Molecular mechanisms in hepatocyte de- and re-differentiation. Background: In vivo, hepatocytes are found most frequently as terminally committed cells with low turnover [ref]. It is in the case of certain injuries that these cells are capable of switching to a less differentiated state in which they are highly dividing. The molecular mechanisms and signatures behind this process are at present largely unexplored. Study aims and methodology: We aim at studying those mechanisms in human liver cells using a 3D cell culture system in which hepatocyte de- and, importantly, also re-differentiation can be observed in vitro. We find that during the initial stages of spheroid formation, the cells acquire a rather undifferentiated phenotype. However, during aggregation and the establishment of cell-to-cell contacts, these cells progressively become committed to a hepatocyte phenotype. Thus, we aim at performing a comprehensive study of the molecular signatures and changes ongoing during these events. We will employ high throughput omics technologies– including transcriptomics by RNAseq and proteomics by mass spectrometry, plus epigenomics and metabolomics. We will obtain data at a high temporal resolution by running cell samples at several time points during the cell de-differentiation and re-aggregation process in spheroid formation. We aim at performing an exhaustive unbiased analysis of the data in order to elucidate key molecular signatures and events.