Genome-wide analysis of DNA methylation and gene expression patterns in purified, uncultured human liver cells and activated hepatic stellate cells

Abstract

Background & Aims: Liver fibrogenesis -scarring of the liver that can lead to cirrhosis and liver cancer -is characterized by hepatocyte impairment, capillarization of liver sinusoidal endothelial cells (LSECs) and hepatic stellate cell (HSC) activation. To date, the molecular determinants of a healthy human liver cell phenotype remain largely uncharacterized. Here, we assess the transcriptome and the genome-wide promoter methylome specific for purified, non-cultured human hepatocytes, LSECs and HSCs, and investigate the nature of epigenetic changes accompanying transcriptional changes associated with activation of HSCs.Material and methods: Gene expression profile and promoter methylome of purified, uncultured human liver cells and culture-activated HSCs were respectively determined using Affymetrix HG-U219 genechips and by methylated DNA immunoprecipitation coupled to promoter array hybridization. Histone modification patterns were assessed at the single-gene level by chromatin immunoprecipitation and quantitative PCR.Results: We unveil a DNA-methylation-based epigenetic relationship between hepatocytes, LSECs and HSCs despite their distinct ontogeny. We show that liver cell type-specific DNA methylation targets early developmental and differentiationassociated functions. Integrative analysis of promoter methylome and transcriptome reveals partial concordance between DNA methylation and transcriptional changes associated with human HSC activation. Further, we identify concordant histone methylation and acetylation changes in the promoter and putative novel enhancer elements of genes involved in liver fibrosis.Conclusions: Our study provides the first epigenetic blueprint of three distinct freshly isolated, human hepatic cell types and of epigenetic changes elicited upon HSC activation.

Hepatic stellate cells as key target in liver fibrosis

“…HSC activation can be suppressed by inhibitors of DNMT1 (e.g., 5-Aza-2′-deoxycytidine) and enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2), a polycomb group-protein complex catalyzing trimethylation of histone 3 lysine 27 (H3K27me3) (e.g., 3-deazaneplanocin), supporting their regulatory role in the activation [256–258]. DNMT1-mediated DNA hypermethylation of phosphatase and tension homolog (PTEN) leads to HSC activation [259].…”

Liver Fibrosis: Mechanistic Concepts and Therapeutic Perspectives

“…Important epigenetic changes are induced by ROS in the HSCs, including chromatin remodeling by histone modification, DNA methylation and gene silencing by microRNAs (miRs) [158]. In-vitro and in-vivo approaches have demonstrated that HSCs show a global demethylation of fibrogenic genes during transdifferentiation into myofibroblasts, which is associated with liver fibrosis development [159][160][161].…”

Massive and Reproducible Production of Liver Buds Entirely from Human Pluripotent Stem Cells

“…Immunostaining of WT1, MIIA, and LHX2 also confirmed the correct cell fate induction in the presumptive STM cells ( Figure 3A). Global gene array analysis suggested iPSC-STM signatures showed a remarkable shift toward reported human adult hepatic mesenchymal cell signatures (Asahina et al, 2009(Asahina et al, , 2011El Taghdouini et al, 2015) ( Figure 2G). We previously demonstrated that one function of mesenchymal cells in LB generation is myosin-IIAdependent self-condensation.…”

“…Error bars represent the SD of values obtained from three independent experiments (n = 4). (G) Hierarchical clustering with AdHep mesenchymal cell signatures in iPSC-STM (Asahina et al, 2009(Asahina et al, , 2011El Taghdouini et al, 2015). directly anastomosed with mouse CD31 endothelial cells (Figure 3H) and were surrounded by iPSC-STMs at a perivascular location ( Figure 3I).…”