barcelona . spain - European Association for the Study of the Liver

BARCELONA . SPAIN
152 POSTGRADUATE COURSE SYLLABUS ALCOHOLIC LIVER DISEASE 153
APRIL 18 - 19/2012 THE INTERNATIONAL LIVER CONGRESS TM 2012
STEM CELLS IN ALCOHOLIC LIVER DISEASE: THE ROAD FROM
ANIMAL MODELS TO HUMAN STUDIES
Laurent Spahr
Geneva, Switzerland
E-mail: Laurent.Spahr@hcuge.ch
Grants: Supported by the Clinical Research Center, University Hospital and Faculty of Medicine, Geneva,
the Louis-Jeantet Foundation, and Foundation for Liver and Gut Studies (FLAGS) in Geneva
KEY POINTS
• The impaired regenerative capacity of alcoholic cirrhosis with ASH (acute-on chronic ALD)
participates to the high morbidity and mortality of the disease.
• Under specific experimental conditions, animal studies show that bone marrow (BM)-derived
stem cells participate in liver regeneration, but mechanisms are incompletely elucidated.
• Some benefits derived from BM-derived cell therapy seem associated with the co-administration
of G-CSF to mobilize hematopoietic stem cells, produce humoral factors, and facilitate cells
homing to the site of injury.
• In patients with cirrhosis and ASH, G-CSF stimulates the short-term proliferation of hepatic
progenitors.
• In patients with decompensated liver disease of mixed aetiology, results of the few RCTs
suggest that autologous BM stem cell transplantation is safe and may transiently improve liver
function in non alcoholic cirrhosis.
• Autologous BM stem cell transplantation in decompensated ALD doesn’t improve liver function
(MELD score) over a 3-month period.
• Future studies should better characterize the efficiency of G-CSF and BM stem cell therapy in
the regeneration and repair of decompensated alcoholic liver, and explore the fate and potential
of transplanted cells.
INTRODUCTION
Cirrhosis is associated with insufficient regeneration in response to injury, due to multiples causes including
extensive fibrosis, inflammation, steatosis and replicative senescence of hepatocytes, all of which constitute
an unsuitable environment for liver cell proliferation. Acute-on-chronic liver disease is a frequent clinical
presentation of ALD, and is associated with a poor outcome. Malnutrition and recent exposure to alcohol
have a negative influence on liver cell regeneration by influencing intracellular proliferative pathways. In
patients with decompensated ALD and biopsy-proven alcoholic ASH, supportive (N-acetylcystein, calories,
vitamins, alcohol abstinence) and anti-inflammatory strategies (corticosteroids) improve short-term survival
but overall prognosis remains poor with a 3-month mortality around 25%. Thus, new strategies to stimulate
liver regeneration and improve function are needed. As liver transplantation is often not an option at this
stage of the disease, a minimally invasive regenerative strategy would be welcome.
BONE MARROW CELLS AND LIVER REGENERATION
It has been shown that a subpopulation of hepatocytes and cholangiocytes may derive from the bone
marrow [1]. Under particular experimental conditions with strong selective pressure (for example
immunodeficient animals subject to sublethal liver injury followed by bone marrow transplantation,
fumarylacetatoacetate hydrolase (FAH) deficiency, exposition to hepatotoxin such as retrorsin or carbon
tetrachloride ), hepatocytes of bone marrow origin may participate in liver cell repopulation. However, the
precise mechanisms underlying the beneficial effect of this promising approach are still under investigation.
Firstly, bone marrow-derived pluripotent cells include hematopoietic and mesenchymal stem cells, as well
as endothelial progenitor cells, and the relative contribution of each of these cellular contingents in liver
repair is ill defined. Secondly, there is much controversy concerning the mechanism by which BM-derived
stem cells contribute to liver regeneration (cell fusion, transdifferentiation, local stimulation of endogenous
liver cell progenitors, production of cytokines and growth factors, remodelling of fibrous tissue). Thirdly, the
mechanisms driving these BM-derived stem cells to the site of liver injury probably involves multiple factors,
including stromal-derived factor-1 (ie: SDF-1 binding to CXR4 present on HSC surface). Fourthly, the rate of
engraftment of these cells in the diseased liver is difficult to determine, and seems to vary according to the
experimental model (< 1% to 50%). Finally, whether the administration of G-CSF (to mobilize BM cells and
promote growth factors production), is crucial in governing successful tissue repair is not yet determined.
Regarding safety, the small size of BM stem cells avoids them to be trapped within the sinusoids with the
potential risk of aggregation, ischemia, and increased portal pressure, but there are some concerns about
the risk of inducing cancer (particularly in culture-expanded cells) and the promotion of fibrosis (due to the
presence of myofibroblasts among BM cells).
IS BM CELLS MOBILIZATION USING G-CSF ASSOCIATED WITH LIVER REGENERATION
Experimental studies
Animals exposed to various, severe, often sublethal liver injury (but not related to alcohol) have demonstrated
an improved survival and less severe histological lesions when G-CSF was co-administered [2]. In a
combined toxic liver injury and partial hepatectomy model, G-CSF induced a strong proliferative activity
of oval cells (the intrahepatic progenitors), as well as the generation of a subpopulation of hepatocytes
originating in the bone marrow [3]. Thus, G-CSF seems to improve liver regeneration via both direct (the
liver stem cell niche) and indirect (contribution to the generation of bone marrow-derived hepatocytes)
mechanisms. It has also been suggested that G-CSF may contribute to the homing of bone marrow cells
to the injured liver.
Clinical studies
Transposition of experimental data with G-CSF into clinical studies with cirrhotics is challenging, with regard
to tolerance (musculoskeletal pain, fever, allergic reactions) and potentially severe side effects (rare cases
of splenic rupture). In a small uncontrolled pilot study [4] on advanced cirrhosis (Child-Pugh > 9) of mixed
aetiology, subcutaneous administration of G-CSF (5 ug/kg twice a day for 3 days) was associated with
increased circulating levels of CD34+ mobilized BM cells. Tolerance was excellent, although transient
increase in spleen size was reported during treatment. These results were confirmed in a larger group of
patients with acute decompensated cirrhosis, using either a 5 ug/kg/day or a 15 ug/kg/day dose for 5 days,
and increased circulating CD34+ cells were associated with a production of stromal cell derived factor
(SDF-1), a potent chemo attractant of hematopoietic progenitor cells, as well as the growth factor VEGF [5].
However, these biological effects didn’t translate into clinical improvement over a 3-week period. To further
explore the role of G-CSF in decompensated cirrhosis, we performed a randomized trial to study the shortterm
effect on liver regeneration in patients with decompensated ALD and superimposed ASH (median
Child-Pugh score of 10) [6]. Using a 5-day course of 10 ug/kg/day G-CSF treatment, circulating CD34+
hematopoietic stem cells were detectable in G-CSF treated patients, but the number of CD34+ cells was
lower than the value obtained in healthy stem cells donors. Using a double immunostaining methods for the
proliferation marker MiB1 and cytokeratin 7 and 18 (immunoreactivity for hepatic progenitors and mature
hepatocyte, respectively) performed on baseline and a repeat liver biopsy at day 7, we demonstrated a
significant increase in MiB1+/CK7+ and MiB1+/CK18+ hepatic cells in liver biopsy specimen limited to
G-CSF-treated patients. In addition, changes in circulating CD34+ and MiB1+/CK7+ in liver tissue between
baseline and day-7 values showed a positive correlation (r = 0.65, p < 0.03). Again, these encouraging
results on liver cell proliferation were not associated with any benefits in terms of liver function over a
4-week study period.