FGF19 in obstructive cholestasis: unveil the signal

Bile salts are potent signalling molecules influencing various metabolic and
functional processes. Bile salts exert these functions by activating nuclear
(e.g. FXR ) and plasma cell membrane-bound receptors (e.g. TGR5) which are
expressed in several tissues (e.g. liver, small intestine, colon, kidney and
gallbladder). Bile salts regulate their own biosynthesis by controlling the
transcription of the hepatic bile salt synthetic enzyme CYP7A1. Two pathways
are involved in the negative feedback control of bile salt synthesis: i) the
hepatic FXR-SHP pathway and ii) the ileal FXR-FGF19 pathway. Studies showed
that the latter is more prominent in controlling CYP7A1 transcript levels (viz.
bile salt synthesis). Thus, bile salts are synthesized in the liver, excreted
in bile and expelled by the gallbladder into the proximal intestine (to aid in
lipid absorption and digestion) and reabsorbed in the terminal ileum to recycle
back to the liver via portal blood. Bile salts reclaimed from the intestinal
lumen by the ileocyte, activate FXR. This induces the expression of an
enterokine, FGF19, which signals via portal blood to the liver to activate its
receptor which initiates downstream signalling to repress bile salt synthesis.
The FXR/FGF19 signalling pathway is the subject of the present study.

Patients with obstructive cholestasis (=accumulation of bile) caused by
malignancies (e.g. pancreatic cancer, cholangiocarcinoma) have a perturbed
enterohepatic cycle. Obstructive cholestasis is associated with i) gut barrier
dysfunction, ii) endotoxemia, iii) bacterial overgrowth and iv) liver injury.
Previous study showed that FGF19 is expressed in the liver of patients with
obstructive cholestasis. However, knowledge about the contribution of FGF19
protein by the gut in obstructive cholestasis has thus far been unexplored.
Preliminary findings revealed that FGF19 is produced by the portal drained
viscera (viz. intestine) of non-cholestatic patients undergoing liver surgery.
The inter-organ signalling of FGF19 in an obstructed entero-hepatic cycle has
not yet been characterized and likewise the metabolic and other functional
effects of inflicted FGF19 signalling during cholestasis have not been
clarified.

The hypothesis is that the FXR-FGF19 pathway is disturbed in patients with
obstructive cholestasis, and this is associated with organ injury and metabolic
dysfunction. We postulate that FGF19 is not produced by the terminal ileum
under conditions of obstructive cholestastic, but production is shifted to the
liver and this affects metabolic processes.

The aim of this study is to investigate FGF19 signalling in patients with
cholestasis compared to non-cholestatic patients or post-cholestatic patients
(drained patients) by calculating fluxes across the portal drained organs.
Secondly, we aim to investigate the metabolic and functional consequences
(glucose, lipid homeostasis, cholestatic itch, gut barrier function) of a
disturbed FXR-FGF19 pathway in humans. This study will provide insights that
may lead to potential therapeutic strategies for patients with a disturbed
enterohepatic cycle (e.g. cholestatic liver diseases).

To investigate FGF19 signalling and metabolic and functional consequences of a
disturbed enterohepatic cycle in patients with obstructive cholestasis

Study design: This is a prospective multi-center observational study comparing
non-cholestatic patients (control group), post-cholestatic patients with a
biliary stent (restored enterohepatic cycle group) with cholestatic patients
(cholestatic group).

Patients planned for a Whipple procedure are included. Informed consent will be
obtained either at the outpatient ward or at the day of admission, generally
one day before surgery. Patients will then also have time to ask questions.

Blood from these patients will be sampled during surgery under general
anesthesia from the portal vein, hepatic vein, superior mesenteric vein,
inferior mesenteric vein, splenic vein, renal vein and the radial artery. The
experimental set-up consists of 1 time arterial blood sampling (10ml) and 1
time intra-abdominal blood sampling (6x10ml) which is maximal 70 ml in total.
During surgery blood flow will be measured of the portal vein and hepatic
artery to precisely calculate organ fluxes. The portal vein and hepatic artery
are easy accessible for flow measurement with the Transonic flow meter. This
device is used routinely to measure actual blood flow. No risks are associated
with measuring of these blood flows.

Additionally, bile will be sampled during surgery (4 ml) from the gallbladder
or hepatic duct during surgery, and biopsies will be taken from the liver (1),
gallbladder (1), jejunum (1), common bile duct (CBD, 1) and white adipose
tissue (WAT, from three sites; subcutaneous, omental and visceral adipose
tissue for gene expression studies of genes.. Moreover, preoperative stool and
urine, and jejunal content during surgery will be collected for detailed
analysis of microbiota/bile salt composition to investigate the effect of
obstructive cholestasis on these parameters. Patients will be assessed for
severity of itch by a questionnaire (visual analog scale and 5D itch scale) on
the day before surgery to correlate fluxes to cholestatic itch.

The methods applied, i.e. arterial and intra-abdominal blood sampling and
collecting liver/jejunal biopsies, have been used previously without any
consequences for the surgical procedures or the patients (MEC 02-045, MEC
03-032, MEC 06-2067 and MEC 11-3-084) as published by Van de Poll et al35. In
comparison to previously approved MEC protocols, bile (4 ml) will be sampled
from the gallbladder or hepatic duct during surgery, and biopsies of the
gallbladder (1), CBD (1) and WAT (3), as well as stool will be collected one
time preoperatively. There are no additional risks related to the collection of
bile and tissues since these are part of the resected tissues. No further risk
is associated with the collection of preoperative stool and urine. Although
the results of this project have no direct positive effects for the patients
involved, they do contribute to the understanding of the role of bile salts and
FGF19 signaling under cholestatic conditions. Insights from this study would
provide novelty and substantial knowledge in possible effects of FGF19 therapy
in cholestatic liver injury.