An automated and multiplexed liver fibrosis screening assay on a microfluidic liver model that replicates the cellular composition and organization of the hepatic lobule
Abstract: Mimicking the complex biology and physiology of the liver in vitro is imperative in order to decipher the mechanisms of liver diseases and develop effective therapies for patients. These models should not only capture the cellular composition and interactions of the native tissue, but also be translated in sensitive and predictive assays for drug discovery and development. Persistent efforts have been dedicated to developing various complex in vitro human liver models, but they often result in intricate microfluidic setups that are incompatible with automated procedures and do not scale well. Alternatively, scalable and automated models often lack biological complexity and therefore, the necessary cellular interactions to predict drug responses in the context of complex diseases. We developed a liver model that captures the cellular organization and interactions found within a native liver lobule. The model was developed by embedding primary or stem cell-derived liver cells, including both parenchymal and non-parenchymal cells in a hydrogel within a microfluidic chip. Cellular reorganization led to polarized hepatocytes with a distinct blood and bile side, functional liver sinusoidal vasculature with associated stellate cells and functional resident immune cells. The platform itself comprises 64 individual chip units, patterned underneath a microtiter plate and is therefore compatible with standard laboratory equipment. We deployed our model for a liver fibrosis screen, a complex disease for which there is yet no drug approved. We induced a fibrotic phenotype, presented as an increased expression of stellate cell activation markers and cytokine release which could be ameliorated after addition of a control compound. With this trigger and control compound, we developed a sensitive fibrosis assay by multiplexing two imaging redouts using high content imaging systems and cytokine release measurements. This yielded a very reproducible assay, with a sufficiently large assay window to be suitable for high-throughput screenings (Z’ > 0.5). We also multiplexed toxicity and viability readouts in order to discriminate between fibrosis inhibition as a result of effective inhibition or cellular toxicity. Finally, we screened a validation library of clinically relevant compounds in a fully automated workflow. In summary, we developed an in vitro liver model comprising a vascular network, polarized hepatocytes and functional hepatic cells with cellular organization reminiscent of the liver lobule. We showed induction of liver fibrosis in an assay that could successfully be screened for both efficacy and toxicity. In conjunction with its high throughput capability, this has the potential to revolutionize drug discovery and develop therapies for complex liver diseases.