Abstract
Rare pediatric liver disorders can be very destructive to the lives of patients and their families. Often, therapeutic options are limited to symptomatic care and precise disease mechanisms remain elusive. The relatively low incidence of each individual disease and the often widespread geographic distribution of patients complicate research and treatment
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development. In this thesis, we have investigated the potential of patient-derived liver organoids to study such rare pediatric liver diseases. We characterized intrahepatic cholangiocyte organoids (ICOs) from several monogenic liver disorders and showed that organoids express affected proteins of diseases such as cystic fibrosis, Wilson disease and methylmalonic acidemia. We uncovered that ICOs possess a hybrid phenotype, combining hepatocyte and cholangiocyte characteristics. This led us to investigate whether patient-derived liver organoids would be interesting to study the rare perinatal disease biliary atresia (BA), in which the hepatobiliary tree is occluded and becomes fibrotic. We found that organoids from different hepatobiliary regions (intrahepatic, extrahepatic and gallbladder) of BA patients can be cultured and biobanked. Further characterization showed that BA patient organoids display BA specific growth behavior and increased sensitivity to viral infections. While we showed that various hepatocyte and cholangiocyte functions can be studied in liver organoids, our data also indicated that several hepatocyte functions are currently limited in this in vitro system. Therefore, we devised a novel culture strategy to increase hepatic maturation in liver organoids. We found that hepatic functions, such as drug metabolism, improve when liver organoid cells are cultured on hollow fiber membranes coated with extracellular matrix proteins of the hepatic niche. Moreover, we demonstrated that hepatic transepithelial transporter defects such as progressive familial intrahepatic cholestasis type 3 can be studied in patient-derived liver organoids.
In summary, we have demonstrated that patient-derived liver organoids are a useful tool to study various rare liver and metabolic disorders affecting the hepatobiliary tract. Several diseases such as BA, cystic fibrosis, Wilson disease, progressive familial intrahepatic cholestasis type 3 and methylmalonic acidemia can currently be studied with ICOs. Although ICOs were previously praised as a new break-through hepatocyte model, we have shown that organoids from all hepatobiliary regions are at least equally useful for researching cholangiopathies, such as BA. The development of new technologies, such as organ chips and co-culture strategies, are exciting not only to study the disease mechanisms of rare diseases but also to improve the hepatic maturity of ICOs. Applying the myriad of culture improvement strategies to ICOs will likely produce a fantastic patient-derived hepatocyte model. We have taken the first steps toward improving hepatic maturation of ICOs and in doing so have demonstrated that the ICO application range can be broadened to facilitate the study of (cholestatic) disease mechanisms. Similarly, we anticipate that these improvements in hepatic maturation of ICOs will aid in the development of safe therapeutics for (rare) liver diseases in a personalized manner in the future.
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