Cellular and Molecular Cascades during Liver Regeneration

Abstract

The demand for organs such as the liver for patients with end stage disease is greater than what is currently available. Thus, there is a dire need to have alternative solutions, for which none exist at the moment. Investigating the key underlying mechanisms involved not only in liver regeneration and repair, but also in development, can give us a better under- standing of how to promote a pro-regenerative phenotype in the liver. This review will focus on the cellular and molecular aspects of liver regeneration and address signaling mechanisms involved in liver development and how they are recapitulated in regeneration after a partial hepatectomy.

Full text

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Cellular and Molecular Cascades during
Liver Regeneration
Ali-Reza Sadri1, Marc G. Jeschke2,3 and Saeid Amini-Nik2,3*
1Sunnybrook Health Sciences Centre, Toronto, Canada
2Department of Surgery, University of Toronto, Canada
3Sunnybrook’s Trauma, Emergency & Critical Care (TECC) Program, Canada
*Corresponding author:
Saeid Amini Nik, MSc, MD, PhD
Assistant Professor
Department of Surgery
University of Toronto
Sunnybrook’s Trauma
Emergency & Critical Care (TECC)
Program
Ofce: M7-161, Lab: M7-140
2075 Bayview Ave. Toronto
ON M4N 3M5, Canada
Tel. 416-480-6100
Fax: 416-480-6763
E-mail: saeid.amininik@utoronto.ca
Article History:
Received: June 22nd, 2015
Accepted: August 3rd, 2015
Published: August 17th, 2015
Citation:
Sadri AR, Jeschke MG, Amini-Nik
S. Cellular and molecular cascades
during liver regeneration. Surg Res
Open J. 2015; 2(2): 53-61.
Copyright:
© 2015 Amini-Nik S. This is an open
access article distributed under the
Creative Commons Attribution Li-
cense, which permits unrestricted
use, distribution, and reproduction
in any medium, provided the origi-
nal work is properly cited.
Volume 2 : Issue 2
Article Ref. #: 1000SROJ2110
Review
Page 53
ABSTRACT
The demand for organs such as the liver for patients with end stage disease is greater
than what is currently available. Thus, there is a dire need to have alternative solutions, for
which none exist at the moment. Investigating the key underlying mechanisms involved not
only in liver regeneration and repair, but also in development, can give us a better under-
standing of how to promote a pro-regenerative phenotype in the liver. This review will focus
on the cellular and molecular aspects of liver regeneration and address signaling mechanisms
involved in liver development and how they are recapitulated in regeneration after a partial
hepatectomy.
KEYWORDS: Liver; Regeneration; Hepatectomy; Stem cell; Healing; Inammation.
ABBREVIATIONS: PHx: Partialhepatectomy; HSC: Hepatic Stellate Cells; BECs: Biliary Epi-
thelial Cells; STM: Septum Transversum Mesenchyme; IL-6: Interleukin-6; FSCs: Facultative
Stem Cells; Ang2: Angiopoietin 2; GFAP: Glial Fibrillary Acidic Protein; GFP: Green Fluores-
cent Protein; Hh: Hedgehog.
PREFACE
The liver’s remarkable regenerative capacity was rst described by the Greeks in
the legend of Prometheus, a Titan who was banished by Zeus to eternal punishment. He was
chained to a rock on a mountain, where an eagle would eat his liver daily, only to have it regen-
erate every night. To this date, we still do not have a clear idea how the liver recovers following
injury.
INTRODUCTION
The liver is known for its imperative roles in metabolic homeostasis, immune regu-
lation, bile secretion, serum protein synthesis and detoxication properties. The majority of
blood ow that enters the liver is from the spleen, pancreas and intestines via the portal vein.
This blood gets ltered from toxins and drugs before entering the heart to be circulated to the
rest of the body. Thus, the liver is subjected to routine exposure to damaging agents. It has
been hypothesized that the liver has evolved to become a highly regenerative organ to counter
these toxins,1 because liver dysfunction and failure can ultimately lead to death. It is yet to be
demonstrated whether the liver’s remarkable regenerative capacity is due to several cell types
or a single cell of origin.
One of the most studied models of cell organ and tissue regeneration is liver regenera-
tion after a 2/3 Partialhepatectomy (PHx). Different methods of liver resection are used to ob-
tain the desired amount of liver mass loss. When performing a PHx, the vessels and ducts at the
pedicel of the particular lobe must be ligated prior to cutting the lobe. Typically, the left lateral

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Page 54
lobe and median lobes are removed, which equates to 67% of the
liver mass.2 There is an impressive increase in hepatocyte pro-
liferation, which peaks at 36 hours,2 followed by reconstitution
of non-parenchymal cells after surgical liver resection as seen in
animals. This surgical model has become popular over the last
few years and gained acceptance by the majority of the research
community for numerous reasons. The rst reason being due to
the multi-lobe structure of the liver, resection of different seg-
ments can be done without disturbing the remnant lobe(s). Thus,
regeneration of the remaining lobes is accomplished through
liver specic mechanisms and not due to acute inammation or
necrosis,3,4 which is observed during liver laceration. Second,
the procedure can be done in 10-15 minutes and regeneration
is triggered almost instantly, which can be tracked temporally
through different phases. Third, the procedure is easily repro-
ducible and if done correctly, all animals will survive.2
Due to the absence of any signicant inammation or
injury to the remaining lobes after a PHx,4 there is no reported
observation of stem cell activation or cellular reprogramming. In
fact, after a PHx, the liver does not regrow the resected lobes but
the remaining lobes, compensate for the loss via proliferation
and increase in hepatocyte size. This process is referred to as
“compensatory hypertrophy”5,6 but we will continue to use “liver
regeneration” as it is still a widely used term in this eld. Previ-
ous studies have shown that during liver regeneration, almost all
the hepatocytes undergo1-2 rounds of replication to restore nor-
mal liver mass.6,7 However, more recent ndings using modern
lineage tracing and imaging techniques demonstrate that cellular
hypertrophy is a signicant contributor to the compensatory re-
sponse and that hepatocytes undergo on average only 0.7 rounds
of cell division in mice. The rst 4 hours after a PHx is known as
the “priming phase” as hepatocytes prepare to respond to vari-
ous cytokines by substantially changing their gene expression,
including up-regulation of anti-proliferative genes.8 It is specu-
lated that it is during this phase that hepatocyte hypertrophy is
initiated.
Considering that healing involves several stages start-
ing with inammation, it is not clear whether the regenerative
capacity of liver is mainly due to the absence of signicant in-
ammation or the internal capacity of liver by itself to deliver
the regeneration capacity. Part of this might be due to its unique
histology and anatomical position, which we will discuss here.
LIVER ANATOMY
The liver is made of liver lobules, which are hexagonal
in shape with a portal triad in each corner and a central vein in
the center9 (Figure 1A). The portal triad consists of a bile duct,
portal venule and portal arteriole. Hepatocytes work to absorb
metabolites and toxins, which have entered the liver through the
portal vein. Bile is secreted from hepatocytes into the bile ducts,
which will eventually enter the gall bladder for storage and re-
leased into the duodenum. Sinusoids are lined with endothelial
cells forming the blood vessels. They drain the blood from the
portal venules and arterioles into the central vein to be taken
back to the heart. Inside the sinusoids are Kupffer cells, which
are the resident macrophages of the liver. These cells work to
cleanse the blood before it enters the central vein. Hepatic Stel-
late Cells (HSC) are located in the area between the sinusoids
and hepatocytes, known as the space of Dissé10 (Figure1B).
OVERVIEW OF LIVER DEVELOPMENT
Hepatocytes make up approximately 70% of the mass
of the adult organ and are derived from embryonic endoderm,
as are Biliary Epithelial Cells (BECs), also known as cholan-
giocytes. Other cells populating the liver include stellate cells,
Kupffer cells and endothelial cells, which are of mesodermal
origin. Through developmental studies on various animal mod-
els such as mouse, chicken, zebrash, and Xenopus, many genes
and molecular pathways have been identied that regulate em-
bryonic development. These studies have enabled scientists to
identify pathways implicated in liver regeneration in adult ani-
mals and humans. The regenerative mechanisms appear to reca-
pitulate what is observed during development.
Figure 1: The functional unit of the liver. (A) The liver lobule. (B) The cell populations between
the portal triad and central vein.

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Page 55
The endoderm germ layer develops during gastrulation
and forms a primitive gut tube that is subdivided into foregut,
mid-gut, and hindgut regions. Fate mapping studies have dem-
onstrated that the embryonic liver originates from the ventral
foregut endoderm at embryonic day 8.0 of gestation (e8.0).11 The
thickening of the ventral foregut epithelium at e9.0 results in a
hepatic diverticulum, which is the rst indicator of liver devel-
opment. The anterior segment of the hepatic diverticulum gives
rise to the liver and intrahepatic biliary tree, while the posterior
segment forms the gallbladder and extra-hepatic bile ducts. Pre-
ceding vascularization of the liver bud, at e9.0 endothelial pre-
cursor cells are situated between the epithelial cells and the Sep-
tum Transversum Mesenchyme (STM). Expression of vascular
endothelial growth factor receptor 2 (Vegfr-2) has been shown to
be essential as embryos that lack this gene fail to produce endo-
thelial cells and hepatoblasts cannot go on to occupy the STM.12
During e9.5, the liver bud forms through the hepatic
endoderm cells, known as hepatoblasts, and occupying the
STM.13,14 The STM provides the hepatic broblasts and stel-
late cells.15 Starting at e10 until e15, liver bud gets invaded by
hematopoietic cells as its development accelerates in order to
become the main hematopoietic organ of the fetus. Thus, liver
development involves contributions from tissues of endoderm
and mesoderm origin. Hepatoblasts have bi-potential proper-
ties. Hepatoblasts that surround the portal vein differentiate to
cholangiocytes, which form the primitive bile ducts also known
as ductal plates. Primitive cholangiocytes express markers: Sry
box containing gene 9 (Sox9), Ostepontin (OPN), and EpCAM.
The remaining hepatoblasts in the parenchyma differentiate into
hepatocytes.16
ESSENTIAL FACTORS DURING LIVER DEVELOPMENT
The regional identity of the endoderm seems to be
contingent upon the spatial gradients of FGF, Wnt, BMP and
retinoic acid secreted from the adjacent mesoderm.17 However,
it is still not understood how these pathways specify regional
identity. Studies on chick and Xenopus suggest that FGF and
Wnts released from the posterior mesoderm suppress foregut
fate and promote hindgut development.18 To establish foregut
identity Wnt and FGF4 signaling needs to be inhibited in the
anterior mesoderm. Inhibiting β-catenin, a downstream effector
molecule in Wnt signaling, results in activation of Hhex, lead-
ing to ectopic liver buds in the intestine.17 Interestingly, by e10,
β-catenin has the opposite effect and promotes hepatic growth.19
The specic Wnt ligands that effect hepatogenesis are still un-
known. Experiments on chick embryos show that Wnt9a ex-
pressed in the sinusoidal wall is essential for liver bud growth
through proliferation of hepatoblast and hepatocytes in culture.12
In zebrash, Wnt2b expression in the lateral plate mesoderm
has been shown to be necessary for liver development. Wnt2
is also expressed in the lateral plate mesoderm and cooperates
with Wnt2bbto control liver specication and proliferation in
zebrash.20 The combined role of these signaling molecules is
essential for liver specication because blocking them causes
liver agenesis.21
In terms of hepatoblast proliferation and differentia-
tion, hedgehog signaling is involved in promoting the prolifera-
tive response and subsequently needs to be shut off for differen-
tiation to occur in a timely manner.22
Jagged-1, a Notch ligand is known to be expressed in
the portal mesenchyme, which activates Notch-2 in neighbour-
ing hepatoblasts, to promote differentiation of hepatoblasts into
bile ducts.23 Loss of Jag1 expression in the portal vein mesen-
chyme causes duct development to stall midway during ductal
plate morphogenesis, leading to a paucity of bile ducts.24
Despite advancements in system biology and cell lin-
eage studies, the cellular and molecular mechanisms of liver
regeneration are still not clear. The information we learn and
gather from regeneration of the liver may be used and applied to
enhance regeneration of other organs. Here, we summarize the
molecular and cellular mechanisms of liver regeneration after a
PHx.
THE CELLULAR RESPONSE AFTER A PHx
Proliferation is the main method of liver regeneration
after a PHx.25 In mice it takes one week for the liver to return to
75% of its original size. The regenerative response involves con-
stitution of hepatocytes rst followed by biliary epithelial cells
and then non-parenchymal cells.26 Although cellular prolifera-
tion is the key regenerative mechanism, cellular hypertrophy is
also observed.6 Impaired hepatocyte proliferation is observed in
aged mice, which is reversed in pregnant mice. Pregnant mice
recover from a PHx at rates comparable to younger mice through
hepatocyte hypertrophy.27 This highlights the role of systemic
factors contributing in hepatocyte hypertrophy.
The liver’s response to a PHx is divided into two main
phases. The rst phase occurs between days 1-3 and is termed
the “inductive phase” (Figure 2A). During this phase hepato-
cytes undergo proliferation. This proliferative response peaks at
36 hours and goes back down at 72 hours.28-30 The “angiogenic
phase” is the next phase which occurs, from day 4 to 8, where
non-parenchymal cells proliferate, returning the liver to its nor-
mal mass and function (Figure 2B). Non-parenchymal cells have
an essential role during these phases of regeneration, which will
be discussed in more detail below.
THE MOLECULAR RESPONSE AFTER A PHx
The ability of the liver to know when to start and stop
regeneration has puzzled scientists for years. However, certain
factors have been shown to be necessary for regeneration post
PHx. For example, Interleukin-6 (IL-6) and the bile acid recep-
tor, FXR, have been shown to be essential for regeneration.31,32

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Page 56
When the genes for IL-6 or FXR are knocked down, there is
a higher mortality rate post PHx compared to their respective
wild-type counterparts. In addition, assessment of proliferation
through BrdU staining shows a poor proliferative response in
hepatocytes. However, there is no change in non-parnechymal
cells such as Kupffer cells, and endothelial cells, suggesting that
non-parenchymal cells do not need IL-6 for this response.
According to transplantation studies, hepatocytes ap-
pear to have intrinsic regenerative mechanisms that are species-
specic. For instance, transplantation of rat hepatocytes into
mice liver, which later are subjected to PHx, has shown irreg-
ular proliferation kinetics. Rat hepatocytes become BrdU+ 24
hours later, as expected while mouse hepatocytes express BrdU
32 hours later.33 Thus, even with the change in cellular environ-
ment, rat hepatocytes stay true to their typical response to a PHx.
This suggests that hepatocytes have a certain level of autonomy
when it comes to regeneration and highlights the intrinsic ca-
pability of hepatocytes rather than micro-environmental niche
effects.
An alternative mechanism to liver regeneration in-
volves a group of cells termed “Facultative Stem Cells” (FSCs)
or “oval cells”. FSCs were rst described in rat studies that in-
volved exposure to several carcinogens that are known to be
toxic to the liver.34 In rats it has been shown that these cells ap-
pear when hepatocyte proliferation is impaired but they are also
observed in mice even with hepatocyte proliferation. However,
the appearance of oval cells or impaired proliferation is not ob-
served when rodents undergo a PHx without any chemical inter-
vention. Further discussion of FSCs is beyond the scope of this
review. Although, it is evident that the liver’s resiliency comes
from the multiple avenues of regeneration at its disposal.
LIVER SINUSOIDAL ENDOTHELIAL CELLS (LSECs)
LSECs are shown to regulate the temporal response
of liver regeneration post-PHx. Angiopoietin 2 (Ang2), is an
angiogenic protein that is down-regulated during the inductive
phase,30 which is associated with decreased TGF-β, an anti-
proliferative factor, and increased expression of cyclin D1, thus
boosting hepatocyte proliferation (Figure 2C). In the angio-
genic phase, Ang2 levels increase, and subsequently promotes
increased VEGFR2 and Wnt2 expression and proliferation of
LSECs initiates29 (Figure 2D).
The liver vasculature has varying responses to whether
there is an acute or chronic injury. During an acute insult, there
is up-regulation of CXCR7 by LSECs and increase in CXCR4,
which together induce transcription factor inhibitor of DNA
binding 1 (Id1).28 This induces production of Wnt2 and HGF,
which are pro-regenerative angiocrine factors and triggers re-
generation. The essential role of CXCR7 was shown when dele-
tion of CXCR7 in LSECs through an inducible system resulted
in a poor regenerative response due to an impaired ID1 mediated
production of angiocrine factors.28 (Table 1)
MACROPHAGES
The powerful role macrophages play in regeneration
has been shown in organisms such as zebrash, which depend
on these cells to regenerate their ns, and portions of the heart.
In addition, macrophages are required for limb re-growth in
salamanders.40 The liver is known to have the highest concentra-
tion of resident macrophages of any organ. Both Kupffer cells
and recruited monocyte-derived macrophages have been impli-
Liver Lineage Signalling pathway Reference
Foregut Endoderm Wnt/β-catenin and FGF4 sup-
pressed
18
Hepatoblast FGF, BMP 35,36
Hepatocyte Wnt/β-catenin 20,37
Cholangiocyte (Bile duct
cell) Notch 38,39
Figure 2: The proliferation kinetics and main signalling pathways involved in liver regeneration
after a PHx. (A) Proliferation of non-parenchymal cells occurs during the inductive phase. (B) The
angiogenic phase involves proliferation of non-parenchymal cells. (C) Role of non-parenchymal
cells during hepatocyte proliferation in the inductive phase. (D) Role of non-parenchymal cells in
inhibiting hepatocyte proliferation arrest and regeneration in the angiogenic phase.
Table 1: Signalling pathways involved in liver development.

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Page 57
cated in liver regeneration after a PHx.41 When macrophages are
ablated using liposomal clodronate followed by a PHx there is
a delayed proliferative response from hepatocytes and the size
of the remnant liver at 96 hours post-surgery is signicantly
less in Kupffer cell depleted rats.41 This suggests that cytokines
and growth factors secreted by macrophages are important for
proliferative responses. Expression of key cytokines involved
in liver regeneration are also down regulated at the mRNA
level, this includes, IL-6, IL-10, TNF, HGF, and TGF-β1 at 4
hours post-PHx. The temporal defect in liver regeneration due
to the absence of Kupffer cells may be associated with a lack
of Wnt ligands that promote Wnt/β-catenin signaling in hepa-
tocytes. When there is macrophage specic knockdown of the
gene Wntless and PHx is performed a temporal deciency in
liver regeneration is observed.42 There is a 1/3 drop in S- phase
hepatocytes and hepatocyte mitosis was observed in Wls-MKO
mice 40 hours after PHx. This was associated with a reduction in
β-catenin-TCF4 complex and Cyclin-D1 expression at 40 hours,
highlighting a role for β-catenin mediated TCF transcription fac-
tor in this process.43 These ndings suggest that Kupffer cells are
essential for initiating hepatocyte proliferation in a timely man-
ner through secretion of Wnt ligands. Other factors thought to be
important for hepatocyte proliferation is interleukin-6 (IL-6) and
tumor necrosis factor-α (TNF-α). Mice decient in either IL-6
or TNF-α receptor type 1 showed impaired hepatocyte prolifera-
tion 40 hours post-surgery and higher mortality. (Table 2)
STELLATE CELLS
In a healthy liver, Hepatic Stellate Cells (HSCs) are
in a quiescent state and store lipids such as Vitamin A. HSCs
encompass approximately 5-8% of cells. Upon chronic liver
injury, impaired hepatocytes and immune cells secrete factors
that cause HSCs to become proliferative and differentiate into
myobroblasts.10 These myobroblasts are well known to be
key producers collagen 1 and promote brosis.49 Thus, it seems
they are associated with an undesirable outcome in liver injury.
However, it is also suggested that HSCs may have pro-regener-
ative properties as well. Spatially, the majority of HSCs reside
in the Canals of Hering, a suggested stem cell niche in the adult
liver.46 More importantly, they are known to produce factors as-
sociated with regeneration such as HGF, Notch and hedgehog
ligands. HSCs isolated from the early phase of regeneration in
rats showed high levels of HGF in conditioned media. Further-
more, it has been argued that HSCs express the stem/progeni-
tor cell marker CD133+ and are able to differentiate into he-
patocyte-like cells with certain cytokines.47 A lineage study was
done on HSCs using a Glial Fibrillary Acidic Protein (GFAP)
promoter and a Green Fluorescent Protein (GFP) reporter gene
showing that after a diet-induced injury GFP+ cells proliferate
and express progenitor markers cytokeratin 7 and 19.48 After-
wards, GFP+ hepatocytes were observed suggesting that HSCs
gave rise to progenitor cells that went on to differentiate into
hepatocytes. They show that HSCs may produce hepatocytes via
mesenchymal to epithelial transition.
HSCs play an essential role during liver regeneration as
their regulatory effect includes stopping regeneration. They se-
crete factors that arrest regeneration once the appropriate mass is
achieved. The dominant arresting factor is TGF-β, which HSCs
are the main producers of in the liver. In mice with the gene
Foxf1 knocked down, the stellate cells were unable to become
activated and impaired liver regeneration ensued along with di-
minished notch-2 production, which promotes regeneration of
biliary epithelial cells.50 Furthermore, in rats with 2-AAF/PHx
injured livers and given L-cysteine in their diets, to impair stel-
late cell activation, there was abnormal regeneration due to poor
progenitor cell response.51 Thus, HSCs appear to have a tem-
poral role in regulating the regenerative response of the liver.
Initially, they promote regeneration through secretion of growth
factors and then put on the brakes once the normal weight and
function is achieved.
THE CRITICAL ROLE OF HEDGEHOG SIGNALING
The importance of Hedgehog (Hh) signaling goes be-
yond just development as it is up regulated during regeneration
after PHx. When Hh signaling is blocked after a PHx, via cy-
clopamine, there is reduced expression of numerous progeni-
tor markers such as α-fetoprotein (AFP), Factor-inducible 14
Role in liver regeneration Reference
Hepatocyte Hyper proliferative response post-PHx 25
Cholangiocyte (Bile duct cell) Hyper proliferative response post-PHx 38,44
Sinusoidal endothelial cell Spatiotemporal regulation in proliferation
kinetics of hepatocytes and endothelial cells
28-30
Kupffer cell
Secrete wnt ligands that control hepato-
cyte proliferation in a timely manner, wnt3a
secretion promotes differentiation of hepatic
progenitor cells into hepatocytes.
41,45
Stellate cell
Secrete factors that promote and stop hepato-
cyte proliferation. May give rise to hepatocytes
through MET, Secretion of Notch ligands
promotes differentiation of hepatic progenitor
cells to cholangiocytes.
46-48
Table 2: Contribution of different cellular components of the liver during liver regeneration.

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
Page 58
(Fn14), and cytokeratin 19 at the mRNA and protein level.52,53
Furthermore, proliferation of hepatocytes was impaired as BrdU
incorporation decreased by 90% in hepatocytes and 40% in
ductular cells.52,53 The nal outcome of this treatment shows a
higher mortality in comparison with the control treated group.
This highlights the importance of Hh signaling pathway in liver
regeneration. It is still not clear which cell type needs activation
of the Hh signaling pathway during liver regeneration, which
can be further elucidated in cell lineage studies.
CLINICAL INSIGHTS INTO LIVER REGENERATION
The human liver, like in rodents, undergoes a hyper-
proliferative response after a PHx.54,55 However, because the
PHx model in animals, when done with precision, is relatively
“clean” it does not fully recapitulate what is observed in the con-
text of human liver disease, where signicant inammation, ne-
crosis, and brosis are commonly observed.
In humans, outcomes of hepatectomy have improved
over time. However, post-hepatectomy liver failure is still one of
the most fatal complications of hepatectomy and occurs in up to
10% of cases. The ability of the remnant liver to regenerate af-
ter hepatectomy is the main factor in determining morbidity and
mortality. If the remnant liver is less than 20%, liver function is
impaired and could lead to post-resection liver failure.56,57 Due to
a scarcity in treatments for numerous liver conditions, liver re-
section remains the sole remedy,54-57 despite the high concern for
morbidity and mortality.58,59 Investigating the pro-regenerative
aspects of the cell types discussed above may assist in enhanc-
ing the recovery and survival of patients’ post-hepatectomy and
possibly after trauma, such as a severe burn.60 Thus, despite the
divergence, the compensatory response after liver resection is
clinically essential and provides a great model to learn about
growth and regeneration. A better understanding of how cells
in the liver interact and respond to their microenvironment will
give us the ability to pinpoint aberrant healing and develop novel
therapies to treat liver disease.
FUTURE OUTLOOK
The regenerative capacity of the liver is unquestion-
able. Whether a single or several cell type(s) give rise to new he-
patocytes during liver regeneration is not yet well dened. While
it is believed that hepatocytes undergo hypertrophy and prolifer-
ate to regenerate the liver, it is not clear whether all hepatocytes
are able to proliferate. Can a group of hepatocytes have higher
capacity to proliferate? Are these hepatic progenitor cells? In ad-
dition, the majority of liver regeneration studies using the PHx
model focus on how regeneration is initiated and what factors
promote it while missing out on how it is stopped once regenera-
tion is complete. Thus, future studies need to focus more on cell
specic studies through lineage tracing to address the plasticity
of liver cells and their fate during regeneration. Furthermore, a
better understanding of how liver regeneration is terminated and
the discrepancies between the PHx model in rodents and what is
observed in the clinic need to be taken into consideration.
CONFLICTS OF INTEREST
The authors declare that they have no conicts of interest.
ACKNOWLEDGMENTS
Thank you to Andrea Datu for assisting in editing the manu-
script.
REFERENCES
1. Michalopoulos GK, DeFrances MC. Liver regeneration. Sci-
ence. 1997; 276(5309): 60-66.
2. Mitchell C, Willenbring H. A reproducible and well-tolerated
method for 2/3 partial hepatectomy in mice. Nat Protoc. 2008.
3(7): 1167-1170. doi: 10.1038/nprot.2008.80
3. Fausto N, Campbell JS, Riehle KJ. Liver regeneration. Hepa-
tology. 2006; 43(2 Suppl 1): S45-S53.
4. Michalopoulos GK. Liver regeneration after partial hepatec-
tomy: critical analysis of mechanistic dilemmas. Am J Pathol.
2010; 176(1): 2-13.
5. Miyajima A, Tanaka M, Itoh T. Stem/progenitor cells in
liver development, homeostasis, regeneration, and reprogram-
ming. Cell Stem Cell. 2014; 14(5): 561-574. doi: 10.1016/j.
stem.2014.04.010
6. Miyaoka Y, Ebato K, Kato H, Arakawa S, Shimizu S, Miya-
jima A. Hypertrophy and unconventional cell division of hepato-
cytes underlie liver regeneration. Curr Biol. 2012; 22(13): 1166-
1175. doi: 10.1016/j.cub.2012.05.016
7. Alison, MR, Islam S, Lim S. Stem cells in liver regeneration,
brosis and cancer: the good, the bad and the ugly. J Pathol.
2009; 217(2): 282-298. doi: 10.1002/path.2453
8. Su AI, Guidotti LG, Pezacki JP, Chisari FV, Schult PG. Gene
expression during the priming phase of liver regeneration after
partial hepatectomy in mice. Proc Natl Acad Sci USA. 2002;
99(17): 11181-11186. doi: 10.1073/pnas.122359899
9. Thomson AW, Knolle PA. Antigen-presenting cell function
in the tolerogenic liver environment. Nat Rev Immunol. 2010;
10(11): 753-766. doi: 10.1038/nri2858
10. Yin, C, Evason KJ, Asahina K, Stainier DY. Hepatic stellate
cells in liver development, regeneration, and cancer. J Clin In-
vest. 2013; 123(5): 1902-1910. doi: 10.1172/JCI66369

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
11. Tremblay KD, Zaret KS. Distinct populations of endoderm
cells converge to generate the embryonic liver bud and ventral
foregut tissues. Dev Biol. 2005; 280(1): 87-99. doi: 10.1016/j.
ydbio.2005.01.003
12. Matsumoto K, Miki R, Nakayama M, Tatsumi N, Yokouchi
Y. Wnt9a secreted from the walls of hepatic sinusoids is essential
for morphogenesis, proliferation, and glycogen accumulation of
chick hepatic epithelium. Dev Biol. 2008; 319(2): 234-247. doi:
10.1016/j.ydbio.2008.04.021
13. Houssaint E. Differentiation of the mouse hepatic primor-
dium. I. An analysis of tissue interactions in hepatocyte differ-
entiation. Cell Differ. 1980; 9(5): 269-279. doi: 10.1016/0045-
6039(80)90026-3
14. Medlock ES, Haar JL. The liver hemopoietic environment:
II. Peroxidase reactive mouse fetal liver hemopoietic cells. Anat
Rec. 1983; 207(1): 43-53.
15. Asahina K, Tsai SY, Li P, et al. Mesenchymal origin of he-
patic stellate cells, submesothelial cells, and perivascular mes-
enchymal cells during mouse liver development. Hepatology.
2009; 49(3): 998-1011. doi: 10.1002/hep.22721
16. Lemaigre FP. Development of the biliary tract. Mech Dev.
2003; 120(1): 81-87. doi: 10.1016/S0925-4773(02)00334-9
17. McLin VA, Rankin SA, Zorn AM. Repression of Wnt/beta-
catenin signaling in the anterior endoderm is essential for liver
and pancreas development. Development. 2007; 134(12): 2207-
2217. doi: 10.1242/dev.001230
18. Dessimoz J, Opokaa R, Kordicha JJ, Grapin-Bottonb A,
Wells JM. FGF signaling is necessary for establishing gut tube
domains along the anterior-posterior axis in vivo. Mech Dev.
2006; 123(1): 42-55. doi: 10.1016/j.mod.2005.10.001
19. Monga SP, Monga HK, Tan X, Mulé K, Pediaditakis P, Mi-
chalopoulos GK. Beta-catenin antisense studies in embryonic
liver cultures: role in proliferation, apoptosis, and lineage speci-
cation. Gastroenterology. 2003; 124(1): 202-216.
20. Ober EA, Verkade H, Field HA, Stainier DYR. Mesodermal
Wnt2b signalling positively regulates liver specication. Nature.
2006; 442(7103): 688-691. doi: 10.1038/nature04888
21. Poulain M, Ober EA. Interplay between Wnt2 and Wnt2bb
controls multiple steps of early foregut-derived organ develop-
ment. Development. 2011; 138(16): 3557-3568. doi: 10.1242/
dev.055921
22. Itoh HYT, Miyajima A. Hedgehog signal activation coordi-
nates proliferation and differentiation of fetal liver progenitor
cells. Exp Cell Res. 2009; 315(15): 2648-2657. doi: 10.1016/j.
yexcr.2009.06.018
23. Zong Y, Panikkar A, Xu J, et al. Notch signaling controls
liver development by regulating biliary differentiation. Develop-
ment. 2009; 136(10): 1727-1739. doi: 10.1242/dev.029140
24. Geisler F, Nagl F, Mazur PK, et al. Liver-specic inactiva-
tion of Notch2, but not Notch1, compromises intrahepatic bile
duct development in mice. Hepatology. 2008; 48(2): 607-616.
doi: 10.1002/hep.22381
25. Yanger K, Knigin D, Zong Y, et al. Adult hepatocytes are
generated by self-duplication rather than stem cell differen-
tiation. Cell Stem Cell. 2014; 15(3): 340-349. doi: 10.1016/j.
stem.2014.06.003
26. Taub R. Liver regeneration: from myth to mechanism. Nat
Rev Mol Cell Biol. 2004; 5(10): 836-847. doi: 10.1038/nrm1489
27. Gielchinsky Y, Laufer N, Weitman E, et al. Pregnancy re-
stores the regenerative capacity of the aged liver via activation
of an mTORC1-controlled hyperplasia/hypertrophy switch.
Genes Dev. 2010; 24(6): 543-548. doi: 10.1101/gad.563110
28. Ding BS, Cao Z, Lis R, et al. Divergent angiocrine signals
from vascular niche balance liver regeneration and brosis. Na-
ture. 2014; 505(7481): 97-102. doi: 10.1038/nature12681
29. Ding BS, Nolan DJ, Butler JM, et al. Inductive angiocrine
signals from sinusoidal endothelium are required for liver re-
generation. Nature. 2010; 468(7321): 310-315. doi: 10.1038/
nature09493
30. Hu J, Srivastava K, Wieland M, et al. Endothelial cell-de-
rived angiopoietin-2 controls liver regeneration as a spatiotem-
poral rheostat. Science. 2014; 343(6169): 416-419. doi: 10.1126/
science.1244880
31. Cressman DE, Greenbaum LE, DeAngelis RA, et al. Liver
failure and defective hepatocyte regeneration in interleukin-
6-decient mice. Science. 1996; 274(5291): 1379-1383. doi:
10.1126/science.274.5291.1379
32. Huang W, Ma K, Zhang J, et al. Nuclear receptor-dependent
bile acid signaling is required for normal liver regeneration. Sci-
ence. 2006; 312(5771): 233-236. doi: 10.1126/science.1121435
33. Weglarz TC, Sandgren EP. Timing of hepatocyte entry into
DNA synthesis after partial hepatectomy is cell autonomous.
Proc Natl Acad Sci USA. 2000; 97(23): 12595-12600. doi:
10.1073/pnas.220430497
34. Factor VM, Radaeva SA, Thorgeirsson SS. Origin and fate of
Page 59

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
oval cells in dipin-induced hepatocarcinogenesis in the mouse.
Am J Pathol. 1994; 145(2): 409-422.
35. Shin D, Shin CH, Tucker J, et al. Bmp and Fgf signaling are
essential for liver specication in zebrash. Development. 2007;
134(11): 2041-2050. doi: 10.1242/dev.000281
36. Jung J, Zheng M, Goldfarb M, Zaret KS. Initiation of mam-
malian liver development from endoderm by broblast growth
factors. Science. 1999; 284(5422): 1998-2003. doi: 10.1126/sci-
ence.284.5422.1998
37. Nejak-BowenK, Monga SP. Wnt/beta-catenin signaling in
hepatic organogenesis. Organogenesis. 2008; 4(2): 92-99.
38. Tanimizu N, Miyajima A. Notch signaling controls hepato-
blast differentiation by altering the expression of liver-enriched
transcription factors. J Cell Sci. 2004; 117(Pt 15): 3165-3174.
doi: 10.1242/ jcs.01169
39. Li L, Krantz ID, Deng Y, et al. Alagille syndrome is caused
by mutations in human Jagged1, which encodes a ligand for
Notch1. Nat Genet. 1997; 16(3): 243-251. doi: 10.1038/ng0797-
243
40. Godwin JW, Pinto AR, Rosenthal NA. Macrophages are re-
quired for adult salamander limb regeneration. Proc Natl Acad Sci
USA. 2013; 110(23): 9415-9420. doi: 10.1073/pnas.1300290110
41. Meijer C, Pintoa AR, 41. Rosenthal NA. Kupffer cell deple-
tion by CI2MDP-liposomes alters hepatic cytokine expression
and delays liver regeneration after partial hepatectomy. Liver.
2000; 20(1): 66-77. doi: 10.1034/j.1600-0676.2000.020001066.x
42. Yang J, Mowry LE, Nejak-Bowen KN, et al. beta-catenin
signaling in murine liver zonation and regeneration: a Wnt-
Wnt situation! Hepatology. 2014; 60(3): 964-976. doi: 10.1002/
hep.27082
43. Amini-Nik S, Elizabeth Cambridge, Winston Yu, et al. beta-
Catenin-regulated myeloid cell adhesion and migration deter-
mine wound healing. J Clin Invest. 2014; 124(6): 2599-2610.
44. Lesage G, Glaser SS, Gubba S, et al. Regrowth of the rat bili-
ary tree after 70% partial hepatectomy is coupled to increased
secretin-induced ductal secretion. Gastroenterology. 1996;
111(6): 1633-1644.
45. Boulter L, Govaere O, Bird TG, et al. Macrophage-derived
Wnt opposes Notch signaling to specify hepatic progenitor cell
fate in chronic liver disease. Nat Med. 2012; 18(4): 572-579.
doi: 10.1038/nm.2667
46. Kordes C, Haussinger D. Hepatic stem cell niches. J Clin
Invest. 2013; 123(5): 1874-1880. doi: 10.1172/JCI66027
47. Kordes C, Sawitza I, Müller-Marbach A, et al. CD133+ hepat-
ic stellate cells are progenitor cells. Biochem Biophys Res Com-
mun. 2007; 352(2): 410-417. doi: 10.1016/j.bbrc.2006.11.029
48. Yang, L, Jung Y, Omenetti A, et al. Fate-mapping evidence
that hepatic stellate cells are epithelial progenitors in adult
mouse livers. Stem Cells. 2008; 26(8): 2104-2113. doi: 10.1634/
stemcells.2008-0115
49. Krizhanovsky V, Yon M, Dickins RA, et al. Senescence of
activated stellate cells limits liver brosis. Cell. 2008; 134(4):
657-667. doi: 10.1016/j.cell.2008.06.049
50. Kalinichenko VV, Bhattacharyya D, Zhou Y, et al. Foxf1 +/-
mice exhibit defective stellate cell activation and abnormal liver
regeneration following CCl4 injury. Hepatology. 2003; 37(1):
107-117. doi: 10.1053/jhep.2003.50005
51. Pintilie DG, Shupe TD, Oh SH, et al. Hepatic stellate cells’
involvement in progenitor-mediated liver regeneration. Lab In-
vest. 2010; 90(8): 1199-1208. doi: 10.1038/labinvest.2010.88
52. Ochoa B, Syn WK, Delgado I, et al. Hedgehog signaling is
critical for normal liver regeneration after partial hepatectomy
in mice. Hepatology. 2010; 51(5): 1712-1723. doi: 10.1002/
hep.23525
53. Hadem J, Bockmeyer CL, Lukasz A, et al. Angiopoietin-2 in
acute liver failure. Crit Care Med. 2012; 40(5): 1499-1505. doi:
10.1097/CCM.0b013e318241e34e
54. Simmonds PC, Primrose JN, Colquitt JL, et al. Surgical re-
section of hepatic metastases from colorectal cancer: a system-
atic review of published studies. Br J Cancer. 2006; 94(7): 982-
999. doi: 10.1038/sj.bjc.6603033
55. Clavien PA, Petrowsky H, DeOliveira ML, Graf R. Strategies
for safer liver surgery and partial liver transplantation. N Engl J
Med. 2007; 356(15): 1545-1559. doi: 10.1056/NEJMra065156
56. Van Cutsem E, Nordlinger B, Adam R, et al. Towards a pan-
European consensus on the treatment of patients with colorectal
liver metastases. Eur J Cancer. 2006; 42(14): 2212-2221.
57. Kishi Y, Abdalla EK, Chun YS, et al. Three hundred and
one consecutive extended right hepatectomies: evaluation of
outcome based on systematic liver volumetry. Ann Surg. 2009;
250(4): 540-548. doi: 10.1097/SLA.0b013e3181b674df
58. Kopetz S, e Chang GJ, Overman MJ, t al. Improved survival
in metastatic colorectal cancer is associated with adoption of he-
patic resection and improved chemotherapy. J Clin Oncol. 2009;
Page 60

surgical research
Open Journal http://dx.doi.org/10.17140/SROJ-2-110
Surg Res Open J
ISSN 2377-8407
27(22): 3677-3683. doi: 10.1200/JCO.2008.20.5278
59. Abdalla EK, Adam R, Bilchik AJ, Jaeck D, Vauthey JN,
Mahvi D. Improving resectability of hepatic colorectal metasta-
ses: expert consensus statement. Ann Surg Oncol. 2006; 13(10):
1271-1280. doi: 10.1245/s10434-006-9045-5
60. Jeschke MG. The hepatic response to thermal injury: is the
liver important for postburn outcomes? Mol Med. 2009; 15(9-
10): 337-351. doi: 10.2119/molmed.2009.00005
Page 61