The bilious bowel: the relation between bile and bowel in development of biliary atresia and liver fibrosis. An explorative study

Biliary atresia (BA) is a rare disease of infancy. An unknown (infectious?)
event leads to bile duct scarring and liver injury. Only surgery -
reconstructing bile ducts via the so-called Kasai procedure - can postpone
liver scarring necessitating transplantation. Surgery is successful in
re-establishing bile flow in some 55% of cases. However, despite successful
surgery liver fibrosis rapidly progresses and liver failure ensues. A liver
transplantation will be necessary in some 70% of children with BA. BA is the
main indication for pediatric liver transplantation. As yet the driving force
of the rapidly developing fibrosis is unknown. Identifying the pro-fibrotic
factors at work in children with biliary atresia might offer novel therapeutic
avenues to prevent or postpone liver transplantation. Results from these
studies might also be applicable to other hepatobiliary diseases in children as
well as in adults.

The liver is continuously exposed to gut-derived products. Healthy individuals
develop tolerance. Susceptible neonates might fail to develop tolerance or be
exposed to higher level of danger signals due to a *leaky gut*. Scarring of
bile ducts diminishes bile flow. The absence of bile in the bowel changes the
gut microbiome and increases bowel permeability, inducing translocation and
further liver scarring: a vicious circle.

Activation of receptors on immune cells and bile duct cells
(Toll-like-receptors, TLR) by exposure to danger signals (e.g. bacterial or
viral products) induces liver scarring (*fibrosis*). Different TLRs respond to
different pathogens. Investigating which TLR is activated points towards the
pathogen inducing BA.

Many consider BA the result of an unknown perinatal (infectious?) insult,
leading to inappropriate immune activation aimed at bile duct cells. I
hypothesize that BA and BA associated liver fibrosis is due to immune
over-activation, triggered by persistent exposure to gut-derived products.
This *leaky gut* may be due to *tight junction* dysfunctioning (gaps in the
bowel) and/or a change in microbiome.

To investigate the origin of BA we will assess which TLR(s) is/are activated.
To test the hypothesis that during development of BA and associated liver
fibrosis there is a *leaky gut* and/or a change in microbiome we will use human
material to assess translocation markers in blood, bowel tight junction
functioning, and the gut microbiome. Data will be related to clinical outcomes
such as grade of fibrosis in pre-/preoperative liver biopsies, clearance of
jaundice and transplantation-free survival. Data from BA patients will be
compared to data from patients with other hepatobiliary diseases such as
patients with choledochal malformation, Progressive Familial Intrahepatic
Cholestasis (PFIC) and Alagille*s disease as well 30 healthy controls (children
with inguinal hernia or jejunal atresia - who are the same age as children with
biliary atresia. We will also compare BA patients who need to undergo liver
transplantation (most of them before their fifth year) and those who survive
for a prolonged time with their own liver. This will shed light on the effects
of the microbiome and bowel wall integrity on the progression of liver
fibrosis.

prospective study, explorative in nature

There will be no risk or burden for the children, who will not have to undergo
extra biopsies or venapunctures for this research. Collection of faeces from
the diaper will also not be a burden. During the Kasai procedure as well as
during surgery for choledochal malformation a Roux Y reconstruction
(anastomosis between two parts of small bowel in Y-form) is performed, which
makes it easy and without any risk to obtain a small part of small bowel.
Obtaining a small piece of bowel during the performance of a partial external
biliary diversion (a *bile-stoma* constructed of a loop of bowel anastomosed
between the gallbladder and the skin) in children with PFIC/Alagille is also
without any risk or burden for the same reasons. Obtaining the jejunal tissue
during the closure of the ostomy in children with jejunal atresia is also
without risk or burden, as the piece of small bowel is otherwise discarded.
Liver biopsies are routinely obtained during or prior to the Kasai procedure as
well as during other hepatobiliary surgery for clinical reasons. This also
holds true for obtaining ascites and a mesenteric lymph node.
Collecting faeces from the diaper of a child undergoing surgery for inguinal
hernia is also not a burden. CHildren undergoing surgery for inguinal hernia
always get an I.V. line prior to surgery: blood will be collected from the I.V.
This wil not be a burden or a risk.

As biliary atresia and most of the other aforementioned diseases are diseases
only occurring in young children, this study can only be performed in this
patient group of young children