Background & Aims:
Submassive hepatic necrosis (SMHN) is the defining histological feature of acute-on-chronic
liver failure (ACLF). In the condition of SMHN, liver progenitor cells (LPC)-mediated
regeneration is crucial for survival and recovery of ACLF patients. LPC-mediated liver
regeneration comprises three consecutive steps: formation of reactive ducts by activated
LPC, LPC differentiation towards hepatocytes, and functional bile canaliculi formation
by newly generated hepatocytes. In this study, we examined which steps play crucial
roles in ACLF recovery.
Methods:
We investigated liver tissue specimens of 55 ACLF patients with a known interval between
the first symptoms of acute decompensation and the time of tissue sampling. Immunostaining
for Cytokeratin 7 and apical molecules, including BSEP, MDR1, CD10 and CD26, were
used to identify LPC and bile canaliculi. Electronic microscopy was performed to detect
the existence of bile canaliculi. Expression of CYP7A1, the rate-limiting enzyme for
bile acid synthesis, and nuclear receptors such as farnesoid X receptor (FXR) and
small heterodimer partner (SHP) was measured by immunohistochemistry. In collagen
sandwich cultured primary hepatocytes, we examined how nuclear receptors regulate
expression of apical molecules.
Results:
Rapid ductular reaction occurs in all ACLF patients regardless of clinical duration
and outcome. LPCs differentiate into hepatocytes over time. However, LPC-derived hepatocytes
do not improve liver function in irreversible ACLF patients. Electronic microscopy
analysis reveals that although these hepatocytes maintain a typical polar structure,
bile canaliculi present without microvilli. Immunostaining analyses further show that
in contrast to recovered patients, hepatocytes in irreversible ACLF patients have
lost functional molecules in the bile canali, including CD26, CD10, and BSEP, whereas
MDR1 expression is maintained. The loss of hepatocyte membrane transporters, e.g.
BSEP, in irreversible ACLF is associated with a failed feedback regulatory mechanism
of bile acid synthesis, which is characterized by high levels of CYP7A1, but lack
of nuclear FXR and SHP in hepatocytes. In vitro, FXR agonist GW4064 up-regulates expression
of apical molecules, including BSEP and MRP2, whereas depleting FXR inhibits expression
of BSEP and MRP2, but not MDR1. Furthermore, the effects of FXR are SHP-dependent.
Conclusions:
Functional bile canaliculi are required to restore intact liver function following
SMHN, thus determining recovery in ACLF. Maintenance of functional hepatocellular
polarity is at least partially controlled by nuclear receptors, in particular the
FXR-SHP axis.
