Introduction Biliary complications are still a major cause for
morbidity and mortality after liver transplantation. Ischemia/reperfusions
injury (IRI) leads to disruption of the biliary epithelium. Yet little is known
about the underlying molecular mechanism. We introduce a novel model to study the
effect of IRI on human cholangiocytes using extrahepatic cholangiocyte organoids
(ECOs).
Method Extrahepatic bile duct tissue was collected during liver
transplantation at static cold storage and after reperfusion (n=15).
Gallbladder tissue was used as control (n=5). ECOs (n=9) were
cultured from extrahepatic biliary tissue. Multiplex immunofluorescence, in-situ
hybridization and qRT-PCR were performed to study programmed cell death. IRI was
induced in ECOs by introducing cells to a hypoxic chamber for 48h, followed by
reoxygenation.
Results After reperfusion an activation of programmed cell death was
observed (p=0.002). Cultured ECOs formed circular structures recreating a
tubular structure similar to that found in the bile duct. Analysis revealed a
cholangiocyte phenotype of the ECOs. After hypoxia ECOs showed increased expression
of ACSL4 (p<0.0001) and VEGF- A (p<0.0001). HIF1-α
expression was increased after reoxygenation (p<0.0001). Expression patterns
were similar to those found in the bile duct biopsies at static cold storage.
Discussion ECOs are in-vitro cellular systems that self-organize
through mechanisms like those found in-vivo. They recapitulate the structure and
exhibit similar patterns of ACSL4, VEGF- A and HIF1-α expression as
extrahepatic bile duct during liver transplantation and thus provide a suitable
model to study IRI in choanocytes after liver transplantation.
