyte cilia decreases levels of [Ca2+]i and
increases cAMP production via a Ca2+-inhibitable adenylyl cyclase 6 (which is
also localized within the cilium) increasing cell proliferation and accelerating
fluid secretion.53,61
șTGR5, the G -protein coupled receptor linked to cAMP signalling, is overexpressed
in cystic cholangiocytes and is mislocalized from cilia.63
șDecreased levels of [Ca2+]i in hepatic cysts are linked to aberrantly expressed
calcium channel TRPV4 in cholangiocyte cilia.65
șGlucosidase 2 subunit  β and Sec63 are required in mice for adequate
expression of a functional complex of polycystin -1 and polycystin -2. Polycystin -1
is the rate-limiting component of this complex.69,70
șIn the absence of polycystin-1 or polycystin-2, the resulting cilium is required
to promote cyst growth, whereas the total absence of the cilium results in
inhibition of cystogenesis.71
Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; ARPKD, autosomal
recessive polycystic kidney disease; cAMP , cyclic adenosine monophosphate; Hnf1 β,
hepatocyte nuclear factor 1 β; PCLD, polycystic liver disease; PKDH1, polycystic kidney and
hepatic disease 1; Sec63, Sec63 homolog; TGR5, g-protein coupled bile acid receptor-1;
TRPV4, transient receptor potential cation subfamily V member 4.
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NATURE REVIEWS | GASTROENTEROLOGY & HEPATOLOGY VOLUME 11 | DECEMBER 2014 | 757Table 3 | Clinical trials in polycystic liver diseases
Study Patient randomization Treatment Liver volume reduction
Dose Duration Route of
administration
Octreotide (targets cAMP)
Caroli et al.
(2010)100ADPKD, n = 12
(5 treated, 7 placebo)40 mg every
28 days6 months Intramuscular 4.5% (vs 0.9% increase in placebo)
Hogan et al.
(2010)101ADPKD and ADPLD, n = 42
(28 treated, 14 placebo)40 mg every
28 (± 5) days12 months Intramuscular 4.95% (vs 0.92% increase in placebo)
Hogan et al.
(2012)102Extension study:
ADPKD and ADPLD, n = 41/42
(all received treatment)40 mg every
28 (± 5) daysAdditional
12 months
(24 months total)Intramuscular No significant changes (–0.77%) after an
additional year of therapy
Lanreotide (targets cAMP)
Temmerman
et al.
(2013)105ADPKD and ADPLD, n = 132
(106 treated, 26 placebo)90 mg (n = 55)
Or 120 mg (n = 51)
every 28 days6 months Subcutaneous 1.4% (LAN 90 mg)
2.8% (LAN 120 mg) (vs 1.1% increase in placebo)
LAN 90 mg had fewer adverse effects
van Keimpema
et al. (2009)103ADPKD and ADPLD, n = 54
(28 treated, 27 placebo)120 mg every
28 days24 weeks Subcutaneous 2.9% (vs 1.6% increase in placebo)
Chrispijn et al.
(2012)104Extension study:
ADPKD and ADPLD, n = 41/54
(all received treatment)120 mg every
28 days12 months Subcutaneous 4%
Sirolimus vs tacrolimus (targets mTOR)
Qian et al.
(2008)85ADPKD (sirolimus n = 7,
tacrolimus n = 9)5–10 mg daily
(sirolimus)
3 mg twice day
(tacrolimus)Retrospective
analysis
19.4 monthsOral 11.9% decrease with sirolimus vs 14.1%
increase with tacrolimus
Everolimus alone or in combination with octreotide (targets cAMP and mTOR)
Chrispijn et al.
(2013)88ADPKD and ADPLD,
n = 44 (23 received
octreotide and 21 received
octreotide + everolimus)40 mg octreotide
every 4 weeks,
2.5 mg
everolimus daily48 weeks Intramuscular
(octreotide)
and oral
(everolimus)3.5 ± 5.2% octreotide monotherapy vs
3.8 ± 4.7% in the octreotide/everolimus group
(in response to octreotide, everolimus does
not further reduce liver volume)
Clinical trial NCT01670110 testing pasireotide in severe polycystic liver disease is underway (Mayo Clinic).107 Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; ADPLD,
autosomal dominant polycystic liver disease; cAMP , cyclic adenosine monophosphate; mTOR, mammalian target of rapamycin.
could have potential therapeutic value in treating patients
with PCLDs.
However, the role of secretin and flux proteins in cyst
progression is more complex than expected. In contrast
to the aforementioned data, human cholangiocytes from
patients with ADPKD exhibit decreased anion exchanger
activity due to diminished expression of mature glyco –
sylated anion exchanger 2 polypeptide and decreased
membrane-localized anion exchanger 2.112 The secre –
tory differences between PCK rat cholangiocytes and
cholangio cytes from patients with ADPKD could be
linked to differences related to the particular form of
PCLD. Thus, as cysts from PCK rats flow into the bile
ducts, cysts in patients with ADPKD might be discon –
nected from the biliary tree,55 which could result in differ –
ent pathological or adaptive secretory processes. Of note,
chronic administration of secretin had negligible effects on
hepatic cystogenesis of PCK rats and Pkd2WS25/– mice and
the absence of the secretin receptor in Pkd2WS25/–:SCTR−/−
double mutant mice did not alter the severity of PCLD.113
Together, these data suggest that secretin could have a
modest role in the pathogenesis of PCLDs.
Cell–matrix interactions
Interactions between cells and the extracellular matrix are
involved in normal ductal plate formation but also in the development and progression of PCLDs. The extracellu –
lar matrix is a complex structure of glycoproteins and is
remodelled by matrix metalloproteases, tissue inhibitors of
metalloproteases and hormones. Free space for cyst growth
is generated by overexpression and hyper secretion of
matrix metalloproteases by cholangiocytes, which is regu –
lated by oestrogens and cytokines in an autocrine and para –
crine fashion (Figure 1g).114 IL-6 and IL-8 present in the
cystic fluid interact with their plasma membrane receptors
(IL-6R and CXCR1, respectively) localized in cholangio –
cytes, which enhances the expression and secretion of
matrix metalloproteases. Oestrogens, present in the cystic
fluid of female patients, have the same effect.114 By contrast,
other factors present in the cystic fluid such as VEGF, EGF,
hepatocyte growth factor, epithelial neutrophil attract –
ant 78 and growth-related oncogene α did not alter the
matrix metalloprotease activity of normal cholangiocytes
or cholangiocytes from patients with ADPKD.114 Altered
cell–extracellular matrix inter actions detailed above are
in agreement with the observation that the expression of
basement membrane proteins such as laminin and colla –
gen type IV around bile ducts is degraded in CHF, Caroli
disease and in PCK rats.115 Finally, the importance of
matrix metalloprotease hyperactivity in PCLDs is high –
lighted by the fact that chronic pharmacological inhibi –
tion of matrix metalloproteases with marimastat halts
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758 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastroTable 4 | Preclinical studies in animal models of polycystic liver diseases
Study Drug Animal model Treatment Results
cAMP
Masyuk et al.
(2007)97Octreotide PCK rat Intraperitoneally (10 μg/kg daily) for
4, 8, 12, and 16 weeks or 100 μg/kg
daily for 4 weeksReduction of liver weight, cyst
volume, hepatic fibrosis, and
mitotic indices
Masyuk et al.
(2013)99Pasireotide PCK rat and
Pkd2WS25/– mouseOsmotic mini-pumps (20 μg/kg daily)
for 6 weeksReduction of liver weight, cyst
volume, hepatic fibrosis and
mitotic indices
Raf kinase
Spirli et al.
(2012)89Sorafenib Pkd2flox/–:pCxCreERTM
(Pkd2cKO mouse)Orally (20–60 mg/kg daily) for 8 weeks Increase in liver cyst area,
cell proliferation and
phosphorylated Erk
mTOR
Spirli et al.
(2010)73Sirolimus Pkd2flox/–:pCxCreERTM
(Pkd2cKO mouse)Intraperitoneally (1.5 mg/kg daily)
for 8 weeksReduction of liver cyst area,
liver weight and Pcna expression
Renken et al.
(2011)84Sirolimus PCK rat In drinking water (2 mg/kg daily)
for 4, 8 or 12 weeksFailed to attenuate hepatorenal
cystogenesis
Cdc25A
Masyuk et al.
(2012)75Vitamin K3 Pkd2WS25/– mouse
and PCK ratIn drinking water (0.15 g/l) for 4
and 8 weeksReduction of both liver and
kidney weights and hepatorenal
cystic and fibrotic areas
Masyuk et al.
(2012)75PM-20 Pkd2WS25/– mouse Intraperitoneally (1 mg/kg) for 4 weeks Reduction of hepatorenal
cystogenesis
MMPs
Urribarri et al.
(2014)114Marimastat PCK rat Orally twice a day (0.29 mg/kg daily)
for 8 weeksReduction of hepatic
cystogenesis, fibrosis and
inflammation
Hdac6
Gradilone et al.
(2014)119ACY-1215 PCK rat Intraperitoneally (30 mg/kg daily)
for 4 weeksReduction of hepatic
cystogenesis and fibrosis
Ppar- γ
Yoshihara et al.
(2011)90Pioglitazone
(full agonist)PCK rat Orally (10 mg/kg daily) for 16 weeks Reduction of liver weight, liver
cystic area and fibrotic index
Yoshihara et
al. (2013)92Telmisartan
(partial agonist)PCK rat Orally (3 mg/kg daily) for 16 weeks Reduction of liver weight, cystic
and fibrotic areas
Vegfr2
Spirli et al.
(2010)74SU5416 Pkd1KO and
Pkd2KO miceIntraperitoneally (12.5 mg/kg) twice
a week for 8 weeksReduction of hepatic
cystogenesis, liver weight and
levels of phosphorylated Erk
and Pcna in Pkd2KO but not
Pkd1KO mice
Amura et al.
(2007)82SU5416 Pkd2WS25/ mouse Subcutaneously (0.75 mg) every
2 weeks for 4 and 8 monthsReduction of liver cystic area
Egfr
Torres et al.
(2004)81EKI-785 and
EKB-569PCK rat Intraperitoneally (EKI -785: 90 mg/kg)
every 3 days
Intraperitoneally (EKB -569: 20 mg/kg)
every 3 days
Orally (EKB -569: 5, 10 or 20 mg/kg)
every 3 days
All administered for 7 weeksNo effects on fibrocystic liver
disease
Trpv4
Gradilone
et al. (2010)65Trpv4 activator
GSK1016790APCK rat Intraperitoneally (0.01 mg/kg daily)
for 8 weeksTrpv4 activation induced a
significant decrease in renal
cystic area and a nonsignificant
decrease in liver cysts
Abbreviations: cAMP , cyclic adenosine monophosphate; Cdc25A, cyclin division cycle 25A; Egfr, epidermal growth factor receptor; Erk, extracellular-regulated
kinase; Hdac6, histone deacetylase 6; MMP , matrix metalloprotease; mTOR, mammalian target of rapamycin; Pcna, proliferating cell nuclear antigen; PPAR-y,
peroxisome proliferator-activated receptor gamma; Raf, rapidly accelerated fibrosarcoma; Trpv4, transient receptor potential cation channel, subfamily V,
member 4; Vegfr2, vascular endothelial growth factor receptor 2.
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NATURE REVIEWS | GASTROENTEROLOGY & HEPATOLOGY VOLUME 11 | DECEMBER 2014 | 759hepatic cystogenesis in PCK rats, decreasing fibrosis and
inflammation.114 Importantly, the dose of marimastat
used was reported as nontoxic in clinical trials for cancer
treatment.116,117 Thus, inhibition of matrix metalloprotease
hyperactivity in cystic cholangio cytes has emerged as a
potential therapeutic tool for the treatment of patients with
PCLDs and needs to be v alidated in future clinical trials.
Epigenetic abnormalities
In addition to genetic mutations, epigenetic alterations
such as abnormal expression of microRNAs (miRNAs)
and histone deacetylases have been associated with
the hyperproliferative phenotype of cystic cholangio –
cytes (Figure 1h). Global changes in miRNA levels were
reported in cholangiocytes isolated from PCK rats com –
pared with control animals. Out of 109 differentially
expressed miRNAs analysed by microarray, 97 (~89%)
were downregulated and 12 (~11%) were overexpressed in
PCK rats.118 Downregulation of the majority of miRNAs
in PCK rat cholangiocytes was associated with overexpres –
sion of their predicted target proteins involved in prolifera –
tion, secretion and cell–extracellular matrix interactions.118
In particular, miRNA -15a was highly underexpressed in
cultured PCK rat cholangiocytes, as well as in cystic tissue
from PCK rats and patients with PCLDs. Downregulation
of miRNA -15a led to overexpression of the cell division
cycle 25A (Cdc25A) protein.118 Experimental upregula –
tion of miRNA -15a in PCK rat cholangiocytes decreased
levels of Cdc25A, which halted cholangiocyte hyper –
proliferation and cyst growth. By contrast, inhibition of
miRNA -15a in normal rat cholangiocytes promoted cell
proliferation, increased Cdc25A levels and accelerated cyst
growth.118 To determine the potential therapeutic value
of targeting Cdc25A in patients with PCLDs, Cdc25A+/–
mice (with decreased Cdc25A expression but normal
liver morphology) were cross-bred with Pkhd1del2/del2 mice
(which overexpress Cdc25A and develop hepatic cysts). As
expected, liver weights, hepatic cystogenesis and fibrosis
were reduced in double mutants compared with Pkhd1del2/
del2 mice.75 In addition, pharmacological inhibition of
Cdc25A with vitamin K3 or PM -20 decreased hepatorenal
cystogenesis and fibrosis in PCK rats and Pkd2WS25/– mice
by affecting cell cycle progression and proliferation.75
Histone deacetylase 6, which is involved in the regu –
lation of the cell cycle and ciliary disassembly, has also
been associated with the pathogenesis of PCLDs. Histone
deacetylase 6 is overexpressed in cystic cholangio –
cytes from both PCK rats and patients with PCLDs,119 and pharmacological inhibition of this complex with
tubastatin -A, tubacin or ACY -1215 all decreased prolifera –
tion of PCK rat cholangio cytes in vitro in a dose-dependen t
and time-dependent manner. Moreover, treatment of
PCK rats with ACY -1215 attenuated hepatic cysto genesis
and fibrosis, which indicates that targeting histone
d eacetylase 6 might be a useful therapeutic approach.119
Conclusions
Despite advances in our understanding of the mecha –
nisms of hepatic cystogenesis and the discovery of poten –
tial therapeutic targets, the available treatment options
(conservative management, surgery and medical thera –
pies) are limited.1,22,25,120,121 Novel approaches are mainly
focused on the cAMP signalling pathway1,61,120,122 and
evaluating the effect of somatostatin analogues on hepatic
cystogenesis. However, given that the benefits of using
somatostatin analogues are modest, identification of new
targets for therapeutic intervention is urgently needed.
Several new targets (for example, histone deacetylase 6,
Cdc25A phosphatase, PPAR-γ and matrix metallopro –
teases)75,90,92,114,119 have already been evaluated in pre –
clinical studies and need to be tested clinically (Table 4).
Moreover, downregulation of the intracellular calcium
levels in cholangiocytes seems to be a central event in
hepatic cystogenesis. Therefore, restoration of these levels
in cystic cholangiocytes seems to be a promising thera –
peutic strategy. Advances in the field of PCLD research
and the discovery of new mutations in genes involved in
disease susceptibility, such as LRP5 in ADPLD, has high –
lighted new signalling pathways, such as Wnt signalling,
that could be pharmacologically important to target.
Finally, the possibility exists that combined therapeutic
strategies might have an additive or synergistic effect.
Review criteria
We searched PubMed (June 2014) using the following
terms: “polycystic liver diseases”, “cystogenesis”,
“ADPKD”, “ADPLD”, “ARPKD”, “congenital hepatic
fibrosis”, “Caroli disease”, “primary cilium”, “ductal
plate malformation”, “cystic fluid”, “cystogenesis”,
“biliary dilatation”, “cystic cholangiocytes”, “molecular
pathways”, “proliferation”, “secretion”, “angiogenesis”,
“cell–extracellular matrix”, “microRNAs”, “epigenetics”,
“oestrogens”, “cAMP”, “calcium”, “treatment” and
“clinical trials”. All selected papers were full-text in
English. We searched the reference lists of identified
papers for further relevant papers.
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management of polycystic liver disease.
Nat. Rev. Gastroenterol. Hepatol. 10, 101–108
(2013).
2. Drenth, J. P ., te Morsche, R. H., Smink, R.,
Bonifacino, J. S. & Jansen, J. B. Germline
mutations in PRKCSH are associated with
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3. Li, A. et al. Mutations in PRKCSH cause
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6. [No authors listed] The polycystic kidney
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lies within a duplicated region on chromosome
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760 | DECEMBER 2014 | VOLUME 11 www.nature.com/nrgastropolycystic kidney disease. J. Am. Soc. Nephrol .
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Jansen, J. B. & Drenth, J. P . Extensive mutational
analysis of PRKCSH and SEC63 broadens the
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Author contributions
All authors contributed equally to all aspects
of this manuscript.
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