In vitro zonation of stem cell-derived hepatic stellate cells as a platform for liver fibrosis modeling
Liver cirrhosis is characterized by extensive fibrosis and represents a worldwide health burden with a significant death rate. Currently, there is no effective antifibrotic treatment available. Hepatic stellate cells are one of the key drivers of hepatic fibrosis and are implicated in inflammation and cancer. Recent studies revealed spatial zonation of HSCs across the hepatic lobule. In mouse model it has been shown that central vein-associated HSCs (CaHSCs) are pathogenic collagen-producing cells (https://doi.org/10.1016/j.celrep.2019.10.024). At the same time data about the zonation of human HSC and its impact in the disease progression is limited, as well as physiologically relevant 3D in vitro models for drug testing and discovery.
The aim of the current project is developing of 3D zonated organoids, generated from induced pluripotent stem cells and consisting of hepatocytes, endothelial cells and hepatic stellate cells for the testing of antifibrotic drugs.
During the project student will have the following tasks:
- Differentiation of induced pluripotent stem cells toward hepatic stellate cells
- Characterization of iPSC-derived HSC (iHSC) by immunofluirescence, qPCR and flow cytometry.
- Organoid formation using iPSC-derived hepatocytes (iHEP) and iHSC
- iHEP-iHSC organoids will be cultured in microfluidic chip with expose to the liver zone-specific morphogens and in contact with central vein endothelial cells.
- Evaluation of iPSC-HSCs capacity to respond to the fibrogenic signals (transforming growth factor beta 1 (TGF-b1), connective tissue growth factor(CTGF) and LPS.
- hPSC culture and differentiation
- Generation of organoids
- Cell culture in microfluidic devices
- Imaging (confocal microscopy, live cell imaging)
- ELISA, Bioluminescent assays
- Flow cytometry
- Data handling and statics