Summary: A heatmap, which focused upon 2458 genes that were involved in stem cell differentiation and liver development, was prepared to visualize the relative relationship between the transcriptomes of iPSCs, PHHs, human liver tissue and HO1. The dendrogram indicates that HO1 is more closely related to liver tissue than to PHHs; this is because cholangiocytes are present in HO1 and liver, but are absent (or rare) in PHHs. Examination of the genes within this heatmap indicated that there was a lack of expression of pluripotent genes in HO1; and that HO1 had an increased level of expression of cellular markers for liver lineages and liver development, which included mature hepatocytes, bile duct cells and NOTCH signaling components. A cluster analysis of 1510 genes indicates that there were major changes in the transcriptome after the cells that were dissociated from an HO1 were transferred from the growth medium into the differentiation medium. As with the HO1s, the gene expression profile of HO2s resembled that of PHHs and human liver. Of importance, the level of expression of mRNAs that are of importance for various human liver functions (ADH4, CYP3A4, TTR, GSRA1, TDO2, GSTA1 and FAH) were markedly increased in HO2s.
Overall Design: Whole transcriptome analysis was performed on iPSCs, primary human hepatocytes (PHH), HO1 (primary hepatic organoids) and human liver tissue. For this analysis, over 100 organoids that were generated from a single donor were analyzed. Next, whole transcriptome analysis was performed to characterize HO2s (secondary organoids) and their relationship to iPSCs, PHHs and human liver tissue. This analysis utilized 1,000 HO2s that were prepared from a single donor.
